Heterocyclic compounds and their use in preventing or treating bacterial infections
11530227 · 2022-12-20
Assignee
Inventors
- Julien Barbion (Sannois, FR)
- Audrey Caravano (Enghien les Bains, FR)
- Sophie Chasset (Nandy, FR)
- Francis Chevreuil (Chantilly, FR)
- Nicolas Lecointe (Paris, FR)
- Frédéric Le Strat (Combs la Ville, FR)
- Chrystelle Oliveira (Saint Prix, FR)
- Christophe Simon (Chevilly Larue, FR)
Cpc classification
C07D495/22
CHEMISTRY; METALLURGY
C07D471/22
CHEMISTRY; METALLURGY
International classification
C07D495/22
CHEMISTRY; METALLURGY
C07D471/22
CHEMISTRY; METALLURGY
Abstract
The present invention relates to compounds of formula (I) and their use for treating or preventing a bacterial infection or as an antibacterial agent and/or as a β-lactamase inhibitor. ##STR00001##
Claims
1. A compound of formula (I) ##STR00033## wherein: W represents an 8- to 10-membered aromatic or partially unsaturated bicycle optionally having one or more heteroatoms chosen independently from the group consisting of O, N, N(T.sup.2), S and/or optionally substituted by one or more T.sup.1; R.sup.1 is chosen from the group consisting of H, (CH.sub.2).sub.mCN, (CH.sub.2).sub.mC(═O)NR.sup.2R.sup.3, (CH.sub.2).sub.mC(═O)NR.sup.4NR.sup.2R.sup.3, (CH.sub.2).sub.mC(═O)NR.sup.2OR.sup.3, (CH.sub.2).sub.pOR.sup.2, (CH.sub.2).sub.pNR.sup.2R.sup.3, (CH.sub.2).sub.pNR.sup.4C(═NR.sup.4)N(R.sup.4).sub.2, (CH.sub.2).sub.mC(═NOZ.sup.4)NZ.sup.1Z.sup.2 and (CH.sub.2).sub.p-(5 to 6-membered heteroaryl 1 or 4 heteroatoms independently chosen from the group consisting of N, O or S); m is an integer from 0 to 6; p is an integer from 1 to 6; R.sup.2 and R.sup.3, identical or different, are chosen from the group consisting of H, linear or branched (C1-C6)alkyl, (C3-C11)cycloalkyl, (C6-C10)aryl, 4- to 6-membered heterocyclyl having 1 to 2 heteroatoms chosen independently from the group consisting of N, O or S, 5- to 10-membered heteroaryl having 1 to 4 heteroatoms chosen independently from the group consisting of N, O or S, C(═O)(linear or branched C1-C6)alkyl, C(═O)(4 to 6-membered heterocyclyl having 1 to 2 heteroatoms chosen independently from the group consisting of N, O or S) or form together with the nitrogen atom to which they are linked a 4- to 6-membered heterocyclyl having 1 to 2 heteroatoms chosen independently from the group consisting of N, O or S, wherein alkyl, cycloalkyl, aryl, heterocyclyl and heteroaryl are optionally substituted by one or more R.sup.5; R.sup.4, each identical or different, is independently chosen from the group consisting of H and linear or branched (C1-C6)alkyl, wherein alkyl is optionally substituted by one or more R.sup.5; R.sup.5, each identical or different, is chosen from the group consisting of OH, O-(linear or branched-C1-C6)alkyl, NH.sub.2, NH(linear or branched C1-C6)alkyl, N[(linear or branched C1-C6)Alkyl].sub.2, C(═O)NH.sub.2, C(═O)NH (linear or branched C1-C6)alkyl and C(═ON[linear or branched(C1-C6)alkyl].sub.2; Y.sup.1 is chosen from the group consisting of SO.sub.3H, CHFC(═O)Y.sup.2, CF.sub.2C(═O)Y.sup.2, and SO.sub.3(C1-C6)alkyl-C(═O)O(C1-C6)alkyl; Y.sup.2 is chosen from the group consisting of OH, O(C1-C6)alkyl, linear or branched, O(C3-C11)cycloalkyl, O-(4 to 6-membered heterocyclyl having 1 or 2 heteroatoms chosen independently from the group consisting of N, O and S), and NY.sup.3Y.sup.4, wherein alkyl, cycloalkyl and heterocyclyl are optionally substituted by one or more Y.sup.5; Y.sup.3 and Y.sup.4, each identical or different, is chosen from the group consisting of linear or branched (C1-C6)alkyl, linear or branched O(C1-C6)alkyl, (C3-C11)cycloalkyl, 4 to 6-membered heterocyclyl having 1 or 2 heteroatoms chosen independently from the group consisting of N, O or S, or form together with the nitrogen atom to which they are linked a 4- to 6-membered heterocyclyl having 1 or 2 heteroatoms chosen independently from the group consisting of N, O or S, wherein alkyl, cycloalkyl and heterocyclyl is optionally substituted by one or more Y.sup.5; and Y.sup.5, each identical or different, is chosen from the group consisting of linear or branched (C1-C6)alkyl, (C3-C6)cycloalkyl, linear or branched O(C1-C6)alkyl, linear or branched O(C1-C6)alkyl-O(C1-C6)alkyl, linear or branched (C1-C6)alkyl-O(C1-C6)alkyl; and O(C3-C6)cycloalkyl; T.sup.1 is chosen from the group consisting of halogen, (CH.sub.2).sub.m-CN, (CH.sub.2).sub.m-OX.sup.1, (CH.sub.2).sub.m-C(═O)NX.sup.1X.sup.2, (CH.sub.2).sub.m-C(═O)NX.sup.1OX.sup.2, (CH.sub.2).sub.m-C(═O)NX.sup.1NX.sup.2X.sup.3, (CH.sub.2).sub.m-C(═NOX.sup.1)X.sup.2, (CH.sub.2).sub.m-C(═NX.sup.1)NX.sup.2X.sup.3, (CH.sub.2).sub.m-NX.sup.1X.sup.2, (CH.sub.2).sub.m-NX.sup.1C(═O)X.sup.2, (CH.sub.2).sub.m-NX.sup.1C(═O)NX.sup.2X.sup.3, (CH.sub.2).sub.m-NX.sup.1S(═O).sub.2NX.sup.2X.sup.3, (CH.sub.2).sub.m-NX.sup.1S(═O).sub.2X.sup.2, (CH.sub.2).sub.m-NX.sup.1C(═NX.sup.2)NHX.sup.3, (CH.sub.2).sub.m-NX.sup.1C(═NX.sup.2)X.sup.2, (CH.sub.2).sub.m-S(═O).sub.2NX.sup.1X.sup.2, linear or branched (C1-C6)alkyl, (C3-C6)cycloalkyl, (CH.sub.2).sub.m-aryl, (CH.sub.2).sub.m-(5- to 6-membered heteroaryl having from 1 to 4 heteroatoms chosen independently from the group consisting of N, O or S), and (CH.sub.2).sub.m-(4- to 6-membered heterocyclyl having from 1 to 2 heteroatoms chosen independently from the group consisting of N, O or S), wherein alkyl, cycloalkyl, aryl, heteroaryl and heterocyclyl are optionally substituted by one or more X.sup.4; T.sup.2 is chosen from the group consisting of H, (CH.sub.2).sub.n-CN, (CH.sub.2).sub.n-OX.sup.1, (CH.sub.2).sub.m-C(═O)X.sup.1, (CH.sub.2).sub.m-C(═O)NX.sup.1X.sup.2, (CH.sub.2).sub.m-C(═O)NX.sup.1OX.sup.2, (CH.sub.2).sub.m-C(═O)NX.sup.1NX.sup.2X.sup.3, (CH.sub.2).sub.m-C(═NOX.sup.1)X.sup.2, (CH.sub.2).sub.m-C(═NX.sup.1)NX.sup.2X.sup.3, (CH.sub.2).sub.n-NX.sup.1X.sup.2, (CH.sub.2).sub.n-NX.sup.1C(═O)X.sup.2, (CH.sub.2).sub.n-NX.sup.1C(═O)NX.sup.2X.sup.3, (CH.sub.2).sub.n-NX.sup.1S(═O).sub.2NX.sup.2X.sup.3, (CH.sub.2).sub.n-NX.sup.1S(═O).sub.2X.sup.2, (CH.sub.2).sub.n-NX.sup.1C(═NX.sup.2)NHX.sup.3, (CH.sub.2).sub.n-NX.sup.1C(═NX.sup.2)X.sup.2, (CH.sub.2).sub.m-S(═O).sub.2NX.sup.2X.sup.3, linear or branched (C1-C6)alkyl, (C3-C6)cycloalkyl, (CH.sub.2).sub.m-aryl, (CH.sub.2).sub.m-(5- to 6-membered heteroaryl having from 1 to 4 heteroatoms chosen independently from the group consisting of N, O or S), and (CH.sub.2).sub.m-(4- to 6-membered heterocyclyl having 1 to 2 heteroatoms chosen independently from the group consisting of N, O or S), wherein alkyl, cycloalkyl, aryl, heteroaryl and heterocyclyl are optionally substituted by one or more X.sup.4; X.sup.1, X.sup.2 and X.sup.3, each identical or different, are chosen from the group consisting of H, linear or branched (C1-C6)alkyl, (C3-C6)cycloalkyl, (C2-C6)alkyl-NZ.sup.1Z.sup.2, (C2-C6)alkyl-NHC(═NZ.sup.1)NHZ.sup.2, (C2-C6)alkyl-NHC(═NZ.sup.1)Z.sup.2, (C2-C6)alkyl-NZ.sup.1C(═O)Z.sup.2, (C2-C6)alkyl-OZ.sup.1, (C1-C6)alkyl-C(═NZ.sup.1)NHZ.sup.2, (C1-C6)alkyl-CONZ.sup.1Z.sup.2, (C1-C6)alkyl-COOZ.sup.1, (CH.sub.2).sub.m-aryl, (CH.sub.2).sub.m-(5- to 6-membered heteroaryl having from 1 to 4 heteroatoms chosen independently from the group consisting of N, O and S), and (CH.sub.2).sub.m-(4- to 6-membered heterocyclyl having from 1 to 2 heteroatoms chosen independently from the group consisting of N, O and S), or form together with the nitrogen atom to which they are linked a 4- to 6-membered heterocyclyl having 1 or 2 heteroatoms chosen independently from the group consisting of N, O or S, wherein alkyl, cycloalkyl, aryl, heteroaryl and heterocyclyl are optionally substituted by one or more Z.sup.3; X.sup.4, each identical or different, is chosen from the group consisting of H, halogen, linear or branched (C1-C6)alkyl, (C3-C6)cycloalkyl, (CH.sub.2).sub.m-NZ.sup.1Z.sup.2, (CH.sub.2).sub.m-NHC(═NZ.sup.1)NHZ.sup.2, (CH.sub.2).sub.m-NHC(═NZ.sup.1)H, (CH.sub.2).sub.m-NZ.sup.1C(═O)Z.sup.2, (CH.sub.2).sub.m-OZ.sup.1, (CH.sub.2).sub.m-C(═NZ.sup.1)NHZ.sup.2, (CH.sub.2).sub.m-CONZ.sup.1Z.sup.2, (CH.sub.2).sub.m-COOZ.sup.1, (CH.sub.2).sub.m-aryl, (CH.sub.2).sub.m-(5- to 6-membered heteroaryl having from 1 to 4 heteroatoms chosen independently from the group consisting of N, O and S), and (CH.sub.2).sub.m-(4- to 6-membered heterocyclyl having from 1 to 2 heteroatoms chosen independently from the group consisting of N, O and S), wherein alkyl, cycloalkyl, aryl, heteroaryl and heterocyclyl are optionally substituted by one or more Z.sup.3; Z.sup.1 and Z.sup.2, identical or different, are chosen from the group consisting of H, linear or branched (C1-C6)alkyl, (C3-C6)cycloalkyl, (C2-C6)alkyl-N(Z.sup.4).sub.2, (C2-C6)alkyl-NZ.sup.4C(═O)Z.sup.4, (C2-C6)alkyl-OZ.sup.4, (C1-C6)alkyl-C(═NH)NHZ.sup.4, (C1-C6)alkyl-CON(Z.sup.4).sub.2, and (C1-C6)alkyl-COOZ.sup.4; Z.sup.3, each identical or different, is chosen from the group consisting of H, halogen, linear or branched (C1-C6)alkyl, (C3-C6)cycloalkyl, (CH.sub.2).sub.m-N(Z.sup.4).sub.2, (CH.sub.2).sub.m-OZ.sup.4C(═O)Z.sup.4, (CH.sub.2).sub.m-OZ.sup.4, (CH.sub.2).sub.m-CON(Z.sup.4).sub.2, and (CH.sub.2).sub.m-COOZ.sup.4; Z.sup.4, each identical or different, is chosen from the group consisting of H, linear or branched (C1-C6)alkyl, and (C3-C6)cycloalkyl, n is an integer from 2 to 6; any sulphur atom present within a heterocycle can be oxidized to form a S═O group or a S(O).sub.2 group; any nitrogen atom present within a heterocycle or present within group wherein it is trisubstituted thus forming a tertiary amino group, can be further quaternized by a methyl group; or a racemate, an enantiomer, a diastereoisomer, or a pharmaceutically acceptable salt thereof.
2. The compound according to claim 1, wherein: R.sup.1 is chosen from the group consisting of H, (CH.sub.2).sub.mCN, (CH.sub.2).sub.mC(═O)NR.sup.2R.sup.3, (CH.sub.2).sub.mC(═O)NR.sup.4NR.sup.2R.sup.3, (CH.sub.2).sub.mC(═O)NR.sup.2OR.sup.3, (CH.sub.2).sub.pOR.sup.2, (CH.sub.2).sub.pNR.sup.2R.sup.3, (CH.sub.2).sub.pNR.sup.4C(═NR.sup.4)N(R.sup.4).sub.2, and (CH.sub.2).sub.p-(5 to 6-membered heteroaryl having 1 or 4 heteroatoms independently chosen from the group consisting of N, O or S); and/or Y.sup.1 is chosen from the group consisting of SO.sub.3H, CHFC(═O)Y.sup.2, and CF.sub.2C(═O)Y.sup.2.
3. The compound according to claim 1, wherein the compound is of formula (IA) ##STR00034## wherein: W1 is a 5- to 6-membered ring, aromatic or partially unsaturated, optionally substituted by one or more T.sup.1 and having optionally one or more heteroatoms independently selected from the group consisting of O, N, N(T.sup.2) and S; and W2 is a 5- to 6-membered ring, aromatic or partially unsaturated, optionally substituted by one or more T.sup.1 and having optionally one or more heteroatoms independently selected from the group consisting of O, N, N(T.sup.2) and S.
