DEPROTECTION METHOD AND RESIN REMOVAL METHOD IN SOLID-PHASE REACTION FOR PEPTIDE COMPOUND OR AMIDE COMPOUND, AND METHOD FOR PRODUCING PEPTIDE COMPOUND

Abstract

The present inventors found that peptide compounds/amide compounds in which the protecting groups of interest are removed and/or which are removed from resins for solid-phase synthesis can be produced without main chain damage by contacting starting peptide compounds/amide compounds with silylating agents.

Claims

1. A method of producing a peptide compound in which a protecting group removable by a silylating agent is removed, the method comprising the step of contacting a starting peptide compound comprising natural amino acid residues and/or amino acid analog residues with the silylating agent in a solvent and thereby removing the protecting group from the starting peptide compound, wherein the silylating agent is prepared by mixing a silyl compound or acid with an electrophilic species scavenger, wherein the starting peptide compound comprises at least one protecting group removable by the silylating agent, and wherein the starting peptide compound comprises at least one N-substituted amino acid residue.

2. A method of producing a peptide compound in which a resin for solid-phase synthesis that is removable by a silylating agent is removed, the method comprising the step of contacting a starting peptide compound comprising natural amino acid residues and/or amino acid analog residues with the silylating agent in a solvent and thereby removing the resin for solid-phase synthesis from the starting peptide compound, wherein the silylating agent is prepared by mixing a silyl compound or acid with an electrophilic species scavenger, wherein the starting peptide compound is linked to the removable resin for solid-phase synthesis, and wherein the starting peptide compound comprises at least one N-substituted amino acid residue.

3. The method of claim 1 or 2, wherein the starting peptide compound comprises at least one structure in which at least two amino acid residues are linked to each other, wherein the structure is represented by general formula (I) below: ##STR00179## wherein R.sub.1 is hydrogen, PG.sub.1, a natural amino acid residue, or an amino acid analog residue; R.sub.2 is selected from the group consisting of hydrogen and C.sub.1-C.sub.6 alkyl, or R.sub.2 and R.sub.4 or R.sub.2 and R.sub.4′, together with the nitrogen atom and carbon atom to which they are attached, form a 3- to 7-membered heterocyclic ring optionally substituted with hydroxy or C.sub.1-C.sub.4 alkoxy, wherein R.sub.4′ is hydrogen when R.sub.2 and R.sub.4 together form the heterocyclic ring, and R.sub.4 is hydrogen when R.sub.2 and R.sub.4′ together form the heterocyclic ring; except when R.sub.2 and R.sub.4, or R.sub.2 and R.sub.4T together form the heterocyclic ring, (a) R.sub.4′ is hydrogen, and R.sub.4 is selected from the group consisting of hydrogen, optionally substituted C.sub.1-C.sub.6 alkyl, C.sub.3-C.sub.6 cycloalkyl, C.sub.3-C.sub.6 cycloalkyl-C.sub.1-C.sub.4 alkyl, optionally substituted phenyl, optionally substituted phenylmethyl, optionally substituted phenylethyl, 2-(methylthio)ethyl, —CH.sub.2SPG.sub.2, N-PG.sub.3-indol-3-ylmethyl, 4-(PG.sub.2O)benzyl, PG.sub.2-O-methyl, 1-(PG.sub.2O)ethyl, 2-(PG.sub.2O)ethyl, PG.sub.2-OCO(CH.sub.2)—, PG.sub.2-OCO(CH.sub.2).sub.2—, PG.sub.3N-n-butyl, —CON(R.sub.14A)(R.sub.14B), —CH2-CON(R.sub.14A)(R.sub.14B), and —(CH2)2CON(R.sub.14A)(R.sub.14B), (b) R4 and R4′ are independently optionally substituted C1-C6 alkyl, or (c) R4 and R4′, together with the carbon atom to which they are attached, form a 3-to 7-membered alicyclic ring; R.sub.5 is a single bond or —C(R.sub.5A)(R.sub.5B)—; R.sub.5A and R.sub.5B are independently selected from the group consisting of hydrogen, C.sub.1-C.sub.6 alkyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted aryl-C.sub.1-C.sub.4 alkyl, and optionally substituted heteroaryl-C.sub.1-C.sub.4 alkyl; R.sub.6 is selected from the group consisting of hydrogen and C.sub.1-C.sub.6 alkyl, or R.sub.6 and R.sub.7 or R.sub.6 and R.sub.7′ together with the nitrogen atom and carbon atom to which they are attached, form a 3- to 7-membered heterocyclic ring optionally substituted with hydroxy or C.sub.1-C.sub.4 alkoxy, wherein R.sub.7′ is hydrogen when R.sub.6 and R.sub.7 together form the heterocyclic ring, and R.sub.7 is hydrogen when R.sub.6 and R.sub.7′ together form the heterocyclic ring; except when R.sub.6 and R.sub.7 or R.sub.6 and R.sub.7′ together form the heterocyclic ring, (a) R.sub.7′ is hydrogen, and R.sub.7 is selected from the group consisting of hydrogen, optionally substituted C.sub.1-C.sub.6 alkyl, C.sub.3-C.sub.6 cycloalkyl, C.sub.3-C.sub.6 cycloalkyl-C.sub.1-C.sub.4 alkyl, optionally substituted phenyl, optionally substituted phenylmethyl, optionally substituted phenylethyl, 2-(methylthio)ethyl, —CH.sub.2SPG.sub.4, N-PG.sub.5-indol-3-ylmethyl, 4-(PG.sub.4O)benzyl, PG.sub.4-O-methyl, 1-(PG.sub.4O)ethyl, 2-(PG.sub.4O)ethyl, PG.sub.4-OCO(CH.sub.2)—, PG.sub.4-OCO(CH.sub.2).sub.2—, PG.sub.5N-n-butyl, —CON(R.sub.15A)(R.sub.15B), —CH.sub.2—CON(R.sub.15A)(R.sub.15B), and —(CH.sub.2).sub.2CON(R.sub.15A)(R.sub.15B), or (b) R.sub.7 and R.sub.7′ are independently optionally substituted C.sub.1-C.sub.6 alkyl, or (c) R.sub.7 and R.sub.7′, together with the carbon atom to which they are attached, form a 3- to 7-membered alicyclic ring; R.sub.8 is a single bond or —C(R.sub.8A)(R.sub.8B)—; R.sub.8A and R.sub.8B are independently selected from the group consisting of hydrogen, C.sub.1-C.sub.6 alkyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted aryl-C.sub.1-C.sub.4 alkyl, and optionally substituted heteroaryl-C.sub.1-C.sub.4 alkyl; R.sub.9 is hydroxy, —O-PG.sub.6, a natural amino acid residue, an amino acid analog residue, —O-RES, or —NH-RES; RES is a resin for solid-phase synthesis; R.sub.14A and R.sub.14B are independently hydrogen or C.sub.1-C.sub.4 alkyl, or R.sub.14A and R.sub.14B, together with the nitrogen atom to which they are attached, form a 4- to 8-membered ring optionally comprising one or more additional heteroatoms; R.sub.15A and R.sub.15B are independently hydrogen or C.sub.1-C.sub.4 alkyl, or R.sub.15A and R.sub.15B, together with the nitrogen atom to which they are attached, form a 4- to 8-membered ring optionally comprising one or more additional heteroatoms; PG.sub.1 is selected from the group consisting of Fmoc, Boc, Alloc, Cbz, Teoc, and trifluoroacetyl; PG.sub.2 and PG.sub.4 are independently selected from the group consisting of hydrogen, t-Bu, trityl, methoxytrityl, cumyl, benzyl, THP, 1-ethoxyethyl, methyl, ethyl, allyl, optionally substituted aryl, optionally substituted aryl-C.sub.1-C.sub.4 alkyl, optionally substituted heteroaryl-C.sub.1-C.sub.4 alkyl, and 2-(trimethylsilyl)ethyl; PG.sub.3 and PG.sub.5 are independently selected from the group consisting of hydrogen, Fmoc, Boc, Alloc, Cbz, Teoc, methoxycarbonyl, t-Bu, trityl, cumyl, and benzyl; and PG.sub.6 is selected from the group consisting of t-Bu, trityl, cumyl, benzyl, methyl, ethyl, allyl, and 2-(trimethylsilyl)ethyl.

4. The method of any one of claims 1 to 3, wherein the starting peptide compound comprises at the C-terminus a structure in which at least two amino acid residues are linked to each other, wherein the structure is represented by general formula (II) below: ##STR00180## wherein R.sub.1′ is a group represented by the formula (III): ##STR00181## * represents the point of attachment; R.sub.1 is hydrogen, PG.sub.1, a natural amino acid residue, or an amino acid analog residue; R.sub.2 is selected from the group consisting of hydrogen and C.sub.1-C.sub.6 alkyl, or R.sub.2 and R.sub.10, or R.sub.2 and R.sub.10′, together with the nitrogen atom and carbon atom to which they are attached, form a 3- to 7-membered heterocyclic ring optionally substituted with hydroxy or C.sub.1-C.sub.4 alkoxy, wherein R.sub.10′ is hydrogen when R.sub.2 and R.sub.10 together form the heterocyclic ring, and R.sub.10 is hydrogen when R.sub.2 and R.sub.10′ together form the heterocyclic ring; except when R.sub.2 and R.sub.10 or R.sub.2 and R.sub.10′ together form the heterocyclic ring, (a) R.sub.10′ is hydrogen, and R.sub.10 is selected from the group consisting of hydrogen, optionally substituted C.sub.1-C.sub.6 alkyl, C.sub.3-C.sub.6 cycloalkyl, C.sub.3-C.sub.6 cycloalkyl-C.sub.1-C.sub.4 alkyl, optionally substituted phenyl, optionally substituted phenylmethyl, optionally substituted phenylethyl, 2-(methylthio)ethyl, —CH.sub.2SPG.sub.8, N-PG.sub.9-indol-3-ylmethyl, 4-(PG.sub.8O)benzyl, PG.sub.8-O-methyl, 1-(PG.sub.8O)ethyl, 2-(PG.sub.8O)ethyl, PG.sub.8-OCO(CH.sub.2)—, PG.sub.8-OCO(CH.sub.2).sub.2—, PG.sub.9N-n-butyl, —CON(R.sub.16A)(R.sub.16B), —CH.sub.2—CON(R.sub.16A)(R.sub.16B), and —(CH.sub.2).sub.2CON(R.sub.16A)(R.sub.16B), or (b) R.sub.10 and R.sub.10′ are independently optionally substituted C.sub.1-C.sub.6 alkyl, C.sub.3-C.sub.6 cycloalkyl, or C.sub.3-C.sub.6 cycloalkyl-C.sub.1-C.sub.4 alkyl, or (c) R.sub.10 and R.sub.10′, together with the carbon atom to which they are attached, form a 3- to 7-membered alicyclic ring; R.sub.11 is a single bond or —C(R.sub.11A)(R.sub.11B)—; R.sub.11A and R.sub.11B are independently selected from the group consisting of hydrogen, C.sub.1-C.sub.6 alkyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted aryl-C.sub.1-C.sub.4 alkyl, and optionally substituted heteroaryl-C.sub.1-C.sub.4 alkyl; R.sub.12 and R.sub.12′ are independently selected from the group consisting of hydrogen, PG.sub.10-O-methyl, —(CH.sub.2).sub.nCOO-PG.sub.10, —(CH.sub.2).sub.nCOO-RES, and —(CH.sub.2).sub.nCONH-RES; RES is a resin for solid-phase synthesis; n is 0, 1, or 2; R.sub.6 is selected from the group consisting of hydrogen and C.sub.1-C.sub.6 alkyl; R.sub.13 is C.sub.1-C.sub.4 alkyl or —(CH.sub.2).sub.mCON(R.sub.17A)(R.sub.17B); m is 0, 1, or 2; R.sub.16A and R.sub.16B are independently hydrogen or C.sub.1-C.sub.4 alkyl, or R.sub.16A and R.sub.16B, together with the nitrogen atom to which they are attached, form a 4- to 8-membered ring optionally comprising one or more additional heteroatoms; R.sub.17A and R.sub.17B are independently hydrogen or C.sub.1-C.sub.4 alkyl, or R.sub.17A and R.sub.17B, together with the nitrogen atom to which they are attached, form a 4- to 8-membered ring optionally comprising one or more additional heteroatoms; PG.sub.1 is independently selected from the group consisting of Fmoc, Boc, Alloc, Cbz, Teoc, and trifluoroacetyl; PG.sub.8 is selected from the group consisting of hydrogen, t-Bu, trityl, methoxytrityl, cumyl, benzyl, THP, 1-ethoxyethyl, methyl, ethyl, allyl, optionally substituted aryl, optionally substituted aryl-C.sub.1-C.sub.4 alkyl, optionally substituted heteroaryl-C.sub.1-C.sub.4 alkyl, and 2-(trimethylsilyl)ethyl; PG.sub.9 is selected from the group consisting of hydrogen, Fmoc, Boc, Alloc, Cbz, Teoc, methoxycarbonyl, t-Bu, trityl, cumyl, and benzyl; and PG.sub.10 is selected from the group consisting of t-Bu, trityl, cumyl, benzyl, methyl, ethyl, allyl, optionally substituted aryl, optionally substituted aryl-C.sub.1-C.sub.4 alkyl, optionally substituted heteroaryl-C.sub.1-C.sub.4 alkyl, and 2-(trimethylsilyl)ethyl.

5. A method of producing an amide compound in which a protecting group removable by a silylating agent is removed, the method comprising the step of contacting a starting amide compound with the silylating agent in a solvent and thereby removing the protecting group from the starting amide compound, wherein the silylating agent is prepared by mixing a silyl compound or acid with an electrophilic species scavenger, wherein the starting amide compound is represented by general formula (II) below: ##STR00182## wherein R.sub.1′ is a hydrogen atom or PG.sub.7; R.sub.12 and R.sub.12′ are independently selected from the group consisting of hydrogen, PG.sub.10-O-methyl, —(CH.sub.2).sub.nCOO-PG.sub.10, —(CH.sub.2).sub.nCOO-RES, and —(CH.sub.2).sub.nCONH-RES; RES is a resin for solid-phase synthesis; n is 0, 1, or 2; R.sub.6 is selected from the group consisting of hydrogen and C.sub.1-C.sub.6 alkyl; R.sub.13 is C.sub.1-C.sub.4 alkyl or —(CH.sub.2).sub.mCON(R.sub.17A)(R.sub.17B); m is 0, 1, or 2; R.sub.17A and R.sub.17B are independently hydrogen or C.sub.1-C.sub.4 alkyl, or R.sub.17A and R.sub.17B, together with the nitrogen atom to which they are attached, form a 4- to 8-membered ring optionally comprising one or more additional heteroatoms; PG.sub.7 is selected from the group consisting of Fmoc, Boc, Alloc, Cbz, Teoc, and trifluoroacetyl; and PG.sub.10 is selected from the group consisting of t-Bu, trityl, cumyl, benzyl, methyl, ethyl, allyl, optionally substituted aryl, optionally substituted aryl-C.sub.1-C.sub.4 alkyl, optionally substituted heteroaryl-C.sub.1-C.sub.4 alkyl, and 2-(trimethylsilyl)ethyl, and wherein the starting amide compound comprises at least one protecting group removable by the silylating agent.

6. A method of producing an amide compound in which a resin for solid-phase synthesis is removed, the method comprising the step of contacting a starting amide compound with a silylating agent in a solvent and thereby removing the starting amide compound from the resin for solid-phase synthesis, wherein the silylating agent is prepared by mixing a silyl compound or acid with an electrophilic species scavenger, and wherein the starting amide compound is represented by general formula (II) below: ##STR00183## wherein R.sub.1′ is a hydrogen atom or PG.sub.7; R.sub.12 and R.sub.12′ are independently selected from the group consisting of hydrogen, PG.sub.10-O-methyl, —(CH.sub.2).sub.nCOO-PG.sub.10, —(CH.sub.2).sub.nCOO-RES, and —(CH.sub.2).sub.nCONH-RES; RES is a resin for solid-phase synthesis, wherein at least one of R.sub.12 and R.sub.12′ is —(CH.sub.2).sub.nCOO-RES or —(CH.sub.2).sub.nCONH-RES; RES is a resin for solid-phase synthesis; n is 0, 1, or 2; R.sub.6 is selected from the group consisting of hydrogen and C.sub.1-C.sub.6 alkyl; R.sub.13 is C.sub.1-C.sub.4 alkyl or —(CH.sub.2).sub.mCON(R.sub.17A)(R.sub.17B); m is 0, 1, or 2; R.sub.17A and R.sub.17B are independently hydrogen or C.sub.1-C.sub.4 alkyl, or R.sub.17A and R.sub.17B, together with the nitrogen atom to which they are attached, form a 4- to 8-membered ring optionally comprising one or more additional heteroatoms; PG.sub.7 is selected from the group consisting of Fmoc, Boc, Alloc, Cbz, Teoc, and trifluoroacetyl; and PG.sub.10 is selected from the group consisting of t-Bu, trityl, cumyl, benzyl, methyl, ethyl, allyl, optionally substituted aryl, optionally substituted aryl-C.sub.1-C.sub.4 alkyl, optionally substituted heteroaryl-C.sub.1-C.sub.4 alkyl, and 2-(trimethylsilyl)ethyl.

7. The method of any one of claims 1 and 3 to 5, wherein the removable protecting group is selected from the group consisting of t-Bu, triphenylmethyl, 2-(trimethylsilyl)-ethyl, Boc, Teoc, Cbz, methoxycarbonyl, tetrahydropyranyl, 1-ethoxyethyl, methoxytrityl, and cumyl.

8. The method of any one of claims 1 to 7, wherein the silyl compound is represented by formula 1 below: ##STR00184## wherein R.sub.AX, R.sub.AY, and R.sub.AZ are independently C.sub.1-C.sub.4 alkyl or phenyl, and X is selected from the group consisting of —OTf, —OClO.sub.3, Cl, Br, and I.

9. The method of claim 8, wherein the silyl compound is selected from the group consisting of TMSOTf, TESOTf, TBSOTf, TIPSOTf, TBDPSOTf, TTMSOTf, TMSCl, TMSBr, TMSOClO.sub.3, and TMSI.

10. The method of any one of claims 1 to 7, wherein the acid is represented by HX, wherein X is selected from the group consisting of —OTf, —OClO.sub.3, Cl, Br, and I.

11. The method of any one of claims 1 to 10, wherein the electrophilic species scavenger is selected from the group consisting of formulas (2) to (10) below: ##STR00185## wherein in formula 2, R.sub.B is a substituted silyl group and R.sub.C is a substituted silyl group, or R.sub.B and R.sub.C, together with the nitrogen atom and carbon atom to which they are attached, form a 5- to 7-membered ring; and R.sub.D is C.sub.1-C.sub.4 alkyl optionally substituted with one or more fluorine atoms or is optionally substituted methylene, wherein when R.sub.D is optionally substituted methylene, formula 2 is dimerized to form a compound represented by the formula below: ##STR00186## wherein in formula 3, R.sub.G is a silyl group substituted with one or more C.sub.1-C.sub.4 alkyl; R.sub.H is hydrogen or C.sub.1-C.sub.4 alkyl; and R.sub.I is hydrogen, or C.sub.1-C.sub.4 alkyl optionally substituted with one or more fluorine atoms; wherein in formula 4, (a-1) R.sub.J is a substituted silyl group, R.sub.K is C.sub.1-C.sub.4 alkoxy, and R.sub.M and R.sub.L are independently hydrogen or C.sub.1-C.sub.4 alkyl; (a-2) R.sub.J is a substituted silyl group, R.sub.M is hydrogen or C.sub.1-C.sub.4 alkyl, and R.sub.K and R.sub.L, together with the carbon atoms to which they are attached, form a 5- to 8-membered ring comprising an oxygen atom; (b-1) R.sub.J is a substituted silyl group, R.sub.K is C.sub.1-C.sub.4 alkyl, and R.sub.M and R.sub.L are independently hydrogen or C.sub.1-C.sub.4 alkyl; (b-2) R.sub.J is a substituted silyl group, R.sub.M is hydrogen or C.sub.1-C.sub.4 alkyl, and R.sub.K and R.sub.L are taken together with the carbon atoms to which they are attached, form a 5- to 8-membered ring; or (c-1) RJ and RM, together with the carbon atoms to which they are attached, form a 5- to 7-membered ring comprising an oxygen atom, RK is hydrogen or C1-C4 alkyl, and RL is C1-C4 alkyl; (c-2) RJ and RM, together with the carbon atoms to which they are attached, form a 5- to 7-membered ring comprising an oxygen atom, and RK and RL, together with the carbon atom to which they are attached, form a 5- to 8-membered ring; (d-1) RJ is C1-C4 alkyl and RM, RK, and RL are independently hydrogen or C1-C4 alkyl; (d-2) RJ is C1-C4 alkyl, RM is hydrogen or C1-C4 alkyl, and RK and RL, together with the carbon atoms to which they are attached, form a 5- to 8-membered ring; (e-1) RJ is C1-C3 alkylcarbonyl and RM, RK, and RL are independently hydrogen or C1-C4 alkyl; (e-2) RJ is C1-C3 alkylcarbonyl, RM is hydrogen or C1-C4 alkyl, and RK and RL, together with the carbon atoms to which they are attached, form a 5- to 8-membered ring; (f-1) RJ is a substituted silyl group or C1-C4 alkyl, RK is optionally substituted di-C1-C4 alkylamino, and RM and RL are independently hydrogen or C1-C4 alkyl; or (f-2) RJ is a substituted silyl group or C1-C4 alkyl, RM is hydrogen or C1-C4 alkyl, and RK and RL, together with the carbon atoms to which they are attached, form a 5- to 8-membered ring comprising a nitrogen atom, wherein the 5- to 8-membered ring is optionally substituted with C.sub.1-C.sub.4 alkyl; wherein in formula 5, R.sub.N, R.sub.N′, and R.sub.O are independently hydrogen or C.sub.1-C.sub.4 alkyl; wherein in formula 6, R.sub.P is a substituted silyl group; and R.sub.Q is a substituted silyl group or C.sub.1-C.sub.4 alkyl and R.sub.R is hydrogen, a substituted silyl group, or C1-C4 alkyl, or R.sub.Q and R.sub.R, together with the nitrogen atom to which they are attached, form a 5- to 8-membered heterocyclic ring optionally comprising one or more additional heteroatoms; wherein in formula 7, X is a single bond or a carbon atom, wherein when X is a single bond, R.sub.S is absent, R.sub.UA and R.sub.R, together with the carbon atom and nitrogen atom to which they are attached, form an optionally substituted 6-membered aromatic heterocyclic ring, and R.sub.UB and R.sub.T, together with the carbon atom and nitrogen atom to which they are attached, form an optionally substituted 6-membered aromatic heterocyclic ring, and when X is a carbon atom, R.sub.UA and R.sub.UB are independently C.sub.1-C.sub.4 alkyl and R.sub.R, R.sub.S, and R.sub.T, together with the carbon atoms to which they are attached, form the structure below: ##STR00187## wherein in formula 8, R.sub.V is C.sub.1-C.sub.4 alkyl or C.sub.3-C.sub.6 cycloalkyl; wherein in formula 9, R.sub.W and R.sub.X are independently C.sub.1-C.sub.4 alkyl or a substituted silyl group; and wherein in formula 10, R.sub.Y and R.sub.Z are independently C.sub.1-C.sub.4 alkyl or a substituted silyl group.

12. The method of claim 11, wherein the electrophilic species scavenger is selected from the group consisting of N,O-bis(trimethylsilyl)acetamide, N,O-bis(trimethylsilyl)trifluoroacetamide, N-methyl-N-trimethylsilylacetamide, N-methyl-N-trimethylsilyltrifluoroacetamide, dimethylketene methyl trimethylsilyl acetal, isopropenyloxytrimethylsilane, 2,2,4,4-tetramethylpentanone imine, 1,1,1,3,3,3-hexamethyldisilazane (HMDS), N-trimethylsilylmorpholine, N-trimethylsilyldiethylamine, and N-tert-butyltrimethylsilylamine.

13. The method of any one of claims 1 to 9 and 11 to 12, wherein per one equivalent of the protecting group to be removed or one equivalent of the resin to be removed, 1 to 5 equivalents of the silyl compound and 1 to 10 equivalents of the electrophilic species scavenger are mixed.

14. The method of any one of claims 1 to 12, wherein per one equivalent of the protecting group to be removed or one equivalent of the resin to be removed, 0.1 to 0.5 equivalent of the silyl compound or acid is mixed, wherein the electrophilic species scavenger is selected from the group consisting of N,O-bis(trimethylsilyl)acetamide, N,O-bis(trimethylsilyl)trifluoroacetamide, N-methyl-N-trimethylsilylacetamide, N-methyl-N-trimethylsilyltrifluoroacetamide, dimethylketene methyl trimethylsilyl acetal, and isopropenyloxytrimethylsilane, wherein the silyl compound is selected from the group consisting of TMSOTf, TESOTf, TBSOTf, TIPSOTf, TBDPSOTf, TTMSOTf, TMSCl, TMSBr, and TMSOClO.sub.3, and wherein the acid is represented by HX, wherein X is selected from the group consisting of —OTf, —OClO.sub.3, Cl, Br, and I.

15. The method of any one of claims 1 to 14, wherein the starting peptide compound comprises 1 to 30 amino acid residues and is linear or cyclic.

16. The method of any one of claims 1 to 4 and 6 to 15, wherein the resin for solid-phase synthesis is CTC resin, Wang resin, or SASRIN resin.

17. The method of any one of claims 1 to 16, wherein the method comprises mixing the starting peptide compound with the solvent, then with the electrophilic species scavenger, and subsequently with the silyl compound or acid.

18. An amide compound represented by general formula (A) below or a salt thereof: ##STR00188## wherein R.sub.1′ is selected from the group consisting of hydrogen, Fmoc, Boc, Alloc, Cbz, Teoc, and trifluoroacetyl; R.sub.17A and R.sub.17B are both methyl, or R.sub.17A and R.sub.17B, together with the nitrogen atom to which they are attached, form piperidine or morpholine; and R.sub.18 is hydrogen or PG.sub.10, wherein PG.sub.10 is selected from the group consisting of t-Bu, trityl, cumyl, benzyl, methyl, ethyl, allyl, optionally substituted aryl, optionally substituted aryl-C.sub.1-C.sub.4 alkyl, optionally substituted heteroaryl-C.sub.1-C.sub.4 alkyl, and 2-(trimethylsilyl)ethyl.

19. The amide compound or salt thereof of claim 18, wherein the amide compound is selected from the group consisting of: (3-1) 3-((((9H-fluoren-9-yl)methoxy)carbonyl)(methyl)amino)-4-(dimethylamino)-4-oxobutanoic acid, (3-2) allyl 3-((((9H-fluoren-9-yl)methoxy)carbonyl)(methyl)amino)-4-(dimethylamino)-4-oxobutanoate, (3-3) tert-butyl 3-((((9H-fluoren-9-yl)methoxy)carbonyl)(methyl)amino)-4-(dimethylamino)-4-oxobutanoate, (3-4) benzyl 3-((((9H-fluoren-9-yl)methoxy)carbonyl)(methyl)amino)-4-(dimethylamino)-4-oxobutanoate, (3-5) 3-((tert-butoxycarbonyl)(methyl)amino)-4-(dimethylamino)-4-oxobutanoic acid, (3-6) allyl 3-((tert-butoxycarbonyl)(methyl)amino)-4-(dimethylamino)-4-oxobutanoate, (3-7) tert-butyl 3-((tert-butoxycarbonyl)(methyl)amino)-4-(dimethylamino)-4-oxobutanoate, (3-8) benzyl 3-((tert-butoxycarbonyl)(methyl)amino)-4-(dimethylamino)-4-oxobutanoate, (3-9) 3-(((benzyloxy)carbonyl)(methyl)amino)-4-(dimethylamino)-4-oxobutanoic acid, (3-10) allyl 3-(((benzyloxy)carbonyl)(methyl)amino)-4-(dimethylamino)-4-oxobutanoate, (3-11) tert-butyl 3-(((benzyloxy)carbonyl)(methyl)amino)-4-(dimethylamino)-4-oxobutanoate, (3-12) benzyl 3-(((benzyloxy)carbonyl)(methyl)amino)-4-(dimethylamino)-4-oxobutanoate, (3-13) 3-(((allyloxy)carbonyl)(methyl)amino)-4-(dimethylamino)-4-oxobutanoic acid, (3-14) allyl 3-(((allyloxy)carbonyl)(methyl)amino)-4-(dimethylamino)-4-oxobutanoate, (3-15) tert-butyl 3-(((allyloxy)carbonyl)(methyl)amino)-4-(dimethylamino)-4-oxobutanoate, (3-16) benzyl 3-(((allyloxy)carbonyl)(methyl)amino)-4-(dimethylamino)-4-oxobutanoate, (3-17) 4-(dimethylamino)-3-(methyl((2-(trimethylsilyl)ethoxy)carbonyl)amino)-4-oxobutanoic acid, (3-18) allyl 4-(dimethylamino)-3-(methyl((2-(trimethylsilyl)ethoxy)carbonyl)amino)-4-oxobutanoate, (3-19) tert-butyl 4-(dimethylamino)-3-(methyl((2-(trimethylsilyl)ethoxy)carbonyl)amino)-4-oxobutanoate, (3-20) benzyl 4-(dimethylamino)-3-(methyl((2-(trimethylsilyl)ethoxy)carbonyl)amino)-4-oxobutanoate, (2-1) 3-((((9H-fluoren-9-yl)methoxy)carbonyl)(methyl)amino)-4-morpholino-4-oxobutanoic acid, (2-2) allyl 3-((((9H-fluoren-9-yl)methoxy)carbonyl)(methyl)amino)-4-morpholino-4-oxobutanoate, (2-3) tert-butyl 3-((((9H-fluoren-9-yl)methoxy)carbonyl)(methyl)amino)-4-morpholino-4-oxobutanoate, (2-4) benzyl 3-((((9H-fluoren-9-yl)methoxy)carbonyl)(methyl)amino)-4-morpholino-4-oxobutanoate, (2-5) 3-((tert-butoxycarbonyl)(methyl)amino)-4-morpholino-4-oxobutanoic acid, (2-6) allyl 3-((tert-butoxycarbonyl)(methyl)amino)-4-morpholino-4-oxobutanoate, (2-7) tert-butyl 3-((tert-butoxycarbonyl)(methyl)amino)-4-morpholino-4-oxobutanoate, (2-8) benzyl 3-((tert-butoxycarbonyl)(methyl)amino)-4-morpholino-4-oxobutanoate, (2-9) 3-(((benzyloxy)carbonyl)(methyl)amino)-4-morpholino-4-oxobutanoic acid, (2-10) allyl 3-(((benzyloxy)carbonyl)(methyl)amino)-4-morpholino-4-oxobutanoate, (2-11) tert-butyl 3-(((benzyloxy)carbonyl)(methyl)amino)-4-morpholino-4-oxobutanoate, (2-12) benzyl 3-(((benzyloxy)carbonyl)(methyl)amino)-4-morpholino-4-oxobutanoate, (2-13) 3-(((allyloxy)carbonyl)(methyl)amino)-4-morpholino-4-oxobutanoic acid, (2-14) allyl 3-(((allyloxy)carbonyl)(methyl)amino)-4-morpholino-4-oxobutanoate, (2-15) tert-butyl 3-(((allyloxy)carbonyl)(methyl)amino)-4-morpholino-4-oxobutanoate, (2-16) benzyl 3-(((allyloxy)carbonyl)(methyl)amino)-4-morpholino-4-oxobutanoate, (2-17) 3-(methyl((2-(trimethylsilyl)ethoxy)carbonyl)amino)-4-morpholino-4-oxobutanoic acid, (2-18) allyl 3-(methyl((2-(trimethylsilyl)ethoxy)carbonyl)amino)-4-morpholino-4-oxobutanoate, (2-19) tert-butyl 3-(methyl((2-(trimethylsilyl)ethoxy)carbonyl)amino)-4-morpholino-4-oxobutanoate, (2-20) benzyl 3-(methyl((2-(trimethylsilyl)ethoxy)carbonyl)amino)-4-morpholino-4-oxobutanoate, (4-1) 3-(methylamino)-4-oxo-4-(piperidin-1-yl)butanoic acid, (4-2) allyl 3-(methylamino)-4-oxo-4-(piperidin-1-yl)butanoate, (4-3) tert-butyl 3-(methylamino)-4-oxo-4-(piperidin-1-yl)butanoate, (4-4) benzyl 3-(methylamino)-4-oxo-4-(piperidin-1-yl)butanoate, (4-5) 3-(methylamino)-4-morpholino-4-oxobutanoic acid, (4-6) allyl 3-(methylamino)-4-morpholino-4-oxobutanoate, (4-7) tert-butyl 3-(methylamino)-4-morpholino-4-oxobutanoate, (4-8) benzyl 3-(methylamino)-4-morpholino-4-oxobutanoate, (4-9) 4-(dimethylamino)-3-(methylamino)-4-oxobutanoic acid, (4-10) allyl 4-(dimethylamino)-3-(methylamino)-4-oxobutanoate, (4-11) tert-butyl 4-(dimethylamino)-3-(methylamino)-4-oxobutanoate, (4-12) benzyl 4-(dimethylamino)-3-(methylamino)-4-oxobutanoate, (1-1) 3-((((9H-fluoren-9-yl)methoxy)carbonyl)(methyl)amino)-4-oxo-4-(piperidin-1-yl)butanoic acid, (1-2) allyl 3-((((9H-fluoren-9-yl)methoxy)carbonyl)(methyl)amino)-4-oxo-4-(piperidin-1-yl)butanoate, (1-3) tert-butyl 3-((((9H-fluoren-9-yl)methoxy)carbonyl)(methyl)amino)-4-oxo-4-(piperidin-1-yl)butanoate, (1-4) benzyl 3-((((9H-fluoren-9-yl)methoxy)carbonyl)(methyl)amino)-4-oxo-4-(piperidin-1-yl)butanoate, (1-5) 3-((tert-butoxycarbonyl)(methyl)amino)-4-oxo-4-(piperidin-1-yl)butanoic acid, (1-6) allyl 3-((tert-butoxycarbonyl)(methyl)amino)-4-oxo-4-(piperidin-1-yl)butanoate, (1-7) tert-butyl 3-((tert-butoxycarbonyl)(methyl)amino)-4-oxo-4-(piperidin-1-yl)butanoate, (1-8) benzyl 3-((tert-butoxycarbonyl)(methyl)amino)-4-oxo-4-(piperidin-1-yl)butanoate, (1-9) 3-(((benzyloxy)carbonyl)(methyl)amino)-4-oxo-4-(piperidin-1-yl)butanoic acid, (1-10) allyl 3-(((benzyloxy)carbonyl)(methyl)amino)-4-oxo-4-(piperidin-1-yl)butanoate, (1-11) tert-butyl 3-(((benzyloxy)carbonyl)(methyl)amino)-4-oxo-4-(piperidin-1-yl)butanoate, (1-12) benzyl 3-(((benzyloxy)carbonyl)(methyl)amino)-4-oxo-4-(piperidin-1-yl)butanoate, (1-13) 3-(((allyloxy)carbonyl)(methyl)amino)-4-oxo-4-(piperidin-1-yl)butanoic acid, (1-14) allyl 3-(((allyloxy)carbonyl)(methyl)amino)-4-oxo-4-(piperidin-1-yl)butanoate, (1-15) tert-butyl 3-(((allyloxy)carbonyl)(methyl)amino)-4-oxo-4-(piperidin-1-yl)butanoate, (1-16) benzyl 3-(((allyloxy)carbonyl)(methyl)amino)-4-oxo-4-(piperidin-1-yl)butanoate, (1-17) 3-(methyl((2-(trimethylsilyl)ethoxy)carbonyl)amino)-4-oxo-4-(piperidin-1-yl)butanoic acid, (1-18) allyl 3-(methyl((2-(trimethylsilyl)ethoxy)carbonyl)amino)-4-oxo-4-(piperidin-1-yl)butanoate, (1-19) tert-butyl 3-(methyl((2-(trimethylsilyl)ethoxy)carbonyl)amino)-4-oxo-4-(piperidin-1-yl)butanoate, and (1-20) benzyl 3-(methyl((2-(trimethylsilyl)ethoxy)carbonyl)amino)-4-oxo-4-(piperidin-1-yl)butanoate.

