SYNTHESIS OF RAS INHIBITORS

Abstract

The present invention relates to Ras inhibitors, intermediates in the synthesis thereto, and methods for preparing the Ras inhibitors and the intermediates.

Claims

1. A method of separating enantiomers of a compound of Formula I ##STR00300## the method comprising the steps of: a) contacting the compound of Formula I with a chiral acid to form a diastereomeric salt of the compound of Formula I; and b) separating each diastereomer of the diastereomeric salt, thereby separating enantiomers of a compound of Formula I wherein n and m are each, independently, 0, 1, 2, 3, 4, or 5; PG is a nitrogen protecting group; and X.sup.1, X.sup.2, and X.sup.3 are each, independently, CH.sub.2, CHF, CF.sub.2, C(O), or O.

2. The method of claim 1, wherein the chiral acid comprises a chiral carboxylic acid.

3. The method of claim 1 or 2, wherein the separating step (b) comprises recrystallization.

4. The method of any one of claims 1 to 3, wherein the compound of Formula I is further described by Formula Ia: ##STR00301##

5. A barium salt of a compound of Compound A: ##STR00302##

6. The barium salt of claim 5, wherein the barium salt is a 2:1 (compound A: barium) salt.

7. A method of preparing the barium salt of claim 5 or 6, the method comprising the steps of: a) condensing a compound of Formula IIa and Compound C to form a compound of Formula IIb: ##STR00303## b) cyclizing the compound of Formula IIb and a compound of Formula IIc to form a compound of Formula IId: ##STR00304## c) hydrolyzing the compound of Formula IId to form a compound of Formula IIe: ##STR00305## d) methylating the compound of Formula IIe to form a compound of Formula IIf: ##STR00306## and e) hydrolyzing the compound of Formula IIf to form the barium salt of Compound A: ##STR00307## wherein R.sup.2 and R.sup.3 are each, independently, optionally substituted C.sub.1-C.sub.6 alkyl, optionally substituted C.sub.3-6 cycloalkyl, or optionally substituted C.sub.6-C.sub.10 aryl, and LG is a leaving group selected from the group consisting of halogen, triflate, mesylate, and tosylate.

8. The method of claim 7, wherein the methylating step (d) comprises contacting the compound of Formula IIe with MeB(OH).sub.2.

9. The method of claim 7 or 8, wherein the hydrolyzing step (e) comprises contacting the compound of Formula IIf with barium hydroxide.

10. A method of preparing a compound, or a salt thereof, of Formula III: ##STR00308## the method comprising the steps of: a) coupling a compound of Formula IIId with a compound of Formula I to form a compound of Formula IIIe: ##STR00309## and b) deprotecting the compound of Formula IIIe to form the compound of Formula III: ##STR00310## wherein n and m are each, independently, 0, 1, 2, 3, 4, or 5; PG is a nitrogen protecting group; each R.sup.4 is independently, optionally substituted C.sub.1-C.sub.6 alkyl or optionally substituted C.sub.6-C.sub.10 aryl; R.sup.5 is optionally substituted C.sub.1-C.sub.6 alkyl, optionally substituted C.sub.1-C.sub.6 heteroalkyl, optionally substituted 3- to 6-membered cycloalkyl, or optionally substituted 3- to 6-membered heterocycloalkyl; and X.sup.1, X.sup.2, and X.sup.3 are each, independently, CH.sub.2, CHF, CF.sub.2, C(O), or O.

11. The method of claim 10, wherein the compound of Formula IIId is prepared by a method comprising the steps of: A1) combining a compound of Formula IIIa and a compound of Formula IIIb to form a compound of Formula IIIc: ##STR00311## B1) reducing the compound of Formula IIIc to form the compound of Formula IId: ##STR00312## wherein Q is MgBr, MgCl, MgI, or Li.

12. The method of claim 11, wherein the reducing step (B1) comprises contacting the compound of Formula IIIc with a ketoreductase enzyme.

13. The method of claim 10, wherein the compound of Formula IIId is prepared by the steps of: A2) converting a compound of Formula IIIf to form a compound of Formula IIIg: ##STR00313## B2) reacting (e.g., esterifying) the compound of Formula IIIg to form the compound of Formula IIId: ##STR00314##

14. The method of claim 13, wherein the converting step (A2) comprises forming a diazonium salt of the compound of Formula IIIf.

15. A method of preparing a compound, or a salt thereof, of Formula IV: ##STR00315## the method comprising the steps of: a) coupling a compound of Formula III and a barium salt of compound A to form a compound of Formula IVa: ##STR00316## and b) hydrolyzing the compound of Formula IVa to form the compound of Formula IV: ##STR00317## wherein n and m are each, independently, 0, 1, 2, 3, 4, or 5; R.sup.4 is optionally substituted C.sub.1-C.sub.6 alkyl or optionally substituted C.sub.6-C.sub.10 aryl; R.sup.5 is optionally substituted C.sub.1-C.sub.6 alkyl, optionally substituted C.sub.1-C.sub.6 heteroalkyl, optionally substituted 3- to 6-membered cycloalkyl, or optionally substituted 3- to 6-membered heterocycloalkyl; and X.sup.1, X.sup.2, and X.sup.3 are each, independently, CH.sub.2, CHF, CF.sub.2, C(O), or O.

16. The method of claim 15, wherein the coupling step (a) comprises contacting the compound of Formula III and the Compound A barium salt with chloro-N,N,N,N-tetramethylformamidinium hexafluorophosphate (TCFH).

17. A method of preparing a compound, or a salt thereof, of Formula VIII: ##STR00318## the method comprising the steps of: a) esterifying and N-alkylating a compound of Formula VIIIa to form a compound of Formula VIIIb: ##STR00319## b) reducing the compound of Formula VIIIb to form a compound of Formula VIIIc as a mixture of atropisomers: ##STR00320## c) separating the mixture of atropisomers of Formula VIIIc to obtain a stereochemically pure compound of Formula VIII; wherein R.sup.12 is halogen; and R.sup.13 is optionally substituted C.sub.1-C.sub.6 alkyl.

18. A method of preparing a compound, or a salt thereof, of Formula VIII: ##STR00321## the method comprising the steps of: a) esterifying a compound of Formula VIIIa to form a compound of Formula VIIId: ##STR00322## b) N-alkylating the compound of Formula VIIId to form the compound of Formula VIIIb: ##STR00323## c) reducing the compound of Formula VIIIb to form a compound of Formula VIIIc as a mixture of atropisomers: ##STR00324## and d) separating the mixture of atropisomers of Formula VIIIc to obtain the stereochemically pure compound of Formula VIII; wherein R.sup.12 is halogen; and R.sup.13 is optionally substituted C.sub.1-C.sub.6 alkyl.

19. A method of preparing a compound, or a salt thereof, of Formula IX: ##STR00325## the method comprising the steps of: a) borylating a compound of Formula VIII to form a compound Formula X: ##STR00326## and b) coupling the compound of Formula X with the compound of Formula XI to form a compound of Formula XII: ##STR00327## wherein R.sup.12 is halogen; R.sup.13 is optionally substituted C.sub.1-C.sub.6 alkyl; and R.sup.14 is optionally substituted 3- to 6-membered cycloalkyl.

20. A method of preparing a compound, or a salt thereof, of Formula XIV: ##STR00328## the method comprising the steps of: a) reacting a compound of Formula XV with a compound of Formula XII to form a compound of Formula XVI: ##STR00329## b) removing PG.sub.a of the compound of Formula XVI to form a compound of Formula XVII: ##STR00330## c) cyclizing the compound of Formula XVII with a transitional metal catalyst to form a compound of Formula XVIII: ##STR00331## d) removing PG.sub.b of the compound of Formula XVIII to form a compound of Formula XIX: ##STR00332## and e) coupling the compound of Formula XIX with the compound of Formula IV to form a compound of Formula XIV: ##STR00333## wherein n and m are each, independently, 0, 1, 2, 3, 4, or 5; X.sup.1, X.sup.2, and X.sup.3 are each, independently, CH.sub.2, CHF, CF.sub.2, C(O), or O; R.sup.12 is halogen; R.sup.13 is optionally substituted C.sub.1-C.sub.6 alkyl; R.sup.14 is optionally substituted 3- to 6-membered cycloalkyl; PG.sub.a is an acid-labile protecting group; and PG.sub.b is an acid-stable protecting group.

21. The method of claim 20, wherein step (a) comprises reacting the compound of Formula XII and the compound of Formula XV in the presence of a carbodiimide coupling reagent, an anti-racemization agent, and a base.

22. The method of claim 20 or 21, wherein step (c) further comprises contacting the compound of Formula XVIII with hydrochloric acid to form a hydrochloride salt of the compound of Formula XVIII.

23. The method of any one of claims 20 to 22, wherein step (e) comprises coupling the compound of Formula XIX and the compound of Formula IV in the presence of a coupling reagent, an anti-racemization reagent, and a base.

24. A compound of Formula IIf-1: ##STR00334## or a salt thereof, wherein R.sup.2 is optionally substituted C.sub.1-6 alkyl, optionally substituted C.sub.3-6 cycloalkyl, or optionally substituted C.sub.6-10 aryl; R.sup.20 is hydrogen or C.sub.1-6 alkyl optionally substituted with 1 or 2 substituents independently selected from CN, OH, C.sub.1-6 alkoxy, and C.sub.1-6 haloalkyl; R.sup.21 is OH, halogen, CN, C.sub.1-6 alkyl, C.sub.1-6 alkoxy or C.sub.1-6 haloalkyl; and q is 0, 1, or 2.

25. A method of preparing a compound of Formula IIf: ##STR00335## the method comprising alkylating a compound of Formula IIe with a methylating agent: ##STR00336## wherein R.sup.2 is optionally substituted C.sub.1-6 alkyl, optionally substituted C.sub.3-6 cycloalkyl, or optionally substituted C.sub.6-10 aryl.

26. The method of claim 25, wherein the methylating agent comprises trimethylboroxine.

27. A method of preparing a compound of Formula IVa: ##STR00337## the method comprising coupling a compound of Formula III with a compound of Formula IIf wherein n and m are each, independently, 0, 1, 2, 3, 4, or 5; each R.sup.4 is independently optionally substituted C.sub.1-C.sub.6 alkyl or optionally substituted C.sub.6-C.sub.10 aryl; each R.sup.5 is independently optionally substituted C.sub.1-C.sub.6 alkyl, optionally substituted C.sub.1-C.sub.6 heteroalkyl, optionally substituted 3- to 6-membered cycloalkyl, or optionally substituted 3- to 6-membered heterocycloalkyl; and X.sup.1, X.sup.2, and X.sup.3 are each, independently, CH.sub.2, CHF, CF.sub.2, C(O), or O.

28. The method of claim 27, wherein the coupling comprises contacting the compound of Formula III and the compound of Formula IIf with a base.

29. A method of preparing a compound of Formula VIIIc: ##STR00338## the method comprising N-alkylating a compound of Formula VIIIe with a compound having the structure of R.sup.13-LG.sup.1: ##STR00339## wherein each R.sup.12 is halogen; each R.sup.13 is optionally substituted C.sub.1-C.sub.6 alkyl; and LG.sup.1 is a leaving group.

30. A method of preparing a compound of Formula Ville: ##STR00340## the method comprising reducing a compound of Formula VIIIf with a reducing agent: ##STR00341## wherein R.sup.12 is halogen; and R.sup.13 is optionally substituted C.sub.1-C.sub.6 alkyl.

31. The method of claim 30, wherein the reducing agent is NaBH.sub.4.

Description

DETAILED DESCRIPTION

[0253] Provided herein are synthetic methods and intermediates for making Ras inhibitors, or salts thereof. The methods and intermediates can be useful for achieving a higher yield, a higher chemical purity, and/or a higher stereoisomeric purity, and a lower cost for the preparation of the Ras inhibitors or the intermediates. Further synthetic details are provided in the Examples.

[0254] The compounds described herein may be prepared using the methods described herein and/or using known organic, inorganic, or enzymatic processes. The synthetic methods may employ the use of commercially available starting materials or starting materials prepared by processes known to those skilled in the art of organic synthesis. These methods include but are not limited to those methods described in the Schemes below and in WO 2021/091967, WO 2022/235870, and WO 2023/060253 the disclosure of each of which is incorporated herein by reference.

Synthetic Methods

[0255] In one aspect, the disclosure provides a method of separating enantiomers of a compound of Formula I

##STR00095##

[0256] The method includes the steps of: [0257] a) contacting the compound of Formula I with a chiral acid to form a diastereomeric salt of the compound of Formula I; and [0258] b) separating each diastereomer of the diastereomeric salt, thereby separating enantiomers of a compound of Formula I
wherein [0259] n and m are each, independently, 0, 1, 2, 3, 4, or 5; [0260] PG is a nitrogen protecting group; [0261] and [0262] X.sup.1, X.sup.2, and X.sup.3 are each, independently, CH.sub.2, CHF, CF.sub.2, C(O), or O.

[0263] In some embodiments, the nitrogen protecting group is Boc (tert-Butyloxycarbonyl). In some embodiments, the chiral acid is a chiral carboxylic acid. In some embodiments, the chiral carboxylic acid is a chiral dicarboxylic acid. In some embodiments, the chiral dicarboxylic acid is tartaric acid. In some embodiments, the chiral carboxylic acid is L(+)-tartaric acid. In some embodiments, the separating step (b) includes recrystallization.

[0264] In some embodiments, the compound of Formula I is further described by the compound of Formula Ia:

##STR00096##

[0265] In some embodiments, the compound of Formula I or Formula Ia is Compound B:

##STR00097##

[0266] In another aspect, the disclosure provides a barium salt of Compound A:

##STR00098##

[0267] In some embodiments, the barium salt is a 2:1 (compound A: barium) salt.

[0268] In a further aspect, the disclosure provides a method of preparing the barium salt of Compound A. The method includes the steps of: [0269] a) condensing a compound of Formula IIa and Compound C to form a compound of Formula IIb:

##STR00099## [0270] b) cyclizing the compound of Formula IIb and a compound of Formula IIc to form a compound of Formula IIId:

##STR00100## [0271] c) hydrolyzing the compound of Formula IIId to form a compound of Formula IIe:

##STR00101## [0272] d) methylating the compound of formula IIe to form a compound of formula IIf:

##STR00102##

and [0273] e) hydrolyzing the compound of Formula IIf to form the barium salt of Compound A:

##STR00103##

wherein R.sup.2 and R.sup.3 are each, independently, optionally substituted C.sub.1-C.sub.6 alkyl, optionally substituted C.sub.3-6 cycloalkyl, or optionally substituted C.sub.6-C.sub.10 aryl, and LG is a leaving group selected from the group consisting of halogen, triflate, mesylate, and tosylate. In some embodiments, R.sup.3 is optionally substituted C.sub.6-C.sub.10 aryl. In some embodiments, R.sup.3 is

##STR00104##

In some embodiments, R.sup.3 is optionally substituted C.sub.1-C.sub.6 alkyl. In some embodiments, R.sup.3 is tert-butyl. In some embodiments, R.sup.2 is optionally substituted C.sub.1-C.sub.6 alkyl, optionally substituted C.sub.3-6 cycloalkyl, or optionally substituted C.sub.6-10 aryl. In some embodiments, R.sup.2 is benzyl, methyl, or t-Butyl. In some embodiments, LG is halogen. In some embodiments, LG is Br.

[0274] In some embodiments of the method of preparing the barium salt of Compound A, the condensing step (a) includes contacting the compound of Formula IIa and Compound C with a base. In some embodiments, the base is an amine base. In some embodiments, the base is pyrrolidine. In some embodiments, the condensing step (a) is carried out according to the following scheme:

##STR00105##

[0275] In some embodiments, the condensing step (a) is carried out according to the following scheme:

##STR00106##

[0276] In some embodiments, the condensing step (a) is carried out according to the following scheme:

##STR00107##

[0277] In some embodiments, the cyclizing step (b) includes contacting the compound of Formula IIb and the compound of Formula IIc with a base. In some embodiments, the base is a lithium base. In some embodiments, the base is LiHMDS.

[0278] In some embodiments, the cyclizing step (b) is carried out according to the following scheme:

##STR00108##

[0279] In some embodiments, the hydrolyzing step (c) includes contacting the compound of formula Ild with an acid. In some embodiments, the acid is a carboxylic acid. In some embodiments, the carboxylic acid is trifluoroacetic acid. In some embodiments, the acid is hydrochloric acid.

[0280] In some embodiments, the hydrolyzing step (c) is carried out according to the following scheme:

##STR00109##

[0281] In some embodiments, the hydrolyzing step (c) is carried out according to the following scheme:

##STR00110##

[0282] In some embodiments, the hydrolyzing step (c) is carried out according to the following scheme:

##STR00111##

[0283] In some embodiments, the methylating step (d) includes contacting the compound of Formula IIe with MeB(OH).sub.2. In some embodiments, the methylating step (d) further includes contacting the compound of Formula IIe with a copper catalyst. In some embodiments, the methylating step (d) is carried out according to the following scheme:

##STR00112##

[0284] In some embodiments, the hydrolyzing step (e) includes contacting the compound of Formula IIf with barium hydroxide.

[0285] In some embodiments, the hydrolyzing step (e) is carried out according to the following scheme:

##STR00113##

[0286] In yet another aspect, the disclosure provides a method of preparing a compound of Formula IIf:

##STR00114##

the method including alkylating a compound of Formula IIe with a methylating agent:

##STR00115## [0287] wherein R.sup.2 is optionally substituted C.sub.1-6 alkyl, optionally substituted C.sub.3-6 cycloalkyl, or optionally substituted C.sub.6-10 aryl.

[0288] In some embodiments, the methylating agent includes trimethylboroxine. In some embodiments, the methylating agent includes trimethylboroxine and Cu(OAc).sub.2. In some embodiments, the methylating agent includes trimethylboroxine and CuOAc.

[0289] In some embodiments of the method of preparing the compound of Formula IIf, the alkylating step includes contacting the compound of Formula IIe with an alkaline salt. In some embodiments, the alkaline salt is potassium carbonate (K.sub.2CO.sub.3) or sodium carbonate (Na.sub.2CO.sub.3). In some embodiments, the alkylating step includes 2,2-bipyridine (bipy). In some embodiments, the alkylating step includes di-tert-butyl peroxide (DTBP). In some embodiments, the alkylating step is carried out a temperature of at least 25 C. (e.g., 25, 30, 35, or 40 C.). In some embodiments, the alkylating step is carried out in an organic solvent (e.g., acetonitrile (ACN)). In some embodiments of the method of preparing the compound of Formula IIf, the alkylating step is carried out according to the following scheme:

##STR00116##

[0290] In some embodiments of the method of preparing the compound of Formula IIf, the alkylating step is carried out according to the following scheme:

##STR00117##

[0291] In another aspect, the disclosure provides a method of preparing a compound, or a salt thereof, of Formula III:

##STR00118##

[0292] The method includes the steps of: [0293] a) coupling a compound of Formula IIId with a compound of Formula I to form a compound of Formula IIIe:

##STR00119##

and [0294] b) deprotecting the compound of Formula IIIe to form the compound of Formula III:

##STR00120##

wherein [0295] n and m are each, independently, 0, 1, 2, 3, 4, or 5; [0296] PG is a nitrogen protecting group; [0297] each R.sup.4 is independently, optionally substituted C.sub.1-C.sub.6 alkyl or optionally substituted C.sub.6-C.sub.10 aryl; and [0298] R.sup.5 is optionally substituted C.sub.1-C.sub.6 alkyl, optionally substituted C.sub.1-C.sub.6 heteroalkyl, optionally substituted 3- to 6-membered cycloalkyl, or optionally substituted 3- to 6-membered heterocycloalkyl; and X.sup.1, X.sup.2, and X.sup.3 are each, independently, CH.sub.2, CHF, CF.sub.2, C(O), or O.

[0299] In some embodiments, the nitrogen protecting group is Boc (tert-Butyloxycarbonyl). In some embodiments, the compound of Formula IIId is prepared by a method including the steps of: [0300] A1) combining a compound of Formula IIIa and a compound of Formula IIIb to form a compound of Formula IIIc:

##STR00121## [0301] B1) reducing the compound of Formula IIIc to form the compound of Formula IIId:

##STR00122##

wherein Q is MgBr, MgCl, MgI, or Li.

[0302] In some embodiments, the combining step (A1) includes contacting the Compound of Formula IIIa with a compound of Formula IIIb, wherein Q is MgBr. In some embodiments, the contacting is carried out in tetrahydrofuran at 78 C.

[0303] In some embodiments, the reducing step (B1) includes contacting the compound of Formula IIIc with a ketoreductase enzyme.

[0304] In some embodiments, the compound of Formula IIId is prepared by a method including the steps of: [0305] A2) converting a compound of Formula IIIf to form a compound of Formula IIIg:

##STR00123## [0306] B2) reacting (e.g., esterifying) the compound of Formula IIIg to form the compound of Formula IIId:

##STR00124##

[0307] In some embodiments, the converting step (A2) includes forming a diazonium salt of the compound of Formula IIIf. In some embodiments, the converting step (A2) and reacting (e.g., esterifying) step (B2) is carried out according to the following scheme

##STR00125##

[0308] In some embodiments, the combining step (a) of Formula IIId with a compound of Formula I to form a compound of Formula IIIe includes contacting the compound of Formula IIId with triflic anhydride followed by contacting the compound of Formula IIId with the compound of Formula I. In some embodiments, the combining step (a) of Formula IIId with a compound of Formula I to form a compound of Formula IIIe is carried out according to the following scheme:

##STR00126##

[0309] In some embodiments, the deprotecting step (b) includes contacting the compound of Formula Ille with an acid. In some embodiments, the acid is hydrochloric acid. In some embodiments, the hydrochloric acid is delivered an ethyl acetate solution of hydrochloric acid.

[0310] In yet another aspect, the disclosure provides a method of preparing a compound of Formula IVa:

##STR00127##

the method including coupling a compound of Formula III with a compound of Formula IIf [0311] wherein n and m are each, independently, 0, 1, 2, 3, 4, or 5; [0312] each R.sup.4 is independently optionally substituted C.sub.1-C.sub.6 alkyl or optionally substituted C.sub.6-C.sub.10 aryl; [0313] each R.sup.5 is independently optionally substituted C.sub.1-C.sub.6 alkyl, optionally substituted C.sub.1-C.sub.6 heteroalkyl, optionally substituted 3- to 6-membered cycloalkyl, or optionally substituted 3- to 6-membered heterocycloalkyl; and [0314] X.sup.1, X.sup.2, and X.sup.3 are each, independently, CH.sub.2, CHF, CF.sub.2, C(O), or O.

[0315] In some embodiments, R.sup.4 is benzyl, methyl, or t-Butyl. In some embodiments, R.sup.5 is isopropyl, cyclopropyl, cyclobutyl, cyclopentyl, or cyclohexyl. In some embodiments, R.sup.5 is cyclopentyl.

[0316] In some embodiments, the coupling includes contacting the compound of Formula III and the compound of Formula IIf with a base. In some embodiments, the base is 1,5,7-triazobicyclo[4,4,0]dec-5-ene (TBD). In some embodiments, the coupling is carried out in an organic solvent (e.g., methyl tert-butyl ether (MTBE)). In some embodiments of the method of preparing the compound of Formula IVa, the coupling is carried out at a temperature of at least 10 C. (e.g., 10, 5, 0, or 5 C.). In some embodiments of the method of preparing the compound of Formula IVa, the coupling is carried out according to the following scheme:

##STR00128##

[0317] In some embodiments of the method of preparing the compound of Formula IVa, the compound of Formula III is a compound of Formula III-1.

[0318] In some embodiments of the method of preparing the compound of Formula IVa, the coupling is carried out according to the following scheme:

##STR00129##

[0319] In some embodiments, the compound of Formula III is Compound D:

##STR00130##

which is prepared by: [0320] a) coupling a compound of Formula IIId-1 with Compound B to form a compound of Formula IIIe-1 having the following structure:

##STR00131##

and [0321] b) deprotecting the compound of Formula IIIe-1 to form Compound D having the following structure:

##STR00132##

[0322] In some embodiments, the combining step (a) of Formula IIId with Compound B to form a compound of Formula IIIe includes contacting the compound of Formula IIId with triflic anhydride followed by contacting the compound of Formula IIId with Compound B.

[0323] In some embodiments, the combining step (a) of Formula IIId with Compound B to form a compound of Formula IIIe is carried out according to the following scheme:

##STR00133##

[0324] In some embodiments, the deprotecting step (b) includes contacting the compound of Formula Ille with an acid. In some embodiments, the acid is hydrochloric acid. In some embodiments, the hydrochloric acid is delivered an ethyl acetate solution of hydrochloric acid.

[0325] In yet another aspect, the disclosure provides a method of preparing a compound, or a salt thereof, of Formula IV:

##STR00134##

[0326] The method includes the steps of: [0327] a) coupling a compound of Formula III and a barium salt of compound A to form a compound of Formula IVa:

##STR00135##

and [0328] b) hydrolyzing the compound of Formula IVa to form the compound of Formula IV:

##STR00136##

wherein [0329] n and m are each, independently, 0, 1, 2, 3, 4, or 5; [0330] R.sup.4 is optionally substituted C.sub.1-C.sub.6 alkyl or optionally substituted C.sub.6-C.sub.10 aryl; [0331] R.sup.5 is optionally substituted C.sub.1-C.sub.6 alkyl, optionally substituted C.sub.1-C.sub.6 heteroalkyl, optionally substituted 3- to 6-membered cycloalkyl, or optionally substituted 3- to 6-membered heterocycloalkyl; and [0332] X.sup.1, X.sup.2, and X.sup.3 are each, independently, CH.sub.2, CHF, CF.sub.2, C(O), or O.

[0333] In some embodiments, the coupling step (a) includes contacting the compound of Formula III and the Compound A barium salt with chloro-N,N,N,N-tetramethylformamidinium hexafluorophosphate (TCFH). In some embodiments, the coupling step (a) further includes contacting the compound of Formula III and the Compound A barium salt with NMI. In some embodiments, the coupling step (a) is carried out according to the following scheme:

##STR00137##

[0334] In some embodiments, the hydrolyzing step (b) includes contacting the compound of Formula IVa with an acid or base. In some embodiments, the hydrolyzing step (b) includes hydrogenating the compound of Formula IVa. In some embodiments, the hydrogenating further includes contacting the compound of Formula IVa with a palladium catalyst.

[0335] In some embodiments, the hydrolyzing step (b) is carried out according to the following scheme:

##STR00138##

[0336] In some embodiments of any of the methods and compounds described herein, m is 0. In some embodiments, m is 1. In some embodiments, m is 2. In some embodiments, n is 0. In some embodiments, n is 1. In some embodiments, n is 2.

[0337] In some embodiments of any of the methods and compounds described herein, each of X.sup.1, X.sup.2, and X.sup.3 is CH.sub.2. In some embodiments, PG is

##STR00139##

In some embodiments, LG is halogen. In some embodiments, LG is Br.

[0338] In some embodiments of any of the methods and compounds described herein, R.sup.2 is optionally substituted C.sub.1-C.sub.6 alkyl, optionally substituted C.sub.3-6 cycloalkyl, or optionally substituted C.sub.6-10 aryl. In some embodiments, R.sup.2 is benzyl, methyl, or t-Butyl. In some embodiments, R.sup.3 is optionally substituted C.sub.6-C.sub.10 aryl. In some embodiments, R.sup.3 is

##STR00140##

In some embodiments, R.sup.3 is optionally substituted C.sub.1-C.sub.6 alkyl. In some embodiments, R.sup.3 is

##STR00141##

In some embodiments, R.sup.4 is optionally substituted C.sub.1-C.sub.6 alkyl or optionally substituted C.sub.6-C.sub.10 aryl. In some embodiments, R.sup.4 is benzyl, methyl, or t-Butyl. In some embodiments, R.sup.5 is optionally substituted C.sub.1-C.sub.6 alkyl or optionally substituted 3- to 6-membered cycloalkyl. In some embodiments, R.sup.5 is optionally substituted C.sub.1-C.sub.6 alkyl. In some embodiments, R.sup.5 is isopropyl. In some embodiments, R.sup.5 is optionally substituted 3- to 6-membered cycloalkyl. In some embodiments, R.sup.5 is cyclopentyl.

[0339] In some embodiments, the method includes preparing Compound E:

##STR00142##

and the method including the steps of: [0340] a) coupling Compound D and a barium salt of compound A to form a compound of Formula IVa having the following structure:

##STR00143##

and [0341] b) hydrolyzing the compound of Formula IVa to form Compound E having the following structure:

##STR00144##

[0342] In some embodiments, the coupling step (a) includes contacting Compound D and the Compound A barium salt with chloro-N,N,N,N-tetramethylformamidinium hexafluorophosphate (TCFH). In some embodiments, the coupling step (a) further includes contacting Compound D and the barium salt of Compound A with NMI. In some embodiments, the coupling step (a) is carried out according to the following scheme:

##STR00145##

[0343] In some embodiments, the hydrolyzing step (b) includes contacting the compound of Formula IVa with an acid or base. In some embodiments, the hydrolyzing step (b) includes hydrogenating the compound of Formula IVa. In some embodiments, the hydrogenating further includes contacting the compound of Formula IVa with a palladium catalyst. In some embodiments, the hydrolyzing step (b) is carried out according to the following scheme:

##STR00146##

[0344] In yet another aspect, the disclosure provides a method of preparing a compound, or a salt thereof, of Formula VIII:

##STR00147##

the method including the steps of: [0345] a) esterifying and N-alkylating a compound of Formula VIIIa to form a compound of Formula VIIIb:

##STR00148## [0346] b) reducing the compound of Formula VIIIb to form a compound of Formula VIIIc as a mixture of atropisomers:

##STR00149##

and [0347] c) separating the mixture of atropisomers of Formula VIIIc to obtain a stereochemically pure compound of Formula VIII; [0348] wherein R.sup.12 is halogen; and [0349] R.sup.13 is optionally substituted C.sub.1-C.sub.6 alkyl.

[0350] In some embodiments, the esterifying and N-alkylating step (a) includes contacting the compound of Formula VIIIa with an alkylating agent. In some embodiments, the alkylating agent is R.sup.13-LG, wherein R.sup.13 is optionally substituted C.sub.1-C.sub.6 alkyl and LG is a leaving group. In some embodiments, the alkylating agent is 2,2,2-trifluoroethyl trifluoromethanesulfonate.

[0351] In some embodiments, the esterifying and N-alkylating step (a) is carried out according to the following scheme:

##STR00150##

[0352] In some embodiments, the reducing step (b) includes contacting the compound of Formula VIIIb with a reducing agent. In some embodiments, the reducing agent is a borohydride. In some embodiments, the borohydride is lithium borohydride. In some embodiments, the reducing step (b) is carried out according to the following scheme:

##STR00151##

[0353] In some embodiments, the compound of Formula VIII is Compound F:

##STR00152##

and the method includes the steps of: [0354] a) esterifying and N-alkylating a compound of Formula VIIIa to form a compound of Formula VIIIb having the following structure:

##STR00153## [0355] b) reducing the compound of Formula VIIIb to form a compound of Formula VIIIc as a mixture of atropisomers having the following structure:

##STR00154##

and [0356] c) separating the mixture of atropisomers of Formula VIIIc to obtain the stereochemically pure compound of Formula VIII.

[0357] In some embodiments, the esterifying and N-alkylating step (a) includes contacting the compound of Formula VIIIa with an alkylating agent. In some embodiments, the alkylating agent is R.sup.13-LG, wherein R.sup.13 is optionally substituted C.sub.1-C.sub.6 alkyl and LG is a leaving group. In some embodiments, the alkylating agent is 2,2,2-trifluoroethyl trifluoromethanesulfonate. In some embodiments, the esterifying and N-alkylating step (a) is carried out according to the following scheme:

##STR00155##

[0358] In some embodiments, the reducing step (b) includes contacting the compound of Formula VIIIb with a reducing agent. In some embodiments, the reducing agent is a borohydride. In some embodiments, the borohydride is lithium borohydride.