4. The compound according to claim 3, wherein: W1 represents a 5-membered ring, aromatic or partially unsaturated, optionally substituted by one or more T.sup.1 and having 1 or 2 heteroatoms independently selected from the group consisting of O, N, N(T.sup.2), S; and W2 represents a 5- to 6-membered ring, aromatic or partially unsaturated, optionally substituted by one or more T.sup.1 and having optionally one or more heteroatoms independently selected from the group consisting of O, N, N(T.sup.2) and S; or W1 represents a 6-membered ring, aromatic or partially unsaturated, optionally substituted by one or more T.sup.1 and having optionally 1 or 2 heteroatoms independently selected from the group consisting of O, N, N(T.sup.2) and S; and W2 represents a 5- to 6-membered ring, aromatic or partially unsaturated, optionally substituted by one or more T.sup.1 and having optionally one or more heteroatoms independently selected from the group consisting of O, N, N(T.sup.2) and S; or W1 represents a thiazole, thiophene, pyrrole, pyrrole for which one N atom is substituted by T.sup.2 or imidazole, optionally substituted by one or more T.sup.1; and W2 represents a 5- to 6-membered ring, aromatic or partially unsaturated, optionally substituted by one or more T.sup.1 and having optionally one or more heteroatoms independently selected from the group consisting of O, N, N(T.sup.2) and S; or W1 represents a phenyl, pyridine, pyrazine or thiazine, optionally substituted by one or more T.sup.1; and W2 represents a 5- to 6-membered ring, aromatic or partially unsaturated, optionally substituted by one or more T.sup.1 and having optionally one or more heteroatoms independently selected from the group consisting of O, N, N(T.sup.2) and S; or W1 represents a thiazole, thiophene, pyrrole, pyrrole for which one N atom is substituted by T.sup.2 or imidazole, optionally substituted by one T.sup.1; and W2 represents a imidazole, imidazole for which one N atom is substituted by T.sup.2, triazole, triazole for which one N atom is substituted by T.sup.2, pyrrole, pyrrole for which one N atom is substituted by T.sup.2, pyrazole, pyrazole for which one N atom is substituted by T.sup.2, dihydropyrrole, dihydropyrrole for which one N atom is substituted by T.sup.2, and thiazole optionally substituted by one or more T.sup.1; or W1 represents a thiazole, thiophene, pyrrole, pyrrole for which one N atom is substituted by T.sup.2 or imidazole, optionally substituted by one T.sup.1; and W2 represents a phenyl, pyridine, pyridazine, pyrimidine, pyrazine or tetrahydropyridine for which one N atom is substituted by T.sup.2, optionally substituted by one T.sup.1; or W1 represents a phenyl, pyridine, pyrazine or thiazine, optionally substituted by one or more T.sup.1; and W2 represents a imidazole, imidazole for which one N atom is substituted by T.sup.2, triazole, triazole for which one N atom is substituted by T.sup.2, pyrrole, pyrrole for which one N atom is substituted by T.sup.2, pyrazole, pyrazole for which one N atom is substituted by T.sup.2, dihydropyrrole, dihydropyrrole for which one N atom is substituted by T.sup.2, and thiazole optionally substituted by one or more T.sup.1; or W1 represents a phenyl, pyridine, pyrazine or thiazine, optionally substituted by one or more T.sup.1; and W2 represents a phenyl, pyridine, pyridazine, pyrimidine, pyrazine or tetrahydropyridine for which one N atom is substituted by T.sup.2, optionally substituted by one T.sup.1.
5. The compound according to claim 1, chosen among the following: ##STR00035## wherein: W2a, W2b and W2c are independently chosen among 5- to 6-membered ring, aromatic or partially unsaturated, optionally substituted by one or more T.sup.1 and having optionally one or more heteroatoms independently selected from the group consisting of O, N, N(T.sup.2) and S; and X is chosen from C or N.
6. The compound according to claim 5, wherein: W2a is chosen from the group consisting of phenyl or pyridinyl; and W2b and W2c are chosen in from the group consisting of phenyl and 5 to 6-membered heterocycle, aromatic or partially unsaturated, optionally substituted by one or more T.sup.1, having 1 to 3 heteroatoms independently chosen from the group consisting of N(T.sup.2), N or S.
7. The compound according to claim 5, chosen among the following: ##STR00036##
8. The compound according to claim 1, wherein: R.sup.1 is chosen from the group consisting of H, CN, C(═O)NR.sup.2R.sup.3, C(═O)NHNHR.sup.2, C(═O)NHOR.sup.2, (CH.sub.2)OR.sup.2, (CH.sub.2)NHR.sup.2, (CH.sub.2)NR.sup.4C(═NR.sup.4)N(R.sup.4).sub.2, C(═NOZ.sup.4)NZ.sup.1Z.sup.2, and (CH.sub.2)(5- to 6-membered heteroaryl having 1 to 4 heteroatoms independently chosen from the group consisting of N, O or S); R.sup.2 and R.sup.3, identical or different, are chosen from the group consisting of H, linear or branched (C1-C6)alkyl, (4- to 6-membered heterocyclyl having 1 or 2 heteroatoms independently chosen from the group consisting of N, O or S), and C(═O)(4- to 6-membered heterocyclyl comprising having 1 or 2 heteroatoms independently chosen from the group consisting of N, O or S); R.sup.4, each identical or different, is independently chosen from the group consisting of H and linear or branched (C1-C6)alkyl, wherein alkyl is optionally substituted by one or more R.sup.5; R.sup.5, each identical or different, is chosen from the group consisting of OH, O(linear or branched-C1-C6)alkyl, NH.sub.2, NH(linear or branched C1-C6)alkyl, N[(linear or branched C1-C6)Alkyl].sub.2, C(═O)NH.sub.2, C(═O)NH(linear or branched C1-C6)alkyl, C(═O)N[linear or branched (C1-C6)alkyl].sub.2; Y.sup.2 is chosen in from the group consisting of OH, O(C1-C6)alkyl, linear or branched, O-(4- to 6-membered heterocyclyl having 1 or 2 heteroatoms chosen from the group consisting of N, O and S, wherein alkyl and heterocyclyl are optionally substituted by one or more Y.sup.5; and Y.sup.5, each identical or different, is chosen from the group consisting of linear or branched (C1-C6)alkyl, linear or branched O(C1-C6)alkyl, linear or branched O(C1-C6)alkyl-O(C1-C6)alkyl, linear or branched (C1-C6)alkyl-O(C1-C6)alkyl.
9. The compound according to claim 1, wherein: R.sup.1 is chosen from the group consisting of H, CN, C(═O)NR.sup.2R.sup.3, C(═O)NHNHR.sup.2, C(═O)NHOR.sup.2, (CH.sub.2)OR.sup.2, (CH.sub.2)NHR.sup.2, (CH.sub.2)NR.sup.4C(═NR.sup.4)N(R.sup.4).sub.2, and (CH.sub.2)(5- to 6-membered heteroaryl having 1 to 4 heteroatoms independently chosen from the group consisting of N, O or S); R.sup.2 and R.sup.3, identical or different, are chosen from the group consisting of H, linear or branched (C1-C6)alkyl, (4- to 6-membered heterocyclyl having 1 or 2 heteroatoms independently chosen from the group consisting of N, O or S), and C(═O)(4- to 6-membered heterocyclyl having 1 or 2 heteroatoms independently chosen from the group consisting of N, O or S); R.sup.4, each identical or different, is independently chosen from the group consisting of H and linear or branched (C1-C6)alkyl, wherein alkyl is optionally substituted by one or more R.sup.5; R.sup.5, each identical or different, is chosen from the group consisting of OH, O(linear or branched-C1-C6)alkyl, NH.sub.2, NH(linear or branched C1-C6)alkyl, N[(linear or branched C1-C6)Alkyl].sub.2, C(═O)NH.sub.2, C(═O)NH(linear or branched C1-C6)alkyl, C(═O)N[linear or branched (C1-C6)alkyl].sub.2; Y.sup.2 is chosen in from the group consisting of OH, O(C1-C6)alkyl, linear or branched, O-(4- to 6-membered heterocyclyl having 1 or 2 heteroatoms chosen from the group consisting of N, O and S, wherein alkyl and heterocyclyl are optionally substituted by one or more Y.sup.5; and Y.sup.5, each identical or different, is chosen from the group consisting of linear or branched (C1-C6)alkyl, linear or branched O(C1-C6)alkyl, linear or branched O(C1-C6)alkyl-O(C1-C6)alkyl, linear or branched (C1-C6)alkyl-O(C1-C6)alkyl.
10. A pharmaceutical composition comprising a compound according to claim 1 and a pharmaceutically acceptable excipient.
11. The pharmaceutical composition according to claim 10 further comprising an antibacterial compound.
12. The pharmaceutical composition according to claim 11, wherein the antibacterial compound is selected from aminoglycosides, β-lactams, glycylcyclines, tetracyclines, quinolones, fluoroquinolones, glycopeptides, lipopeptides, macrolides, ketolides, lincosamides, streptogramins, oxazolidinones, polymyxins and mixtures thereof.
13. The pharmaceutical composition according to claim 10 further comprising a β-lactam compound.
14. The pharmaceutical composition according to claim 13, wherein the β-lactam compound is selected from penicillin, cephalosporins, penems, carbapenems, monobactam, and combinations thereof.
15. A pharmaceutical composition comprising at least a compound according to claim 1 and ceftazidime.
16. A kit comprising: a first pharmaceutical composition that comprises a first pharmaceutically active compound and a first pharmaceutically acceptable excipient; and a second pharmaceutical composition that comprises a second pharmaceutically active compound and second pharmaceutically acceptable excipient; wherein the first and second pharmaceutically active compounds are different compounds according to claim 1.
17. A kit comprising: a first pharmaceutical composition comprising at least a compound according to claim 1; and a second pharmaceutical composition comprising ceftazidime.
18. A method for treating a bacterial infection caused by bacteria producing one or more β-lactamase, the method comprising the administration to a patient in need thereof an effective amount of a compound according to claim 1.
19. The method according to claim 18, wherein the bacteria is a gram-positive bacteria or gram-negative bacteria.
20. The method according to claim 18, wherein the bacteria is a gram-negative bacteria.
21. A method for treating a bacterial infection, the method comprising the simultaneous, separate or sequential administration to a patient in need thereof an effective amount of the first and second pharmaceutically acceptable active compounds of a kit according to claim 16.
22. The compound according to claim 1, chosen among the following: ##STR00037## wherein: W1 is a 5- to 6-membered ring, aromatic or partially unsaturated, optionally substituted by one or more T.sup.1 and having optionally one or more heteroatoms independently selected from the group consisting of O, N, N(T.sup.2), and S; and W2 is a 5- to 6-membered ring, aromatic or partially unsaturated, optionally substituted by one or more T.sup.1 and having optionally one or more heteroatoms independently selected from the group consisting of O, N, N(T.sup.2), and S.
Description
EXAMPLES
(1) The following examples are provided for the purpose of illustrating the present invention and by no means should be interpreted to limit the scope of the present invention.
(2) Preparation of the compounds and biological activity: Abbreviations or symbols used herein include: ACHN: 1,1′-azobis(cyclohexanecarbonitrile) ACN: acetonitrile AcOH: acetic acid Bn: benzyl Boc: tert-butoxycarbonyl Boc.sub.2O: tert-butoxycarbonyl anhydride BocON: [2-(tert-butoxycarbonyloxyimino)-2-phenylacetonitrile] bs: broad singlet Burgess reagent: methyl N-(triethylammoniosulfonyl)carbamate CDI: 1,1′-Carbonyldiimidazole CFU: colony-forming units CLSI: clinical laboratory standards institute d: doublet DBU: 1,8-diazabicyclo[5.4.0]undec-7-ene DCM: dichloromethane dd: double doublet ddd: double double doublet ddt: double double triplet dq: double quartet dt: double triplet DTAD: di-tert-butylazodicarboxylate DEAD: diethyl azodicarboxylate Dess-Martin periodinane: 1,1,1-tris(acetyloxy)-1,1-dihydro-1,2-benziodoxol-3-(1H)-one DHP 3,4-dihydro-2H-pyran DIAD: diisopropyl azodicarboxylate DIPEA: N,N-diisopropylethylamine DMAP: 4-dimethylaminopyridine DMF: N,N-dimethylformamide DMSO: dimethylsulfoxide EtOAc: ethyl acetate Et.sub.2O: diethyl ether Fmoc-Cl: 9-Fluorenylmethyl chloroformate h: hours HATU: 1-[Bis(dimethylamino)methylene]-1H-1,2,3-triazolo[4,5-b]pyridinium-3-oxid hexafluorophosphate m: multiplet min: minutes MeOH: methanol MeONa: sodium methoxide MIC: minimum inhibitory concentration MS: mass spectrometry MsCl: methanesulfonyl chloride MTBE: tert-butyl methyl ether NBS: N-bromosuccinimide NMR: nuclear magnetic resonance spectroscopy Ns: nosyl, nitrobenzenesulfonyl Pd(Ph.sub.3).sub.4: tetrakis(triphenylphosphine)palladium(0) PG: protective group PhSH: thiophenol PMe.sub.3: trimethylphosphine PPh.sub.3: triphenylphosphine Ppm: parts per million q: quartet rt: room temperature s: singlet SEM: [2-(trimethylsilyl)ethoxy]methyl t: triplet TBAF: tetra-n-butylammonium fluoride TBDMSCl: tert-butyldimethylsilyl chloride TBDMSOTf: trifluoromethanesulfonic acid tert-butyldimethylsilyl ester TBSOTf: trimethylsilyl trifluoromethanesulfonate tBuOK: potassium tert-butoxide TEA: triethylamine TFA: trifluoroacetic acid THF: tetrahydrofuran THP: tetrahydropyranyl TLC: thin layer chromatography TMSI: Iodotrimethylsilane
Example 1: Synthesis of Sodium (2,5-methano-3-oxo-1,5-dihydrobenzothiopheno[2,3-e][1,3]diazepin-4-yl) sulfate
(3) ##STR00020##
Step 1: Preparation of Intermediate 1-(benzothiophen-2-yl)-2-bromo-ethanone (1a)
(4) 1-(benzothiophen-2-yl)ethanone (5 g, 28.36 mmol) was solubilized into a mixture of CHCl.sub.3/MeOH (5/1) under argon atmosphere. Copper(II) bromide (12.7 g, 56.74 mmol), was added and the mixture was warmed at 60° C. for 16 h. The reaction was evaporated and the residue was triturated with MTBE and pentane. The precipitate was filtrated to give intermediate (1a) (6.9 g, 27.04 mmol, 95%) slightly contaminated by di-brominated byproduct. MS m/z ([M+H].sup.+) 255 (bromine isotopy).