Description

DESCRIPTION OF EMBODIMENTS

[0408] The abbreviations used in the present invention are listed below. [0409] AcOEt: Ethyl acetate [0410] Alloc: Allyloxycarbonyl [0411] Allyl: Allyl [0412] BEP: 2-Bromo-1-ethylpyridinium tetrafluoroborate [0413] Bn: Benzyl [0414] Boc: tert-Butoxycarbonyl [0415] BSA: N,O-Bis(trimethylsilyl)acetamide [0416] BSTFA: N,O-Bis(trimethylsilyl)trifluoroacetamide [0417] Bu: Butyl [0418] Cbz: Benzyloxycarbonyl [0419] cHx: Cyclohexyl [0420] Cl-CTC resin: 2-Chlorotrityl chloride polymer resin [0421] CPME: Cyclopentyl methyl ether [0422] CTC: 2-Chlorotrityl chloride [0423] DBU: 2,3,4,6,7,8,9,10-Octahydropyrimido[1,2-a]azepine [0424] DdZ: 3,5-Dimethoxyphenylisopropoxycarbonyl [0425] DIC: N,N′-Diisopropylcarbodiimide [0426] DIPEA: N,N-Diisopropylethylamine [0427] DMA: N,N-Dimethylacetamide [0428] DME: 1,2-Dimethoxyethane [0429] DMF: N,N-Dimethylformamide [0430] DMSO: Dimethyl sulfoxide [0431] DMT-MM: 4-(4,6-Dimethoxy-1,3,5-triazin-2-yl)-4-methylmorpholinium chloride [0432] EDCI: 1-(3-Dimethylaminopropyl)-3-ethylcarbodiimide [0433] eq.: Equivalent [0434] EtOH: Ethanol [0435] Et: Ethyl [0436] Fmoc: 9-Fluorenylmethyloxycarbonyl [0437] HATU: 1-[Bis(dimethylamino)methylene]-1H-1,2,3-triazolo[4,5-b]pyridinium 3-oxide hexafluorophosphate [0438] HMDS: Hexamethyldisilazane [0439] HOAt: 3H-1,2,3-Triazolo[4,5-b]pyridin-3-ol [0440] HOBt: 1,2,3-Benzotriazol-1-ol [0441] HPLC: High performance liquid chromatography [0442] i-PrOAc: Isopropyl acetate [0443] i-Pr: Isopropyl [0444] LCMS: Liquid chromatography mass spectrometry [0445] MeCN: Acetonitrile [0446] Me: Methyl [0447] MS: Mass spectroscopy [0448] MsOH: Methanesulfonic acid [0449] MSTFA: N-Methyl-N-trimethylsilyltrifluoroacetamide [0450] MTBE: Methyl tert-butyl ether [0451] ND: Not determined [0452] NMI: 1-Methylimidazole [0453] NMP: N-Methylpyrrolidone [0454] oxyma: Ethyl cyano(hydroxyimino)acetate [0455] Pd/C: Palladium on carbon [0456] Ph: Phenyl [0457] pip: Piperidinyl [0458] prep.: Preparation [0459] T3P: Propylphosphonic anhydride [0460] TBDPS: tert-Butyldiphenylsilyl [0461] TBS: tert-Butyldimethylsilyl [0462] t-Bu: tert-Butyl [0463] TEA: Triethylamine [0464] Teoc: 2-(Trimethylsilyl)ethoxycarbonyl [0465] TES: Triethylsilyl [0466] TFA: Trifluoroacetic acid [0467] TFE: 2,2,2-Trifluoroethanol [0468] TfOH: Trifluoromethanesulfonic acid [0469] Tf: Trifluoromethanesulfonyl [0470] THF: Tetrahydrofuran [0471] 2-MeTHF: 2-Methyltetrahydrofuran [0472] TIPS: Triisopropylsilyl [0473] TMSOTf: Trimethylsilyl trifluoromethanesulfonate [0474] TMS: Trimethylsilyl [0475] Tr: Trityl [0476] TTMS: Tris(trimethylsilyl)silyl [0477] vol.: volume [0478] Gly: Glycine [0479] Ala: Alanine [0480] Ser: Serine [0481] Thr: Threonine [0482] Val: Valine [0483] Leu: Leucine [0484] Ile: Isoleucine [0485] Phe: Phenylalanine [0486] Tyr: Tyrosine [0487] Trp: Tryptophan [0488] His: Histidine [0489] Glu: Glutamic acid [0490] Asp: Aspartic acid [0491] Gln: Glutamine [0492] Asn: Asparagine [0493] Cys: Cysteine [0494] Met: Methionine [0495] Lys: Lysine [0496] Arg: Arginine [0497] Pro: Proline [0498] MeGly: N-Me Glycine [0499] MeAla: N-Me Alanine [0500] MeSer: N-Me Serine [0501] MeThr: N-Me Threonine [0502] MeVal: N-Me Valine [0503] MeLeu: N-Me Leucine [0504] MeIle: N-Me Isoleucine [0505] MePhe: N-Me Phenylalanine [0506] MeTyr: N-Me Tyrosine [0507] MeTrp: N-Me Tryptophan [0508] MeHis: N-Me Histidine [0509] MeGlu: N-Me Glutamic acid [0510] MeAsp: N-Me Aspartic acid [0511] MeGln: N-Me Glutamine [0512] MeAsn: N-Me Asparagine [0513] MeCys: N-Me Cysteine [0514] MeMet: N-Me Methionine [0515] MeLys: N-Me Lysine [0516] MeArg: N-Me Arginine

Definitions of Functional Groups

[0517] The term “alkyl” as used herein refers to a monovalent group derived by removing any one hydrogen atom from an aliphatic hydrocarbon, and covers a subset of hydrocarbyl or hydrocarbon group structures that contain hydrogen and carbon atoms, but do not contain a heteroatom (which refers to an atom other than carbon and hydrogen atoms) or an unsaturated carbon-carbon bond in the skeleton. The “alkyl” includes linear or branched alkyl. The alkyl is an alkyl having 1 to 20 carbon atoms (C.sub.1-C.sub.20; hereinafter, “C.sub.p-C.sub.q” means that it has p to q carbon atoms), examples of which include C.sub.1-C.sub.6 alkyl and C.sub.1-C.sub.4 alkyl. Specific examples of the alkyl include methyl, ethyl, propyl, butyl, pentyl, hexyl, isopropyl, tert-butyl, and sec-butyl.

[0518] The term “cycloalkyl” as used herein refers to a saturated or partially saturated cyclic monovalent aliphatic hydrocarbon group, including single rings, bicyclo rings, and spiro rings. The cycloalkyl may be partially unsaturated. Preferred examples of the cycloalkyl include C.sub.3-C.sub.6 cycloalkyl, which include, for example, cyclopropyl, cyclobutyl, cyclopentyl, and cyclohexyl.

[0519] As used herein, the term “cycloalkylalkyl” refers to a group in which any hydrogen atom in the above-defined “alkyl” is replaced by the above-defined “cycloalkyl.” Preferred examples of the cycloalkylalkyl include C.sub.3-6 cycloalkyl-C.sub.1-6 alkyl and C.sub.3-6 cycloalkyl-C.sub.1-4 alkyl. Specific examples include cyclopropylmethyl, cyclopropylethyl, cyclobutylmethyl, cyclopentylmethyl, and cyclohexylmethyl.

[0520] As used herein, the term “alkoxy” refers to an oxy group to which the above-defined “alkyl” is bonded. Preferred examples include C.sub.1-4 alkoxy and C.sub.1-3 alkoxy. Specific examples of the alkoxy include methoxy, ethoxy, 1-propoxy, 2-propoxy, n-butoxy, i-butoxy, sec-butoxy, and tert-butoxy.

[0521] As used herein, the term “alicyclic ring” refers to a monovalent non-aromatic hydrocarbon ring. The alicyclic ring may have an unsaturated bond in the ring, and may be a polycyclic ring having two or more rings. Carbon atoms constituting the ring may be oxidized to form carbonyl. The number of atoms forming the alicyclic ring is preferably 3 to 7 (3- to 7-membered alicyclic ring). Specific examples of the alicyclic ring include a cycloalkyl ring, a cycloalkenyl ring, and a cycloalkynyl ring.

[0522] As used herein, the term “heterocyclic ring” refers to a non-aromatic monovalent or divalent heterocyclic ring comprising preferably 1 to 5 heteroatoms in the ring-forming atoms. The heterocyclic ring may have a double bond and/or a triple bond in the ring, may have a carbon atom in the ring that is oxidized to form carbonyl, and may be a single ring, a fused ring, or a spiro ring. The number of the ring-forming atoms is preferably 3 to 12 (3- to 12-membered heterocyclic ring), more preferably 4 to 7 (4- to 7-membered heterocyclic ring), and still more preferably 5 to 6 (5- to 6-membered heterocyclic ring).

[0523] Specific examples of the heterocyclic ring include azetidine, piperazine, pyrrolidine, piperidine, morpholine, homomorpholine, (R)-hexahydropyrrolo[1,2-a]pyrazine, (S)-hexahydropyrrolo[1,2-a]pyrazine, 3-oxopiperazine, 2-oxopyrrolidine, azetidine, 2-oxoimidazolidine, oxetane, dihydrofuran, tetrahydrofuran, dihydropyran, tetrahydropyran, tetrahydropyridine, thiomorpholine, pyrazolidine, imidazoline, oxazolidine, isoxazolidine, thiazolidine, imidazolidine, isothiazolidine, thiadiazolidine, oxazolidone, benzodioxane, benzoxazoline, dioxolane, dioxane, and tetrahydrothiopyran.

[0524] As used herein, the term “aromatic heterocyclic ring” refers to an aromatic monovalent or divalent heterocyclic ring comprising preferably 1 to 5 heteroatoms in the ring-forming atoms. The aromatic heterocyclic ring may be partially saturated, and may be a single ring, a fused ring (such as a bicyclic aromatic heterocyclic ring in which a monocyclic aromatic heterocyclic ring is fused with a benzene ring or a monocyclic aromatic heterocyclic ring), or a spiro ring. The number of the ring-forming atoms is preferably 4 to 10 (4- to 10-membered aromatic heterocyclic ring).

[0525] Specific examples of the aromatic heterocyclic ring include furan, thiophene, pyrrole, imidazole, pyrazole, thiazole, isothiazole, oxazole, isoxazole, oxadiazole, thiadiazole, triazole, tetrazole, pyridine, pyrimidine, pyridazine, pyrazine, triazine, benzofuran, benzothiophene, benzothiadiazole, benzothiazole, benzoxazole, benzoxadiazole, benzimidazole, indole, isoindole, indazole, quinoline, isoquinoline, cinnoline, quinazoline, quinoxaline, indolizine, and imidazopyridine.

[0526] The term “aryl” as used herein refers to a monovalent aromatic hydrocarbon ring, preferred examples of which include C.sub.6-C.sub.10 aryl. Specific examples of the aryl include phenyl and naphthyl (e.g., 1-naphthyl or 2-naphthyl).

[0527] The term “heteroaryl” as used herein refers to a monovalent aromatic heterocyclic group comprising preferably 1 to 5 heteroatoms in the ring-forming atoms“ ”. The heteroaryl may be partially saturated, and may be a single ring or fused rings (such as bicyclic heteroaryl in which heteroaryl is fused with benzene ring or monocyclic heteroaryl ring). The number of the ring-forming atoms is preferably 5 to 10 (5- to 10-membered heteroaryl).

[0528] Specific examples of the heteroaryl include furyl, thienyl, pyrrolyl, imidazolyl, pyrazolyl, thiazolyl, isothiazolyl, oxazolyl, isoxazolyl, oxadiazolyl, thiadiazolyl, triazolyl, tetrazolyl, pyridyl, pyrimidyl, pyridazinyl, pyrazinyl, triazinyl, benzofuranyl, benzothienyl, benzothiadiazolyl, benzothiazolyl, benzoxazolyl, benzoxadiazolyl, benzimidazolyl, indolyl, isoindolyl, azaindolyl, indazolyl, quinolyl, isoquinolyl, cinnolinyl, quinazolinyl, quinoxalinyl, benzodioxolyl, indolizinyl, and imidazopyridyl.

[0529] As used herein, the term “heterocyclyl” refers to a non-aromatic monovalent heterocyclic group comprising preferably 1 to 5 heteroatoms in the ring-forming atoms. The heterocyclyl may have a double or triple bond in the ring, may have a carbon atom oxidized to form carbonyl, and may be a single ring or a fused ring. The number of the ring-forming atoms is preferably 3 to 10 (3- to 10-membered heterocyclyl).

[0530] Specific examples of the heterocyclyl include oxetanyl, dihydrofuryl, tetrahydrofuryl, dihydropyranyl, tetrahydropyranyl, tetrahydropyridyl, morpholinyl, thiomorpholinyl, pyrrolidinyl, piperidinyl, piperazinyl, pyrazolidinyl, imidazolinyl, imidazolidinyl, oxazolidinyl, isoxazolidinyl, thiazolidinyl, isothiazolidinyl, thiadiazolidinyl, azetidinyl, oxazolidone, benzodioxanyl, benzoxazolyl, dioxolanyl, and dioxanyl.

[0531] As used herein, the term “arylalkyl” refers to a group in which any hydrogen atom in the above-defined “alkyl” is replaced by the above-defined “aryl.” Preferred examples of the arylalkyl include C6-10 aryl-C1-4 alkyl and C6-10 aryl-C1-3 alkyl. Specific examples include benzyl, phenylmethyl, phenethyl (phenylethyl), and naphthylmethyl.

[0532] As used herein, the term “heteroarylalkyl” refers to a group in which any hydrogen atom in the above-defined “alkyl” is replaced by the above-defined “heteroaryl.” Preferred examples of the heteroarylalkyl include 5- to 10-membered heteroaryl-C1-3 alkyl. Specific examples include pyrrolylmethyl, imidazolylmethyl, thienylmethyl, pyridylmethyl, pyrimidylmethyl, quinolylmethyl, and pyridylethyl.

[0533] As used herein, the term “substituted silyl” refers to a silyl group substituted with 1 to 3 substituents. The substituents may be the same or different. Such substituents are preferably C1-C6 alkyl, aryl, and tri-C1-C6 alkylsilyl. Specific examples include trimethylsilyl, triethylsilyl, triisopropylsilyl, t-butyldimethylsilyl, t-butyldiphenylsilyl, and tris(trimethylsilyl)silyl.

[0534] When the modifier “optionally substituted” is provided herein, examples of such substituents include alkyl, alkoxy, alkenyl, alkenyloxy, alkynyl, alkynyloxy, cycloalkyl, aryl, heteroaryl, heterocyclyl, arylalkyl, heteroarylalkyl, substituted silyl, halogen atoms, nitro, amino, monoalkylamino, dialkylamino, cyano, carboxyl, alkoxycarbonyl, and formyl.

[0535] As used herein, the term “deprotection/removal of a protection group” refers to converting a protected functional group back to an original functional group by removing the protecting group.

[0536] As used herein, the term “resin removal/removal of a resin” refers to cleaving a peptide compound from a resin for solid-phase synthesis that is bound to the peptide compound. The resin for solid-phase synthesis is preferably bound to the C-terminal amino acid residue of the starting peptide compound.

[0537] In one embodiment, the present invention provides methods of producing peptide pharmaceuticals comprising amino acid analogs useful for peptide pharmaceuticals. In another embodiment, the present invention provides methods of producing peptides comprising high-quality amino acid analogs for supplying pharmaceutical drug substances. In still another embodiment, the present invention provides novel amide compounds useful for producing peptide compounds.

Deprotection/Resin Removal Methods

[0538] In some aspects, the present invention relates to a method of producing a peptide compound in which a protecting group removable by a silylating agent is removed, the method comprising the step of contacting a starting peptide compound comprising natural amino acid residues and/or amino acid analog residues with the silylating agent in a solvent and thereby removing the protecting group from the starting peptide compound.

[0539] In some aspects, the present invention relates to a method of producing a peptide compound in which a resin for solid-phase synthesis is removed, the method comprising the step of contacting a starting peptide compound comprising natural amino acid residues and/or amino acid analog residues with a silylating agent in a solvent and thereby removing the starting peptide compound from the resin for solid-phase synthesis.

[0540] As used herein, the term “starting peptide compound” refers to a “peptide compound” which is a starting material to be subjected to the deprotection reaction and/or resin removal reaction of the present invention. The starting peptide compound preferably comprises at least one N-substituted amino acid residue.

[0541] “Peptide compounds” in the present invention include linear or cyclic peptide compounds comprising natural amino acid residues and/or amino acid analog residues. Cyclic peptide compounds are synonymous with “peptide compounds having a cyclic moiety.”

[0542] As used in the present invention, “linear peptide compounds” are not particularly limited as long as they are peptide compounds that are formed by natural amino acids or amino acid analogs forming amide or ester bonds and that do not have a cyclic moiety. Such a linear peptide compound can be formed by a total of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 20, 25, or 30 natural amino acids or amino acid analogs. The number of amino acids constituting the linear peptide compound preferably ranges from 1 to 30, from 6 to 20, from 7 to 19, from 7 to 18, from 7 to 17, from 7 to 16, from 7 to 15, from 8 to 14, or from 9 to 13.

[0543] As used in the present invention, “peptide compounds having a cyclic moiety” are not particularly limited as long as they are peptide compounds that are formed by natural amino acids or amino acid analogs forming amide or ester bonds and that have a cyclic moiety. Such cyclic moieties are preferably formed through covalent bonds such as amide bonds, carbon-carbon bonds, S—S bonds, thioether bonds, and triazole bonds (WO2013/100132, WO2012/026566, WO2012/033154, WO2012/074130, WO2015/030014, Comb Chem High Throughput Screen. 2010; 13:75-87, Nature Chem. Bio. 2009, 5, 502, Nat Chem Biol. 2009, 5, 888-90, Bioconjugate Chem., 2007, 18, 469-476, ChemBioChem, 2009, 10, 787-798, Chemical Communications (Cambridge, United Kingdom) (2011), 47(36), 9946-9958). Compounds obtained by further chemically modifying such compounds are also included in the peptide compounds of the present invention. The peptide compounds of the present invention having a cyclic moiety may also have a linear moiety. The number of amide or ester bonds (the number or length of natural amino acids or amino acid analogs) is not particularly limited, but when the peptide compound has a linear moiety, the cyclic and linear moieties combined preferably have 30 residues or less. The total number of amino acids is more preferably 9 or more in order to achieve high metabolic stability. In addition to above, the cyclic moiety is preferably formed by 5 to 12, 6 to 12, or 7 to 12, still more preferably 7 to 11 or 8 to 11, and particularly preferably 9 to 11 (10 or 11), natural amino acids and amino acid analogs. The linear moiety preferably has 0 to 8, 0 to 7, 0 to 6, 0 to 5, or 0 to 4, and more preferably 0 to 3, amino acids and amino acid analogs. The total number of natural amino acids and amino acid analogs is preferably 1 to 30, 6 to 20, 7 to 19, 7 to 18, 7 to 17, 7 to 16, 7 to 15, 8 to 14, or 9 to 13.

[0544] Although there are no particular limitations on the types of natural amino acid residues and amino acid analog residues forming the cyclic moiety of the peptide compound of the present invention having a cyclic moiety, the cyclic moiety is preferably formed by natural amino acid residues and amino acid analog residues having a functional group with high metabolic stability. The method of cyclizing the peptide compound of the present invention having a cyclic moiety is not particularly limited as long as the method can form such a cyclic moiety. Examples of the cyclization method include amide bond formation from carboxylic acids and amines; and carbon-carbon bond formation reactions using transition metals as catalysts such as Suzuki reaction, Heck reaction, and Sonogashira reaction. Accordingly, the peptide compound of the present invention contains at least one set of functional groups allowing such bond formation reactions before cyclization. In particular, in terms of metabolic stability, it preferably contains functional groups that form an amide bond by bond formation reaction.

[0545] Preferably, the cyclic moiety formed does not include a bond that contains a heteroatom which may be readily oxidized and that hinders metabolic stability, for example. Examples of bonds produced by cyclization include amide bonds formed by active esters and amines and bonds produced by Heck reaction products from carbon-carbon double bonds and aryl halides.

[0546] The “peptide compound” of the present invention may be a linear or cyclic peptide comprising at least one N-substituted amino acid (preferably 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 20, 25, or 30, particularly preferably 5, 6, or 7, N-substituted amino acids) and at least one N-unsubstituted amino acid, under or independently of the above-described conditions of the total number of natural amino acids and amino acid analogs. The number of N-substituted amino acids is preferably within the range of 1 to 30, 6 to 20, 7 to 19, 7 to 18, 7 to 17, 7 to 16, 7 to 15, 8 to 14, or 9 to 13.

[0547] In some aspects, the starting peptide compound comprises 1 to 30 amino acid residues and is linear or cyclic. Amino acids contained in the starting peptide compound may be either “natural amino acids” or “amino acid analogs.” “Amino acids”, “natural amino acids”, and “amino acid analogs” may be referred to herein as “amino acid residues,” “natural amino acid residues,” and “amino acid analog residues,” respectively. The starting peptide compound may be formed by secondary amides, tertiary amides, or a mixture of secondary and tertiary amides.

[0548] The term “natural amino acid” refers to Gly, Ala, Ser, Thr, Val, Leu, Ile, Phe, Tyr, Trp, His, Glu, Asp, Gln, Asn, Cys, Met, Lys, Arg, or Pro.

[0549] “Amino acid analogs” are not particularly limited, and include β-amino acids, γ-amino acids, D-amino acids, N-substituted amino acids, α,α-disubstituted amino acids, hydroxycarboxylic acids, and unnatural amino acids (amino acids whose side chains are different from those of natural amino acids: for example, unnatural α-amino acids, β-amino acids, and γ-amino acids). An α-amino acid may be a D-amino acid, or an α,α-dialkylamino acid. In a similar manner to an α-amino acid, a β-amino acid and a γ-amino acid are also allowed to have any configuration. Examples of N-substituted amino acids include amino acids of which the amino groups are substituted with any substituents. Examples of such substituents include, but are not particularly limited to, an alkyl group, an aryl group, and an aralkyl group. N-substituted amino acids include N-alkylamino acids, N-arylamino acids, and N-methylamino acids. There is no particular limitation on the selection of amino acid side chain, but in addition to a hydrogen atom, it can be freely selected from, for example, an alkyl group, an alkenyl group, an alkynyl group, an aryl group, a heteroaryl group, an aralkyl group, and a cycloalkyl group. One or two non-adjacent methylene groups in such a group are optionally substituted with an oxygen atom, a carbonyl group (—CO—), or a sulfonyl group (—SO2-). Each group may have one, two or more substituent(s). For example, the substituents are freely selected from any functional groups including a halogen atom, an N atom, an O atom, an S atom, a B atom, an Si atom, or a P atom (i.e., an optionally substituted alkyl group, alkenyl group, alkynyl group, aryl group, heteroaryl group, aralkyl group, and cycloalkyl group).

[0550] “Natural amino acids” and “amino acid analogs” as used herein which constitute a peptide compound include all isotopes corresponding to each amino acid. The isotope of the “natural amino acid” or “amino acid analog” refers to one having at least one atom replaced with an atom of the same atomic number (number of protons) and different mass number (total number of protons and neutrons). Examples of isotopes contained in the “natural amino acid” or “amino acid analog” constituting the peptide compounds of the present invention include a hydrogen atom, a carbon atom, a nitrogen atom, an oxygen atom, a phosphorus atom, a sulfur atom, a fluorine atom, and a chlorine atom, which respectively include .sup.2H and .sup.3H; .sup.13C and .sup.14C; .sup.15N; .sup.17O and .sup.11O; .sup.32P; .sup.35S; .sup.18F; and .sup.36Cl.

[0551] Examples of substituents containing a halogen atom as used herein include a halogen-substituted alkyl, cycloalkyl, alkenyl, alkynyl, aryl, heteroaryl, or aralkyl. More specific examples include fluoroalkyl, difluoroalkyl, and trifluoroalkyl.

[0552] Substituents containing an O atom include groups such as hydroxy (—OH), oxy (—OR), carbonyl (—C═O—R), carboxy (—CO2H), oxycarbonyl (—C═O—OR), carbonyloxy (—O—C═O—R), thiocarbonyl (—C═O—SR), carbonylthio (—S—C═O—R), aminocarbonyl (—C═O—NHR), carbonylamino (—NH—C═O—R), oxycarbonylamino (—NH—C═O—OR), sulfonylamino (—NH—SO2-R), aminosulfonyl (—SO2-NHR), sulfamoylamino (—NH—SO2-NHR), thiocarboxyl (—C(═O)—SH), and carboxylcarbonyl (—C(═O)—CO2H).

[0553] Examples of oxy (—OR) include alkoxy, cycloalkoxy, alkenyloxy, alkynyloxy, aryloxy, heteroaryloxy, and aralkyloxy. The alkoxy is preferably C1-C4 alkoxy and C1-C2 alkoxy, and particularly preferably methoxy or ethoxy.

[0554] Examples of carbonyl (—C═O—R) include formyl (—C═O—H), alkylcarbonyl, cycloalkylcarbonyl, alkenylcarbonyl, alkynylcarbonyl, arylcarbonyl, heteroarylcarbonyl, and aralkylcarbonyl.

[0555] Examples of oxycarbonyl (—C═O—OR) include alkyloxycarbonyl, cycloalkyloxycarbonyl, alkenyloxycarbonyl, alkynyloxycarbonyl, aryloxycarbonyl, heteroaryloxycarbonyl, and aralkyloxycarbonyl.

[0556] Examples of carbonyloxy (—O—C═O—R) include alkylcarbonyloxy, cycloalkylcarbonyloxy, alkenylcarbonyloxy, alkynylcarbonyloxy, arylcarbonyloxy, heteroarylcarbonyloxy, and aralkylcarbonyloxy.

[0557] Examples of thiocarbonyl (—C═O—SR) include alkylthiocarbonyl, cycloalkylthiocarbonyl, alkenylthiocarbonyl, alkynylthiocarbonyl, arylthiocarbonyl, heteroarylthiocarbonyl, and aralkylthiocarbonyl.

[0558] Examples of carbonylthio (—S—C═O—R) include alkylcarbonylthio, cycloalkylcarbonylthio, alkenylcarbonylthio, alkynylcarbonylthio, arylcarbonylthio, heteroarylcarbonylthio, and aralkylcarbonylthio.

[0559] Examples of aminocarbonyl (—C═O—NHR) include alkylaminocarbonyl (examples of which include C1-C6 or C1-C4 alkylaminocarbonyl, in particular, ethylaminocarbonyl and methylaminocarbonyl), cycloalkylaminocarbonyl, alkenylaminocarbonyl, alkynylaminocarbonyl, arylaminocarbonyl, heteroarylaminocarbonyl, and aralkylaminocarbonyl. Additional examples include compounds in which the H atom bonded to the N atom in —C═O—NHR is further replaced with alkyl, cycloalkyl, alkenyl, alkynyl, aryl, heteroaryl, or aralkyl.

[0560] Examples of carbonylamino (—NH—C═O—R) include alkylcarbonylamino, cycloalkylcarbonylamino, alkenylcarbonylamino, alkynylcarbonylamino, arylcarbonylamino, heteroarylcarbonylamino, and aralkylcarbonylamino. Additional examples include compounds in which the H atom bonded to the N atom in —NH—C═O—R is further replaced with alkyl, cycloalkyl, alkenyl, alkynyl, aryl, heteroaryl, or aralkyl.

[0561] Examples of oxycarbonylamino (—NH—C═O—OR) include alkoxycarbonylamino, cycloalkoxycarbonylamino, alkenyloxycarbonylamino, alkynyloxycarbonylamino, aryloxycarbonylamino, heteroaryloxycarbonylamino, and aralkyloxycarbonylamino. Additional examples include compounds in which the H atom bonded to the N atom in —NH—C═O—OR is further replaced with alkyl, cycloalkyl, alkenyl, alkynyl, aryl, heteroaryl, or aralkyl.

[0562] Examples of sulfonylamino (—NH—SO2-R) include alkylsulfonylamino, cycloalkylsulfonylamino, alkenylsulfonylamino, alkynylsulfonylamino, arylsulfonylamino, heteroarylsulfonylamino, and aralkylsulfonylamino. Additional examples include compounds in which the H atom attached to the N atom in —NH—SO2-R is further replaced with alkyl, cycloalkyl, alkenyl, alkynyl, aryl, heteroaryl, or aralkyl.

[0563] Examples of aminosulfonyl (—SO2-NHR) include alkylaminosulfonyl, cycloalkylaminosulfonyl, alkenylaminosulfonyl, alkynylaminosulfonyl, arylaminosulfonyl, heteroarylaminosulfonyl, and aralkylaminosulfonyl. Additional examples include compounds in which the H atom attached to the N atom in —SO2-NHR is further replaced with alkyl, cycloalkyl, alkenyl, alkynyl, aryl, heteroaryl, or aralkyl.

[0564] Examples of sulfamoylamino (—NH—SO2-NHR) include alkylsulfamoylamino, cycloalkylsulfamoylamino, alkenylsulfamoylamino, alkynylsulfamoylamino, arylsulfamoylamino, heteroarylsulfamoylamino, and aralkylsulfamoylamino. The two H atoms bonded to the N atoms in —NH—SO2-NHR may be further replaced with substituents independently selected from the group consisting of alkyl, cycloalkyl, alkenyl, alkynyl, aryl, heteroaryl, and aralkyl, and these two substituents may form a ring.

[0565] Substituents containing an S atom include groups such as thiol (—SH), thio (—S—R), sulfinyl (—S═O—R), sulfonyl (—SO2-R), and sulfo (—SO3H).

[0566] Examples of thio (—S—R) include alkylthio, cycloalkylthio, alkenylthio, alkynylthio, arylthio, heteroarylthio, and aralkylthio.

[0567] Examples of sulfonyl (—SO2-R) include alkylsulfonyl, cycloalkylsulfonyl, alkenylsulfonyl, alkynylsulfonyl, arylsulfonyl, heteroarylsulfonyl, and aralkylsulfonyl.

[0568] Substituents containing an N atom include groups such as azido (—N3, also called “azido group”), cyano (—CN), primary amino (—NH2), secondary amino (—NH—R; also called monosubstituted amino), tertiary amino (—NR(R′); also called disubstituted amino), amidino (—C(═NH)—NH2), substituted amidino (—C(═NR)—NR′R″), guanidino (—NH—C(═NH)—NH2), substituted guanidino (—NR—C(═NR″′)—NR′R″), aminocarbonylamino (—NR—CO—NR′R″), pyridyl, piperidino, morpholino, and azetidinyl.

[0569] Examples of secondary amino (—NH—R; monosubstituted amino) include alkylamino, cycloalkylamino, alkenylamino, alkynylamino, arylamino, heteroarylamino, and aralkylamino.

[0570] Examples of tertiary amino (—NR(R′); disubstituted amino) include amino groups having any two substituents each independently selected from alkyl, cycloalkyl, alkenyl, alkynyl, aryl, heteroaryl, and aralkyl, such as alkyl(aralkyl)amino, where any two such substituents may form a ring. Specific examples include dialkylamino, in particular, C1-C6 dialkylamino, C1-C4 dialkylamino, dimethylamino, and diethylamino. The term “Cp-Cq dialkylamino group” as used herein refers to an amino group substituted with two Cp-Cq alkyl groups, where the two Cp-Cq alkyl groups may be the same or different.

[0571] Examples of substituted amidino (—C(═NR)—NR′R″) include groups in which three substituents R, R′, and R″ on the N atom are each independently selected from alkyl, cycloalkyl, alkenyl, alkynyl, aryl, heteroaryl, and aralkyl, such as alkyl(aralkyl)(aryl)amidino.

[0572] Examples of substituted guanidino (—NR—C(═NR″′)—NR′R″) include groups in which R, R′, R″, and R″′ are each independently selected from alkyl, cycloalkyl, alkenyl, alkynyl, aryl, heteroaryl, and aralkyl, or groups in which these substituents form a ring.

[0573] Examples of aminocarbonylamino (—NR—CO—NR′R″) include groups in which R, R′, and R″ are each independently selected from a hydrogen atom, alkyl, cycloalkyl, alkenyl, alkynyl, aryl, heteroaryl, and aralkyl, or groups in which these substituents form a ring.

[0574] In some aspects, the starting peptide compound to be subjected to deprotection and/or resin removal comprises at least one N-substituted amino acid residue. Examples of the number of N-substituted amino acid residues contained in the starting peptide compound include 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 20, 25, or 30. The number of N-substituted amino acid residues is preferably within the range of 6 to 20, 7 to 19, 7 to 18, 7 to 17, 7 to 16, 7 to 15, 8 to 14, or 9 to 13. Two or more N-substituted amino acid residues may be linked to each other in the starting peptide compound.

[0575] In some aspects, half or more of the amino acids constituting the peptide compound of the present invention (for example, n or more amino acids (where n is an integer) when the peptide compound is constituted by 2n amino acids, or n+1 or more amino acids when the peptide compound is constituted by 2n+1 amino acids) are preferably N-substituted amino acids.

[0576] As used herein, the term “N-substitution” in N-substituted amino acids refers to, but is not limited to, replacement of a hydrogen atom attached to an N atom with a methyl group, an ethyl group, a propyl group, a butyl group, or a hexyl group. Preferred N-substituted amino acids include amino acids in which amino groups contained in natural amino acids are N-methylated, N-ethylated, N-propylated, N-butylated, and N-pentylated. Such amino acids are called N-methylamino acid, N-ethylamino acid, N-propylamino acid, N-butylamino acid, and N-pentylamino acid.

[0577] When the starting peptide compound comprises at least one N-substituted amino acid residue (for example, when the starting peptide compound comprises one or more partial structures in which an N-substituted amino acid residue is linked to the adjacent amino acid residue, more specifically, one or more partial structures each comprising at least two amino acid residues, which structure is represented by general formulas (I) and/or (II)), conventional deprotection/resin removal methods easily cause main-chain damage such as amide bond cleavage or peptide main-chain rearrangement. Even in case of such starting peptide compounds, use of the methods of the present invention is able to remove protecting groups of interest efficiently in high yield and purity without involving main-chain damage, and to cleave peptide compounds from resins in solid-phase reactions efficiently in high yield and purity.

[0578] In some embodiments, the starting peptide compound of the present invention may not comprise N-substituted amino acid residues or a partial structure in which an N-substituted amino acid residue is linked to the adjacent amino acid residue.

[0579] In some aspects, the starting peptide compound to be subjected to deprotection and/or resin removal comprises at least one partial structure in which at least two amino acid residues are linked to each other, wherein the structure is represented by general formula (I) below.

##STR00017##

[0580] In formula (I), R.sub.1 is hydrogen, PG.sub.1, a natural amino acid residue, or an amino acid analog residue.

[0581] When the structure represented by formula (I) is at the N-terminus of the starting peptide compound, R.sub.1 is preferably hydrogen or PG.sub.1. On the other hand, when the structure represented by formula (I) is at a position other than the N-terminus of the starting peptide compound, R.sub.1 is preferably a natural amino acid residue or an amino acid analog residue.

[0582] In formula (I), R.sub.2 is selected from the group consisting of hydrogen and C.sub.1-C.sub.6 alkyl, or R.sub.2 and R.sub.4 or R.sub.2 and R.sub.4′, together with the nitrogen atom and carbon atom to which they are attached, form a 3- to 7-membered heterocyclic ring optionally substituted with hydroxy or C.sub.1-C.sub.4 alkoxy.

[0583] In some aspects, when R.sub.2 is C.sub.1-C.sub.6 alkyl, the C.sub.1-C.sub.6 alkyl is preferably methyl, ethyl, propyl, butyl, or pentyl.

[0584] In some aspects, when R.sub.2 and R.sub.4 or R.sub.2 and R.sub.4′, together with the nitrogen atom and carbon atom to which they are attached, form a 3- to 7-membered heterocyclic ring optionally substituted with hydroxy or C.sub.1-C.sub.4 alkoxy, the 3- to 7-membered heterocyclic ring formed is preferably an azetidine ring, a pyrrolidine ring, or a piperidine ring. R.sub.4′ is hydrogen when R.sub.2 and R.sub.4 together form the heterocyclic ring, and R.sub.4 is hydrogen when R.sub.2 and R.sub.4′ together form the heterocyclic ring.