[0359] In some embodiments, the reducing step (b) is carried out according to the following scheme:

##STR00156##

[0360] In yet another aspect, the disclosure provides a method of preparing a compound, or a salt thereof, of Formula VIII:

##STR00157##

the method including the steps of: [0361] a) reacting (e.g., esterifying) a compound of Formula VIIIa to form a compound of Formula VIIId:

##STR00158## [0362] b) N-alkylating the compound of Formula VIIId to form the compound of Formula VIIIb:

##STR00159## [0363] c) reducing the compound of Formula VIIIb to form a compound of Formula VIIIc as a mixture of atropisomers:

##STR00160##

and [0364] d) separating the mixture of atropisomers of Formula VIIIc to obtain the stereochemically pure compound of Formula VIII; [0365] wherein R.sup.12 is halogen; and [0366] R.sup.13 is optionally substituted C.sub.1-C.sub.6 alkyl.

[0367] In some embodiments, step (a) includes contacting the compound of Formula VIIIa with R.sup.13OH, wherein R.sup.13 is optionally substituted C.sub.1-C.sub.6 alkyl. In some embodiments, the alcohol is methanol. In some embodiments, step (a) further includes contacting the compound of Formula VIIIa with an acid. In some embodiments, the acid is sulfuric acid. In some embodiments, step (a) is carried out according to the following scheme:

##STR00161##

[0368] In some embodiments, the N-alkylating step (b) includes contacting the compound of Formula VIIId with an alkylating agent. In some embodiments, the alkylating agent is R.sup.13-LG, wherein R.sup.13 is optionally substituted C.sub.1-C.sub.6 alkyl and LG is a leaving group. In some embodiments, the alkylating agent is 2,2,2-trifluoroethyl trifluoromethanesulfonate. In some embodiments, the N-alkylating step (b) is carried out according to the following scheme:

##STR00162##

[0369] In some embodiments, the reducing step (c) includes contacting the compound of Formula VIIIb with a reducing agent. In some embodiments, the reducing agent is a borohydride. In some embodiments, the borohydride is lithium borohydride.

[0370] In some embodiments, the reducing step (c) is carried out according to the following scheme:

##STR00163##

[0371] In some embodiments, the compound of Formula VIII is Compound G:

##STR00164##

and the method includes the steps of: [0372] a) reacting (e.g., esterifying) a compound of Formula VIIIa to form a compound of Formula VIIId having the following structure:

##STR00165## [0373] b) N-alkylating the compound of Formula VIIId to form the compound of Formula VIIIb having the following structure:

##STR00166## [0374] c) reducing the compound of Formula VIIIb to form Compound G as a mixture of atropisomers having the following structure:

##STR00167##

[0375] In some embodiments, step (a) includes contacting the compound of Formula VIIIa with R.sup.13OH, wherein R.sup.13 is optionally substituted C.sub.1-C.sub.6 alkyl. In some embodiments, the alcohol is methanol. In some embodiments, step (a) further includes contacting the compound of Formula VIIIa with an acid. In some embodiments, the acid is sulfuric acid. In some embodiments, step (a) is carried out according to the following scheme:

##STR00168##

[0376] In some embodiments, the N-alkylating step (b) includes contacting the compound of Formula VIIId with an alkylating agent. In some embodiments, the alkylating agent is R.sup.13-LG, wherein R.sup.13 is optionally substituted C.sub.1-C.sub.6 alkyl and LG is a leaving group. In some embodiments, the alkylating agent is 2,2,2-trifluoroethyl trifluoromethanesulfonate. In some embodiments, the N-alkylating step (b) is carried out according to the following scheme:

##STR00169##

[0377] In some embodiments, the reducing step (c) includes contacting the compound of Formula VIIIb with a reducing agent. In some embodiments, the reducing agent is a borohydride. In some embodiments, the borohydride is lithium borohydride. In some embodiments, the reducing step (c) is carried out according to the following scheme:

##STR00170##

[0378] In some embodiments, the mixture of atropisomers is separated by column chromatography or chemical resolution. In some embodiments, the chemical resolution includes contacting the compound of Formula VIIIc with an acid. In some embodiments, the acid is methanesulfonic acid. In some embodiments, the acid is camphor sulfonic acid (e.g., D-camphor-10-sulfonic acid).

[0379] In yet another aspect, the disclosure provides a method of preparing a compound of Formula VIIIc:

##STR00171##

the method including N-alkylating a compound of Formula VIIIe with a compound having the structure of R.sup.13-LG.sup.1:

##STR00172## [0380] wherein each R.sup.12 is independently halogen; [0381] each R.sup.13 is independently optionally substituted C.sub.1-C.sub.6 alkyl; and [0382] LG.sup.1 is a leaving group.

[0383] In some embodiments, each R.sup.12 is Br. In some embodiments, each R.sup.13 is CH.sub.2CF.sub.3. In some embodiments, LG.sup.1 is halogen, triflate, mesylate, or tosylate. In some embodiments, R.sup.13-LG is 2,2,2-trifluoroethyl trifluoromethanesulfonate. In some embodiments, the method includes contacting the compound of Formula VIIIe and R.sup.13-LG.sup.1 with a base. In some embodiments, the base is tripotassium phosphate (K.sub.3PO.sub.4). In some embodiments, the In some embodiments, the method is carried out at a temperature of at least 10 C. (e.g., 10, 15, 20, or 25 C.). In some embodiments, the method produces a mixture of atropisomers of the compound of Formula VIIIc. In some embodiments, the N-alkylating is carried out according to the following scheme:

##STR00173##

[0384] In some embodiments of the method of preparing the compound of Formula VIIIc, the compound of Formula Ville is Compound K. In some embodiments, the compound of Formula VIIIc is Compound G. In some embodiments, the N-alkylating is carried out according to the following scheme:

##STR00174##

[0385] In yet another aspect, the disclosure provides a method of preparing a compound of Formula VIIIe:

##STR00175##

the method including reducing a compound of Formula VIIIf with a reducing agent:

##STR00176## [0386] wherein R.sup.12 is halogen; and [0387] R.sup.13 is optionally substituted C.sub.1-C.sub.6 alkyl.

[0388] In some embodiments, each R.sup.12 is Br. In some embodiments, each R.sup.13 is CH.sub.2CF.sub.3. In some embodiments, the reducing agent is NaBH.sub.4. In some embodiments, the method includes contacting the compound of Formula VIIIf with calcium chloride (CaCl.sub.2). In some embodiments, the reducing is carried out at a temperature of at least 20 C. (e.g., 20, 25, or 30 C.). In some embodiments, the reducing is carried out according to the following scheme:

##STR00177##

[0389] In some embodiments, the reducing is carried out according to the following scheme:

##STR00178##

[0390] In yet another aspect, the disclosure provides a method of preparing a compound, or a salt thereof, of Formula IX:

##STR00179##

the method including the steps of: [0391] a) borylating a compound of Formula VIII to form a compound Formula X:

##STR00180##

and [0392] b) coupling the compound of Formula X with the compound of Formula XI to form a compound of Formula XII:

##STR00181## [0393] wherein R.sup.12 is halogen; [0394] R.sup.13 is optionally substituted C.sub.1-C.sub.6 alkyl; and [0395] R.sup.14 is optionally substituted 3- to 6-membered cycloalkyl.

[0396] In some embodiments, the borylating step (a) includes contacting the compound of Formula VIII with a borylating agent. In some embodiments, the borylating agent is bis(pinacolato)diboron (B.sub.2Pin.sub.2). In some embodiments, the borylating step (a) further includes contacting the compound of Formula VIII with 4,4-di-tert-butyl-2,2-dipyridyl (dtbpy). In some embodiments, the borylating step (a) further includes contacting the compound of Formula VIII with an iridium catalyst. In some embodiments, the iridium catalyst is [Ir(OMe)(COD)]2.

[0397] In some embodiments, the borylating step (a) is carried out according to the following scheme:

##STR00182##

[0398] In some embodiments, the coupling step (b) includes contacting the compound of Formula X and the compound of Formula XI with a copper catalyst. In some embodiments, the copper catalyst is Cu(OAc).sub.2. In some embodiments, the coupling step (b) includes contacting the compound of Formula X and the compound of Formula XI with a base. In some embodiments, the base is an amine base. In some embodiments, the base is triethylamine or tetramethylpiperidine. In some embodiments, the coupling step (b) is carried out according to the following scheme:

##STR00183##

[0399] In some embodiments, the compound of Formula IX is Compound H:

##STR00184##

the method includes the steps of: [0400] a) borylating a compound of Formula VIII to form a compound Formula X having the following structure:

##STR00185## [0401] b) coupling the compound of Formula X with the compound of Formula XI to form Compound H having the following structure:

##STR00186##

[0402] In some embodiments, the borylating step (a) includes contacting the compound of Formula VIII with a borylating agent. In some embodiments, the borylating agent is bis(pinacolato)diboron (B.sub.2Pin.sub.2). In some embodiments, the borylating step (a) further includes contacting the compound of Formula VIII with 4,4-di-tert-butyl-2,2-dipyridyl (dtbpy). In some embodiments, the borylating step (a) further includes contacting the compound of Formula VIII with an iridium catalyst. In some embodiments, the iridium catalyst is [Ir(OMe)(COD)].sub.2. In some embodiments, the borylating step (a) is carried out according to the following scheme:

##STR00187##

[0403] In some embodiments, the coupling step (b) includes contacting the compound of Formula X and the compound of Formula XI with a copper catalyst. In some embodiments, the copper catalyst is Cu(OAc).sub.2. In some embodiments, the coupling step (b) includes contacting the compound of Formula X and the compound of Formula XI with a base. In some embodiments, the base is an amine base. In some embodiments, the base is triethylamine or tetramethylpiperidine. In some embodiments, the coupling step (b) is carried out according to the following scheme:

##STR00188##

[0404] In some embodiments, the compound, or a salt thereof, of Formula X:

##STR00189##

is further prepared by: [0405] a) borylating the compound of Formula VIII to form a compound of Formula XIII:

##STR00190##

and [0406] b) hydrolyzing and borylating the compound of Formula XIII in one step to form Formula X:

##STR00191## [0407] wherein R.sup.12 is halogen; and [0408] R.sup.13 is optionally substituted C.sub.1-C.sub.6 alkyl.

[0409] In some embodiments, the borylating step (a) includes contacting the compound of Formula VIII with a borylating agent. In some embodiments, the borylating agent is pinacolborane (HBPin). In some embodiments, the borylating step (a) is carried out according to the following scheme:

##STR00192##

[0410] In some embodiments, the hydrolyzing and borylating step (b) includes contacting the compound of Formula XIII with a borylating agent. In some embodiments, the borylating agent is bis(pinacolato)diboron (B.sub.2Pin.sub.2). In some embodiments, the hydrolyzing and borylating step (b) further includes contacting the compound of Formula XIII with 4,4-di-tert-butyl-2,2-dipyridyl (dtbpy). In some embodiments, the hydrolyzing and borylating step (b) further includes contacting the compound of Formula XIII with an iridium catalyst. In some embodiments, the iridium catalyst is [Ir(OMe)(COD)].sub.2. In some embodiments, the hydrolyzing and borylating step (b) is carried out according to the following scheme:

##STR00193##

[0411] In yet another aspect, the disclosure provides a method of preparing a compound, or a salt thereof, of Formula XIV:

##STR00194##

the method including the steps of: [0412] a) reacting (e.g., esterifying) a compound of Formula XV with a compound of Formula XII to form a compound of Formula XVI:

##STR00195## [0413] b) removing (e.g., deprotecting) PG.sub.a of the compound of Formula XVI to form a compound of Formula XVII:

##STR00196## [0414] c) cyclizing the compound of Formula XVII with a transition metal catalyst to form a compound of Formula XVII:

##STR00197## [0415] d) removing (e.g., deprotecting) PG.sub.b of the compound of Formula XVIII to form a compound of Formula XIX:

##STR00198##

and [0416] e) coupling the compound of Formula XIX with the compound of Formula IV to form a compound of Formula XIV:

##STR00199## [0417] wherein n and m are each, independently, 0, 1, 2, 3, 4, or 5; [0418] X.sup.1, X.sup.2, and X.sup.3 are each, independently, CH.sub.2, CHF, CF.sub.2, C(O), or O; [0419] R.sup.12 is halogen; [0420] R.sup.13 is optionally substituted C.sub.1-C.sub.6 alkyl; [0421] R.sup.14 is optionally substituted 3- to 6-membered cycloalkyl; [0422] PG.sub.a is an acid-labile protecting group; and [0423] PG.sub.b is an acid-stable protecting group.

[0424] In some embodiments, step (a) includes contacting the compound of Formula XV and the compound of Formula XII with a carbodiimide coupling reagent, an anti-racemization agent, and a base. In some embodiments, the carbodiimide coupling reagent is EDCl, the anti-racemization agent is HOBt, and the base is N,N-diisopropylethylamine.

[0425] In some embodiments, step (a) is carried out according to the following scheme:

##STR00200##

[0426] In some embodiments, the deprotecting step (b) includes contacting the compound of Formula XVI with an acid. In some embodiments, the acid is trifluoroacetic acid. In some embodiments, the deprotecting step (b) is carried out according to the following scheme:

##STR00201##

[0427] In some embodiments, the cyclizing step (c) includes contacting the compound of Formula XVII with a transition metal catalyst, a phosphine ligand, and a base. In some embodiments, the transition metal catalyst includes palladium, platinum, nickel, copper, and iron catalysts. In some embodiments, the transition metal catalyst is a palladium catalyst. In some embodiments, the palladium catalyst is P(tBu).sub.3 Pd G3 or Pd.sub.2(dba).sub.3. In some embodiments, the phosphine ligand is Q-Phos or [(tBu).sub.3PH]BF.sub.4. In some embodiments, the base is K.sub.3PO.sub.4. In some embodiments, the cyclizing step (c) is carried out according to the following scheme:

##STR00202##

[0428] In some embodiments, the cyclizing step (c) is carried out according to the following scheme:

##STR00203##

[0429] In some embodiments, the cyclizing step (c) further includes contacting the compound of Formula XVIII with hydrochloric acid to form a hydrochloride salt of the compound of Formula XVIII.

[0430] In some embodiments, the deprotecting step (d) includes contacting the compound of Formula XVIII with a palladium catalyst and hydrogen gas in an organic solvent. In some embodiments, the palladium catalyst is Pd/C. In some embodiments, the organic solvent is methyl tert-butyl ether. In some embodiments, the deprotecting step (d) is carried out according to the following scheme:

##STR00204##

[0431] In some embodiments, the deprotecting step (d) further includes washing the compound of Formula XIX with N-acetyl cysteine.

[0432] In some embodiments, the deprotecting step (e) includes contacting the compound of Formula XIX and the compound of Formula IV with a coupling reagent, an anti-racemization reagent, and a base. In some embodiments, the coupling reagent is PyBOP or EDCl. In some embodiments, the anti-racemization agent is selected from the group consisting of Oxyma, HOBt, or HOPO. In some embodiments, the base is N,N-diisopropylethylamine.

[0433] In some embodiments, the coupling step (e) is carried out according to the following scheme:

##STR00205##

[0434] In some embodiments, the coupling step (e) is carried out according to the following scheme:

##STR00206##

[0435] In some embodiments, the coupling step (e) is carried out according to the following scheme:

##STR00207##

In some embodiments, the method further includes the step of purifying the compound of Formula XIV. In some embodiments, the purifying includes recrystallizing the compound of Formula XIV. In some embodiments, the recrystallizing includes adding a first solvent, followed by adding a second solvent. In some embodiments, the first solvent is dioxane or 2-methyltetrahydrofuran. In some embodiments, the second solvent is diisopropyl ether or isopropyl alcohol. In some embodiments, the recrystallizing further includes adding a third solvent. In some embodiments, the third solvent is heptane. In some embodiments, R.sup.12 is Br. In some embodiments, R.sup.13 is CH.sub.2CF.sub.3. In some embodiments, PG.sub.a is Boc. In some embodiments, PG.sub.b is CBz.

[0436] In some embodiments, the compound of Formula XIV is Compound 1:

##STR00208##

and the method includes the steps of: [0437] a) reacting (e.g., esterifying) a compound of Formula XV with a compound of Formula XII to form a compound of Formula XVI having the following structure:

##STR00209## [0438] b) deprotecting the Boc of the compound of Formula XVI to form a compound of Formula XVII having the following structure:

##STR00210## [0439] c) cyclizing the compound of Formula XVII with a transition metal catalyst to form a compound of Formula XVII having the following structure:

##STR00211## [0440] d) removing (e.g., deprotecting) PG.sub.b of the compound of Formula XVIII to form a compound of Formula XIX having the following structure:

##STR00212## [0441] and [0442] e) coupling the compound of Formula XIX with the compound of Formula IV to form Compound 1 having the following structure:

##STR00213##

[0443] In some embodiments, step (a) includes contacting the compound of Formula XV and the compound of Formula XII with a carbodiimide coupling reagent, an anti-racemization agent, and a base. In some embodiments, the carbodiimide coupling reagent is EDCl, the anti-racemization agent is HOBt, and the base is N,N-diisopropylethylamine. In some embodiments, step (a) is carried out according to the following scheme:

##STR00214##

[0444] In some embodiments, the deprotecting step (b) includes contacting the compound of Formula XVI with an acid. In some embodiments, the acid is trifluoroacetic acid. In some embodiments, the deprotecting step (b) is carried out according to the following scheme:

##STR00215##

[0445] In some embodiments, the cyclizing step (c) includes contacting the compound of Formula XVII with a transition metal catalyst, a phosphine ligand, and a base. In some embodiments, the transition metal catalyst includes palladium, platinum, nickel, copper, and iron catalysts. In some embodiments, the transition metal catalyst is a palladium catalyst. In some embodiments, the palladium catalyst is P(tBu).sub.3 Pd G3 or Pd.sub.2(dba).sub.3. In some embodiments, the phosphine ligand is Q-Phos or [(tBu).sub.3PH]BF.sub.4. In some embodiments, the base is K.sub.3PO.sub.4.

[0446] In some embodiments, the cyclizing step (c) is carried out according to the following scheme:

##STR00216##

[0447] In some embodiments, the cyclizing step (c) is carried out according to the following scheme:

##STR00217##

[0448] In some embodiments, the cyclizing step (c) further includes contacting the compound of Formula XVIII with hydrochloric acid to form a hydrochloride salt of the compound of Formula XVIII.

[0449] In some embodiments, the deprotecting step (d) includes contacting the compound of Formula XVIII with a palladium catalyst and hydrogen gas in an organic solvent. In some embodiments, the palladium catalyst is Pd/C. In some embodiments, the organic solvent is methyl tert-butyl ether. In some embodiments, the deprotecting step (d) is carried out according to the following scheme:

##STR00218##

[0450] In some embodiments, the deprotecting step (d) further includes washing the compound of Formula XIX with N-acetyl cysteine.

[0451] In some embodiments, the deprotecting step (e) includes contacting the compound of Formula XIX and the compound of Formula IV with a coupling reagent, an anti-racemization reagent, and a base.

[0452] In some embodiments, the coupling reagent is PyBOP or EDCl. In some embodiments, the anti-racemization agent is selected from the group consisting of Oxyma, HOBt, or HOPO. In some embodiments, the base is N,N-diisopropylethylamine.

[0453] In some embodiments, the coupling step (e) is carried out according to the following scheme:

##STR00219##

[0454] In some embodiments, the coupling step (e) is carried out according to the following scheme:

##STR00220##

[0455] In some embodiments, the coupling step (e) is carried out according to the following scheme:

##STR00221##

[0456] In some embodiments, the method further includes the step of purifying Compound 1. In some embodiments, the purifying includes recrystallizing Compound 1. In some embodiments, the recrystallizing includes adding a first solvent, followed by adding a second solvent. In some embodiments, the first solvent is dioxane or 2-methyltetrahydrofuran. In some embodiments, the second solvent is diisopropyl ether or isopropyl alcohol. In some embodiments, the recrystallizing further includes adding a third solvent. In some embodiments, the third solvent is heptane.

Aziridine-containing Compounds

[0457] The compounds prepared by the methods described herein may be useful as cross-linking groups.

[0458] For example, a compound of Formula IV contains an aziridine moiety that may act as an

##STR00222##

[0459] Persons of skill in the art will be familiar with nucleophiles that can react with an aziridine electrophile. For example, a nucleophilic amino acid (e.g., cysteine, aspartic acid, glutamic acid, tyrosine, arginine, histidine, or lysine) may react with the aziridine in the Compound of Formula IV.

[0460] The compound of Formula IV may be combined with a monovalent organic moiety. Those of skill in the art are familiar with organic moieties. The monovalent organic moiety may be, for example, a small molecule (e.g., a macrocyclic small molecule), a polymer, a nucleic acid (e.g., a DNA or RNA oligonucleotide), a polypeptide, an oligosaccharide, an organometallic, or a protein, such as a mutated protein. The organic moiety may be bound to the Compound of Formula IV as disclosed herein in a variety of ways, and persons of skill in the art are familiar with methodologies of installing a synthetic intermediate of Formula IV as described herein to a monovalent organic moiety. Exemplary methods are disclosed in WO 2021/091967, WO 2022/235870, and WO 2023/060253, the disclosure of each of which is incorporated herein by reference.

[0461] Persons of skill in the art will be familiar with methods of combining carboxylic acid-containing moieties with other compounds. For example, in some embodiments, a compound of Formula IV may be reacted with an alcohol in an esterification reaction. In some embodiments, a compound of Formula IV may be reacted with an amine in a peptide coupling reaction. Those of skill in the art will be familiar with peptide coupling reactions. Nonlimiting examples of peptide coupling reagents that may be used to react the carboxylic acid moiety of Formula IV with an amine include carbodiimide coupling reagents (e.g., DCC, DIC, and EDC), uronium coupling reagents (e.g., COMU, HATU, HBTU, HCTU, TATU, TOTU, TBTU), carbonyl diimidazole (CDI), and phosphonium coupling reagents (e.g., BOP, PyBOP, BOPCI, PyAOP, PyBOP). In some embodiments, 1-hydroxybenzotriazole (HOBt) may be added to the peptide coupling reaction.

[0462] In some embodiments, the carboxylic acid may first be converted to an ester that can react directly with a nucleophile.

[0463] For example, the compound of Formula IV may be converted to a compound of Formula V:

##STR00223##

wherein R.sup.6 is

##STR00224##

[0464] In some embodiments, the compound of Formula IV or Formula V is reacted with an amine of Formula VI, or a salt thereof:

##STR00225## [0465] wherein the dotted lines represent zero, one, two, three, or four non-adjacent double bonds; [0466] A is N(H or CH.sub.3)C(O)(CH.sub.2) where the amino nitrogen is bound to the carbon atom of CH(R.sup.10), optionally substituted 3 to 6-membered cycloalkylene, optionally substituted 3 to 6-membered heterocycloalkylene, optionally substituted 6-membered arylene, or optionally substituted 5 to 10-membered heteroarylene; [0467] X.sup.1 is optionally substituted C.sub.1-C.sub.2 alkylene, NR, O, or S(O).sub.n; [0468] X.sup.2 is O or NH; [0469] X.sup.3 is N or CH; [0470] n is 0, 1, or 2; [0471] R is hydrogen, cyano, optionally substituted C.sub.1-C.sub.4 alkyl, optionally substituted C.sub.2-C.sub.4 alkenyl, optionally substituted C.sub.2-C.sub.4 alkynyl, C(O)R, C(O)OR, C(O)N(R).sub.2, S(O)R, S(O).sub.2R, or S(O).sub.2N(R).sub.2; [0472] each R is, independently, H or optionally substituted C.sub.1-C.sub.4 alkyl; [0473] Y.sup.1 is C, CH, or N; [0474] Y.sup.2, Y.sup.3, Y.sup.4, and Y.sup.7 are, independently, C or N; [0475] Y.sup.5 is CR.sup.x, CH.sub.2, or N; [0476] Y.sup.6 is C(O), CR.sup.z, CH.sub.2, or N; [0477] R.sup.x is hydrogen, halogen, optionally substituted C.sub.1-C.sub.3 alkyl, optionally substituted C.sub.1-C.sub.3 alkoxy, optionally substituted 3 to 6-membered cycloalkyl, optionally substituted 3 to 6-membered heterocycloalkyl; [0478] R.sup.y is hydrogen, halogen, optionally substituted C.sub.1-C.sub.3 alkyl, optionally substituted C.sub.1-C.sub.3 alkoxy, optionally substituted 3 to 6-membered cycloalkyl, optionally substituted 3 to 6-membered heterocycloalkyl; [0479] R.sup.1 is cyano, optionally substituted C.sub.1-C.sub.6 alkyl, optionally substituted C.sub.1-C.sub.6 heteroalkyl, optionally substituted 3 to 6-membered cycloalkyl, optionally substituted 3 to 6-membered cycloalkenyl, optionally substituted 3 to 6-membered heterocycloalkyl, optionally substituted 6 to 10-membered aryl, or optionally substituted 5 to 10-membered heteroaryl, or [0480] R.sup.1 and R.sup.2 combine with the atoms to which they are attached to form an optionally substituted 3 to 14-membered heterocycloalkyl; [0481] R.sup.2 is absent, hydrogen, optionally substituted C.sub.1-C.sub.6 alkyl, optionally substituted C.sub.2-C.sub.6 alkenyl, optionally substituted C.sub.2-C.sub.6 alkynyl, optionally substituted 3 to 6-membered cycloalkyl, optionally substituted 3 to 7-membered heterocycloalkyl, optionally substituted 6-membered aryl, optionally substituted 5 or 6-membered heteroaryl; R.sup.3 is absent, or [0482] R.sup.2 and R.sup.3 combine with the atom to which they are attached to form an optionally substituted 3 to 8-membered cycloalkyl or optionally substituted 3 to 14-membered heterocycloalkyl; [0483] R.sup.4 is absent, hydrogen, halogen, cyano, or methyl optionally substituted with 1 to 3 halogens; [0484] R.sup.5 is hydrogen, C.sub.1-C.sub.4 alkyl optionally substituted with halogen, cyano, hydroxy, or C.sub.1-C.sub.4 alkoxy, cyclopropyl, or cyclobutyl; [0485] R.sup.6 is hydrogen or methyl; R.sup.7 is hydrogen, halogen, or optionally substituted C.sub.1-C.sub.3 alkyl, or [0486] R.sup.6 and R.sup.7 combine with the carbon atoms to which they are attached to form an optionally substituted 3 to 6-membered cycloalkyl or optionally substituted 3 to 7-membered heterocycloalkyl; [0487] R.sup.8 is hydrogen, halogen, hydroxy, cyano, optionally substituted C.sub.1-C.sub.3 alkoxyl, optionally substituted C.sub.1-C.sub.3 alkyl, optionally substituted C.sub.2-C.sub.6 alkenyl, optionally substituted C.sub.2-C.sub.6 alkynyl, optionally substituted 3 to 8-membered cycloalkyl, optionally substituted 3 to 14-membered heterocycloalkyl, optionally substituted 5 to 10-membered heteroaryl, or optionally substituted 6 to 10-membered aryl, or [0488] R.sup.7 and R.sup.8 combine with the carbon atom to which they are attached to form CCR.sup.7R.sup.8; CN(OH), CN(OC.sub.1-C.sub.3 alkyl), CO, CS, CNH, optionally substituted 3 to 6-membered cycloalkyl, or optionally substituted 3 to 7-membered heterocycloalkyl; [0489] R.sup.7a and R.sup.8a are, independently, hydrogen, halo, optionally substituted C.sub.1-C.sub.3 alkyl, or combine with the carbon to which they are attached to form a carbonyl; [0490] R.sup.7 is hydrogen, halogen, or optionally substituted C.sub.1-C.sub.3 alkyl; R.sup.8 is hydrogen, halogen, hydroxy, cyano, optionally substituted C.sub.1-C.sub.3 alkoxyl, optionally substituted C.sub.1-C.sub.3 alkyl, optionally substituted C.sub.2-C.sub.6 alkenyl, optionally substituted C.sub.2-C.sub.6 alkynyl, optionally substituted 3 to 8-membered cycloalkyl, optionally substituted 3 to 14-membered heterocycloalkyl, optionally substituted 5 to 10-membered heteroaryl, or optionally substituted 6 to 10-membered aryl, or [0491] R.sup.7 and R.sup.8 combine with the carbon atom to which they are attached to form optionally substituted 3 to 6-membered cycloalkyl or optionally substituted 3 to 7-membered heterocycloalkyl; [0492] R.sup.9 is hydrogen, F, optionally substituted C.sub.1-C.sub.6 alkyl, optionally substituted C.sub.1-C.sub.6 heteroalkyl, optionally substituted 3 to 6-membered cycloalkyl, or optionally substituted 3 to 7-membered heterocycloalkyl, or [0493] R.sup.9 and L combine with the atoms to which they are attached to form an optionally substituted 3 to 14-membered heterocycloalkyl; [0494] R.sup.9 is hydrogen or optionally substituted C.sub.1-C.sub.6 alkyl; [0495] R.sup.10 is hydrogen, halo, hydroxy, C.sub.1-C.sub.3 alkoxy, or C.sub.1-C.sub.3 alkyl; [0496] R.sup.10a is hydrogen or halo; [0497] R.sup.11 is hydrogen or C.sub.1-C.sub.3 alkyl; and [0498] R.sup.34 is hydrogen or C.sub.1-C.sub.3 alkyl (e.g., methyl).

[0499] In some embodiments, the compound of Formula IV or Formula V is reacted with an amine of Formula VII, or a salt thereof:

##STR00226##

wherein A is optionally substituted 3 to 6-membered heterocycloalkylene, optionally substituted 3 to 6-membered cycloalkylene, optionally substituted 6-membered arylene, or optionally substituted 5 to 10-membered heteroarylene; R.sup.1 is hydrogen or optionally substituted 3 to 10-membered heterocycloalkyl; and R.sup.2 is optionally substituted C.sub.1-C.sub.6 alkyl, optionally substituted C.sub.3-6 cycloalkyl, or optionally substituted C.sub.6-10 aryl.

EXAMPLES

[0500] The disclosure is further illustrated by the following examples and synthesis examples, which are not to be construed as limiting this disclosure in scope or spirit to the specific procedures herein described. It is to be understood that the examples are provided to illustrate certain embodiments and that no limitation to the scope of the disclosure is intended thereby. It is to be further understood that resort may be had to various other embodiments, modifications, and equivalents thereof which may suggest themselves to those skilled in the art without departing from the spirit of the present disclosure or scope of the appended claims.

[0501] All variables described in the Examples below have the same definitions as defined in the Summary, above.

Example 1Synthesis of a Compound of Formula I

##STR00227##

[0502] To a reactor was charged a compound of Formula I racemate (8.40 mol, 1.0 equiv), EtOH (9.6 V), L(+)-Tartaric acid (0.5 equiv) and H.sub.2O (0.35 V). The reaction mixture was purged with N.sub.2 gas for three times and then stirred at 80 C. for 3 h. Then the reaction was cooled down to 25 C. slowly in 12 h. The reaction mixture was stirred at 20-25 C. for 4 h. The slurry was filtered, and the cake was washed with EtOH (1 V). The wet cake was charged with EtOH (3 V) and H.sub.2O (0.105 V) into a reactor. The mixture was purged with N.sub.2 gas for three times and then stirred at 80 C. for 3 h. Then the reaction was cooled down to 25 C. slowly in 12 h. The reaction mixture was stirred at 20-25 C. for 4 h. The slurry was filtered, and the cake was washed with EtOH (1 V) and MTBE (1 V) respectively. The cake was dried under reduced pressure (40 C., 50-80 mbar) to afford a compound of Formula I tartrate as off-white solids (100% a/a purity, 93.3% ee, 39% yield). Table 1 summarizes the HPLC method for this synthesis.

TABLE-US-00001 TABLE 1 HPLC Method for Example 1 Achiral HPLC method: Column: Agilent Poroshell 120 EC-C18 (4.6 100 mm, 2.7 m) Mobile Phase: A: 0.05% TFA in water B: 0.05% TFA in MeCN Time (min) A % B % Gradient: 0.0 95 5 10.0 5 95 12.0 5 95 12.1 95 5 17.0 95 5 Flow Rate: 1.2 mL/min UV Detector Wavelength: 210 nm Column Temperature: 40 C. Chiral HPLC method: Column: CHIRALCEL IG-3 (4.6 250 mm, 3 m) Mobile Phase: A: hexane (0.1% DEA) B: IPA (0.1% DEA) Time (min) A % B % Gradient: 0.0 75 15 60.0 75 15 Flow Rate: 0.8 mL/min UV Detector Wavelength: 250 nm Column Temperature: 40 C.