Step 2: Preparation of Intermediate 2-azido-1-(benzothiophen-2-yl)ethanone (1b)
(5) Intermediate (1a) (6.9 g, 27.04 mmol) was solubilized into acetone (200 mL). Sodium azide (1.76 g, 27.04 mmol) was added and the mixture was stirred for 16 h at rt. The precipitate was filtered on Celite and the filtrate was evaporated. The residue was triturated successively with MTBE and pentane and then filtered to give intermediate (1b) (4.04 g, 18.42 mmol, 68%). MS m/z ([M+Na].sup.+) 240. .sup.1H NMR (400 MHz, CDCl.sub.3): δ (ppm) 4.57 (s, 2H), 7.42-7.47 (m, 1H), 7.49-7.54 (m, 1H), 7.88-7.94 (m, 2H), 7.98 (s, 1H).
Step 3: Preparation of Intermediate 2-amino-1-(benzothiophen-2-yl)ethanol (1c)
(6) To a solution of intermediate (1 b) (1.92 g, 8.76 mmol) in THF (78 mL) under inert atmosphere at 0° C. was dropped a solution of LAH (2N in THF) (9.6 mL, 19.2 mmol). After stirring for 1 h, the mixture was quenched with ice and HCl 1N. The colored impurities were extracted with DCM. Then the aqueous layer was neutralized with NaOH 1N and the product was extracted with EtOAc. The organic layer was dried over Na.sub.2SO.sub.4, filtered and concentrated. The residue was triturated with Et.sub.2O to give intermediate (1c) as an off-white solid (1.40 g, 7.24 mmol, 82%). MS m/z ([M-OH].sup.+) 176. .sup.1H NMR (400 MHz, CDCl.sub.3: δ (ppm) 2.02 (bs, 2H), 3.03 (dd, J=6.7, 12.8 Hz, 1H), 3.14 (dd, J=4.1, 12.8 Hz, 1H), 4.92-4.99 (m, 1H), 7.21 (s, 1H), 7.29-7.38 (m, 2H), 7.73 (d, J=7.3 Hz, 1H), 7.83 (d, J=7.3 Hz, 1H).
Step 4: Preparation of Intermediate 1,2,3,4-tetrahydrobenzothiopheno[3,2-c]pyridin-4-ol hydrochloride (1d)
(7) To a solution of intermediate (1c) (0.250 g, 1.29 mmol) in EtOH (20 mL) were added formaldehyde (37% wt solution in water, 0.145 mL, 1.94 mmol) and HCl 4N/dioxane (0.323 mL, 1.29 mmol). After stirring for 16 h at 80° C., conversion was not completed. So, formaldehyde 37% wt solution in water (0.250 mL, 3.36 mmol) was added and the mixture was heated at 80° C. for 16 h (operation was repeated until complete conversion). The mixture was cold and dried under nitrogen flux. The residue was triturated with Et.sub.2O to give intermediate (1d) (0.312 g, 1.29 mmol, quantitative). MS m/z ([M+H].sup.+) 206.
Step 5: Preparation of Intermediate tert-butyl 4-hydroxy-3,4-dihydro-1H-benzothiopheno[3,2-c]pyridine-2-carboxylate (1e)
(8) To a solution of intermediate (1d) (0.312 g, 1.29 mmol) in DCM (13 mL) under argon atmosphere at 0° C. were successively added Boc.sub.2O (0.310 mg, 1.42 mmol) and TEA (0.450 mL, 3.23 mmol). After stirring for 18 h at rt, the mixture was evaporated under nitrogen flux and the residue was purified by column chromatography on silica gel (DCM/acetone 10/0 to 9/1) to give intermediate (1e) as a white solid (0.357 g, 1.17 mmol, 90%). MS m/z ([M+Na].sup.+) 328. .sup.1H NMR (400 MHz, CDCl.sub.3): δ (ppm) 1.52 (s, 9H), 3.81 (d, J=13.6 Hz, 1H), 3.99 (dd, J=4.8, 13.6 Hz, 1H), 4.53 (d, J=16.7 Hz, 1H), 4.84-4.93 (m, 2H), 7.35-7.41 (m, 2H), 7.59-7.64 (m, 1H), 7.83-7.85 (m, 1H).
Step 6: Preparation of Intermediate tert-butyl 4-(allyloxyamino)-3,4-dihydro-1H-benzothiopheno[3,2-c]pyridine-2-carboxylate (1f)
(9) To a solution of intermediate (1e) (0.355 g, 1.16 mmol) in DCM (13 mL) under argon atmosphere at −78° C. were successively added TEA (0.650 mL, 4.64 mmol) and methanesulfonic anhydride (0.606 g, 3.48 mmol). After stirring for 1 h at −78° C., 0-allylhydroxylamine 60% (0.990 g, 8.14 mmol) in DCM (3.5 mL) was dropped into the mixture which was stirred for 20 min at −78° C. The temperature was then raised at rt for 2 h30. The mixture was quenched with saturated aqueous NaHCO.sub.3 and extracted with DCM. The organic extract was dried over Na.sub.2SO.sub.4, filtered and concentrated. The residue was purified by column chromatography on silica gel (cyclohexane/EtOAc 100/0 to 95/5) to give intermediate (1f) (0.338 g, 0.938 mmol, 80%). MS m/z ([M+H].sup.+) 361. .sup.1H NMR (300 MHz, CDCl.sub.3): δ (ppm) 1.52 (s, 9H), 3.60-3.70 (m, 1H), 4.20-4.50 (m, 5H), 4.80-4.95 (m, 1H), 5.22 (d, J=10.4 Hz, 1H), 5.32 (d, J=16.3 Hz, 1H), 5.97 (ddt, J=5.9, 10.4, 16.3 Hz, 1H), 7.31-7.40 (m, 2H), 7.59-7.61 (m, 1H), 7.80-7.82 (m, 1H).
Step 7: Preparation of Intermediate tert-butyl 4-[allyloxy(chlorocarbonyl)amino]-3,4-dihydro-1H-benzothiopheno[3,2-c]pyridine-2-carboxylate (1q)
(10) To a solution of intermediate (1f) (0.338 g, 0.94 mmol) in ACN (17 mL) under argon atmosphere at −10° C. were successively added TEA (0.393 mL, 2.82 mmol) and a solution of diphosgene (0.147 mL, 1.22 mmol) in ACN (2.3 mL). After stirring for 30 min at −10° C., the mixture was evaporated under nitrogen flux. The residue was triturated with DCM and evaporated under nitrogen flux again. The residue was triturated with Et.sub.2O and the insoluble was filtered off. The filtrate was concentrated to give intermediate (1 g) (0.280 g, 0.662 mmol, 70%) which was used in the next step without further purification. MS m/z ([M+Na].sup.+) 445.
Step 8: Preparation of Intermediate 4-allyloxy-2,5-methano-1,5-dihydrobenzothiopheno[2,3-e][1,3]diazepin-3-one (1 h)
(11) To a solution of intermediate (1 g) (0.280 g, 0.662 mmol) in DCM (2.2 mL) under argon atmosphere at 0° C. was dropped HCl 4N in dioxane (6.6 mL, 40 mmol). After stirring for 1 h at rt, the mixture was evaporated under nitrogen flux. The residue was triturated with Et.sub.2O to give intermediate (1 h) as a brown oil (0.040 g, 0.140 mmol, 21%). MS m/z ([M+H].sup.+) 287.
Step 9: Preparation of Intermediate 4-hydroxy-2,5-methano-1,5-dihydrobenzothiopheno[2,3-e][1,3]diazepin-3-one (1i)
(12) A solution of intermediate (1 h) (0.040 g, 0.140 mmol) in DCM (2 mL) was degassed 10 min under argon atmosphere. AcOH (0.016 mL, 0.278 mmol) and Pd(PPh.sub.3).sub.4 (0.081 g, 0.07 mmol) were successively added. After stirring for 30 min at rt, the mixture was evaporated under nitrogen flux. The residue was purified by column chromatography on silica gel (DCM/acetone 10/0 to 8/2). The fractions of interest are combined and concentrated. The residue was triturated with Et.sub.2O to give intermediate (1i) as a yellow solid (0.013 g, 0.053 mmol, 38%). MS m/z ([M+H].sup.+) 247. .sup.1H NMR (400 MHz, acetone-d.sub.6): δ (ppm) 3.32 (d, J=10.9 Hz, 1H), 3.78 (dd, J=2.8, 10.9 Hz, 1H), 4.39 (d, J=16.6 Hz, 1H), 4.50 (d, J=2.8 Hz, 1H), 4.63 (d, J=16.6 Hz, 1H), 7.41-7.31 (m, 2H), 7.62-7.70 (m, 1H), 7.79-7.81 (m, 1H).
Step 10: Preparation of Sodium (2,5-methano-3-oxo-1,5-dihydrobenzothiopheno[2,3-e][1,3]diazepin-4-yl) sulfate, Example 1
(13) To a solution of intermediate (1i) (0.013 g, 0.053 mmol) in anhydrous pyridine (0.6 mL) under inert atmosphere was added sulfur trioxide pyridine complex (0.050 g, 0.317 mmol). After stirring for 18 h, the heterogeneous mixture was concentrated in vacuum. DCM was added and the insoluble was filtered off. The filtrate concentrated. The residue was purified by flash chromatography on silica gel (DCM/acetone 10/0 to 0/10) to give Example 1 as pyridinium salt (6 mg, 0.018 mmol). The pyridinium salt was applied on a Dowex sodium form column (Dowex® 50WX8 hydrogen form stored with an aqueous solution of 2N NaOH and washed until neutral pH with water). The fractions containing the desired compound were combined, frozen and lyophilized. The solid was triturated with DCM to give Example 1 (2.7 g, 0.0083 mmol, 15%). MS m/z ([M−H].sup.−) 325. .sup.1H NMR (400 MHz, D.sub.2O): δ (ppm) 3.70 (d, J=11.4 Hz, 1H), 4.0 (dd, J=3.0, 11.4 Hz, 1H), 4.68 (s, 2H), 5.11 (d, J=2.9 Hz, 1H), 7.44-7.52 (m, 2H), 7.68-7.74 (m, 1H), 7.97-8.03 (m, 1H).
Example 2: Synthesis of sodium trans-(1-ethoxycarbonyl-2,5-methano-3-oxo-1,5-dihydrobenzothiopheno[2,3-e][1,3]diazepin-4-yl) sulfate
(14) ##STR00021##
Step 1: Preparation of Intermediate ethyl 4-hydroxy-1,2,3,4-tetrahydrobenzothiopheno[3,2-c]pyridine-1-carboxylate Hydrochloride (2a)
(15) To a solution of intermediate (1c) (1 g, 5.17 mmol) into EtOH (51 mL) were added ethyl glyoxylate 50% solution in toluene (1.54 mL, 7.76 mmol) and HCl 4N in dioxane (1.30 mL, 5.17 mmol). After stirring for 16 h at 80° C., the mixture was cold and the precipitate was filtered to give intermediate (2a) as a white solid (1.19 g, 3.79 mmol, 73%). MS m/z ([M+H].sup.+) 278. .sup.1H NMR (300 MHz, D.sub.2O): δ (ppm) 1.23 (t, J=7.1 Hz, 3H), 3.64 (dd, J=9.0, 12.7 Hz, 1H), 3.95-4.03 (m, 1H), 4.24-4.42 (m, 2H), 5.40 (ddd, J=1.2, 6.0, 9.0 Hz, 1H), 5.84 (s, 1H), 7.51-7.60 (m, 2H), 8.02-8.04 (m, 2H).
Step 2: Preparation of Intermediate O2-tert-butyl O1-ethyl 4-hydroxy-3,4-dihydro-1H-benzothiopheno[3,2-c]pyridine-1,2-dicarboxylate (2b)
(16) To a solution of intermediate (2a) (1.13 g, 3.60 mmol) in DCM (36 mL) under argon atmosphere at 0° C. were successively added Boc.sub.2O (0.864 mg, 3.96 mmol) and TEA (1.25 mL, 9 mmol). After 16 h at rt, the mixture was washed with NaOH 1N and extracted with EtOAc. The organic layer was dried over Na.sub.2SO.sub.4, filtered and concentrated to give intermediate (2b) as a white solid (1.36 g, 3.60 mmol, quantitative, mixture of diastereoisomers). MS m/z ([M+Na].sup.+) 400. .sup.1H NMR (400 MHz, CDCl.sub.3): δ (ppm) 1.22 (q, J=6.9 Hz, 3H), 1.53 (s, 9H), 3.7-3.84 (m, 1H), 4.14-4.21 (m, 3H), 4.92-4.97 (m, 1H), 5.68 and 5.79 (2s, 1H), 7.35-7.45 (m, 2H), 7.84 (d, J=7.8 Hz, 1H), 8.00 (d, J=7.8 Hz, 1H).
Step 3: Preparation of Intermediate O2-tert-butyl O1-ethyl 4-(allyloxyamino)-3,4-dihydro-1H-benzothiopheno[3,2-c]pyridine-1,2-dicarboxylate (2c)
(17) To a solution of intermediate (2b) (800 mg, 2.12 mmol) in DCM (24 mL) under argon atmosphere at −78° C. were successively added TEA (1.19 mL, 8.48 mmol) and methanesulfonic anhydride (1.11 g, 6.36 mmol). After stirring for 1 h at −78° C., 0-allylhydroxylamine 60% (1.80 g, 14.84 mmol) in DCM (6.4 mL) was dropped. The mixture was stirred for 1 h20 at −78° C. then the temperature was raised at rt for 1 h30. The mixture was diluted with DCM and washed with saturated aqueous NaHCO.sub.3. The organic extract was dried over Na.sub.2SO.sub.4, filtered and concentrated. The residue was purified by column chromatography on silica gel (cyclohexane/EtOAc 10/0 to 95/5) to give intermediate (2c) as a colorless oil (0.762 g, 1.76 mmol, 83%). MS m/z ([M+H].sup.+) 433. .sup.1H NMR (400 MHz, CDCl.sub.3): δ (ppm) 1.22-1.28 (m, 3H), 1.50-1.54 (m, 9H), 3.68-4.76 (m, 7H), 5.21-5.43 (m, 2H), 5.69-6.00 (m, 2H), 7.32-7.44 (m, 2H), 7.78-7.83 (m, 1H), 8.01-8.08 (m, 1H).
Step 4: Preparation of Intermediate O2-tert-butyl O1-ethyl 4-[allyloxy(chlorocarbonyl)amino]-3,4-dihydro-1H-benzothiopheno[3,2-c]pyridine-1,2-dicarboxylate (2d)
(18) To a solution of intermediate (2c) (0.758 g, 1.75 mmol) in ACN (32 mL) under argon atmosphere at −10° C. were successively added TEA (0.732 mL, 5.25 mmol) and a solution of diphosgene (0.275 mL, 2.28 mmol) in ACN (4.3 mL). After stirring for 30 min at −10° C., the mixture was evaporated under nitrogen flux. The residue was successively triturated with DCM and Et.sub.2O. The residue was filtered off. The filtrate was evaporated to give intermediate (2d) which was used in the next step without further purification (0.866 g, 1.75 mmol, quantitative). MS m/z ([M+Na].sup.+) 417.