[0585] In formula (I), except when R.sub.2 and R.sub.4 or R.sub.2 and R.sub.4, together form the heterocyclic ring, [0586] (a) R.sub.4′ is hydrogen, and R.sub.4 is selected from the group consisting of hydrogen, optionally substituted C.sub.1-C.sub.6 alkyl, C.sub.3-C.sub.6 cycloalkyl, C.sub.3-C.sub.6 cycloalkyl-C.sub.1-C.sub.4 alkyl, optionally substituted phenyl, optionally substituted phenylmethyl, optionally substituted phenylethyl, 2-(methylthio)ethyl, —CH.sub.2SPG.sub.2, N-PG.sub.3-indol-3-ylmethyl, 4-(PG.sub.2O)benzyl, PG.sub.2-O-methyl (i.e., —CH.sub.2—O-PG.sub.2), 1-(PG.sub.2O)ethyl, 2-(PG.sub.2O)ethyl, PG.sub.2-OCO(CH.sub.2)—, PG.sub.2-OCO(CH.sub.2).sub.2—, PG.sub.3N-n-butyl, —CON(R.sub.14A)(R.sub.14B), —CH.sub.2—CON(R.sub.14A)(R.sub.14B), and —(CH.sub.2).sub.2CON(R.sub.14A)(R.sub.14B), [0587] (b) R.sub.4 and R.sub.4′ are independently optionally substituted C.sub.1-C.sub.6 alkyl, or [0588] (c) R.sub.4 and R.sub.4′, together with the carbon atom to which they are attached, form a 3- to 7-membered alicyclic ring.

[0589] Combinations of R.sub.4 and R.sub.4′ are preferably a hydrogen atom and a hydrogen atom, methyl and a hydrogen atom, ethyl and a hydrogen atom, isopropyl and a hydrogen atom, isobutyl and a hydrogen atom, cyclopropyl and a hydrogen atom, cyclopropylmethyl and a hydrogen atom, cyclopentyl and a hydrogen atom, cyclohexyl and a hydrogen atom, optionally substituted phenyl and a hydrogen atom, optionally substituted phenylmethyl and a hydrogen atom, optionally substituted phenylethyl and a hydrogen atom, 2-(methylthio)ethyl and a hydrogen atom, —CH.sub.2SPG.sub.2 and a hydrogen atom, N-PG.sub.3-indol-3-ylmethyl and a hydrogen atom, 4-(PG.sub.2O)benzyl and a hydrogen atom, PG.sub.2-O-methyl and a hydrogen atom, 1-(PG.sub.2O)ethyl and a hydrogen atom, 2-(PG.sub.2O)ethyl and a hydrogen atom, PG.sub.2-OCO(CH.sub.2)— and a hydrogen atom, PG.sub.2-OCO(CH.sub.2).sub.2— and a hydrogen atom, PG.sub.3N-n-butyl and a hydrogen atom, —CON(R.sub.14A)(R.sub.14B) and a hydrogen atom, —CH.sub.2—CON(R.sub.14A)(R.sub.14B) and a hydrogen atom, —(CH.sub.2).sub.2CON(R.sub.14A)(R.sub.14B) and a hydrogen atom, methyl and methyl, and methyl and ethyl. When R.sub.4 and R.sub.4′, together with the carbon atom to which they are attached, form a 3- to 7-membered alicyclic ring, the alicyclic ring is preferably a cyclopropyl ring, a cyclobutyl ring, a cyclopentyl ring, or a cyclohexyl ring.

[0590] In formula (I), R.sub.5 is a single bond or —C(R.sub.5A)(R.sub.5B)—; and R.sub.5A and R.sub.5B are independently selected from the group consisting of hydrogen, C1-C.sub.6 alkyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted aryl-C1-C.sub.4 alkyl, and optionally substituted heteroaryl-C.sub.1-C.sub.4 alkyl.

[0591] R.sub.5 is preferably a single bond or —C(R.sub.5A)(R.sub.5B)— where the combinations of R.sub.5A and R.sub.5B is a hydrogen atom and a hydrogen atom, methyl and a hydrogen atom, ethyl and a hydrogen atom, isopropyl and a hydrogen atom, isobutyl and a hydrogen atom, cyclopropyl and a hydrogen atom, cyclopropylmethyl and a hydrogen atom, optionally substituted phenyl and a hydrogen atom, optionally substituted phenylmethyl and a hydrogen atom, optionally substituted phenylethyl and a hydrogen atom, methyl and methyl, or methyl and ethyl.

[0592] In formula (I), R.sub.6 is selected from the group consisting of hydrogen and C.sub.1-C.sub.6 alkyl, or R.sub.6 and R.sub.7 or R.sub.6 and R.sub.7′, together with the nitrogen atom and carbon atom to which they are attached, form a 3- to 7-membered heterocyclic ring optionally substituted with hydroxy or C.sub.1-C.sub.4 alkoxy.

[0593] In some aspects, when R.sub.6 is C.sub.1-C.sub.6 alkyl, the C.sub.1-C.sub.6 alkyl is preferably methyl, ethyl, propyl, butyl, or pentyl.

[0594] In some aspects, when R.sub.6 and R.sub.7 or R.sub.6 and R.sub.7′, together with the nitrogen atom and carbon atom to which they are attached, form a 3- to 7-membered heterocyclic ring optionally substituted with hydroxy or C1-C4 alkoxy, the 3- to 7-membered heterocyclic ring formed is preferably an azetidine ring, a pyrrolidine ring, or a piperidine ring. R7′ is hydrogen when R6 and R7 together form the heterocyclic ring, and R7 is hydrogen when R6 and R7′ together form the heterocyclic ring.

[0595] In formula (I), preferably, either or both of R2 and R6 are other than hydrogen, and more preferably, either or both of R2 and R6 are C1-C6 alkyl.

[0596] In formula (I), except when R.sub.6 and R.sub.7, or R.sub.6 and R.sub.7′ together form the heterocyclic ring, [0597] (a) R.sub.7′ is hydrogen, and R.sub.7 is selected from the group consisting of hydrogen, optionally substituted C.sub.1-C.sub.6 alkyl, C.sub.3-C.sub.6 cycloalkyl, C.sub.3-C.sub.6 cycloalkyl-C.sub.1-C.sub.4 alkyl, optionally substituted phenyl, optionally substituted phenylmethyl, optionally substituted phenylethyl, 2-(methylthio)ethyl, —CH.sub.2SPG.sub.4, N-PG.sub.5-indol-3-ylmethyl, 4-(PG.sub.4O)benzyl, PG.sub.4-O-methyl (i.e., —CH.sub.2—O-PG.sub.4), 1-(PG.sub.4O)ethyl, 2-(PG.sub.4O)ethyl, PG.sub.4-OCO(CH.sub.2)—, PG.sub.4-OCO(CH.sub.2).sub.2—, PG.sub.5N-n-butyl, —CON(R.sub.15A)(R.sub.15B), —CH.sub.2—CON(R.sub.15A)(R.sub.15), and —(CH.sub.2).sub.2CON(R.sub.15A)(R.sub.15B), [0598] (b) R.sub.7 and R.sub.7′ are independently optionally substituted C.sub.1-C.sub.6 alkyl, or [0599] (c) R.sub.7 and R.sub.7′, together with the carbon atom to which they are attached, form a 3- to 7-membered alicyclic ring.

[0600] Combinations of R.sub.7 and R.sub.7′ are preferably a hydrogen atom and a hydrogen atom, methyl and a hydrogen atom, ethyl and a hydrogen atom, isopropyl and a hydrogen atom, isobutyl and a hydrogen atom, cyclopropyl and a hydrogen atom, cyclopropylmethyl and a hydrogen atom, cyclopentyl and a hydrogen atom, cyclohexyl and a hydrogen atom, optionally substituted phenyl and a hydrogen atom, optionally substituted phenylmethyl and a hydrogen atom, optionally substituted phenylethyl and a hydrogen atom, 2-(methylthio)ethyl and a hydrogen atom, —CH.sub.2SPG.sub.4 and a hydrogen atom, N-PG.sub.5-indol-3-ylmethyl and a hydrogen atom, 4-(PG.sub.4O)benzyl and a hydrogen atom, PG.sub.4-O-methyl and a hydrogen atom, 1-(PG.sub.4O)ethyl and a hydrogen atom, 2-(PG.sub.4O)ethyl and a hydrogen atom, PG.sub.4-OCO(CH.sub.2)— and a hydrogen atom, PG.sub.4-OCO(CH.sub.2).sub.2— and a hydrogen atom, PG.sub.5N-n-butyl and a hydrogen atom, —CON(R.sub.15A)(R.sub.15B) and a hydrogen atom, —CH.sub.2—CON(R.sub.15A)(R.sub.15B) and a hydrogen atom, —(CH.sub.2).sub.2CON(R.sub.15A)(R.sub.15B) and a hydrogen atom, methyl and methyl, and methyl and ethyl. When R.sub.7 and R.sub.7′, together with the carbon atom to which they are attached, form a 3- to 7-membered alicyclic ring, the alicyclic ring is preferably a cyclopropyl ring, a cyclobutyl ring, a cyclopentyl ring, or a cyclohexyl ring.

[0601] In formula (I), R.sub.8 is a single bond or —C(R.sub.8A)(R.sub.8B)—; and R.sub.8A and R.sub.8B are independently selected from the group consisting of hydrogen, C.sub.1-C.sub.6 alkyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted aryl-C.sub.1-C.sub.4 alkyl, and optionally substituted heteroaryl-C.sub.1-C.sub.4 alkyl.

[0602] R.sub.8 is preferably a single bond or —C(R.sub.8A)(R.sub.8B)— where the combination of R.sub.8A and R.sub.8B is a hydrogen atom and a hydrogen atom, methyl and a hydrogen atom, ethyl and a hydrogen atom, isopropyl and a hydrogen atom, isobutyl and a hydrogen atom, cyclopropyl and a hydrogen atom, cyclopropylmethyl and a hydrogen atom, optionally substituted phenyl and a hydrogen atom, optionally substituted phenylmethyl and a hydrogen atom, optionally substituted phenylethyl and a hydrogen atom, methyl and methyl, or methyl and ethyl.

[0603] In formula (I), R.sub.9 is hydroxy, —O-PG.sub.6, a natural amino acid residue, an amino acid analog residue, —O-RES, or —NH-RES, where RES is a resin for solid-phase synthesis.

[0604] When the structure represented by formula (I) is at the C-terminus of the starting peptide compound, R.sub.9 is preferably hydroxy, —O-PG.sub.6, —O-RES, or —NH-RES. On the other hand, when the structure represented by formula (I) is at a position other than the C-terminus of the starting peptide compound, R.sub.9 is preferably a natural amino acid residue or an amino acid analog residue.

[0605] In formula (I), R.sub.14A and R.sub.14B are independently hydrogen or C.sub.1-C.sub.4 alkyl, or R.sub.14A and R.sub.14B, together with the nitrogen atom to which they are attached, form a 4- to 8-membered ring optionally comprising one or more additional heteroatoms. When R.sub.14A and/or R.sub.14B are C.sub.1-C.sub.4 alkyl, the C.sub.1-C.sub.4 alkyl is preferably methyl, ethyl, or propyl. When R.sub.14A and R.sub.14B, together with the nitrogen atom to which they are attached, form a 4- to 8-membered ring optionally comprising one or more additional heteroatoms, the 4- to 8-membered ring is preferably an azetidine ring, a pyrrolidine ring, a piperidine ring, a piperazine ring, or a morpholine ring.

[0606] In formula (I), R.sub.15A and R.sub.15B are independently hydrogen or C.sub.1-C.sub.4 alkyl, or R.sub.15A and R.sub.15B, together with the nitrogen atom to which they are attached, form a 4- to 8-membered ring optionally comprising one or more additional heteroatoms. When R.sub.15A and/or R.sub.15B are C.sub.1-C.sub.4 alkyl, the C.sub.1-C.sub.4 alkyl is preferably methyl, ethyl, or propyl. When R.sub.15A and R.sub.15B, together with the nitrogen atom to which they are attached, form a 4- to 8-membered ring optionally comprising one or more additional heteroatoms, then the 4- to 8-membered ring is preferably an azetidine ring, a pyrrolidine ring, a piperidine ring, a piperazine ring, or a morpholine ring.

[0607] In formula (I), PG.sub.1 is selected from the group consisting of Fmoc, Boc, Alloc, Cbz, Teoc, and trifluoroacetyl.

[0608] PG.sub.1 is preferably Fmoc, Boc, or Cbz.

[0609] In formula (I), PG.sub.2 and PG.sub.4 are independently selected from the group consisting of hydrogen, t-Bu, trityl, methoxytrityl, cumyl, benzyl, THP, 1-ethoxyethyl, methyl, ethyl, allyl, optionally substituted aryl, optionally substituted aryl-C.sub.1-C.sub.4 alkyl, optionally substituted heteroaryl-C.sub.1-C.sub.4 alkyl, and 2-(trimethylsilyl)ethyl.

PG.sub.2 and PG.sub.4 are preferably methyl, allyl, t-Bu, trityl, methoxytrityl, cumyl, THP, optionally substituted aryl, optionally substituted aryl-C.sub.1-C.sub.4 alkyl, or optionally substituted heteroaryl-C.sub.1-C.sub.4 alkyl.

[0610] In formula (I), PG.sub.3 and PG.sub.5 are independently selected from the group consisting of hydrogen, Fmoc, Boc, Alloc, Cbz, Teoc, methoxycarbonyl, t-Bu, trityl, cumyl, and benzyl.

[0611] PG.sub.3 and PG.sub.5 are preferably Fmoc, Boc, Cbz, t-Bu, or trityl.

[0612] In formula (I), PG.sub.6 is selected from the group consisting of t-Bu, trityl, cumyl, benzyl, methyl, ethyl, allyl, and 2-(trimethylsilyl)ethyl.

[0613] PG.sub.6 is preferably t-Bu, trityl, cumyl, benzyl, methyl, or allyl.

[0614] Structures represented by general formula (I), in which at least two amino acid residues are linked to each other, include many such structures easily cleaved or damaged when known deprotection or resin removal conditions are used. Only the protecting groups of interest or resins for solid-phase synthesis can be selectively and efficiently removed without such cleavage or damage by using the reaction conditions of the present invention.

[0615] In some aspects, structures represented by general formula (I), in which at least two amino acid residues are linked to each other, include those in which one N-substituted amino acid is linked to one N-unsubstituted amino acid and those in which two N-substituted amino acids are linked to each other. Specific examples of such amino acid residues include structures represented by general formula (I′), in which two amino acid residues are linked to each other, wherein R.sub.5 and R.sub.8 in general formula (I) are single bonds.

##STR00018##

[0616] Each group in formula (I′) may be the same as that in formula (I) above.

[0617] In some aspects, in formula (I′), R1 is preferably hydrogen, PG1, a natural amino acid residue, or an amino acid analog residue.

[0618] In some aspects, in formula (I′), preferably, either or both of R2 and R6 are other than hydrogen, and more preferably, either or both of R2 and R6 are C1-C6 alkyl.

[0619] In some aspects, in formula (I′), R2 and R6 may be independently C1-C6 alkyl, which is preferably methyl, ethyl, propyl, butyl, or pentyl.

[0620] In some aspects, in formula (I′), R2 and R4 or R2 and R4′, or R6 and R7 or R6 and R7′ may each independently, together with the nitrogen atom and carbon atom to which they are attached, form a 3- to 7-membered heterocyclic ring optionally substituted with hydroxy or C1-C4 alkoxy. R4′ is hydrogen when R2 and R4 together form the heterocyclic ring, and R4 is hydrogen when R2 and R4′ together form the heterocyclic ring. R7′ is hydrogen when R6 and R7 together form the heterocyclic ring, and R7 is hydrogen when R6 and R7′ together form the heterocyclic ring.

[0621] In some aspects, in formula (I′), R9 is hydroxy, —O-PG6, a natural amino acid residue, an amino acid analog residue, —O-RES, or —NH-RES, where RES is a resin for solid-phase synthesis.

[0622] In some aspects, in formula (I′), R4 and R4′, or R7 and R7′ may each be C1-C6 alkyl, or R4 and R4′, or R7 and R7′ may, together with the carbon atom to which they are attached, form a 3- to 7-membered alicyclic ring.

[0623] In some aspects, in formula (I′), R4′ may be hydrogen, and R4 may be selected from the group consisting of hydrogen, optionally substituted C1-C6 alkyl, C3-C6 cycloalkyl, C3-C6 cycloalkyl-C1-C4 alkyl, optionally substituted phenyl, optionally substituted phenylmethyl, optionally substituted phenylethyl, 2-(methylthio)ethyl, —CH2SPG2, N-PG3-indol-3-ylmethyl, 4-(PG2O)benzyl, PG2-O-methyl, 1-(PG2O)ethyl, 2-(PG2O)ethyl, PG2-OCO(CH2)-, PG2-OCO(CH2)2-, PG3N-n-butyl, —CON(R14A)(R14B), —CH2-CON(R14A)(R14B), and —(CH2)2CON(R14A)(R14B).

[0624] In some aspects, in formula (I′), R7′ may be hydrogen, and R7 may be selected from the group consisting of hydrogen, optionally substituted C1-C6 alkyl, C3-C6 cycloalkyl, C3-C6 cycloalkyl-C1-C4 alkyl, optionally substituted phenyl, optionally substituted phenylmethyl, optionally substituted phenylethyl, 2-(methylthio)ethyl, —CH2SPG4, N-PG5-indol-3-ylmethyl, 4-(PG4O)benzyl, PG4-O-methyl, 1-(PG4O)ethyl, 2-(PG4O)ethyl, PG4-OCO(CH2)-, PG4-OCO(CH2)2-, PG5N-n-butyl, —CON(R15A)(R15B), —CH2-CON(R15A)(R15B), and —(CH2)2CON(R15A)(R15B).

[0625] More preferably, in the structure represented by general formula (I′), R.sub.1 is hydrogen, PG.sub.1, a natural amino acid residue, or an amino acid analog residue; [0626] R.sub.2 and R.sub.6 are methyl or ethyl; and/or [0627] R.sub.2 and R.sub.4 or R.sub.2 and R.sub.4′, or R.sub.6 and R.sub.7 or R.sub.6 and R.sub.7′ each independently, together with the nitrogen atom and carbon atom to which they are attached, form a 4- to 6-membered heterocyclic ring optionally substituted with hydroxy or C.sub.1-C.sub.4 alkoxy. R.sub.4′ is hydrogen when R.sub.2 and R.sub.4 together form the heterocyclic ring, and R.sub.4 is hydrogen when R.sub.2 and R.sub.4′ together form the heterocyclic ring. R.sub.7′ is hydrogen when R.sub.6 and R.sub.7 together form the heterocyclic ring, and R.sub.7 is hydrogen when R.sub.6 and R.sub.7′ together form the heterocyclic ring. [0628] R.sub.9 may be —O-PG.sub.6, a natural amino acid residue, an amino acid analog residue, or —O-RES, where RES is a solid-phase synthesis CTC, Wang, or SASRIN resin. [0629] R.sub.4 and R.sub.4′, or R.sub.7 and R.sub.7′ may be each independently methyl or ethyl, and R.sub.4 and R.sub.4′ or R.sub.7 and R.sub.7′ may, together with the nitrogen atom and carbon atom to which they are attached, form a 5- or 6-membered alicyclic ring. [0630] When R.sub.4, and R.sub.7′ are hydrogen, R.sub.4 and R.sub.7 may be each independently hydrogen, or alkyl selected from the group consisting of methyl, ethyl, isopropyl, isobutyl, and sec-butyl, or optionally substituted phenylmethyl, optionally substituted phenylethyl, benzyloxymethyl, 1-benzyloxyethyl, 2-benzyloxyethyl, tert-butoxycarbonylmethyl, methoxycarbonylmethyl, tert-butoxycarbonylethyl, methoxycarbonylethyl, tert-butoxycarbamoylbutyl, N,N-dimethylaminocarbonyl, piperidylcarbonyl, pyrrolidylcarbonyl, N,N-dimethylaminocarbonylmethyl, piperidylcarbonylmethyl, pyrrolidylcarbonylmethyl, N,N-dimethylaminocarbonylethyl, piperidylcarbonylethyl, or pyrrolidylcarbonylethyl.

[0631] In some aspects, the starting peptide compound may comprise one or more additional natural amino acid residues and/or amino acid analog residues, in addition to one or more structures represented by general formula (I).

[0632] In some aspects, the starting peptide compound comprises at least one protecting group removable by the method of the present invention. Such a protecting group may be contained in a structure represented by general formula (I), or may be contained in an amino acid residue other than the structure represented by general formula (I).

[0633] In some aspects, the starting peptide compound comprises at least one resin for solid-phase synthesis that is removable by the method of the present invention. Such a resin may be contained in a structure represented by general formula (I), or may be contained in an amino acid residue other than the structure represented by general formula (I).

[0634] In some aspects, the resin for solid-phase synthesis is bound to the carboxyl group contained in the C-terminal amino acid residue of the starting peptide compound.

[0635] In some aspects, the starting peptide compound to be subjected to deprotection and/or resin removal comprises at the C-terminus a structure in which at least two amino acid residues are linked to each other, wherein the structure is represented by general formula (II) below.

##STR00019##

[0636] R.sub.1′ is a group represented by formula (III):

##STR00020## [0637] wherein * represents the point of attachment.

[0638] In formula (III), R1 is hydrogen, PG1, a natural amino acid residue, or an amino acid analog residue.

[0639] In formula (III), R2 is selected from the group consisting of hydrogen and C1-C6 alkyl, or R2 and R10 or R2 and R10′, together with the nitrogen atom and carbon atom to which they are attached, form a 3- to 7-membered heterocyclic ring optionally substituted with hydroxy or C1-C4 alkoxy. R10′ is hydrogen when R.sub.2 and R.sub.10 together form the heterocyclic ring, or R.sub.10 is hydrogen when R.sub.2 and R.sub.10, together form the heterocyclic ring.

[0640] In some aspects, when R.sub.2 is C.sub.1-C.sub.6 alkyl, the C.sub.1-C.sub.6 alkyl is preferably methyl, ethyl, propyl, butyl, or pentyl.

[0641] In some aspects, when R.sub.2 and R.sub.10, or R.sub.2 and R.sub.10′, together with the nitrogen atom and carbon atom to which they are attached, form a 3- to 7-membered heterocyclic ring optionally substituted with hydroxy or C.sub.1-C.sub.4 alkoxy, the 3- to 7-membered heterocyclic ring formed is preferably an azetidine ring, a pyrrolidine ring, or a piperidine ring.

[0642] In formula (III), except when R.sub.2 and R.sub.10 or R.sub.2 and R.sub.10′ together form the heterocyclic ring, [0643] (a) R.sub.10′ is hydrogen, and R.sub.10 is selected from the group consisting of hydrogen, optionally substituted C.sub.1-C.sub.6 alkyl, C.sub.3-C.sub.6 cycloalkyl, C.sub.3-C.sub.6 cycloalkyl-C.sub.1-C.sub.4 alkyl, optionally substituted phenyl, optionally substituted phenylmethyl, optionally substituted phenylethyl, 2-(methylthio)ethyl, —CH.sub.2SPG.sub.8, N-PG.sub.9-indol-3-ylmethyl, 4-(PG.sub.8O)benzyl, PG.sub.8-O-methyl (i.e., —CH.sub.2—O-PG.sub.8), 1-(PG.sub.8O)ethyl, 2-(PG.sub.8O)ethyl, PG.sub.8-OCO(CH.sub.2)—, PG.sub.8-OCO(CH.sub.2).sub.2—, PG.sub.9N-n-butyl, —CON(R.sub.16A)(R.sub.16B), —CH.sub.2—CON(R.sub.16A)(R.sub.16B), and —(CH.sub.2).sub.2CON(R.sub.16A)(R.sub.16B), [0644] (b) R.sub.10 and R.sub.10′ are independently optionally substituted C.sub.1-C.sub.6 alkyl, C.sub.3-C.sub.6 cycloalkyl, or C.sub.3-C.sub.6 cycloalkyl-C.sub.1-C.sub.4 alkyl, or [0645] (c) R.sub.10 and R.sub.10′, together with the carbon atom to which they are attached, form a 3- to 7-membered alicyclic ring.

[0646] The combination of R.sub.10 and R.sub.10′ is preferably a hydrogen atom and a hydrogen atom, methyl and a hydrogen atom, ethyl and a hydrogen atom, isopropyl and a hydrogen atom, isobutyl and a hydrogen atom, cyclopropyl and a hydrogen atom, cyclopropylmethyl and a hydrogen atom, optionally substituted phenyl and a hydrogen atom, optionally substituted phenylmethyl and a hydrogen atom, optionally substituted phenylethyl and a hydrogen atom, 2-(methylthio)ethyl and a hydrogen atom, —CH.sub.2SPG.sub.8 and a hydrogen atom, N-PG.sub.9-indol-3-ylmethyl and a hydrogen atom, 4-(PG.sub.8O)benzyl and a hydrogen atom, PG.sub.8-O-methyl and a hydrogen atom, 1-(PG.sub.8O)ethyl and a hydrogen atom, 2-(PG.sub.8O)ethyl and a hydrogen atom, PG.sub.8-OCO(CH.sub.2)— and a hydrogen atom, PG.sub.8-OCO(CH.sub.2).sub.2— and a hydrogen atom, PG.sub.9N-n-butyl and a hydrogen atom, —CON(R.sub.16A)(R.sub.16B) and a hydrogen atom, —CH.sub.2—CON(R.sub.16A)(R.sub.16B) and a hydrogen atom, —(CH.sub.2).sub.2CON(R.sub.16A)(R.sub.16B) and a hydrogen atom, methyl and methyl, or methyl and ethyl. When R.sub.10 and R.sub.10′, together with the carbon atom to which they are attached, form a 3- to 7-membered alicyclic ring, the alicyclic ring is preferably a cyclopropyl ring, a cyclobutyl ring, a cyclopentyl ring, or a cyclohexyl ring.

[0647] In formula (III), R.sub.11 is a single bond or —C(R.sub.11A)(R.sub.11B)—; and

[0648] R.sub.11A and R.sub.11B are independently selected from the group consisting of hydrogen, C1-C.sub.6 alkyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted aryl-C.sub.1-C.sub.4 alkyl, and optionally substituted heteroaryl-C.sub.1-C.sub.4 alkyl.

[0649] R.sub.11 is preferably a single bond or —C(R.sub.11A)(R.sub.11B)— where the combination of R.sub.11A and R.sub.11B is a hydrogen atom and a hydrogen atom, methyl and a hydrogen atom, ethyl and a hydrogen atom, isopropyl and a hydrogen atom, isobutyl and a hydrogen atom, cyclopropyl and a hydrogen atom, cyclopropylmethyl and a hydrogen atom, optionally substituted phenyl and a hydrogen atom, optionally substituted phenylmethyl and a hydrogen atom, optionally substituted phenylethyl and a hydrogen atom, methyl and methyl, or methyl and ethyl.

[0650] In formula (II), R.sub.12 and R.sub.12′ are independently selected from the group consisting of hydrogen, PG.sub.10-O-methyl (i.e., —CH.sub.2—O-PG.sub.10), —(CH.sub.2).sub.nCOO-PG.sub.10, —(CH.sub.2).sub.nCOO-RES, and —(CH.sub.2).sub.nCONH-RES. RES is a resin for solid-phase synthesis, and n is 0, 1, or 2.

[0651] In some aspects, when used for the resin removal method of the present invention, either one of R12 and R12′ is preferably selected from —(CH2)nCOO-RES and —(CH2)nCONH-RES.

[0652] In formula (II), R6 is selected from the group consisting of hydrogen and C1-C6 alkyl. In some aspects, when R6 is C1-C6 alkyl, the C1-C6 alkyl is preferably methyl, ethyl, propyl, butyl, or pentyl.

[0653] In formulas (II) and (III), preferably, either or both of R2 and R6 are other than hydrogen, and more preferably, either or both of R2 and R6 are C1-C6 alkyl.

[0654] In formula (II), R13 is C1-C4 alkyl or —(CH2)mCON(R17A)(R17B), wherein m is 0, 1, or 2, and R17A and R17B are independently hydrogen or C1-C4 alkyl, or R17A and R17B, together with the nitrogen atom to which they are attached, form a 4- to 8-membered ring optionally comprising one or more additional heteroatoms.

[0655] In some aspects, when R13 is C1-C4 alkyl, the C1-C4 alkyl is preferably methyl or ethyl.

[0656] In some aspects, when R17A and R17B, together with the nitrogen atom to which they are attached, form a 4- to 8-membered ring optionally comprising one or more additional heteroatoms, the ring formed is preferably an azetidine ring, a pyrrolidine ring, a piperidine ring, a piperazine ring, or a morpholine ring.

[0657] In formula (II), R16A and R16B are independently hydrogen or C1-C4 alkyl, or R16A and R16B, together with the nitrogen atom to which they are attached, form a 4- to 8-membered ring optionally comprising one or more additional heteroatoms.

[0658] When R16A and R16B are C1-C4 alkyl, the C1-C4 alkyl is preferably methyl, ethyl, or propyl.

[0659] When R16A and R16B, together with the nitrogen atom to which they are attached, form a 4- to 8-membered ring optionally comprising one or more additional heteroatoms, the 4- to 8-membered ring is preferably an azetidine ring, a pyrrolidine ring, a piperidine ring, a piperazine ring, or a morpholine ring.

[0660] PG1 is selected from the group consisting of Fmoc, Boc, Alloc, Cbz, Teoc, and trifluoroacetyl.

[0661] PG.sub.1 is preferably Fmoc, Boc, or Cbz.

[0662] In formula (II), PG.sub.8 is selected from the group consisting of hydrogen, t-Bu, trityl, methoxytrityl, cumyl, benzyl, THP, 1-ethoxyethyl, methyl, ethyl, allyl, optionally substituted aryl, optionally substituted aryl-C.sub.1-C.sub.4 alkyl, optionally substituted heteroaryl-C.sub.1-C.sub.4 alkyl, and 2-(trimethylsilyl)ethyl.

[0663] PG.sub.8 is preferably methyl, allyl, t-Bu, trityl, methoxytrityl, cumyl, THP, optionally substituted aryl, optionally substituted aryl-C.sub.1-C.sub.4 alkyl, or optionally substituted heteroaryl-C.sub.1-C.sub.4 alkyl.

[0664] In formula (II), PG9 is selected from the group consisting of hydrogen, Fmoc, Boc, Alloc, Cbz, Teoc, methoxycarbonyl, t-Bu, trityl, cumyl, and benzyl. PG9 is preferably Fmoc, Boc, Cbz, t-Bu, or trityl.

[0665] In formula (II), PG10 is selected from the group consisting of t-Bu, trityl, cumyl, benzyl, methyl, ethyl, allyl, optionally substituted aryl, optionally substituted aryl-C1-C4 alkyl, optionally substituted heteroaryl-C1-C4 alkyl, and 2-(trimethylsilyl)ethyl. PG10 is preferably t-Bu, trityl, cumyl, benzyl, methyl, or allyl.

[0666] In some aspects, PG1 to PG10 that can be comprised in general formulas (I) to (III) may be each independently a protecting group that can be removed by the deprotection method of the present invention. Such protecting groups include t-Bu, triphenylmethyl, 2-(trimethylsilyl)-ethyl, Boc, Teoc, Cbz, methoxycarbonyl, tetrahydropyranyl, and 1-ethoxyethyl.

[0667] In some aspects, PG1 to PG10 may be each independently a protecting group that cannot be removed by the deprotection method of the present invention. Such protecting groups include Fmoc and Alloc.

[0668] In some aspects, PG1 to PG10 may be each independently a group that does not function as a protecting group, for example, a group that cannot be removed or can be structurally transformed. Examples of such protecting groups include those described in Greene's “Protective Groups in Organic Synthesis” (5th ed., John Wiley & Sons 2014).

[0669] Structures represented by general formula (II), in which at least two amino acid residues are linked to each other, include many such structures easily subjected to cleavage or rearrangement when known deprotection or resin removal conditions are used. Such cleavage and rearrangement may be called damage. Only the protecting groups of interest or resins for solid-phase synthesis can be selectively and efficiently removed without such damage using the reaction conditions of the present invention.

[0670] In some aspects, structures represented by general formula (II), in which two amino acid residues are linked to each other, include those in which one N-substituted amino acid is linked to one N-unsubstituted amino acid and those in which two N-substituted amino acids are linked to each other. Specific examples of such structures include structures in which R1′ of general formula (II) is represented by general formula (III) and in which R.sub.11 of formula (III) is —C(R.sub.11A)(R.sub.11B)— (formula IV) or R.sub.11 of formula (III) is a single bond (formula V).

##STR00021##

[0671] Each group in formulas (IV) and (V) may be the same as that in formulas (II) and (III) above.

[0672] In some aspects, in formulas (IV) and (V), R1 is hydrogen, PG1, a natural amino acid residue, or an amino acid analog residue.

[0673] In some aspects, in formulas (IV) and (V), R1 is hydrogen, PG1, a natural amino acid residue, or an amino acid analog residue, and R2 and R10 are preferably methyl, ethyl, propyl, butyl, or pentyl.

[0674] In some aspects, in formulas (IV) and (V), R1 is hydrogen, PG1, a natural amino acid residue, or an amino acid analog residue, and R2 and R10 or R2 and R10′ preferably each independently, together with the nitrogen atom and carbon atom to which they are attached, form a 3- to 7-membered heterocyclic ring optionally substituted with hydroxy or C1-C4 alkoxy. R10′ is hydrogen when R2 and R10 together form the heterocyclic ring, and R10 is hydrogen when R2 and R10′ together form the 3- to 7-membered heterocyclic ring.

[0675] In some aspects, in formulas (IV) and (V), R10′ may be hydrogen, and R10 may be selected from the group consisting of hydrogen, optionally substituted C1-C6 alkyl, C3-C6 cycloalkyl, C3-C6 cycloalkyl-C1-C4 alkyl, optionally substituted phenyl, optionally substituted phenylmethyl, optionally substituted phenylethyl, 2-(methylthio)ethyl, —CH2SPG8, N-PG9-indol-3-ylmethyl, 4-(PG8O)benzyl, PG8-O-methyl, 1-(PG8O)ethyl, 2-(PG8O)ethyl, PG8-OCO(CH2)-, PG8-OCO(CH2)2-, PG9N-n-butyl, —CON(R16A)(R16B), —CH2-CON(R16A)(R16B), and —(CH2)2CON(R16A)(R16B).

[0676] In some aspects, in formulas (IV) and (V), preferably, R6 may be C1-C6 alkyl, which is preferably methyl, ethyl, propyl, butyl, or pentyl.

[0677] In some aspects, in formulas (IV) and (V), R13 may be C1-C4 alkyl, which is preferably methyl, ethyl, propyl, or butyl, and more preferably methyl or ethyl.

[0678] In some aspects, in formulas (IV) and (V), R.sub.13 may be —(CH.sub.2).sub.mCON(R.sub.17A)(R.sub.17B), and m is 0, 1, or 2. In this case, R17A and/or R17B may be independently C1-C4 alkyl, and the C1-C4 alkyl is preferably methyl or ethyl. R17A and R17B may, together with the nitrogen atom to which they are attached, form a 4- to 8-membered ring optionally comprising one or more additional heteroatoms, and the 4- to 8-membered ring is preferably an azetidine ring, a pyrrolidine ring, a piperidine ring, a piperazine ring, or a morpholine ring.

[0679] In some aspects, in formulas (IV) and (V), R12 and R12′ are independently selected from the group consisting of hydrogen, PG10-O-methyl, —(CH2)nCOO-PG10, —(CH2)nCOO-RES, and —(CH2)nCONH-RES. When R12 and R12′ is PG10-O-methyl, —(CH2)nCOO-PG10, —(CH2)nCOO-RES, or —(CH2)nCONH-RES, n is 0, 1, or 2, and PG10 is preferably t-Bu, trityl, cumyl, benzyl, methyl, or allyl. RES is a resin for solid-phase synthesis, and preferably CTC resin or Wang resin.