Example 2Synthetic Procedure for benzyl (2R,3R)-3-cyclopropyl-1-methylaziridine-2-carboxylate and (2R,3R)-3-cyclopropyl-1-methylaziridine-2-carboxylic acid (Compound A barium salt)

##STR00228##

Part 1Synthesis of (R,E)-N-(cyclopropylmethylene)-4-methylbenzenesulfinamide

##STR00229##

[0503] To a reactor were charged cyclopropanecarbaldehyde (5.78 kg, 82.46 mol, 2.0 equiv), pyrrolidine (14.66 g, 0.41 mol, 0.5 mol %) and DCM (128.00 L, 20 V). The reaction mixture was stirred at 20-25 C. for 30 min. The cyclopropanecarbaldehyde solution (64.00 L, 41.2 mol, 10 V) was transferred to a jacketed reactor with distillation device, followed by adding the starting material (6.4 kg, 1.0 equiv). The reaction mixture was distilled at 40-45 C. to remove H.sub.2O. When the reaction solution was less than 7 V, another portion of cyclopropanecarbaldehyde solution (19.20 L, 3 V) was added. The cyclopropanecarbaldehyde solution was added completely within 53 h. The reaction was monitored by HPLC (IPC1% a/a). After completion, the reaction was cooled to 20-25 C. and the reaction solution was filtered. The filtrate was concentrated to 4 V under reduced pressure (35-40 C.) and n-heptane (10.00 L, 1.5 V) was charged. The mixture was concentrated to 4 V under reduced pressure (45-49 C.) and n-heptane (10.00 L, 1.5 V) was charged. The process was repeated 4 times until the solvent was swapped to heptane. The mixture was stirred at 15-20 C. for 14 h. The slurry was filtered, and the wet cake was dried under reduced pressure (35-40 C., 100-200 mbar) for 16 h. The product was obtained as light-yellow solids (95.8% a/a, 93.7% w/w, 84% yield).

[0504] LCMS (ESI+): Calculated for C.sub.11H.sub.13NOS [M+H].sup.+: 208.1; found: 208.1.

[0505] .sup.1H NMR (400 MHz, DMSO-d.sub.6) 7.58-7.50 (m, 3H), 7.38 (d, J=8.0 Hz, 2H), 2.37 (s, 3H), 2.00-1.87 (m, 1H), 1.13-0.92 (m, 4H).

Part 2Synthesis of benzyl (2R,3R)-3-cyclopropyl-1-((R)-p-tolylsulfinyl)aziridine-2-carboxylate

##STR00230##

[0506] To a reactor was charged (R,E)-N-(cyclopropylmethylene)-4-methylbenzenesulfinamide (1.0 kg, 4.82 mol, 1.0 equiv), benzyl bromoacetate (1.44 kg, 6.29 mol, 1.3 equiv), THF (10.00 L, 10 V) under the protection of N.sub.2. The reaction mixture was cooled to 50 C. to 40 C. and stirred for 30 min. LiHMDS (1.0 M in THF, 4.82 mol, 4.82 L, 1.0 equiv) was added dropwise to the reaction mixture in 2.5 h at 455 C. After completion the reaction was quenched by pouring the reaction mixture into ice water (20.00 L, 20 V). MTBE (10.00 L, 10 V) was added, and the mixture was stirred at 10-20 C. for 10 min. The mixture was settled for 30 min before organic phase was separated. The aqueous phase was extracted with MTBE (10.00 L, 10 V). The combined organic phase was washed with aq. NaCl solution (10 wt %, 10.00 L, 10 V). The organic phase was concentrated under reduced pressure (25-35 C., 50-100 mbar). The residue was dissolved in MTBE (10.00 L, 10 V) and charged DABCO (81.10 g, 0.72 mol, 0.15 equiv). The mixture was stirred for 9 h and was sampled for HPLC analysis (IPC: benzyl bromoacetate=0% a/a). The mixture was washed with aq. NaCl solution (10 wt %, 10.00 L, 10 V). The organic phase was concentrated under reduced pressure (25-35 C., 50-100 mbar) to afford the crude product as brown oil (80.4% a/a purity).

[0507] The crude product combined with different batches (7.5 kg, 1.0 equiv) and isopropyl ether (5.25 L, 0.7 V) were added to a reactor. The mixture was stirred at 20-25 C. for 30 min before it was cooled to 05 C. Product seed (1.0 g) was added. The mixture was cooled to 15 C. to 20 C. and stirred at this temperature for 2 h. The slurry was filtered, and the cake was rinsed with isopropyl ether (pre-cooled to 20 C., 0.75 L, 0.1 V). The wet cake was dried to afford light yellow solids (3.1 kg, 89.1% a/a purity). The solids were dissolved in isopropyl ether (2.70 L, 0.9 V). The solution was cooled to 15 C. to 20 C. and stirred at this temperature for 1 h. The slurry was filtered, and the cake was rinsed with isopropyl ether (pre-cooled to 20 C., 0.60 L, 0.2 V). The wet cake was dried to afford light yellow solids (2.48 kg, 98.3% a/a purity) of the product.

[0508] LCMS (ESI+): Calculated for C.sub.20H.sub.21NO.sub.3S [M+H].sup.+: 356.1; found: 356.3.

[0509] .sup.1H NMR (400 MHz, DMSO-d.sub.6) 7.63-7.56 (m, 2H), 7.40-7.29 (m, 5H), 7.19-7.10 (m, 2H), 5.14-5.02 (m, 2H), 3.15 (d, J=7.1 Hz, 1H), 2.38 (s, 3H), 2.35 (d, J=7.5 Hz, 1H), 1.05-0.92 (m, 1H), 0.61-0.44 (m, 3H), 0.44-0.34 (m, 1H).

Part 3Synthesis of benzyl (2R,3R)-3-cyclopropylaziridine-2-carboxylate

##STR00231##

[0510] To a reactor were charged benzyl (2R,3R)-3-cyclopropyl-1-((R)-p-tolylsulfinyl)aziridine-2-carboxylate (3.00 kg, 8.44 mol, 1.0 equiv), acetone (30.00 L, 10 V) and H.sub.2O (15.00 L, 5 V). The reaction mixture was cooled to 10 C. to 5 C. TFA (4.81 kg, 42.20 mol, 5.0 equiv) was added to the reaction dropwise in 90 min at 15 C. to 5 C. The reaction mixture was stirred at 10 C. to 0 C. for 3 h.

[0511] The reaction was quenched with aq. ammonia solution (25% w/w, 1 V, 7.0 equiv). MTBE (45.00 L, 15 V) was added, and the phases were separated. The aqueous phase was extracted with MTBE (130.00 L, 110 V; 315.00 L, 35 V). The combined organic phase was washed with aq. NaCl solution (13% w/w, 230.00 L, 210 V). The organic phase was concentrated and swapped with MTBE (10.00 L3) to remove acetone. The MTBE solution was washed with water (30.00 L, 10 V). The organic phase was concentrated to dryness to afford oil (25-35 C., 50-100 mbar). The residue was dissolved in isopropyl ether (2.50 L, 0.42 V) and n-heptane (2.00 L, 0.33 V) was added to the solution. The solution was cooled to 10 C. and product seed (20 g) was added. The mixture was stirred for 1 h and then cooled to 5-10 C. n-heptane (2.00 L, 0.33 V) was added to the solution and the mixture was stirred at 5-10 C. for 2 h. The slurry was filtered at 5-10 C. and the cake was rinsed with isopropyl ether/n-heptane (1:5 v/v, 0.2 V). The cake was dried under reduced pressure (100-200 mbar, 25 C.) to afford the product (1.50 kg, 92.1% a/a, 89.6% w/w assay, 100% ee, 99.4% de).

[0512] LCMS (ESI+): Calculated for C.sub.13H.sub.15NO.sub.2 [M+H].sup.+: 218.1; found: 218.2.

[0513] .sup.1H NMR (400 MHz, CDCl.sub.3) 7.43-7.31 (m, 5H), 5.32-5.20 (m, 2H), 2.73 (d, J=6.0 Hz, 1H), 1.68 (dd, J=8.2, 6.0 Hz, 1H), 0.83-0.70 (m, 1H), 0.65-0.56 (m, 1H), 0.54-0.40 (m, 2H), 0.36-0.25 (m, 1H).

Part 4Synthesis of benzyl (2R,3R)-3-cyclopropyl-1-methylaziridine-2-carboxylate

##STR00232##

[0514] To a reactor were charged benzyl (2R,3R)-3-cyclopropylaziridine-2-carboxylate (10.90 kg, 50.17 mol, 1.0 equiv), DMF (109.00 L, 10 V), MeB(OH).sub.2 (9.00 kg, 150.35 mol, 3.0 equiv), Cu(OAc).sub.2 (10.10 kg, 50.17 mol, 1.0 equiv), 2,2-dipyridine (7.90 kg, 50.17 mol, 1.0 equiv), anhydrous Na.sub.2CO.sub.3 (16.00 kg, 150.96 mol, 3.0 equiv) and 4 molecular sieves (43.60 kg, 4 w/w). The reaction mixture was bubbled with 5% O.sub.2/95% N.sub.2 (10-14 L/min) while maintaining the temperature at 2010 C. The reaction was heated to 30-35 C. and stirred at the temperature for 15 h under 5% O.sub.2/95% N.sub.2. A sample was taken for HPLC analysis.

[0515] The reaction was cooled to 20 C. The reaction mixture was filtered, and the cake was rinsed with MTBE (109 L, 10 V). The filtrate was transferred to a reactor, MTBE (109 L, 10 V) and H.sub.2O (109 L, 10V) were added. The phases were separated, and the aqueous phase was extracted with MTBE (2109 L, 210 V). The combined organic phase was washed with aq. NaCl solution (5109 L, 51.0V) and H.sub.2O (163.50 L, 15 V) respectively. The organic phase was concentrated to 6-7 V under reduced pressure (s 45 C.). The concentrated solution of the product was used for the next step (92.49% a/a, 15.67% HPLC assay, 9.9 kg of product, 78.8% yield, 99.6% ee, 100% de).

Part 4bAlternative Synthesis of benzyl (2R,3R)-3-cyclopropyl-1-methylaziridine-2-carboxylate

##STR00233##

[0516] To a reactor were charged benzyl (2R,3R)-3-cyclopropylaziridine-2-carboxylate (10 g, 0.046 mole, 1.0 equiv), acetonitrile (118.5 g, 12), trimethylboroxine (8.09 g 50% THE solution, 0.7 equiv, 0.032 mol), Cu(OAc).sub.2 (8.36 kg, 0.046 mol, 1.0 equiv), 2,2-dipyridine (7.2 g, 0.046 mol, 1.0 equiv), anhydrous K.sub.2CO.sub.3 (12.72 g, 0.092 mol, 2.0 equiv), and di-tert-butyl peroxide (6.73 g, 0.046 mol, 1 equiv). The reaction was heated to 35 C. and stirred at the temperature for 23 h A sample was taken for HPLC analysis. The reaction was cooled to 20 C. The reaction mixture was filtered, and the cake was rinsed with MTBE (37 g, 3.7). The filtrate was transferred to a reactor and cooled to 0 C., after which 30% sodium thiosulfate (145.43 g, 14.5) was added dropwise. The mixture was concentrated to 10 V at 35 C. under vacuum. The residual mixture was extracted with MTBE (111 g, 11.1). The organic phase was concentrated to 2 V under vacuum. The concentrated solution of the product was used for the next step (94.0% purity, 94.0% yield).

[0517] LCMS (ESI+): Calculated for C.sub.14H.sub.17NO.sub.2 [M+H].sup.+: 232.1; found: 232.1.

[0518] .sup.1H NMR (400 MHz, DMSO-d.sub.6) 7.41-7.30 (m, 5H), 5.22-5.07 (m, 2H), 2.26 (s, 3H), 2.20 (d, J=6.6 Hz, 1H), 1.39 (t, J=6.9 Hz, 1H), 0.82-0.69 (m, 1H), 0.49-0.28 (m, 3H), 0.22-0.11 (m, 1H).

Part 5Synthesis of (2R,3R)-3-cyclopropyl-1-methylaziridine-2-carboxylic acid, % Ba salt

##STR00234##

[0519] To a reactor were charged benzyl (2R,3R)-3-cyclopropyl-1-methylaziridine-2-carboxylate (1.50 kg, 6.48 mol, 1.00 equiv), H.sub.2O (1.50 L, 1 V) and THE (15.00 L, 10 V) at 25-30 C. The reaction was cooled to 0-5 C. Ba(OH).sub.2.Math.8H.sub.2O (1.02 kg, 3.24 mol, 0.5 equiv) was added to the reaction mixture and the reaction was warmed to 25-30 C. The reaction was stirred at 25-30 C. for 16 h. A sample was taken for HPLC analysis.

[0520] The reaction was concentrated under reduced pressure (25-30 C., 50-100 mbar) and the solvent was swapped with MeCN (215.00 L, 210 V). MeCN (15.00 L, 10 V) was charged, and the mixture was stirred at 25-30 C. for 16 h. The reaction mixture was concentrated under reduced pressure (25-30 C., 50-100 mbar) and the solvent was swapped with MTBE (315.00 L, 310 V). MTBE (15.00 L, 10 V) was charged, and the mixture was stirred at 25-30 C. for 16 h. The slurry was filtered, and the cake was rinsed with MTBE (23.00 L, 22 V). The wet cake was dried under reduced pressure (25-30 C., 50-100 mbar) to afford crude (2R,3R)-3-cyclopropyl-1-methylaziridine-2-carboxylic acid, M Ba salt (1.38 kg, 58.9% w/w assay). Table 2 summarizes the HPLC method for this synthesis.

[0521] LCMS (ESI+): Calculated for C.sub.7H.sub.11NO.sub.2 [M+H].sup.+: 142.1; found: 142.1.

[0522] .sup.1H NMR (400 MHz, Methanol-d.sub.4) 2.38 (s, 3H), 2.11 (d, J=7.3 Hz, 1H), 1.23-1.12 (m, 1H), 1.00-0.86 (m, 1H), 0.67-0.53 (m, 1H), 0.52-0.40 (m, 2H), 0.37-0.24 (m, 1H).

TABLE-US-00002 TABLE 2 HPLC Method for Example 2 IPC method: Column: XBridge C18 (4.6 150 mm, 3.5 m) Mobile Phase: A: 10 mM NH.sub.4OAc in water B: MeCN Time (min) A % B % Gradient: 0.0 65 35 10.0 5 95 12.0 5 95 13.0 65 35 Flow Rate: 1.0 mL/min UV Detector Wavelength: 210 nm Column Temperature: 40 C.

Example 3Alternate Synthetic Route for benzyl (2R,3R)-3-cyclopropylaziridine-2-carboxylate

##STR00235##

Part 1Synthesis of (R,E)-N-(cyclopropylmethylene)-2-methylpropane-2-sulfinamide

##STR00236##

[0523] To a reactor were charged (R)-t-butyl-sulfinylamine (2.00 kg, 16.50 mol, 1.0 equiv), cyclopropanecarbaldehyde (1.20 kg, 17.12 mol, 1.05 equiv), 4 MS (1.60 kg, 0.8 w/w), DCM (10.00 L, 5 V) and pyrrolidine (5.87 g, 0.08 mmol). The reaction mixture was stirred at rt for 16 h at which point HPLC analysis showed reaction completion. After completion, the mixture was filtered through celite pad and rinsed with DCM (3.00 L, 1.5 V). The filtrate was concentrated to afford crude product (2.89 kg, 99.6% a/a, 94.7% assay, 95.8% yield).

[0524] LCMS (ESI+): Calculated for C.sub.8H.sub.15NOS [M+H].sup.+: 174.1; found: 174.1.

[0525] .sup.1H NMR (400 MHz, CDCl.sub.3) 7.42 (d, J=7.9 Hz, 1H), 1.94 (qt, J=8.0, 4.6 Hz, 1H), 1.13 (s, 9H), 1.08-1.01 (m, 2H), 0.95-0.86 (m, 2H).

Part 2Synthesis of benzyl (2R,3R)-1-((R)-tert-butylsulfinyl)-3-cyclopropylaziridine-2-carboxylate

##STR00237##

[0526] To a reactor were added the crude product from part 1 (1.10 kg, 6.35 mol, 1.0 equiv), benzyl bromoacetate (1.89 kg, 8.25 mol, 1.3 equiv) and THF (11.00 L, 10 V). The reaction was cooled to 70 C.-75 C. LiHMDS (1.0 M in THF, 6.35 mol, 1.0 equiv) was added dropwise in 2 h at 70 C.-75 C. and the reaction mixture was stirred for 1 h. The reaction was warmed gradually to rt and stirred for overnight at which point HPLC analysis showed reaction completion (IPC: starting material s 1% a/a, 210 nm). After completion, the reaction mixture was poured into ice water (6.05 L, 5.5 V). MTBE (11.00 L, 10 V) was added, and the mixture was stirred for 10 min at 20-25 C. The mixture was settled for 30 min and organic phase was separated. The aqueous phase was extracted with MTBE (211.00 L, 210 V). The combined organic phase was washed with aq. NaCl solution (10 wt %, 11.00 L, 10 V). The organic phase was concentrated under reduced pressure (25-35 C., 50-100 mbar). The residue was dissolved in MTBE (11.00 L, 10 V) and charged DABCO (0.21 kg, 1.90 mol, 0.3 equiv). The mixture was stirred for 2 h at 20-25 C. and was sampled for HPLC analysis (IPC: benzyl bromoacetate=0% a/a). The mixture was washed with H.sub.2O (11.00 L, 10 V). The organic phase was added aq. NaOH solution (0.1 M, 11.00 L, 10 V) and the mixture was stirred at 20-25 C. for 2 h. The reaction mixture was sampled for HPLC analysis. The organic phase was separated, and the aqueous phase was extracted with MTBE (10 V). The combined organic phase was washed with aq. NaCl solution (10 wt %, 11.00 L, 10 V) and H.sub.2O (11.00 L, 10 V) respectively. The organic solvent was concentrated under reduced pressure and MTBE (1.10 L, 1 V) was added to the residue. The mixture was cooled to 10 C. and the product seed was charged, followed by adding n-heptane (8.80 L, 8 V) in 2 h. The mixture was cooled to 5 C. and stirred for 16 h at 5 C. The slurry was filtered, and the cake was dried to afford the product (1.25 kg, 96.4% LCAP, 94.1% assay, 57.7% yield).

[0527] LCMS (ESI+): Calculated for C.sub.17H.sub.23NO.sub.3S [M+H].sup.+: 322.2; found: 322.2.

[0528] .sup.1H NMR (400 MHz, CDCl.sub.3) 7.43-7.33 (m, 5H), 5.24 (q, J=12.3 Hz, 2H), 3.41 (d, J=7.1 Hz, 1H), 2.07 (t, J=6.9 Hz, 1H), 1.24 (s, 9H), 0.55-0.47 (m, 3H), 0.35-0.26 (m, 1H).

Part 3Synthesis of benzyl (2R,3R)-3-cyclopropylaziridine-2-carboxylate

##STR00238##

[0529] To a reactor were added benzyl (2R,3R)-1-((R)-tert-butylsulfinyl)-3-cyclopropylaziridine-2-carboxylate (0.10 kg, 0.31 mol, 1.0), MTBE (1.50 L, 15 V). The reaction mixture was cooled to 10 C.5 C. HCl in dioxane (4.0 M, 1.56 mol, 5.0 equiv) was added dropwise in 35 min at 10 C. to 5 C. The reaction was stirred at 10 C. to 5 C. for 30 min and the reaction was sampled for HPLC analysis. The reaction mixture was filtered at 10 C. to 5 C. and the cake was rinsed with pre-cooled MTBE (20 C., 0.50 L, 5 V) to give the crude product as an HCl salt.

[0530] To a reactor were added MTBE (1.00 L, 10 V) and triethylamine (62.90 g, 0.62 mol, 2.0 equiv). The solution was cooled to 10 C.5 C. The crude product HCl salt was added to the reaction in 10 min at 10 C. to 5 C. The reaction was stirred at 10 C. to 5 C. for 3 h and then warmed to 0-5 C. H.sub.2O (0.5 L, 5 V) was added to the reaction and stirred for 10 min. The organic phase was separated, and the aqueous phase was extracted with MTBE (0.50 L2, 5 V2). The combined organic phase was concentrated under reduced pressure at 30 C. to give crude product. The crude was dissolved in MTBE (0.10 L, 1 V) and the solution was cooled to 15 C. n-Heptane (0.70 L, 7 V) was added dropwise over 2 h at 15 C. The mixture was cooled to 5 C. and stirred at 5 C. overnight. The slurry was filtered and dried under vacuum to afford benzyl (2R,3R)-3-cyclopropylaziridine-2-carboxylate as off-white solids (53.30 g, 99.8% LCAP, 78.8% yield).

[0531] LCMS (ESI+): Calculated for C.sub.13H.sub.15NO.sub.2 [M+H].sup.+: 218.1; found: 218.2.

[0532] .sup.1H NMR (400 MHz, CDCl.sub.3) 7.43-7.31 (m, 5H), 5.32-5.20 (m, 2H), 2.73 (d, J=6.0 Hz, 1H), 1.68 (dd, J=8.2, 6.0 Hz, 1H), 0.83-0.70 (m, 1H), 0.65-0.56 (m, 1H), 0.54-0.40 (m, 2H), 0.36-0.25 (m, 1H).

Example 4Synthetic Procedure for benzyl (S)-2-cyclopentyl-2-((S)-2,7-diazaspiro[4.4]nonan-2-yl)acetate (a compound of Formula IIIe-1)

##STR00239##

Part 1Synthesis of a Compound of Formula IIIc-1

##STR00240##

[0533] To a reactor were charged dibenzyl oxalate (1.00 kg, 3.70 mol, 1.0 equiv) and THF (30.00 L, 30.0 V) at 20-25 C. as solution A. To another reactor was charged R.sup.5MgBr (0.22 M in THF, 1.3 equiv) at 20-25 C. under N.sub.2 flow as solution B. Solution A and solution B were mixed with a flat-push flow reactor (flow rate was A: 250 mL/min, B: 182.2 mL/min, temperature was 75 to 70 C., retention time in reactor was 15 s). The reaction was monitored by HPLC at which point HPLC analysis showed reaction completion. The reaction was quenched by sat. NH.sub.4Cl (10.00 L, 10 V) aqueous solution. The mixture was stirred at 20-25 C. for 30 min. The phases were separated, and the organic phase was washed with aq. citric acid (0.5 wt %, 10.00 L, 10 V). The organic phase was concentrated under vacuum to dryness and isopropyl alcohol (2.00 L, 2 V) was added to the residue to give the product as an IPA solution. To a reactor was charged aq. NaHSO.sub.3 solution (40 wt %, 1.50 L, 1.5 V), followed by adding the product IPA solution. The mixture was stirred at 20-25 C. for 2 h. IPA (13.5 V) was charged to the mixture in 2 h. The mixture was continued to stir at 20-25 C. for 10 h. The slurry was filtered, and the cake was washed with IPA (3 V). The cake was dissolved in H.sub.2O (7 V) and aq. Na.sub.2CO.sub.3 (20 wt %, 2-2.5 V) was added dropwise to the solution to adjust pH9 while stirring at 5 C.-5 C. MTBE (5 V) was added to the solution and the mixture was stirred at rt for 30 min. The organic phase was separated and washed with aq. citric acid (0.3 V) and brine (5 V) respectively. The organic phase was concentrated under vacuum to dryness to the compound of Formula IIIc-1 as a colorless oil (99.4% LCAP, 96.8% assay, 58% yield).

Part 2Synthesis of a Compound of Formula IIId-1

##STR00241##

[0534] To a reactor were charged H.sub.2O (2.50 L, 10 V), Na.sub.2HPO.sub.4 (84.80 g, 33.9% w/w), NaH.sub.2PO.sub.4 (2.00 g, 0.82% w/w) at 20-25 C. The reaction was warmed to 30-35 C. Glucose (0.58 kg, 3.23 mol, 3.0 equiv) and NADP (6.25 g, 2.5% w/w) were added to the reaction mixture. EW-KRED-R122 (5.00 g, 2% w/w) and GDH (12.50 g, 5% w/w) were added to the reaction mixture. The reaction mixture was stirred at 30-35 C. for at least 10 min. A compound of Formula IIIc-1 (1.08 mol, 1.0 equiv) in n-heptane (0.25 L, 1 V) was added dropwise to the reaction mixture over 10 min and the reaction mixture was stirred at 30-35 C. for 20 h during which pH was checked after 8 h (the pH was controlled at 7.3-8.2 with 1M aq. NaOH solution). The reaction was monitored by HPLC at which point HPLC analysis showed reaction completion. After completion of reaction, MTBE (5.00 L, 20 V) was added, and the mixture was kept stirring for 10 min. The mixture was filtered and the cake was washed with MTBE (0.50 L, 2 V). The filtrate was settled for 10 min and the organic phase was separated. The aqueous phase was washed with MTBE (5.00 L, 20 V). The combined organic phase was washed with aq. NaCl (20 wt %, 5.00 L, 20 V). The organic phase was concentrated under vacuum to dryness to afford colorless oil as a compound of Formula IIId-1 (97.8% LCAP, 97.2% assay, 100% ee, 97.5% yield).

Part 3Synthesis of a Compound of Formula IIIe-1

##STR00242##

[0535] To a reactor were charged a compound of Formula IIId-1 (83.18 mol, 1.0 equiv) and DCM (234.00 L, 12 V) at 15-25 C., followed by adding DIPEA (21.50 kg, 166.36 mol, 2.0 equiv). The reaction mixture was cooled to 10-0 C. under N.sub.2 flow. Tf.sub.2O (31.85 kg, 116.45 mol, 1.4 equiv) was added dropwise to the reaction mixture over 1 h. The reaction mixture was kept stirring for 1 h at 10-0 C. and a sample was taken for HPLC analysis. H.sub.2O (19.50 L, 1 V) was added to the reaction at 0-5 C., followed by adding a compound of Formula I ( tartaric salt, 91.92 mol, 1.1 equiv) and K.sub.3PO.sub.4 (45.90 kg, 216.23 mol, 2.6 equiv). The reaction was warmed to 20-25 C. over 1 h and was continued to stir for 6 h. After completion, aq. HCl (0.5 M, 293.00 L, 15 V) was added to the reaction at 105 C. The mixture was stirred for 30 min and the organic phase was separated. To the organic phase was added H.sub.2O (195.00 L, 10 V) at 105 C. The mixture was stirred for 30 min and the organic phase was separated. The organic phase was concentrated to dryness to afford the product as yellow oil (93.2% LCAP, 91.6% yield). Table 3 summarizes the HPLC method for this synthesis.

TABLE-US-00003 TABLE 3 HPLC Method IPC method: Column: XBridge C18 (4.6 150 mm, 3.5 m) Mobile Phase: A: 0.05% TFA in water (v/v) B: 0.05% TFA in MeCN (v/v) Time (min) A % B % Gradient: 0.0 60 40 12.0 5 95 15.0 5 95 16.0 60 40 Flow Rate: 1.0 mL/min UV Detector Wavelength: 210 nm Column Temperature: 40 C.

Part 4Synthesis of a Compound of Formula III-1

##STR00243##

[0536] To a reactor were charged a compound of Formula IIIe-1 (68.00 mol, 1.0 equiv) and EtOAc (150.50 L, 5 V). The solution was cooled to 0-5 C. HCl in EtOAc solution (4 M, 150.50 L, 5 V) was added dropwise to the reaction. The reaction was warmed to 20-30 C. and was stirred at 20-30 C. for 6 h. The reaction was monitored by HPLC at which point HPLC analysis showed reaction completion. After completion, the reaction was cooled to 0-10 C., aq. HCl (0.5 M, 301.00 L, 10 V) was added to the reaction. The mixture was stirred at 20-30 C. for 20 min, settled and phases are separated. The organic phase was extracted with aq. HCl (0.5 M, 301.00 L, 10 V) at 0-20 C. The combined aqueous phase was washed with MTBE (2301.00 L, 210 V). The pH of aqueous phase was adjusted to 11-12 at 5-10 C. using aq. NaOH solution (20 wt %). The aqueous phase was extracted with MTBE (2301.00 L, 210 V). The combined organic phase was washed with aq. NaCl solution (26 wt %, 301.00 L, 10 V). The organic phase was concentrated to dryness under vacuum to afford a Compound of Formula III-1 crude as colorless oil (assay 90.1% yield).

[0537] To a reactor were added the compound of Formula III-1 crude (61.29 mol, 1.0 equiv) and IPA (210.00 L, 10 V) at 2010 C. In another reactor, a solution of D-(+)-2,3-dibenzoyl tartaric acid (21.96 kg, 61.29 mol, 1.0 equiv) in IPA solution (210.00 L, 10 V) was prepared. of the IPA solution of D-(+)-2,3-dibenzoyl tartaric acid (42.00 L, 2 V) was added to the solution dropwise over 1.5 h at 40-45 C. A seed of the compound of Formula III-1 (0.01 w/w) was charged to the reaction at 40-45 C. The mixture was stirred at 40-45 C. for 1.5 h. The rest of the portion () of D-(+)-2,3-dibenzoyl tartaric acid in IPA solution (168.00 L, 8 V) was added dropwise to the reaction over 6 h. The mixture was stirred at 40-45 C. for 3 h and then was cooled to 0-5 C. The mixture was continued to stir at 0-5 C. for 6 h. The slurry was filtered, and the cake was washed with IPA (42.00 L, 2 V) and MTBE (42.00 L, 2 V). The cake was dissolved in H.sub.2O (210.00 L, 10 V) and MTBE (420.00 L, 20 V) was added to the solution. The pH of aqueous phase was adjusted to 11-12 with aq. NaOH solution (10 wt %). The mixture was stirred for 45 min, then settled and the phase was separated. The organic phase was washed with aq. NaCl solution (26 wt %, 210.00 L, 10 V). The organic phase was concentrated to dryness under vacuum to afford the product as colorless oil (99.96% LCAP, 99.01% ee, 85.8% yield). Table 4 summarizes the HPLC method for this synthesis.

TABLE-US-00004 TABLE 4 HPLC Method IPC method: Column: XBridge C18 (4.6 150 mm, 3.5 m) Mobile Phase: A: 0.05% TFA in water (v/v) B: 0.05% TFA in MeCN (v/v) Time (min) A % B % Gradient: 0.0 60 40 12.0 5 95 15.0 5 95 16.0 60 40 Flow Rate: 0.8 mL/min UV Detector Wavelength: 210 nm Column Temperature: 40 C.

Example 5Alternate Synthesis for a Compound of Formula IIId-1

##STR00244##

Part 1Synthesis of a Compound of Formula IIIg

##STR00245##

[0538] To a reactor were charged H.sub.2SO.sub.4 (56.76 kg, 578.78 mol, 2.5 equiv) and H.sub.2O (660.00 L, 20 V) at 20-25 C., followed by a compound of Formula IIIf (231.51 mol, 1.0 equiv). The reaction mixture was stirred at 20-25 C. for 30 min to form a homogeneous solution. The solution was filtered via a microporous filter (0.22 m) to tank A as solution A. To another reactor were charged NaNO.sub.2 (102.45 kg, 1484.88 mol, 6.4 equiv) and H.sub.2O (800.00 L, 24.1 V) at 20-25 C. The reaction mixture was stirred at 20-25 C. for 30 min and transferred to tank B as solution B. Solution A and solution B were mixed with a continuous micro reactor. Flow rate was A: 37.7 L/h, B: 27.7 L/h, temperature was 50 C., retention time in micro reactor was 2 min. The solution was purged with N.sub.2 gas for 12 h to push most of the N.sub.02 gas into the exhaust gas absorption reactor. NaCl (198.9 kg, 6 w/w) was charged to the reactor and was kept agitating to dissolve the solids. The solution was extracted with MTBE (2994.50 L, 230 V). The organic phase was concentrated to 2-3 V under reduced pressure (0.08 MPa, 20-30 C.). The MTBE residue was swapped with n-heptane to 2-3 V under reduced pressure (0.08 Mpa, 20-30 C.) until MTBE residue is less than 3 wt % and KF is less than 0.5 wt %. EtOAc (19.90 L, 0.6 V) was added to the n-heptane residue and the mixture was heated to 452 C. The reaction mixture was stirred at 452 C. for at least 8 h and then gradually cooled to 30 C. by the speed of 1.5 C./h. The slurry was filtered, and the cake was washed with EtOAc/n-heptane (1:5 v/v, 33.00 L, 1 V). The wet cake was dried under vacuum (0.09 Mpa, 455 C.) to afford a compound of Formula IIIg as off-white solids (91.4% LCAP, 44.9% yield). Table 5 summarizes the HPLC method for this synthesis.