Step 5: Preparation of Intermediate ethyl trans-4-allyloxy-2,5-methano-3-oxo-1,5-dihydrobenzothiopheno[2,3-e][1,3]diazepine-1-carboxylate (2e)
(19) To a solution of intermediate (2d) (0.866 g, 1.75 mmol) in DCM (6 mL) under argon atmosphere at 0° C. was dropped HCl 4N in dioxane (17.5 mL, 70 mmol). After stirring for 1 h at 0° C. and for 1 h at rt, the mixture was evaporated under nitrogen flux. The residue was triturated with Et.sub.2O to intermediate (2e) as a yellow oil (0.627 g, 1.75 mmol, quantitative). MS m/z ([M+H].sup.+) 359. .sup.1H NMR (400 MHz, CDCl.sub.3): δ (ppm) 1.35 (t, 3H), 3.67-3.70 (m, 1H), 4.02 (d, J=11.4 Hz, 1H), 4.27-5.35 (m, 2H), 4.40-4.54 (m, 3H), 5.28-5.38 (m, 3H), 5.94-6.08 (m, 1H), 7.32-7.39 (m, 2H), 7.75-7.81 (m, 2H).
Step 6: Preparation of Intermediate ethyl trans-4-hydroxy-2,5-methano-3-oxo-1,5-dihydrobenzothiopheno[2,3-e][1,3]diazepine-1-carboxylate (2f)
(20) A solution of intermediate (2e) (0.250 g, 0.697 mmol) in DCM (15 mL) was degassed 10 min under argon atmosphere. AcOH (0.080 mL, 1.394 mmol) and Pd(PPh.sub.3).sub.4 (0.403 g, 0.349 mmol) were successively added. After stirring for 30 min at rt, the mixture was evaporated under nitrogen flux. The residue was purified by column chromatography on silica gel (DCM/acetone 10/1 to 9/1). Fractions of interest are combined and concentrated. The residue was triturated with Et.sub.2O to give intermediate (2f) as an orange solid (0.054 g, 0.167 mmol, 24%). MS m/z ([M+H].sup.+) 319. .sup.1H NMR (300 MHz, acetone-d.sub.6): δ (ppm) 1.32 (t, J=7.1 Hz, 3H), 3.66 (dd, J=3.0, 11.5 Hz, 1H), 3.95 (dd, J=0.3, 8.6 Hz, 1H), 4.30 (q, J=7.1 Hz, 2H), 4.69 (d, J=3.0 Hz, 1H), 5.29 (s, 1H), 7.45-7.36 (m, 2H), 7.81-7.85 (m, 1H), 7.94-7.97 (m, 1H), 8.89 (s, 1H).
Step 7: Preparation of sodium trans-(1-ethoxycarbonyl-2,5-methano-3-oxo-1,5-dihydrobenzothiopheno[2,3-e][1,3]diazepin-4-yl) sulfate, Example 2
(21) To a solution of intermediate (2f) (0.044 g, 0.138 mmol) in anhydrous pyridine (1.5 mL) under inert atmosphere was added sulfur trioxide pyridine complex (0.132 g, 0.829 mmol). After stirring for 16 h, the mixture was concentrated in vacuum. DCM was added and the insoluble was filtered off. The filtrate was purified by flash chromatography on silica gel (DCM/acetone 10/0 to 0/10) to give Example 2 as pyridinium salt (0.074 g, 0.185 mmol). The sulfated product was applied on a Dowex sodium form column (Dowex® 50WX8 hydrogen form stored with an aqueous solution of 2N NaOH and washed until neutral pH with water). The fractions containing the desired compound were combined, frozen and lyophilized. The solid was triturated with DCM to give Example 2 as sodium salt (0.035 g, 0.083 mmol, 49%). MS m/z ([M+H].sup.+) 399. .sup.1H NMR (400 MHz, D.sub.2O): δ (ppm) 1.28 (t, J=7.1 Hz, 3H), 3.79 (d, J=11.9 Hz, 1H), 3.93 (dd, J=3.0, 11.9 Hz, 1H), 4.24-4.42 (m, 2H), 5.13 (d, J=3.0 Hz, 1H), 5.63 (s, 1H), 7.40-7.48 (m, 2H), 7.77-7.84 (m, 1H), 7.90-7.96 (m, 1H).
Example 3: synthesis of Sodium trans-(1-carbamoyl-2,5-methano-3-oxo-1,5-dihydrobenzothiopheno[2,3-e][1,3]diazepin-4-yl) sulfate
(22) ##STR00022##
Step 1: Preparation of Intermediate tert-butyl 4-(allyloxyamino)-1-carbamoyl-3,4-dihydro-1H-benzothiopheno[3,2-c]pyridine-2-carboxylate (3a)
(23) To a solution of intermediate (2c) (500 mg, 1.16 mmol) in MeOH (2 mL) was added NH.sub.3 7N in MeOH (15 mL, 105 mmol). After stirring for 18 h at 66° C., NH.sub.3 aqueous 30% was added (4 mL) and the mixture was stirred for 6 days at 66° C. The solution was evaporated under nitrogen flux. The residue was diluted with DCM and washed with NaOH 1N. The organic layer was dried over Na.sub.2SO.sub.4, filtered and concentrated. The residue was purified by column chromatography on silica gel (DCM/acetone 10/0 to 9/1). The solid was triturated with Et.sub.2O to give intermediate (3a) as a mixture of both diastereoisomers (244 mg, 0.605 mmol, 52%). MS m/z ([M+H].sup.+) 404.
Step 2: Preparation of Intermediate 4-(allyloxyamino)-1,2,3,4-tetrahydrobenzothiopheno[3,2-c]pyridine-1-carboxamide (3b)
(24) To a solution of intermediate (3a) (234 mg, 0.58 mmol) in DCM (6 mL) under argon atmosphere at 0° C. was dropped TFA (1.5 mL, 19.72 mmol). After stirring for 1 h at rt, the mixture was evaporated under nitrogen flux. The residue was purified by column chromatography on silica gel (DCM/MeOH 10/0 to 9/1) to give intermediate (3b) as a mixture of both diastereoisomers (249 mg, 0.821 mmol, quantitative). MS m/z ([M+H].sup.+) 304. .sup.1H NMR (400 MHz, CDCl.sub.3): δ (ppm) 3.47-3.50 (m, 0.5H), 3.75-3.83 (m, 1.5H), 4.15 (t, J=5.4 Hz, 2H), 4.44-4.47 (m, 1H), 5.18 (d, J=9.6 Hz, 1H), 5.21-5.29 (m, 1H), 5.56 (d, J=15.7 Hz, 1H), 5.80-5.90 (m, 1H), 6.66 and 6.84 (2S, 1H), 7.31-7.45 (m, 2H), 7.62-7.76 (m, 1H), 7.79-7.87 (m, 1H).
Step 3: Preparation of Intermediate trans-4-allyloxy-2,5-methano-3-oxo-1,5-dihydrobenzothiopheno[2,3-e][1,3]diazepine-1-carboxamide (3c)
(25) To a solution of intermediate (3b) (175 mg, 0.577 mmol) in chlorobenzene (1.6 mL) under argon atmosphere were successively added DIPEA (0.110 mL, 0.635 mmol) and a solution of Fmoc-Cl (151 mg, 0.582 mmol) in chlorobenzene (0.8 mL). After 1 h at rt, CDI (117 mg, 0.721 mmol) was added and the mixture was heated at 45° C. for 1 h. CDI was added and the mixture was heated at 45° C. until complete conversion. Then, the mixture was cold and Et.sub.2NH (0.240 mL, 2.31 mmol) was added. After 3 h at rt, the mixture was diluted with DCM. The organic layer was washed with HCl 1N and brine, dried over Na.sub.2SO.sub.4, filtered and concentrated. The residue was purified by column chromatography on silica gel (DCM/MeOH 10/0 to 9/1) to give intermediate (3c) (130 mg, 1.75 mmol, 22%). MS m/z ([M+H].sup.+) 330. .sup.1H NMR (400 MHz, CDCl.sub.3): δ (ppm) 3.53 (d, J=11.1 Hz, 1H), 3.66 (dd, J=3.0, 11.1 Hz, 1H), 4.39-4.49 (m, 2H), 4.53 (d, J=2.7 Hz, 1H), 5.26 (s, 1H), 5.30-5.35 (m, 1H), 5.38 (dq, J=1.3, 17.3 Hz, 1H), 6.02 (ddt, J=6.3, 10.4, 16.9 Hz, 1H), 6.10 (bs, 1H), 6.94 (bs, 1H), 7.33 (dq, J=1.3, 7.1 Hz, 2H), 7.72-7.77 (m, 2H).
Step 4: Preparation of Intermediate trans-4-hydroxy-2,5-methano-3-oxo-1,5-dihydrobenzothiopheno[2,3-e][1,3]diazepine-1-carboxamide (3d)
(26) A solution of intermediate (3c) (43 mg, 0.130 mmol) in DCM (2 mL) was degassed 10 min under argon atmosphere. AcOH (0.015 mL, 0.26 mmol) and Pd(PPh.sub.3).sub.4 (0.075 g, 0.065 mmol) were successively added. After stirring for 30 min at rt, the mixture was dried under nitrogen flux. The residue was purified by column chromatography on silica gel (DCM/acetone 10/0 to 0/10) to give intermediate (3d) (23 mg, 0.079 mmol, 60%). MS m/z ([M+H].sup.+) 290. .sup.1H NMR (300 MHz, acetone-d.sub.6): δ (ppm) 3.60 (dd, J=3.0, 11.2 Hz, 1H), 3.79 (d, J=11.2 Hz, 1H), 4.60 (d, J=3.0 Hz, 1H), 5.18 (s, 1H), 7.32-7.38 (m, 2H), 7.82-7.90 (m, 2H).
Step 5: Preparation of Sodium trans-(1-carbamoyl-2,5-methano-3-oxo-1,5-dihydrobenzothiopheno[2,3-e][1,3]diazepin-4-yl) sulfate, Example 3
(27) To a solution of intermediate (3d) (23 mg, 0.079 mmol) in anhydrous pyridine (0.8 mL) under inert atmosphere was added sulfur trioxide pyridine complex (0.076 g, 0.476 mmol). After stirring for 18 h, the heterogeneous mixture was concentrated in vacuum. The residue was filtered and washed with DCM, acetone and IPA. The residue was solubilized with water and the aqueous solution was partially evaporated by nitrogen flux. The aqueous solution was applied on a Dowex sodium form column (Dowex® 50WX8 hydrogen form stored with an aqueous solution of 2N NaOH and washed until neutral pH with water). The fractions containing the desired compound were combined, frozen and lyophilized to give Example 3 as a white solid (0.011 g, 0.028 mmol, 35%). MS m/z ([M+H].sup.+) 370. MS m/z ([M−H].sup.−) 368. .sup.1H NMR (400 MHz, D.sub.2O): δ (ppm) 3.75 (d, J=11.9 Hz, 1H), 3.93 (dd, J=3.0, 11.9 Hz, 1H), 5.16 (d, J=2.6 Hz, 1H), 5.55 (s, 1H), 7.46-7.53 (m, 2H), 7.73-7.80 (m, 1H), 7.98-8.04 (m, 1H).
Example 4: Synthesis of sodium 1,4-methano-3-oxo-1,5-dihydro-[1,3]diazepino[5,6-b]quinolin-2-yl) sulfate
(28) ##STR00023##
Step 1: Preparation of Intermediate tert-butyl 4-oxo-1,3-dihydrobenzo[b][1,7]naphthyridine-2-carboxylate (4a)
(29) A mixture of 2-nitrobenzaldehyde (600 mg, 3.97 mmol), N-Boc-piperidine-3,5-dione (1.01 g, 4.76 mmol) and iron (1.11 g, 19.85 mmol) in acetic acid (20 mL) was heated at 50° C. for 1 h. The mixture was concentrated in vacuo. The residue was dissolved in AcOEt and filtered over Celite®. The filtrate was carefully washed twice with a saturated solution of NaHCO.sub.3. The organic layer was dried over Na.sub.2SO.sub.4, filtered and concentrated in vacuo. The residue was purified by flash chromatography on silica gel (Heptane/AcOEt 10/0 to 0/10) to provide intermediate (4a) as a yellow solid (448 mg, 1.50 mmol, 37%). MS m/z ([M+H].sup.+) 299. .sup.1H NMR (400 MHz, CDCl.sub.3): δ (ppm) 1.49 (s, 9H), 4.48 (s, 2H), 5.07 (s, 2H), 7.62 (ddd, J=1.2, 6.8, 8.1 Hz, 1H), 7.88 (ddd, J=1.5, 6.9, 8.5 Hz, 1H), 7.98 (d, J=8.0 Hz, 1H), 8.13 (d, J=8.6 Hz, 1H), 8.91 (s, 1H).
Step 2: Preparation of Intermediate tert-butyl 4-hydroxy-3,4-dihydro-1H-benzo[b][1,7]naphthyridine-2-carboxylate (4b)
(30) To a solution of intermediate (4a) (445 mg, 1.49 mmol) in MeOH (5 mL) at 0° C. was portionwise added NaBH.sub.4 (68 mg, 1.79 mmol). The reaction mixture was stirred for 45 min at 0° C., then hydrolyzed with water and concentrated in vacuo. The residue was solubilized in AcOEt. The layers were separated. The aqueous layer was extracted with AcOEt. The organic layers were combined, washed with brine, dried over Na.sub.2SO.sub.4, filtered and concentrated in vacuo to provide intermediate (4b) as a yellow foam (412 mg, 1.37 mmol, 91%). MS m/z ([M+H].sup.+) 301. .sup.1H NMR (400 MHz, CDCl.sub.3): δ (ppm) 1.51 (s, 9H), 3.77 (dd, J=6.4, 13.3 Hz, 1H), 3.94 (d, J=13.3 Hz, 1H), 4.78-5.08 (m, 3H), 7.53 (t, J=7.4 Hz, 1H), 7.72 (ddd, J=1.4, 6.8, 8.4 Hz, 1H), 7.83 (d, J=8.1 Hz, 1H), 8.05 (d, J=8.6 Hz, 1H), 8.34 (s, 1H).
Step 3: Preparation of Intermediate tert-butyl 4-(allyloxyamino)-3,4-dihydro-1H-benzo[b][1,7]naphthyridine-2-carboxylate (4c)
(31) To a solution of intermediate (4b) (405 mg, 1.35 mmol) in DCM (5 mL) under inert atmosphere at −78° C. were successively added methanesulfonic anhydride (352 mg, 2.02 mmol) and TEA (0.552 mL, 3.96 mmol). The mixture was stirred at −78° C. for 90 min. A solution of 0-allylhydroxylamine 50% in DCM (985 mg, 6.74 mmol) was added and the mixture was stirred at rt overnight. Water was added and the layers were separated. The aqueous layer was extracted with DCM. The organic layers were combined, dried over Na.sub.2SO.sub.4, filtered and concentrated in vacuo. The residue was purified by flash chromatography on silica gel (heptane/AcOEt 10/0 to 0/10) to provide intermediate (4c) as a yellow oil (205 mg, 0.57 mmol, 42%). MS m/z ([M+H].sup.+) 356.