[0680] In some aspects, when used for the resin removal method of the present invention, either one of R12 and R12′ is preferably selected from —(CH2)nCOO-RES and —(CH2)nCONH-RES.

[0681] In some aspects, in formulas (IV) and (V), more preferably, R1 is hydrogen, PG.sub.1, a natural amino acid residue, or an amino acid analog residue; [0682] R.sub.2 is methyl and ethyl; and/or [0683] R.sub.2 and R.sub.10, together with the nitrogen atom and carbon atom to which they are attached, form a 4- to 6-membered heterocyclic ring optionally substituted with hydroxy or C.sub.1-C.sub.4 alkoxy. [0684] R.sub.10 and R.sub.10′ may be each independently selected from methyl or ethyl, or R.sub.10 and R.sub.10′ may together form a 5- or 6-membered alicyclic ring. Alternatively, R.sub.10′ may be hydrogen, and R.sub.10 may be hydrogen, or alkyl selected from methyl, ethyl, isopropyl, isobutyl, and sec-butyl, or optionally substituted phenylmethyl, optionally substituted phenylethyl, benzyloxymethyl, 1-benzyloxyethyl, 2-benzyloxyethyl, tert-butoxycarbonylmethyl, methoxycarbonylmethyl, tert-butoxycarbonylethyl, methoxycarbonylethyl, tert-butoxycarbamoylbutyl, N,N-dimethylaminocarbonyl, piperidylcarbonyl, pyrrolidylcarbonyl, N,N-dimethylaminocarbonylmethyl, piperidylcarbonylmethyl, pyrrolidylcarbonylmethyl, N,N-dimethylaminocarbonylethyl, piperidylcarbonylethyl, or pyrrolidylcarbonylethyl. [0685] R.sub.6 may be methyl or ethyl, and [0686] R.sub.13 may be methyl or —(CH.sub.2).sub.mCON(R.sub.17A)(R.sub.17B). When R.sub.13 is —(CH.sub.2).sub.mCON(R.sub.17A)(R.sub.17B), R.sub.17A and R.sub.17B, together with the nitrogen atom to which they are attached, form a piperidine ring. [0687] R.sub.12 and R.sub.12′ are independently selected from the group consisting of hydrogen, PG.sub.10-O-methyl, —(CH.sub.2).sub.nCOO-PG.sub.10, —(CH.sub.2).sub.nCOO-RES, and —(CH.sub.2).sub.nCONH-RES.

[0688] In some aspects, the starting peptide compound may comprise one or more additional natural amino acid residues and/or amino acid analog residues, in addition to a structure represented by general formula (II). The starting peptide compound may comprise one or more structures represented by general formula (I), may comprise a structure represented by general formula (II) at the C-terminus, and optionally may further comprise one or more additional natural amino acid residues and/or amino acid analog residues. The peptide compound may be formed by secondary amides, tertiary amides, or a mixture of secondary and tertiary amides.

[0689] In some aspects, the starting peptide compound comprises at least one protecting group removable by the method of the present invention. Such a protecting group may be contained in a structure represented by general formula (II), or may be contained in an amino acid residue other than the structure represented by general formula (II).

[0690] In some aspects, the starting peptide compound comprises at least one resin for solid-phase synthesis that is removable by the method of the present invention. When the starting peptide compound comprises a structure represented by general formula (II), such a resin is included in the structure represented by general formula (II).

[0691] In some aspects, the present invention relates to a method of producing an amide compound in which a protecting group removable by a silylating agent is removed, the method comprising the step of contacting a starting amide compound with the silylating agent in a solvent and thereby removing the protecting group from the starting amide compound.

[0692] In some aspects, the present invention relates to a method of producing an amide compound in which a resin for solid-phase synthesis is removed, the method comprising the step of contacting a starting amide compound with a silylating agent in a solvent and thereby removing the starting amide compound from the resin for solid-phase synthesis

[0693] In some aspects, the starting amide compound to be subjected to deprotection/resin removal is represented by general formula (II).

##STR00022##

[0694] In formula (II), R.sub.1′ is a hydrogen atom or PG.sub.7, and the other groups are the same as defined above. R.sub.6 is preferably C.sub.1-C.sub.6 alkyl. PG.sub.7 is selected from the group consisting of Fmoc, Boc, Alloc, Cbz, Teoc, and trifluoroacetyl and is preferably Fmoc, Boc, or Cbz.

[0695] Starting amide compounds represented by general formula (II), in particular, N-substituted compounds are susceptible to damage such as rearrangement when known deprotection or resin removal conditions are used. Only the protecting groups of interest or resin for solid-phase synthesis can be selectively and efficiently removed without such damage using the reaction conditions of the present invention.

[0696] Herein, the “silylating agent” is not particularly limited as long as it can be used for the deprotection or resin removal reaction of the present invention, and the term refers to an agent that can function as a deprotecting agent or reagent and/or a resin-removing agent or reagent. Herein, “silylating agents” may be called deprotecting agents or reagents and/or resin-removing agents or reagents. The silylating agent can be prepared, for example, by mixing the silyl compound with an electrophilic species scavenger or mixing an acid with an electrophilic species scavenger having a silyl group in a solvent. The mixing of the silyl compound with an electrophilic species scavenger or the mixing of an acid with an electrophilic species scavenger having a silyl group may be performed previously, or in a solvent, or in the presence or absence of the starting peptide compound.

[0697] Examples of the “silyl compound” as used herein include silyl compounds having a leaving group (X) represented by formula 1:

##STR00023##

wherein R.sub.AX, R.sub.AY, and R.sub.AZ are independently C.sub.1-C.sub.4 alkyl or phenyl, and X is selected from the group consisting of —OTf, —OClO.sub.3, Cl, Br, and I.
Preferably, R.sub.AX, R.sub.AY, and R.sub.AZ may be independently selected from methyl, ethyl, i-propyl, t-butyl, and phenyl.

[0698] More specific examples of such silyl compounds include TMSOTf, TESOTf, TBSOTf, TIPSOTf, TBDPSOTf, TTMSOTf, TMSCl, TMSBr, TMSOClO.sub.3, and TMSI.

[0699] The “acid” that can be used herein for preparing a silylating agent is not particularly limited as long as it can generate the silyl compound, and examples thereof include acids represented by HX (wherein X is as defined for X of formula (1)).

[0700] As used herein, the term “electrophilic species scavenger” refers to a compound that can form an adduct or salt with a proton or is reactive with a cationic species and that is less susceptible to silylation. Such electrophilic species scavengers include imidates (formula 2), amides (formula 3), ketene acetals (formula 4), ketene alkoxy hemiaminals (formula 4), enol ethers (formula 4), enol esters (formula 4), imines (formula 5), amines (formula 6), diamines (formula 7), dialkylcarbodiimides (formula 8), ureas (formula 9), or urethanes (formula 10). They may have a substituted silyl group, specific examples of the substituted silyl group include TMS (trimethylsilyl), TES (triethylsilyl), TBS (tributylsilyl), TIPS (triisopropylsilyl), and TBDPS (t-butyl-dimethylsilyl).

[0701] The imidates are represented by formula 2:

##STR00024##

wherein [0702] R.sub.B is a substituted silyl group and R.sub.C is a substituted silyl group, or [0703] R.sub.B and R.sub.C, together with the nitrogen atom and carbon atom to which they are attached, form a 5- to 7-membered ring; and [0704] R.sub.D is C.sub.1-C.sub.4 alkyl optionally substituted with one or more fluorine atoms or is optionally substituted methylene, wherein when R.sub.D is optionally substituted methylene, formula 2 is dimerized to form a compound represented by the formula below:

##STR00025##

[0705] Preferred imidates include N-silyl imidates represented by formula 2-1 and bisoxazolines represented by formula 2-2 below:

##STR00026##

wherein [0706] R.sub.BX, R.sub.BY, and R.sub.BZ are independently C.sub.1-C.sub.4 alkyl or phenyl, R.sub.C and R.sub.D are as defined above; and [0707] R.sub.E and R.sub.F are independently C.sub.1-C.sub.4 alkyl. [0708] Preferably, R.sub.BX, R.sub.BY, and R.sub.BZ may be independently selected from methyl, ethyl, i-propyl, t-butyl, and phenyl.

[0709] Preferred N-silyl imidates include N,O-bis(trimethylsilyl)acetamides represented by formula 2-1-1. Preferred bisoxazolines include 4-substituted bisoxazolines represented by formula 2-2-1.

##STR00027##

[0710] More preferred N-silyl imidates include N,O-bis(trimethylsilyl)acetamide represented by formula 2-1-1-1 and N,O-bis(trimethylsilyl)trifluoroacetamide represented by formula 2-1-1-2. More preferred bisoxazolines include N,O-bis(trimethylsilyl)trifluoroacetamide 2,2′-isopropylidenebis[(4S)-4-tert-butyl-2-oxazoline] represented by formula 2-2-1-1.

##STR00028##

[0711] The amides are represented by formula 3:

##STR00029##

wherein [0712] R.sub.G is a silyl group substituted with one or more C.sub.1-C.sub.4 alkyl; [0713] R.sub.H is hydrogen or C.sub.1-C.sub.4 alkyl; and [0714] R.sub.I is hydrogen, or C.sub.1-C.sub.4 alkyl optionally substituted with one or more fluorine atoms.

[0715] Preferred amides include N-silylamides represented by formula 3-1:

##STR00030##

wherein [0716] RGX, RGY, and RGZ are independently C1-C4 alkyl or phenyl; and [0717] R.sub.H and R.sub.I are as defined above.

[0718] Preferably, R.sub.GX, R.sub.GY, and R.sub.GZ may be independently selected from methyl, ethyl, i-propyl, t-butyl, and phenyl.

[0719] Preferred N-silylamides include N-trimethylsilylacetamides represented by formula 3-1-1:

##STR00031##

wherein RH and RI are as defined above.

[0720] More preferably, N-trimethylsilylacetamides include N-methyl-N-trimethylsilylacetamide represented by formula 3-1-1-1 and N-methyl-N-trimethylsilyltrifluoroacetamide represented by formula 3-1-1-2.

##STR00032##

[0721] The ketene acetals, ketene alkoxy hemiaminals, enol ethers, and enol esters are each represented by formula 4:

##STR00033##

wherein [0722] (a-1) RJ is a substituted silyl group, RK is C1-C4 alkoxy, and RM and RL are independently hydrogen or C.sub.1-C.sub.4 alkyl; [0723] (a-2) R.sub.J is a substituted silyl group, R.sub.M is hydrogen or C.sub.1-C.sub.4 alkyl, and R.sub.K and R.sub.L, together with the carbon atoms to which they are attached, form a 5- to 8-membered ring comprising an oxygen atom; [0724] (b-1) R.sub.J is a substituted silyl group, R.sub.K is C.sub.1-C.sub.4 alkyl, and R.sub.M and R.sub.L are independently hydrogen or C.sub.1-C.sub.4 alkyl; [0725] (b-2) R.sub.J is a substituted silyl group, R.sub.M is hydrogen or C.sub.1-C.sub.4 alkyl, and R.sub.K and R.sub.L, together with the carbon atoms to which they are attached, form a 5- to 8-membered ring; [0726] (c-1) R.sub.J and R.sub.M, together with the carbon atoms to which they are attached, form a 5- to 7-membered ring comprising an oxygen atom, R.sub.K is hydrogen or C.sub.1-C.sub.4 alkyl, and R.sub.L is C.sub.1-C.sub.4 alkyl; [0727] (c-2) R.sub.J and R.sub.M, together with the carbon atoms to which they are attached, form a 5- to 7-membered ring comprising an oxygen atom, and R.sub.K and R.sub.L, together with the carbon atoms to which they are attached, form a 5- to 8-membered ring; [0728] (d-1) R.sub.J is C.sub.1-C.sub.4 alkyl and R.sub.M, R.sub.K, and R.sub.L are independently hydrogen or C.sub.1-C.sub.4 alkyl; [0729] (d-2) R.sub.J is C.sub.1-C.sub.4 alkyl, R.sub.M is hydrogen or C.sub.1-C.sub.4 alkyl, and R.sub.K and R.sub.L, together with the carbon atoms to which they are attached, form a 5- to 8-membered ring; [0730] (e-1) R.sub.J is C.sub.1-C.sub.3 alkylcarbonyl and R.sub.M, R.sub.K, and R.sub.L are independently hydrogen or C1-C4 alkyl; [0731] (e-2) R.sub.J is C.sub.1-C.sub.3 alkylcarbonyl, R.sub.M is hydrogen or C1-C4 alkyl, and R.sub.K and R.sub.L, together with the carbon atoms to which they are attached, form a 5- to 8-membered ring; [0732] (f-1) RJ is a substituted silyl group or C1-C4 alkyl, RK is optionally substituted di-C1-C4 alkylamino, and RM and RL are independently hydrogen or C1-C4 alkyl; or [0733] (f-2) RJ is a substituted silyl group or C1-C4 alkyl, RM is hydrogen or C1-C4 alkyl, and RK and RL, together with the carbon atoms to which they are attached, form a 5- to 8-membered ring comprising a nitrogen atom, and the 5- to 8-membered ring is optionally substituted with C.sub.1-C.sub.4 alkyl.

[0734] Here, preferred examples of the C.sub.1-C.sub.4 alkyl include methyl, ethyl, n-propyl, isopropyl, n-butyl, i-butyl, and t-butyl. Preferred examples of the C.sub.1-C.sub.3 alkylcarbonyl include acetyl and propionyl.

[0735] Preferred ketene acetals include silylketene acetals represented by formula 4-1:

##STR00034##

wherein [0736] R.sub.JX1, R.sub.JY1, and R.sub.JZ1 are independently C.sub.1-C.sub.4 alkyl or phenyl; [0737] R.sub.KX1 is C.sub.1-C.sub.4 alkyl; and [0738] R.sub.L and R.sub.M are as defined above. [0739] Here, preferred examples of the C.sub.1-C.sub.4 alkyl include methyl, ethyl, n-propyl, isopropyl, n-butyl, i-butyl, and t-butyl. [0740] Preferably, R.sub.JX1, R.sub.JY1, and R.sub.JZ1 may be independently selected from methyl, ethyl, i-propyl, t-butyl, and phenyl.

[0741] Preferred ketene silyl acetals include ketene trimethylsilyl acetals represented by formula 4-1-1:

##STR00035##

wherein R.sub.KX1, R.sub.L, and R.sub.M are as defined above.

[0742] Preferred R.sub.KX1, R.sub.L, and R.sub.M include methyl, ethyl, n-propyl, isopropyl, n-butyl, i-butyl, and t-butyl.

[0743] Preferred ketene trimethylsilyl acetals include dimethylketene methyl trimethylsilyl acetal represented by formula 4-1-1-1.

##STR00036##

[0744] Preferred enol ethers include silylenol ethers represented by formula 4-2:

##STR00037##

wherein [0745] R.sub.JX2, R.sub.JY2, and R.sub.JZ2 are independently C.sub.1-C.sub.4 alkyl or phenyl; [0746] R.sub.KX2 is hydrogen or C.sub.1-C.sub.4 alkyl; and [0747] R.sub.L and R.sub.M are as defined above. [0748] Preferred C.sub.1-C.sub.4 alkyl of R.sub.KX1, R.sub.L, and R.sub.M include methyl, ethyl, n-propyl, isopropyl, n-butyl, i-butyl, and t-butyl. [0749] Preferably, R.sub.JX2, R.sub.JY2, and R.sub.JZ2 are independently selected from the group consisting of methyl, ethyl, i-propyl, t-butyl, and phenyl.

[0750] Preferred silyl enol ethers include trimethylsilyl enol ethers represented by formula 4-2-1:

##STR00038##

wherein [0751] R.sub.KX2, R.sub.L, and R.sub.M are as defined above.

[0752] Preferred trimethylsilyl enol ethers include isopropenyloxytrimethylsilane represented by formula 4-2-1-1.

##STR00039##

[0753] In another embodiment, preferred enol ethers include cyclic enol ethers represented by formula 4-3:

##STR00040##

wherein [0754] R.sub.KX3 and RU are independently hydrogen or C.sub.1-C.sub.4 alkyl.

[0755] Preferred cyclic enol ethers include dihydropyran represented by formula 4-3-1.

##STR00041##

[0756] In another embodiment, preferred enol ethers include linear enol ethers, and preferred linear enol ethers include ethyl vinyl ether represented by formula 4-4.

##STR00042##

[0757] The imines are represented by formula 5:

##STR00043##

wherein [0758] R.sub.N, R.sub.N′, and R.sub.O are independently hydrogen or C.sub.1-C.sub.4 alkyl. [0759] Preferred C.sub.1-C.sub.4 alkyl of R.sub.N, R.sub.N′, and R.sub.O include methyl, ethyl, n-propyl, isopropyl, n-butyl, i-butyl, and t-butyl.

[0760] Preferred imines include ketimines represented by formula 5-1:

##STR00044## [0761] wherein R.sub.N is as defined above. [0762] Preferred R.sub.N and R.sub.N′ include methyl, ethyl, n-propyl, isopropyl, n-butyl, i-butyl, and t-butyl.

[0763] Preferred ketimines include 2,2,4,4-tetramethylpentanone imine represented by formula 5-1-1.

##STR00045##

[0764] The amines are represented by formula 6:

##STR00046## [0765] wherein [0766] R.sub.P is a substituted silyl group; and [0767] R.sub.Q is a substituted silyl group or C.sub.1-C.sub.4 alkyl and R.sub.R is hydrogen, a substituted silyl group, or C.sub.1-C.sub.4 alkyl, or [0768] R.sub.Q and R.sub.R, together with the nitrogen atom to which they are attached, form a 5- to [0769] 8-membered heterocyclic ring comprising an oxygen atom.

[0770] Preferred amines include disilylamines represented by formula 6-1:

##STR00047## [0771] wherein [0772] R.sub.PX1, R.sub.PY1, R.sub.PZ1, R.sub.QX1, R.sub.QY1, and R.sub.QZ1 are independently C.sub.1-C.sub.4 alkyl or phenyl. [0773] Preferably, R.sub.PX1, R.sub.PY1, R.sub.PZ1, R.sub.QX1, R.sub.QY1, and R.sub.QZ1 are independently selected from the group consisting of methyl, ethyl, i-propyl, t-butyl, and phenyl.

[0774] Preferred disilylamines include 1,1,1,3,3,3-hexamethyldisilazane (HMDS) represented by formula 6-1-1.

##STR00048##

[0775] In another embodiment, preferred amines include N-dialkyl-N-silylamines represented by formula 6-2:

##STR00049## [0776] wherein [0777] R.sub.PX2, R.sub.PY2, and R.sub.PZ2 are independently C.sub.1-C.sub.4 alkyl or phenyl; and [0778] R.sub.Q and R.sub.R are as defined above. [0779] Preferred C.sub.1-C.sub.4 alkyl of R.sub.PX2, R.sub.PY2, and R.sub.PZ2 include methyl, ethyl, n-propyl, isopropyl, n-butyl, i-butyl, and t-butyl. [0780] Preferably, R.sub.PX2, R.sub.PY2, and R.sub.PZ2 are independently selected from the group consisting of methyl, ethyl, i-propyl, t-butyl, and phenyl.

[0781] Preferred N-dialkyl-N-silylamines include N-dialkyl-N-trimethylsilylamines represented by formula 6-2-1:

##STR00050##

wherein R.sub.Q and R.sub.R are as defined above.

[0782] Preferred N-dialkyl-N-trimethylsilylamines include N-trimethylsilylmorpholine represented by formula 6-2-1-1 and N-trimethylsilyldiethylamine represented by formula 6-2-1-2.

##STR00051##

[0783] In another embodiment, preferred amines include N-alkyl-N-disilylamines represented by formula 6-3:

##STR00052## [0784] wherein [0785] R.sub.QX3, R.sub.QY3, R.sub.QZ3, R.sub.RX1, R.sub.RY1, and R.sub.RZ1 are independently C.sub.1-C.sub.4 alkyl or phenyl; and [0786] R.sub.R is C.sub.1-C.sub.4 alkyl. [0787] Specifically, for example, R.sub.QX3, R.sub.QY3, R.sub.QZ3, R.sub.RX1, R.sub.RY1, and R.sub.RZ1 may be independently selected from methyl, ethyl, i-propyl, t-butyl, and phenyl. [0788] Preferred C.sub.1-C.sub.4 alkyl of R.sub.R include methyl, ethyl, n-propyl, isopropyl, n-butyl, i-butyl, and t-butyl.

[0789] Preferred N-alkyl-N-disilylamines include N-alkyl-N-bistrimethylsilylamines represented by formula 6-3-1:

##STR00053## [0790] wherein [0791] R.sub.R is C.sub.1-C.sub.4 alkyl. [0792] Preferred C.sub.1-C.sub.4 alkyl of R.sub.R include methyl, ethyl, n-propyl, isopropyl, n-butyl, i-butyl, and t-butyl.

[0793] In another embodiment, preferred amines include N-alkyl-N-silylamines represented by formula 6-4:

##STR00054## [0794] wherein [0795] R.sub.PX2, R.sub.PY2, and R.sub.PZ2 are independently C.sub.1-C.sub.4 alkyl or phenyl; and [0796] R.sub.Q is C.sub.1-C.sub.4 alkyl. [0797] Preferably, R.sub.PX2, R.sub.PY2, and R.sub.PZ2 are independently selected from the group consisting of methyl, ethyl, i-propyl, t-butyl, and phenyl. [0798] Preferred C.sub.1-C.sub.4 alkyl of R.sub.Q include methyl, ethyl, n-propyl, isopropyl, n-butyl, i-butyl, and t-butyl.

[0799] Preferred N-alkyl-N-silylamines include N-alkyl-N-trimethylsilylamines represented by formula 6-4-1:

##STR00055## [0800] wherein R.sub.Q is C.sub.1-C.sub.4 alkyl. [0801] Preferred C1-C4 alkyl of R.sub.Q include methyl, ethyl, n-propyl, isopropyl, n-butyl, i-butyl, and t-butyl.

[0802] Preferred N-alkyl-N-trimethylsilylamines include N-tert-butyltrimethylsilylamine represented by formula 6-4-1-1.

##STR00056##

[0803] The diamines are represented by formula 7:

##STR00057## [0804] wherein [0805] X is a single bond or a carbon atom, [0806] wherein when X is a single bond, R.sub.S is absent, R.sub.UA and R.sub.R, together with the carbon atom and nitrogen atom to which they are attached, form an optionally substituted 6-membered aromatic heterocyclic ring, and R.sub.UB and R.sub.T, together with the carbon atom and nitrogen atom to which they are attached, form an optionally substituted 6-membered aromatic heterocyclic ring, and [0807] when X is a carbon atom, R.sub.UA and R.sub.UB are independently C.sub.1-C.sub.4 alkyl and R.sub.R, R.sub.S, and R.sub.T, together with the carbon atoms to which they are attached, form the structure below:

##STR00058##

[0808] When X in formula 7 is a carbon atom, preferred diamines include tetraalkylnaphthalenediamines represented by formula 7-1:

##STR00059## [0809] wherein R.sub.UA and R.sub.UB are independently C.sub.1-C.sub.4 alkyl.

[0810] Preferred tetraalkylnaphthalenediamines include N,N,N′,N′-tetramethyl-1,8-naphthalenediamine represented by formula 7-1-1.

##STR00060##

[0811] In another embodiment, when X in formula 7 is a single bond, preferred diamines include 2,2′-bipyridine represented by formula 7-2-1.

##STR00061##

[0812] The dialkylcarbodiimides are represented by formula 8:

##STR00062## [0813] wherein R.sub.V is C.sub.1-C.sub.4 alkyl or C.sub.3-C.sub.6 cycloalkyl. [0814] Preferred C.sub.1-C.sub.4 alkyl of R.sub.V include methyl, ethyl, n-propyl, isopropyl, n-butyl, i-butyl, t-butyl, and cyclohexyl.

[0815] Preferred dialkylcarbodiimides include diisopropylcarbodiimide represented by formula 8-1.

##STR00063##

[0816] The ureas are represented by formula 9:

##STR00064## [0817] wherein R.sub.W and R.sub.X are independently C1-C4 alkyl or a substituted silyl group.

[0818] Preferred ureas include N,N′-bis(trimethylsilyl)urea represented by formula 9-1.

##STR00065##

[0819] The urethanes are represented by formula 10:

##STR00066## [0820] wherein R.sub.Y and R.sub.Z are independently C.sub.1-C.sub.4 alkyl or a substituted silyl group.

[0821] Preferred urethanes include N,O-bis(trimethylsilyl)urea represented by formula 10-1.

##STR00067##

[0822] In some aspects, silylating agents can be prepared by mixing a peptide compound, a solvent, an electrophilic species scavenger, and a silyl compound or acid in any order. It is preferred to mix the peptide compound with the solvent, then with the electrophilic species scavenger, and subsequently with the silyl compound or acid.

[0823] In some aspects, the solvent used may be an aprotic solvent, examples of which include esters, ethers, alkylnitriles, halogenated hydrocarbons, and hydrocarbons. Among them, ethyl acetate, isopropyl acetate, 2-methyltetrahydrofuran, tetrahydrofuran, diethyl ether, methyl tert-butyl ether, dichloromethane, 1,2-dichloroethane, toluene, or acetonitrile is preferred, ethyl acetate, isopropyl acetate, 2-methyltetrahydrofuran, tetrahydrofuran, or 1,2-dichloroethane is more preferred, and ethyl acetate or 2-methyltetrahydrofuran is particularly preferred.

[0824] In some aspects, a stoichiometric amount or more of a silyl compound, and an electrophilic species scavenger can be used in order to produce a silylating agent in the present invention to remove a protecting group and/or remove a resin from a peptide compound. In this case, 1 to 5 equivalents, preferably 2 to 4 equivalents, of a silyl compound and 1 to 10 equivalents, preferably 1 to 8 equivalents, of an electrophilic species scavenger can be used for one equivalent of the protecting group to be removed or one equivalent of the resin to be removed which is contained in a peptide, for example.

[0825] In some aspects, a silylating agent can be prepared by combining an electrophilic species scavenger with a catalytic amount of, for example, 0.1 to 0.5 equivalent, preferably 0.3 to 0.4 equivalent, of a silyl compound, per one equivalent of the protecting group to be removed or one equivalent of the resin to be removed. In this case, TMSOTf, TESOTf, TBSOTf, TIPSOTf, TBDPSOTf, TTMSOTf, TMSCl, TMSBr, or TMSOClO.sub.3 is preferably used as such a silylating agent, and N,O-bis(trimethylsilyl)acetamide, N,O-bis(trimethylsilyl)trifluoroacetamide, N-methyl-N-trimethylsilylacetamide, N-methyl-N-trimethylsilyltrifluoroacetamide, dimethylketene methyl trimethylsilyl acetal, or isopropenyloxytrimethylsilane is preferably used as such an electrophilic species scavenger. In this case, 1 to 10 equivalents, preferably 1 to 8 equivalents, of the electrophilic species scavenger can be used per one equivalent of the protecting group to be removed or one equivalent of the resin to be removed.

[0826] In some aspects, a silylating agent can be prepared by combining an electrophilic species scavenger with a catalytic amount of, for example, 0.1 to 0.5 equivalent, preferably 0.3 to 0.4 equivalent, of an acid per one equivalent of the protecting group to be removed or one equivalent of the resin to be removed. In this case, TfOH, HOClO.sub.3, HCl, HBr, or HI is preferably used, and TfOH is more preferably used as such an acid. N-silyl imidates (formula 2-1), N-silylamides (formula 3-1), ketene silyl acetals (formula 4-1), and silyl enol ethers (formula 4-2) are preferably used, and N,O-bis(trimethylsilyl)acetamide (formula 2-1-1-1), N,O-bis(trimethylsilyl)trifluoroacetamide (formula 2-1-1-2), N-methyl-N-trimethylsilylacetamide (formula 3-1-1-1), N-methyl-N-trimethylsilyltrifluoroacetamide (formula 3-1-1-2), dimethylketene methyl trimethylsilyl acetal (formula 4-1-1-1), and isopropenyloxytrimethylsilane (formula 4-2-1-1) are more preferably used as such electrophilic species scavengers. Among them, N,O-bis(trimethylsilyl)trifluoroacetamide and N-methyl-N-trimethylsilyltrifluoroacetamide are particularly preferred. In this case, 1 to 10 equivalents, preferably 1 to 8 equivalents, of the electrophilic species scavenger can be used per one equivalent of the protecting group to be removed or one equivalent of the resin to be removed.

[0827] In some aspects, the “protecting group removable by a silylating agent” is not particularly limited, as long as the protecting group can be removed by the silylating agent. Specific examples of such protecting groups include protecting groups for carboxyl groups, such as t-Bu, benzyl, triphenylmethyl, cumyl, and 2-(trimethylsilyl)-ethyl; protecting groups for amino groups, such as Boc, Teoc, Cbz, methoxycarbonyl, and cumyl; protecting groups for hydroxy groups, such as tetrahydropyranyl and 1-ethoxyethyl; and protecting groups for SH groups, such as triphenylmethyl and methoxytrityl.

[0828] Chemical synthesis methods for start peptide compounds herein include, for example, liquid phase synthesis methods, solid phase synthesis methods using Fmoc synthesis, Boc synthesis, or such, and combinations thereof. In Fmoc synthesis, an amino acid in which the main chain amino group is protected with an Fmoc group, the side-chain functional groups are protected when necessary with protecting groups that are not cleaved by a base such as piperidine, and the main chain carboxylic acid group is not protected, is used as a basic unit. The basic unit is not particularly limited and may be any other combination as long as it has an Fmoc-protected amino group and a carboxylic acid group. For example, a dipeptide may be used as a basic unit. The basic unit to be positioned at the N terminus may be one that is not an Fmoc amino acid. For example, it may be a Boc amino acid, or a carboxylic acid analog that does not have an amino group. The main chain carboxylic acid group is immobilized onto a solid phase by a chemical reaction with a functional group on a solid-phase carrier. Next, the Fmoc group is deprotected by a base such as piperidine or DBU, and the newly generated amino group and a subsequently added basic unit, i.e. a protected amino acid carrying a carboxylic acid, are subjected to a condensation reaction to generate an amide bond. In the condensation reaction, various combinations such as DIC and HOBt, DIC and HOAt, and HATU and DIPEA are possible. Repeating the Fmoc group deprotection and the subsequent amide bond-forming reaction enables generation of the desired peptide sequence. After the desired sequence is obtained, removal of the resin from the solid phase is performed, and the protecting groups introduced as necessary to the side-chain functional groups are deprotected. The deprotection and resin removal methods of the present invention can be provided for removal from solid-phase resins and removal of protecting groups from side chain functional groups. Steps such as cyclization may also be conducted during the solid-phase resin removal step or the deprotection step or finally. For example, side chain carboxylic acids may be condensed with N-terminal amino groups of main chains, and side chain amino groups may be condensed with C-terminal carboxylic acids of main chains. In this case, reaction orthogonality is needed between the C-terminal carboxylic acid and the side chain carboxylic acid to be cyclized, or between the N-terminal amino or hydroxy group of the main chain and the side chain amino group to be cyclized, and the protecting group is selected taking orthogonality of the protecting group into consideration. Reaction products thus obtained can be purified by reverse phase columns, molecular sieve columns, or the like. The details are described in Solid-Phase Synthesis Handbook published by Merck K.K. on May 1, 2002, for example.

[0829] Herein, the “resin for solid-phase synthesis” is not particularly limited, as long as it can be used for the synthesis of peptide compounds by the solid-phase method and can be removed by the silylating agent of the present invention. Specific examples of such resins for solid-phase synthesis include those removable under acidic conditions, such as CTC resins, Wang resins, SASRIN resins, trityl chloride resins (Trt resins), 4-methyltrityl chloride resins (Mtt resins), and 4-methoxytrityl chloride resins (Mmt). The resin can be appropriately selected according to the functional group on the amino acid to be used. For example, when using a carboxylic acid (main-chain carboxylic acid or side-chain carboxylic acid represented by Asp or Glu) or a hydroxy group on an aromatic ring (phenol group represented by Tyr) as the functional group on the amino acid, use of trityl chloride resin (Trt resin) or 2-chlorotritylchloride resin (CTC resin) as the resin is preferred. When using an aliphatic hydroxy group (aliphatic alcohol group represented by Ser or Thr) as the functional group on the amino acid, use of tritylchloride resin (Trt resin), 2-chlorotritylchloride resin (CTC resin), or 4-methyltritylchloride resin (Mtt resin) as the resin is preferred.

[0830] Furthermore, the types of polymers constituting the resins are also not particularly limited. For resins composed of polystyrenes, either 100 to 200 mesh or 200 to 400 mesh may be used. The cross-link percentage is also not particularly limited, but those cross-linked with 1% divinylbenzene (DVB) are preferred. Types of the polymer forming the resin include Tentagel or Chemmatrix.

[0831] In some aspects, the deprotection reaction and/or resin removal reaction of the present invention can be performed at a reaction temperature of −50 to 100° C., preferably −20 to 50° C., and more preferably 0 to 30° C.

[0832] In some aspects, the deprotection reaction and/or resin removal reaction of the present invention can be performed for a reaction time of 10 minutes to one week, preferably 10 minutes to 6 hours, and more preferably 1 to 3 hours.

[0833] In some aspects, in the deprotection reaction and/or resin removal reaction of the present invention, a target product can be obtained by adding an alcohol or water to a reaction solution to quench the reaction, and causing the target product to precipitate or washing an organic layer with water or saline and then concentrating the organic layer under reduced pressure. The alcohol is not particularly limited, but is preferably a water-soluble and low-boiling alcohol, and particularly preferably methanol. The water used for quenching the reaction is not particularly limited, but is preferably alkaline water, and particularly preferably aqueous sodium bicarbonate or dipotassium hydrogenphosphate. When the organic layer is washed, the saline is not particularly limited in terms of concentration, but is preferably brine or 5% saline.

[0834] In some aspects, when a starting peptide compound comprises both a protecting group that can be removed by the method of the present invention and a resin for solid-phase synthesis that can be removed by the method of the present invention, deprotection and resin removal reactions can be conducted at the same time. Specifically, the present invention also relates to a method of producing a peptide compound in which a protecting group removable by a silylating agent is removed from a starting peptide compound comprising natural amino acid residues and/or amino acid analog residues and in which a resin for solid-phase synthesis that is removable by the silylating agent is removed from the starting peptide compound, the method comprising the step of contacting the starting peptide compound with the silylating agent in a solvent.

[0835] In some aspects, the present invention relates to an amide compound represented by formula (A) below or a salt thereof:

##STR00068##

[0836] In formula (A), R.sub.1′ is selected from the group consisting of hydrogen, Fmoc, Boc, Alloc, Cbz, Teoc, and trifluoroacetyl. R.sub.1′ is preferably hydrogen, Fmoc, Boc, or Cbz.

[0837] In formula (A), R.sub.17A and R.sub.17B are both methyl, or R.sub.17A and R.sub.17B, together with the nitrogen atom to which they are attached, form piperidine or morpholine.

[0838] In formula (A), R.sub.18 is hydrogen or PG.sub.10. PG.sub.10 is selected from the group consisting of t-Bu, trityl, cumyl, benzyl, methyl, ethyl, allyl, optionally substituted aryl, optionally substituted aryl-C.sub.1-C.sub.4 alkyl, optionally substituted heteroaryl-C.sub.1-C.sub.4 alkyl, and 2-(trimethylsilyl)ethyl. R.sub.18 is preferably a hydrogen atom, t-Bu, benzyl, or allyl.

[0839] Preferred combinations of R.sub.1′ and R.sub.18 in formula (A) include Fmoc and a hydrogen atom, Fmoc and allyl, Fmoc and t-Bu, Fmoc and benzyl, Boc and a hydrogen atom, Boc and allyl, Boc and t-Bu, Boc and benzyl, Cbz and a hydrogen atom, Cbz and allyl, Cbz and t-Bu, Cbz and benzyl, Alloc and a hydrogen atom, Alloc and allyl, Alloc and t-Bu, Alloc and benzyl, Teoc and a hydrogen atom, Teoc and allyl, Teoc and t-Bu, Teoc and benzyl, a hydrogen atom and a hydrogen atom, a hydrogen atom and allyl, a hydrogen atom and t-Bu, and a hydrogen atom and benzyl.