TABLE-US-00005 TABLE 5 HPLC Method IPC method: Column: XBridge C18 (4.6 150 mm, 3.5 m) Mobile Phase: A: 0.05% TFA in water B: 0.0% TFA in MeCN Time (min) A % B % Gradient: 0.0 90 10 10.0 50 50 13.0 5 95 16.0 5 95 17.0 90 10 Flow Rate: 0.8 mL/min UV Detector Wavelength: 210 nm Column Temperature: 40 C.

Part 2Synthesis of a Compound of Formula IIId-1

##STR00246##

[0539] A reactor was charged with dimethylacetamide (128.00 L, 5 V), DIPEA (34.44 kg, 266.46 mol, 1.5 equiv) and a compound of Formula IIIg (177.57 mol, 1.0 equiv) at 20-25 C. under N.sub.2 atmosphere. The reaction mixture was cooled to 0-10 C. and BnBr (36.45 kg, 213.11 mol, 1.2 equiv) was added dropwise. The reaction mixture was warmed to 15-25 C. and was continued to stir at 15-25 C. for 6 h. The reaction was monitored by GC at which point GC analysis showed reaction completion. DMAP (13.01 kg, 106.49 mol, 0.6 equiv) was added and the mixture was stirred for 18 h at 20-25 C. and a sample was taken for GC analysis to make sure BnBr is not detected. H.sub.2O (256.00 L, 10V) was added to the solution and the resulting solution was extracted with MTBE (2128.00 L, 25 V). The combined organic phase was washed with aq. citric acid (2128.00 L, 25 V) and H.sub.2O (128.00 L, 5 V) respectively. The organic phase was concentrated to 2-3 V under reduced pressure (0.08 Mpa, 20-35 C.). The MTBE residue was swapped with DCM (256.00 L, 10 V) to 2-3 V under reduced pressure (0.08 Mpa, 20-35 C.) until MTBE residue was less than 3% LCAP by GC analysis. A compound of Formula IIId-1 was obtained as solution in DCM (95.6% LCAP, 105% yield). Table 6 summarizes the HPLC method for this synthesis.

TABLE-US-00006 TABLE 6 HPLC Method IPC method: Column: Rtx-35 30 m*0.32 mm, 1 m Flow 2 mL/min Inject temperature 230 C. Oven temperature 40 C. for 0 min, then 20 C./min to 280 C. for 3 min Detection temperature 280 C. Split 30:1 Air flow 300 mL/min N.sub.2 flow 30 mL/min H.sub.2 flow 30 mL/min

Example 6Synthesis Procedure for a Compound of Formula IV

##STR00247##

Part 1Synthesis of a Compound of Formula IV-1

##STR00248##

[0540] In a reactor Compound A barium salt (8.5 kg, 40.88 mol, 1.0 equiv) was dissolved in DMF (56.00 L, 4 V) at 1510 C. NMI (13.4 kg, 163.22 mol, 4.0 equiv) and a compound of Formula III-1 (40.88 mol, 1.0 equiv) were added to the solution, followed by adding TCFH (13.80 kg, 49.18 mol, 1.2 equiv) solution in DMF (4.00 L, 1 V) dropwise over 1 h at 1510 C. The reaction mixture was stirred at 1510 C. for at least 30 min. The reaction was monitored by HPLC at which point HPLC analysis showed reaction completion.

[0541] The reaction mixture was cooled to 55 C. and quenched with H.sub.2O (28.00 L, 2 V). The mixture was warmed to 205 C. and stirred for 10 min. MTBE (280.00 L, 20 V), H.sub.2O (182.00 L, 13 V) and Na.sub.2CO.sub.3 (1.12 kg, 0.08 w/w) were added to the mixture and stirred for 20 min. The organic phase was separated, and the aqueous phase was extracted with MTBE (140.00 L, 10 V). The combined organic phase was washed with aq. N.sub.2CO.sub.3 solution (15 wt %, 2164.60 kg, 211.76 w/w). The organic phase was concentrated under reduced pressure (0.08 MPa, 20-40 C.) to 4-5 V. The MTBE was swapped with EtOAc by adding EtOAc (280.00 L, 20 V) and concentrated under reduced pressure (0.08 MPa, 20-40 C.) to 4-5 V, the process was repeated twice. A portion of EtOAc (210.00 L, 15 V) was added to the residue and the mixture was stirred for 40 min. The solution was cooled to 55 C. and maleic acid (2.37 kg, 20.42 mol, 0.5 equiv) was added portion-wise to the solution. A compound of Formula IVa-1 maleate seed was added to the reaction and stirred for 15 min. The solution of maleic acid (6.17 kg, 1.3 equiv) in EtOAc (280.00 L, 20 V) was added dropwise to the reaction over 7.5 h and continued to stir at 55 C. for 7.5 h. The slurry was filtered, and the cake was washed with pre-cooled EtOAc (42.00 L, 3 V) at 55 C. The wet cake was collected.

[0542] To a reactor were added MTBE (140.00 L, 10 V) and the wet cake at 05 C. aq. Na.sub.2CO.sub.3 solution (15 wt %) was added at 05 C. until the pH of aqueous layer is 9-11. The mixture was warmed to 255 C. and stirred for 30 min. The organic phase was separated, and the aqueous phase was extracted with MTBE (140.00 L, 10 V). The combined organic phase was washed with aq. Na.sub.2CO.sub.3 solution (15% wt %, 140.00 L, 10 V). The organic phase was concentrated under reduced pressure (0.08 MPa, 20-40 C.) to 4-5 V. The MTBE was swapped with MeOH by adding MeOH (280.00 L, 20 V) and concentrated under reduced pressure (0.08 MPa, 20-40 C.) to 4-5 V, the process was repeated twice. A portion of MeOH (112.00 L, 8 V) was added to the residue and the mixture was stirred for 30 min at 2010 C. A sample was taken for GC analysis (MTBE residue s 3 wt %). A compound of Formula IVa-1 was obtained as MeOH solution (97.8% LCAP, 50.7% yield).

Part 2Synthesis of a Compound of Formula IV

##STR00249##

[0543] MeOH solution of a compound of Formula IVa-1 (13.00 mol, 1.0 equiv) and TEA (4.00 kg, 39.00 mol, 3.0 equiv) were added into a reactor. The mixture was cooled to 05 C. and PdC12 (60.50 g, 1% w/w) was added to the reaction. The reaction mixture was stirred for at least 3 h at 05 C. under pressure of 0-0.15 MPa with H.sub.2. A sample was taken for HPLC analysis. The reaction mixture was filtered via diatomite and washed the filter cake with MeOH (42.35 L, 7 V). The filtrate was concentrated to 8-10 V under reduced pressure (20-30 C., 0.08 Mpa). The solvent was swapped with THE (60.50 L, 10 V) to 8-10 V under reduced pressure (20-30 C., 0.08 Mpa) twice. To the residue was added THE (60.50 L, 10 V) and TEA (2.00 kg, 19.76 mol, 1.5 equiv). The resulting solution was concentrated to 8-10 V under reduced pressure (20-30 C., 0.08 Mpa) and THE (18.15 L, 3 V) was added to the residue. The solution was stirred for at least 30 min at 2010 C. and slurry formed. MTBE (60.50 L, 10 V) was added to the slurry over 1.2 h at 2010 C. The mixture was concentrated to 10-13 V under reduced pressure (20-30 C., 0.08 Mpa). The solvent was swapped with MTBE (48.40 L, 8 V) to 10-13 V for 6 times by charging MTBE dropwise over 1 h and then concentration under reduced pressure (20-30 C., 0.08 Mpa). MTBE (48.40 L, 8 V) was added to the suspension dropwise over 1 h at 2010 C. and a sample was taken for analysis (THF residue7% w/w). The slurry was filtered, and the cake was washed with (18.15 L, 3 V). The cake was dried under vacuum (305 C., 0.08 Mpa) for at least 8 h until LOD10% w/w. The compound of Formula IV was obtained as off-white solids (95.9% LCAP, 90.2% w/w assay, 76.4% yield).

Compound of Formula IV Na Salt

[0544] MeOH solution of a compound of Formula IV (1.0 equiv) and TEA (3.0 equiv) were added into a reactor. The mixture was cooled to 05 C. and PdCl.sub.2 (1% w/w) was added to the reaction. The reaction mixture was stirred for at least 3 h at 05 C. under pressure of 0-0.15 MPa with H.sub.2. A sample was taken for HPLC analysis. The reaction mixture was filtered via diatomite and washed the filter cake with MeOH (7 V). The filtrate was concentrated to 3-4 V under reduced pressure (20-30 C., 0.08 Mpa). MeOH (7 V) was added to the residue and the solution was stirred for 10 min at rt. NaOMe (1.03 equiv, 30 wt % MeOH solution) was added to the solution at rt and the resulting mixture was stirred for at least 3 h at rt. The reaction solution was concentrated to 3-4 V under reduced pressure (20-30 C., 0.08 Mpa) to afford solution A. In another reactor was charged MTBE (60 V) and the reactor was cooled to 5-5 C. The seeds of the Compound of Formula IV Na salt (2 wt %) was added to MTBE solution and the suspension was stirred for 30 min. Solution A of the compound of Formula IV was added to MTBE over 5 h at 5-5 C. and the suspension was stirred for 2 h. The slurry was filtered, and the wet cake was washed with MTBE (2 V). The wet cake was dried at 30-40 C. for 5 h under vacuum to afford the compound of Formula IV Na salt. Table 7 summarizes the HPLC method for this synthesis.

TABLE-US-00007 TABLE 7 HPLC Method IPC method: Column: XBridge C18 (4.6 150 mm, 3.5 m) Mobile Phase: A: 10 mM NH.sub.4OAc in water B: MeOH Time (min) A % B % Gradient: 0.0 90 10 13.0 5 95 16.0 5 95 17.0 90 10 Flow Rate: 1.0 mL/min UV Detector Wavelength: 210 nm Column Temperature: 40 C.

Example 7Alternate Synthesis Route for a Compound of Formula IVa-1

##STR00250##

[0545] To a reactor were charged MTBE (90.50 mL, 5 V), a compound of Formula III-1 (52.85 mmol, 1.0 equiv), benzyl (2R,3R)-3-cyclopropyl-1-methylaziridine-2-carboxylate (12.84 g, 55.49 mmol, 1.05 equiv) and 1,5,7-triazobicyclo[4,4,0]dec-5-ene (TBD) (2.21 g, 15.86 mmol, 0.3 equiv). The reaction mixture was cooled to 5-0 C. The reaction mixture was stirred at 5-0 C. for 24 hours at which point HPLC analysis showed reaction completion. Aq. Na.sub.2CO.sub.3 solution (5 wt %, 181.00 mL, 10 V) was added to quench the reaction and the mixture was stirred at 15-25 C. for at least 10 min. The phases were separated, and the organic phase was washed with aq. Na.sub.2CO.sub.3 solution (5 wt %, 181.00 mL, 10 V). The organic phase was concentrated to dryness (T s 40 C.). The residue was dissolved in EtOAc (145.00 mL, 8 V) and the solution was cooled to 10-0 C. To the solution was added maleic acid (3.07 g, 26.42 mmol, 0.5 equiv), followed by adding a compound of Formula IVa-1 maleate seed (1 wt %) at 10-0 C. The mixture was stirred for at least 10 min before the solution of maleic acid in MTBE/EtOAc (v/v=3:2, 181.00 mL, 10 V) was added dropwise over 3 h. The suspension was stirred for 12 h at 10-0 C. The slurry was filtered, and the cake was washed with pre-cooled MTBE/EtOAc (v/v=1:2, 54.30 mL, 3 V). The filter cake was transferred to a reactor where MTBE (181.00 mL, 10 V) was cooled to 5-0 C. Aq. Na.sub.2CO.sub.3 solution (15 wt %, 181.00 mL, 10 V) was added to the solution to adjust the pH=8-10. The mixture was stirred for at least 10 min at 15-25 C. The phases were separated, and the aqueous phase was extracted with MTBE (90.50 mL, 5 V). The combined organic phase was washed with aq. Na.sub.2CO.sub.3 solution (5 wt %, 181.00 mL, 10 V). The organic phase was concentrated to dryness to afford the compound of Formula IVa-1 maleate (97.2% LCAP, 92.7% QNMR assay). Table 8 summarizes the HPLC method for this synthesis.

TABLE-US-00008 TABLE 8 HPLC Method IPC method: Column: XBridge C18 (4.6 150 mm, 3.5 m) Mobile Phase: A: 0.05% TFA in water (v/v) B: 0.05% TFA in MeCN (v/v) Time (min) A % B % Gradient: 0.0 90 10 12.0 45 55 15.0 5 95 17.0 5 95 18.0 90 10 Flow Rate: 0.8 mL/min UV Detector Wavelength: 210 nm Column Temperature: 40 C.

Example 8Synthetic Procedure for Compound Btert-butyl (S)-2,7-diazaspiro[4.4]nonane-2-carboxylate hemi((2R,3R)-2,3-dihydroxysuccinate)

##STR00251##

Part 1Synthesis of 1-(tert-butyl) 3-methyl 3-(cyanomethyl)pyrrolidine-1,3-dicarboxylate

##STR00252##

[0546] To a reactor was charged THE (16.00 L, 8 V). The reactor was cooled to 20 to 15 C. n-BuLi (2.5 M in hexane, 4.54 L, 1.3 equiv) was added to the reactor dropwise while maintaining the inner temperature between 20 to 15 C. Then di-isopropylethylamine (DIPEA, 1.24 kg, 12.25 mol, 1.4 equiv) was added dropwise at 20 to 15 C. The reaction mixture was stirred at 20 to 15 C. for 30 min under N.sub.2. The reaction mixture was cooled to 70 to 80 C. 1-(tert-butyl) 3-methyl pyrrolidine-1,3-dicarboxylate (2.00 kg, 8.72 mol, 1.0 equiv) in THE (2.00 L, 1 V) was added dropwise to the reaction mixture at 70 to 80 C. The reaction mixture was stirred at 70 to 80 C. for 30 min under N.sub.2. 2-bromoacetonitrile (1.36 kg, 11.34 mol, 1.3 equiv) in THE (2.00 L, 1 V) was added dropwise to the reaction mixture at 70 to 80 The reaction mixture was warmed to 20-30 C. The reaction was monitored by HPLC. The reaction mixture was quenched by adding 15% w/w aq. NH.sub.4Cl (16.00 L, 8V) dropwise while maintaining temperature at 2010 00 The mixture was extracted with EtOAc (20.00 L, 1 BV) and EtOAc (10.00 L, 5V). The combined organic layer was washed with 15% w/w aq. NaCl (8.00 L3, 4V). The organic layer was then concentrated (40-50 00, 10050 mbar) to afford 1-(tert-butyl) 3-methyl 3-(cyanomethyl)pyrrolidine-1,3-dicarboxylate as dark red oil (7.70 kg, 99.66% a/a purity, 51.3% w/w assay, 60% yield). LCMS (ESI): Calculated for C.sub.13H.sub.20N.sub.2O.sub.4 [M+H.sub.2O].sup.+: 286.2; found: [M+H.sub.2O]: 286.2. .sup.1H NMR (300 MHz, DMSO-d.sub.6, 25 C.) 3.71 (s, 3H), 3.68-3.64 (d, J=12 Hz, 1H), 3.38-3.25 (m, 3H), 3.00 (t, J=2.1 Hz, 2H), 2.31-2.21 (m, 1H), 2.02-1.90 (m, 1H), 1.90 (s, 9H). Table 9 summarizes the HPLC method for this synthesis.

TABLE-US-00009 TABLE 9 HPLC Method for Part 1 HPLC method: Column: Agilent Poroshell 120 EC-C18 (3 50 mm, 1.9 m) Mobile Phase: A: 0.1% H.sub.3PO.sub.4 in water B: MeCN Time (min) A % B % Gradient: 0.0 95 5 6.0 5 95 8.0 5 95 8.1 95 5 10.0 95 5 Flow Rate: 0.8 mL/min UV Detector 210 nm Wavelength: Column Temperature: 40 C. Retention Times: 1-(tert-butyl) 3-methyl pyrrolidine-1,3- dicarboxylate: 3.781 min 1-(tert-butyl) 3-methyl 3-(cyanomethyl)pyrrol- idine-1,3-dicarboxylate: 3.689 min

Part 2Synthesis of tert-butyl 6-oxo-2, 7-diazaspiro[4.4]nonane-2-carboxylate

##STR00253##

[0547] To a reactor were charged MeOH (59.00 L, 10 V), 1-(tert-butyl) 3-methyl 3-(cyanomethyl)pyrrolidine-1,3-dicarboxylate (2) (5.90 kg, 21.99 mol, 1.0 equiv) and Raney Ni (5.90 kg, 100% w/w). The reaction mixture was purged with N.sub.2 gas three times, followed by purging with H.sub.2 for three times to 1 atm. The reaction mixture was stirred at 20-30 C. for 16 hours at which point HPLC analysis showed reaction completion (IPC: 2/(2+3)1% a/a, 210 nm). The resulting mixture was filtered through diatomite (5.90 kg, 100% w/w). The cake was washed with MCOH (23.60 L, 4 V) and H.sub.2O (23.60 L, 4 V) respectively. The filtrate was concentrated (40-50 C., 80-150 mbar) to a volume of 4-5 V. DCM (47.20 L, 8 V) was added to the residue. The organic layer was separated, and the aqueous layer was extracted with CM (23.60 L, 4 V). The combined organic layers were dried over anhydrous Na.sub.2SO.sub.4 (11.80 kg) for 30 min at 20-30 C. The organic phase was filtered and concentrated under reduced pressure to 2 V (40-50 C., 80-150 mbar). MTBE (11.80 L, 2 V) was added to the residue and the mixture was concentrated to obtain solid (40-50 C., 80-150 mbar). MTBE (35.40 L, 6 V) was added to the residue and the mixture was stirred at 255 00 for 1 h. n-Heptane (11.80 L, 2 V) was added dropwise to the mixture and the mixture was stirred at 255 C. for 3-5 h. The resulting slurry was filtered and the cake was washed with n-heptane/MTBE (v/v, 1:1, 5.90 L, 1 V). The wet cake was dried under reduced pressure (40-50 C., 80-150 mbar) for 16 h. tert-butyl 6-oxo-2,7-diazaspiro[4.4]nonane-2-carboxylate was obtained as yellow solids (2.25 kg, 98.9% a/a purity, 97% w/w assay, 61% yield). LCMS (ESI): Calculated for C.sub.12H.sub.20N.sub.2O.sub.3 [M-.sup.tBu+H]: 185.1; found: [M-.sup.tBu+H]: 185.2. .sup.1H NMR (300 MHz, DMSO-d.sub.6, 25 C.) 6.38 (s, 1H), 3.66-3.25 (m, 6H), 2.23-2.12 (m, 3H), 1.80-1.77 (m, 1H), 1.45 (s, 9H). Table 10 summarizes the HPLC method for this synthesis.

TABLE-US-00010 TABLE 10 HPLC Method for Part 2 HPLC method: Column: Agilent Poroshell 120 EC-C18 (3 50 mm, 1.9 m) Mobile Phase: A: 0.1% H.sub.3PO.sub.4 in water B: MeCN Time (min) A % B % Gradient: 0.0 95 5 6.0 5 95 8.0 5 95 8.1 95 5 10.0 95 5 Flow Rate: 0.8 mL/min UV Detector 210 nm Wavelength: Column Temperature: 40 C. Retention Times: 1-(tert-butyl) 3-methyl 3-(cyanomethyl)pyrrol- idine-1,3-dicarboxylate: 3.698 min tert-butyl 6-oxo-2,7-diazaspiro[4.4]nonane-2- carboxylate: 2.906 min

Part 3Synthesis of tert-butyl 2,7-diazaspiro[4.4]nonane-2-carboxylate

##STR00254##

[0548] To a sealed autoclave reactor were charged toluene (10.00 L, 5 V), tert-butyl 6-oxo-2,7-diazaspiro[4.4]nonane-2-carboxylate (2.00 kg, 8.32 mol, 1.0 equiv), NiCl.sub.2(DME) (0.013 kg, 0.058 mol, 0.007 equiv) and PhSiH.sub.3 (1.98 kg, 18.30 mol, 2.2 equiv). The reaction mixture was purged with N.sub.2 gas three times. The reaction was stirred at 115-120 C. for 24 h at which point HPLC analysis showed reaction completion (IPC: 3/(3+4) s 2% a/a, 210 nm). The reaction mixture was cooled to room temperature. MTBE (20.00 L, 10 V) and aq. citric acid (20% w/w, 12.00 L, 6 V) were added. The mixture was stirred at 20-30 C. for 2-3 h. The aqueous layer was separated, and the organic layer was extracted with aq. citric acid (20% w/w, 2.00 L, 1 V). To the combined aqueous phase was added MTBE (10.00 L, 5 V). The aqueous layer was separated and adjusted to pH=13-14 with aq. KOH (20% w/w, 14.00 L, 7 V). The aqueous phase was extracted with MTBE (30.00 L4, 15 V). The combined organic phases were dried over anhydrous Na.sub.2SO.sub.4, and filtered. The filtrate was concentrated under reduced pressure (40-50 C., 50-300 mbar) to afford tert-butyl 2,7-diazaspiro[4.4]nonane-2-carboxylate (1.95 kg, 97.7% a/a purity, 80.8% w/w assay, 86% yield) as a brown oil. LCMS (ESI+): Calculated for C.sub.12H.sub.22N.sub.2O.sub.2 [M+H].sup.+: 227.2; found: 227.1. .sup.1H NMR (400 MHz, CDCl.sub.3) 3.52-3.31 (m, 2H), 3.31-3.16 (m, 2H), 3.01 (t, J=7.1 Hz, 2H), 2.85 (dd, J=11.0, 6.0 Hz, 1H), 2.77 (dd, J=11.0, 4.5 Hz, 1H), 1.99 (s, 1H), 1.87-1.64 (m, 4H), 1.46 (s, 9H). Table 11 summarizes the HPLC method for this synthesis.

TABLE-US-00011 TABLE 11 HPLC Method for Part 3 HPLC method: Column: Agilent Poroshell 120 EC-C18 (3 50 mm, 1.9 m) Mobile Phase: A: 0.1% H.sub.3PO.sub.4 in water B: MeCN Time (min) A % B % Gradient: 0.0 95 5 6.0 5 95 8.0 5 95 8.1 95 5 10.0 95 5 Flow Rate: 0.8 mL/min UV Detector 210 nm Wavelength: Column Temperature: 40 C. Retention Times: tert-butyl 6-oxo-2,7-diazaspiro[4.4]nonane- 2-carboxylate: 2.906 min Compound B racemate: 2.121 min

Part 4tert-butyl (S)-2,7-diazaspiro[4.4]nonane-2-carboxylate % tartaric acid (Compound B % tartaric acid salt)

##STR00255##

[0549] A reactor was charged Compound B-racemate (1.90 kg, 8.40 mol, 1.0 equiv), EtOH (18.24 L, 9.6 V), L(+)-Tartaric acid (0.63 kg, 4.20 mol, 0.5 equiv) and H.sub.2O (0.66 L, 0.35 V). The reaction mixture was purged with N.sub.2 gas for three times and then stirred at 80 C. for 3 h. Then the reaction was cooled down to 25 C. slowly in 12 h. The reaction mixture was stirred at 20-25 C. for 4 h. The slurry was filtered, and the cake was washed with EtOH (1.90 L, 1 V). The wet cake was charged with EtOH (5.70 L, 3 V) and H.sub.2O (0.20 L, 0.105 V) into a reactor. The mixture was purged with N.sub.2 gas for three times and then stirred at 80 C. for 3 h. Then the reaction was cooled down to 25 C. slowly in 12 h. The reaction mixture was stirred at 20-25 C. for 4 h. The slurry was filtered, and the cake was washed with EtOH (1.90 L, 1 V) and MTBE (1.90 L, 1 V) respectively. The cake was dried under reduced pressure (40 C., 50-80 mbar) to afford Compound B % tartrate as off-white solids (0.979 kg, 100% a/a purity, 93.3% ee, 39% yield). LCMS (ESI+): Calculated for C.sub.12H.sub.22N.sub.2O.sub.2 [M+H].sup.+: 227.2; found: 227.1. .sup.1H NMR (400 MHz, D.sub.2O) 4.28 (s, 1H), 3.44 (dt, J=12.1, 7.4 Hz, 3H), 3.40-3.30 (m, 3H), 3.25 (s, 2H), 2.02 (dt, J=11.0, 7.0 Hz, 2H), 1.96 (td, J=7.0, 3.0 Hz, 2H), 1.42 (s, 9H). Tables 12 and 13 summarize the HPLC method for this synthesis.

TABLE-US-00012 TABLE 12 Achiral HPLC Method for Part 4 Column: Agilent Poroshell 120 EC-C18 (4.6 100 mm, 2.7 m) Mobile Phase: A: 0.05% TFA in water B: 0.05% TFA in MeCN Time (min) A % B % Gradient: 0.0 95 5 10.0 5 95 12.0 5 95 12.1 95 5 17.0 95 5 Flow Rate: 1.2 mL/min UV Detector Wavelength: 210 nm Column Temperature: 40 C. Retention Times: Compound B: 4.253 min

TABLE-US-00013 TABLE 13 Chiral HPLC Method for Part 4 Column: CHIRALCEL IG-3 (4.6 250 mm, 3 m) Mobile Phase: A: hexane (0.1% DEA) B: IPA (0.1% DEA) Time (min) A % B % Gradient: 0.0 75 15 60.0 75 15 Flow Rate: 0.8 mL/min UV Detector Wavelength: 250 nm Column Temperature: 40 C. Retention Times: Compound B enantiomer: 32.026 min Compound B: 41.782 min

Example 9Alternative Synthesis of Compound Btert-butyl (S)-2,7-diazaspiro[4.4]nonane-2-carboxylate hemi((2R,3R)-2,3-dihydroxysuccinate)

##STR00256##

Part 1Synthesis of methyl 5-oxo-1-((S)-1-phenylethyl)pyrrolidine-3-carboxylate

##STR00257##

[0550] To a reactor was charged MeOH (10 L, 4 V), (S)-()-1-phenylethylamine (2.50 kg, 20.63 mol, 1.0 equiv), and dimethyl itaconate (3.26 kg, 20.61 mol, 1.0 equiv). The reaction mixture was stirred at 60-70 C. for 22 h. After completion, the reaction mixture was concentrated under vacuum to remove MeOH. Toluene (12.5 L, 5 V) was added to the residue and the mixture was stirred at 100-110 C. for 20 h. The mixture was cooled to 20-25 C. and aq. citric acid solution (20% w/w, 1.5 L) was added. The mixture was stirred at 20-25 C. for 3-5 h. The organic layer was separated and concentrated under vacuum to afford methyl 5-oxo-1-((S)-1-phenylethyl)pyrrolidine-3-carboxylate as orange oil (5.1 kg).

Part 2Synthesis of methyl (R)-3-(cyanomethyl)-5-oxo-1-((S)-1-phenylethyl)pyrrolidine-3-carboxylate

##STR00258##

[0551] To a reactor was charged THE (16 L, 10 V), and methyl 5-oxo-1-((S)-1-phenylethyl)pyrrolidine-3-carboxylate (1.70 kg, 6.87 mol, 1.0 equiv). The mixture was cooled to 75 to 85 C. LiHMDS (1.0 M, 1.15 kg, 1.0 equiv) was added and the reaction mixture was stirred at 75 to 85 C. for 2 h. BrCH.sub.2CN (0.82 kg, 6.84 mol, 1.0 equiv) was added and the reaction mixture was stirred at 75 to 85 C. for 2 h. After completion, HOAc (0.82 kg, 13.65 mol, 2.0 equiv) was added at 15-20 C. The mixture was concentrated and EtOAc (5.1 L, 3 V) was added. To the mixture was added aq. NaCl solution (25% w/w, 2.5 L, 1.5 V) and the mixture was stirred for 0.5 h. The organic phase was separated and concentrated. MTBE (3.5 L, 2 V) was charged to the residue and the mixture was stirred for 2-4 h. The resulting slurry was filtered and the wet cake was collected. The wet cake was dried to afford methyl (R)-3-(cyanomethyl)-5-oxo-1-((S)-1-phenylethyl)pyrrolidine-3-carboxylate as orange powder (0.63 kg).

Part 3Synthesis of (R)-7-((S)-1-phenylethyl)-2,7-diazaspiro[4.4]nonane-1,8-dione

##STR00259##

[0552] To a reactor was charged MeOH (4.5 L, 5 V) and Raney Ni (0.225 kg, 25% w/w) at 15-25 C. To the mixture was added methyl (R)-3-(cyanomethyl)-5-oxo-1-((S)-1-phenylethyl)pyrrolidine-3-carboxylate (0.90 kg, 3.14 mol, 1.0 equiv) and TEA (0.32 kg, 3.16 mol, 1.0 equiv). The mixture was degassed with N.sub.2 gas and then H.sub.2 (1.0 MPa) was charged to the reactor. The reaction was stirred at 50-60 C. for 24 h under the atmosphere of H.sub.2. After completion, the mixture was filtered and the filtrate was concentrated. To the concentrated residue was added 2-MeTHF (0.8 L, 1 V) and n-heptane (0.4 L, 0.5 V). The mixture was stirred at 60-70 C. for 2-4 h. The mixture was cooled and the slurry was filtered. The wet cake was dried to give (R)-7-((S)-1-phenylethyl)-2,7-diazaspiro[4.4]nonane-1,8-dione as yellow solids (0.62 kg).

Part 4Synthesis of (S)-2-((S)-1-phenylethyl)-2,7-diazaspiro[4.4]nonane

##STR00260##

[0553] To a reactor was charged 2-MeTHF (6.0 L, 5 V) and LiAlH.sub.4 (2.5 M in THF, 3.72 L, 9.29 mol, 2.0 equiv). (R)-7-((S)-1-phenylethyl)-2,7-diazaspiro[4.4]nonane-1,8-dione (1.2 kg, 4.64 mol, 1.0 equiv) was charged to the reactor dropwise at 50-60 C. The reaction mixture was stirred at 50-60 C. for 12 h. The reaction mixture was cooled to 15-25 C. and diluted with 2-MeTHF (3.6 L, 3 V). To the mixture was added H.sub.2O (0.35 L, 0.3 V), aq. NaOH solution (0.35 L, 0.3 V) and H.sub.2O (1.06 L, 0.9 V) sequentially at 15-25 C. The mixture was further stirred for 1 h. The resulting slurry was filtered and the filtrate was collected as (S)-2-((S)-1-phenylethyl)-2,7-diazaspiro[4.4]nonane in 2-MeTHF solution, which was used directly for next step (0.86 kg assay).

Part 5Synthesis of tert-butyl (S)-7-((S)-1-phenylethyl)-2,7-diazaspiro[4.4]nonane-2-carboxylate

##STR00261##

[0554] To a reactor was added 2-MeTHF solution of (S)-2-((S)-1-phenylethyl)-2,7-diazaspiro[4.4]nonane (0.86 kg assay, 3.73 mol, 1.0 equiv) and Boc.sub.2O (0.85 kg, 3.9 mol, 1.05 equiv). The resulting mixture was stirred at 15-25 C. for 1 h and then stirred at 45-55 C. for 12 h. The organic solution was washed with aq. NaCl solution (25% w/w, 2.4 L, 2.8 V). The organic layer was concentrated to afford tert-butyl (S)-7-((S)-1-phenylethyl)-2,7-diazaspiro[4.4]nonane-2-carboxylate as orange oil (1.24 kg).