Step 4: Preparation of Intermediate 2-allyloxy-1,4-methano-1,5-dihydro-[1,3]diazepino[5,6-b]quinolin-3-one (4d)
(32) To a solution of intermediate (4c) (200 mg, 0.563 mmol) in DCM (5 mL) at 0° C. were successively added TEA (118 μL, 0.844 mmol) and diphosgene (48 μL, 0.394 mmol). The mixture was stirred at 0° C. for 30 min before adding a saturated solution of NaHCO.sub.3. The mixture was stirred for 5 min and the layers were separated. The aqueous layer was extracted with DCM. The organic layers were combined, dried over Na.sub.2SO.sub.4, filtered and concentrated in vacuo. The residue was dissolved in dioxane (1.5 mL) and a solution of hydrochloric acid 4N in dioxane (4 mL) was added. The mixture was stirred at rt for 50 min and concentrated in vacuo. The residue was solubilized in AcOEt (10 mL) and a saturated solution of NaHCO.sub.3 (10 mL) was added. The aqueous layer was extracted with AcOEt. The organic layers were combined, dried over Na.sub.2SO.sub.4, filtered and concentrated in vacuo. The residue was purified by flash chromatography on silica gel (DCM/AcOEt 10/0 to 3/7) to provide intermediate (4d) as a yellow foam (102 mg, 0.362 mmol, 64%). MS m/z ([M+H].sup.+) 282. .sup.1H NMR (400 MHz, CDCl.sub.3): δ (ppm) 3.30 (d, J=11.3 Hz, 1H), 3.82 (ddd, J=1.3, 3.0, 11.4 Hz, 1H), 4.40-4.60 (m, 4H), 4.75 (dd, J=1.3, 17.8 Hz, 1H), 5.29-5.44 (m, 2H), 5.97-6.12 (m, 1H), 7.54 (ddd, J=1.2, 6.9, 8.1 Hz, 1H), 7.71 (ddd, J=1.5, 6.9, 8.5 Hz, 1H), 7.80 (dd, J=1.5, 8.2 Hz, 1H), 7.93 (s, 1H), 8.00 (d, J=8.5 Hz, 1H).
Step 5: Preparation of sodium 1,4-methano-3-oxo-1,5-dihydro-[1,3]diazepino[5,6-b]quinolin-2-yl) sulfate, Example 4
(33) To a solution of intermediate (4d) (102 mg, 0.36 mmol) in anhydrous DCM (5 mL) under inert atmosphere were successively added AcOH (42 μL, 0.72 mmol) and Pd(PPh.sub.3).sub.4 (210 mg, 0.18 mmol). After stirring at rt for 2 h, further Pd(PPh.sub.3).sub.4 (105 mg, 0.09 mmol) was added and the stirring resumed for 1 h. Pyridine (3 mL) and sulfur trioxide-pyridine complex (289 mg, 1.81 mmol) were added. The mixture was stirred at rt overnight then concentrated in vacuo. DCM was added to the residue and the solids were filtered off. The filtrate was concentrated in vacuo. The crude was purified by flash chromatography on silica gel (DCM/acetone 10/0 to 0/10). The fractions containing the sulfated intermediate were combined and applied on a Dowex sodium form column (Dowex® 50WX8 hydrogen form stored with a 2N aqueous NaOH solution and washed until neutral pH with water). The fractions containing the desired compound were combined, frozen and lyophilized. The residue was purified by flash chromatography on C18 silica gel (water/ACN 10/0 to 0/10) to provide Example 4 as a white solid (38 mg, 0.11 mmol, 30%). MS m/z ([M+H].sup.+) 322. MS m/z ([M−H].sup.−) 320. .sup.1H NMR (400 MHz, DMSO-d.sub.6): δ (ppm) 3.51 (d, J=11.6 Hz, 1H), 3.67 (ddd, J=1.2, 3.1, 11.6 Hz, 1H), 4.33 (d, J=17.4 Hz, 1H), 4.57 (d, J=17.4 Hz, 1H), 4.95 (d, J=2.8 Hz, 1H), 7.60 (ddd, J=1.2, 6.8, 8.1 Hz, 1H), 7.75 (ddd, J=1.5, 6.9, 8.4 Hz, 1H), 7.93 (d, J=8.3 Hz, 1H), 8.03 (dd, J=1.2, 8.0 Hz, 1H), 8.13 (s, 1H).
Example 5: Synthesis of sodium (1,4-methano-3-oxo-1,5-dihydro-[1,3]diazepino[5,6-b][1,7]naphthyridin-2-yl) sulfate
(34) ##STR00024##
Step 1: Preparation of Intermediate tert-butyl 4-oxo-1,3,5,10-tetrahydropyrido[3,4-b][1,7]naphthyridine-2-carboxylate (5a)
(35) A mixture of 3-nitropyridine-4-carboxaldehyde (1.50 g, 9.86 mmol), N-Boc-piperidine-3,5-dione (2.52 g, 11.83 mmol) and iron (2.75 g, 49.3 mmol) in acetic acid (50 mL) was heated at 50° C. for 3 h. The mixture was concentrated in vacuo. The solution was co-evaporated twice with toluene. The residue was dissolved in AcOEt and a saturated solution of NaHCO.sub.3. The mixture was filtered over Celite®. The layers of the filtrate was separated. The aqueous layer was extracted with AcOEt. The organic layers were combined, dried over Na.sub.2SO.sub.4, filtered and concentrated in vacuo. Acetone (20 mL) was added to the residue and the mixture refluxed for 30 min. The mixture was cooled at rt and filtrated to provide intermediate (5a) as an off-white solid (1.21 g, 4.01 mmol, 40%). MS m/z ([M+H].sup.+) 302. .sup.1H NMR (400 MHz, DMSO-d.sub.6): δ (ppm) 1.43 (s, 9H), 3.55 (s, 2H), 3.94 (s, 2H), 4.29 (s, 2H), 7.13 (d, J=4.9 Hz, 1H), 7.91-8.19 (m, 2H), 9.76 (s, 1H).
Step 2: Preparation of Intermediate tert-butyl 4-oxo-1,3-dihydropyrido[3,4-b][1,7]naphthyridine-2-carboxylate (5b)
(36) To a suspension of intermediate (5a) (1.20 g, 3.98 mmol) in DCM (80 mL) at rt was added activated manganese dioxide (3.16 g, 35.8 mmol) in three portions every 20 min. The mixture was stirred at rt for 20 min then filtered over a pad of Celite®. The filtrate was concentrated in vacuo. The residue was purified by flash chromatography on silica gel (DCM/AcOEt 10/0 to 2/8) to provide intermediate (5b) as a yellow solid (645 mg, 2.15 mmol). MS m/z ([M+H].sup.+) 300.
(37) .sup.1H NMR (400 MHz, DMSO-d.sub.6): δ (ppm) 1.41 (s, 9H), 4.48 (s, 2H), 5.01 (s, 2H), 8.14 (dd, J=1.0, 5.6 Hz, 1H), 8.71 (d, J=5.6 Hz, 1H), 9.05 (d, J=0.9 Hz, 1H), 9.48 (s, 1H).
Step 3: Preparation of Intermediate tert-butyl 4-hydroxy-3,4-dihydro-1H-pyrido[3,4-b][1,7]naphthyridine-2-carboxylate (5c)
(38) Using the procedure described in example 4 (step 2), intermediate (5b) (543 mg, 1.81 mmol) was converted into intermediate (5c) as a pale yellow solid (497 mg, 1.65 mmol, 90%) which was used without purification. MS m/z ([M+H].sup.+) 302. .sup.1H NMR (400 MHz, Acetone-d.sub.6): δ (ppm) 1.50 (s, 9H), 3.27-3.58 (m, 1H), 4.16 (bs, 1H), 4.73 (d, J=18.2 Hz, 1H), 4.84-5.18 (m, 3H), 7.84 (dd, J=1.0, 5.6 Hz, 1H), 8.47 (s, 1H), 8.56 (d, J=5.6 Hz, 1H), 9.30 (d, J=1.0 Hz, 1H).
Step 4: Preparation of Intermediate tert-butyl 4-(allyloxyamino)-3,4-dihydro-1H-benzo[b][1,7]naphthyridine-2-carboxylate (5d)
(39) To a solution of intermediate (5c) (250 mg, 0.83 mmol) in DCM (10 mL) under inert atmosphere at −20° C. were successively added methanesulfonic anhydride (217 mg, 1.24 mmol) and TEA (0.226 mL, 1.66 mmol). The mixture was stirred at −20° C. for 20 min. Water was added. The layers were separated. The aqueous layer was extracted with DCM. The organic layers were combined, dried over Na.sub.2SO.sub.4, filtered and concentrated in vacuo. A solution of 0-allylhydroxylamine 50% in DCM (1.21 g, 8.3 mmol) was added to the residue and the mixture was heated at 40° C. for 30 min. The mixture was diluted with DCM and washed with a saturated solution of NaHCO.sub.3. The organic layer was dried over Na.sub.2SO.sub.4, filtered and concentrated in vacuo. The residue was purified by flash chromatography on silica gel (DCM/AcOEt 10/0 to 0/10) to provide intermediate (5d) as a yellow oil (158 mg, 0.44 mmol, 53%). MS m/z ([M+H].sup.+) 357. 1H NMR (400 MHz, CDCl.sub.3): δ (ppm) 1.49 (s, 9H), 3.44-3.59 (m, 1H), 4.14-4.44 (m, 4H), 4.67 (d, J=18.3 Hz, 1H), 5.01-5.20 (m, 2H), 5.24 (d, J=17.3 Hz, 1H), 5.66 (bs, 1H), 5.81-5.97 (m, 1H), 7.61 (d, J=5.6 Hz, 1H), 8.21 (s, 1H), 8.55 (d, J=5.6 Hz, 1H), 9.40 (s, 1H).
Step 5: Preparation of Intermediate 2-allyloxy-1,4-methano-1,5-dihydro-[1,3]diazepino[5,6-b][1,7]naphthyridin-3-one (5e)
(40) To a solution of intermediate (5d) (158 mg, 0.443 mmol) in DCM (5 mL) at 0° C. were successively added TEA (93 μL, 0.664 mmol) and diphosgene (37 μL, 0.31 mmol). The mixture was stirred at 0° C. for 30 min before adding a saturated solution of NaHCO.sub.3. The mixture was stirred for 5 min and the layers were separated. The aqueous layer was extracted with DCM. The organic layers were combined, dried over Na.sub.2SO.sub.4, filtered and concentrated in vacuo. The residue was dissolved in DCM (3 mL) and TFA (1 mL) was added. The mixture was stirred at rt for 3 h then concentrated in vacuo. The residue was solubilized in AcOEt (10 mL) and a saturated solution of NaHCO.sub.3 (10 mL) was added. The aqueous layer was extracted with AcOEt. The organic layers were combined, dried over Na.sub.2SO.sub.4, filtered and concentrated in vacuo. The residue was purified by flash chromatography on silica gel (DCM/acetone 10/0 to 0/10) to provide intermediate (5e) as an orange oil (13 mg, 0.046 mmol, 10%). MS m/z ([M+H].sup.+) 283. .sup.1H NMR (400 MHz, CDCl.sub.3): δ (ppm) 3.30 (d, J=11.5 Hz, 1H), 3.84 (ddd, J=1.3, 3.0, 11.6 Hz, 1H), 4.40-4.51 (m, 2H), 4.54 (d, J=18.0 Hz, 1H), 4.59 (d, J=2.9 Hz, 1H), 4.74 (dd, J=1.2, 18.0 Hz, 1H), 5.30-5.43 (m, 2H), 5.96-6.09 (m, 1H), 7.64 (dd, J=0.8, 5.6 Hz, 1H), 7.92 (s, 1H), 8.61 (d, J=5.6 Hz, 1H), 9.39 (s, 1H).
Step 6: Preparation of sodium (1,4-methano-3-oxo-1,5-dihydro-[1,3]diazepino[5,6-b][1,7]naphthyridin-2-yl) sulfate, Example 5
(41) To a solution of intermediate (5e) (13 mg, 0.046 mmol) in anhydrous DCM (1 mL) under inert atmosphere were successively added AcOH (5 μL, 0.092 mmol) and Pd(PPh.sub.3).sub.4 (27 mg, 0.023 mmol). After stirring at rt for 30 min, further Pd(PPh.sub.3).sub.4 (27 mg, 0.023 mmol) and AcOH (5 μL, 0.092 mmol) were added. The stirring resumed for 1 h. The precipitate was filtered and solubilized in pyridine (0.7 mL) before adding sulfur trioxide-pyridine complex (37 mg, 0.23 mmol). The mixture was stirred at rt for 4 h then concentrated in vacuo. DCM was added to the residue and the solids were filtered off. The filtrate was concentrated in vacuo. The crude was applied on a Dowex sodium form column (Dowex® 50WX8 hydrogen form stored with a 2N aqueous NaOH solution and washed until neutral pH with water). The fractions containing the desired compound were combined, frozen and lyophilized. The residue was purified by flash chromatography on C18 silica gel (water/ACN 99/1 to 10/90) to provide Example 5 as a white solid (1.5 mg, 0.0043 mmol, 9%). MS m/z ([M−H].sup.−) 321. .sup.1H NMR (400 MHz, D.sub.2O): δ (ppm) 3.69 (d, J=12.0 Hz, 1H), 4.03 (ddd, J=1.3, 3.1, 12.0 Hz, 1H), 4.65 (d, J=17.9 Hz, 1H), 4.77 (d, J=17.9 Hz, 1H), 5.21 (d, J=3.0 Hz, 1H), 7.82 (dd, J=0.8, 5.7 Hz, 1H), 8.33 (s, 1H), 8.48 (d, J=5.7 Hz, 1H), 9.14 (s, 1H).
Example 6: Synthesis of sodium [trans-1-ethoxycarbonyl-2,5-methano-6-methyl-3-oxo-1,5-dihydro-[1,3]diazepino[5,6-b]indol-4-yl] sulfate
(42) ##STR00025##
Step 1: Preparation of Intermediate 2-(1-methylindol-2-yl)-2-trimethylsilyloxy-acetonitrile
(43) In a sealed tube, TMSCN (4 mL) was added to 1-methylindole-2-carbaldehyde (1.25 g, 7.85 mmol) and mixture was stirred at 80° C. for 48 h. The mixture was concentrated to give crude intermediate (6a) which was used without purification in the next step. MS m/z ([M+H].sup.+) 259.