[0840] Specific examples of the amide compound represented by formula (A) include the following compounds: [0841] (1-1) 3-((((9H-fluoren-9-yl)methoxy)carbonyl)(methyl)amino)-4-oxo-4-(piperidin-1-yl)butanoic acid, [0842] (1-2) allyl 3-((((9H-fluoren-9-yl)methoxy)carbonyl)(methyl)amino)-4-oxo-4-(piperidin-1-yl)butanoate, [0843] (1-3) tert-butyl 3-((((9H-fluoren-9-yl)methoxy)carbonyl)(methyl)amino)-4-oxo-4-(piperidin-1-yl)butanoate, [0844] (1-4) benzyl 3-((((9H-fluoren-9-yl)methoxy)carbonyl)(methyl)amino)-4-oxo-4-(piperidin-1-yl)butanoate, [0845] (1-5) 3-((tert-butoxycarbonyl)(methyl)amino)-4-oxo-4-(piperidin-1-yl)butanoic acid, [0846] (1-6) allyl 3-((tert-butoxycarbonyl)(methyl)amino)-4-oxo-4-(piperidin-1-yl)butanoate, [0847] (1-7) tert-butyl 3-((tert-butoxycarbonyl)(methyl)amino)-4-oxo-4-(piperidin-1-yl)butanoate, [0848] (1-8) benzyl 3-((tert-butoxycarbonyl)(methyl)amino)-4-oxo-4-(piperidin-1-yl)butanoate, [0849] (1-9) 3-(((benzyloxy)carbonyl)(methyl)amino)-4-oxo-4-(piperidin-1-yl)butanoic acid, [0850] (1-10) allyl 3-(((benzyloxy)carbonyl)(methyl)amino)-4-oxo-4-(piperidin-1-yl)butanoate, [0851] (1-11) tert-butyl 3-(((benzyloxy)carbonyl)(methyl)amino)-4-oxo-4-(piperidin-1-yl)butanoate, [0852] (1-12) benzyl 3-(((benzyloxy)carbonyl)(methyl)amino)-4-oxo-4-(piperidin-1-yl)butanoate, [0853] (1-13) 3-(((allyloxy)carbonyl)(methyl)amino)-4-oxo-4-(piperidin-1-yl)butanoic acid, [0854] (1-14) allyl 3-(((allyloxy)carbonyl)(methyl)amino)-4-oxo-4-(piperidin-1-yl)butanoate, [0855] (1-15) tert-butyl 3-(((allyloxy)carbonyl)(methyl)amino)-4-oxo-4-(piperidin-1-yl)butanoate, [0856] (1-16) benzyl 3-(((allyloxy)carbonyl)(methyl)amino)-4-oxo-4-(piperidin-1-yl)butanoate, [0857] (1-17) 3-(methyl((2-(trimethylsilyl)ethoxy)carbonyl)amino)-4-oxo-4-(piperidin-1-yl)butanoic acid, [0858] (1-18) allyl 3-(methyl((2-(trimethylsilyl)ethoxy)carbonyl)amino)-4-oxo-4-(piperidin-1-yl)butanoate, [0859] (1-19) tert-butyl 3-(methyl((2-(trimethylsilyl)ethoxy)carbonyl)amino)-4-oxo-4-(piperidin-1-yl)butanoate, [0860] (1-20) benzyl 3-(methyl((2-(trimethylsilyl)ethoxy)carbonyl)amino)-4-oxo-4-(piperidin-1-yl)butanoate, [0861] (2-1) 3-((((9H-fluoren-9-yl)methoxy)carbonyl)(methyl)amino)-4-morpholino-4-oxobutanoic acid, [0862] (2-2) allyl 3-((((9H-fluoren-9-yl)methoxy)carbonyl)(methyl)amino)-4-morpholino-4-oxobutanoate, [0863] (2-3) tert-butyl 3-((((9H-fluoren-9-yl)methoxy)carbonyl)(methyl)amino)-4-morpholino-4-oxobutanoate, [0864] (2-4) benzyl 3-((((9H-fluoren-9-yl)methoxy)carbonyl)(methyl)amino)-4-morpholino-4-oxobutanoate, [0865] (2-5) 3-((tert-butoxycarbonyl)(methyl)amino)-4-morpholino-4-oxobutanoic acid, [0866] (2-6) allyl 3-((tert-butoxycarbonyl)(methyl)amino)-4-morpholino-4-oxobutanoate, [0867] (2-7) tert-butyl 3-((tert-butoxycarbonyl)(methyl)amino)-4-morpholino-4-oxobutanoate, [0868] (2-8) benzyl 3-((tert-butoxycarbonyl)(methyl)amino)-4-morpholino-4-oxobutanoate, [0869] (2-9) 3-(((benzyloxy)carbonyl)(methyl)amino)-4-morpholino-4-oxobutanoic acid, [0870] (2-10) allyl 3-(((benzyloxy)carbonyl)(methyl)amino)-4-morpholino-4-oxobutanoate, [0871] (2-11) tert-butyl 3-(((benzyloxy)carbonyl)(methyl)amino)-4-morpholino-4-oxobutanoate, [0872] (2-12) benzyl 3-(((benzyloxy)carbonyl)(methyl)amino)-4-morpholino-4-oxobutanoate, [0873] (2-13) 3-(((allyloxy)carbonyl)(methyl)amino)-4-morpholino-4-oxobutanoic acid, [0874] (2-14) allyl 3-(((allyloxy)carbonyl)(methyl)amino)-4-morpholino-4-oxobutanoate, [0875] (2-15) tert-butyl 3-(((allyloxy)carbonyl)(methyl)amino)-4-morpholino-4-oxobutanoate, [0876] (2-16) benzyl 3-(((allyloxy)carbonyl)(methyl)amino)-4-morpholino-4-oxobutanoate, [0877] (2-17) 3-(methyl((2-(trimethylsilyl)ethoxy)carbonyl)amino)-4-morpholino-4-oxobutanoic acid, [0878] (2-18) allyl 3-(methyl((2-(trimethylsilyl)ethoxy)carbonyl)amino)-4-morpholino-4-oxobutanoate, [0879] (2-19) tert-butyl 3-(methyl((2-(trimethylsilyl)ethoxy)carbonyl)amino)-4-morpholino-4-oxobutanoate, [0880] (2-20) benzyl 3-(methyl((2-(trimethylsilyl)ethoxy)carbonyl)amino)-4-morpholino-4-oxobutanoate, [0881] (3-1) 3-((((9H-fluoren-9-yl)methoxy)carbonyl)(methyl)amino)-4-(dimethylamino)-4-oxobutanoic acid, [0882] (3-2) allyl 3-((((9H-fluoren-9-yl)methoxy)carbonyl)(methyl)amino)-4-(dimethylamino)-4-oxobutanoate, [0883] (3-3) tert-butyl 3-((((9H-fluoren-9-yl)methoxy)carbonyl)(methyl)amino)-4-(dimethylamino)-4-oxobutanoate, [0884] (3-4) benzyl 3-((((9H-fluoren-9-yl)methoxy)carbonyl)(methyl)amino)-4-(dimethylamino)-4-oxobutanoate, [0885] (3-5) 3-((tert-butoxycarbonyl)(methyl)amino)-4-(dimethylamino)-4-oxobutanoic acid, [0886] (3-6) allyl 3-((tert-butoxycarbonyl)(methyl)amino)-4-(dimethylamino)-4-oxobutanoate, [0887] (3-7) tert-butyl 3-((tert-butoxycarbonyl)(methyl)amino)-4-(dimethylamino)-4-oxobutanoate, [0888] (3-8) benzyl 3-((tert-butoxycarbonyl)(methyl)amino)-4-(dimethylamino)-4-oxobutanoate, [0889] (3-9) 3-(((benzyloxy)carbonyl)(methyl)amino)-4-(dimethylamino)-4-oxobutanoic acid, [0890] (3-10) allyl 3-(((benzyloxy)carbonyl)(methyl)amino)-4-(dimethylamino)-4-oxobutanoate, [0891] (3-11) tert-butyl 3-(((benzyloxy)carbonyl)(methyl)amino)-4-(dimethylamino)-4-oxobutanoate, [0892] (3-12) benzyl 3-(((benzyloxy)carbonyl)(methyl)amino)-4-(dimethylamino)-4-oxobutanoate, [0893] (3-13) 3-(((allyloxy)carbonyl)(methyl)amino)-4-(dimethylamino)-4-oxobutanoic acid, [0894] (3-14) allyl 3-(((allyloxy)carbonyl)(methyl)amino)-4-(dimethylamino)-4-oxobutanoate, [0895] (3-15) tert-butyl 3-(((allyloxy)carbonyl)(methyl)amino)-4-(dimethylamino)-4-oxobutanoate, [0896] (3-16) benzyl 3-(((allyloxy)carbonyl)(methyl)amino)-4-(dimethylamino)-4-oxobutanoate, [0897] (3-17) 4-(dimethylamino)-3-(methyl((2-(trimethylsilyl)ethoxy)carbonyl)amino)-4-oxobutanoic acid, [0898] (3-18) allyl 4-(dimethylamino)-3-(methyl((2-(trimethylsilyl)ethoxy)carbonyl)amino)-4-oxobutanoate, [0899] (3-19) tert-butyl 4-(dimethylamino)-3-(methyl((2-(trimethylsilyl)ethoxy)carbonyl)amino)-4-oxobutanoate, [0900] (3-20) benzyl 4-(dimethylamino)-3-(methyl((2-(trimethylsilyl)ethoxy)carbonyl)amino)-4-oxobutanoate, [0901] (4-1) 3-(methylamino)-4-oxo-4-(piperidin-1-yl)butanoic acid, [0902] (4-2) allyl 3-(methylamino)-4-oxo-4-(piperidin-1-yl)butanoate, [0903] (4-3) tert-butyl 3-(methylamino)-4-oxo-4-(piperidin-1-yl)butanoate, [0904] (4-4) benzyl 3-(methylamino)-4-oxo-4-(piperidin-1-yl)butanoate, [0905] (4-5) 3-(methylamino)-4-morpholino-4-oxobutanoic acid, [0906] (4-6) allyl 3-(methylamino)-4-morpholino-4-oxobutanoate, [0907] (4-7) tert-butyl 3-(methylamino)-4-morpholino-4-oxobutanoate, [0908] (4-8) benzyl 3-(methylamino)-4-morpholino-4-oxobutanoate, [0909] (4-9) 4-(dimethylamino)-3-(methylamino)-4-oxobutanoic acid, [0910] (4-10) allyl 4-(dimethylamino)-3-(methylamino)-4-oxobutanoate, [0911] (4-11) tert-butyl 4-(dimethylamino)-3-(methylamino)-4-oxobutanoate, or [0912] (4-12) benzyl 4-(dimethylamino)-3-(methylamino)-4-oxobutanoate.

[0913] The amide compound of the present invention represented by formula (A) can be synthesized according to the following scheme, for example.

##STR00069##

[0914] The amidation step in the above scheme can be achieved by condensing any amine ((R.sub.17A)(R.sub.17B)NH) with a carboxyl group using a dehydration condensation agent such as a carbodiimide compound according to Solid-Phase Synthesis Handbook (publisher: Merck K.K., date of publication: May 1, 2002) or the method of Albert et al. (Synthesis, 1987, 7, 635-637), for example.

[0915] The deprotection step in the above scheme can be achieved using the method described in Greene's “Protective Groups in Organic Synthesis” (5th ed., John Wiley & Sons 2014) or the deprotection method described herein, for example.

[0916] The compounds of the present invention may be either free forms or salts thereof, both of which are included in the present invention. Examples of such “salts” include inorganic acid salts, organic acid salts, inorganic base salts, organic base salts, and acidic or basic amino acid salts.

[0917] Examples of the inorganic acid salts include hydrochlorides, hydrobromides, sulfates, nitrates, and phosphates. Examples of the organic acid salts include acetates, succinates, fumarates, maleates, tartrates, citrates, lactates, stearates, benzoates, methanesulfonates, benzenesulfonates, and p-toluenesulfonates.

[0918] Examples of the inorganic base salts include alkali metal salts such as sodium salts and potassium salts, alkaline earth metal salts such as calcium salts and magnesium salts, aluminum salts, and ammonium salts. Examples of the organic base salts include diethylamine salts, diethanolamine salts, meglumine salts, and N,N-dibenzylethylenediamine salts.

[0919] Examples of the acidic amino acid salts include aspartates and glutamates. Examples of the basic amino acid salts include arginine salts, lysine salts, and ornithine salts.

[0920] The compounds of the present invention may absorb water to form hydrates when left in the air, for example. Such hydrates are also included in the present invention.

[0921] Further, the compounds of the present invention may absorb certain other solvents to form solvates. Such solvates are also included in the present invention.

[0922] Herein, although structural formulas of the compounds of the present invention may represent certain isomers for convenience, the present invention includes all isomers and mixtures of isomers possible in terms of compound structure, such as geometric isomers, optical isomers, and tautomers, and the compounds are not limited to the formulas described for convenience. For example, when the compounds have an asymmetric carbon atom in the molecule and exist as optically active isomers and racemates, both are included in the present invention.

[0923] All prior art references cited herein are incorporated by reference into this description.

EXAMPLE

[0924] The present invention will be further illustrated with reference to the following Examples but is not limited thereto.

[0925] Solvents such as methylene chloride, ethyl acetate, 2-MeTHF, dichloroethane, or DMF used in the practice of the present invention were those from commercial suppliers, used without purification. Solvents such as dehydrated solvents, ultradehydrated solvents, or anhydrous solvents used for reactions without addition of water as a solvent were those from commercial suppliers, used without purification.

[0926] Unless otherwise stated, silyl compounds such as TMSOTf used in the practice of the present invention, or reagents like electrophilic species scavengers such as BSA and BSTFA used in the practice of the present invention were those from commercial suppliers, used without purification.

[0927] Unless otherwise stated, starting peptide or amide compounds used in the practice of the present invention were those from commercial suppliers, used without purification. As necessary, such compounds were produced by known methods and used.

[0928] In the following examples, analysis was performed by any one or more of the HPLC methods 1 to 4 or methods A, B, C, D, E, F, G, or H.

Analysis: HPLC (reaction conversion rate, purity)

[0929] HPLC Method 1 [0930] Instrument: Waters ACQUITY UPLC H-Class [0931] Column: Ascentis Express C18 (2.7 μm, 4.6 mm×50 mm), Supelco [0932] Eluent: A) 0.05% TFA/water, B) 0.05% TFA/CH.sub.3CN [0933] Gradient (B): 5% (0 min.).Math.100% (6 min.).Math.5% (7 min.).Math.5% (9 min.) [0934] Flow rate: 1.0 mL/min. [0935] Detection: 210 nm [0936] Injection vol.: 5 μL [0937] Sample prep.: 5 μL/1.00 mL

[0938] HPLC Method 2 [0939] Instrument: Waters LCT Premier [0940] Column: Ascentis Express C18 (2.7 μm, 4.6 mm×50 mm), Supelco [0941] Column temp.: 35 deg. [0942] Eluent: A) 0.05% TFA/water, B) 0.05% TFA/CH3CN [0943] Gradient (B): 50% (0 min.).Math.100% (6 to 11 min.).Math.50% (11 min.).Math.50% (13 min.) [0944] Flow rate: 1.0 mL/min. [0945] Detection: 210 nm [0946] Injection vol.: 5 μL [0947] Sample prep.: 5 μL/1.00 mL

[0948] HPLC Method 3 [0949] Instrument: Waters ACQUITY UPLC H-Class [0950] Column: Ascentis Express C18, (2.7 μm, 2.1 mm×50 mm), Supelco [0951] Column temp.: 35 deg. [0952] Eluent: A) 0.05% TFA/water, B) 0.05% TFA/CH3CN [0953] Gradient (A): 95%(0 min).fwdarw.0%(4.0 min).fwdarw.0%(4.5 min).fwdarw.95%(4.6 min).fwdarw.95%(6 min) [0954] Flow rate: 0.25 mL/min [0955] Detection: PDA 210 nm (200-400 nm PDA total) [0956] Injection vol.: 0.3 μL [0957] Sample prep.: 50 μL/1.00 mL

[0958] HPLC Method 4 [0959] Instrument: Waters ACQUITY UPLC H-Class [0960] Column: CAPCELL CORE ADME, (2.7 μm, 3.0 mm×150 mm) [0961] Column temp.: 30 deg. [0962] Solvents: A) 0.05% TFA/water, B) 0.05% TFA/CH3CN [0963] Gradient (A): 70%(0 min).fwdarw.30%(20.0 min).fwdarw.0%(20.1 min).fwdarw.0%(22.0 min).fwdarw.70%(22.1 min).fwdarw.70%(24 min) [0964] Flow rate: 0.3 mL/min [0965] Detection: PDA 254 nm (200-400 nm PDA total) [0966] Injection vol.: 0.3 μL [0967] Sample prep.: amorphous 10.0 mg in CH.sub.3CN 10 ml; this solution 0.3 mL/0.7 mL CH.sub.3CN

A. Boc Removal Reaction Experiments

Example 1

[0968] Boc Removal Reaction of Boc-MeAla-MePhe-Leu-MeLeu-Thr(OBn)-MeGly-OAllyl (Compound 3a) (6-mer: TfOH-BSTFA Method)

##STR00070##

[0969] 77.7 mg of the raw material was weighed into a reaction vessel and dissolved in 5 v/w of ethyl acetate, and the reaction vessel was cooled with ice. After 10 minutes, 0.055 ml (2.4 eq) of BSTFA and 3.0 μl (0.4 eq) of TfOH were sequentially added under a nitrogen atmosphere, and the reaction solution was stirred. Three hours after the addition of the reagents, the reaction was analyzed by LCMS and the raw material was confirmed to disappear.

[0970] The reaction was quenched with saturated aqueous sodium bicarbonate. The organic layer was washed with saturated aqueous sodium bicarbonate and 5% saline. The resulting organic layer was concentrated under reduced pressure to give 77.3 mg of a deprotected product quantitatively.

[0971] Amide bond cleavage was not confirmed.

TABLE-US-00001 TABLE 1 Weight (mg) Yield (%) Raw material (Compound 3a) 77.7 — Product (Compound 3b) 77.3 Quantitative

TABLE-US-00002 TABLE 2 Analysis (HPLC method 1) MW m/z rt Purity LC A % Raw material 906.55 807.55 5.976 93.83 (Compound 3a) ( [M − Boc + H]+) Product 806.49 807.49 ( [M + H]+) 3.968 94.36 (Compound 3b)

Example 2

[0972] Boc Removal Reaction of Boc-MeIle-Ala-MePhe-MeVal-Asp(pip)-MeAla-MePhe-Leu-MeLeu-Thr(OBn)-MeGly-OAllyl (Compound 1a) (11-mer: TMSOTf-HMDS Method)

##STR00071##

[0973] 0.0810 g of the raw material was weighed into a reaction vessel and dissolved in 10 v/w of ethyl acetate, and the reaction vessel was cooled with ice. After 10 minutes, 0.079 ml (7.2 eq) of HMDS and 34 μl (3.6 eq) of TMSOTf were sequentially added under a nitrogen atmosphere, and the reaction solution was stirred. Four hours after the addition of the reagents, the reaction was analyzed by LCMS and the raw material was confirmed to disappear. Amide bond cleavage was not confirmed.

[0974] After confirming the disappearance of the raw material, the reaction solution was stirred for another one hour and the reaction was analyzed by LCMS. Also at this time, amide bond cleavage was not confirmed.

[0975] To the reaction vessel was added 1 ml of brine to quench the reaction. 5 ml of ethyl acetate was then added and the two layers were separated. The organic layer was washed with a mixture of 1 ml of saturated aqueous sodium bicarbonate and 1 ml of brine. The resulting organic layer was concentrated to give 68.0 mg of a deprotected product in 90% yield as a transparent film.

TABLE-US-00003 TABLE 3 Weight (mg) Yield (%) Raw material (Compound 1a) 81.0 — Product (Compound 1b) 68.0 90

TABLE-US-00004 TABLE 4 Analysis (HPLC method 1: Determination of purities of the raw material and target product) Purity MW m/z rt LC A % Raw material (Compound 1a) 1560.96 1242.05 6.519 96.680 Product (Compound 1b) 1460.90  731.78 4.635 98.003

TABLE-US-00005 TABLE 5 Analysis (HPLC method 2: Mass spectroscopy) MW m/z rt Raw material (Compound 1a) 1560.96 1561.4097 ( [M + H]+) 5.883 Product (Compound 1b) 1460.90 1461.5303 ( [M + H]+) 4.817

Example 3

[0976] Boc Removal Reaction of Boc-MeIle-Ala-MePhe-MeVal-Asp(pip)-MeAla-MePhe-Leu-MeLeu-Thr(OBn)-MeGly-OAllyl (Compound 1a) (11-mer: TMSOTf-HMDS Method)

##STR00072##

[0977] The reaction was carried out according to Example 2. 19 hours after the reaction was started, the reaction was analyzed by LCMS. However, decomposition of the product was not confirmed.

TABLE-US-00006 TABLE 6 Weight (mg) Yield (%) Raw material (Compound 1a) 21.7 — Product (Compound 1b) Not isolated Not isolated

TABLE-US-00007 TABLE 7 Analysis (HPLC method 1: Determination of purities of the raw material and target product) MW m/z rt Purity LC A % Raw material (Compound 1a) 1560.96 1241.89 6.476 98.408 Product (Compound 1b) 1460.90  731.76 4.562 97.256

Example 4

[0978] Boc Removal Reaction of Boc-MeAla-MePhe-Leu-MeLeu-Thr(OBn)-MeGly-MeIle-Ala-MePhe-MeVal-Asp(OBn)-pip (Compound 2a) (11-mer: TfOH-BSTFA Method)

##STR00073##

[0979] The reaction was carried out according to Example 1.

TABLE-US-00008 TABLE 9 Weight (mg) Yield (%) Raw material (Compound 2a) 48.1 — Product (Compound 2b) Not isolated Not isolated

TABLE-US-00009 TABLE 10 Analysis (HPLC method 1: Determination of purities of the raw material and target product) MW m/z rt Purity LC A % Raw material (Compound 2a) 1610.97 ND 6.492 84.00 Product (Compound 2b) 1510.92 ND 4.776 81.54

TABLE-US-00010 TABLE 11 Analysis (HPLC method 2: Mass spectroscopy) MW m/z rt Raw material (Compound 2a) 1610.97 1611.43 ( [M + H]+) 5.96 Product (Compound 2b) 1510.92 1511.53 ( [M + H]+) 2.84

Example 5

[0980] Boc Removal Reaction of Boc-MePhe-Leu-MeLeu-Thr(OBn)-MeGly-OAllyl (Compound 4a) (5-mer: TfOH-BSTFA Method)

##STR00074##

[0981] The reaction was carried out according to Example 1.

TABLE-US-00011 TABLE 12 Weight (mg) Yield (%) Raw material (Compound 4a) 98.5 — Product (Compound 4b) 78.7 91

TABLE-US-00012 TABLE 13 Analysis (HPLC method 1) Purity LC MW m/z rt Area % Raw material 821.49 844.4 ( [M + Na]+) 6.033 90.51 (Compound 4a) Product (Compound 4b) 721.44 722.46 ( [M + H]+) 3.879 93.30

Example 6

[0982] Reactions for Boc Removal of Boc-MeLeu-Thr(OBn)-MeGly-OAllyl (Compound 8a) and its elongation to form Boc-Leu-MeLeu-Thr(OBn)-MeGly-OAllyl (3 mer: TfOH-BSTFA Method)

##STR00075##

[0983] 73.3 mg of the starting material was weighed into a reaction vessel and dissolved in 5 v/w of isopropyl acetate. After cooling the reaction vessel with ice, 0.085 ml of BSTFA and 0.0047 ml of TfOH were added and the reaction solution was stirred for two hours. After confirming by LC that deprotection was completed without main chain cleavage, 5 v/w of water and 186.9 mg of dipotassium hydrogenphosphate were added. 40.1 mg of Boc-Leu-OH monohydrate and 63.1 mg of DMT-MM were then added and the reaction solution was stirred with ice-cooling. 17 hours after the addition of the reagents, completion of the reaction was confirmed by LC. The reaction was quenched by adding 5 v/w of a 1 N aqueous sodium hydroxide solution. The organic layer was separated by liquid separation treatment, then washed once with a 1 N aqueous sodium hydroxide solution, twice with a 5% aqueous potassium bisulfate solution, and once with a 5% aqueous sodium chloride solution, and subsequently concentrated under reduced pressure. 82.0 mg (93% yield) of a crude product was obtained as an oily liquid.

TABLE-US-00013 TABLE 14 Weight (mg) Yield (%) Raw material (Compound 8a) 73.3 — Product (Compound 7a) 82.0 93

TABLE-US-00014 TABLE 15 Analysis of deprotection reaction (HPLC method 1) Purity MW m/z rt LC A % Raw material 547.33 448.30 5.513 96.17 (Compound 8a) ( [M − Boc + H]+) Boc-removed product 447.27 ND 2.938 95.95

TABLE-US-00015 TABLE 16 Analysis of elongation reaction (HPLC method 1) MW m/z rt Purity LC A % Product (Compound 8b) 660.85 561.28 5.657 95.36 ( [M − Boc + H]+)

[0984] Reactions for Teoc removal of Teoc-MeLeu-Thr(OBn)-MeGly-OAllyl (Compound 80b) and its elongation to form Boc-Leu-MeLeu-Thr(OBn)-MeGly-OAllyl (7a) (3-mer: TfOH-BSTFA Method)

##STR00076##

[0985] Compound 7a can be obtained without main chain cleavage by removing the Teoc group in the same operation as in Example 6 and then reacting with Boc-Leu-OH.

Example 7

[0986] Boc Removal Reaction of Boc-MeIle-Ala-MePhe-MeVal-Asp(OBn)-pip (Compound 5a) (5-mer: TfOH-BSTFA Method)

##STR00077##

[0987] Deprotection and elongation reactions were performed in a one-pot operation according to the experimental method described in Example 6.

TABLE-US-00016 TABLE 17 Weight (mg) Yield (%) Raw material (Compound 5a) 105.3 — Product (Compound 2a) 153.6 78%

TABLE-US-00017 TABLE 18 Analysis of deprotection reaction (HPLC method 1) MW m/z rt Purity LC A % Raw material 862.52 573.33 5.978 95.01 (Compound 5a) Deprotected 762.47 763.47 ([M + H]+) 3.779 94.05 product (Compound 5b)

TABLE-US-00018 TABLE 19 Analysis of the product after elongation (HPLC method 1: Determination of purities of the raw material and target product) MW m/z rt Purity LC A % Product (Compound 2a) 1610.97 ND 6.699 85.93

TABLE-US-00019 TABLE 20 Analysis of the product after elongation (HPLC method 2: Mass spectroscopy) MW m/z rt Product 1610.97 1611.43 ([M + H]+) 5.96 (Compound 2a)

Example 8

[0988] Boc Removal and Elongation Reactions of Boc-Ala-MePhe-MeVal-Asp(OBn)-pip (Compound 6a) (4-mer: TfOH-BSTFA Method)

##STR00078##

[0989] Deprotection and elongation reactions were performed in a one-pot operation according to the experimental method described in Example 6.

TABLE-US-00020 TABLE 21 Weight (mg) Yield (%) Raw material (Compound 6a) 277 — Product (Compound 5a) 308.9 95%

TABLE-US-00021 TABLE 22 Analysis of the deprotected product (HPLC method 1) MW m/z rt Purity LC A % Raw material 735.42  758.4 ([M + Na]+) 5.357 98.921 (Compound 6a) Deprotected 635.37 636.40 ([M + H]+) 3.383 97.893 product (Compound 6b)

TABLE-US-00022 TABLE 23 Analysis after elongation reaction (HPLC method 1) MW m/z rt Purity LC A % Product (Compound 5a) 862.52 573.33 5.978 96.938

Example 9

[0990] Boc Removal Reaction of Boc-Leu-MeLeu-Thr(OBn)-MeGly-OAllyl (Compound 7a) (4-mer: TfOH-BSTFA Method)

##STR00079##

[0991] Deprotection and elongation reactions were performed in a one-pot operation according to the experimental method described in Example 6.

TABLE-US-00023 TABLE 24 Weight (mg) Yield (%) Raw material (Compound 7a) 82.0 — Product (Compound 4a) 98.5 97

TABLE-US-00024 TABLE 25 Analysis (HPLC method 1) MW m/z rt Purity LC A % Raw material 660.41 561.28 ([M − 5.657 95.36 (Compound 7a) 100 + H]+) Intermediate 560.36 561.40 ([M + H]+) 3.545 94.04 (Compound 7b)

TABLE-US-00025 TABLE 26 Analysis (HPLC method 1) MW m/z rt Purity LC A % Product 821.49 844.4 ([M + Na]+) 6.034 90.36 (Compound 4a)

Example 10

[0992] Boc Removal Reaction of Boc-Leu-MeLeu-Thr(OBn)-MeGly-OAllyl (Compound 7a) Under Known Conditions (4-Mer: HCl Method) (Russ. J. Bioorg. Chem., 2016, 42, 143.)

##STR00080##

[0993] 18.7 mg of the raw material was weighed into a reaction vessel and dissolved in 10 v/w of trifluoroethanol. 8.5 μl (1.2 eq) of 4 N HCl (ethyl acetate solution) was then added and the reaction solution was stirred. Two hours after the addition of the reagent, the reaction was analyzed by LCMS (HPLC method 1) and the conversion rate (=target product/(target product+starting material)) was confirmed to be 81%. At this time, 0.861% of H-Thr(OBn)-MeGly-OAllyl (Compound 7c) and 1.275% of H-Leu-MeLeu-Thr(OBn)-OH (Compound 7d) were respectively detected as compounds in which amide bonds were cleaved.

TABLE-US-00026 TABLE 27 Analysis (HPLC method 1) MW m/z rt LC A % Raw material 660.41 561.28 ([M − 5.657 18.437 (Compound 7a) Boc + H]+) Product 560.36 561.40 ([M + H]+) 3.363 79.258 (Compound 7b) Cleaved 320.17 321.08 ([M + H]+) 2.500 0.861% product (Compound 7c) Cleaved 449.29 450.17 ([M + H]+) 2.759 1.275% product (Compound 7d)

Example 11

[0994] Reactions for Deprotection of Boc-MePhe-MeVal-Asp(OBn)-pip (Compound 9a) and its Elongation to Form Boc-Ala-MePhe-MeVal-Asp(OBn)-pip (3-mer: TMSOTf-BSTFA Method)

##STR00081##

[0995] Deprotection and elongation reactions were performed in a one-pot operation according to the experimental method described in Example 6.

TABLE-US-00027 TABLE 28 Weight (mg) Yield (%) Raw material (Compound 9a) 253.2 — Product (Compound 6a) 319.4 Quantitative

TABLE-US-00028 TABLE 29 Analysis of deprotection reaction (HPLC method 1) MW m/z rt Purity LC A % Raw material 664.38  687.5 ([M + Na]+) 5.647 99.206 (Compound 9a) Deprotected 564.33 565.31 ([M + H]+) 3.338 98.885 product

TABLE-US-00029 TABLE 30 Analysis of elongation reaction (HPLC method 1) MW m/z rt Purity LC A % Product 735.92 758.4 ([M + Na]+) 5.377 99.538 (Compound 6a)

Example 12

[0996] Deprotection of Boc-MePhe-MeVal-Asp(OBn)-pip (Compound 9a) (3-Mer: TMSOTf-MSTFA Method)

##STR00082##

[0997] Deprotection and elongation reactions were performed in a one-pot operation according to the experimental method described in Example 6.

TABLE-US-00030 TABLE 31 Weight (mg) Yield (%) Raw material (Compound 9a) 200.6 — Product (Compound 6a) 220.6 95

TABLE-US-00031 TABLE 32 Analysis of deprotection reaction (HPLC method 1) MW m/z rt Purity LC A % Raw material 664.38  687.5 ([M + Na]+) 5.647 99.206 (Compound 9a) Deprotected 564.33 565.31 ([M + H]+) 3.338 97.921 product (Compound 9b)

TABLE-US-00032 TABLE 33 Analysis of elongation reaction (HPLC method 1) MW m/z rt Purity LC A % Product 735.92 758.4 ([M + Na]+) 5.308 99.279 (Compound 6a)

Example 13

[0998] Deprotection of Boc-MePhe-MeVal-Asp(OBn)-pip (Compound 9a) (3-mer: TMSOTf-BSA Method, TMSOTf 2.4 Eq, BSA 2.4 Eq)

##STR00083##

[0999] Deprotection and elongation reactions were performed in a one-pot operation according to the experimental method described in Example 6.

TABLE-US-00033 TABLE 34 Weight (mg) Yield (%) Raw material (Compound 9a) 204.6 — Product (Compound 6a) 219.0 97

TABLE-US-00034 TABLE 35 Analysis of deprotection reaction (HPLC method 1) MW m/z rt Purity LC A % Raw material 664.38  687.5 ([M + Na]+) 5.647 99.206 (Compound 9a) Deprotected 564.33 565.31 ([M + H]+) 3.338 98.970 product (Compound 9b)

TABLE-US-00035 TABLE 36 Analysis of elongation reaction (HPLC method 1) MW m/z rt Purity LC A % Product 735.92 758.5 ([M + Na]+) 5.301 99.573 (Compound 6a)

Example 14

[1000] Deprotection of Boc-MePhe-MeVal-Asp(OBn)-pip (Compound 9a) (3-mer: TMSOTf-HMDS Method)

##STR00084##

[1001] 107.4 mg of the raw material was weighed into a reaction vessel and dissolved in 5 v/w of ethyl acetate, and the reaction vessel was cooled with ice. After 10 minutes, 0.085 ml (2.4 eq) of HMDS (1,1,1,3,3,3-hexamethyldisilazane) and 35 μl (1.2 eq) of TMSOTf were sequentially added under a nitrogen atmosphere, and the reaction solution was stirred. Two hours after the addition of the reagents, the reaction was analyzed by LCMS to find that the conversion rate was 55.8%. Therefore, 20 μl (0.69 eq) of TMOSTf was further added. One hour after the further addition of the reagent, the raw material was confirmed to disappear by LCMS. Amide bond cleavage was not confirmed.

[1002] To the reaction vessel was added 373.4 mg (13.3 eq) of dipotassium hydrogenphosphate, after which 0.89 ml of water was added and the reaction solution was stirred for 20 minutes.

[1003] To the reaction vessel was then added 3 ml of ethyl acetate, and the two layers were separated. The organic layer was washed with a mixture of 1 ml of saturated aqueous sodium bicarbonate and 1 ml of brine. The resulting organic layer was concentrated.

[1004] The resulting oily liquid was then dissolved in 1 ml of ethyl acetate, and 251.0 mg of dipotassium hydrogenphosphate was added. After adding 0.89 ml of water, the reaction vessel was cooled with ice. 36.8 mg of Boc-Ala-OH and 79.5 mg DMT-MM were added, and the reaction solution was stirred overnight in an ice bath. After 15.5 hours, the raw material was confirmed to disappear by LCMS and liquid separation treatment was then conducted. The resulting organic layer was washed twice with 5% potassium carbonate, once with water, twice with 5% potassium bisulfate, and once with brine, and concentrated. 0.1075 g (90% yield) of a crude product was obtained as a white solid.