Part 6Synthesis of Compound B L-tartratetert-butyl (S)-2,7-diazaspiro[4.4]nonane-2-carboxylate L-tartrate

##STR00262##

[0555] To a reactor was added IPA (4.2 L, 3.4 V) and H.sub.2O (2.1 L), tert-butyl (S)-7-((S)-1-phenylethyl)-2,7-diazaspiro[4.4]nonane-2-carboxylate (1.23 kg, 3.72 mol, 1.0 equiv), and L-tartaric acid (0.28 kg, 1.86 mol, 0.5 equiv) were added to the reactor at 15-25 C. Pd/C (0.06 kg, 5% w/w) was added to the reaction mixture under N.sub.2. The mixture was degassed with H.sub.2 (0.5 MPa) and stirred at 15-25 C. for 12 h. The mixture was filtered and the filtrate was collected. The filtrate was concentrated to 1-2 V under vacuum at 40-50 C. To the concentrated residue was added IPA (3.6 L, 3 V). The resulting mixture was stirred at 40-50 C. for 2 h. The mixture was cooled to 15 to 25 C. and stirred for 16 h. The slurry was filtered and the wet cake was collected. The wet cake was dried to afford Compound B % tartrate as white solids (0.90 kg).

Example 10Synthetic Procedure for Compound Dbenzyl (S)-2-cyclopentyl-2-((S)-2,7-diazaspiro[4.4]nonan-2-yl)acetate

##STR00263##

Part 1Synthesis of benzyl 2-cyclopentyl-2-oxoacetate

##STR00264##

[0556] To a reactor were charged dibenzyl oxalate (1.00 kg, 3.70 mol, 1.0 equiv) and THF (30.00 L, 30.0 V) at 20-25 C. as solution A. To another reactor was charged cyclopentyl magnesium bromide (0.22 M in THF, 4.81 mol, 1.3 equiv) at 20-25 C. under N.sub.2 flow as solution B. Solution A and solution B were mixed with a flat-push flow reactor (flow rate was A: 250 mL/min, B: 182.2 mL/min, temperature was 75 to 70 C., retention time in reactor was 15 s). The reaction was monitored by HPLC at which point HPLC analysis showed reaction completion. The reaction was quenched by sat. NH.sub.4Cl (10.00 L, 10 V) aqueous solution. The mixture was stirred at 20-25 C. for 30 min. The phases were separated, and the organic phase was washed with aq. citric acid (0.5 wt %, 10.00 L, 10 V). The organic phase was concentrated under vacuum to dryness and isopropyl alcohol (2.00 L, 2 V) was added to the residue to give benzyl 2-cyclopentyl-2-oxoacetate IPA solution. To a reactor was charged aq. NaHSO.sub.3 solution (40 wt %, 1.50 L, 1.5 V), followed by adding the benzyl 2-cyclopentyl-2-oxoacetate IPA solution. The mixture was stirred at 20-25 C. for 2 h. IPA (13.50 L, 13.5 V) was charged to the mixture in 2 h. The mixture was continued to stir at 20-25 C. for 10 h. The slurry was filtered, and the cake was washed with IPA (3.00 L, 3 V). The cake was dissolved in H.sub.2O (7.00 L, 7 V) and aq. Na.sub.2CO.sub.3 (20 wt %, 2.00-2.50 L, 2-2.5 V) was added dropwise to the solution to adjust pH9 while stirring at 5 C.-5 C. MTBE (5.00 L, 5 V) was added to the solution and the mixture was stirred at rt for 30 min. The organic phase was separated and washed with aq. citric acid (0.5 wt %, 3.00 L, 3 V) and brine (5.00 L, 5 V) respectively. The organic phase was concentrated under vacuum to dryness to afford benzyl 2-cyclopentyl-2-oxoacetate as a colorless oil (543 g, 99.4% LCAP, 96.8% assay, 58% yield).

[0557] LCMS (ESI+): Calculated for C.sub.14H.sub.16O.sub.3 [M+NH.sub.4].sup.+: 250.1; found: 250.0.

[0558] .sup.1H NMR (400 MHz, DMSO-d.sub.6) 7.59-7.26 (m, 5H), 5.28 (s, 2H), 3.47 (tt, J=8.9, 6.8 Hz, 1H), 1.97-1.77 (m, 2H), 1.77-1.63 (m, 2H), 1.63-1.43 (m, 4H).

Part 2Synthesis of benzyl (R)-2-cyclopentyl-2-hydroxyacetate

##STR00265##

[0559] To a reactor were charged H.sub.2O (2.50 L, 10 V), Na.sub.2HPO.sub.4 (84.80 g, 33.9% w/w), NaH.sub.2PO.sub.4 (2.00 g, 0.82% w/w) at 20-25 C. The reaction was warmed to 30-35 C. Glucose (0.58 kg, 3.23 mol, 3.0 equiv) and NADP (6.25 g, 2.5% w/w) were added to the reaction mixture. EW-KRED-R122 (5.00 g, 2% w/w) and GDH (12.50 g, 5% w/w) were added to the reaction mixture. The reaction mixture was stirred at 30-35 C. for at least 10 min. Benzyl 2-cyclopentyl-2-oxoacetate (0.25 kg, 1.08 mol, 1.0 equiv) in n-heptane (0.25 L, 1 V) was added dropwise to the reaction mixture over 10 min and the reaction mixture was stirred at 30-35 C. for 20 h during which pH was checked after 8 h (the pH was controlled at 7.3-8.2 with 1M aq. NaOH solution). The reaction was monitored by HPLC at which point HPLC analysis showed reaction completion. After completion of reaction, MTBE (5.00 L, 20 V) was added, and the mixture was kept stirring for 10 min. The mixture was filtered and the cake was washed with MTBE (0.50 L, 2 V). The filtrate was settled for 10 min and the organic phase was separated. The aqueous phase was washed with MTBE (5.00 L, 20 V). The combined organic phase was washed with aq. NaCl (20 wt %, 5.00 L, 20 V). The organic phase was concentrated under vacuum to dryness to afford benzyl (R)-2-cyclopentyl-2-hydroxyacetate as a colorless oil (245 g, 97.8% LCAP, 97.2% assay, 100% ee, 97.5% yield).

[0560] LCMS (ESI+): Calculated for C.sub.14H.sub.18O.sub.3 [M+Na].sup.+: 257.1; found: 257.2.

[0561] .sup.1H NMR (400 MHz, DMSO-d.sub.6) 7.47-7.27 (m, 5H), 5.38 (d, J=6.1 Hz, 1H), 5.13 (d, J=1.2 Hz, 2H), 3.93 (t, J=6.2 Hz, 1H), 2.28-2.05 (m, 1H), 1.65-1.49 (m, 4H), 1.49-1.27 (m, 4H).

Part 3Synthesis of tert-butyl (S)-7-((S)-2-(benzyloxy)-1-cyclopentyl-2-oxoethyl)-2,7-diazaspiro[4.4]nonane-2-carboxylate

##STR00266##

[0562] A reactor was charged with benzyl (R)-2-cyclopentyl-2-hydroxyacetate (19.45 kg, 83.18 mol, 1.0 equiv) and DCM (234.00 L, 12 V) at 15-25 C., followed by adding DIPEA (21.50 kg, 166.36 mol, 2.0 equiv). The reaction mixture was cooled to 10-0 C. under N.sub.2 flow. Tf.sub.2O (31.85 kg, 116.45 mol, 1.4 equiv) was added dropwise to the reaction mixture over 1 h. The reaction mixture was kept stirring for 1 h at 10-0 C. and a sample was taken for HPLC analysis. H.sub.2O (19.50 L, 1 V) was added to the reaction at 0-5 C., followed by adding Compound B ( tartaric salt, 27.70 kg, 91.92 mol, 1.1 equiv) and K.sub.3PO.sub.4 (45.90 kg, 216.23 mol, 2.6 equiv). The reaction was warmed to 20-25 C. over 1 h and was continued to stir for 6 h. After completion, aq. HCl (0.5 M, 293.00 L, 15 V) was added to the reaction at 105 C. The mixture was stirred for 30 min and the organic phase was separated. To the organic phase was added H.sub.2O (195.00 L, 10 V) at 105 C. The mixture was stirred for 30 min and the organic phase was separated. The organic phase was concentrated to dryness to afford tert-butyl (S)-7-((S)-2-(benzyloxy)-1-cyclopentyl-2-oxoethyl)-2,7-diazaspiro[4.4]nonane-2-carboxylate as yellow oil (33.76 kg, 93.2% LCAP, 91.6% yield). Table 14 summarizes the HPLC method for this synthesis.

[0563] LCMS (ESI+): Calculated for C.sub.26H.sub.38N.sub.2O.sub.4 [M+H].sup.+: 443.3; found: 443.4.

[0564] .sup.1H NMR (400 MHz, DMSO-d.sub.6) 7.63-7.15 (m, 5H), 5.13 (d, J=1.3 Hz, 2H), 3.24 (dt, J=11.5, 5.1 Hz, 1H), 3.19 (d, J=8.0 Hz, 1H), 3.14 (d, J=10.4 Hz, 1H), 3.09 (d, J=10.6 Hz, 1H), 3.04 (d, J=10.4 Hz, 1H), 2.79 (dt, J=12.8, 5.4 Hz, 1H), 2.68 (t, J=7.7 Hz, 1H), 2.61 (dd, J=8.8, 3.1 Hz, 1H), 2.52 (d, J=1.8 Hz, 1H), 2.29-2.11 (m, 1H), 1.84-1.64 (m, 3H), 1.60 (t, J=7.0 Hz, 2H), 1.57-1.44 (m, 5H), 1.39 (s, 10H), 1.16-1.03 (m, 1H).

TABLE-US-00014 TABLE 14 HPLC Method for Parts 2 and 3 IPC method: Column: XBridge C18 (4.6 150 mm, 3.5 m) Mobile Phase: A: 0.05% TFA in water (v/v) B: 0.05% TFA in MeCN (v/v) Time (min) A % B % Gradient: 0.0 60 40 12.0 5 95 15.0 5 95 16.0 60 40 Flow Rate: 1.0 mL/min UV Detector Wavelength: 210 nm Column Temperature: 40 C. Retention Times: benzyl (R)-2-cyclopentyl-2-hydroxyacetate: 6.79 min benzyl (R)-2-cyclopentyl-2-trifylacetate: 11.18 min tert-butyl (S)-7-((S)-2-(benzyloxy)-1-cyclopentyl-2-oxoethyl)- 2,7-diazaspiro[4.4]nonane-2-carboxylate: 5.62 min

Part 4Synthesis of benzyl (S)-2-cyclopentyl-2-((S)-2,7-diazaspiro[4.4]nonan-2-yl)acetate

##STR00267##

[0565] To a reactor were charged tert-butyl (S)-7-((S)-2-(benzyloxy)-1-cyclopentyl-2-oxoethyl)-2,7-diazaspiro[4.4]nonane-2-carboxylate (30.1 kg, 68.00 mol, 1.0 equiv) and EtOAc (150.50 L, 5 V). The solution was cooled to 0-5 C. HCl in EtOAc solution (4 M, 150.50 L, 5 V) was added dropwise to the reaction. The reaction was warmed to 20-30 C. and was stirred at 20-30 C. for 6 h. The reaction was monitored by HPLC at which point HPLC analysis showed reaction completion. After completion, the reaction was cooled to 0-10 C., aq. HCl (0.5 M, 301.00 L, 10 V) was added to the reaction. The mixture was stirred at 20-30 C. for 20 min, settled and phases are separated. The organic phase was extracted with aq. HCl (0.5 M, 301.00 L, 10 V) at 0-20 C. The combined aqueous phase was washed with MTBE (2301.00 L, 210 V). The pH of aqueous phase was adjusted to 11-12 at 5-10 C. using aq. NaOH solution (20 wt %). The aqueous phase was extracted with MTBE (2301.00 L, 210 V). The combined organic phase was washed with aq. NaCl solution (26 wt %, 301.00 L, 10 V). The organic phase was concentrated to dryness under vacuum to afford crude benzyl (S)-2-cyclopentyl-2-((S)-2,7-diazaspiro[4.4]nonan-2-yl)acetate as colorless oil (assay 20.99 kg, 90.1% yield).

[0566] To a reactor were added crude benzyl (S)-2-cyclopentyl-2-((S)-2,7-diazaspiro[4.4]nonan-2-yl)acetate (20.99 kg, 61.29 mol, 1.0 equiv) and IPA (210.00 L, 10 V) at 2010 C. In another reactor, a solution of D-(+)-2,3-dibenzoyl tartaric acid (21.96 kg, 61.29 mol, 1.0 equiv) in IPA solution (210.00 L, 10 V) was prepared. One fifth of the IPA solution of D-(+)-2,3-dibenzoyl tartaric acid (42.00 L, 2 V) was added to the benzyl (S)-2-cyclopentyl-2-((S)-2,7-diazaspiro[4.4]nonan-2-yl)acetate solution dropwise over 1.5 h at 40-45 C. A seed of benzyl (S)-2-cyclopentyl-2-((S)-2,7-diazaspiro[4.4]nonan-2-yl)acetate (0.21 kg, 0.01 w/w) was charged to the reaction at 40-45 C. then. The mixture was stirred at 40-45 C. for 1.5 h. The rest of portion () of D-(+)-2,3-dibenzoyl tartaric acid in IPA solution (168.00 L, 8 V) was added dropwise to the reaction over 6 h. The mixture was stirred at 40-45 C. for 3 h and then was cooled to 0-5 C. The mixture was continued to stir at 0-5 C. for 6 h. The slurry was filtered, and the cake was washed with IPA (42.00 L, 2 V) and MTBE (42.00 L, 2 V). The cake was dissolved in H.sub.2O (210.00 L, 10 V) and MTBE (420.00 L, 20 V) was added to the solution. The pH of aqueous phase was adjusted to 11-12 with aq. NaOH solution (10 wt %). The mixture was stirred for 45 min, then settled and the phase was separated. The organic phase was washed with aq. NaCl solution (26 wt %, 210.00 L, 10 V). The organic phase was concentrated to dryness under vacuum to afford benzyl (S)-2-cyclopentyl-2-((S)-2,7-diazaspiro[4.4]nonan-2-yl)acetate as colorless oil (19.98 kg, 99.96% LCAP, 99.01% ee, 85.8% yield). Table 15 summarizes the HPLC method for this synthesis.

[0567] LCMS (ESI+): Calculated for C.sub.21H.sub.30N.sub.2O.sub.2 [M+H].sup.+: 343.2; found: 343.3.

[0568] .sup.1H NMR (400 MHz, CDCl.sub.3) 7.82-7.30 (m, 4H), 5.15 (s, 2H), 3.13 (d, J=10.6 Hz, 1H), 2.92 (dtd, J=16.7, 9.4, 5.1 Hz, 3H), 2.82 (d, J=10.8 Hz, 1H), 2.79-2.65 (m, 3H), 2.60 (d, J=8.6 Hz, 1H), 2.27 (dq, J=10.4, 7.8 Hz, 1H), 1.86-1.74 (m, 1H), 1.69 (tdd, J=10.0, 6.2, 4.2 Hz, 4H), 1.63-1.48 (m, 4H), 1.48-1.38 (m, 1H), 1.20 (s, 2H).

TABLE-US-00015 TABLE 15 HPLC Method for Part 4 IPC method: Column: XBridge C18 (4.6 150 mm, 3.5 m) Mobile Phase: A: 0.05% TFA in water (v/v) B: 0.05% TFA in MeCN (v/v) Time (min) A % B % Gradient: 0.0 60 40 12.0 5 95 15.0 5 95 16.0 60 40 Flow Rate: 0.8 mL/min UV Detector Wavelength: 210 nm Column Temperature: 40 C. Retention Times: tert-butyl (S)-7-((S)-2-(benzyloxy)-1-cyclopentyl-2-oxoethyl)- 2,7-diazaspiro[4.4]nonane-2-carboxylate: 10.65 min benzyl (S)-2-cyclopentyl-2-((S)-2,7-diazaspiro[4.4]nonan-2- yl)acetate: 7.44 min

Example 11Alternative Synthetic Route for benzyl (R)-2-cyclopentyl-2-hydroxyacetate

##STR00268##

Part 1Synthesis of (R)-2-cyclopentyl-2-hydroxyacetic acid

##STR00269##

[0569] A reactor was charged with H.sub.2SO.sub.4 (56.76 kg, 578.78 mol, 2.5 equiv) and H.sub.2O (660.00 L, 20 V) at 20-25 C., followed by (R)-2-amino-2-cyclopentylacetic acid (33.15 kg, 231.51 mol, 1.0 equiv). The reaction mixture was stirred at 20-25 C. for 30 min to form a homogeneous solution. The solution was filtered via a microporous filter (0.22 m) to tank A as solution A. To another reactor were charged NaNO.sub.2 (102.45 kg, 1484.88 mol, 6.4 equiv) and H.sub.2O (800.00 L, 24.1 V) at 20-25 C. The reaction mixture was stirred at 20-25 C. for 30 min and transferred to tank B as solution B. Solution A and solution B were mixed with a continuous micro reactor. Flow rate was A: 37.7 L/h, B: 27.7 L/h, temperature was 50 C., retention time in micro reactor was 2 min. The solution was purged with N.sub.2 gas for 12 h to push most of the NO.sub.2 gas into the exhaust gas absorption reactor. NaCl (198.9 kg, 6 w/w) was charged to the reactor and was kept agitating to dissolve the solids. The solution was extracted with MTBE (2994.50 L, 230 V). The organic phase was concentrated to 2-3 V under reduced pressure (0.08 MPa, 20-30 C.). The MTBE residue was swapped with n-heptane to 2-3 V under reduced pressure (0.08 Mpa, 20-30 C.) until MTBE residue is less than 3 wt % and KF is less than 0.5 wt %. EtOAc (19.90 L, 0.6 V) was added to the n-heptane residue and the mixture was heated to 452 C. The reaction mixture was stirred at 452 C. for at least 8 h and then gradually cooled to 30 C. by the speed of 1.5 C./h. The slurry was filtered, and the cake was washed with EtOAc/n-heptane (1:5 v/v, 33.00 L, 1 V). The wet cake was dried under vacuum (0.09 Mpa, 455 C.) to afford (R)-2-cyclopentyl-2-hydroxyacetic acid as off-white solids (13.3 kg, 91.4% LCAP, 44.9% yield). Table 16 summarizes the HPLC method for this synthesis.

[0570] LCMS (ESI+): Calculated for C.sub.7H.sub.12O.sub.3 [MH.sub.2O].sup.+: 127.1; found: 127.3.

[0571] .sup.1H NMR (400 MHz, DMSO-d.sub.6) 3.80 (d, J=5.9 Hz, 1H), 2.13 (td, J=7.9, 5.9 Hz, 1H), 1.71-1.50 (m, 4H), 1.50-1.30 (m, 4H).

TABLE-US-00016 TABLE 16 HPLC Method for Part 1 IPC method: Column: XBridge C18 (4.6 150 mm, 3.5 m) Mobile Phase: A: 0.05% TFA in water B: 0.0% TFA in MeCN Time (min) A % B % Gradient: 0.0 90 10 10.0 50 50 13.0 5 95 16.0 5 95 17.0 90 10 Flow Rate: 0.8 mL/min UV Detector 210 nm Wavelength: Column Temperature: 40 C. Retention Times: (R)-2-amino-2-cyclopentylacetic acid: 3.63 min (R)-2-cyclopentyl-2-hydroxyacetic acid: 7.32 min

Part 2Synthesis of benzyl (R)-2-cyclopentyl-2-hydroxyacetate

##STR00270##

[0572] A reactor was charged with dimethylacetamide (128.00 L, 5 V), DIPEA (34.44 kg, 266.46 mol, 1.5 equiv) and (R)-2-cyclopentyl-2-hydroxyacetic acid (25.6 kg, 177.57 mol, 1.0 equiv) at 20-25 C. under N.sub.2 atmosphere. The reaction mixture was cooled to 0-10 C. and BnBr (36.45 kg, 213.11 mol, 1.2 equiv) was added dropwise. The reaction mixture was warmed to 15-25 C. and was continued to stir at 15-25 C. for 6 h. The reaction was monitored by GC at which point GC analysis showed reaction completion. DMAP (13.01 kg, 106.49 mol, 0.6 equiv) was added and the mixture was stirred for 18 h at 20-25 C. and a sample was taken for GC analysis to make sure BnBr is not detected. H.sub.2O (256.00 L, 10V) was added to the solution and the resulting solution was extracted with MTBE (2128.00 L, 25 V). The combined organic phase was washed with aq. citric acid (2128.00 L, 25 V) and H.sub.2O (128.00 L, 5 V) respectively. The organic phase was concentrated to 2-3 V under reduced pressure (0.08 Mpa, 20-35 C.). The MTBE residue was swapped with DCM (256.00 L, 10 V) to 2-3 V under reduced pressure (0.08 Mpa, 20-35 C.) until MTBE residue was less than 3% LCAP by GC analysis. Benzyl (R)-2-cyclopentyl-2-hydroxyacetate was obtained as solution in DCM (44.02 kg assay, 95.6% LCAP, 105% yield). Table 17 summarizes the HPLC method for this synthesis.

[0573] LCMS (ESI+): Calculated for C.sub.14H.sub.18O.sub.3 [M+Na].sup.+: 257.1; found: 257.2.

[0574] .sup.1H NMR (400 MHz, DMSO-d.sub.6) 7.47-7.27 (m, 5H), 5.38 (d, J=6.1 Hz, 1H), 5.13 (d, J=1.2 Hz, 2H), 3.93 (t, J=6.2 Hz, 1H), 2.28-2.05 (m, 1H), 1.65-1.49 (m, 4H), 1.49-1.27 (m, 4H).

TABLE-US-00017 TABLE 17 HPLC Method for Parts 1 and 2 IPC method: Column: Rtx-35 30 m*0.32 mm, 1 m Flow 2 mL/min Inject temperature 230 C. Oven temperature 40 C. for 0 min, then 20 C./min to 280 C. for 3 min Detection temperature 280 C. Split 30:1 Air flow 300 mL/min N.sub.2 flow 30 mL/min H.sub.2 flow 30 mL/min Retention times (R)-2-cyclopentyl-2-hydroxyacetic acid: 8.48 min benzyl (R)-2-cyclopentyl-2-hydroxy- acetate: 12.02 min

Example 12Synthetic Procedure for (S)-2-cyclopentyl-2-((S)-7-((2R,3R)-3-cyclopropyl-1-methylaziridine-2-carbonyl)-2,7-diazaspiro[4.4]nonan-2-yl)acetic acid (Compound E) and the sodium salt thereof

##STR00271##

Part 1Synthesis of benzyl (S)-2-cyclopentyl-2-((S)-7-((2R,3R)-3-cyclopropyl-1-methylaziridine-2-carbonyl)-2,7-diazaspiro[4.4]nonan-2-yl)acetate

##STR00272##

[0575] In a reactor, Compound A Ba salt (8.5 kg, 40.88 mol, 1.0 equiv) was dissolved in DMF (56.00 L, 4 V) at 1510 C. NMI (13.4 kg, 163.22 mol, 4.0 equiv) and Compound D (14.00 kg, 40.88 mol, 1.0 equiv) were added to the solution, followed by adding TCFH (13.80 kg, 49.18 mol, 1.2 equiv) solution in DMF (4.00 L, 1 V) dropwise over 1 h at 1510 C. The reaction mixture was stirred at 1510 C. for at least 30 min. The reaction was monitored by HPLC at which point HPLC analysis showed reaction completion. The reaction mixture was cooled to 55 C. and quenched with H.sub.2O (28.00 L, 2 V). The mixture was warmed to 205 C. and stirred for 10 min. MTBE (280.00 L, 20 V), H.sub.2O (182.00 L, 13 V) and Na.sub.2CO.sub.3 (1.12 kg, 0.08 w/w) were added to the mixture and stirred for 20 min. The organic phase was separated, and the aqueous phase was extracted with MTBE (140.00 L, 10 V). The combined organic phase was washed with aq. N.sub.2CO.sub.3 solution (15 wt %, 2164.60 kg, 211.76 w/w). The organic phase was concentrated under reduced pressure (0.08 MPa, 20-40 C.) to 4-5 V. The MTBE was swapped with EtOAc by adding EtOAc (280.00 L, 20 V) and concentrated under reduced pressure (0.08 MPa, 20-40 C.) to 4-5 V, the process was repeated twice. A portion of EtOAc (210.00 L, 15 V) was added to the residue and the mixture was stirred for 40 min. The solution was cooled to 55 C. and maleic acid (2.37 kg, 20.42 mol, 0.5 equiv) was added portion-wise to the solution. A seed of benzyl (S)-2-cyclopentyl-2-((S)-7-((2R,3R)-3-cyclopropyl-1-methylaziridine-2-carbonyl)-2,7-diazaspiro[4.4]nonan-2-yl)acetate as the maleate salt was added to the reaction and stirred for 15 min. The solution of maleic acid (6.17 kg, 1.3 equiv) in EtOAc (280.00 L, 20 V) was added dropwise to the reaction over 7.5 h and continued to stir at 55 C. for 7.5 h. The slurry was filtered, and the cake was washed with pre-cooled EtOAc (42.00 L, 3 V) at 55 C. The wet cake was collected. To a reactor were added MTBE (140.00 L, 10 V) and the wet cake at 05 C. aq. Na.sub.2CO.sub.3 solution (15 wt %) was added at 05 C. until the pH of aqueous layer is 9-11. The mixture was warmed to 255 C. and stirred for 30 min. The organic phase was separated, and the aqueous phase was extracted with MTBE (140.00 L, 10 V). The combined organic phase was washed with aq. Na.sub.2CO.sub.3 solution (15% wt %, 140.00 L, 10 V). The organic phase was concentrated under reduced pressure (0.08 MPa, 20-40 C.) to 4-5 V. The MTBE was swapped with MeOH by adding MeOH (280.00 L, 20 V) and concentrated under reduced pressure (0.08 MPa, 20-40 C.) to 4-5 V, the process was repeated twice. A portion of MeOH (112.00 L, 8 V) was added to the residue and the mixture was stirred for 30 min at 2010 C. A sample was taken for GC analysis (MTBE residue s 3 wt %). Benzyl (S)-2-cyclopentyl-2-((S)-7-((2R,3R)-3-cyclopropyl-1-methylaziridine-2-carbonyl)-2,7-diazaspiro[4.4]nonan-2-yl)acetate was obtained as MeOH solution (128.7 kg solution, 9.65 kg benzyl (S)-2-cyclopentyl-2-((S)-7-((2R,3R)-3-cyclopropyl-1-methylaziridine-2-carbonyl)-2,7-diazaspiro[4.4]nonan-2-yl)acetate by assay, 97.8% LCAP, 50.7% yield).

[0576] LCMS (ESI+): Calculated for C.sub.28H.sub.39N.sub.3O.sub.3 [M+H].sup.+: 466.3; found: 466.6.

[0577] .sup.1H NMR (400 MHz, Methanol-d.sub.4) 7.49-7.30 (m, 5H), 6.27 (s, 4H), 5.41-5.20 (m, 2H), 3.87-3.69 (m, 2H), 3.69-3.32 (m, 7H), 2.83 (d, J=2.4 Hz, 3H), 2.55 (dt, J=12.0, 8.2 Hz, 1H), 2.45 (t, J=7.5 Hz, 1H), 2.27-1.96 (m, 4H), 1.96-1.84 (m, 1H), 1.77-1.63 (m, 1H), 1.53 (d, J=4.8 Hz, 5H), 1.37-1.23 (m, 1H), 0.85-0.61 (m, 4H), 0.53 (tq, J=6.3, 4.3 Hz, 1H).

Part 2Synthesis of (S)-2-cyclopentyl-2-((S)-7-((2R,3R)-3-cyclopropyl-1-methylaziridine-2-carbonyl)-2,7-diazaspiro[4.4]nonan-2-yl)acetic acid (Compound E)

##STR00273##

[0578] A MeOH solution of benzyl (S)-2-cyclopentyl-2-((S)-7-((2R,3R)-3-cyclopropyl-1-methylaziridine-2-carbonyl)-2,7-diazaspiro[4.4]nonan-2-yl)acetate (6.05 kg assay, 13.00 mol, 1.0 equiv) and TEA (4.00 kg, 39.00 mol, 3.0 equiv) were added into a reactor. The mixture was cooled to 05 C. and PdCl.sub.2 (60.50 g, 1% w/w) was added to the reaction. The reaction mixture was stirred for at least 3 h at 05 C. under pressure of 0-0.15 MPa with H.sub.2. A sample was taken for HPLC analysis. The reaction mixture was filtered via diatomite and washed the filter cake with MeOH (42.35 L, 7 V). The filtrate was concentrated to 8-10 V under reduced pressure (20-30 C., 0.08 Mpa). The solvent was swapped with THE (60.50 L, 10 V) to 8-10 V under reduced pressure (20-30 C., 0.08 Mpa) twice. To the residue was added THE (60.50 L, 10 V) and TEA (2.00 kg, 19.76 mol, 1.5 equiv). The resulting solution was concentrated to 8-10 V under reduced pressure (20-30 C., 0.08 Mpa) and THE (18.15 L, 3 V) was added to the residue. The solution was stirred for at least 30 min at 2010 C. and slurry formed. MTBE (60.50 L, 10 V) was added to the slurry over 1.2 h at 2010 C. The mixture was concentrated to 10-13 V under reduced pressure (20-30 C., 0.08 Mpa). The solvent was swapped with MTBE (48.40 L, 8 V) to 10-13 V for 6 times by charging MTBE dropwise over 1 h and then concentration under reduced pressure (20-30 C., 0.08 Mpa). MTBE (48.40 L, 8 V) was added to the suspension dropwise over 1 h at 2010 C. and a sample was taken for analysis (THF residue7% w/w). The slurry was filtered, and the cake was washed (18.15 L, 3 V). The cake was dried under vacuum (305 C., 0.08 Mpa) for at least 8 h until LOD10% w/w. Compound E was obtained as off-white solids (4.13 kg, 95.9% LCAP, 90.2% w/w assay, 76.4% yield).

[0579] LCMS (ESI+): Calculated for C.sub.21H.sub.33N.sub.3O.sub.3 [M+H].sup.+: 376.3; found: 376.4.

[0580] .sup.1H NMR (400 MHz, Methanol-d.sub.4) 3.87-3.76 (m, 1H), 3.73-3.63 (m, 1H), 3.63-3.38 (m, 7H), 2.44-2.25 (m, 5H), 2.19 (dq, J=13.1, 6.7, 6.2 Hz, 1H), 2.14-1.95 (m, 3H), 1.90 (qt, J=6.9, 2.9 Hz, 1H), 1.82-1.51 (m, 7H), 1.46 (dt, J=9.0, 7.0 Hz, 1H), 0.73-0.56 (m, 1H), 0.46 (dddd, J=17.8, 7.5, 5.7, 4.3 Hz, 3H), 0.28 (dtt, J=11.5, 6.2, 2.7 Hz, 1H).

Sodium Salt of Compound E

[0581] A MeOH solution of benzyl (S)-2-cyclopentyl-2-((S)-7-((2R,3R)-3-cyclopropyl-1-methylaziridine-2-carbonyl)-2,7-diazaspiro[4.4]nonan-2-yl)acetate (1.0 equiv) and TEA (3.0 equiv) were added into a reactor. The mixture was cooled to 05 C. and PdCl.sub.2 (1% w/w) was added to the reaction. The reaction mixture was stirred for at least 3 h at 05 C. under pressure of 0-0.15 MPa with H.sub.2. A sample was taken for HPLC analysis. The reaction mixture was filtered via diatomite and washed the filter cake with MeOH (7 V). The filtrate was concentrated to 3-4 V under reduced pressure (20-30 C., 0.08 Mpa). MeOH (7 V) was added to the residue and the solution was stirred for 10 min at rt. NaOMe (1.03 equiv, 30 wt % MeOH solution) was added to the solution at rt and the resulting mixture was stirred for at least 3 h at rt. The reaction solution was concentrated to 3-4 V under reduced pressure (20-30 C., 0.08 Mpa) to afford solution A. In another reactor was charged MTBE (60 V) and the reactor was cooled to 5-5 C. The seeds of Compound E Na salt (2 wt %) was added to MTBE solution and the suspension was stirred for 30 min. Solution A of Compound E was added to MTBE over 5 h at 5-5 C. and the suspension was stirred for 2 h. The slurry was filtered, and the wet cake was washed with MTBE (2 V). The wet cake was dried at 30-40 C. for 5 h under vacuum to afford Compound E Na salt. Table 18 summarizes the HPLC method for this synthesis.

[0582] LCMS (ESI+): Calculated for C.sub.21H.sub.33N.sub.3O.sub.3 [M+H].sup.+: 376.26; found: 376.28.