Step 2: Preparation of Intermediate 2-amino-1-(1-methylindol-2-yl)ethanol (6b)
(44) At 0° C., a solution of LiAlH.sub.4 2M in THF (7.85 mL, 15.7 mmol) was added dropwise to a solution of intermediate (6a) (7.85 mmol) in THF (5 mL) and the mixture was stirred at rt for 2 h. At 0° C., the mixture was quenched by addition of AcOEt (1 mL), water (0.6 mL), NaOH 10% (0.9 mL), and water (1.8 mL). After 15 min of stirring, solids were filtered off and washed with AcOEt. The filtrate was concentrated and the residue was triturated with DCM. The solid was filtered and dried under vacuum to give intermediate (6b) (890 mg, 4.68 mmol, 60% on 2 steps). MS m/z ([M−OH].sup.+) 173. .sup.1H NMR (400 MHz, DMSO-d.sub.6): δ (ppm) 1.50 (bs, 2H), 2.82-2.96 (m, 2H), 3.76 (s, 3H), 4.65 (t, J=6.3 Hz, 1H), 5.31 (bs, 1H), 6.34 (s, 1H), 6.99 (ddd, J=1.0, 7.0, 7.9 Hz, 1H), 7.10 (ddd, J=1.2, 7.0, 8.3 Hz, 1H), 7.40 (dd, J=1.0, 8.2 Hz, 1H), 7.48 (d, J=7.8 Hz, 1H).
Step 3: Preparation of Intermediate ethyl 4-hydroxy-5-methyl-1,2,3,4-tetrahydropyrido[4,3-b]indole-1-carboxylate (6c)
(45) A solution of Ethyl glyoxylate 50% in Toluene (1.04 mL, 5.26 mmol) was added to a solution of intermediate (6b) (1.0 g, 5.26 mmol) in MeOH (5.2 mL). The mixture was stirred at rt for 18 h. The precipitate was filtered, washed with MeOH and dried under vacuum to give intermediate (6c) as a white solid (820 mg, 2.99 mmol, 57%, mixture of both diastereoisomers). MS m/z ([M+H].sup.+) 275. .sup.1H NMR (400 MHz, DMSO-d.sub.6): δ (ppm) 1.24 (t, J=7.1 Hz, 3H), 2.70 (bs, 1H), 3.03 (dd, J=4.4, 13.2 Hz, 1H), 3.15 (dd, J=4.5, 13.3 Hz, 1H), 3.76 (s, 3H), 4.10-4.24 (m, 2H), 4.64 (s, 1H), 4.65-4.72 (m, 1H), 5.41 (d, J=6.5 Hz, 1H), 6.98-7.05 (m, 1H), 7.11-7.17 (m, 1H), 7.43 (d, J=8.2 Hz, 1H), 7.53 (d, J=8.0 Hz, 1H).
Step 4: Preparation of Intermediate O2-tert-butyl O1-ethyl 4-hydroxy-5-methyl-3,4-dihydro-1H-pyrido[4,3-b]indole-1,2-dicarboxylate (6d)
(46) TEA (0.84 mL, 5.98 mmol) and Boc.sub.2O (652 mg, 2.99 mmol) were added to a solution of intermediate (6c) (820 mg, 2.99 mmol) in DCM (15 mL). After stirring for 2 h at rt, the mixture was diluted with DCM and washed with H.sub.2O and brine. The organic layer was dried over Na.sub.2SO.sub.4, filtered and concentrated in vacuo. The crude product was purified by column chromatography on silica gel (DCM/Acetone: 10/0 to 6/4) to give intermediate (6d) (1.06 g, 2.83 mmol, 95%, mixture of both diastereoisomers). MS m/z ([M-OH].sup.+) 357. .sup.1H NMR (400 MHz, CDCl.sub.3): δ (ppm) 1.23 (t, J=7.1 Hz, 3H), 1.51 (s, 9H), 3.59-3.68 (m, 1H), 3.82 (s, 3H), 4.08-4.20 (m, 2H), 4.24-4.42 (m, 1H), 4.91-5.01 (m, 1H), 5.57-5.69 (m, 1H), 7.13-7.19 (m, 1H), 7.24-7.30 (m, 1H), 7.34 (d, J=8.3 Hz, 1H), 7.73 (d, J=8.0 Hz, 1H).
Step 5: Preparation of Intermediate O2-tert-butyl O1-ethyl 4-(allyloxyamino)-5-methyl-3,4-dihydro-1H-pyrido[4,3-b]indole-1,2-dicarboxylate (6e)
(47) At −78° C., a solution of Ms.sub.2O (700 mg, 4.02 mmol) in DCM (4 mL) was slowly added to a solution of TEA (0.75 mL, 5.36 mmol) and intermediate (6d) (500 mg, 1.34 mmol) in DCM (16 mL). The mixture was stirred for 45 min at −78° C. A solution of O-allylhydroxylamine (683 mg, 9.36 mmol) in DCM (4 mL) was then added. The mixture was slowly warmed to rt for 1 h. The mixture was diluted with DCM and washed with H.sub.2O and brine. The organic layer was dried over Na.sub.2SO.sub.4, filtered and concentrated in vacuo. The crude product was purified by column chromatography on silica gel (DCM/Acetone: 10/0 to 8/2) to give intermediate (6e) (530 mg, 1.24 mmol, 93%, mixture of both diastereoisomers). MS m/z ([M+H].sup.+) 430.
Step 6: Preparation of Intermediate ethyl trans-4-allyloxy-2,5-methano-6-methyl-3-oxo-1,5-dihydro-[1,3]diazepino[5,6-b]indole-1-carboxylate (6f)
(48) At 0° C., diphosgene (157 μL, 1.3 mmol) was added to a solution of TEA (279 μL, 2.0 mmol) and intermediate (6e) (430 mg, 1.0 mmol) in DCM (5 mL). The mixture was stirred at 0° C. for 45 min then a solution of MeSO.sub.3H (973 μL, 15.0 mmol) in DCM (5 mL) was added. After 1 h at 0° C., TEA (6.95 mL, 50 mmol) was added and mixture was stirred at 70° C. for 18 h. The mixture was diluted with DCM and washed with H.sub.2O and brine. The organic layer was dried over Na.sub.2SO.sub.4, filtered and concentrated in vacuo. The crude product was purified by column chromatography on silica gel (DCM/Acetone: 10/0 to 8/2) to give intermediate (6f) (252 mg, 0.71 mmol, 71%). MS m/z ([M+H].sup.+) 356. .sup.1H NMR (400 MHz, CDCl.sub.3): δ (ppm) 1.35 (t, J=7.1 Hz, 3H), 3.73 (s, 3H), 3.73-3.78 (m, 2H), 3.92 (d, J=11.4 Hz, 1H), 4.20-4.38 (m, 2H), 4.39-4.52 (m, 2H), 4.64 (d, J=2.5 Hz, 1H), 5.30-5.34 (m, 1H), 5.34-5.41 (m, 1H), 5.97-6.09 (m, 1H), 7.12 (ddd, J=1.1, 7.0, 8.0 Hz, 1H), 7.23 (ddd, J=1.2, 6.9, 8.3 Hz, 1H), 7.28-7.32 (m, 1H), 7.66-7.70 (m, 1H).
Step 7: Preparation of sodium [trans-1-ethoxycarbonyl-2,5-methano-6-methyl-3-oxo-1,5-dihydro-[1,3]diazepino[5,6-b]indol-4-yl] sulfate, Example 6
(49) AcOH (15 μL, 0.25 mmol) and Pd(PPh.sub.3).sub.4 (72 mg, 0.06 mmol) were successively added to a solution of intermediate (6f) (44 mg, 0.12 mmol) in anhydrous DCM (1.2 mL). After 2 h at rt, further AcOH (15 μL, 0.25 mmol) and Pd(PPh.sub.3).sub.4 (72 mg, 0.06 mmol) were added and mixture was stirred for 18 h. Mixture was evaporated under flux of nitrogen. The residue was purified by flash chromatography on silica gel (DCM/acetone 10/0 to 6/4) to give the de-allylated intermediate (16 mg). Pyridine (1 mL) and sulfur trioxide pyridine complex (45 mg, 0.28 mmol) were added to the de-allylated intermediate and mixture was stirred at rt for 3 h. The mixture was diluted with DCM and the solids were filtered off. The filtrate was concentrated and the residue was purified by flash chromatography on silica gel (DCM/acetone 10/0 to 0/10). Desired fractions were concentrated in vacuo and residue was applied on a Dowex sodium form column (Dowex® 50WX8 hydrogen form stored with an aqueous solution of 2N NaOH and washed until neutral pH with water). The fractions containing the desired compound were combined, frozen and lyophilized to provide Example 6 as sodium salt (12 mg, 0.029 mmol, 24%). MS m/z ([M−H].sup.−) 394. .sup.1H NMR (400 MHz, D.sub.2O): δ (ppm) 1.33 (t, J=6.1 Hz, 3H), 3.75-3.77 (m, 1H), 3.82 (s, 3H), 3.95-3.97 (m, 1H), 4.25-4.45 (m, 2H), 5.32 (s, 1H), 5.61 (s, 1H), 7.25 (t, J=6.7 Hz, 1H), 7.37 (t, J=6.7 Hz, 1H), 7.56 (d, J=8.1 Hz, 1H), 7.77 (d, J=7.8 Hz, 1H).
Example 7: Synthesis of sodium [9-(diethylcarbamoyl)-2,5-methano-3-oxo-5,7,8,10-tetrahydro-1H-pyrido[3,4]thieno[1,3-d][1,3]diazepin-4-yl] sulfate
(50) ##STR00026## ##STR00027## ##STR00028##
Step 1: Preparation of Intermediate O5-tert-butyl O3-ethyl 2-amino-6,7-dihydro-4H-thieno[3,2-c]pyridine-3,5-dicarboxylate (7a)
(51) To a solution of N-tert-butoxycarbonyl-4-piperidone (1.0 g, 5.02 mmol) and ethylcyanoacetate (0.53 mL, 5.02 mmol) in absolute ethanol (25 mL) was added sulfur powder (160 mg, 5.02 mmol) and morpholine (437 mg, 5.02 mmol). The mixture was refluxed for 1 hour and then concentrated under reduced pressure to give intermediate 7a as yellow solid (1.60 g, 5.02 mmol, 100%) which was used without further purification. MS m/z ([M+H].sup.+) 327. .sup.1H NMR (400 MHz, CDCl.sub.3) δ (ppm): 1.33 (t, J=7.1 Hz, 3H), 1.47 (s, 9H), 2.80 (t, J=5.9 Hz, 2H), 3.61 (t, J=5.9 Hz, 2H), 4.26 (q, J=7.1 Hz, 2H), 4.35 (bs, 2H), 6.01 (bs, 2H).
Step 2: Preparation of Intermediate O5-tert-butyl O3-ethyl 6,7-dihydro-4H-thieno[3,2-c]pyridine-3,5-dicarboxylate (7b)
(52) At 0° C., isoamyl nitrite (1.49 mL, 11.06 mmol) was added dropwise to a solution of intermediate (7a) (1.6 g, 5.02 mmol) in THF (8 mL). The mixture was then refluxed for 2 h, cooled to room temperature and concentrated under reduced pressure. The crude was purified by column chromatography on silica gel (petroleum ether/EtOAc: 10/1) to give intermediate (7b) as yellow powder (140 mg, 0.45 mmol, 10%). MS m/z ([M+Na].sup.+) 334. .sup.1H NMR (400 MHz, CDCl.sub.3) δ (ppm): 1.35 (m, J=7.1 Hz, 3H), 1.48 (s, 9H), 2.98 (t, J=5.9 Hz, 2H), 3.66 (t, J=5.8 Hz, 2H), 4.29 (q, J=7.1 Hz, 2H), 4.60 (s, 2H), 7.97 (s, 1H).
Step 3: Preparation of Intermediate tert-butyl 3-(hydroxymethyl)-6,7-dihydro-4H-thieno[3,2-c]pyridine-5-carboxylate (7c)
(53) A suspension of intermediate (7b) (140 mg, 0.45 mmol) and lithium borohydride (40 mg, 1.8 mmol) in THF (0.5 mL) and toluene (1 mL) was heated at 100° C. for 1 h. The mixture was then cooled to room temperature and concentrated. The crude was purified by column chromatography on silica gel (heptane/ethyl acetate: 1/1) to give intermediate (7c) as colorless oil (90 mg, 0.33 mmol, 75%). MS m/z ([M+Na].sup.+) 292.
Step 4: Preparation of Intermediate tert-butyl 3-[(2,2-dimethoxyethylamino)methyl]-6,7-dihydro-4H-thieno[3,2-c]pyridine-5-carboxylate (7d)
(54) At 0° C., to a solution of intermediate (7c) (900 mg, 3.66 mmol) in DCM (20 mL) were successively added TEA (1.5 mL, 10.98 mmol) and methanesulfonic anhydride (1.27 g, 7.33 mmol). The mixture was stirred 2 h at 0° C. Then, a solution of dimethoxyethylamine (2.4 mL, 21.96 mmol) in DCM (30 mL) was added dropwise. The mixture was stirred for 30 minutes then 16 h at rt. The mixture was diluted with DCM, washed with water and extracted twice with DCM. Organics were dried over Na.sub.2SO.sub.4, filtered and concentrated. The crude product was purified by column chromatography on silica gel (DCM/MeOH: 1/1) to give intermediate (7d) (0.84 g, 2.36 mmol, 64%). MS m/z ([M+H].sup.+) 357.
Step 5: Preparation of Intermediate 2-hydroxy-2,3,4,5,6,7,8,9-octahydrothieno[3,2-c:4,5-c]dipyridinium dihydrochloride (7e)
(55) A solution of intermediate (7d) (0.840 g, 2.36 mmol) in HCl 6N (15 mL) was stirred for 1 h30 at 60° C. The mixture was concentrated to give intermediate (7e) (1.02 g) which was used without further purification. MS m/z ([M+H].sup.+) 211.
Step 6: Preparation of Intermediate 4,7-di-tert-butyl 2-hydroxy-2,3,8,9-tetrahydro-5,6H-thieno[3,2-c:4,5-c]dipyridine-4,7-dicarboxylate (7f)
(56) To a solution of intermediate (7e) (1.02 g, 2.36 mmol) in DCM (40 mL) under argon atmosphere were successively added Boc.sub.2O (2.7 g, 12.75 mmol) and DIPEA (4.95 mL, 28.32 mmol). After stirring for 2 days at rt, the mixture was concentrated. The residue was purified by column chromatography on silica gel (heptane/EtOAc: 1/1) to give intermediate (7f) as colorless oil (0.30 g, 0.73 mmol, 31%). MS m/z=([M+Na.sup.+]) 433.