TABLE-US-00036 TABLE 37 Weight (mg) Yield (%) Raw material (Compound 9a) 107.4 — Product (Compound 6a) 107.5 90

TABLE-US-00037 TABLE 38 Analysis of deprotection reaction (HPLC method 1) MW m/z rt Purity LC A % Raw material 664.38  687.5 ([M + Na]+) 5.647 99.206 (Compound 9a) Intermediate 564.33 565.31 ([M + H]+) 3.360 98.783 (Compound 9b)

TABLE-US-00038 TABLE 39 Analysis of elongation reaction (HPLC method 1) MW m/z rt Purity LC A % Product 735.42 758.4 ([M + Na]+) 5.268 98.031 (Compound 9a)
B. Screening of electrophilic species scavengers

Example 15

[1005] Deprotection of Boc-MePhe-MeVal-Asp(OBn)-pip (Compound 9a) (Base: N-(Trimethylsilyl)diethylamine)

##STR00085##

[1006] 99.5 mg of the raw material was weighed into a reaction vessel and dissolved in 5 v/w of ethyl acetate, and the reaction vessel was cooled with ice. 0.070 ml (2.4 eq) of N-((trimethylsilyl)diethylamine) and 0.032 ml (1.2 eq) of TMSOTf were sequentially added under a nitrogen atmosphere, and the reaction solution was stirred. 1.5 hours after the addition of the reagents, the reaction was analyzed by LCMS to find that the conversion rate was 30%. Therefore, 0.029 ml (1.1 eq) of TMOSTf was further added. Four hours after the start of the reaction, the reaction was analyzed by LCMS and the raw material was confirmed to disappear. At this time, amide bond cleavage was not confirmed. The reaction solution was stirred overnight, and 20 hours after the start of the reaction, the reaction was analyzed again by LCMS. Also at this time, amide bond cleavage was not confirmed. 21 hours after the start of the reaction, 0.2032 g of dipotassium hydrogenphosphate and 0.50 mL of water were added to the reaction solution which was then stirred for 30 min.

[1007] The reaction solution was diluted with 4 ml of ethyl acetate, and the two layers were then separated to provide an organic layer. The resulting organic layer was washed twice with a mixed solvent of 0.5 ml of brine and 0.5 ml of saturated aqueous sodium bicarbonate. The organic layer was then washed with 0.5 ml of brine and concentrated. The residue was dissolved in 4 ml of isopropyl acetate and washed twice with a mixed solvent of 1 ml of 0.5 M aqueous sodium hydroxide and 0.5 ml of brine. Then, the organic layer was washed with 0.5 ml of 10% saline and concentrated under reduced pressure to give 81.7 mg of a deprotected product in 97% yield.

TABLE-US-00039 TABLE 40 Weight (mg) Yield (%) Raw material (Compound 9a) 99.5 — Product (Compound 9b) 81.7 97

TABLE-US-00040 TABLE 41 Analysis (HPLC method 1) MW m/z rt Purity LC A % Raw material 664.38  687.5 ([M + Na]+) 5.647 99.206 (Compound 9a) Product 564.33 565.27 ([M + H]+) 3.364 95.547 (Compound 9b)

Example 16

[1008] Deprotection of Boc-MePhe-MeVal-Asp(OBn)-pip (Compound 9a)

[1009] (Base: N-(Trimethylsilyl)morpholine)

##STR00086##

[1010] The experiment was performed according to the method described in Example 15.

TABLE-US-00041 TABLE 42 Weight (mg) Yield (%) Raw material (Compound 9a) 98.4 — Product (Compound 9b) 117.8 Quantitative

TABLE-US-00042 TABLE 43 Analysis (HPLC method 1) MW m/z rt Purity LC A % Raw material 664.38  687.5 ([M + Na]+) 5.647 99.206 (Compound 9a) Product 564.33 565.30 ([M + H]+) 3.350 98.652 (Compound 9b)

Example 17

[1011] Deprotection of Boc-MePhe-MeVal-Asp(OBn)-pip (Compound 9a)

[1012] (Base: N-tert-Butyltrimethylsilylamine)

##STR00087##

[1013] The experiment was performed according to the method described in Example 15.

TABLE-US-00043 TABLE 44 Weight (mg) Yield (%) Raw material (Compound 9a) 100.9 — Product (Compound 9b) 128.6 Quantitative

TABLE-US-00044 TABLE 45 Analysis (HPLC method 1) MW m/z rt Purity LC A % Raw material 664.38  687.5 ([M + Na]+) 5.647 99.206 (Compound 9a) Product 564.33 565.28 ([M + H]+) 3.354 98.631 (Compound 9b)

Example 18

[1014] Deprotection of Boc-MePhe-MeVal-Asp(OBn)-pip (Compound 9a)

[1015] (Base: 2,2,4,4-Tetramethylpentanoneimine)

##STR00088##

[1016] The experiment was performed according to the method described in Example 15.

TABLE-US-00045 TABLE 46 Weight (mg) Yield (%) Raw material (Compound 9a) 88.0 — Product (Compound 9b) 102.5 Quantitative

TABLE-US-00046 TABLE 47 Analysis (HPLC method 1) MW m/z rt Purity LC A % Raw material 664.38  687.5 ([M + Na]+) 5.647 99.206 (Compound 9a) Product 564.33 565.28 ([M + H]+) 3.338 98.433 (Compound 9b)

Example 19

[1017] Deprotection of Boc-MePhe-MeVal-Asp(OBn)-pip (Compound 9a)

[1018] (Base: Isopropenyloxytrimethylsilane)

##STR00089##

[1019] The experiment was performed according to the method described in Example 15.

TABLE-US-00047 TABLE 48 Weight (mg) Yield (%) Raw material (Compound 9a) 44.3 — Product (Compound 9b) 44.2 Quantitative

TABLE-US-00048 TABLE 49 Analysis (HPLC method 1) MW m/z rt Purity LC A % Raw material 664.38  687.5 ([M + Na]+) 5.647 99.206 (Compound 9a) Product 564.33 565.27 ([M + H]+) 3.340 97.587 (Compound 9b)

Example 20

[1020] Deprotection of Boc-MePhe-MeVal-Asp(OBn)-pip (Compound 9a)

[1021] (Base: Dimethylketene methyl trimethylsilyl acetal)

##STR00090##

[1022] The experiment was performed according to the method described in Example 15.

TABLE-US-00049 TABLE 50 Weight (mg) Yield (%) Raw material (Compound 9a) 44.9 — Product (Compound 9b) 44.7 Quantitative

TABLE-US-00050 TABLE 51 Analysis (HPLC method 1) MW m/z rt Purity LC A % Raw material 664.38  687.5 ([M + Na]+) 5.647 99.206 (Compound 9a) Product 564.33 565.27 ([M + H]+) 3.348 98.352 (Compound 9b)

Example 21

[1023] Deprotection of Boc-MePhe-MeVal-Asp(OBn)-pip (Compound 9a)

[1024] (Base: 3,4-Dihydro-2H-pyran)

##STR00091##

[1025] The experiment was performed according to the method described in Example 15.

TABLE-US-00051 TABLE 52 Weight (mg) Yield (%) Raw material (Compound 9a) 53.7 — Product (Compound 9b) 69.2 Quantitative

TABLE-US-00052 TABLE 53 Analysis (HPLC method 1) MW m/z rt Purity LC A % Raw material 664.38  687.5 ([M + Na]+) 5.647 99.206 (Compound 9a) Product 564.33 565.27 ([M + H]+) 3.325 98.503 (Compound 9b)

Example 22

[1026] Deprotection of Boc-MePhe-MeVal-Asp(OBn)-pip (Compound 9a)

[1027] (Base: N,N,N′,N′-Tetramethyl-1,8-naphthalenediamine)

##STR00092##

[1028] The experiment was performed according to the method described in Example 15.

TABLE-US-00053 TABLE 54 Weight (mg) Yield (%) Raw material (Compound 9a) 50.6 — Product (Compound 9b) 57.6 Quantitative

TABLE-US-00054 TABLE 55 Analysis (HPLC method 1) MW m/z rt Purity LC A % Raw material 664.38  687.5 ([M + Na]+) 5.647 99.206 (Compound 9a) Product 564.33 565.28 ([M + H]+) 3.353 98.409 (Compound 9b)

[1029] (LCA % of the product described above is a value obtained by excluding the base remaining after post-treatment.)

Example 23

[1030] Deprotection of Boc-MePhe-MeVal-Asp(OBn)-pip (Compound 9a)

[1031] (Base: 2,2′-Isopropylidenebis[(4S)-4-tert-butyl-2-oxazoline])

##STR00093##

[1032] The experiment was performed according to the method described in Example 15.

TABLE-US-00055 TABLE 56 Weight (mg) Yield (%) Raw material (Compound 9a) 45.7 — Product (Compound 9b) 128.5 Quantitative

TABLE-US-00056 TABLE 57 Analysis (HPLC method 1) MW m/z rt LC A % Raw material 664.38  687.5 ([M + Na]+) 5.647 99.206 (Compound 9a) Product 564.33 565.28 ([M + H]+) 3.353 90.099 (Compound 9b)

[1033] (LCA % of the product described above is a value obtained by excluding the base remaining after post-treatment.)

Example 24

[1034] Deprotection of Boc-MePhe-MeVal-Asp(OBn)-pip (Compound 9a)

[1035] (Base: Diisopropylcarbodiimide)

##STR00094##

[1036] The experiment was performed according to the method described in Example 15.

TABLE-US-00057 TABLE 58 Weight (mg) Yield (%) Raw material (Compound 9a) 36.8 — Product (Compound 9b) 31.2 85

TABLE-US-00058 TABLE 59 Analysis (HPLC method 1) MW m/z rt LC A % Raw material 664.38  687.5 ([M + Na]+) 5.647 99.206 (Compound 9a) Product 564.33 565.28 ([M + H]+) 3.347 89.920 (Compound 9b)

[1037] (LCA % of the product described above is a value obtained by excluding the base remaining after post-treatment.)

C. Screening of Electrophilic Species Scavengers (Experiments in 11-Mer Peptides)

Example 25

[1038] Boc Removal Reaction of Boc-MeIle-Ala-MePhe-MeVal-Asp(pip)-MeAla-MePhe-Leu-MeLeu-Thr(OBn)-MeGly-OAllyl (Compound 1a) (11-mer: TMSOTf-BSTFA Method)

##STR00095##

[1039] The experiment was performed according to Example 2.

TABLE-US-00059 TABLE 60 Weight (mg) Yield (%) Raw material (Compound 1a) 19.7 — Product (Compound 1b) 14.8 80

TABLE-US-00060 TABLE 61 Analysis (HPLC method 1: Determination of purities of the raw material and target product) MW m/z rt Purity LC A % Raw material (Compound 1a) 1560.96 1242.05 6.519 96.680 Product (Compound 1b) 1460.90 731.67 4.638 96.275

Example 26

[1040] Boc Removal Reaction of Boc-MeIle-Ala-MePhe-MeVal-Asp(pip)-MeAla-MePhe-Leu-MeLeu-Thr(OBn)-MeGly-OAllyl (Compound 1a) (11-mer: TMSOTf-BSA Method)

##STR00096##

[1041] The experiment was performed according to Example 2.

TABLE-US-00061 TABLE 62 Weight (mg) Yield (%) Raw material (Compound 1a) 18.0 — Product (Compound 1b) 11.8 70

TABLE-US-00062 TABLE 63 Analysis (HPLC method 1: Determination of purities of the raw material and target product) MW m/z rt Purity LC A % Raw material (Compound 1a) 1560.96 1242.05 6.519 96.680 Product (Compound 1b) 1460.90 731.67 4.633 94.636

Example 27

[1042] Boc Removal Reaction of Boc-MeIle-Ala-MePhe-MeVal-Asp(pip)-MeAla-MePhe-Leu-MeLeu-Thr(OBn)-MeGly-OAllyl (Compound 1a) (11-mer: TMSOTf-silylamine Method)

##STR00097##

[1043] The experiment was performed according to Example 2.

TABLE-US-00063 TABLE 64 Weight (mg) Yield (%) Raw material (Compound 1a) 19.3 — Product (Compound 1b) 16.1 89

TABLE-US-00064 TABLE 65 Analysis (HPLC method 1: Determination of purities of the raw material and target product) MW m/z rt Purity LC A % Raw material (Compound 1a) 1560.96 1242.05 6.519 96.680 Product (Compound 1b) 1460.90 731.68 4.627 96.267

Example 28

[1044] Boc Removal Reaction of Boc-MeIle-Ala-MePhe-MeVal-Asp(pip)-MeAla-MePhe-Leu-MeLeu-Thr(OBn)-MeGly-OAllyl (Compound 1a) (11-mer: TMSOTf-silylamine Method)

##STR00098##

[1045] The experiment was performed according to Example 2.

TABLE-US-00065 TABLE 66 Weight (mg) Yield (%) Raw material (Compound 1a) 20.2 — Product (Compound 1b) 15.3 81

TABLE-US-00066 TABLE 67 Analysis (HPLC method 1: Determination of purities of the raw material and target product) MW m/z rt Purity LC A % Raw material (Compound 1a) 1560.96 1242.05 6.519 96.680 Product (Compound 1b) 1460.90 731.65 4.626 97.574

Example 29

[1046] Boc Removal Reaction of Boc-MeIle-Ala-MePhe-MeVal-Asp(pip)-MeAla-MePhe-Leu-MeLeu-Thr(OBn)-MeGly-OAllyl (Compound 1a) (11-mer: TMSOTf-imine Method)

##STR00099##

[1047] The experiment was performed according to Example 2.

TABLE-US-00067 TABLE 68 Weight (mg) Yield (%) Raw material (Compound 1a) 18.7 — Product (Compound 1b) 15.7 90

TABLE-US-00068 TABLE 69 Analysis (HPLC method 1: Determination of purities of the raw material and target product) MW m/z rt Purity LC A % Raw material (Compound 1a) 1560.96 1242.05 6.519 96.680 Product (Compound 1b) 1460.90 731.70 4.631 97.089

Example 30

[1048] Boc Removal Reaction of Boc-MeIle-Ala-MePhe-MeVal-Asp(pip)-MeAla-MePhe-Leu-MeLeu-Thr(OBn)-MeGly-OAllyl (Compound 1a) (11-mer: TMSOTf-MSTFA Method)

##STR00100##

[1049] The experiment was performed according to Example 2.

TABLE-US-00069 TABLE 70 Weight (mg) Yield (%) Raw material (Compound 1a) 21.7 — Product (Compound 1b) 15.7 77

TABLE-US-00070 TABLE 71 Analysis (HPLC method 1: Determination of purities of the raw material and target product) MW m/z rt Purity LC A % Raw material (Compound 1a) 1560.96 1242.05 6.519 96.680 Product (Compound 1b) 1460.90 731.61 4.616 96.490

Example 31

[1050] Boc Removal Reaction of Boc-MeIle-Ala-MePhe-MeVal-Asp(pip)-MeAla-MePhe-Leu-MeLeu-Thr(OBn)-MeGly-OAllyl (Compound 1a) (11-mer: TMSOTf-silylamine Method)

##STR00101##

[1051] The experiment was performed according to Example 2.

TABLE-US-00071 TABLE 72 Weight (mg) Yield (%) Raw material (Compound 1a) 17.5 — Product (Compound 1b) 13.5 81

TABLE-US-00072 TABLE 73 Analysis (HPLC method 1: Determination of purities of the raw material and target product) MW m/z rt Purity LC A % Raw material (Compound 1a) 1560.96 1242.05 6.519 96.680 Product (Compound 1b) 1460.90 731.71 4.629 97.390

Example 32

[1052] Boc Removal Reaction of Boc-MeIle-Ala-MePhe-MeVal-Asp(pip)-MeAla-MePhe-Leu-MeLeu-Thr(OBn)-MeGly-OAllyl (Compound 1a) (11-mer: TMSOTf-ketene silyl acetal Method)

##STR00102##

[1053] The experiment was performed according to Example 2.

TABLE-US-00073 TABLE 74 Weight (mg) Yield (%) Raw material (Compound 1a) 23.6 — Product (Compound 1b) 19.8 90

TABLE-US-00074 TABLE 75 Analysis (HPLC method 1: Determination of purities of the raw material and target product) MW m/z rt Purity LC A % Raw material (Compound 1a) 1560.96 1242.05 6.519 96.680 Product (Compound 1b) 1460.90 731.63 4.620 97.488

Example 33

[1054] Boc Removal Reaction of Boc-MeIle-Ala-MePhe-MeVal-Asp(pip)-MeAla-MePhe-Leu-MeLeu-Thr(OBn)-MeGly-OAllyl (Compound 1a) (11-mer: TMSOTf-Proton Sponge Method)

##STR00103##

[1055] The experiment was performed according to Example 2.

TABLE-US-00075 TABLE 76 Weight (mg) Yield (%) Raw material (Compound 1a) 17.5 — Product (Compound 1b) 16.2 97

[1056] (The base remains even after post-treatment.)

TABLE-US-00076 TABLE 77 Analysis (HPLC method 1: Determination of purities of the raw material and target product) MW m/z rt Purity LC A % Raw material (Compound 1a) 1560.96 1242.05 6.519 96.680 Product (Compound 1b) 1460.90 731.68 4.628 96.025

[1057] (LCA % of the product described above is a value obtained by excluding the base remaining after post-treatment.)

Example 34

[1058] Boc Removal Reaction of Boc-MeIle-Ala-MePhe-MeVal-Asp(pip)-MeAla-MePhe-Leu-MeLeu-Thr(OBn)-MeGly-OAllyl (Compound 1a) (11-mer: TMSOTf-lutidine Method)

##STR00104##

[1059] 18.8 mg of the raw material was weighed into a reaction vessel and dissolved in 10 v/w of ethyl acetate. 0.005 ml (3.6 eq) of 2,6-lutidine and 0.0078 ml (3.6 eq) of TMSOTf were then added and the reaction solution was stirred. Four hours after the addition of the reagents, the reaction was analyzed by LCMS (HPLC method 1) and the conversion rate (=target product/(target product+starting material)) was confirmed to be 14%. Next, the reaction was analyzed by LCMS 19 hours after the addition of the reagents to find that the conversion rate was still 14%. Amide bond cleavage was not confirmed.

TABLE-US-00077 TABLE 78 Weight (mg) Yield (%) Raw material (Compound 1a) 18.8 — Product (Compound 1b) Not isolated Not isolated

TABLE-US-00078 TABLE 79 Analysis (HPLC method 1: Determination of purity of the raw material) MW m/z rt Purity LC A % Raw material (Compound 1a) 1560.96 1242.05 6.519 96.680

TABLE-US-00079 TABLE 80 Analysis (HPLC method 1: Determination of conversion rate) MW m/z rt LC A % Raw material (Compound 1a) 1560.96 1242.05 6.519 75.059 Product (Compound 1b) 1460.90 731.67 4.620 11.831

Example 35

[1060] Boc Removal Reaction of Boc-MeIle-Ala-MePhe-MeVal-Asp(pip)-MeAla-MePhe-Leu-MeLeu-Thr(OBn)-MeGly-OAllyl (Compound 1a) (11-mer: TMSOTf-lutidine Method)

##STR00105##

[1061] 89.3 mg of the raw material was weighed into a reaction vessel and dissolved in 1 ml of ethyl acetate. 0.016 ml (2.4 eq) of 2,6-lutidine and 0.012 ml (1.2 eq) of TMSOTf were then added and the reaction solution was stirred. 1.25 hours after the addition of the reagents, the reaction was analyzed by LCMS (HPLC method 1) to find that the conversion rate (=target product/(target product+starting material)) was 0%. Therefore, 0.032 ml (4.8 eq) of 2,6-lutidine and 0.024 ml (2.4 eq) of TMSOTf were further added. The reaction was analyzed by LCMS, 1.25 hours and 14 hours after the further addition of the reagents, respectively, to find that the conversion rate was still 0%. Amide bond cleavage was not confirmed.

TABLE-US-00080 TABLE 81 Weight (mg) Yield (%) Raw material (Compound 1a) 89.3 — Product (Compound 1b) Not isolated Not isolated

TABLE-US-00081 TABLE 82 Analysis (HPLC method 1: Determination of purity of the raw material) MW m/z rt Purity LC A % Raw material (Compound 1a) 1560.96 1242.05 6.545 89.564

TABLE-US-00082 TABLE 83 Analysis (HPLC method 1: Determination of conversion rate) MW m/z rt LC A % Raw material (Compound 1a) 1560.96 1242.05 6.571 89.649 Product (Compound 1b) 1460.90 ND ND 0

Example 36

[1062] Boc Removal Reaction of Boc-MeIle-Ala-MePhe-MeVal-Asp(pip)-MeAla-MePhe-Leu-MeLeu-Thr(OBn)-MeGly-OAllyl (Compound 1a) (11-mer: TMSOTf-2,6-di-tert-butylpyridine Method)

##STR00106##

[1063] 14.9 mg of the raw material was weighed into a reaction vessel and dissolved in 0.2 ml of ethyl acetate. 0.011 ml (5.4 eq) of 2,6-di-t-butylpyridine and 0.0062 ml (3.6 eq) of TMSOTf were then added and the reaction solution was stirred. Four hours after the addition of the reagents, the reaction was analyzed by LCMS (HPLC method 1) to find that the conversion rate (=target product/(target product+starting material)) was 59%. The reaction was analyzed again by LCMS 22 hours after the start of the reaction to find that the conversion rate was still 59%. Amide bond cleavage was not confirmed.

TABLE-US-00083 TABLE 84 Weight (mg) Yield (%) Raw material (Compound 1a) 14.9 — Product (Compound 1b) Not isolated Not isolated

TABLE-US-00084 TABLE 85 Analysis (HPLC method 1: Determination of purity of the raw material) MW m/z rt Purity LC A % Raw material (Compound 1a) 1560.96 1242.05 6.545 96.680

TABLE-US-00085 TABLE 86 Analysis (HPLC method 1: Determination of conversion rate) MW m/z rt LC A % Raw material (Compound 1a) 1560.96 1242.05 6.571 26.863 Product (Compound 1b) 1460.90 731.67 4.620 38.429 Ethyl acetate 1.850 12.893 2,6-di(tBu)pyridine 3.844 19.629

Example 37

[1064] Boc Removal Reaction of Boc-MeIle-Ala-MePhe-MeVal-Asp(pip)-MeAla-MePhe-Leu-MeLeu-Thr(OBn)-MeGly-OAllyl (Compound 1a) (11-mer: TMSOTf-tert-amine Method)

##STR00107##

[1065] 17.7 mg of the raw material was weighed into a reaction vessel and dissolved in 0.20 ml of ethyl acetate. 0.0057 ml (3.6 eq) of triethylamine and 0.0074 ml (3.6 eq) of TMSOTf were then added and the reaction solution was stirred. The reaction was analyzed by LCMS (HPLC method 1) one hour and four hours after the addition of the reagents to find that the conversion rate (=target product/(target product+starting material)) was merely 0.7% at each time. Amide bond cleavage was not confirmed.

TABLE-US-00086 TABLE 87 Weight (mg) Yield (%) Raw material (Compound 1a) 17.7 — Product (Compound 1b) Not isolated Not isolated

TABLE-US-00087 TABLE 88 Analysis (HPLC method 1: Determination of purity of the raw material) MW m/z rt Purity LC A % Raw material (Compound 1a) 1560.96 1242.05 6.519 96.680

TABLE-US-00088 TABLE 89 Analysis (HPLC method 1: Determination of conversion rate) MW m/z rt LC A % Raw material (Compound 1a) 1560.96 1242.05 6.579 84.391 Product (Compound 1b) 1460.90 731.67 4.639 0.599 Ethyl acetate (Solvent) 1.844 13.474

Example 38

[1066] Boc Removal Reaction of Boc-MeIle-Ala-MePhe-MeVal-Asp(pip)-MeAla-MePhe-Leu-MeLeu-Thr(OBn)-MeGly-OAllyl (Compound 1a) (11-mer: TMSOTf-tert-amine Method)

##STR00108##

[1067] 19.1 mg of the raw material was weighed into a reaction vessel and dissolved in 0.20 ml of ethyl acetate. 0.0074 ml (3.6 eq) of diisopropylethylamine and 0.0077 ml (3.6 eq) of TMSOTf were then added and the reaction solution was stirred. The reaction was analyzed by LCMS (HPLC method 2) two hours and four hours after the addition of the reagents to find that the conversion rate (=target product/(target product+starting material)) was merely 0.8% or less at each time. Amide bond cleavage was not confirmed.

TABLE-US-00089 TABLE 90 Weight (mg) Yield (%) Raw material (Compound 1a) 19.1 — Product (Compound 1b) Not isolated Not isolated

TABLE-US-00090 TABLE 91 Analysis (HPLC method 1: Determination of purity of the raw material) MW m/z rt Purity LC A % Raw material (Compound 1a) 1560.96 1242.05 6.519 96.680

TABLE-US-00091 TABLE 92 Analysis (HPLC method 1: Determination of conversion rate) MW m/z rt LC A % Raw material (Compound 1a) 1560.96 1241.79 6.568 85.820 Product (Compound 1b) 1460.90 731.68 4.629 0.458 Ethyl acetate (Solvent) 1.836 12.495

Example 39

[1068] Boc Removal Reaction of Boc-MeIle-Ala-MePhe-MeVal-Asp(pip)-MeAla-MePhe-Leu-MeLeu-Thr(OBn)-MeGly-OAllyl (Compound 1a) (11-mer: TMSOTf-tert-amine Method)

##STR00109##

[1069] 92.9 mg of the raw material was weighed into a reaction vessel and dissolved in 1 ml of ethyl acetate, and the reaction vessel was then cooled with ice. 0.015 ml (2.4 eq) of diisopropylethylamine and 0.013 ml (1.2 eq) of TMSOTf were then added under a nitrogen atmosphere, and the reaction solution was stirred. The reaction was analyzed two hours after the addition of the reagents to find that the conversion rate (=target product/(target product+starting material)) was 0%. Therefore, 0.030 ml (4.8 eq) of diisopropylethylamine and 0.026 ml (2.4 eq) of TMSOTf were further added three hours after the addition of the reagents. Three hours after the further addition of the reagents, the reaction was analyzed again to find that the conversion rate was 0%. Amide bond cleavage was not confirmed.

TABLE-US-00092 TABLE 93 Weight (mg) Yield (%) Raw material (Compound 1a) 92.9 — Product (Compound 1b) Not isolated Not isolated

TABLE-US-00093 TABLE 94 Analysis (HPLC method 1: Determination of purity of the raw material) MW m/z rt Purity LC A % Raw material (Compound 1a) 1560.96 1241.93 6.538 89.465

TABLE-US-00094 TABLE 95 Analysis (HPLC method 1: Determination of conversion rate) MW m/z rt LC A % Raw material (Compound 1a) 1560.96 1242.05 6.568 89.432 Product (Compound 1b) 1460.90 ND ND 0

Example 40

[1070] Boc Removal Reaction of Boc-MeIle-Ala-MePhe-MeVal-Asp(pip)-MeAla-MePhe-Leu-MeLeu-Thr(OBn)-MeGly-OAllyl (Compound 1a) (11-mer: conducted using only TMSOTf in the absence of an organic base.)

##STR00110## ##STR00111##

[1071] 18.3 mg of the raw material was weighed into a reaction vessel and dissolved in 0.2 ml of ethyl acetate, and the reaction vessel was then cooled with ice. 0.0077 ml (3.6 eq) of TMSOTf was then added under a nitrogen atmosphere, and the reaction solution was stirred. The reaction was analyzed 1.5 hours after the addition of the reagent to confirm that multiple amide bond-cleaved products were produced at a conversion rate (=target product/(target product+starting material)) of 68%.

TABLE-US-00095 TABLE 96 Weight (mg) Yield (%) Raw material (Compound 1a) 18.3 — Product (Compound 1b) Not isolated Not isolated

TABLE-US-00096 TABLE 97 Analysis (HPLC method 1: Determination of purity of the raw material) MW m/z rt Purity LC A % Raw material (Compound 1a) 1560.96 1242.05 6.519 96.680

TABLE-US-00097 TABLE 98 Analysis (HPLC method 1: Determination of conversion rate and rates of production of by-products) MW m/z rt LC A % Raw material (Compound 1a) 1560.96 1241.77 6.575 15.543 Product (Compound 1b) 1460.90 731.72 4.630 32.712 Cleaved product (Compound 1c) 377.23 378.22 ([M + H]+) 2.269 0.986 Cleaved product (Compound 1d) 757.47 758.40 ([M + H]+) 2.758 9.731 Cleaved product (Compound 1e) 721.44 722.43 ([M + H]+) 3.893 15.859 Cleaved product (Compound 1f) 1101.68 1102.56 ([M + H]+)  4.220 4.701 Cleaved product (Compound 1g) Unknown 573.27 4.788 4.808 Ethyl acetate 1.840 15.660
The structure of Compound 1g is unknown.

Example 41

[1072] Boc Removal Reaction of Boc-MeIle-Ala-MePhe-MeVal-Asp(pip)-MeAla-MePhe-Leu-MeLeu-Thr(OBn)-MeGly-OAllyl (Compound 1a) (11-mer: conducted with TMSOTf and triisopropylsilane in the absence of an organic base.)

##STR00112## ##STR00113##

[1073] 20.5 mg of the raw material was weighed into a reaction vessel and dissolved in 0.2 ml of ethyl acetate, and the reaction vessel was then cooled with ice. 0.0097 ml (3.6 eq) of triisopropylsilane and 0.0085 ml (3.6 eq) of TMSOTf were then added under a nitrogen atmosphere, and the reaction solution was stirred. The reaction was analyzed one hour after the addition of the reagents to confirm that multiple e bond-cleaved products were produced at a conversion rate (=target product/(target product+starting material)) of 43%.

TABLE-US-00098 TABLE 99 Weight (mg) Yield (%) Raw material (Compound 1a) 20.5 — Product (Compound 1b) Not isolated Not isolated

TABLE-US-00099 TABLE 100 Analysis (HPLC method 1: Determination of purity of the raw material) MW m/z rt Purity LC A % Raw material (Compound 1a) 1560.96 1242.05 6.538 96.680

TABLE-US-00100 TABLE 101 Analysis (HPLC method 1: Determination of conversion rate and rates of production of by- products) MW m/z rt LC A % Raw material (Compound 1a) 1560.96 1241.77 6.575 28.216 Product (Compound 1b) 1460.90 731.59 4.624 20.995 Cleaved product (Compound 1c) 377.23 378.24 ([M + H]+) 2.259 0.479 Cleaved product (Compound 1d) 757.47 758.44 ([M + H]+) 2.750 5.875 Cleaved product (Compound 1e) 721.44 722.36 ([M + H]+) 3.885 15.802 Cleaved product (Compound 1f) 1101.68 1102.55 ([M + H]+)  4.212 5.239 Impurity (Compound 1g) Unknown 573.24 4.780 8.178 Ethyl acetate 1.836 14.570
The structure of Compound 1g is unknown.

Example 42

[1074] Boc Removal Reaction of Boc-MeIle-Ala-MePhe-MeVal-Asp(pip)-MeAla-MePhe-Leu-MeLeu-Thr(OBn)-MeGly-OAllyl (Compound 1a) (11-mer: TFA Method)

##STR00114##

[1075] 35.3 mg of the raw material was weighed into a reaction vessel and dissolved in 10 v/w of methylene chloride, and the reaction vessel was then cooled with ice. 0.004 ml (2.3 eq) of trifluoroacetic acid was then added under a nitrogen atmosphere, and the reaction solution was stirred. The reaction was analyzed 10 minutes after the addition of the reagent to find that the conversion rate (=target product/(target product+starting material)) was 0%. Therefore, 0.004 ml (2.3 eq) of trifluoroacetic acid was further added. The reaction was analyzed again 10 minutes after the further addition of the reagent to find that the conversion rate was 0%. Therefore, 0.0094 ml (5.4 eq) of trifluoroacetic acid was further added. 10 minutes after the further addition of the reagent, the reaction solution was warmed to room temperature and continued to be stirred. Four hours after warming to room temperature, the reaction was analyzed to confirm that an amide bond-cleaved product 1e was produced at a conversion rate of 2%.

TABLE-US-00101 TABLE 102 Weight (mg) Yield (%) Raw material (Compound 1a) 35.3 — Product (Compound 1b) Not isolated Not isolated

TABLE-US-00102 TABLE 103 Analysis (HPLC method 1: Determination of purity of the raw material) MW m/z rt Purity LC A % Raw material (Compound 1a) 1560.96 1242.05 6.519 96.680

TABLE-US-00103 TABLE 104 Analysis (HPLC method 1: Determination of conversion rate and rates of production of by-products) MW m/z rt LC A % Raw material (Compound 1a) 1560.96 1242.05 6.568 93.488 Product (Compound 1b) 1460.90 731.67 4.637 2.048 Cleaved product (Compound 1e) 721.44 722.47 ([M + H]+) 3.900 0.880 Impurity (Compound 1g) Unknown 573.29 4.789 1.020 Methylene chloride 2.375 1.059
The structure of Compound 1g is unknown.

Example 43

[1076] Boc Removal Reaction of Boc-MeIle-Ala-MePhe-MeVal-Asp(pip)-MeAla-MePhe-Leu-MeLeu-Thr(OBn)-MeGly-OAllyl (Compound 1a) (11-mer: TFA-TIPS-H.sub.2O-PhOH Method)

[1077] Reference: J. Am. Chem. Soc., 2015, 137, 13488

##STR00115## ##STR00116##

[1078] 15.9 mg of the raw material was weighed into a reaction vessel, 0.0125 ml of water and 0.0125 ml of triisopropylsilane were added, and the reaction vessel was cooled to −10° C. 0.48 ml of trifluoroacetic acid was then added and the reaction solution was stirred. The reaction was analyzed two hours after the addition of the reagent to confirm that amide bond-cleaved products 1d and 1e were produced at a conversion rate (=target product/(target product+starting material)) of 89%. The reaction was analyzed again 14.5 hours after the addition of the reagent to confirm that the raw material was consumed and the amide bond-cleaved products 1d and 1e were increasingly produced.

TABLE-US-00104 TABLE 105 Weight (mg) Yield (%) Raw material (Compound 1a) 15.9 — Product (Compound 1b) Not isolated Not isolated

TABLE-US-00105 TABLE 106 Analysis (HPLC method 1: Determination of purity of the raw material) MW m/z rt Purity LC A % Raw material (Compound 1a) 1560.96 1242.05 6.519 96.680

TABLE-US-00106 TABLE 107 Analysis (HPLC method 1: Determination of conversion rate and rates of production of by-products at two hours after the reaction) LC MW m/z rt A % Raw material 1560.96 1241.57 6.527 9.531 (Compound 1a) Product (Compound 1b) 1460.90 731.67 4.629 77.361 Cleaved product 757.47 758.41 ([M + H]+) 2.761 5.716 (Compound 1d) Impurity (Compound 1e) 721.44 722.47 ([M + H]+) 3.902 7.392

TABLE-US-00107 TABLE 108 Analysis (HPLC method 1: Rates of production of the target product and by-products at 14.5 hours after the reaction) LC MW m/z rt A % Raw material 1560.96 ND ND 0 (Compound 1a) Product (Compound 1b) 1460.90 731.67 4.631 29.383 Cleaved product 757.47 758.39 ([M + H]+) 2.757 37.117 (Compound 1d) Cleaved product 721.44 722.41 ([M + H]+) 3.892 32.950 (Compound 1e)

Example 44

[1079] Boc Removal Reaction of Boc-MeIle-Ala-MePhe-MeVal-Asp(pip)-MeAla-MePhe-Leu-MeLeu-Thr(OBn)-MeGly-OAllyl (Compound 1a) (11-mer: TFA-TIPS-H.sub.2O-PhOH Method) Reference: J. Am. Chem. Soc., 2012, 134, 13244

##STR00117## ##STR00118##

[1080] 10.3 mg of phenol was weighed into a reaction vessel, 0.034 ml of water and 0.025 ml of triisopropylsilane were added, and the reaction vessel was cooled to −10° C. 0.50 ml of trifluoroacetic acid was then added. Six minutes after the addition, 13.4 mg of the raw material was added and the reaction solution was stirred. The reaction was analyzed two hours after the addition of the raw material to confirm that amide bond-cleaved products 1d and 1e were produced at a conversion rate (=target product/(target product+starting material)) of 82%. The reaction was analyzed again seven hours after the addition of the raw material to confirm that the raw material was consumed and the amide bond-cleaved products 1d and 1e were increasingly produced.