[0583] .sup.1H NMR (400 MHz, Methanol-d.sub.4) 3.70-3.65 (m, 1H), 3.57-3.35 (m, 3H), 3.01-2.79 (m, 4H), 2.39 (s, 3H), 2.34 (t, J=7.2 Hz, 1H), 2.29-2.16 (m, 1H), 2.10-1.70 (m, 6H), 1.70-1.47 (m, 5H), 1.42 (p, J=7.2 Hz, 2H), 0.69-0.21 (m, 5H).

TABLE-US-00018 TABLE 18 HPLC Method for Parts 1 and 2 IPC method: Column: XBridge C18 (4.6 150 mm, 3.5 m) Mobile Phase: A: 10 mM NH.sub.4OAc in water B: MeOH Time (min) A % B % Gradient: 0.0 90 10 13.0 5 95 16.0 5 95 17.0 90 10 Flow Rate: 1.0 mL/min UV Detector 210 nm Wavelength: Column Temperature: 40 C. Retention Times: Compound E: 6.46 min benzyl (S)-2-cyclopentyl-2-((S)-7-((2R,3R)-3- cyclopropyl-1-methylaziridine-2-carbonyl)-2,7- diazaspiro[4.4]nonan-2-yl)acetate: 14.05 min

Example 13Alternative Synthetic Route for benzyl (S)-2-cyclopentyl-2-((S)-7-((2R,3R)-3-cyclopropyl-1-methylaziridine-2-carbonyl)-2,7-diazaspiro[4.4]nonan-2-yl)acetate

##STR00274##

[0584] To a reactor were charged MTBE (90.50 mL, 5 V), Compound D (18.10 g, 52.85 mmol, 1.0 equiv), Compound A (12.84 g, 55.49 mmol, 1.05 equiv) and 1,5,7-triazobicyclo[4,4,0]dec-5-ene(TBD) (2.21 g, 15.86 mmol, 0.3 equiv). The reaction mixture was cooled to 5-0 C. The reaction mixture was stirred at 5-0 C. for 24 hours at which point HPLC analysis showed reaction completion. Aq. Na.sub.2CO.sub.3 solution (5 wt %, 181.00 mL, 10 V) was added to quench the reaction and the mixture was stirred at 15-25 C. for at least 10 min. The phases were separated, and the organic phase was washed with aq. Na.sub.2CO.sub.3 solution (5 wt %, 181.00 mL, 10 V). The organic phase was concentrated to dryness (T s 40 C.). The residue was dissolved in EtOAc (145.00 mL, 8 V) and the solution was cooled to 10-0 C. To the solution was added maleic acid (3.07 g, 26.42 mmol, 0.5 equiv), followed by adding seeds of the maleate salt of benzyl (S)-2-cyclopentyl-2-((S)-7-((2R,3R)-3-cyclopropyl-1-methylaziridine-2-carbonyl)-2,7-diazaspiro[4.4]nonan-2-yl)acetate (181.00 mg, 1 wt %) at 10-0 C. The mixture was stirred for at least 10 min before the solution of maleic acid in MTBE/EtOAc (v/v=3:2, 181.00 mL, 10 V) was added dropwise over 3 h. The suspension was stirred for 12 h at 10-0 C. The slurry was filtered and the cake was washed with pre-cooled MTBE/EtOAc (v/v=1:2, 54.30 mL, 3 V). The filter cake was transferred to a reactor where MTBE (181.00 mL, 10 V) was cooled to 5-0 C. Aq. Na.sub.2CO.sub.3 solution (15 wt %, 181.00 mL, 10 V) was added to the solution to adjust the pH=8-10. The mixture was stirred for at least 10 min at 15-25 C. The phases were separated, and the aqueous phase was extracted with MTBE (90.50 mL, 5 V). The combined organic phase was washed with aq. Na.sub.2CO.sub.3 solution (5 wt %, 181.00 mL, 10 V). The organic phase was concentrated to dryness to afford the maleate salt of benzyl (S)-2-cyclopentyl-2-((S)-7-((2R,3R)-3-cyclopropyl-1-methylaziridine-2-carbonyl)-2,7-diazaspiro[4.4]nonan-2-yl)acetate (19.3 g, 97.2% LCAP, 92.7% QNMR assay).

Besylate salt of benzyl (S)-2-cyclopentyl-2-((S)-7-((2R,3R)-3-cyclopropyl-1-methylaziridine-2-carbonyl)-2,7-diazaspiro[4.4]nonan-2-yl)acetate

[0585] A reactor was charged with MTBE (5 V), Compound D (1.0 equiv), Compound A (1.05 equiv), and 1,5,7-triazobicyclo[4,4,0]dec-5-ene(TBD) (0.3 equiv). The reaction mixture was cooled to 5-0 C. The reaction mixture was stirred at 5-0 C. for 24 hours at which point HPLC analysis showed reaction completion. Aq. Na.sub.2CO.sub.3 solution (5 wt %, 10 V) was added to quench the reaction and the mixture was stirred at 15-25 C. for at least 10 min. The phases were separated, and the organic phase was washed with aq. Na.sub.2CO.sub.3 solution (5 wt %, 10 V). The organic phase was concentrated to dryness (T s 40 C.). Benzenesulfonic acid (2.1 equiv) was dissolved in THE (4.53 V) and set aside for later use. The concentrated benzyl (S)-2-cyclopentyl-2-((S)-7-((2R,3R)-3-cyclopropyl-1-methylaziridine-2-carbonyl)-2,7-diazaspiro[4.4]nonan-2-yl)acetate residue was diluted with acetone (10 V). The solution was cooled to 73 C. The premade benzenesulfonic acid solution in THE (3.8/21) was added portion-wise to benzyl (S)-2-cyclopentyl-2-((S)-7-((2R,3R)-3-cyclopropyl-1-methylaziridine-2-carbonyl)-2,7-diazaspiro[4.4]nonan-2-yl)acetate and a seed of the besylate salt of benzyl (S)-2-cyclopentyl-2-((S)-7-((2R,3R)-3-cyclopropyl-1-methylaziridine-2-carbonyl)-2,7-diazaspiro[4.4]nonan-2-yl)acetate (0.5% w/w) was then added to the mixture. The mixture was stirred at 73 C. for 1 h before another portion of benzenesulfonic acid THE solution (15.2/21) was added. The resulting mixture was stirred at 73 C. for 16 h. Then the rest of benzenesulfonic acid THE solution (0.5/21) was added to the mixture and continued to stir for 6 h. The slurry was filtered and the wet cake was washed with THE/acetone (v/v=1:2, 3 V). The wet cake was added in MTBE (10 V) and aq. Na.sub.2CO.sub.3 solution (15% w/w) was added to adjust pH to 7.5-9.0 at 05 C. To the mixture was added H.sub.2O (5 V) and the biphasic solution was stirred for 0.5 h before the organic phase was separated. The aqueous phase was washed with MTBE (5 V). The organic phases were combined and washed with aq. Na.sub.2CO.sub.3 solution (5% w/w, 10 V). The organic phase was concentrated under vacuum to 4-5 V at NMT 40 C. The concentrated residue was diluted with acetone (10 V) and the resulting mixture was concentrated under vacuum to 4-5 V at NMT 40 C. The solvent wash with acetone was repeated once. The solution of benzyl (S)-2-cyclopentyl-2-((S)-7-((2R,3R)-3-cyclopropyl-1-methylaziridine-2-carbonyl)-2,7-diazaspiro[4.4]nonan-2-yl)acetate in acetone was used for next step. Table 19 summarizes the HPLC method for this synthesis.

[0586] LCMS (ESI+): Calculated for C.sub.28H.sub.39N.sub.3O.sub.3 [M+H].sup.+: 466.3; found: 466.6.

[0587] .sup.1H NMR (400 MHz, Methanol-d.sub.4) 7.49-7.30 (m, 5H), 6.27 (s, 4H), 5.41-5.20 (m, 2H), 3.87-3.69 (m, 2H), 3.69-3.32 (m, 7H), 2.83 (d, J=2.4 Hz, 3H), 2.55 (dt, J=12.0, 8.2 Hz, 1H), 2.45 (t, J=7.5 Hz, 1H), 2.27-1.96 (m, 4H), 1.96-1.84 (m, 1H), 1.77-1.63 (m, 1H), 1.53 (d, J=4.8 Hz, 5H), 1.37-1.23 (m, 1H), 0.85-0.61 (m, 4H), 0.53 (tq, J=6.3, 4.3 Hz, 1H).

TABLE-US-00019 TABLE 19 HPLC Method for besylate salt formation IPC method: Column: XBridge C18 (4.6 150 mm, 3.5 m) Mobile Phase: A: 0.05% TFA in water (v/v) B: 0.05% TFA in MeCN (v/v) Time (min) A % B % Gradient: 0.0 90 10 12.0 45 55 15.0 5 95 17.0 5 95 18.0 90 10 Flow Rate: 0.8 mL/min UV Detector 210 nm Wavelength: Column Temperature: 40 C. Retention Times: Compound D: 9.35 min benzyl (S)-2-cyclopentyl-2-((S)-7-((2R,3R)-3- cyclopropyl-1-methylaziridine-2-carbonyl)-2,7- diazaspiro[4.4]nonan-2-yl)acetate: 10.04 min

Example 14Synthesis of (S)-3-(5-bromo-2-(5-(4-cyclopropylpiperazin-1-yl)-2-(1-methoxyethyl)pyridin-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indol-3-yl)-2,2-dimethyipropan-1-ol (Compound H)

##STR00275##

Part 1Synthesis of (S)-5-(2-(1-methoxyethyl)pyridin-3-yl)-2,2-dimethyl-5-oxopentanoic acid

##STR00276##

[0588] Reactor A was charged with THE (230.5 kg, 2 V) and i-PrMgCl.Math.LiCl (465.0 kg, 1.3 mol/L, 1.03 equiv) under nitrogen protection with stirring. The reactor was cooled to 205 C. A solution of 3-bromo-2-[(1S)-1-methoxyethyl]pyridine (129.7 kg, 600.2 mol, 1.0 equiv) in THE (350.4 kg, 3 V) was added to the reactor dropwise (21 kg/min) while maintaining the inner temperature at 205 C. The reaction mixture was stirred for 2 h at 205 C. and sampled for IPC. Reactor B was charged with THE (577.4 kg, 5 V) and 2,2-dimethylglutaric anhydride (89.2 kg, 627.5 mol, 1.05 equiv) with stirring at 105 C. The solution of reactor A was charged to reactor B dropwise at 105 C. (3.9 kg/min). The reaction mixture was sampled for HPLC analysis. A solution of 2,2-dimethylglutaric anhydride (0.1 equiv) in THE (0.3 V) was added to the reaction mixture and stirred for additional 5 h at 105 C. The reaction mixture was sampled for HPLC analysis. The reaction mixture was quenched by adding H.sub.2O (3 V) at 55 C. The mixture temperature was adjusted to 205 C. and stirred for at least 0.5 h. n-heptane (3 V) and 4% NaOH aq. solution (3 V) were added to the organic phase. After stirring for 0.5 h, the aqueous phase was separated. The combined aqueous layer was adjusted to pH=7.50.3 with KHSO.sub.4 at 205 C. The aqueous phase was washed with n-heptane (5 V1, 2 V1). The aqueous phase was filtered and the filtrate was adjusted to pH=5.7-7.0 with KHSO.sub.4. The crystal seed of (S)-5-(2-(1-methoxyethyl)pyridin-3-yl)-2,2-dimethyl-5-oxopentanoic acid (14.76% w/w) was added to the solution. The solution was acidified to pH=5.50.2 with KHSO.sub.4. The slurry was stirred for at least 5 h at 15-25 C. and filtered. The filter cake of the product was obtained as light yellow solids.

[0589] LCMS (ESI+): Calculated for C.sub.15H.sub.22NO.sub.4 (M+H): 280.1; Found: 280.1

[0590] .sup.1H NMR (400 MHz, DMSO-d.sub.6, 25 C.) 12.2 (s, 1H), 8.59 (dd, J=4.8, 5.2 Hz, 1H), 7.89 (dd, J=7.6, 8.0 Hz, 1H), 7.40-7.37 (m, 1H), 4.58-4.53 (m, 1H), 3.11 (s, 3H), 2.81-2.77 (m, 2H), 1.82-1.78 (m, 2H), 1.42 (d, J=6.4 Hz, 3H), 1.12 (s, 6H).

Part 2Synthesis of (S)-3-(5-bromo-2-(2-(1-methoxyethyl)pyridin-3-yl)-1H-indol-3-yl)-2,2-dimethylpropanoic acid

##STR00277##

[0591] To a reactor was charged H.sub.2O (522 L, 2.5 V). H.sub.2SO.sub.4 (150.7 kg, 1537 mol, 2.0 equiv) was added dropwise to the reactor at 2020 C. (S)-5-(2-(1-methoxyethyl)pyridin-3-yl)-2,2-dimethyl-5-oxopentanoic acid (215.6 kg, 771.8 mol, 1.0 equiv) and 4-bromophenylhydrazine hydrochloride (189.5 kg, 847.9 mol, 1.1 equiv) were added to the reactor at 2010 C. The reaction mixture was heated to 70-75 C. and stirred for at least 0.5 h. The reaction mixture was then heated at 955 C. for 32 h. The mixture was sampled for HPLC analysis. After completion, the reaction was cooled to 60-65 C. Water (1185.8 kg, 5.5 V) and 30% NaOH aq. solution (189.3 kg, 1.9 equiv) were added, and the mixture was stirred at 60-65 C. for 5 h. The mixture was cooled to 255 C. and stirred for 2 h. The mixture was filtered and washed with H.sub.2O (800 L, 3 V). The wet cake was added to H.sub.2O (1022.5 kg) and H.sub.2SO.sub.4 (53.9 kg) at 255 C. The mixture was warmed to 80-85 C. and stirred for 5 h. The mixture was then cooled to 25-30 C. and stirred for 16 h. The slurry was filtered and washed with H.sub.2O (1020 L, 3 V). The filter cake was dried under 505 C. for 36 h to afford the product as yellow solids.

[0592] LCMS (ESI+): Calculated for C.sub.21H.sub.24BrN.sub.2O.sub.3 (M+H): 433.1; Found: 433.1

[0593] .sup.1H NMR (400 MHz, DMSO-d.sub.6, 25 C.) 12.1 (s, 1H), 11.4 (s, 1H), 8.70 (s, 1H), 7.82-7.77 (m, 2H), 7.45-7.20 (m, 3H), 4.19 (d, J=5.6 Hz 1H), 2.93 (s, 4H), 2.78 (s, 1H), 1.36 (d, J=4.0 Hz, 3H), 0.88 (d, J=5.2 Hz, 6H).

Part 3methyl (S)-3-(5-bromo-2-(2-(1-methoxyethyl)pyridin-3-yl)-1H-indol-3-yl)-2,2-dimethylpropanoate

##STR00278##

[0594] To a reactor were charged MeOH (5.45 L, 5 V) and (S)-3-(5-bromo-2-(2-(1-methoxyethyl)pyridin-3-yl)-1H-indol-3-yl)-2,2-dimethylpropanoic acid.Math.H.sub.2SO.sub.4 (1.09 kg, 2.27 mol, 1.0 equiv). SOCl.sub.2 (331 g, 2.78 mol, 1.2 equiv) was then added at 50-60 C. The reaction mixture was stirred at 65 C. for 12 h. After completion, the reaction mixture was cooled to 10 C. The mixture was adjusted to pH=7-8 with 5% Na.sub.2CO.sub.3 at 10-15 C. The mixture was filtered and the filter cake was washed with H.sub.2O (2.2 L, 2 V). The wet cake was triturated with H.sub.2O (5.4 L, 5 V) at 25 C. for 6 h. The mixture was filtered and the filter cake was washed with H.sub.2O (2.2 L, 2 V). The filter cake was dried in blast air oven (N.sub.2, 45-50 C.) for 24 h to afford the product.

[0595] LCMS (ESI+): Calculated for C.sub.22H.sub.26BrN.sub.2O.sub.3 (M+H+): 447.1; Found: 446.9

[0596] 1H NMR (300 MHz, DMSO-d6) 11.41 (s, 1H), 8.72 (dd, J=4.7, 1.8 Hz, 1H), 7.81 (dd, J=7.7, 1.8 Hz, 1H), 7.65 (d, J=1.9 Hz, 1H), 7.47 (dd, J=7.8, 4.7 Hz, 1H), 7.32 (d, J=8.6 Hz, 1H), 7.22 (dd, J=8.6, 1.9 Hz, 1H), 4.23 (q, J=6.3 Hz, 1H), 3.32 (s, 9H), 2.95 (s, 4H), 1.39 (d, J=6.3 Hz, 3H), 0.94 (s, 6H).

Part 4Synthesis of methyl (S)-3-(5-bromo-2-(2-(1-methoxyethyl)pyridin-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indol-3-yl)-2,2-dimethylpropanoate

##STR00279##

[0597] To a reactor were charged DMF (1163.1 kg, 5 V), methyl (S)-3-(5-bromo-2-(2-(1-methoxyethyl)pyridin-3-yl)-1H-indol-3-yl)-2,2-dimethylpropanoate (239.04 kg, 536.7 mol, 1.0 equiv), and Cs.sub.2CO.sub.3 (245.94, 754.8 mol, 1.4 equiv). The reactor was rinsed with DMF (454.6 kg, 2 V). The reaction mixture was cooled to 15-20 C. and 2,2,2-trifluoroethyl trifluoromethanesulfonate (170 kg, 732.4 mol, 1.35 equiv) was added. The reaction mixture was stirred at 15-20 C. for 16 h. The reaction mixture was sampled for HPLC analysis. After completion the reaction was quenched with AcOH (64.4 kg, 2.0 equiv) and H.sub.2O (1435.0 kg, 6 V) at 15-20 C. The resulting mixture was extracted with MTBE two times (5 V, 2 V). The combined organic layers were concentrated to 3-4 V at 35-45 C. The concentrated mixture was diluted with EtOH (4 V) and further concentrated to about 3-4 V under reduced pressure at 35-45 C. The previous unit operation was repeated until the MTBE content in EtOH solution was below 0.5% w/w. The resulting EtOH solution of methyl (S)-3-(5-bromo-2-(2-(1-methoxyethyl)pyridin-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indol-3-yl)-2,2-dimethylpropanoate, as a 1.4:1 mixture of diastereomers, was used directly in the next step.

[0598] LCMS (ESI+): Calculated for C.sub.24H.sub.27BrF.sub.3N.sub.2O.sub.3 (M+H+): 529.1; Found: 529.9

[0599] 1H NMR (300 MHz, DMSO-d.sub.6) 8.77 (dt, J=4.8, 1.4 Hz, 1H), 7.86-7.75 (m, 1H), 7.73-7.62 (m, 2H), 7.55 (ddd, J=7.8, 6.0, 4.7 Hz, 1H), 7.40 (ddd, J=8.8, 4.0, 1.9 Hz, 1H), 5.37 (dd, J=16.5, 8.6 Hz, 1H), 4.82 (dd, J=17.9, 9.0 Hz, 1H), 4.46 (dd, J=16.4, 9.5 Hz, OH), 3.93 (dt, J=18.4, 6.2 Hz, 1H), 3.66-3.56 (m, 2H), 3.46 (d, J=13.2 Hz, 3H), 3.06 (s, 1H), 2.94 (s, 2H), 1.84-1.70 (m, 2H), 1.32 (dd, J=37.8, 6.2 Hz, 3H), 1.05-0.87 (m, 7H).

Part 5Synthesis of Mixture of Atropisomers of Compound G(S)-3-(5-bromo-2-(2-(1-methoxyethyl)pyridin-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indol-3-yl)-2,2-dimethylpropan-1-ol

##STR00280##

[0600] To a reactor was charged the EtOH solution of methyl (S)-3-(5-bromo-2-(2-(1-methoxyethyl)pyridin-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indol-3-yl)-2,2-dimethylpropanoate (1.0 equiv, 5 V) at 15-20 C. under N.sub.2. To the mixture was charged CaCl.sub.2) (60.04 kg, 541.0 mol, 1.0 equiv) at 20-30 C. batchwise, followed by addition of NaBH.sub.4 (51.04 kg, 1349.2 mol, 2.5 equiv) at 20-30 C. The reaction mixture was agitated at 20-30 C. under for 14 h. The reaction mixture was sampled for HPLC analysis. The reaction was quenched by dropwise addition of aq. HCl (3 M) to the point of pH=1-2. Aq. NaOH (30% w/w) was added to the mixture to adjust the pH=4-4.5. The resulting mixture was concentrated 3-5 V. The concentrated mixture was further diluted with MTBE (891.2 kg, 5 V) and H.sub.2O (1197.5 kg, 5 V). The organic phase was separated. The organic layer was washed with H.sub.2O (718.5 kg, 3 V), aq. NaOH (484.0 kg, 2 V) and H.sub.2O (718.5 kg, 3 V) sequentially. The organic layer was concentrated to 3-5 V at 45-60 C. The concentrated mixture was diluted with IPA (756.3 kg, 4 V) and further concentrated to about 3-4 V under reduced pressure at 45-60 C. The previous unit operation was repeated until the MTBE content in IPA solution was below 2% w/w. The resulting IPA solution of Compound G mixture, as a 1.46:1 mixture of diastereomers (247.6 kg, 98.8% a/a, 92% isolated yield) was purified by column purification or by chemical resolution.

Part 6Purification of Compound G Mixture

##STR00281##

Condition 1: Column Chromatography

[0601] The mixture of Compound G (19.0 kg) was treated with SiO.sub.2 (30.0 kg). The silica gel was loaded on column chromatography (240 kg SiO.sub.2, n-heptane/EtOAc=1/0 to 0/1) to obtain the pure fractions containing Compound G (1000 L). The fractions were concentrated to 25 L. To the concentrated residue was added n-heptane (100 L) and the resulting mixture was concentrated to 25 L. The n-heptane wash was repeated once. The residue was diluted with n-heptane (100 L) and the mixture was stirred at 15-20 C. for 12 h. The slurry was filtered and the wet cake was dried in the oven under vacuum to obtain Compound G (18.0 kg, 98.9% a/a) as white solids.

Condition 2: Resolution by CSA

[0602] The IPA solution of the Compound G mixture was added to a reactor, to which was charged IPA (1314.0 kg, 1.2 V). The IPA solution of Compound G mixture (assay weight 247.6 kg, 495.8 mol, 1.0 equiv) was then pumped through the continuous reactor& quenching reactor (the continuous reactor: 22010 C., quenching reactor: 0-20 C., back pressure value: 4.000.20 MPa) with flow rate of pump 80.0 L/h. The equilibrated IPA solution of Compound G was charged to reactor while remaining agitated at 2010 C. The mixture was sampled for HPLC analysis (criterion: diastereomers ratio1.77). D-camphor-10-sulfonic acid (CSA, 150.0 kg, 645.7 mol, 1.3 equiv) was added to the reaction mixture followed by adding IPA (21.5 kg, 0.1 V) to rinse the charging port. The reaction mixture was stirred at 255 C. for 2 h and then stirred at 533 C. for 2 h. Compound G CSA seed (0.1% w/w) was charged to the reaction mixture at 433 C. and then stirred at 433 C. for 2 h. The solution was further cooled to 63 C. and stirred at the temperature for 4 h. The resulting suspension was filtered and washed with IPA (212.0 kg, 1.0 V). To further control the atropisomer impurity of Compound G, the wet cake is re-suspended in IPA (10 V) and agitated at 67-70 C. for 3-4 h. The mixture was cooled to 5 C. and stirred at 5 C. for 2 h. The resulting wet cake (assay weight 200.0 kg, 1.0 equiv) was suspended in MTBE (3.5 V) at 205 C., followed by adding H.sub.2O (4 V). The pH was adjusted to pH=8-10 by slowly charging aq. NaOH (30% w/w). The mixture was agitated at 205 C. for 0.5 h. The organic phase was separated and washed with H.sub.2O (33 V). The organic layer was concentrated to 1-2 V. The concentrated mixture was diluted with n-heptane (3 V) and further concentrated to about 2-3 V under reduced pressure. The solvent swap with n-heptane was repeated two times. The concentrated mixture was diluted with n-heptane (3 V) and the mixture was agitated at 555 C. for 3 h. The mixture was cooled to 205 C. and stirred for 1 h at 205 C. The slurry was filtered and washed with n-heptane (1.2 V). The wet cake was dried at 45-55 C. to afford the product as off-white solids (143.6 kg, 99.05% a/a, 58% yield). To further control the atropisomer impurity of Compound G, the solid was re-suspended in THF/n-heptane (v/v=1:4, 5 V) and agitated at 67-70 C. for 3-4 h. The mixture was cooled to 5 C. and stirred at 5 C. for 2 h. The slurry was filtered and washed with n-heptane (1.5V). The wet cake was dried at 45-55 C. to afford Compound G as off-white solids. To recover Compound G, the mother liquor from resolution process was charged into a reactor. The solution was concentrated to 4-5 V under vacuum at NMT 50 C. 30% w/w aq. NaOH solution was added to the concentrated residue to adjust the pH to 8-10 at 15-30 C. The mixture was concentrated to 0.8-1.2 V under vacuum at NMT 50 C. MTBE (5 V) and H.sub.2O (5 V) were added to the residue. The biphasic system was stirred at 255 C. for 0.5 h. The organic phase was separated and washed with H.sub.2O (5 V). The organic phase was then concentrated to 1.2-1.6 V at NMT 50 C. The concentrated residue was diluted with IPA (1.6 V) and the mixture was then concentrated to 1.2-1.6 V at NMT 50 C. The solvent swap with IPA was repeated once and IPA (3 V) was added to the concentrated residue. The IPA solution of the Compound G mixture was subjected to the equilibrium process and CSA salt formation again. The IPA solution of the Compound G mixture was pumped through the continuous reactor and quenching reactor (the continuous reactor: 22010 C.; quenching reactor: 0-20 C.; back pressure: 4.000.20 MPa) with flow rate of pump 80.0 L/h. The equilibrated IPA solution of Compound G was charged to reactor while remaining agitated at 2010 C. The mixture was sampled for HPLC analysis (criterion: diastereomers ratio1.77). D-camphor-10-sulfonic acid (1.3 equiv) was added to the reaction mixture followed by adding IPA (0.1 V) to rinse the charging port. The reaction mixture was stirred at 255 C. for 2 h and then stirred at 533 C. for 2 h. A CSA seed of Compound G (0.1% w/w) was charged to the reaction mixture at 433 C. and then stirred at 433 C. for 2 h. The solution was further cooled to 63 C. and stirred at the temperature for 4 h. The resulting suspension was filtered and washed with IPA (1.0 V). The resulting wet cake (1.0 equiv) was suspended in MTBE (3.5 V) at 205 C., followed by adding H.sub.2O (4 V). The pH was adjusted to pH=8-10 by slowly charging aq. NaOH (30% w/w). The mixture was agitated at 205 C. for 0.5 h. The organic phase was separated and washed with H.sub.2O (33 V). The organic layer was concentrated to 1-2 V. The concentrated mixture was diluted with n-heptane (3 V) and further concentrated to about 2-3 V under reduced pressure. The solvent swap with n-heptane was repeated two times. The concentrated mixture was diluted with n-heptane (3 V) and the mixture was agitated at 555 C. for 3 h. The mixture was cooled to 205 C. and stirred for 1 h at 205 C. The slurry was filtered and washed with n-heptane (1.2 V). The wet cake was dried at 45-80 C. to afford atropisomerically pure Compound G as off-white solids.

Condition 3: Resolution by Methanesulfonic Acid (MsOH)

[0603] To the toluene solution of the Compound G mixture was charged xylene (3 V). The mixture was concentrated to 2.5-3.5 V. The residue was agitated at 1375 C. for 24 h. The mixture was cooled to 60-70 C. and a sample was taken for HPLC analysis. The residue was diluted with 2-MeTHF (5 V) at 205 C. The solution was washed with aq. HCl (0.2 M, 3 V) twice. The organic phase was separated and extracted with aq. HCl (3 M, 3 V) twice. The combined aqueous phase was washed with MTBE (3 V). The aqueous phase was adjusted to pH=8-10 with 30% aq. NaOH. The aqueous phase was extracted with IPAc (5 V). The organic phase was extracted and washed with H.sub.2O (3 V). The organic phase was concentrated to 2-3 V under reduced pressure. The solvent swap with IPAc was repeated twice. The concentrated residue was diluted with IPAc (5 V). To the IPAc solution was added methanesulfonic acid (MSA, 0.35 equiv) at 2010 C. The mixture was stirred at 305 C. for 4 h. The slurry was filtered and the cake was washed with IPAc (2V). The cake containing the undesired atropisomer of Compound G as a mesylate salt is collected and subject to recovery. The filtrate was added 5% aq. NaHCO.sub.3 (3 V) and the pH of aqueous phase should be 8. To the biphasic system was added H.sub.2O (3 V). The organic phase was separated and concentrated to 1.8-2.3 V under vacuum at NMT 45 C. The concentrated residue was diluted with MeOH (7 V). The resulting solution was concentrated to 1.8-2.3 V under vacuum at NMT 45 C. To the MeOH solution was added MSA (1.0 equiv) at 2010 C. The mixture was stirred at 40 C. for 0.5 h until all solids are dissolved. To the solution was added H.sub.2O (2.4 V) over 2 h followed by adding Compound G seed (10% w/w). The mixture was stirred at 40 C. for 4 h before H.sub.2O (1.8 V) was added dropwise. Another portion of H.sub.2O (6 V) was added dropwise and the mixture was stirred for 1 h. The mixture was cooled to 20 C. and the slurry was stirred for 4 h. The slurry was filtered and the wet cake was washed with MeOH/H.sub.2O (v/v=1:3.5, 1.8 V). The wet cake was dried in the oven at 50-55 C. for 16 h to afford Compound G as white solids. To recover Compound G, the filtrate was concentrated under vacuum. The residue was combined with the wet cake of the undesired atropisomer of Compound G as the mesylate salt. Aq. NaOH (30% w/w) was added to adjust the pH to 8-10. MTBE (5 V) was added to the reaction mixture. The mixture was stirred for 1 h at 205 C. The organic phase was separated and concentrated to 1.5-3 V. The concentrated residue was diluted with MTBE (5 V). The resulting mixture was concentrated to 1.5-3 V. The solvent swap with MTBE was repeated twice. The Compound G MTBE solution was concentrated to 1.5-2.5 V and to the residue was added xylene (3 V). The mixture was stirred at 138-143 C. for 24 h before it was cooled to 60 C. To the mixture was added IPA (6.3 V) and the resulting solution was concentrated to 2-3 V under vacuum. The solvent swap with IPA was repeated twice. To the IPA solution was added D-camphor-10-sulfonic acid (1.3 equiv) at 155 C. The mixture was cooled to 5-0 C. and stirred for 14 h at 5-0 C. The slurry was filtered and washed with IPA (2 V). The wet cake was dried at 45-80 C. to afford Compound G as off-white solids.

[0604] LCMS (ESI+): Calculated for C.sub.23H.sub.27BrF.sub.3N.sub.2O.sub.2 (M+H.sup.+): 499.1; Found: 499.1

[0605] 1H NMR (400 MHz, DMSO-d6) 8.75 (dd, J=4.8, 5.2 Hz, 1H), 7.98 (d, J=1.6 Hz, 1H), 7.78-7.76 (m, 1H), 7.66-7.63 (m, 1H), 7.54-7.53 (m, 1H), 7.40-7.38 (m, 1H), 5.39-5.30 (m, 1H), 4.60-4.56 (m, 1H), 4.51-4.42 (m, 1H), 3.98-3.93 (m, 1H), 3.33 (s, 2H), 3.03-2.94 (m, 3H), 2.60-2.50 (m, 1H), 2.29-2.26 (m, 1H), 1.36 (d, J=6.4 Hz, 2H), 0.60 (s, 6H).