Step 7: Preparation of Intermediate 4,7-di-tert-butyl 2-(N-allyloxy)-2,3,8,9-tetrahydro-5,6H-thieno[3,2-c:4,5-c]dipyridine-4,7-dicarboxylate (7 g)
(57) At −78° C., methanesulfonic anhydride (0.382 g, 2.19 mmol) was added to a solution of intermediate (7f) (0.30 g, 0.73 mmol) and TEA (0.408 mL, 2.92 mmol) in DCM (2 mL). After stirring for 1 h20 at −78° C., 0-allylhydroxylamine (0.534 g, 7.3 mmol) in DCM (2 mL) was added dropwise. The mixture was stirred for 20 min at −78° C. then 1 h at rt. The mixture was diluted with DCM, washed twice with water, dried over Na.sub.2SO.sub.4, filtered and concentrated. The crude was purified by column chromatography on silica gel (Heptane/EtOAc: 1/1) to give intermediate (7 g) (0.15 g, 0.32 mmol, 44%). MS m/z ([M+H].sup.+) 466.
Step 8: Preparation of 4-allyloxy-2,5-methano-1,5,7,8,9,10-hexahydropyrido[2,3]thieno[2,4-d][1,3]diazepin-3-one (7 h)
(58) To a solution of intermediate (7 g) (0.15 g, 0.32 mmol) in DCM (2.5 mL) were successively added TEA (0.44 mL, 1.18 mmol) and triphosgene (49 mg, 0.46 mmol). The mixture was stirred for 5 min at rt. The mixture was cooled to 0° C. and methanesulfonic acid (0.62 mL, 3.9 mmol) was then added. The mixture was stirred for 1 h at rt. The mixture was cooled to 0° C. and TEA (0.73 mL, 1.95 mmol) was added. The mixture was stirred at rt for 1 h. The mixture was then diluted with DCM, washed with aq. NaHCO.sub.3, dried over Na.sub.2SO.sub.4, filtered and concentrated. The crude was purified by column chromatography on silica gel (DCM/Acetone: 9/1) to give intermediate (7 h) as white powder (0.09 g, 0.31 mmol, 72%). MS m/z ([M+H].sup.+) 292.
(59) .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ (ppm): 2.98 (t, J=6.1 Hz, 2H), 3.14 (q, J=7.0 Hz, 1H), 3.25 (d, J=11.0 Hz, 1H), 3.40 (dt, J=5.7, 12.0 Hz, 1H), 3.54 (td, J=3.0, 11.2 Hz, 1H), 3.88-4.13 (m, 2H), 4.13-4.25 (m, 2H), 4.38 (d, J=6.0 Hz, 2H), 4.70-4.77 (m, 1H), 5.25 (dq, J=11.3, 10.5 Hz, 1H), 5.34 (dq, J=1.6, 17.3 Hz, 1H), 5.87-5.98 (m, 1H), 9.05 (bs, 1H).
Step 9: Preparation of Intermediates 4-allyloxy-N,N-diethyl-2,5-methano-3-oxo-5,7,8,10-tetrahydro-1H-pyrido[3,4]thieno[1,3-d][1,3]diazepine-9-carboxamide (7i) and tert-butyl 4-allyloxy-2,5-methano-3-oxo-5,7,8,10-tetrahydro-1H-pyrido[3,4]thieno[1,3-d][1,3]diazepine-9-carboxylate (7j)
(60) To a solution of intermediate (7 h) (90 mg, 0.3 mmol) in DCM (4 mL) were successively added Boc.sub.2O (120 mg, 0.55 mmol) and DIPEA (0.21 mL, 1.2 mmol). After stirring for 12 h at rt, the mixture was diluted with DCM. Organics were washed with water, dried over Na.sub.2SO.sub.4, filtered and concentrated. The residue was purified by column chromatography on silica gel (DCM/Acetone: 95/5) to intermediates (7i) (0.05 g, 0.13 mmol, 42%) and (7j) (0.04 g, 0.10 mmol, 34%).
(61) Intermediate (7i): MS m/z ([M+H].sup.+) 391; .sup.1H NMR (400 MHz, CDCl.sub.3) δ (ppm): 1.13 (t, J=7.1 Hz, 6H), 2.84 (dtt, J=1.9, 3.9, 5.9 Hz, 2H), 3.16-3.27 (m, 5H), 3.37-3.54 (m, 2H), 3.71 (dd, J=3.1, 10.9 Hz, 1H), 3.99-4.12 (m, 3H), 4.27 (d, J=16.5 Hz, 1H), 4.36-4.49 (m, 3H), 5.25-5.32 (m, 1H), 5.36 (dq, J=1.4, 17.2 Hz, 1H), 6.01 (ddt, J=6.3, 10.3, 16.9 Hz, 1H).
(62) Intermediate (7j): MS m/z ([M+H].sup.+) 392; .sup.1H NMR (400 MHz, CDCl.sub.3) δ (ppm): 1.46 (s, 9H), 2.75 (d, J=6.2 Hz, 2H), 3.21 (d, J=10.8 Hz, 1H), 3.71 (dd, J=3.1, 10.9 Hz, 2H), 4.05-4.31 (m, 4H), 4.34-4.49 (m, 4H), 5.26-5.31 (m, 1H), 5.35 (dq, J=1.5, 17.2 Hz, 1H), 5.90-6.12 (m, 1H).
Step 10: Preparation of Intermediate N,N-diethyl-4-hydroxy-2,5-methano-3-oxo-5,7,8,10-tetrahydro-1H-pyrido[3,4]thieno[1,3-d][1,3]diazepine-9-carboxamide (7k)
(63) A solution of intermediate (7i) (0.050 g, 0.1128 mmol) in DCM (4 mL) was degassed 5 min under argon atmosphere. Acetic acid (0.030 mL, 0.512 mmol) and Pd(PPh.sub.3).sub.4 (0.1035 g, 0.09 mmol) were successively added. After stirring for 1 h at rt, the mixture was concentrated under nitrogen flux. The residue was purified on silica gel (DCM to DCM/acetone 8/2) to give intermediate (7k) (0.020 g, 0.057 mmol, 45%). MS m/z ([M+H].sup.+) 351.
Step 11: Preparation of sodium [9-(diethylcarbamoyl)-2,5-methano-3-oxo-5,7,8,10-tetrahydro-1H-pyrido[3,4]thieno[1,3-d][1,3]diazepin-4-yl] sulfate, Example 7
(64) To a solution of intermediate (7k) (0.020 g, 0.057 mmol) in anhydrous pyridine (2.5 mL) was added sulfur trioxide pyridine complex (0.102 g, 0.64 mmol). After stirring for 18 h, the heterogeneous mixture was concentrated under nitrogen flux. DCM was added and the insoluble was filtered off. The filtrate was concentrated and the residue was applied on a Dowex sodium form column (Dowex® 50WX8 hydrogen form stored with an aqueous solution of 2N NaOH and washed until neutral pH with water). The fractions containing the desired compound were combined, frozen and lyophilized. The solid was purified on C18 reversed-phase chromatography (H2O/MeCN 99/1 up to 90/10). The fractions containing the desired compound were combined, frozen and lyophilized to give Example 7 (0.0043 g, 0.009 mmol, 17%). MS m/z ([M+H].sup.+) 431. MS m/z ([M−H].sup.−) 429. .sup.1H NMR (400 MHz, D.sub.2O) δ (ppm): 1.13 (t, J=7.1 Hz, 6H), 2.88-2.95 (m, 2H), 3.28 (q, J=7.1 Hz, 4H), 3.47-3.60 (m, 3H), 3.90 (dd, J=3.1, 11.4 Hz, 1H), 4.10-4.22 (m, 2H), 4.26-4.42 (m, 2H), 4.96 (d, J=2.9 Hz, 1H).
Example 8: Synthesis of (2,5-methano-3-oxo-1,5,7,8,9,10-hexahydropyrido[2,3]thieno[2,4-d][1,3]diazepin-4-yl) hydrogen Sulfate
(65) ##STR00029##
Step 1: Preparation of Intermediate tert-butyl 4-hydroxy-2,5-methano-3-oxo-5,7,8,10-tetrahydro-1H-pyrido[3,4]thieno[1,3-d][1,3]diazepine-9-carboxylate (8a)
(66) A solution of intermediate (7j) (0.040 g, 0.10 mmol) in DCM (4 mL) was degassed 5 min under argon atmosphere. Acetic acid (0.023 mL, 0.40 mmol) and Pd(PPh.sub.3).sub.4 (0.116 g, 0.10 mmol) were successively added. After stirring for 1 h at rt, the mixture was concentrated under nitrogen flux. The residue was purified on silica gel (DCM to DCM/acetone 8/2) to give intermediate (8a) (0.035 g, 0.01 mmol, 100%). MS m/z ([M+H].sup.+) 352.
Step 2: Preparation of Intermediate tert-butyl 2,5-methano-3-oxo-4-sulfooxy-5,7,8,10-tetrahydro-1H-pyrido[3,4]thieno[1,3-d][1,3]diazepine-9-carboxylate (8b)
(67) To a solution of intermediate (8a) (35 mg, 0.01 mmol) in anhydrous pyridine (2.5 mL) was added sulfur trioxide pyridine complex (0.079 g, 0.50 mmol). After stirring for 18 h, the heterogeneous mixture was concentrated under nitrogen flux. DCM was added and the insoluble was filtered off. The filtrate was concentrated. The residue was purified by column chromatography on silica gel (DCM/acetone 8/2 to 2/8) to give intermediate (8b) (18.5 mg, 0.04 mmol, 43%). MS m/z ([M−H].sup.−) 430.
Step 3: Preparation of (2,5-methano-3-oxo-1,5,7,8,9,10-hexahydropyrido[2,3]thieno[2,4-d][1,3]diazepin-4-yl) hydrogen sulfate, Example 8
(68) At 0° C., acetic acid (1 mL) was added to a solution of intermediate (8b) in DCM (1 ml. The mixture was stirred at 0° C. for 30 minutes. Heptane (4 mL) was added and mixture was concentrated. The residue was triturated four times with ACN (2 mL). The solid was filtered and dried under vacuum to give Example 8 (0.004 g, 0.015 mmol, 37%). MS m/z ([M+H].sup.+) 332. MS m/z ([M−H].sup.−) 330. .sup.1H NMR (400 MHz, D.sub.2O) δ (ppm): 3.16-3.22 (m, 2H), 3.51-3.68 (m, 3H), 3.91 (dd, J=3.2, 11.5 Hz, 1H), 4.13-4.20 (m, 2H), 4.30 (d, J=16.6 Hz, 1H), 4.38 (d, J=16.6 Hz, 1H), 4.99 (d, J=2.9 Hz, 1H).
Example 9: Synthesis of sodium (6,9-methano-4-methyl-7-oxo-5,9-dihydrothiazolo[3,4]pyrrolo[1,3-d][1,3]diazepin-8-yl) sulfate
(69) ##STR00030## ##STR00031## ##STR00032##
Step 1: Preparation of Intermediate 4H-pyrrolo[2,3-d]thiazol-5-ylmethanol (9a)
(70) At 0° C., LiAlH4 (2M in THF, 19.11 mL) was slowly added to a solution of ethyl 4H-pyrrolo[2,3-d][1,3]thiazole-5-carboxylate (5 g, 25.48 mmol) in THF (100 mL). The mixture was stirred at 0° C. for 5 h. The mixture was quenched with NaOH 2N (4.5 mL). The precipitate was filtered off and washed with Et.sub.2O. The filtrate was concentrated to give intermediate (9a) which was used without further purification (2.4 g, 15.56 mmol, 61%). MS m/z ([M+H].sup.+) 155. .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 4.49 (dd, J=0.6, 5.5 Hz, 2H), 5.08 (t, J=5.5 Hz, 1H), 6.27 (d, J=1.8 Hz, 1H), 8.66 (s, 1H), 11.75 (bs, 1H).
Step 2: Preparation of Intermediate 4H-pyrrolo[2,3-d]thiazole-5-carbaldehyde (9b)
(71) At rt, a solution of intermediate (9a) (2.4 g, 15.56 mmol) and manganese oxide(II) (9.47 g, 108.96 mmol) in THF (100 mL) was stirred for 16 h. The mixture was filtered on a pad of Célite® which was washed with THF. The filtrate was concentrated to give intermediate (9b) which was used without further purification. MS m/z ([M+H].sup.+) 153.
Step 3: Preparation of Intermediate N-(2,2-diethoxyethyl)-1-(4H-pyrrolo[2,3-d]thiazol-5-yl)methanimine (9c)
(72) Intermediate (9b) was solubilized in THF (50 mL) and stirred in presence of aminoacetaldehyde diethyl acetal (3.39 mL, 23.35 mmol) and magnesium sulfate (13.1 g, 108.96 mmol) for 24 h. The mixture was filtered on a pad of Célite®. The filtrate containing intermediate (9c) was directly used without further work-up.
Step 4: Preparation of Intermediate 2,2-diethoxy-N-(4H-pyrrolo[2,3-d]thiazol-5-ylmethyl)ethanamine (9d)
(73) The filtrate of step 3 containing intermediate (9c) was diluted with ethanol (10 mL). Sodium borohydride (883 mg, 23.35 mmol) was slowly added at 0° C. The mixture was stirred for 2 h, then quenched with acetone and water and concentrated. The crude was purified by chromatography on silica gel (DCM/Acetone: 7/3 to 3/7) to give intermediate (9d) (1.05 g, 3.89 mmol, 25%). MS m/z ([M+H].sup.+) 270. .sup.1H NMR (400 MHz, CDCl.sub.3) δ 1.19-1.23 (m, 6H), 2.78 (d, J=5.3 Hz, 2H), 3.39 (d, J=3.9 Hz, 1H), 3.49-3.74 (m, 4H), 3.95 (s, 2H), 4.59 (d, J=5.3 Hz, 1H), 6.25 (s, 1H), 8.44 (s, 1H), 9.63 (bs, 1H).
Step 5: Preparation of Intermediate 5,6,7,8-tetrahydro-4H-thiazolo[3,4]pyrrolo[1,3-b]pyridin-8-ol hydrochloride salt (9e)
(74) A solution of intermediate (9d) (1.05 g, 3.9 mmol) in HCl 4N (12 mL) was stirred at rt for 3 days. The mixture was then concentrated by co-evaporation with EtOH. The crude containing intermediate (9e) as hydrochloride salt was used in the next step without further purification.
Step 6: Preparation of Intermediate tert-butyl 8-hydroxy-4,5,7,8-tetrahydrothiazolo[3,4]pyrrolo[1,3-b]pyridine-6-carboxylate (9f)
(75) The crude containing intermediate (9e) obtained in step 5 was suspended in DCM (50 mL). TEA (8.8 mL, 63.36 mmol) was added and then Boc.sub.2O (851 mg, 3.9 mmol). The mixture was stirred at rt for 10 min. The mixture was washed with NaHCO.sub.350% aq, dried over Na.sub.2SO.sub.4, filtered and concentrated. The crude containing intermediate (9f) was used in the next step without further purification.