TABLE-US-00108 TABLE 109 Weight (mg) Yield (%) Raw material (Compound 1a) 10.3 — Product (Compound 1b) Not isolated Not isolated

TABLE-US-00109 TABLE 110 Analysis (HPLC method 1: Determination of purity of the raw material) MW m/z rt Purity LC A % Raw material (Compound 1a) 1560.96 1242.05 6.538 96.680

TABLE-US-00110 TABLE 111 Analysis (HPLC method 1: Determination of conversion rate and rates of production of by-products at two hours after the reaction) LC MW m/z rt A % Raw material 1560.96 1242.05 6.523 6.500 (Compound 1a) Product (Compound 1b) 1460.90 731.67 4.628 30.640 Cleaved product 757.47 758.38 ([M + H]+) 2.769 1.583 (Compound 1d) Cleaved product 721.44 722.37 ([M + H]+) 3.884 2.659 (Compound 1e) Phenol 2.172 58.619

TABLE-US-00111 TABLE 112 Analysis (HPLC method 1: Rates of production of the target product and by-products at five hours after the reaction) LC MW m/z rt A % Raw material 1560.96 ND ND 0 (Compound 1a) Product (Compound 1b) 1460.90 731.67 4.633 25.258 Cleaved product 757.47 758.40 ([M + H]+) 2.769 8.506 (Compound 1d) Cleaved product 721.44 722.39 ([M + H]+) 3.884 8.035 (Compound 1e) Phenol 2.172 58.200

[1081] As shown in Examples 2, 3, and 26 to 44, it was found that imidates, amides, ketene acetals, enol ethers, imines, amines, diamines, and dialkylcarbodiimides are excellent as electrophilic species scavengers, suppress main chain damage, and allow efficient progress of Boc removal reaction of peptides having a long main chain such as 11-mers.

D. Experiment of Comparison Between the TFA Method and the TMSOTf-HMDS Method in tBu Removal Reactions (Table 113)

[1082] ##STR00119##

Example 45

[1083] t-Bu Removal Reaction of Compound 13a (TFA Method)

[1084] 29.8 mg of Substrate 13a was weighed into a reaction vessel and dissolved in 10 v/w of dichloromethane. To the solution was added TFA (10 eq.) at room temperature. Four hours after the addition of the reagent, TFA (10 eq.) was further added. 1.5 hours after the further addition of the reagent, the reaction was analyzed by LCMS to find that the reaction conversion rate was 52%. The purity was reduced by 22%, while 10% of a by-product due to amide bond cleavage was observed.

Example 46

[1085] t-Bu Removal Reaction of Compound 13a (TMSOTf-HMDS Method)

[1086] 30.0 mg of Substrate 13a was weighed into a reaction vessel and dissolved in 10 v/w of ethyl acetate. HMDS (3.0 eq.) and TMSOTf (2.0 eq) were sequentially added to the solution at 0° C. Two hours after the addition of the reagents, the reaction solution was warmed to room temperature. 2.5 hours after warming, HMDS (3 eq.) and TMSOTf (2 eq.) were further added. 30 minutes after the further addition of the reagents, the reaction was analyzed by LCMS to confirm that the conversion rate was 99% or more. At this time, amide bond cleavage was not confirmed.

##STR00120##

Example 47

[1087] t-Bu Removal Reaction of Compound 14a (TFA Method)

[1088] 29.7 mg of Substrate 14a was weighed into a reaction vessel and dissolved in 10 v/w of dichloromethane. To the solution was added TFA (20 eq.) at room temperature. Seven hours after the addition of the reagent, the reaction was analyzed by LCMS to find that the reaction conversion rate was 92%. The purity was reduced by 46%, while 24% of a by-product due to amide bond cleavage was observed.

Example 48

[1089] t-Bu Removal Reaction of Compound 14a (TMSOTf-HMDS Method)

[1090] 30.0 mg of Substrate 14a was weighed into a reaction vessel and dissolved in 10 v/w of ethyl acetate. HMDS (3.0 eq.) and TMSOTf (2.0 eq) were sequentially added to the solution at 0° C. Six hours after the addition of the reagents, the reaction was analyzed by LCMS and the raw material was confirmed to disappear. At this time, amide bond cleavage was not confirmed. After confirming the completion of the reaction, 44.5 mg (8.0 eq) of potassium hydrogenphosphate and 0.30 mL of water were added to the reaction solution, which was then stirred for 30 minutes with ice-cooling. 1 mL of ethyl acetate was added to the reaction solution, which was then washed with 0.30 mL of brine. The organic layer was filtered through celite, then concentrated, and dried under reduced pressure to give 26.8 mg of 2b in a purity of 99%.

##STR00121##

Example 49

[1091] t-Bu Removal Reaction of Compound 15a (TFA Method)

[1092] 29.9 mg of Substrate 15a was weighed into a reaction vessel and dissolved in 10 v/w of dichloromethane. To the solution was added TFA (20 eq.) at room temperature. 5.5 hours after the addition of the reagent, the reaction was analyzed by LCMS to find that the reaction conversion rate was 50%. 28% of a by-product due to amide bond cleavage was observed.

Example 50

[1093] t-Bu Removal Reaction of Compound 15a (TMSOTf-HMDS Method)

[1094] 30.0 mg of Substrate 15a was weighed into a reaction vessel and dissolved in 10 v/w of ethyl acetate. HMDS (3.6 eq) and TMSOTf (2.4 eq) were sequentially added to the solution at room temperature. Five hours after the addition of the reagents, HMDS (1.8 eq.) and TMSOTf (1.2 eq.) were further added. One hour after the further addition of the reagents, the reaction was analyzed by LCMS and the raw material was confirmed to disappear. At this time, amide bond cleavage was not confirmed.

##STR00122##

Example 51

[1095] t-Bu Removal Reaction of Compound 16a (TFA Method)

[1096] 30.2 mg of Substrate 16a was weighed into a reaction vessel and dissolved in 10 v/w of dichloromethane. To the solution was added TFA (20 eq.) at room temperature. 4.5 hours after the addition of the reagent, TFA (10 eq.) was further added. One hour after the further addition of the reagent, the reaction was analyzed by LCMS to find that the reaction conversion rate was 65%. 7.0% of a by-product due to amide bond cleavage was observed.

Example 52

[1097] t-Bu Removal Reaction of Compound 16a (TMSOTf-HMDS Method)

[1098] 29.9 mg of Substrate 16a was weighed into a reaction vessel and dissolved in 10 v/w of ethyl acetate. HMDS (3.6 eq) and TMSOTf (2.4 eq) were sequentially added to the solution at room temperature. Four hours after the addition of the reagents, the reaction was analyzed by LCMS and the raw material was confirmed to disappear. At this time, amide bond cleavage was not confirmed.

##STR00123##

Example 53

[1099] t-Bu Removal Reaction of Compound 17a (TFA Method)

[1100] 50.0 mg of Substrate 17a was weighed into a reaction vessel and dissolved in 10 v/w of dichloromethane. To the solution was added TFA (5 eq.) at 0° C. Two hours after the addition of the reagent, TFA (5 eq.) was added. After another 15.5 hours, the reaction solution was warmed to room temperature. Three hours after warming, TFA (10 eq.) was added. After another 1.5 hours, TFA (20 eq.) was added. 1.5 hours after the further addition of the reagent, the reaction was analyzed by LCMS to find that the reaction conversion rate was 54%. 26% of a by-product due to amide bond cleavage was observed.

Example 54

[1101] t-Bu Removal Reaction of Compound 17a (TMSOTf-HMDS Method)

[1102] 30.0 mg of Substrate 17a was weighed into a reaction vessel and dissolved in 10 v/w of ethyl acetate. HMDS (5.4 eq.) and TMSOTf (3.6 eq) were sequentially added to the solution at 0° C. Four hours after the addition of the reagents, the reaction solution was warmed to room temperature. Three hours after warming, the reaction was analyzed by LCMS and the raw material was confirmed to disappear. At this time, amide bond cleavage was not confirmed.

##STR00124##

Example 55

[1103] t-Bu Removal Reaction of Compound 18a (TFA Method)

[1104] 29.9 mg of Substrate 18a was weighed into a reaction vessel and dissolved in 10 v/w of dichloromethane. To the solution was added TFA (20 eq.) at 0° C. 4.5 hours after the addition of the reagent, TFA (10 eq.) was further added and the reaction solution was warmed to room temperature. Two hours after the further addition of the reagent, the reaction was analyzed by LCMS to find that the reaction conversion rate was 82%. 34% of a by-product due to amide bond cleavage was observed.

Example 56

[1105] t-Bu Removal Reaction of Compound 18a (TMSOTf-HMDS Method)

[1106] 29.8 mg of Substrate 18a was weighed into a reaction vessel and dissolved in 10 v/w of ethyl acetate. HMDS (7.2 eq.) and TMSOTf (4.8 eq) were sequentially added to the solution at 0° C. Three hours after the addition of the reagents, the reaction solution was warmed to room temperature. 3.5 hours after warming, the reaction was analyzed by LCMS and the raw material was confirmed to disappear. At this time, amide bond cleavage was not confirmed.

##STR00125##

Example 57

[1107] t-Bu Removal Reaction of Compound 19a (TFA Method)

[1108] 50.0 mg of Substrate 19a was weighed into a reaction vessel and dissolved in 10 v/w of dichloromethane. To the solution was added TFA (5 eq.) at 0° C. Two hours after the addition of the reagent, TFA (5 eq.) was added. After another 15.5 hours, the reaction solution was warmed to room temperature. Three hours after warming, TFA (10 eq.) was added. After another 1.5 hours, TFA (20 eq.) was added. 1.5 hours after the further addition of the reagent, the reaction was analyzed by LCMS to find that the reaction conversion rate was 82%. 14% of a by-product due to amide bond cleavage was observed.

Example 58

[1109] t-Bu Removal Reaction of Compound 19a (TMSOTf-HMDS Method)

[1110] 30.0 mg of Substrate 19a was weighed into a reaction vessel and dissolved in 10 v/w of ethyl acetate. HMDS (3.6 eq.) and TMSOTf (2.4 eq) were sequentially added to the solution at 0° C. 3.5 hours after the addition of the reagents, the reaction solution was warmed to room temperature and HMDS (3.6 eq.) and TMSOTf (2.4 eq.) were further added. One hour after the further addition of the reagents, the reaction was analyzed by LCMS and the raw material was confirmed to disappear. At this time, amide bond cleavage was not confirmed.

As shown in Examples 45 to 58, it was found that in tBu removal reactions, the TMSOTf-HMDS method using a combination of silylating agents and electrophilic species scavengers does not cause main chain damage and allows deprotection reactions to proceed more efficiently, as compared with the conventional TFA method.

TABLE-US-00112 TABLE 113 Reaction conditions and experimental results Example Conversion Purity reduction Main chain cleavage Substrate No. Reaction condition rate (LCMS Area %) (LCMS Area %) 13a 45 TFA (10 to 20 eq.) 52% 22% MePhe-MeVal: 10.0% CH.sub.2Cl.sub.2 (10 v/w) rt, 6 h 46 TMSOTf (2.4 to 4.8 eq.) >99%  1.2%  Not detected. HMDS (3.6 to 7.2 eq.) EtOAc (10 v/w %) 0° C. to rt, 5 h 14a 47 TFA (20 eq.) 92% 46% MePhe-MeVal: 22.0% CH.sub.2Cl.sub.2 (10 v/w) MeVal-Asp(pip): 1.5% rt, 7 h 48 TMSOTf (2 eq.) 100%  0.7%  Not detected. HMDS (3 eq.) EtOAc (10 v/w %) 0° C., 6 h 15a 49 TFA (20 eq.) 50% 33% MeIle-Ser(tBu): 6.4% CH.sub.2Cl.sub.2 (10 v/w) MePhe-MeVal: 16.3% rt, 6 h MeVal-Asp(pip): 5. 0% 50 TMSOTf (2.4 to 3.6 eq.) 100%  0.2%  Not detected. HMDS (3.6 to 5.4 eq.) EtOAc (10 v/w %) rt, 6 h 16a 51 TFA (20 to 30 eq.) 65% 26% MeIle-tBuSer: 7.0% CH.sub.2Cl.sub.2 (10 v/w) rt, 6 h 52 TMSOTf (2.4 eq.) 100%  ≈0% Not detected. HMDS (3.6 eq.) EtOAc (10 v/w %) rt, 4 h 17a 53 TFA (5 to 40 eq.) 54% 25% MeIle-Ser(tBu): 5.8% CH.sub.2Cl.sub.2 (10 v/w) MeVal-Asp(pip): 20.5% 0° C. to rt, 24 h 54 TMSOTf (3.6 eq.) 100%  2.6%  Not detected. HMDS (5.4 eq.) EtOAc (10 v/w %) 0° C. to rt, 7 h 18a 55 TFA (20 to 30 eq.) 82% 33% MePhe-MeVal: 15.1% CH.sub.2Cl.sub.2 (10 v/w) MeVal-Asp(pip): 12.3% 0° C. to rt, 7 h MeLeu-Val: 6.9% 56 TMSOTf (4.8 eq.) 100%  2.7%  Not detected. HMDS (7.2 eq.) EtOAc (10 v/w %) 0° C. to rt, 7 h 19a 57 TFA (5 to 40 eq.) 54% 17% MeIle-Ser(tBu): 0.8% CH.sub.2Cl.sub.2 (10 v/w) MePhe-Leu: 5.4% 0° C. to rt, 24 h MePhe-MeVal: 6. 6% MeLeu-Val: 1.6% 58 TMSOTf (2.4 to 4.8 eq.) 100%  0.6%  Not detected. HMDS (3.6 to 7.2 eq.) EtOAc (10 v/w %) 0° C. to rt, 5 h

Identification of Products

[1111]

TABLE-US-00113 TABLE 114 (HPLC method 1) MS (major peaks) and retention times of the target products (in Examples of the TMSOTf/HMDS method) Compound data Retention Example Compound time No. No. MW m/z (min) 45, 46 13b 751.92 752.42 ([M + H]+) 4.723 47, 48 14b 879.11 879.52 ([M + H]+) 5.313 49, 50 15b 1049.32 1049.63 ([M + H]+) 4.985 51, 52 16b 1176.51 1198.67 ([M + Na]+) 5.527 53, 54 17b 1275.64 1089.62 5.652 55, 56 18b 1450.87 536.23 5.964 57, 58 19b 1535.98 1222.41 6.183

TABLE-US-00114 TABLE 115 MS and retention times of decomposed products Example Compound data No. Compound No. MW m/z Retention time (min) 45 13c + its isomer 456.54 457.28 ([M + H]+) 4.504 439.30 ([M—H20 + H]+) 4.894 13d + its isomer 369.51 370.29 ([M + H]+) 2.691 370.31 ([M + H]+) 2.731 47 14c 696.89 679.35 ([M—H20 + H]+) 4.935 14d + its isomer 583.73 584.20 ([M + H]+) 5.118 566.19 ([M—H20 + H]+) 5.477 49 15c + its isomer 449.55 432.28 ([M—H20 + H]+) 5.287 450.30 ([M + H]+) 3.928 15d 673.90 674.40 ([M + H]+) 3.792 15e 753.94 754.29 ([M + H]+) 4.863 15f + its isomer 697.83 698.35 ([M + H]+) 4.101 736.93 736.30 ([M—H20 + H]+) 5.123 15g + its isomer 810.99 793.37 ([M—H20 + H]+) 4.487 793.40 ([M—H20 + H]+) 5.444 51 16c 673.90 674.48 ([M + H]+) 3.591 16d + its isomer 576.74 577.32 ([M + H]+) 4.500 559.28 ([M—H20 + H]+) 5.267 53 17c + its isomers 689.90 690.45 ([M + H]+) 4.764 672.38 ([M—H20 + H]+) 5.432 672.59 ([M—H20 + H]+) 5.463 17d + its isomer 1107.45 1129.61 ([M + Na]+) 5.627 17e + its isomers 1051.34 1033.66 ([M—H20 + H]+) 5.116 1033.77 ([M—H20 + H]+) 5.944 1033.64 ([M—H20 + H]+) 5.985 55 18c 553.70 536.31 ([M—H20 + H]+) 5.199 18d 1212.54 1194.58 ([M—H20 + H]+) 5.469 18e + its isomers 1155.49 1177.57 ([M + Na]+) 5.746 1177.39 ([M + Na]+) 5.788 1137.73 ([M—H20 + H]+) 5.904 57 19c 673.90 674.38 ([M + H]+) 3.633 19d + its isomer 1155.53 1155.73 ([M + H]+) 3.863, 3.906 19e 1211.64 1211.65 ([M + H]+) 4.619 19f 398.46 381.16 ([M—H20 + H]+) 4.898 19g 638.81 621.31 ([M—H20 + H]+) 5.246 19h + its isomers 1240.60 1240.51 ([M + H]+) 5.794, 5.831 1222.59 ([M—H20 + H]+) 5.953

[1112] The structures of Compounds 13c to 19h are provided below.

##STR00126## ##STR00127## ##STR00128##

Example 59

[1113] t-Bu Removal Reaction of Fmoc-MeAsp(OtBu)-pip (Compound 12a) (1-mer: TFA Method)

##STR00129##

[1114] 48.0 mg of the raw material was weighed into a reaction vessel and dissolved in 9 v/w of dichloromethane. 1 v/w of trifluoroacetic acid was then added and the reaction solution was stirred. 19 hours after the addition of the reagents, the reaction was analyzed by LCMS (HPLC method 4) and the conversion rate (=target product/(target product+starting material)) was confirmed to be 77%. At this time, a rearrangement product 12c and a hydrolysate 12d were observed as compounds in which amide bonds were damaged. Impurities of unknown structure were also produced.

TABLE-US-00115 TABLE 116 Weight (mg) Yield (%) Raw material (Compound 12a) 48.0 — Product (Compound 12b) Not isolated Not isolated

[1115] Raw Material Purity Analysis (HPLC Method 1)

TABLE-US-00116 TABLE 117 Analysis (HPLC method 1) MW m/z rt Purity LC A % Raw material (Compound 12a) 492.26 493.24 ([M + H]+) 5.527 98.268

TABLE-US-00117 TABLE 118 Analysis (HPLC method 4) MW m/z rt Purity LC A % Raw material (Compound 12a) 492.26 437.25 ([M-tBu + H]+) 23.660 98.268 Product (Compound 12b) 436.20 437.22 ([M + H]+) 15.194 53.776 Rearrangement product (Compound 12c) 436.20 437.28 ([M + H]+) 15.553 1.474 Hydrolysate (Compound 12d) 369.12 250.91 10.522 0.901 Impurity of unknown structure (12e) 102.07 17.949 28.043

Example 60

[1116] t-Bu removal reaction of Fmoc-MeAsp(OtBu)-pip (Compound 12a) (1-mer: TMSOTf-HMDS Method)

##STR00130##

[1117] The experiment was performed according to Example 2.

TABLE-US-00118 TABLE 119 Weight (g) Yield (%) Raw material (Compound 12a) 59.9 — Product (Compound 12b) 55.8 Quantitative

TABLE-US-00119 TABLE 120 Analysis (HPLC method 1) MW m/z rt Purity LC A % Raw material (Compound 12a) 492.26 493.24 ([M + H]+) 5.527 98.268

TABLE-US-00120 TABLE 121 Analysis (HPLC method 4) MW m/z rt Purity LC A % Product (Compound 12b) 436.20 437.22 ([M + H]+) 15.194 98.999 Rearrangement product (Compound 12c) 436.20 437.28 ([M + H]+) 15.553 0.379 Hydrolysate (Compound 12d) 369.12 250.91 10.522 0.15 Impurity of unknown structure (12e) 102.07 17.949 0.474
E. Experiment of Comparison Between the TFA Method and the TMSOTf-HMDS Method in t-Bu Removal Reactions (Table 122)

##STR00131##

Example 61

[1118] t-Bu Removal Reaction of Compound 20a (TFA Method)

[1119] 27.1 mg of Substrate 20a was weighed into a reaction vessel and dissolved in 10 v/w of dichloromethane. To the solution was added TFA (10 eq.) at room temperature. Eight hours after the addition of the reagent, the reaction was analyzed by LCMS to find that the reaction conversion rate was 76%. The purity was reduced by 11%, while 8.1% of a by-product due to amide bond cleavage was confirmed. The same reaction was performed for Substrate 21a (Table 122).

Example 62

[1120] t-Bu Removal Reaction of Compound 20a (TMSOTf-HMDS Method)

[1121] 27.6 mg of Substrate 20a was weighed into a reaction vessel and dissolved in 10 v/w of ethyl acetate. HMDS (2.4 eq) and TMSOTf (2.4 eq) were sequentially added to the solution at room temperature. Two hours after the addition of the reagents, the reaction was analyzed by LCMS and the raw material was confirmed to disappear. At this time, amide bond cleavage was not confirmed.

[1122] The same reaction was performed for Substrate 21a (Table 122).

TABLE-US-00121 TABLE 122 Purity Amide bond Example Raw Reaction Conversion reduction cleavage Substrate No. material time rate (LCMS Area %) (LCMS Area %) 20a 61 27.1 mg 8 h  76% 11%  8.1% 62 27.6 mg 2 h 100% 0% Not detected. 21a 63 26.0 mg 8 h  76% 9% 2.7% 64 26.2 mg 2 h 100% 0% Not detected.

[1123] Identification of Products

TABLE-US-00122 TABLE 123 (HPLC method 1) Major peaks and retention times of products and target products Example Compound data No. Compound MW m/z Retention time (min) 61 20b 410.47 433.16 ([M + Na]+)    3.858 20c 325.36 104.01 ([M-Fmoc + H]+)*  3.976 63 21b 486.57 487.17 ([M + Na]+)    4.485 21c 401.46 180.02 ([M-Fmoc + H]+)** 4.594 [00132][00133]

F. Experiment of Comparison Between the TFA Method and the TMSOTf-HMDS Method in Resin Removal Reactions (Table 124)

[1124] ##STR00134##

Example 65

[1125] Resin Removal Reaction of Resin 22a (TFA Method)

[1126] 30.2 mg of Resin 22a was weighed into a reaction vessel, and the resin was swollen by adding 10 v/w of dichloromethane. After removing the dichloromethane from the reaction solution, 10 v/w % of TFA (5% dichloromethane solution) was added at room temperature and the vessel was shaken at 25° C. One hour after the shaking started, the reaction was analyzed by LCMS to find that 7.8% of a by-product due to amide bond cleavage was produced. The same reaction was performed for Resins 23a to 24a (Table 124).

Example 66

[1127] Resin Removal Reaction of Resin 22a (TMSOTf-HMDS Method)

[1128] 23.0 mg of Substrate 22a was weighed into a reaction vessel, and the resin was swollen by adding 10 v/w of 1,2-dichloroethane. After removing the 1,2-dichloroethane from the reaction solution, a solution of HMDS (3.6 eq.) and TMSOTf (2.4 eq.) in 10 v/w of 1,2-dichloroethane was added to the reaction vessel. After stirring at 25° C. for four hours, the reaction was analyzed by LCMS and no amide bond cleavage was confirmed. The same reaction was performed for Resins 23a to 24a (Table 124).

TABLE-US-00123 TABLE 124 Amide bond Example Raw Reaction cleavage Substrate No. material Reagent amount time (LCMS Area %) 22a 65 30.2 mg 5% TFA in 1 h 7.9% CH.sub.2Cl.sub.2 (10 v/w) 66 23.0 mg TMSOTf (2.4 eq.) 4 h .sup. 0% HMDS (4.8 eq.) 23a 67 30.0 mg 5% TFA in 1 h 2.3% CH.sub.2Cl.sub.2 (10 v/w) 68 24.0 mg TMSOTf (2.4 eq.) 4 h .sup. 0% HMDS (4.8 eq.) 24a 69 30.3 mg 5% TFA in 1 h 0.3% CH.sub.2Cl.sub.2 (10 v/w) 70 23.6 mg TMSOTf (2.4 eq.) 4 h .sup. 0% HMDS (4.8 eq.)

[1129] Identification of Products

[1130] HPLC Method A [1131] Instrument: Waters ACQUITY UPLC H-Class [1132] Column: Ascentis Express C18 (2.7 μm, 4.6 mm×50 mm), Supelco [1133] Eluent: A) 0.05% TFA/water, B) 0.05% TFA/CH3CN [1134] Gradient (B): 5% (0 min.).Math.100% (4 min.).Math.100% (4.5 min.).Math.5% (4.6 min.) .Math.5% (6 min.) [1135] Flow rate: 1.0 mL/min. [1136] Detection: 210 nm [1137] Injection vol.: 5 μL [1138] Sample prep.: 25 μL/0.975 mL MeCN

[1139] HPLC Method B [1140] Instrument: Waters ACQUITY UPLC H-Class [1141] Column: Ascentis Express C18 (2.7 μm, 4.6 mm×50 mm), Supelco [1142] Eluent: A) 0.05% TFA/water, B) 0.05% TFA/CH3CN [1143] Gradient (B): 5% (0 min.).Math.100% (4 min.).Math.100% (4.5 min.).Math.5% (4.6 min.).Math. [1144] 5% (6 min.) [1145] Flow rate: 1.0 mL/min. [1146] Detection: 210 nm [1147] Injection vol.: 5 μL [1148] Sample prep.: 5 μL/1.00 mL (MeCN 0.950 ml+0.1 M phosphate buffer (pH 8.0) 0.050 ml)

[1149] HPLC Method C [1150] Instrument: Waters ACQUITY UPLC H-Class [1151] Column: Ascentis Express C18 (2.7 μm, 4.6 mm×50 mm), Supelco [1152] Eluent: A) 0.05% TFA/water, B) 0.05% TFA/CH3CN Gradient (B): 5% (0 min.).Math.100% (4 min.).Math.100% (4.5 min.).Math.5% (4.6 min.).Math.5% (6 min.) [1153] Flow rate: 1.0 mL/min. [1154] Detection: 210 nm [1155] Injection vol.: 5 μL [1156] Sample prep.: 5 μL/1.00 mL MeCN

[1157] HPLC Method D [1158] Instrument: Shimadzu LCMS-2020 [1159] Column: Ascentis Express C18 (2.7 μm, 2.1 mm×50 mm), Supelco [1160] Eluent: A) 0.1% FA/water, B) 0.1% FA/CH3CN [1161] Gradient (B): 5% (0 min.).Math.100% (4.5 min.).Math.100% (5 min.) [1162] Flow rate: 1.0 mL/min. [1163] Detection: 210 nm-400 nm [1164] Injection vol.: 1 μL

[1165] HPLC Method E [1166] Instrument: SHIMADZU LCMS-2020 [1167] Column: Ascentis Express C18 2.1 mm×50 mm, Supelco [1168] Eluent: A) 0.1% FA/water, B) 0.1% FA/CH3CN [1169] Gradient (B): 5% (0 min.).Math.100% (1.5 min.).Math.100% (2 min.) [1170] Flow rate: 1.0 mL/min. [1171] Detection: PDA 210 nm-400 nm [1172] Injection vol.: 1 μL

[1173] HPLC Method F [1174] Instrument: Waters Acquity UPLC/SQD2 [1175] Column: Ascentis Express C18 (2.7 μm, 2.1 mm×50 mm), Supelco [1176] Eluent: A) 0.1% FA/water, B) 0.1% FA/CH3CN [1177] Gradient (B): 5% (0 min.).Math.100% (1 min.).Math.100% (1.4 min.) [1178] Flow rate: 1.0 mL/min. [1179] Detection: 210 nm-400 nm [1180] Injection vol.: 1 μL

[1181] HPLC Method G [1182] Instrument: Waters Acquity UPLC/SQD2 [1183] Column: Ascentis Express C18 (2.7 μm, 2.1 mm×50 mm), Supelco [1184] Eluent: A) 0.1% FA/water, B) 0.1% FA/CH3CN [1185] Gradient (B): 5% (0 min.).Math.100% (1 min.).Math.100% (1.4 min.) [1186] Flow rate: 1.0 mL/min. [1187] Detection: 210 nm-400 nm [1188] Injection vol.: 1 μL

[1189] HPLC Method H [1190] Instrument: Waters Acquity UPLC/SQD [1191] Column: Ascentis Express C18 (2.7 μm, 2.1 mm×50 mm), Supelco [1192] Eluent: A) 0.1% FA/water, B) 0.1% FA/CH3CN [1193] Gradient (B): 5% (0 min.).Math.100% (4.5 min.).Math.100% (5.0 min.) [1194] Flow rate: 1.0 mL/min. [1195] Detection: 210 nm-400 nm [1196] Injection vol.: 2 μL

TABLE-US-00124 TABLE 125 Example HPLC Compound data No. method Compound MW m/z Retention time (min) 65 A 20b 410.47 411.18 ([M + H]+)    3.034 66 A 22c 339.39 118.01 ([M-Fmoc + H]+) 3.648 67 A 23b 438.52 439.24 ([M + H]+)    3.310 68 A 23c 353.42 354.16 ([M + H]+)    3.472 69 B 24b 422.48 423.19 ([M + H]+)    2.898 70 B 24c 337.38 116.01 ([M-Fmoc + H]+) 3.049 [00135][00136][00137]

G. Experiment of Comparison Between the TFA Method and the TMSOTf-HMDS Method in Resin Removal Reactions (Table 126)

[1197] ##STR00138##

Example 71

[1198] CTC Resin Removal and t-Bu Removal Reactions of Resin 25a (TFA Method)

[1199] 20.0 mg of Resin 25a was weighed into a reaction vessel, and the resin was swollen by adding 10 v/w of dichloromethane. After removing the dichloromethane from the reaction solution, 10 v/w % of TFA (10% dichloromethane solution) was added at room temperature and the vessel was shaken at 25° C. One hour after the shaking started, the reaction was analyzed by LCMS to find that the reaction conversion rate was 17%. 3.8% of a by-product due to amide bond cleavage was produced. The same reaction was performed for Resin 26a (Table 126).

Example 72

[1200] CTC Resin Removal and tBu Deprotection Reactions of Resin 25a (TMSOTf-HMDS Method)

[1201] 23.0 mg of Substrate 25a was weighed into a reaction vessel, and the resin was swollen by adding 10 v/w of 1,2-dichloroethane. After removing the 1,2-dichloroethane from the reaction solution, a solution of HMDS (10.7 eq.) and TMSOTf (7.1 eq.) in 10 v/w of 1,2-dichloroethane was added to the reaction vessel. After shaking at 25° C. for four hours, the reaction was analyzed by LCMS and the t-Bu ester was confirmed to disappear. The same reaction was performed for Resin 26a (Table 126).

TABLE-US-00125 TABLE 126 Amide bond Example Raw Reaction Conversion cleavage Substrate No. material Reagent time rate (LCMS Area %) 25a 71 20.0 mg 10% TFA 1 h  17% 3.8% in CH.sub.2Cl.sub.2 (10 v/w) 72 20.3 mg TMSOTf (7.1 eq.) 4 h 100%   0% HMDS (10.7 eq.) 26a 73 19.8 mg 10% TFA 1 h  13% 1.1% in CH.sub.2Cl.sub.2 (10 v/w) 74 20.0 mg TMSOTf (7.1 eq.) 3 h 100%   0% HMDS (14.2 eq.)

[1202] Identification of Products

TABLE-US-00126 TABLE 127 (HPLC method 1) Major peaks and retention times of products and target products Compound data Example No. Compound MW m/z Retention time (min) 71, 72 25b 454.48 477.10 ([M + Na]+)   2.649 25c 339.39 118.07 ([M-Fmoc + H]+) 3.560 73, 74 26b 496.56 497.24 ([M + H]+)    3.117 26c 367.45 146.00 ([M-Fmoc + H]+) 3.661 [00139][00140]
H. Synthesis of Raw Materials for Boc Removal, t-Bu Removal, and Resin Removal Reactions

Example 75

[1203] Synthesis of 3-Mer Raw Material for Boc Removal Reaction (Boc-Asp(OBn)-pip)

##STR00141##

[1204] The synthesis was performed according to Example 88.

TABLE-US-00127 TABLE 128 Weight (g) Yield (%) Raw material (27) 34.6 — Product (28) 37.6 90

TABLE-US-00128 TABLE 129 Analysis (HPLC method 1) MW m/z rt Purity LC A % Product (28) 390.48 291.19 ([M-Boc + H]+) 4.530 97.619

Example 76

[1205] Synthesis of 2-mer (Boc-MeVal-Asp(OBn)-pip)

##STR00142##

[1206] 5.714 g of the raw material was weighed into a reaction vessel and 29 ml of MeCN was added. After adding 2.3 ml of methanesulfonic acid, the oil bath was set at 45° C. and the reaction vessel was heated. After 70 minutes, the reaction vessel was brought back to room temperature and then immersed in an ice bath. To the reaction solution was added 14 ml of diisopropylethylamine, followed by addition of 4.046 g of Boc-MeVal-OH and 5.284 g of DMT-MM. One hour after the addition of the reagents, the reaction solution was concentrated. The reaction was quenched by adding ethyl acetate and 5% aqueous potassium carbonate to the concentrate. The organic layer was separated by liquid separation treatment and then washed twice with a 5% aqueous potassium carbonate solution, once with water, three times with a 5% aqueous potassium bisulfate solution, once with a 10% aqueous sodium chloride solution, and once with brine. The organic layer was concentrated under reduced pressure to give 7.055 g (96% yield) of a pale yellow solid.

TABLE-US-00129 TABLE 130 Weight (g) Yield (%) Raw material (28) 5.714 — Product (29) 7.055 96

TABLE-US-00130 TABLE 131 Analysis (HPLC method 1) MW m/z rt Purity LC A % Product (29) 503.64 526.29 ([M + Na]+) 5.178 96.889

Example 77

[1207] Synthesis of 3-mer (Boc-MePhe-MeVal-Asp(OBn)-pip)

##STR00143##

[1208] 7.055 g of the raw material was weighed into a reaction vessel and 35 ml of MeCN was added. After adding 2.2 ml of methanesulfonic acid, the oil bath was set at 45° C. and the reaction vessel was heated. After 70 minutes, the reaction vessel was brought back to room temperature and then the reaction solution was concentrated. After the concentration, 35 ml of ethyl acetate was added, the reaction vessel was immersed in an ice bath, and 9.466 g of sodium bicarbonate was then added. 35 ml of water was then added, followed by addition of 4.61 g of Boc-MePhe-OH and 7.73 g of DMT-MM. After six hours, the reaction was followed by LCMS to confirm that the raw material consumption rate was 99%. One hour after the analysis, liquid separation treatment was conducted by adding ethyl acetate and water to give an organic layer. The organic layer was washed three times with a 5% aqueous potassium carbonate solution, once with water, three times with a 5% aqueous potassium bisulfate solution, and once with brine. The organic layer was concentrated under reduced pressure and the resulting crude product was purified by silica gel column chromatography to provide 6.9757 g (75% yield) of the target product as a white solid.

TABLE-US-00131 TABLE 132 Weight (g) Yield (%) Raw material (29) 7.055 — Product (9a) 6.9757 75

TABLE-US-00132 TABLE 133 Analysis (HPLC method 1) MW m/z rt Purity LC A % Product (9a) 664.84 687.43 ([M + Na]+) 5.684 99.206

Example 78

[1209] Synthesis of Boc-MeGly-OAllyl

##STR00144##

[1210] 9.286 g of the raw material was weighed into a reaction vessel and 46 ml of DMF was added. The reaction vessel was cooled with ice, and 10.20 g of potassium carbonate and 4.15 ml of allyl bromide were then added. After 15 minutes, the reaction vessel was brought back to room temperature and then stirred overnight. To the reaction vessel was added MTBE, and the reaction was quenched with water. After liquid separation, the organic layer was washed once with brine. The organic layer was dried over magnesium sulfate, filtered, and concentrated under reduced pressure. The resulting crude product was purified by silica gel column chromatography to provide 10.083 g (90% yield) of the target product 31 as a transparent oily liquid.

TABLE-US-00133 TABLE 134 Weight (g) Yield (%) Raw material (30) 9.286 — Product (31) 10.083 90

Example 79

[1211] Synthesis of Boc-Thr(OBn)-MeGly-OAllyl

##STR00145##

[1212] Deprotection and condensation reactions were performed according to Example 76.

TABLE-US-00134 TABLE 135 Weight (g) Yield (%) Raw material (31) 6.5224 — Product (32) 13.5594 Quantitative

TABLE-US-00135 TABLE 136 Analysis (HPLC method 1) MW m/z rt Purity LC A % Product 420.51 321.10 ([M-Boc + H]+) 4.770 98.54 (Compound 32)

Example 80

[1213] Synthesis of Boc-MeLeu-Thr(OBn)-MeGly-OAllyl

##STR00146##

[1214] Deprotection and condensation reactions were performed by the same method as described above.