Part 7aTwo-step synthesis of (S)-3-(5-bromo-2-(2-(1-methoxyethyl)-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridin-3-yl)-1-(2, 2, 2-trifluoroethyl)-1H-indol-3-yl)-2, 2-dimethylpropan-1-ol

##STR00282##

[0606] Compound G (200.0 kg, 400.5 mol, 1.0 equiv) was dissolved in THE (500.0 L, 2.5 V) and heptane (500.0 L, 2.5 V) at 205 c. HBPin (53.8 kg, 420.4 mol, 1.05 equiv) was added to the reaction mixture dropwise at 205 C. and the mixture was stirred at 30-35 for 2 h. The reaction mixture was sampled for .sup.1HNMR to indicate that Compound G was consumed completely. The reaction mixture was cooled to 20 C. B.sub.2Pin.sub.2 (119.0 kg, 468.6 mol, 1.2 equiv), dtbpy (2.1 kg, 8.01 mol, 0.02 equiv) and [Ir(OMe)(COD)]2 (1.3 kg, 2.0 mol, 0.005 equiv) were added to the reaction mixture sequentially. The resulting mixture was stirred at 30-35 for 12 h. A sample was taken for HPLC analysis. The reaction was quenched by charging H.sub.2O (0.1 V) and THE (0.5 V) at 0-5 C., followed by charging MTBE (4 V) and H.sub.2O (4 V). The mixture was filtered and the cake was washed with MTBE (3 V). The filtrate was separated and the organic layer was washed with H.sub.2O (3 V). The organic layer was concentrated to 1-2 V at 40-45 C. The concentrated mixture was diluted with DCM (4 V) and further concentrated to 1-2 V. The previous unit operation was repeated twice. The resulting DCM solution of (S)-3-(5-bromo-2-(2-(1-methoxyethyl)-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridin-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indol-3-yl)-2,2-dimethylpropan-1-ol was used directly for next step (479 kg, 86% assay yield, 43.1% w/w, 85.1% a/a).

Part 7bOne-Step Synthesis of (S)-3-(5-bromo-2-(2-(1-methoxyethyl)-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridin-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indol-3-yl)-2,2-dimethylpropan-1-ol

##STR00283##

[0607] To a reactor was charged n-heptane (4 V), dtbpy (1.6 mol %) at 25 C., followed by adding Compound G (1.0 equiv) and B.sub.2Pin.sub.2 (2.0 equiv). N.sub.2 was bubbled under surface of reaction mixture for 1 h. [Ir(OMe)(COD)]2 (0.8 mol %) was added to the suspension under the atmosphere of N.sub.2. The reaction mixture was stirred under N.sub.2 gas until all the solids were dissolved. The reaction mixture was then stirred at 55 for 2 h. After completion, the reaction solution was washed with H.sub.2O (3 V) twice. The organic phase was concentrated to dryness and used for next step.

[0608] LCMS (ESI+): Calculated for C.sub.23H.sub.27BBrF.sub.3N.sub.2O.sub.4 (M-2,3-dimethylbutane): 544.1; Found: 544.0

[0609] 1H NMR (300 MHz, chloroform-d) 9.11 (d, J=1.7 Hz, 1H), 8.10 (t, J=1.5 Hz, 1H), 7.92 (d, J=1.9 Hz, 1H), 7.40 (dd, J=8.7, 1.9 Hz, 1H), 7.29 (s, 1H), 4.73 (s, 1H), 4.47 (q, J=8.6 Hz, 2H), 4.03 (q, J=6.2 Hz, 1H), 3.82-3.72 (m, 1H), 3.58-3.45 (m, 2H), 3.04 (s, 3H), 2.74 (d, J=14.1 Hz, 1H), 2.23 (d, J=14.1 Hz, 1H), 1.91-1.83 (m, 1H), 1.46 (d, J=6.2 Hz, 4H), 1.36 (s, 12H), 1.28 (d, J=5.2 Hz, 35H), 0.95-0.85 (m, 5H), 0.76 (d, J=10.4 Hz, 6H).

Part 8Synthesis of Compound H(S)-3-(5-bromo-2-(5-(4-cyclopropylpiperazin-1-yl)-2-(1-methoxyethyl)pyridin-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indol-3-yl)-2,2-dimethylpropan-1-ol

##STR00284##

Condition 1: TMP as Base

[0610] A reactor was charged with DCM (1173 L, 30 V), 2,2,6,6-tetramethylpiperidine (35.3 kg, 250.0 mol, 4.0 equiv), 1-cyclopropylpiperazine (19.7 kg, 156.1 mol, 2.5 equiv), and Cu(OAc).sub.2 (14.2 kg, 78.2 mol, 1.25 equiv) at 15-20 C. with agitation. 21% 02 was bubbled under the surface for 1 h at 20-25 C. (S)-3-(5-bromo-2-(2-(1-methoxyethyl)-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridin-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indol-3-yl)-2,2-dimethylpropan-1-ol in DCM (39.1 kg assay weight, 62.5 mol, 1.0 equiv) was added to the reaction mixture at 10-20 C. The reaction mixture was heated at 25-35 C. during which 21% 02 was bubbled under surface and was stirred at this condition for 16 h. A sample was taken for HPLC analysis. The reaction was cooled to 20 C. The reaction mixture was quenched with a solution of NH.sub.40H (1.5 V) and H.sub.2O (3 V). The organic phase was separated and washed with a solution of NH.sub.40H (1.5 V) and H.sub.2O (3 V) for two additional times. The organic phase was washed with aq. EDTA (5% w/w, 3 V) and aq. NaCl (5% w/w, 4 V) sequentially. The organic phase was concentrated to about 1-2 V under reduced pressure at 40-45 C. The concentrated mixture was diluted by 2-MeTHF (6 V) and H.sub.2O (3 V). The pH of the system was adjusted to pH=1.1-1.3 by aq. HCl (2 N). The water layer was washed by 2-MeTHF (4 V) two times (Compound G contained in the 2-MeTHF solution can be recycled, the detailed procedure is described below). The water phase pH was adjusted to 8-9 by adding aq. NaOH (30% w/w) slowly. The water phase was concentrated under reduced pressure until no fraction to obtain the Compound H crude. Compound H crude was dissolved in MeCN (2 V) and was stirred at 40-50 C. for 3 h. To the solution was added H.sub.2O (1 V) at 35-45 C., followed by adding a seed of Compound H (0.5% w/w). The mixture was stirred at 35-45 C. for 2 h, to which was added H.sub.2O (2 V). The mixture was cooled to 15-25 C. The slurry was filtered and the cake was washed with MeCN/H.sub.2O (2 V, v/v=2:3). The wet cake was dried in blast air oven (N2, 45-50 C.) for 12 h to afford Compound H as brown solids (51.9% yield from Compound G, 98.8% a/a).

[0611] To recover Compound G, the 2-MeTHF solution containing Compound G is added to a reactor and was washed with aq. NaOH (0.1 N, 1 V) twice. The organic phase was separated, and H.sub.2O (1 V) was added. The aqueous phase was adjusted to pH=1-1.2 with aq. HCl (3N). The organic phase was separated and concentrated under vacuum at 45 C. until dryness. The residue was dissolved in IPA (1 V) and concentrated to 1-2 V. The concentrated residue was diluted with IPA (8 V). D-camphor-10-sulfonic acid (CSA, 1.2 equiv) in IPA (2 V) was added to the reaction mixture dropwise at 15-20 C. The reaction mixture was stirred at 15-20 C. for 2 h and then stirred at 50-55 C. for 2 h. Compound G CSA seed (0.1% w/w) was charged to the reaction mixture at 433 C. and then stirred at 433 C. for 2 h. The solution was further cooled to 63 C. and stirred at the temperature for 4 h. The resulting suspension was filtered and washed with IPA (1.0 V). The wet cake was suspended in IPA (3 V) and stirred at 40 C. for 60 min. The mixture was cooled to 5 C. and stirred at 5 C. for 60 min. The slurry was filtered and washed with IPA (1 V) to afford wet cake as Compound G CSA salt. The resulting wet cake (1.0 equiv) was suspended in MTBE (3.5 V) at 20+5 C., followed by adding H.sub.2O (4 V). The pH was adjusted to pH=8-10 by slowly charging aq. NaOH (30% w/w). The mixture was agitated at 205 C. for 0.5 h. The organic phase was separated and washed with H.sub.2O (33 V). The organic layer was concentrated to 1-2 V. The concentrated mixture was diluted with n-heptane (3 V) and further concentrated to about 2-3 V under reduced pressure. The solvent swap with n-heptane was repeated twice. The concentrated mixture was diluted with n-heptane (3 V) and the mixture was agitated at 555 C. for 3 h. The mixture was cooled to 205 C. and stirred for 1 h at 205 C. The slurry was filtered and washed with n-heptane (1.2 V). The wet cake was dried in Cone dryer at 45-80 C. to afford the Compound G as off-white solids.

Condition 2: TEA as Base

[0612] A reactor was charged with DCM (15 V), TEA (4.0 equiv), 1-cyclopropylpiperazine (2.5 equiv), and Cu(OAc).sub.2 (1.25 equiv) at 15-20 C. with agitation. 21% 02 was bubbled under surface for 1 h at 20-25 C. (S)-3-(5-bromo-2-(2-(1-methoxyethyl)-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridin-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indol-3-yl)-2,2-dimethylpropan-1-ol in DCM (1.0 equiv) was added to the reaction mixture at 10-20 C. The reaction mixture was heated at 25-35 C. during which 21% 02 was bubbled under surface and was stirred at this condition for at least 5 h. A sample was taken for HPLC analysis. The reaction was cooled to 20 C. The reaction mixture was quenched with aq. solution of NH.sub.40H (1.5 V) and H.sub.2O (3 V). The organic phase was separated and washed with N-acetyl cysteine (1 N, 3 V) and H.sub.2O (2 V) sequentially. The organic phase was concentrated to about 1-2 V under reduced pressure at 40-45 C. The concentrated mixture was diluted by 2-MeTHF (6 V) and H.sub.2O (3 V). The pH of the system was adjusted to pH=1.1-1.3 by aq. HCl (2 N). The water layer was washed by 2-MeTHF (4 V) for two times. The water phase pH was adjusted to 8-9 by adding aq. NaOH (2 N) slowly. The water phase was concentrated under reduced pressure until no fraction to obtain crude Compound H. The crude Compound H was dissolved in MeCN (2 V) and was stirred at 40-50 C. for 3 h. To the solution was added H.sub.2O (3 V) slowly at 35-45 C. The mixture was cooled to 15-25 C. and was further stirred for 3 h. The slurry was filtered and the cake was washed with MeCN/H.sub.2O (2 V, v/v=2:3). The wet cake was dried under vacuum to afford Compound H as brown solids.

[0613] LCMS (ESI+): Calculated for C.sub.30H.sub.39BrF.sub.3N.sub.4O.sub.2 (M+H): 623.2; Found: 622.9

[0614] 1H NMR (400 MHz, DMSO-d6) 8.46 (d, J=2.0 Hz, 1H), 7.95 (s, 1H), 7.62 (d, J=4.4 Hz, 1H), 7.36 (d, J=2.0 Hz, 1H), 7.18 (s, 1H), 5.32-5.26 (m, 1H), 4.61-4.47 (m, 2H), 3.85-3.83 (m, 1H), 3.19 (s, 4H), 3.03-3.02 (m, 2H), 2.91 (s, 3H), 2.68 (s, 4H), 2.60 (s, 1H), 2.30-2.27 (m, 1H), 1.66 (s, 1H), 1.32 (d, J=2.0 Hz, 3H), 0.62 (s, 6H), 0.45-0.35 (m, 4H).

Alternative Synthesis of the Mixture of Compound G Atropisomers(S)-3-(5-bromo-2-(2-(1-methoxyethyl)pyridin-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indol-3-yl)-2,2-dimethylpropan-1-ol

##STR00285##

Part 1Synthesis of 2,2,2-trifluoroethyl (S)-3-(5-bromo-2-(2-(1-methoxyethyl)pyridin-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indol-3-yl)-2,2-dimethylpropanoate

[0615] To a reactor were added DMF (770 L, 7 V) and (S)-3-(5-bromo-2-(2-(1-methoxyethyl)pyridin-3-yl)-1H-indol-3-yl)-2,2-dimethylpropanoic acid (110 kg, 255.0 mol, 1.0 equiv) at 10-20 C. The reaction mixture was cooled to 0-5 C., and Cs.sub.2CO.sub.3 (208.0 kg, 637.0 mol, 2.5 equiv) was added, followed by dropwise addition of 2,2,2-trifluoroethyl trifluoromethanesulfonate (148.0 kg, 637.0 mol, 2.5 equiv). The reaction mixture was stirred at 0-5 C. for 16 h. After completion, the reaction was quenched by adding H.sub.2O at 0-20 C. The mixture was extracted with MTBE (550 L, 5 V). The organic layer was separated and the aqueous layer was extracted with MTBE (550 L, 5 V). The combined organic layers were washed with brine (330 L, 3 V). The organic layer was concentrated to 190-380 L under reduced pressure at 40-50 C. The concentrated residue was diluted with THE (220 L, 2 V) and further concentrated to about 1-2V under reduced pressure. The previous unit operation was repeated until the MTBE content in THE solution was below 1% w/w. The resulting THE solution of 2,2,2-trifluoroethyl (S)-3-(5-bromo-2-(2-(1-methoxyethyl)pyridin-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indol-3-yl)-2,2-dimethylpropanoate was used directly for next step (123.6 kg assay weight, 97.5% yield).

[0616] LCMS (ESI+): Calculated for C.sub.25H.sub.26BrF.sub.6N.sub.2O.sub.3 (M+H): 597.1; Found: 597

[0617] 1H NMR (400 MHz, DMSO-d6) 8.76 (d, J=4.8 Hz, 1H), 7.83-7.77 (m, 2H), 7.68-7.64 (m, 1H), 7.57-7.52 (m, 1H), 7.43-7.39 (m, 1H), 4.50-4.43 (m, 2H), 3.59 (t, J=12 Hz, 1H), 3.04 (s, 1H), 2.92 (s, 2H), 2.88 (s, 2H), 2.73 (s, 2H), 1.76-7.73 (m, 1H), 1.37 (d, J=8.0 Hz, 1H), 1.23 (d, J=8.0 Hz, 1H), 0.98 (d, J=20 Hz, 6H).

Part 2Synthesis of the Mixture of Compound G Atropisomers(S)-3-(5-bromo-2-(2-(1-methoxyethyl)pyridin-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indol-3-yl)-2,2-dimethylpropan-1-ol

[0618] To a reactor was added THE solution of 2,2,2-trifluoroethyl (S)-3-(5-bromo-2-(2-(1-methoxyethyl)pyridin-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indol-3-yl)-2,2-dimethylpropanoate (123.6 kg assay weight, 207.0 mol, 1.0 equiv) at 15-20 C. The reaction mixture was warmed to 50-60 C. and LiBH.sub.4 (2M in THF, 155.7 L, 311.0 mol, 1.5 equiv) was added. The reaction mixture was stirred at 60-65 C. for 24 h. A sample was taken for HPLC analysis. After completion, the reaction was quenched with aq. HCl (0.5 M, 618.0 L, 5 V) at 10-20 C. Aq. HCl (0.5 M) was continued to add to adjust the pH to 4-5. MTBE (618 L, 5 V) was added to the mixture and stirred at 10-20 C. for 0.5 h. The organic layer was separated and the aqueous layer was washed with MTBE (370 L, 3 V). The combined organic layers were washed with aq. NaOH (247 L, 2 V) and brine (247 L, 2 V) sequentially. The organic layer was concentrated under reduced pressure at 40-45 C. to afford the mixture of Compound G atropisomers (93.5 kg, 90.1% a/a).

Alternative Synthesis of the Mixture of Compound G Atropisomers(S)-3-(5-bromo-2-(2-(1-m ethoxyethyl)pyridin-3-yl)-1-(2, 2, 2-trifluoroethyl)-1H-indol-3-yl)-2, 2-dim ethylpropan-1-ol

##STR00286##

[0619] To a reactor was charged the EtOH solution of methyl (S)-3-(5-bromo-2-(2-(1-methoxyethyl)pyridin-3-yl)-1H-indol-3-yl)-2,2-dimethylpropanoate (1.0 equiv, 5 V) at 15-20 C. under N.sub.2. To the mixture was charged CaCl.sub.2 (1.25 equiv) at 5 C. batchwise, followed by addition of NaBH.sub.4 (1.5 equiv) at 5 C. The reaction mixture was agitated at 20-30 C. under for 14 h. The reaction mixture was sampled for HPLC analysis. The reaction was quenched by dropwise addition of aq. HCl (3 M) until a pH of 1-2 was achieved. Aq. NaOH (30% w/w) was added to the mixture to adjust the pH between 4-5. The resulting mixture was concentrated and precipitated from water. The resulted cake was washed with water and dried at 45 C. under vacuum.

[0620] To a reactor were charged DMF (7 V), Compound K (1.0 equiv), and K.sub.3PO.sub.4 (1.4 equiv). The reactor was rinsed with DMF (454.6 kg, 7 V). The reaction mixture was cooled to 15-20 C. and 2,2,2-trifluoroethyl trifluoromethanesulfonate (2 equiv) was added. The reaction mixture was stirred at 15-20 C. for 16 h. The reaction mixture was sampled for HPLC analysis. After completion the reaction was quenched with AcOH (64.4 kg, 2.0 equiv) and H.sub.2O (1435.0 kg, 6 V) at 15-20 C. The resulting mixture was extracted with MTBE two times (5 V, 2 V). The combined organic layers were concentrated to 3-4 V at 35-45 C. The concentrated mixture was diluted with IPA (4 V) and further concentrated to about 3-4 V under reduced pressure at 35-45 C. The resulting EtOH solution of methyl (S)-3-(5-bromo-2-(2-(1-methoxyethyl)pyridin-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indol-3-yl)-2,2-dimethylpropanoate (Compound G), as a 1.4:1 mixture of diastereomers, was used directly in the next step.

Example 15Synthesis of Compound 1(2S)-2-cyclopentyl-2-((S)-7-((2R,3R)-3-cyclopropyl-1-methylaziridine-2-carbonyl)-2,7-diazaspiro[4.4]nonan-2-yl)-N-((22S,63S,4S)-12-(5-(4-cyclopropylpiperazin-1-yl)-2-((S)-1-methoxyethyl)pyridin-3-yl)-10,10-dimethyl-5,7-dioxo-11-(2,2,2-trifluoroethyl)-61,62,63,64,65,66-hexahydro-11H-8-oxa-2(4,2)-morpholina-1(5,3)-indola-6(1,3)-pyridazinacycloundecaphane-4-yl)acetamide

##STR00287##

Part 1Synthesis of tert-butyl (S)-2-((S)-2-(((benzyloxy)carbonyl)amino)-3-((S)-3-((3-(5-bromo-2-(5-(4-cyclopropylpiperazin-1-yl)-2-((S)-1-methoxyethyl)pyridin-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indol-3-yl)-2,2-dimethylpropoxy)carbonyl)tetrahydropyridazin-1(2H)-yl)-3-oxopropyl)morpholine-4-carboxylate

##STR00288##

[0621] To a reactor was charged DCM (746.15 kg, 9 V), followed by addition of Compound H (67.35 kg, 108.0 mol, 1.0 equiv), Compound J (64.57 kg, 124.0 mol, 1.15 equiv) DCM solution (1 V) of 1-hydroxybenzotriazole (14.02 kg, 103.8 mol, 1.0 equiv), DMAP (6.50 kg, 53.2 mol, 0.5 equiv), DIPEA (27.00 kg, 208.9 mol, 2.0 equiv) and N-(3-Dimethylaminopropyl)-N-ethylcarbodiimide hydrochloride (29.65 kg, 154.7 mol, 1.5 equiv) at 205 C. The reaction mixture was stirred at 155 C. for 16 h. Reaction monitoring by HPLC showed that the reaction is complete. The reaction mixture was quenched with aq. NaHCO.sub.3 solution (7% w/w, 7.3% w/w of Compound H) at 155 C. and the organic layer was separated. The organic layer was washed with 5% w/w citric acid aqueous solution (731.64 kg). The crude organic solution was concentrated under reduced pressure at NMT 45 C. to 3.5-4.5 V. The concentrated residue was further diluted with DCM (511.85 kg, 6 V). The resulting solution was concentrated under reduced pressure at NMT 45 C. to 3.5-4.5 V. The DCM solution of tert-butyl (S)-2-((S)-2-(((benzyloxy)carbonyl)amino)-3-((S)-3-((3-(5-bromo-2-(5-(4-cyclopropylpiperazin-1-yl)-2-((S)-1-methoxyethyl)pyridin-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indol-3-yl)-2,2-dimethylpropoxy)carbonyl)tetrahydropyridazin-1(2H)-yl)-3-oxopropyl)morpholine-4-carboxylate was used for next step without further purification (90.4% a/a).

[0622] LCMS (ESI+): Calculated for C.sub.55H.sub.72BrF.sub.3N.sub.8O.sub.9 (M+H): 1125.5; Found: 1125.9

[0623] .sup.1H NMR (400 MHz, DMSO-d.sub.6, 25 C.) 8.48 (d, J=2.8 Hz, 1H), 7.87 (d, J=2.0 Hz, 1H), 7.65 (d, J=8.7 Hz, 1H), 7.39 (dd, J=8.8, 1.9 Hz, 1H), 7.33 (m, 5H), 7.22 (d, J=2.7 Hz, 1H), 7.12 (d, J=8.8 Hz, 1H), 5.33 (dd, J=17.0, 8.6 Hz, 1H), 5.07 (m, 2H), 4.98 (s, 2H), 4.51 (m, 1H), 3.97 (s, 1H), 2.88 (s, 3H), 1.37 (s, 9H), 1.32 (d, J=6.2 Hz, 3H), 0.78 (s, 3H), 0.73 (s, 3H), 0.42 (m, 2H), 0.34 (m, 2H).

Part 2Synthesis of 3-(5-bromo-2-(5-(4-cyclopropylpiperazin-1-yl)-2-((S)-1-methoxyethyl)pyridin-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indol-3-yl)-2,2-dimethylpropyl (S)-1-((S)-2-(((benzyloxy)carbonyl)amino)-3-((S)-morpholin-2-yl)propanoyl)hexahydropyridazine-3-carboxylate

##STR00289##

Condition 1: Work-Up with K.SUB.2.CO.SUB.3

[0624] To a reactor was charged with tert-butyl (S)-2-((S)-2-(((benzyloxy)carbonyl)amino)-3-((S)-3-((3-(5-bromo-2-(5-(4-cyclopropylpiperazin-1-yl)-2-((S)-1-methoxyethyl)pyridin-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indol-3-yl)-2,2-dimethylpropoxy)carbonyl)tetrahydropyridazin-1(2H)-yl)-3-oxopropyl)morpholine-4-carboxylate DCM solution (64.1 kg, 1.0 eq) and it was cooled to 55 C. under nitrogen. TFA (237.10 kg, 2.4 V) was charged to the reactor dropwise while maintaining the inner temperature at 55 C. The reaction mixture was stirred at 105 C. for 1 h and then stirred at 155 C. for no less than 4 h. Reaction monitoring by HPLC showed reaction is complete. Aqueous solution of K.sub.2CO.sub.3 (731.65 kg, 40% w/w) was charged to reaction mixture to adjust the pH to the range of 7-10 while maintaining the temperature at 1010 C. H.sub.2O (57.50 kg, 4 V) was added to the mixture and stirred for 0.5 h. The organic layer was separated and was washed with H.sub.2O (452.5 kg, 7 V). The solvent in the solution was exchanged with MTBE (470.9 kg, 10 V) by distillation. The resulting MTBE solution of crude product was added to n-heptane (872.05 kg, 20 V) in another reactor over 6 h at 155 C. The mixture was stirred at 155 C. for at least 3 h. The slurry was filtered, and the wet cake was washed with n-heptane (87 kg, 2V). The crude solid was dried at 255 C. under vacuum for 10 h to give 3-(5-bromo-2-(5-(4-cyclopropylpiperazin-1-yl)-2-((S)-1-methoxyethyl)pyridin-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indol-3-yl)-2,2-dimethylpropyl (S)-1-((S)-2-(((benzyloxy)carbonyl)amino)-3-((S)-morpholin-2-yl)propanoyl)hexahydropyridazine-3-carboxylate as a white solid (103.72 kg, 94.1% a/a, 90.1% yield over 2 steps).

Condition 2: Work-Up with NaOH

[0625] To a reactor was charged tert-butyl (S)-2-((S)-2-(((benzyloxy)carbonyl)amino)-3-((S)-3-((3-(5-bromo-2-(5-(4-cyclopropylpiperazin-1-yl)-2-((S)-1-methoxyethyl)pyridin-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indol-3-yl)-2,2-dimethylpropoxy)carbonyl)tetrahydropyridazin-1(2H)-yl)-3-oxopropyl)morpholine-4-carboxylate DCM solution (1.0 eq) and it was cooled to 55 C. under nitrogen. TFA (237.10 kg, 2.4 V) was charged to the reactor dropwise while maintaining the inner temperature at 55 C. The reaction mixture was stirred at 105 C. for 1 h and then stirred at 155 C. for no less than 4 h. Reaction monitoring by HPLC showed reaction is complete. Aqueous solution of NaOH (20% w/w, 5 of the starting material) was charged to reaction mixture and stirred for 1 h at 5 C. 5% aq. NaOH solution (4f the starting material) was added to adjust the pH to the range of 8-10 while maintaining the temperature at 0-10 C. H.sub.2O (4 V) was added to the mixture and stirred for 0.5 h. The organic layer was separated and was washed with H.sub.2O (7 V). The solvent in the solution was exchanged with MTBE (470.9 kg, 10 V) by evaporation. The resulting MTBE solution of crude product was added to n-heptane (20 V) in another reactor over 6 h at 155 C. The mixture was stirred at 155 C. for at least 3 h. The slurry was filtered, and the wet cake was washed with n-heptane (2V). The crude solid was dried at 255 C. under vacuum for 10 h to give 3-(5-bromo-2-(5-(4-cyclopropylpiperazin-1-yl)-2-((S)-1-methoxyethyl)pyridin-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indol-3-yl)-2,2-dimethylpropyl (S)-1-((S)-2-(((benzyloxy)carbonyl)amino)-3-((S)-morpholin-2-yl)propanoyl)hexahydropyridazine-3-carboxylate as a white solid.

[0626] LCMS (ESI+): Calculated for C.sub.50H.sub.64BrF.sub.3N.sub.8O.sub.7 (M+H): 1025.4; Found: 1025.8

[0627] .sup.1H NMR (400 MHz, DMSO-d.sub.6, 25 C.) 8.48 (d, J=2.9 Hz, 1H), 7.87 (d, J=2.0 Hz, 1H), 7.65 (d, J=8.7 Hz, 1H), 7.39 (dd, J=8.8, 1.9 Hz, 1H), 7.35 (m, 5H), 7.21 (d, J=2.8 Hz, 1H), 7.03 (d, J=8.8 Hz, 1H), 5.33 (dd, J=17.1, 8.6 Hz, 1H), 5.05 (m, 2H), 4.98 (s, 2H), 4.49 (dq, J=15.7, 9.3 Hz, 1H), 3.98 (s, 1H), 2.88 (s, 3H), 1.32 (d, J=6.2 Hz, 3H), 0.78 (s, 3H), 0.73 (s, 3H), 0.43 (m, 2H), 0.34 (m, 2H).

Part 3Synthesis of benzyl ((22S,63S,4S)-12-(5-(4-cyclopropylpiperazin-1-yl)-2-((S)-1-methoxyethyl)pyridin-3-yl)-10,10-dimethyl-5,7-dioxo-11-(2,2,2-trifluoroethyl)-61,62,63,64,65,66-hexahydro-11H-8-oxa-2(4,2)-morpholina-1(5,3)-indola-6(1,3)-pyridazinacycloundecaphane-4-yl)carbamate

Condition 1: P(t-Bu).SUB.3..Math.HBF.SUB.4 .Ligand

##STR00290##

[0628] To a reactor was charged toluene (591.90 kg, 25 V), 3-(5-bromo-2-(5-(4-cyclopropylpiperazin-1-yl)-2-((S)-1-methoxyethyl)pyridin-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indol-3-yl)-2,2-dimethylpropyl (S)-1-((S)-2-(((benzyloxy)carbonyl)amino)-3-((S)-morpholin-2-yl)propanoyl)hexahydropyridazine-3-carboxylate (27.00 kg, 1.0 eq), anhydrous K.sub.3PO.sub.4 (27.90 kg, 5.0 eq). The mixture was degassed by bubbling nitrogen for 1 h under agitation at 205 C. (the content of 02 s 1000 ppm). Pd.sub.2(dba).sub.3.Math.CHCl.sub.3 (1.41 kg, 0.047 eq), P(t-Bu).sub.3.Math.HBF.sub.4 (1.98 kg, 0.105 eq) and H.sub.2O (4.35 kg, 9.0 equiv) were charged to the reactor and the reaction mixture was further degassed by bubbling nitrogen for 1 h. The reaction mixture was agitated at 905 C. for 11 h. Reaction monitoring by HPLC showed the reaction was complete. The crude reaction mixture was cooled to 2010 C. and the mixture was filtered. To the filtrate EtOAc (449.90 kg, 10 V) was charged. The solvent in the resulting solution was concentrated to 3-5 V under reduced pressure at NMT 55 C. The solvent swap with EtOAc was repeated twice. The concentrated solution was washed with H.sub.2O (99.50 kg, 2 V), aq. NaCl (10% w/w, 26.95 kg, 0.5 V), and aq. HCl (1% w/w, 17.50 kg, 0.4 V) successively at 510 C. The organic phase was separated and extracted with aq. HCl solution (1% w/w, 4 V and 3 V) twice. To the combined aqueous phases was charged EtOAc (183.25 kg, 4 V) and n-heptane (35.05 kg, 1 V) at 510 C. Aqueous NaCl solution (25% w/w, 33.45 kg, 0.6 V) was added dropwise to the mixture over 0.5 h. After stirring for 30 min, the aqueous phase was separated. The aqueous phase was concentrated under vacuum at NMT 35 C. to completely remove the organic solvent residue. The resulting suspension was stirred at 510 C. for at least 2 h. The slurry was filtered and the wet cake was washed with aq. NaCl solution (10% w/w, 10.35 kg, 0.2 V). The filter cake was collected as the HCl salt of the product. To a reactor was added EtOAc (468.10 kg, 10 V) and H.sub.2O (262.00 kg, 5 V). The filter cake was added to the reactor at 510 C. with agitation. Aqueous K.sub.2CO.sub.3 solution (40% w/w, 20.95 kg) was added to the solution to adjust the pH to 8-10. The organic phase was separated, and the aqueous phase was washed with EtOAc (234.00 kg, 5 V). The combined organic phase was washed with aq. NaCl solution (10% w/w, 278.00 kg, 5 V). The organic phase was concentrated to 5.5-6.5 V under vacuum at NMT 45 C. The concentrated organic phase was extracted with aq. HCl (1% w/w, 103.70 kg, 2 V) two times and the aqueous phases were collected. The aqueous phase was concentrated at NMT 35 C. under reduced pressure (0.09 mpa) and solid precipitates crashed out. The residue was agitated for 1 h at 510 C. before aq. NaCl solution was added (25% w/w, 15.50 kg, 0.25 V). The resulting mixture was stirred for 2 h at 510 C. The slurry was filtered and the filter cake was washed with aq. NaCl solution (10% w/w, 16.10 kg, 0.3 V). The filter cake was collected as the HCl salt of the product. To a reactor was added EtOAc (234.00 kg, 5 V) and H.sub.2O (130.50 kg, 2.5 V). The filter cake was added to the reactor at 510 C. with agitation. Aqueous K.sub.2CO.sub.3 solution (40% w/w) was added to the solution to adjust the pH to 8-10. The organic phase was separated, and the aqueous phase was washed with EtOAc (140.60 kg, 3 V). The combined organic phase was washed with aq. NaCl solution (10% w/w, 154.85 kg). The organic phase was concentrated to 1.5-2.5 V under vacuum at NMT 45 C. To a reactor with n-heptane (704.55 kg, 21 V) was charged the concentrated residue of the product at 205 C. over 2 h. The slurry was agitated for 2 h at 205 C. The slurry was filtered and the filter cake was washed with n-heptane (34.85 kg, 2 V). The wet cake was dried in oven at 305 C. for 10 h under vacuum to afford benzyl ((22S,63S,4S)-12-(5-(4-cyclopropylpiperazin-1-yl)-2-((S)-1-methoxyethyl)pyridin-3-yl)-10,10-dimethyl-5,7-dioxo-11-(2,2,2-trifluoroethyl)-61,62,63,64,65,66-hexahydro-11H-8-oxa-2(4,2)-morpholina-1(5,3)-indola-6(1,3)-pyridazinacycloundecaphane-4-yl)carbamate (98.6% a/a, 20.60 kg, 44.6% yield) as white solid.

[0629] LCMS (ESI+): Calculated for C.sub.50H.sub.63F.sub.3N.sub.8O.sub.7 (M+H): 945.5; Found: 945.9

[0630] .sup.1H NMR (300 MHz, DMSO-d.sub.6, 25 C.) 8.46 (dd, J=2.8 Hz, 1H), 7.57 (d, J=9.0 Hz, 1H), 7.35 (m, 5H), 7.17 (m, 2H), 7.08 (d, J=9.1 Hz, 1H), 6.98 (s, 1H), 5.28 (m, 2H), 4.97 (m, 2H), 4.71 (m, 1H), 3.33 (s, 3H), 1.33 (d, J=6.1 Hz, 3H), 0.83 (s, 3H), 0.43 (m, 7H), 0.34 (s, 2H).

Condition 2: Qphos Ligand

##STR00291##

[0631] To a reactor was charged toluene (496.90 kg, 35 V), 3-(5-bromo-2-(5-(4-cyclopropylpiperazin-1-yl)-2-((S)-1-methoxyethyl)pyridin-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indol-3-yl)-2,2-dimethylpropyl (S)-1-((S)-2-(((benzyloxy)carbonyl)amino)-3-((S)-morpholin-2-yl)propanoyl)hexahydropyridazine-3-carboxylate (15.01 kg, 14.6 mol, 1.0 eq), anhydrous K.sub.3PO.sub.4 (16.21 kg, 76.4 mol, 5.0 eq). The mixture was degassed by bubbling nitrogen for 1 h under agitation at 205 C. (the content of 02 s 1000 ppm). Pd.sub.2(dba).sub.3.Math.CHCl.sub.3 (745.60 g, 0.7 mol, 0.047 eq), 1,2,3,4,5-Pentaphenyl-1-(di-tert-butylphosphino)ferrocene (1.09 kg, 1.5 mol, 0.105 eq) and H.sub.2O (1.65 kg, 91.7 mol, 6.3 equiv) were charged to the reactor and the reaction mixture was further degassed by bubbling nitrogen for 1 h (the content of O.sub.21000 ppm). The reaction mixture was agitated at 905 C. for 10 h. Reaction monitoring by HPLC showed the reaction was complete. The crude reaction mixture was cooled to 2010 C. and the mixture was filtered. The filtrate was concentrated to 3-5 V. To the concentrated residue EtOAc (358.00 kg, 10 V) was charged. The resulting solution was concentrated to 3-5 V under reduced pressure at NMT 55 C. The solvent swap with EtOAc was repeated twice. The concentrated solution was washed with H.sub.2O (95.00 kg, 2.5 V) and aq. HCl (1% w/w, 25.00 kg, 0.5 V) successively at 05 C. The organic phase was separated and extracted with aq. HCl solution (1% w/w, 4 V and 3 V) twice. To the combined aqueous phases was charged 25% aq. NaCl (27.10 kg, 0.80 V), EtOAc (137.15 kg, 4 V) and n-heptane (25.85 kg, 1 V) at 510 C. After stirring for 30 min, the aqueous phase was separated. The aqueous phase was concentrated under vacuum at NMT 35 C. to completely remove the organic solvent residue. The resulting solution was stirred at 510 C. for at least 2 h. The mother liquor was sampled to get the concentration of the product. The slurry was filtered and the wet cake was washed with aq. NaCl solution (10% w/w, 10.00 kg, 0.3 V). The filter cake was collected as the HCl salt of the product. To a reactor was added EtOAc (340.45 kg, 10 V) and H.sub.2O (185.50 kg, 5 V). The filter cake was added to the reactor at 510 C. with agitation. Aqueous K.sub.2CO.sub.3 solution (40% w/w, 16.00 kg) was added to the solution to adjust the pH to 8-10 range. The organic phase was separated and the aqueous phase was washed with EtOAc (174.00 kg, 5 V). The combined organic phases was washed with aq. NaCl solution (10% w/w, 190.50 kg, 5 V). The organic phase was concentrated to 2-3 V under vacuum at NMT 45 C. To a reactor with n-heptane (539.60 kg, 21 V) was charged the concentrated residue of the product at 205 C. over 2 h. The slurry was agitated for 2 h at 205 C. The slurry was filtered and the filter cake was washed with n-heptane (2 V). The wet cake was dried in oven at 305 C. for 10 h under reduced pressure to afford crude product. To a reactor with H.sub.2O (91.50 kg, 6.8% w/w) and HCl (2.59 kg, 0.19% w/w) was charged crude product (13.38 kg, 14.2 mol, 1.0 equiv) at 55 C. The mixture was stirred at 55 C. for 2 h and then was charged 25% aq. NaCl (23.90 kg, 1 V) dropwise. The resulting slurry was filtered and the wet cake was washed with 10% aq. NaCl (31.40 kg, 2 V). The wet cake was suspended in H.sub.2O (94.00 kg, 7 V) and EtOAc (96.45 kg, 8 V) at 510 C. The pH of aqeuous layer was adjusted to 8-10 with aq.Math.K.sub.2CO.sub.3 solution (40% w/w, 10.65 kg). The organic layer was separated and the aqueous layer was washed with EtOAc (36.40 kg, 3 V). The organic layers were combined and washed with 10% w/w aq. NaCl solution (67.10 kg, 5% w/w). The organic phase was concentrated to 2-3 V under vacuum at NMT 45 C. The concentrated residue was diluted with EtOAc (60.75 kg, 5 V) and the resulting mixture was concentrated to 2-3 V under vacuum at NMT 45 C. The concentrated residue was added to n-heptane (208.10 kg, 21 V) at 205 C. dropwise and the mixture was stirred for 2 h. The slurry was filtered and wet cake was rinsed with n-heptane (18.65 kg, 2 V). The wet cake was dried in the oven under vacuum at 305 C. for 6 h to afford benzyl ((22S,63S,4S)-12-(5-(4-cyclopropylpiperazin-1-yl)-2-((S)-1-methoxyethyl)pyridin-3-yl)-10,10-dimethyl-5,7-dioxo-11-(2,2,2-trifluoroethyl)-61,62,63,64,65,66-hexahydro-11H-8-oxa-2(4,2)-morpholina-1(5,3)-indola-6(1,3)-pyridazinacycloundecaphane-4-yl)carbamate (11.91 kg, 87.6% yield) as white solid.

Condition 3: Using Dioxane as the Solvent

##STR00292##

[0632] To a reactor was charged 1,4-dioxane (58.0 L, 35 V), 3-(5-bromo-2-(5-(4-cyclopropylpiperazin-1-yl)-2-((S)-1-methoxyethyl)pyridin-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indol-3-yl)-2,2-dimethylpropyl (S)-1-((S)-2-(((benzyloxy)carbonyl)amino)-3-((S)-morpholin-2-yl)propanoyl)hexahydropyridazine-3-carboxylate (1.50 kg, 1.5 mol, 1.0 eq), and anhydrous K.sub.3PO.sub.4 (1.03 kg, 4.8 mol, 3.3 eq). The mixture was degassed by bubbling nitrogen for 1 h under agitation at 205 C. (the content of 02 s 1000 ppm). P(t-Bu).sub.3PdG.sub.3 (93 g, 0.15 mol, 0.11 eq), P(t-Bu).sub.3.Math.HBF.sub.4 (46.2 g, 0.15 mol, 0.11 eq) and H.sub.2O (205 mL, 6.6 equiv) were charged to the reactor and the reaction mixture was further degassed by bubbling nitrogen for 1 h (the content of O.sub.21000 ppm). The reaction mixture was agitated at 905 C. for 10 h. Reaction monitoring by HPLC showed the reaction was complete. The crude reaction mixture was cooled to 2010 C. and the mixture was filtered. The filter cake was washed with 1,4-dioxane (1.6 L, 1 V). The filtrate was concentrated to afford the crude product as yellow solids. The solids were dissolved in 2-MeTHF (13.8 L, 6 V) and aq. HCl (0.08 M, 5 V). The organic layer was separated and washed with aq. HCl (0.08 M, 4 V). The organic layer was extracted with aq. HCl (0.3 M, 5 V) for two times at 15 C. The combined aqueous layer was washed with 2-MeTHF (10.5 L, 5 V) at 15 C. twice. Na.sub.2HCO.sub.3 solids were added to the aqueous layer to adjust pH to 7 at 10-20 C. The resulting aqueous solution was extracted with 2-MeTHF (10.5 L, 10 V) two times. The combined organic layer was washed with aq. acetic acid (0.04 M, 10.5 L, 10 V) and 25% w/w aq. NaCl (10.5 L, 10 V) in sequence. The resulting organic layer was concentrated under reduced pressure to afford benzyl ((22S,63S,4S)-12-(5-(4-cyclopropylpiperazin-1-yl)-2-((S)-1-methoxyethyl)pyridin-3-yl)-10,10-dimethyl-5,7-dioxo-11-(2,2,2-trifluoroethyl)-61,62,63,64,65,66-hexahydro-11H-8-oxa-2(4,2)-morpholina-1(5,3)-indola-6(1,3)-pyridazinacycloundecaphane-4-yl)carbamate as yellow solids (1.056 kg, 91.6% a/a).

Condition 4: Pd(t-Bu.SUB.3.P).SUB.2 .Catalyst and New Isolation Process

##STR00293##

[0633] To a reactor was charged anisole (20 V) and 3-(5-bromo-2-(5-(4-cyclopropylpiperazin-1-yl)-2-((S)-1-methoxyethyl)pyridin-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indol-3-yl)-2,2-dimethylpropyl (S)-1-((S)-2-(((benzyloxy)carbonyl)amino)-3-((S)-morpholin-2-yl)propanoyl)hexahydropyridazine-3-carboxylate (1.0 eq). The mixture was degassed by bubbling nitrogen for 1 h under agitation at 205 C. (the content of O.sub.21000 ppm). Cs.sub.2CO.sub.3 (3 equiv) and bis(tri-t-butylphosphine)palladium (0.1 equiv) were charged to the reactor and the reaction mixture was further degassed by bubbling nitrogen for 1 h (the content of 02 S 1000 ppm). The reaction mixture was agitated at 905 C. for 11 h. Reaction monitoring by HPLC showed the reaction was complete. The crude reaction mixture was cooled to 2010 C. and the mixture was filtered. The filter cake was rinsed with anisole (3.5% w/w) and EtOAc (3.6% w/w). The filtrate was concentrated to 3-5 V under reduced pressure at NMT 55 C. The solvent swap with EtOAc was repeated twice. The EtOAc solution was extracted with aq. HCl solution (1% w/w, 4 V and 3 V) twice to form the HCl salt of the product. Alternatively, the EtOAc solution was extracted with p-toluenesulfonic acid or trichloroacetic acid to form the corresponding tosylate salt of the product or the corresponding trichloroacetate salt of the product. The combined aqueous phase was seeded with the HCl salt of the product (0.01% w/w) at 5 C. The mixture was stirred for 5 h at 5 C. before 25% aq. NaCl (0.6% w/w) was added. The resulting mixture was continued to stir for 5 h at 5 C. The slurry was filtered, and the wet cake was washed with 10% aq. NaCl solution (0.2% w/w). The wet cake was suspended in EtOAc (9% w/w) and H.sub.2O (6.5% w/w). A 40% K.sub.2CO.sub.3 solution was added to adjust the pH to 8-10 at 0-10 C. After agitating for 1 h, the organic phase was separated, and the aqueous phase was extracted with EtOAc (4.5% w/w). The organic layers were combined and purified through a pad of silica gel (100-200 mesh, 0.7% w/w). The eluent after silica gel was concentrated to 2.5 V under reduced pressure NMT 40 C. The concentrated solution was added to n-heptane (1.43% w/w) dropwise at 20 C. The resulting mixture was stirred for 5 h at 20 C. The slurry was filtered and the filter cake was washed with n-heptane (1.4% w/w). The wet cake was dried in the oven at 30 C. for 8 h under reduced pressure to afford benzyl ((22S,63S,4S)-12-(5-(4-cyclopropylpiperazin-1-yl)-2-((S)-1-methoxyethyl)pyridin-3-yl)-10,10-dimethyl-5,7-dioxo-11-(2,2,2-trifluoroethyl)-61,62,63,64,65,66-hexahydro-11H-8-oxa-2(4,2)-morpholina-1(5,3)-indola-6(1,3)-pyridazinacycloundecaphane-4-yl)carbamate as white solid.

Part 4Synthesis of (22S,63S,4S)-4-amino-12-(5-(4-cyclopropylpiperazin-1-yl)-2-((S)-1-methoxyethyl)pyridin-3-yl)-10,10-dimethyl-11-(2,2,2-trifluoroethyl)-61,62,63,64,65,66-hexahydro-11H-8-oxa-2(4,2)-morpholina-1(5,3)-indola-6(1,3)-pyridazinacycloundecaphane-5,7-dione

##STR00294##

Condition 1: Using K.SUB.2.CO.SUB.3 .Base

[0634] A pressure reactor was charged with THE (211.60 kg, 12 V), benzyl ((22S,63S,4S)-12-(5-(4-cyclopropylpiperazin-1-yl)-2-((S)-1-methoxyethyl)pyridin-3-yl)-10,10-dimethyl-5,7-dioxo-11-(2,2,2-trifluoroethyl)-61,62,63,64,65,66-hexahydro-11H-8-oxa-2(4,2)-morpholina-1(5,3)-indola-6(1,3)-pyridazinacycloundecaphane-4-yl)carbamate (19.89 kg, 21 mol, 1.0 equiv), 5% w/w Pd/C (20% w/w, 4.25 kg), and K.sub.2CO.sub.3 (2.97 kg, 21 mol, 1 equiv). It was agitated at 0.2 mpa H.sub.2, 20-25 C. The atmosphere was exchanged by venting and refilling with fresh H.sub.2 gas several times during the course of the reaction. After reaction is complete as monitored by HPLC, the atmosphere was exchanged with N.sub.2 gas and the reaction mixture was filtered through a pad of diatomite. The filtrates were then concentrated to 3-5 V under reduced pressure at NMT 40 C. and diluted with EtOAc (124.65 kg, 7 V). The mixture was further concentrated under reduced pressure at NMT 40 C. to 3-5 V. The concentrated residue was diluted with EtOAc (127.40 kg, 7 V) and the resulting solution was washed with 5% w/w aq. N-acetyl-L-cysteine solution (198.05 kg, 10% w/w). The mixture was filtered via fluid filter to remove the Pd complex. The filtrate was added 5% w/w aq. Na.sub.2CO.sub.3 solution to adjust the pH of aqueous layer to 7.4-8.0. The organic layer was separated and washed with 5% w/w aq. Na.sub.2CO.sub.3 (5% w/w) and 5% w/w aq. NaCl (5% w/w) in sequence. The organic layer was concentrated under reduced pressure at NMT 40 C. to 3-5 V. The concentrated residue was diluted with EtOAc (5 V) and the resulting solution was further concentrated under reduced pressure at NMT 40 C. to 3-5 V. The solvent swap process was repeated two times. The resulting crude product solution in EtOAc was added MTBE (58.65 kg, 4 V) over 2 h, followed by adding a seed of the product (0.01% w/w, 200 g) at 2010 C. The mixture was agitated at 2010 C. for 2 h. The mixture was diluted with MTBE (5 V) and concentrated to 4-6 V under reduced pressure at NMT 40 C. The solvent swap with MTBE was repeated three times. The concentrated solution was cooled to 05 C. and stirred for 2 h. The slurry was filtered and the wet cake was washed with MTBE (20.85 kg, 1 V). The filter cake was dried in the oven at 305 C. under vacuum to afford (22S,63S,4S)-4-amino-12-(5-(4-cyclopropylpiperazin-1-yl)-2-((S)-1-methoxyethyl)pyridin-3-yl)-10,10-dimethyl-11-(2,2,2-trifluoroethyl)-61,62,63,64,65,66-hexahydro-11H-8-oxa-2(4,2)-morpholina-1(5,3)-indola-6(1,3)-pyridazinacycloundecaphane-5,7-dione (13.79 kg, 99.7% a/a, 84% yield) as white solid.

[0635] LCMS (ESI+): Calculated for C.sub.42H.sub.57F.sub.3N.sub.8O.sub.5 (M+H): 811.4; Found: 811.5

[0636] .sup.1H NMR (400 MHz, DMSO-d.sub.6, 25 C.) 8.45 (d, J=2.8 Hz, 1H), 7.58 (d, J=8.9 Hz, 1H), 7.15 (d, J=2.7 Hz, 1H), 7.09 (d, J=8.9 Hz, 1H), 6.99 (s, 1H), 5.35 (dd, J=16.5, 8.5 Hz, 1H), 5.06 (d, J=12.2 Hz, 1H), 4.77 (dd, J=16.6, 9.1 Hz, 1H), 3.33 (s, 3H), 1.33 (d, J=6.1 Hz, 3H), 0.79 (s, 3H), 0.42 (m, 4H), 0.34 (s, 3H).

Condition 2: No K.SUB.2.CO.SUB.3 .Base and Inclusion of Citric Acid Wash

##STR00295##

[0637] To a pressure reactor was charged and THE (12 V), benzyl ((22S,63S,4S)-12-(5-(4-cyclopropylpiperazin-1-yl)-2-((S)-1-methoxyethyl)pyridin-3-yl)-10,10-dimethyl-5,7-dioxo-11-(2,2,2-trifluoroethyl)-61,62,63,64,65,66-hexahydro-11H-8-oxa-2(4,2)-morpholina-1(5,3)-indola-6(1,3)-pyridazinacycloundecaphane-4-yl)carbamate (1.0 equiv) and 5% w/w Pd/C (40% w/w, 4.25 kg). It was agitated at 0.2 mpa H.sub.2, 20-25 C. The atmosphere was exchanged by venting and refilling with fresh H.sub.2 gas several times during the course of the reaction. After reaction is complete as monitored by HPLC, the atmosphere was exchanged with N.sub.2 gas and the reaction mixture was filtered through a pad of diatomite. MTBE (14 V) and 0.2% aq. citric acid solution (6 V) were added to the filtrate. The organic phase was separated and washed with 3% w/w aq. Na.sub.2CO.sub.3 solution (6 V). The organic layer was separated and washed with H.sub.2O (3 V). The organic phase was separated and charged with MTBE (10 V). The resulting mixture was concentrated to 3-4 V. The solvent swap with MTBE was repeated two times. The concentrated residue was diluted with MTBE (1 V). The slurry was stirred for 3 h before it was filtered. The filter cake was washed with MTBE (1 V) and the wet cake was dried in the oven at 30-35 C. under reduced pressure to afford (22S,63S,4S)-4-amino-12-(5-(4-cyclopropylpiperazin-1-yl)-2-((S)-1-methoxyethyl)pyridin-3-yl)-10,10-dimethyl-11-(2,2,2-trifluoroethyl)-61,62,63,64,65,66-hexahydro-11H-8-oxa-2(4,2)-morpholina-1(5,3)-indola-6(1,3)-pyridazinacycloundecaphane-5,7-dione as white solid.

Condition 3: Using 2-MeTHF as Solvent, No Solvent Switch

##STR00296##

[0638] To an autoclave was charged 2-MeTHF (12 V), benzyl ((22S,63S,4S)-12-(5-(4-cyclopropylpiperazin-1-yl)-2-((S)-1-methoxyethyl)pyridin-3-yl)-10,10-dimethyl-5,7-dioxo-11-(2,2,2-trifluoroethyl)-61,62,63,64,65,66-hexahydro-11H-8-oxa-2(4,2)-morpholina-1(5,3)-indola-6(1,3)-pyridazinacycloundecaphane-4-yl)carbamate (1.0 equiv), 5% w/w Pd/C (20% w/w), and K.sub.2CO.sub.3 (1 equiv). It was agitated at 20-25 C. with the presence of 0.2 mpa H.sub.2. The atmosphere was exchanged by venting and refilling with fresh H.sub.2 gas several times during the course of the reaction. After reaction is complete as monitored by HPLC, the atmosphere was exchanged with N.sub.2 gas and the reaction mixture was filtered through a pad of diatomite and the wet cake was washed with 2-MeTHF (5-7 times). The filtrate was washed with 5% w/w aq. N-acetyl-L-cysteine solution (10 V) at 10 C. The mixture was filtered. The filtrate was adjusted to pH=7.4-8.0 with 5% w/w aq. Na.sub.2CO.sub.3 solution. The organic phase was separated. The aqueous phase was extracted with 2-MeTHF (3 V). The organic phase was combined and washed with 5% w/w aq. Na.sub.2CO.sub.3 solution (5 V) and 5% w/w aq. NaCl solution (5 V) successively. The organic phase was concentrated to 7 V at NMT 35 C. and swapped with 2-MeTHF (10 V) twice. To the concentrated solution of product crude was added a seed of the product (0.01) and then n-heptane (14 V) dropwise at 25 C. The resulting slurry was stirred at 5 C. for 9 h. The slurry was filtered and the wet cake was rinsed with 2-MeTHF/n-heptane (v/v=1:2, 3 V). The wet cake was dried in the oven at 30 C. for 39 h under vacuum. (22S,63S,4S)-4-amino-12-(5-(4-cyclopropylpiperazin-1-yl)-2-((S)-1-methoxyethyl)pyridin-3-yl)-10,10-dimethyl-11-(2,2,2-trifluoroethyl)-61,62,63,64,65,66-hexahydro-11H-8-oxa-2(4,2)-morpholina-1(5,3)-indola-6(1,3)-pyridazinacycloundecaphane-5,7-dione was obtained as white solids (611.53 g, 97.1 assay, 85.5% yield).

Part 5Synthesis of Compound 1(2S)-2-cyclopentyl-2-((S)-7-((2R,3R)-3-cyclopropyl-1-methylaziridine-2-carbonyl)-2,7-diazaspiro[4.4]nonan-2-yl)-N-((22S,63S,4S)-12-(5-(4-cyclopropylpiperazin-1-yl)-2-((S)-1-methoxyethyl)pyridin-3-yl)-10,10-dimethyl-5,7-dioxo-11-(2,2,2-trifluoroethyl)-61,62,63,64,65,66-hexahydro-11H-8-oxa-2(4,2)-morpholina-1(5,3)-indola-6(1,3)-pyridazinacycloundecaphane-4-yl)acetamide

##STR00297##

Condition 1: HOPO/EDCl Coupling, 2-MeTHF/IPA/Heptane Crystallization

[0639] To a reactor was charged EtOAc (94.80 kg, 10 V), (22S,63S,4S)-4-amino-12-(5-(4-cyclopropylpiperazin-1-yl)-2-((S)-1-methoxyethyl)pyridin-3-yl)-10,10-dimethyl-11-(2,2,2-trifluoroethyl)-61,62,63,64,65,66-hexahydro-11H-8-oxa-2(4,2)-morpholina-1(5,3)-indola-6(1,3)-pyridazinacycloundecaphane-5,7-dione (10.50 kg, 1.0 equiv), Compound E (5.30 kg, 1.1 equiv), DIPEA (6.60 kg, 4.0 equiv), DMAP (0.80 kg, 0.5 equiv), HOPO (0.23 kg, 0.15 equiv) and N-(3-Dimethylaminopropyl)-N-ethylcarbodiimide hydrochloride (3.50 kg, 1.40 equiv) in sequence. The mixture was agitated at 20-30 C. for 9 h until reaction is complete. The crude reaction mixture was added to a reactor with H.sub.2O (84.00 kg, 8 V) at 105 C. The biphasic mixture was agitated at 105 C. for 1 h and the organic phase was separated. The organic layer was washed with aq. NaHCO.sub.3 (5 wt %, 105.20 kg), and aq. NaCl (2% w/w, 105.20 kg) in sequence. To the organic phase was charged H.sub.2O (5 V) and the aqueous phase was adjusted to pH=5.5-6.1 with 5% w/w citric acid aq. solution (32.65 kg) at 55 C. The organic phase was separated. To the organic phase was added H.sub.2O (10.05 kg, 1 V) and 1% w/w aq. Na.sub.2CO.sub.3 solution (7.70 kg) was added to adjust pH to 9-10 at 55 C. The organic phase was separated and washed with 2% w/w aq. NaCl solution (84.00 kg). The organic phase was concentrated to 3-5 V under vacuum at NMT 40 C. The concentrated residue was diluted with 2-MeTHF (10 V) and the mixture was concentrated to 3-5 V under vacuum at NMT 40 C. The solvent swap with 2-MeTHF was repeated three times. The crude product was obtained as 2-MeTHF solution which is further purified by recrystallization. To the 2-MeTHF solution of Compound 1 was added IPA (9.70 kg, 1.0 V) and n-heptane (20.85 kg, 2.5 V) at 5+5 C., followed by addition of Compound 1 seed crystal (5% w/w, 0.92 kg). The slurry was stirred at 55 C. for 12 h. The slurry was filtered and the wet cake was dried in the oven at 3510 C. under vacuum to afford Compound 1 (10.84 kg, 71.6% yield).

[0640] LCMS (ESI+): Calculated for C.sub.63H.sub.88F.sub.3N.sub.11O.sub.7 (M+H): 1168.7; Found: 1169.2

[0641] .sup.1H NMR (400 MHz, CD3OD, 25 C.) 8.42 (d, J=2.4 Hz, 1H), 7.47 (d, J=9.2 Hz, 1H), 7.28 (d, J=1.2 Hz, 1H), 7.12 (m, 1H), 7.10 (m, 1H), 5.67 (d, J=9.2 Hz, 1H), 5.00 (m, 1H), 4.71 (m, 1H), 4.44 (d, J=12.8 Hz, 1H), 4.06 (d, J=2.0 Hz, 1H), 3.91-3.28 (m, 15H), 3.19 (s, 3H), 2.89-2.66 (m, 14H), 2.34 (d, J=14.0 Hz, 3H), 2.24-1.41 (m, 25H), 0.88 (s, 3H), 0.64 (m, 1H), 0.54 (s, 3H), 0.50-0.26 (m, 8H).

Condition 2: PyBOP/Oxyma Coupling, 4-Dioxane/IPE Crystallization

##STR00298##

[0642] To a reactor was charged DMF (60 mL, 6 V), Compound E (5.10 g, 1.05 equiv), DIPEA (7.95 g, 5.0 equiv), (22S,63S,4S)-4-amino-12-(5-(4-cyclopropylpiperazin-1-yl)-2-((S)-1-methoxyethyl)pyridin-3-yl)-10,10-dimethyl-11-(2,2,2-trifluoroethyl)-61,62,63,64,65,66-hexahydro-11H-8-oxa-2(4,2)-morpholina-1(5,3)-indola-6(1,3)-pyridazinacycloundecaphane-5,7-dione (10 g, 1.0 equiv) at 10-20 C. The mixture was cooled to 05 C. and oxyma (1.58 g, 0.9 equiv) was added. The reaction mixture was cooled to 105 C. and PyBOP (7.69 g, 1.2 equiv) in DMF (20 mL, 2 V) was added. The mixture was agitated at 105 C. for 1 h until reaction is complete. To the reaction mixture was added H.sub.2O (35 mL, 3.5 V) and the mixture was filtered through microporous filter. Another portion of H.sub.2O (54 V) was added to the filtrate at 205 C. and the mixture was stirred for 0.5 h. The slurry was filtered and the cake was washed with H.sub.2O (5 V). The wet cake was dried under N.sub.2 flow to afford pink solids. The pink solids were dissolved in IPOAc (7 V) and MTBE (5 V). The solution was washed with H.sub.2O (12 V), 0.83% w/w aq. NaH.sub.2PO.sub.4 solution (12 V) and 3% w/w aq. NaHCO.sub.3 solution (12 V) twice successively. The organic layer was concentrated to dryness. The residue was dissolved in IPOAc (15 V) and was slurried with 9% charcoal for 1 h at rt. The mixture was filtered. The filtrate was concentrated to dryness to afford crude Compound 1. The crude product was dissolved in 1,4-dioxane (2.5 V) and IPE (1.5 V) at 25 C. The solution was cooled to 10 C. and stirred for 2 h. IPE (1 V) was added dropwise to the solution over 8 h and then another portion of IPE (5 V) was added over 10 h. The resulting slurry was filtered and the filter cake was rinsed with 1,4-dioxane/IPE (v/v=2.5:7.5, 2 V). The solids were subjected to the second round of recrystallization by dissolving in 1,4-dioxane (2.5 V) and IPE (1.5 V) at 25 C. IPE (1 V) was dropped to the solution over 0.5 h at 25 C. Then the solution was cooled to 10 C. Compound 1 was seeded and the mixture was stirred for 3 h at 10 C. Another portion of IPE (3.5 V) was added to the mixture over 12 h and the slurry was stirred for 6 h. The slurry was filtered and the filter cake was rinsed with 1,4-dioxane/IPE (v/v=2.5:5.5, 2 V). The wet cake was dried under vacuum at 30 C. to afford Compound 1 as light yellow solids.

Condition 3: HOBt/EDCl Coupling

##STR00299##

[0643] To a reactor was charged EtOAc (58.11 kg, 10 V), (22S,63S,4S)-4-amino-12-(5-(4-cyclopropylpiperazin-1-yl)-2-((S)-1-methoxyethyl)pyridin-3-yl)-10,10-dimethyl-11-(2,2,2-trifluoroethyl)-61,62,63,64,65,66-hexahydro-11H-8-oxa-2(4,2)-morpholina-1(5,3)-indola-6(1,3)-pyridazinacycloundecaphane-5,7-dione (6.40 kg, 1.0 equiv), Compound E (3.50 kg, 1.1 equiv), DIPEA (4.20 kg, 4.0 equiv), DMAP (481.9 g, 0.5 equiv), HOBt (74.9 g, 0.07 equiv) and N-(3-Dimethylaminopropyl)-N-ethylcarbodiimide hydrochloride (2.15 kg, 1.40 equiv) in sequence. The mixture was agitated at 20-30 C. for 9 h until reaction is complete. The crude reaction mixture was added to a reactor with H.sub.2O (52.60 kg, 8 V) at 105 C. The biphasic mixture was agitated at 105 C. for 1 h and the organic phase was separated. The organic layer was washed with aq. NaHCO.sub.3 (5 wt %, 64.00 kg, 6.4% w/w) and aq. NaCl (2% w/w, 64.28 kg, 6.4% w/w) in sequence. To the organic phase was charged H.sub.2O (5 V) and the aqueous phase was adjusted to pH=5.0-5.6 with 5% w/w citric acid aq. solution (1.35 kg) at 55 C. The organic phase was separated. To the organic phase was added H.sub.2O (6.55 kg, 1 V) and 1% w/w aq. NaCO.sub.3 solution (10.30 kg) was added to adjust pH to 9-10 at 55 C. The organic phase was separated and washed with 2% w/w aq. NaCl solution (51.20 kg, 8% w/w). The organic phase was concentrated to 3-5 V under vacuum at NMT 40 C. The concentrated residue was diluted with 2-MeTHF (10 V) and the mixture was concentrated to 3-5 V under vacuum at NMT 40 C. The solvent swap with 2-MeTHF was repeated three times. The crude product was obtained as a 2-MeTHF solution which is further purified by recrystallization. To the 2-MeTHF solution of Compound 1 was added n-heptane (15.00 kg, 2.5 V) at 55 C., followed by addition of a seed crystal of Compound 1 (5% w/w, 0.92 kg). The anti-solvent consisted of 2-MeTHF (7.90 kg, 1.33 V), IPA (3.70 kg, 0.67 V) and n-heptane (122.25 kg, 26 V) was added to the solution of Compound 1. The slurry was stirred at 55 C. for 12 h. The slurry was filtered and the wet cake was dried in the oven at 3510 C. under vacuum to afford Compound 1 (6.24 kg, 98.4% a/a, 67.7% yield) as a white solid.

OTHER EMBODIMENTS

[0644] While the invention has been described in connection with specific embodiments thereof, it will be understood that it is capable of further modifications and this application is intended to cover any variations, uses, or adaptations of the invention following, in general, the principles of the invention and including such departures from the present disclosure come within known or customary practice within the art to which the invention pertains and may be applied to the essential features set forth herein.

[0645] All publications, patents and patent applications are herein incorporated by reference in their entirety to the same extent as if each individual publication, patent, or patent application was specifically and individually indicated to be incorporated by reference in its entirety.