Example 80a

[1215] Synthesis of Teoc-MeLeu-OPfp by Esterification Reaction of Teoc-MeLeu-OH with Pentafluorophenol (Pfp-OH)

##STR00147##

926 mg of the raw material and 736 mg of pentafluorophenol (Pfp-OH) were respectively weighed into a reaction vessel, and 7.8 mL of isopropyl acetate was added. 767 mg of EDCI hydrochloride was added and the reaction solution was stirred at room temperature for two hours. The organic layer was washed twice with 8 mL of 0.5 N hydrochloric acid and twice with 8 mL of a 5% aqueous potassium carbonate solution, and 2 g of sodium sulfate was added to the organic layer. The solid was removed by filtration and the organic layer was concentrated under reduced pressure to give the target product as a transparent oily liquid. The obtained compound is used for the next reaction without purification.

TABLE-US-00136 TABLE 137 Analysis (HPLC method G) Retention time MW m/z (min) Product 455.16 428.3 ([M—C2H4 + H]+) 1.19 (Compound 80a)

Example 80b

[1216] Synthesis of Teoc-MeLeu-Thr(OBn)-MeGly-OAllyl

##STR00148##

[1217] Compound 80b can be synthesized by condensation reaction using the method of Meneses et al. (J. Org. Chem., 2010, 75, 564-569) in which the terminal protecting group of Compound 33 is removed and then reacted with pentafluorophenyl ester (Compound 80a) to form an amide bond.

TABLE-US-00137 TABLE 138 Weight (g) Yield (%) Raw material (33) 13.4338 — Product (8a) 15.5319 100

TABLE-US-00138 TABLE 139 Analysis (HPLC method 1) MW m/z rt Purity LC A % Product (8a) 660.41 561.28 ([M-Boc + H]+) 5.657 95.36

Example 81

[1218] Synthesis of Boc-MeAla-MePhe-Leu-MeLeu-Thr(OBn)-MeGly-OAllyl

##STR00149##

[1219] The reaction was performed according to Example 88.

TABLE-US-00139 TABLE 140 Weight (g) Yield (%) Raw material (4b) 5.0118 — Product (3a) 5.68 90

TABLE-US-00140 TABLE 141 Analysis (HPLC method 1) MW m/z rt Purity LC A % Product (3a) 906.55 807.58 ([M-Boc] + H)+ 5.974 93.06

Example 82

[1220] Synthesis of Boc-MeAla-MePhe-Leu-MeLeu-Thr(OBn)-MeGly-OH

##STR00150##

[1221] 1.1180 g of the raw material was weighed into a reaction vessel and 2.2 ml each of THF, methanol, and water were added. 102.8 mg of lithium hydroxide monohydrate was then added and the reaction solution was stirred at room temperature for five hours. The completion of the reaction was confirmed by LC, and the reaction solution was then concentrated under reduced pressure. After adding ethyl acetate to the concentrate, 0.5 N hydrochloric acid was added to quench the reaction. The organic layer was separated by liquid separation treatment and then washed twice with 0.5 N hydrochloric acid and twice with a 5% aqueous sodium chloride solution. The solvent was evaporated by concentration under reduced pressure to give 918.8 mg (86% yield) of a white solid.

TABLE-US-00141 TABLE 142 Weight (g) Yield (%) Raw material (3a) 1.1180 — Product (34) 0.9188 86

TABLE-US-00142 TABLE 143 Analysis (HPLC method 1) MW m/z rt Purity LC A % Product (34) 866.52 767.5 ([M-Boc] + H)+ 5.361 92.03

Example 83

[1222] Synthesis of Fmoc-Asp(OtBu)-pip

##STR00151##

[1223] The reaction was performed according to Example 88.

TABLE-US-00143 TABLE 144 Weight (g) Yield (%) Raw material (11) 40 — Product (12) 47 Quantitative

TABLE-US-00144 TABLE 145 Analysis (HPLC method 1) MW m/z rt Purity LC A % Product (12) 478.25 423.24 ([M-tBu] + H)+ 4.019 99.318

Example 84

[1224] Synthesis of FmocAsp(OH)pip

##STR00152##

[1225] 45.4 g of the raw material was weighed into a reaction vessel and 454 ml of trifluoroethanol was added. 24 ml of chlorotrimethylsilane was then added and the reaction solution was stirred at room temperature. Two and a half hours after the start of the reaction, 910 ml of water was added in two portions and crystals were allowed to precipitate. The crystals were filtered and then dried to give 25.8 g (61% yield) of the target product 36.

TABLE-US-00145 TABLE 146 Weight (g) Yield (%) Raw material (35) 40 — Product (36) 25.8 61

TABLE-US-00146 TABLE 147 Analysis (HPLC method 1) MW m/z rt Purity LC A % Product (36) 422.18 423.21 ([M] + H)+ 3.101 98.259

Example 85

[1226] Loading of Fmoc-Asp(OH)-pip onto CTC Resin

##STR00153##

[1227] The same loading step as in the method described in Example 112 was conducted. The product was analyzed after 5-mer elongation and the following resin removal step.

TABLE-US-00147 TABLE 148 Weight (g) Raw material (36) 4.09

Example 86

[1228] Solid-Phase Synthesis (2-mer Elongation)

##STR00154##

[1229] The reactions were performed in a solid-phase synthesis column. [1230] 1) Deprotection step: To Fmoc-MeAla-OCTC resin (37, theoretical amino acid loading amount: 9.49 mmol) was added DMF (104 mL). A 20% solution of piperidine in DMF (104 mL) was added at 30° C., and the column was shaken at 30° C. for 15 minutes. After the solution was discharged, a 20% solution of piperidine in DMF (104 mL) was added again and the column was shaken at 30° C. for 15 minutes. After the solution was discharged, the resin was washed with DMF (130 ml) for 2 min×7. [1231] 2) Elongation step: In a reaction vessel separate from the solid-phase column, 6.62 g of Fmoc-MeVal-OH and 2.93 g of Oxyma were dissolved in 52 ml of DMF, and 5.87 ml of DIC was then added. The vessel was shaken at 30° C. for 30 minutes. This solution was added to the solid-phase synthesis column, which was then shaken at 30° C. for two hours. After the solution was discharged, the resin was washed with DMF (130 ml) for 2 min×6.

Example 87

[1232] Solid-Phase Synthesis (Sequential Elongation from 2-mer to 5-mer and Resin Removal Reaction)

##STR00155##

[1233] Deprotection and elongation steps were repeated according to the method described in Example 86 to provide Boc-MeIle-Ala-MePhe-MeVal-Asp(OCTC)-pip. To the resulting resin was added 147 ml of a mixed solvent of diisopropylethylamine, trifluoroethanol, and methylene chloride (2% diisopropylethylamine, solvent volume ratio 1:1), followed by shaking at 30° C. for two and a half hours. The discharged and resin-removed peptide solution was collected. After the resin removal solution was discharged, the resin was washed twice with 25 ml of a mixed solvent of trifluoroethanol and methylene chloride (volume ratio 1:1), and the solutions were concentrated. The concentrate was dissolved in 130 ml of ethyl acetate and washed twice with an aqueous potassium carbonate solution, twice with potassium bisulfate, and once with an aqueous sodium chloride solution, and the organic layer was then concentrated under reduced pressure. After drying, 4.9821 g (69% yield) of a white solid was obtained.

TABLE-US-00148 TABLE 149 Weight (g) Yield (%) Fmoc-MeAsp(OH)-pip (37) 4.09 — Product (38) 4.9821 69

TABLE-US-00149 TABLE 150 Analysis (HPLC method 1) MW m/z rt Purity LC A % Product (38) 772.47 795.5 ([M + Na]+) 4.856 96.471

Example 88

[1234] Synthesis of Fmoc-MeAsp(OtBu)-pip

##STR00156##

[1235] 49.6 g of EDCI hydrochloride was weighed into a reaction vessel and 400 ml of DMF was added. The reaction vessel was cooled to 0° C., and 100.050 g of Fmoc-MeAsp(OtBu)-OH was then added. Next, a solution of 40.1 g of Oxyma in 100 ml of DMF was added, followed by 34.9 ml of piperidine, using a dropping funnel. Four hours and 30 minutes after the dropwise addition of piperidine was completed, the reaction was analyzed by LCMS and the raw material was confirmed to disappear. The reaction solution was diluted with ethyl acetate, and 500 ml of 0.5 N hydrochloric acid was then added to quench the reaction. The organic layer was separated and then washed twice with water, twice with a 5% aqueous sodium carbonate solution, and twice with a 5% aqueous sodium chloride solution. The resulting organic layer was concentrated under reduced pressure. The resulting crude product was recrystallized from ethyl acetate and heptane to give 86.230 g (75% yield) of the target product.

TABLE-US-00150 TABLE 151 Analysis (HPLC method 1) MW m/z rt Purity LC A % Product 492.26 493.24 ([M + H]+) 5.527 98.268 (Compound 11)

Example 89

[1236] Synthesis of Synthetic Intermediate (Cbz-Asp(OtBu)-pip (41)) for 11-Residue Peptide (tBu form, Cbz-MeAla-MePhe-Leu-MeLeu-Val-MeGly-MeIle-Ser(OtBu)-MePhe-MeVal-Asp(OtBu)-pip (22))

##STR00157##

[1237] 17.5 g of Cbz-Asp(OtBu)-OH was weighed into a reaction vessel and 175 mL of ethyl acetate was added. After cooling the reaction solution to 0° C., piperidine (3 eq.), diisopropylethylamine (6 eq.), and a 1.7 M solution of T3P in ethyl acetate (3 eq.) were added, respectively. The reaction solution was warmed to room temperature and stirred at room temperature for 10 minutes, and 175 mL of a 5% aqueous potassium carbonate solution was then added. The aqueous layer was removed, and the organic layer was then washed twice with 175 mL of a 5% aqueous potassium bisulfate solution. The resulting organic layer was concentrated and dried under reduced pressure to afford 20 g of Cbz-Asp(OtBu)-pip (41) in 100% yield.

Example 90

Synthesis of H-Asp(OtBu)-pip (42)

[1238] ##STR00158##

[1239] 9.5 g of Cbz-Asp(OtBu)-pip (41) was weighed into each of two reaction vessels, and 50 mL of CPME was added to each vessel. 10% Pd/C (20 w/w %) was added to both vessels and then reacted at 30° C. under a 3 bar hydrogen atmosphere. Three hours after the start of the reaction, the two reaction solutions were mixed, filtered, and washed with 100 mL of CPME. The resulting mixture was concentrated and dried under reduced pressure to afford 12.3 g of H-Asp(OtBu)-pip (42) in 99% yield.

[1240] 11-mer (22) was synthesized from H-Asp(OtBu)-pip (42) by the 19 steps described below. The synthesized intermediates are provided as follows.

TABLE-US-00151 (42) H-Asp(OtBu)-pip (43) Cbz-MeVal-Asp(OtBu)-pip (44) H-MeVal-Asp(OtBu)-pip (45) Cbz-MePhe-MeVal-Asp(OtBu)-pip (46) H-MePhe-MeVal-Asp(OtBu)-pip (13) Cbz-Ser(OtBu)-MePhe-MeVal-Asp(OtBu)-pip (47) H-Ser(OtBu)-MePhe-MeVal-Asp(OtBu)-pip (14) Cbz-Melle-Ser(OtBu)-MePhe-MeVal-Asp(OtBu)-pip (48) H-Melle-Ser(OtBu)-MePhe-MeVal-Asp(OtBu)-pip (49) Cbz-MeGly-Melle-Ser(OtBu)-MePhe-MeVal-Asp(OtBu)-pip (50) H-MeGly-Melle-Ser(OtBu)-MePhe-MeVal-Asp(OtBu)-pip (15) Cbz-Val-MeGly-Melle-Ser(OtBu)-MePhe-MeVal- Asp(OtBu)-pip (51) H-Val-MeGly-Melle-Ser(OtBu)-MePhe-MeVal-Asp(OtBu)- pip (16) Cbz-MeLeu-Val-MeGly-Melle-Ser(OtBu)-MePhe-MeVal- Asp(OtBu)-pip (52) H-MeLeu-Val-MeGly-Melle-Ser(OtBu)-MePhe-MeVal- Asp(OtBu)-pip (17) Cbz-Leu-MeLeu-Val-MeGly-Melle-Ser(OtBu)-MePhe- MeVal-Asp(OtBu)-pip (53) H-Leu-MeLeu-Val-MeGly-Melle-Ser(OtBu)-MePhe-MeVal- Asp(OtBu)-pip (18) Cbz-MePhe-Leu-MeLeu-Val-MeGly-Melle-Ser(OtBu)- MePhe-MeVal-Asp(OtBu)-pip (54) H-MePhe-Leu-MeLeu-Val-MeGly-Melle-Ser(OtBu)-MePhe- MeVal-Asp(OtBu)-pip (19) Cbz-MeAla-MePhe-Leu-MeLeu-Val-MeGly-Melle- Ser(OtBu)-MePhe-MeVal-Asp(OtBu)-pip

Example 91

Elongation of Peptide Chain (Elongation Method A)

[1241] 8.6 g of 42 was weighed into a reaction vessel and 108 mL of CPME was added. Cbz-MeVal-OH (1.1 eq.) and diisopropylethylamine (3 eq.) were added, and a solution of BEP (1.5 eq.) in 21.5 mL of MeCN was then added. After stirring at room temperature for three minutes, 15 mL of a 10% aqueous sodium bisulfate solution was added. The aqueous layer was removed, and 15 mL of a 5% aqueous potassium carbonate solution and trimethylamine hydrochloride (3 eq.) were then added to the organic layer at room temperature. The mixture was warmed to 40° C. and then stirred at 40° C. for 90 minutes. After cooling to room temperature, the aqueous layer was removed and the resulting organic layer was then washed with 15 mL of a 5% aqueous potassium carbonate solution. The resulting organic layer was concentrated and dried under reduced pressure to give 17 g of 43 quantitatively.

Example 92

[1242] Removal of N-Terminal Cbz (Deprotection Method A)

[1243] 9.5 g of 43 was weighed into each of two reaction vessels, and 50 mL of CPME was added to each vessel. 10% Pd/C (20 w/w %) was added to both vessels and then reacted at 35° C. under a 3 bar hydrogen atmosphere. Two hours after the start of the reaction, the two reaction solutions were mixed, filtered, and washed with 100 mL of CPME. The resulting mixture was concentrated and dried under reduced pressure to give 14 g of 44 in 100% yield.

Example 93

[1244] Elongation of Peptide Chain (Elongation Method B)

[1245] 14 g of 44 was weighed into a reaction vessel, and 126 mL of CPME and 14 mL of acetonitrile were added, respectively. Cbz-MePhe-OH (1.1 eq.), diisopropylethylamine (8 eq.), and a 1.7 M solution of T3P in ethyl acetate (3 eq.) was sequentially added at room temperature. After stirring at room temperature for one hour, 140 mL of a 5% aqueous potassium bisulfate solution was added. The aqueous layer was removed, and 140 mL of a 5% aqueous potassium carbonate solution and trimethylamine hydrochloride (3 eq.) were then added at room temperature. After stirring at room temperature for 30 minutes, the aqueous layer was removed. The resulting organic layer was then washed with 140 mL of a 5% aqueous potassium carbonate solution. The resulting organic layer was concentrated and dried under reduced pressure to give 24.1 g of 45 in 96% yield.

Example 94

[1246] Removal of N-Terminal Cbz (Deprotection Method B)

[1247] 9.2 g of 15a was weighed into each of two reaction vessels, and 46 mL of CPME was added to each vessel. 10% Pd/C (20 w/w %) was added to both vessels and then reacted at 35° C. under a 5 bar hydrogen atmosphere. Six hours after the start of the reaction, the reaction vessels were cooled to room temperature and stored at room temperature overnight under an air atmosphere. Further reaction was performed at 45° C. for four hours under a 5 bar hydrogen atmosphere. The two reaction solutions were mixed, filtered, and washed with 92 mL of CPME. The resulting mixture was concentrated and dried under reduced pressure to give 15.9 g of 51 in 98% yield.

[1248] The peptides of 2-mer to 11-mer were synthesized by the same technique. The reaction conditions different from those described above, except for the reaction time and the mass of the raw material used, are further described in the column “Reaction method.”

TABLE-US-00152 TABLE 152 (HPLC method 1) Example Raw Reaction Product Purity No. Product Reaction method material time amount Yield (LCMS Area %) 91 43 Elongation A 8.6 g 3 min 17 g Quantitative 99.7% 92 44 Deprotection A 9.5 g × 2 2 h 14 g 100%  ND 93 45 Elongation B 14 g 30 min 24.1 g 96% 99.6% 95 46 Deprotection A 11.5 g × 2 2 h 18.1 g 99% ND 96 13 Elongation B 17.3 g 15 min 26.5 g Quantitative 98.9% 97 47 Deprotection A 12 g × 2 4 h 19.5 g 97% ND 98 14 Elongation A 16 g 5 min 22.2 g 100%  99.4% 99 48 Deprotection A 9.5 g × 2 2 h 15.6 g 96% ND 100 49 Elongation B 15.3 g 15 min 19.5 g Quantitative 99.6% 101 50 Deprotection A 9.5 g × 2 3 h 16.3 g 99% ND (5 bar) 102 15 Elongation B 16 g 30 min 20 g 99% 99.6% 94 51 Deprotection B 9.2 g × 2 10 h 15.9 g 98% ND 103 16 Elongation A 14.5 g 1 min 18.0 g 98% 96.0% (40° C.) 104 52 Deprotection A 8 g × 2 4 h 14.3 g 100%  ND (45° C., 5 bar) 105 17 Elongation B 13 g 30 min 15.6 g 98% 97.2% 106 53 Deprotection A 10 g 4 h 8.9 g 99% ND (45° C., 5 bar) 107 18 Elongation A 7 g 3 min 8.6 g 99% 97.0% 108 54 Deprotection A 7.6 g 4 h 6.8 g 98% ND (45° C., 5 bar) 109 19 Elongation B 500 mg 2 h 555 mg 96% 95.3%

[1249] Among the synthesized intermediates, 13a, 14a, 15a, 16a, 17a, 18a, and 19a were used for tBu removal experiments. The compounds used for tBu removal reactions were analyzed by LCMS as described in Table 153.

TABLE-US-00153 TABLE 153 Analysis Compound data Example HPLC Retention time No. Compound Method MW m/z (min) 46 13a 1 808.03 808.49 ([M + H]+) 5.938 48 14a 1 935.22 935.59 ([M + H]+) 6.233 50 15a 1 1105.43 1105.69 ([M + H]+) 5.924 52 16a 1 1232.62 1232.79 ([M + H]+) 6.374 54 17a 1 1345.78 1369 ([M + Na]+) 6.727 56 18a 1 1506.98 1530 ([M + Na]+) 7.096 58 19a 1 1592.09 1615 ([M + Na]+) 6.987
I. Synthesis of C-Terminal tBu-Protected Dipeptides

##STR00159##

Example 110

[1250] Synthesis of Fmoc-MeAla-MeAla-OtBu (20a)

[1251] 99.7 mg of H-MeAla-OtBu hydrochloride and 710 mg of dipotassium hydrogenphosphate were weighed in a reaction vessel, respectively, and 10 v/w each of 2-MeTHF and water relative to H-MeAla-OtBu hydrochloride were added. Fmoc-MeAla-OH (167 mg) and DMT-MM (211 mg) were sequentially added at 0° C. and the reaction solution was then warmed to room temperature. After shaking at room temperature for 16 hours, the reaction was analyzed by LCMS and Fmoc-MeAla-OH was confirmed to disappear. The reaction solution was transferred to a separatory funnel using 4 mL of water and 6 mL of 2-MeTHF, and the aqueous layer was then removed. The resulting organic layer was washed with brine (3 mL), 15% aqueous sodium bisulfate (3 mL), and 5% aqueous sodium carbonate (3 mL), respectively, and the solvent was then removed by concentration under reduced pressure. The resulting crude product was purified by silica gel column chromatography using the conditions described below, and the fractions containing the target product were then concentrated and dried under reduced pressure to provide 227.6 mg of 20a in a yield of 96% and a purity of 99.5%.

Example 111

[1252] Synthesis of Fmoc-MePhe-MeAla-OtBu(21a)

[1253] H-MeAla-OtBu hydrochloride (99.8 mg) and Fmoc-MePhe-OH (205 mg) were weighed into a reaction vessel, respectively, and 8 v/w of isopropyl acetate and 2 v/w of acetonitrile relative to H-MeAla-OtBu hydrochloride as well as diisopropylethylamine (4 eq.) were sequentially added. A 1.7 M solution of T3P in ethyl acetate (2.5 eq.) was added at room temperature. After stirring at room temperature for four hours, a 1.7 M solution of T3P in ethyl acetate (0.5 eq.) and diisopropylethylamine (0.8 eq.) were added. 30 minutes after the addition of the reagents, the reaction was analyzed by LCMS to find that the reaction conversion rate was 98.5%. NMI (2 eq.) was added at room temperature, and the reaction solution was then stirred at 50° C. for five minutes. The reaction solution was transferred to a separatory funnel using 6 mL of ethyl acetate, and the aqueous layer was then removed. The resulting organic layer was washed with a 5% aqueous sodium carbonate solution (3 mL), a 5% aqueous potassium bisulfate solution (3 mL), and a 5% aqueous sodium carbonate solution (3 mL), respectively. The organic layer was dehydrated with sodium sulfate for 30 minutes, then filtered, and concentrated under reduced pressure to remove the solvent. The resulting crude product was purified by silica gel column chromatography using the conditions described below, and the fractions containing the target product were then concentrated and dried under reduced pressure to provide 227.6 mg of 21a in a yield of 96% and a purity of 99.5%.

TABLE-US-00154 TABLE 154 Analysis (HPLC method 1) Compound data Example Retention time No. Compound MW m/z (min) 110 20a 466.58 489.2 ([M + Na]+) 5.243 111 21a 542.68 543.2 ([M + H]+) 5.754

J. Synthesis of CTC Resin-Loaded Dipeptides

[1254] ##STR00160##

Example 112

[1255] Loading of Fmoc-MeAla-OH onto CTC Resin

[1256] 4.47 g of Cl-CTC resin was weighed into a reaction column, dichloromethane (36 mL) was added, and the column was shaken at 30° C. for 60 minutes. After discharging the dichloromethane, a solution of Fmoc-MeAla-OH (1.24 g) and diisopropylethylamine (1.4 mL) in dichloromethane (36 mL) was added at room temperature. After stirring at 30° C. for three hours, the reaction was analyzed by LCMS to find that the reaction conversion rate was 96.0%. After discharging the reaction solution, a solution of methanol (3.6 mL) and diisopropylethylamine (1.8 mL) in DMF (30 mL) was added at room temperature. After shaking at 30° C. for 1.5 hours, the reaction solution was discharged. The resin was washed with 36 mL of DMF four times and then dried under reduced pressure to give 4.80 g of 55. 56 was also synthesized by the same technique. The products were analyzed in the next step.

TABLE-US-00155 TABLE 155 Analysis (HPLC method 1) [00161] [00162] [00163] [00164] [00165] [00166] [00167] [00168] [00169] [00170] Reaction Example No. Product CI-CTC resin Amino acid conversion rate Product amount 112 55 4.47 g 1.24 g 96% 4.80 g 113 10 10.0 g 3.41 g 99% 12.4 g

Example 114

[1257] Synthesis of Fmoc-MeAla-MeAla-OCTC (22a), Fmoc-MeVal-MeAla-OCTC (23a), and Fmoc-Pro-MeAla-OCTC (24a)

[1258] Dipeptides 22a to 24a were simultaneously synthesized using a solid-phase peptide synthesizer Prelude X. 676 mg, 687 mg, and 705 mg of resin Fmoc-MeAla-OCTC (55) were weighed into three reaction vessels, respectively. DMF (8 mL) was added thereto, and the resin was swollen by allowing to stand at room temperature for one hour. After the DMF was discharged, a 20% solution of piperidine in DMF (8 mL) was added and the vessels were shaken at room temperature for 15 minutes. After the solution was discharged, a 20% solution of piperidine in DMF (8 mL) was added again and the vessels were shaken at room temperature for 15 minutes. The cocktails 1 to 3 described below were added to RV 1 to RV 3, respectively, and a 12.5% solution of DIC in DMF (4 eq.) was then added. The vessels were shaken at room temperature for three hours with nitrogen bubbling. After discharging the solution, the resin was washed with DMF (8 v/w) for 2 min×5 and with MTBE (8 v/w) for 2 min×4, which each wash was carried out by shaking at room temperature. The resulting resin was dried under reduced pressure to provide 30a, 31a, and 32a, respectively. The obtained compounds were identified by the resin removal reactions in Examples 65, 67, and 69.

[1259] About 20 mg each of the obtained three resins were weighed into three reaction vessels, a 20% solution of piperidine in DMF (100 mL) was added to each vessel, and the reaction solution was stirred at room temperature for two hours. Dibenzofulvene was quantitatively determined from the absorbance of the solution to calculate the loading rate of each dipeptide (Table 156).

Example 115

[1260] Synthesis of Fmoc-MeAla-MeAsp(OtBu)-OCTC (25a) and Fmoc-MeLeu-MeAsp(OtBu)-OCTC (26a)

[1261] Dipeptides 25a to 26a were simultaneously synthesized using a solid-phase peptide synthesizer Prelude X. 340 mg of resin Fmoc-MeAsp(OtBu)-OCTC (10) was weighed into each of two reaction vessels. DMF (8 mL) was added thereto, and the resin was swollen by allowing to stand at room temperature for 30 minutes. After the DMF was discharged, a 20% solution of piperidine in DMF (8 v/w) was added and the vessels were shaken at room temperature for 5 minutes. After the solution was discharged, a 20% solution of piperidine in DMF (8 v/w) was added again and the vessels were shaken at room temperature for 20 minutes. After discharging the solution, the cocktails 1 and 2 described below were added to the corresponding reaction vessels, respectively, a 12.5% solution of DIC in DMF (4 eq.) was then added, and the vessels were shaken at room temperature for three hours with nitrogen bubbling. After discharging the solution, the resin was washed with DMF (8 v/w) for 2 min×4 and with MTBE (8 v/w) for 2 min×4, which each wash was carried out by shaking at room temperature. The resulting resin was dried under reduced pressure to provide 994 mg of 25a and 1.00 g of 26a, respectively. The obtained compounds were identified by the resin removal reactions in Examples 71 and 73.

[1262] About 20 mg each of the obtained two resins were weighed into two reaction vessels, a 20% solution of piperidine in DMF (100 mL) was added to each vessel, and the reaction solution was stirred at room temperature for two hours. Dibenzofulvene was quantitatively determined from the absorbance of the solution to calculate the loading rate of each dipeptide (Table 156).

TABLE-US-00156 TABLE 156 Analysis (HPLC method 1) Example Reaction No. Product Raw material conversion rate Loading rate 114 22a 676 mg 100% 0.41 mmol/g 23a 687 mg 100% 0.31 mmol/g 24a 705 mg 100% 0.42 mmol/g 115 25a 340 mg 100% 0.27 mmol/g 26a 340 mg 100% 0.27 mmol/g

Example 116

[1263] Synthesis of Fmoc-MeAsp(OAllyl)-mor

##STR00171##

[1264] Fmoc-MeAsp(OAllyl)-OH (56, 87.9 g) was added to a reaction vessel and dissolved in DMF (430 ml). HOBt (31.9 g) and EDCI hydrochloride (49.4 g) were then added at ambient temperature, and the reaction solution was cooled to 0° C. Morpholine (20.4 ml) was gradually added to the reaction solution, which was then stirred at 0° C. for 45 minutes. Water (180 ml) was added at 0° C. to the reaction solution, which was then stirred at ambient temperature for one hour. Water (180 ml) was further added and the reaction solution was stirred at ambient temperature for 1.75 hours. The generated solid was collected by filtration using a Kiriyama funnel, and the resulting solid was washed twice with water (450 ml). The solid after washing with water was dried under reduced pressure to afford 86.8 g (85% yield) of the target product as a colorless solid.

Example 117

[1265] Synthesis of Fmoc-MeAsp(OAllyl)—NMe2

##STR00172##

[1266] Under a nitrogen stream, EDCI hydrochloride (27.4 g) and DMF (217 ml) were added to a reaction vessel, and HOBt (17.7 g) and a solution of Fmoc-MeAsp(OAllyl)-OH (56, 48.8 g) in dichloromethane-DMF (90 ml-90 ml) were then added at 0° C. After stirring the reaction solution at 0° C. for 30 minutes, a dimethylamine-THF solution (2 N, 65.6 ml) was added dropwise over two minutes to the reaction solution, which was then stirred at 0° C. for 30 minutes. The reaction solution was diluted with ethyl acetate (488 ml), and the organic layer was washed with 1 N hydrochloric acid (twice with 391 ml), water (488 ml), a 5% aqueous sodium bicarbonate solution (twice with 488 ml), and a 18% aqueous sodium chloride solution (488 ml) and then dried over sodium sulfate. The drying agent was removed by filtration and then the filtrate was concentrated under reduced pressure to give 51.2 g (98% yield) of the target product as a colorless oil.

Example 118

[1267] Synthesis of Fmoc-MeAsp(OH)-mor

##STR00173##

[1268] Under a nitrogen stream, Fmoc-MeAsp(OAllyl)-mor (57, 22.8 g) and dichloromethane (50 ml) were added to a reaction vessel, and tetrakistriphenylphosphine palladium (0.55 g) was then added at ambient temperature. Phenylsilane (3.61 g) was added dropwise and the reaction solution was then stirred at ambient temperature for 30 minutes. The reaction solution was diluted with MTBE (228 ml) and then extracted with a 5% aqueous sodium bicarbonate solution (228 ml). The aqueous layer was acidified to about pH 3 with a 85% aqueous phosphoric acid solution (12 ml) and extracted with MTBE (228 ml). The organic layer was washed with a 18% aqueous sodium chloride solution (twice with 228 ml) and then dried over sodium sulfate. The drying agent was removed by filtration and then the filtrate was concentrated under reduced pressure to give 20.3 g (97% yield) of the target product as colorless amorphous crystals.

Example 119

[1269] Synthesis of Fmoc-MeAsp(OH)—NMe2

##STR00174##

[1270] Under a nitrogen stream, Fmoc-MeAsp(OAllyl)-NMe2 (58, 32.0 g) and tetrakistriphenylphosphine palladium (0.847 g) were added to a reaction vessel, and dichloromethane (73.3 ml) was then added. Phenylsilane (5.55 g) was added dropwise to the reaction solution, which was then stirred at ambient temperature for 30 minutes. The reaction solution was diluted with MTBE (320 ml) and then extracted with a 5% aqueous sodium bicarbonate solution (308 ml). The aqueous layer was acidified to about pH 2 with a 85% aqueous phosphoric acid solution (30.1 ml) and extracted with MTBE (320 ml). The organic layer was washed with a 18% aqueous sodium chloride solution (twice with 320 ml) and then dried over sodium sulfate. The drying agent was removed by filtration and then the filtrate was concentrated under reduced pressure to give 25.1 g (86% yield) of the target product as pale brown amorphous crystals.

TABLE-US-00157 TABLE 157 Analysis Compound data Example Retention time No. Compound HPLC Method MW m/z (min) 116 57 HPLC method D 478.21 479 ([M + H]+) 2.570 117 58 HPLC method E 436.20 437 ([M + H]+) 1.262 118 59 HPLC method F 438.18 439 ([M + H]+) 0.670 119 60 HPLC method G 396.17 397 ([M + H]+) 0.680

Example 120

[1271] Synthesis of Fmoc-MeAsp(OtBu)-mor (61)

##STR00175##

[1272] Under a nitrogen stream, Fmoc-MeAsp(OtBu)-OH (39, 3.0 g) and Oxyma (1.1 g) were added to a reaction vessel, and dimethylformamide (15 mL) was then added. 1-(3-Dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (1.5 g) and morpholine (0.74 mL) were added to the reaction solution while maintaining the temperature of the reaction solution at 10° C. or lower, and it was stirred for three hours. AcOEt (15 mL) and 0.5 N hydrochloric acid (15 mL) were added to the reaction solution. The organic layer was washed with water (15 mL) and further washed with a 5% aqueous sodium carbonate solution (15 mL). The organic layer was washed with a 5% aqueous sodium chloride solution (15 mL) and then concentrated under reduced pressure to afford 3.50 g (100%) of the target product as yellow amorphous crystals. The obtained compound was used for the next reaction without purification.

Example 121

[1273] Synthesis of Fmoc-MeAsp(OtBu)—NMe2 (62)

##STR00176##

[1274] Under a nitrogen stream, Fmoc-MeAsp(OtBu)-OH (39, 3.0 g) and Oxyma (1.1 g) were added to a reaction vessel, and dimethylformamide (15 mL) was then added. 1-(3-Dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (1.5 g), dimethylamine hydrochloride (0.69 g), and N,N-diisopropylethylamine (1.5 ml) were added to the reaction solution while maintaining the temperature of the reaction solution at 10° C. or lower, and it was stirred for three hours. To the reaction solution was added ethyl acetate (15 mL), followed by 0.5 N hydrochloric acid (15 mL). The organic layer was washed with water (15 mL) and further washed with a 5% aqueous sodium carbonate solution (15 mL). The organic layer was washed with a 5% aqueous sodium chloride solution (15 mL) and then concentrated under reduced pressure to afford 3.17 g (95%) of the target product as a colorless oily substance. The obtained compound was used for the next reaction without purification.

Example 122

[1275] Synthesis of Fmoc-MeAsp(OH)-mor (59)

##STR00177##

[1276] Under a nitrogen stream, Fmoc-MeAsp(OtBu)-mor (61, 1.8 g) and 2-methyltetrahydrofuran (9.2 mL) were added to a reaction vessel, and HMDS (0.86 mL) was then added. The reaction vessel was cooled to 0° C., TMSOTf (0.81 mL) was added, and the reaction solution was stirred at 25° C. for one hour. The reaction vessel was cooled to 0° C., a 5% aqueous potassium dihydrogenphosphate solution (9.2 mL) was added to the reaction solution, and the organic layer was separated. A 5% aqueous sodium carbonate solution (9.2 mL) was added, and the aqueous layer was separated. MTBE (9.2 mL) and 2 N hydrochloric acid (5.0 mL) were added to the aqueous layer, and the organic layer was separated. The organic layer was washed with a 5% aqueous potassium dihydrogenphosphate solution (9.2 mL) and then with a 10% aqueous sodium chloride solution (9.2 mL). The organic layer was concentrated under reduced pressure to afford 1.31 g (97%) of the target product as yellow amorphous crystals.

Example 123

[1277] Synthesis of Fmoc-MeAsp(OH)—NMe2 (60)

##STR00178##

[1278] Under a nitrogen stream, Fmoc-MeAsp(OtBu)—NMe2 (62, 1.6 g) and 2-methyltetrahydrofuran (7.9 mL) were added to a reaction vessel, and HMDS (0.80 mL) was then added. The reaction vessel was cooled to 0° C., TMSOTf (0.75 mL) was added, and the reaction solution was stirred at 25° C. for one hour. A 5% aqueous potassium dihydrogenphosphate solution (7.9 mL) was added to the reaction solution, and the organic layer was separated. A 5% aqueous sodium carbonate solution (7.9 mL) was added, and the aqueous layer was separated. MTBE (7.9 mL) and 2 N hydrochloric acid (5.0 mL) were added to the aqueous layer, and the organic layer was separated. The organic layer was washed with a 5% aqueous potassium dihydrogenphosphate solution (7.9 mL) and then with a 10% aqueous sodium chloride solution (7.9 mL). The organic layer was concentrated under reduced pressure to afford 1.31 g (95%) of the target product as a colorless oily substance.

TABLE-US-00158 TABLE 158 Analysis Compound data Example Retention time No. Compound HPLC Method MW m/z (min) 120 61 HPLC method H 494.24 495.6 ([M + H]+) 2.75 121 62 HPLC method H 452.23 453.6 ([M + H]+) 2.77 122 59 HPLC method H 438.18 439.5 ([M + H]+) 1.84 123 60 HPLC method H 396.17 397.5 ([M + H]+) 1.89

INDUSTRIAL APPLICABILITY

[1279] The present invention provides methods of producing peptide compounds by novel deprotection and/or resin removal methods using silylating agents. The present invention can provide industrially applicable and efficient techniques of peptide synthesis.