Step 7: Preparation of Intermediate tert-butyl 8-[tert-butyl(dimethyl)silyl]oxy-4,5,7,8-tetrahydrothiazolo[3,4]pyrrolo[1,3-b]pyridine-6-carboxylate (9 g)
(76) The crude containing intermediate (9f) obtained in step 6 was suspended in DMF (10 mL). TBDMSCl (705 mg, 4.7 mmol) and imidazole (531 mg, 7.8 mmol) were added and the mixture was stirred for 16 h at rt. The mixture was diluted with AcOEt (60 mL), washed with NaCl 25% (6×10 mL), dried over Na.sub.2SO.sub.4, filtered and concentrated. The crude was purified by column chromatography on silica gel (eluent: n-heptane/AcOEt 7/3 isochratic) to give intermediate (9 g) (1.1 g, 2.68 mmol, 69% over steps 5 to 7). MS m/z ([M+H].sup.+) 410. .sup.1H NMR (400 MHz, CDCl.sub.3) δ 0.22 and 0.25 (2s, 6H), 0.97 (s, 9H), 1.51 (s, 9H), 3.29-3.39 (m, 1H), 4.01-4.15 (m, 1H), 4.51 (d, J=16.1 Hz, 1H), 4.68-4.81 (m, 1H), 4.93 (bs, 1H), 8.08 and 8.44 (2s, 1H), 9.07 and 9.40 (2bs, 1H).
Step 8: Preparation of Intermediate tert-butyl 8-[tert-butyl(dimethyl)silyl]oxy-4-methyl-7,8-dihydro-5H-thiazolo[3,4]pyrrolo[1,3-b]pyridine-6-carboxylate (9 h)
(77) At rt, sodium hydride 60% in mineral oil (161 mg, 4.03 mmol) was added to a solution of intermediate (9 g) (1.1 g, 2.69 mmol) in DMF (5 mL). The mixture was stirred for 15 min then methyl iodide (502 μL, 8.06 mmol) was added. The mixture was stirred for 1 h40. The mixture was quenched with aq. NaCl 25% and diluted with AcOEt. The organic layer was washed with aq. NaCl 25% (4×10 mL), dried over Na.sub.2SO.sub.4, filtered and concentrated. The crude containing intermediate (9 h) was used in the next step without further purification.
Step 9: Preparation of Intermediate tert-butyl 8-hydroxy-4-methyl-7,8-dihydro-5H-thiazolo[3,4]pyrrolo[1,3-b]pyridine-6-carboxylate (9i)
(78) The crude containing intermediate (9 h) obtained in step 8 was diluted with THF (5 mL) and a solution of tetrabutylammonium fluoride 1M in THF (3.22 mL, 3.22 mmol) was added. The mixture was stirred for 20 min. The mixture was hydrolyzed with water (50 μL) and concentrated. The crude was purified by column chromatography on silica gel (eluent: n-heptane/AcOEt 5/5 isochratic) to give intermediate (9i) (650 mg, 2.1 mmol, 78% over steps 8 and 9). MS m/z ([M+H].sup.+) 310. .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 1.44 (s, 9H), 3.35 (bs, 1H), 3.71 (s, 3H), 3.75-3.79 (m, 1H), 4.48-4.57 (m, 2H), 4.63 (q, J=5.9 Hz, 1H), 5.26 (d, J=6.1 Hz, 1H), 8.70 (s, 1H).
Step 10: Preparation of Intermediate tert-butyl 8-[[tert-butyl(dimethyl)silyl]oxyamino]-4-methyl-7,8-dihydro-5H-thiazolo[3,4]pyrrolo[1,3-b]pyridine-6-carboxylate (9j)
(79) At −78° C., a solution of methanesulfonic anhydride (1.098 g, 6.3 mmol) in DCM (8 mL) was dropped to a solution of intermediate (9i) (650 mg, 2.1 mmol) in DCM (24 mL). The mixture was stirred at −78° C. for 1 h. At −78° C., a solution of O-(tert-Butyldimethylsilyl)hydroxylamine (2.47 g, 16.8 mmol) in DCM (8 mL) was dropped to the mixture which was then warmed to rt for 2 h. The mixture was washed with Na.sub.2CO.sub.3 50%, dried over Na.sub.2SO.sub.4, filtered and concentrated. The crude was purified by column chromatography on silica gel (n-heptane/AcOEt 7/3 isochratic) to give intermediate (9j) (730 mg, 1.66 mmol, 79%). MS m/z ([M+Na].sup.+) 461. .sup.1H NMR (400 MHz, CDCl.sub.3) δ 0.15 and 0.17 (2, 6H), 0.97 (s, 9H), 1.50 (s, 9H), 3.30-3.44 (m, 1H), 3.75 (s, 3H), 3.96-4.09 (m, 1H), 4.26-4.44 (m, 2H), 4.89-4.94 (m, 1H), 8.42 (s, 1H).
Step 11: Preparation of Intermediate N-[tert-butyl(dimethyl)silyl]oxy-4-methyl-5,6,7,8-tetrahydrothiazolo[3,4]pyrrolo[1,3-b]pyridin-8-amine (9k)
(80) At rt, zinc(II) bromide (1.5 g, 6.65 mmol) was added to a solution of intermediate (9j) (730 mg, 1.66 mmol) in DCM (20 mL). The mixture was stirred at rt for 48 h. The mixture was filtered and the solid was diluted with water. The solution was basified with NaHCO.sub.3 and extracted twice with DCM. Organics were dried over Na.sub.2SO.sub.4, filtered and concentrated. The crude was purified by column chromatography on silica gel (eluent: DCM/MeOH 95/5 isochratic) to give intermediate (9k) (330 mg, 0.97 mmol, 59%). MS m/z ([M+H].sup.+) 339. .sup.1H NMR (400 MHz, CDCl.sub.3) δ 0.10 and 0.11 (2, 6H), 0.93 (s, 9H), 2.63 (bs, 1H), 2.77 (dd, J=3.2, 13.5 Hz, 1H), 3.15 (d, J=13.5 Hz, 1H), 3.64 (s, 3H), 3.71-3.75 (m, 2H), 3.83 (d, J=16.1 Hz, 1H), 6.09 (d, J=11.4 Hz, 1H), 8.62 (s, 1H).
Step 12: Preparation of Intermediate 8-[tert-butyl(dimethyl)silyl]oxy-6,9-methano-4-methyl-5,9-dihydrothiazolo[3,4]pyrrolo[1,3-d][1,3]diazepin-7-one (91)
(81) At 0° C., intermediate (9k) (330 mg, 0.975 mmol) and DIPEA (679 μL, 3.9 mmol) were solubilized in ACN (7 mL). A solution of triphosgene (101 mg, 0.341 mmol) in ACN (1 mL) was slowly added at 0° C. and the mixture was then stirred at rt for the night. The mixture was concentrated. The crude was solubilized in DCM and washed with NaHCO.sub.325%, dried over Na.sub.2SO.sub.4, filtered and concentrated. The crude was purified by column chromatography on silica gel (eluent: DCM/acetone 95/5 to 45/55) to give intermediate (91) (65 mg, 0.178 mmol, 18%). MS m/z ([M+H].sup.+) 365. .sup.1H NMR (400 MHz, CDCl.sub.3) δ 0.20 (s, 3H), 0.21 (s, 3H), 0.94 (s, 9H), 3.27 (d, J=10.8 Hz, 1H), 3.70 (s, 3H), 3.71 (dd, J=2.9, 10.8 Hz, 1H), 4.35 (d, J=16.1 Hz, 1H), 4.36 (d, J=2.8 Hz, 1H), 4.56 (d, J=16.1 Hz, 1H), 8.38 (s, 1H).
Step 13: Preparation of Intermediate 8-hydroxy-6,9-methano-4-methyl-5,9-dihydrothiazolo[3,4]pyrrolo[1,3-d][1,3]diazepin-7-one (9m)
(82) At rt, a solution of tetrabutylammonium fluoride 1N in THF (123 μL, 0.123 mmol) was added to a solution of intermediate (91) (45 mg, 0.123 mmol) and formic acid (9 μL, 0.247 mmol) in AcOEt (2 mL). A white precipitate was appeared after 10 min. The precipitate was filtered, washed with AcOEt and dried under vacuum to give intermediate (9m) as white powder (23 mg, 0.091 mmol, 75%). MS m/z ([M+H].sup.+) 251.
Step 14: Preparation of Intermediate n-tetrabutylammonium (6,9-methano-4-methyl-7-oxo-5,9-dihydrothiazolo[3,4]pyrrolo[1,3-d][1,3]diazepin-8-yl) sulfate (9n)
(83) At rt, a suspension of intermediate (9m) (23 mg, 0.091 mmol) and sulfur trioxide pyridine complex (98 mg, 0.617 mmol) in pyridine (5 mL) was stirred for 4 h. The mixture was concentrated. The crude was suspended in DCM and filtered. The solid was washed with DCM and dried under vacuum. The solid was solubilized in water (5 mL) and tetrabutylammonium hydrogensulfate (42 mg, 0.123 mmol) was added for pyridinium/tetrabutylammonium cation exchange. Organics were extracted twice with DCM, dried over Na.sub.2SO.sub.4, filtered and concentrated to give intermediate (9n) as tetrabutylammonium salt which was used in the next step without further purification.
Step 15: Preparation of sodium (6,9-methano-4-methyl-7-oxo-5,9-dihydrothiazolo[3,4]pyrrolo[1,3-d][1,3]diazepin-8-yl) sulfate, Example 9
(84) Intermediate (9n) obtained in step 14 was applied on a Dowex sodium form column (Dowex® 50WX8 hydrogen form stored with an aqueous solution of 2N NaOH and washed until neutral pH with H.sub.2O). Fractions of interest are combined, concentrated under nitrogen flux to remove ACN, frozen and lyophilized to give Example 9 as sodium salt (6 mg, 0.017 mmol, 14%). MS m/z ([M−H].sup.−) 329. .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 3.28 (d, J=10.9 Hz, 1H), 3.54 (dd, J=3.0, 10.9 Hz, 1H), 3.66 (s, 3H), 4.37 (d, J=16.3 Hz, 1H), 4.44 (d, J=16.3 Hz, 1H), 4.79 (d, J=3.0 Hz, 1H), 8.70 (s, 1H).
(85) Biological Activity
Method 1: β-Lactamase Inhibitory Activity, Determination of IC.SUB.50 .(Table 1)
(86) Enzyme activity was monitored by spectrophotometric measurement of nitrocefin (NCF—TOKU-E, N005) hydrolysis at 485 nm, at room temperature and in assay buffer A: 100 mM Phosphate pH7, 2% glycerol and 0.1 mg/mL Bovine serum albumin (Sigma, B4287). Enzymes were cloned in E. coli expression vector, expressed and purified in house using classical procedures. To a transparent polystyrene plate (Corning, 3628) were added in each well 5 μL DMSO or inhibitor dilutions in DMSO and 80 μL enzyme in buffer A. Plates were immediately read at 485 nm in a microplate spectrophotometer (BioTek, PowerWave HT) to enable background subtraction. After 30 min of pre-incubation at room temperature, 15 μL of NCF (200 μM final) were finally added in each well. Final enzyme concentrations were 0.1 nM (TEM-1), 0.075 nM (SHV-1), 0.4 nM (CTX-M-15), 1 nM (KPC-2), 0.2 nM (P99 AmpC), 0.2 nM (CMY-37), 0.4 nM (AmpC P. aeruginosa), 0.2 nM (OXA-1), 1.2 nM (OXA-11), 0.4 nM (OXA-15) and 0.3 nM (OXA-48). After 20 min incubation at room temperature, plates were once again read at 485 nm. Enzyme activity was obtained by subtracting the final signal by the background, and was converted to enzyme inhibition using non inhibited wells. IC.sub.50 curves were fitted to a classical Langmuir equilibrium model with Hill slope using XLFIT (IDBS).
(87) TABLE-US-00001 TABLE 1 IC.sub.50 (μM) for β-lactamase Inhibitory Activity BLA IC50 compounds of the invention (μM) (A) (C) CTX- AmpC AmpC (D) Example TEM-1 SHV-1 M-15 KPC-2 (P99) CMY-37 (PAE) OXA-1 OXA-11 OXA-15 OXA-48 1 0.00065 0.0041 0.0010 0.026 0.017 0.013 0.0088 0.12 0.041 3.4 0.0011 2 0.00022 0.00043 0.0026 0.00094 0.00051 0.0012 0.0025 0.037 0.0022 0.83 0.00066 3 0.0013 0.0022 0.0025 0.0093 0.0045 0.0094 0.062 0.19 0.031 7.1 0.0048 4 0.036 0.15 0.0099 0.37 0.12 0.36 2.8 4.1 0.59 13 0.0015 5 0.014 0.077 0.014 0.16 0.10 0.81 3.6 1.7 0.19 1.8 0.0010 6 0.00020 0.00025 0.0015 0.0042 0.0020 0.0043 0.0036 1.3 0.16 6.5 0.0046 7 0.043 0.013 0.0013 0.026 0.078 0.18 0.55 5.1 0.11 14 0.0068 8 0.0096 0.026 0.0061 1.3 1.6 1.8 8.6 15 0.84 30 0.018 9 0.00049 0.00057 0.0011 0.026 0.12 0.089 0.59 0.29 0.062 0.94 0.0012
Method 2: MIC of Compounds and Synergy with Ceftazidime Against a Bacterial Isolate (Tables 3 and 4)
(88) Compounds of the present invention were assessed against a genotyped bacterial strain alone or in combination with the β-lactam ceftazidime. In the assays, MICs of said compounds, or of ceftazidime at fixed concentrations of said compounds were determined by the broth microdilution method according to the Clinical Laboratory Standards Institute (CLSI—M7-A7). Briefly, compounds alone according to the invention were prepared in DMSO and spotted (2 μL each) on sterile polystyrene plates (Corning, 3788). Compounds and ceftazidime dilutions were prepared in DMSO and spotted (1 μL each) on sterile polystyrene plates (Corning, 3788). Log phase bacterial suspensions were adjusted to a final density of 5×10.sup.5 cfu/mL in cation-adjusted Mueller-Hinton broth (Becton-Dickinson) and added to each well (98 μL). Microplates were incubated for 16-20 h at 35° C. in ambient air. The MIC of the compounds was defined as the lowest concentration of said compounds that prevented bacterial growth as read by visual inspection. The MIC of ceftazidime at each compound concentration was defined as the lowest concentration of ceftazidime that prevented bacterial growth as read by visual inspection.
(89) TABLE-US-00002 TABLE 2 Bacterial isolate used in MIC determination Strain Resistance mechanism E. coli ECO 190317 TEM-1, SHV-12, CTX-M-15, OXA-1
(90) TABLE-US-00003 TABLE 3 MIC of Ceftazidime (CAZ) and compounds alone (μg/mL) MIC compounds of the invention alone (μg/mL) Compounds ECO 190317 CAZ 128 1 16 2 >32 3 8 4 8 5 4 6 >32 7 >32 8 1 9 >32
(91) TABLE-US-00004 TABLE 4 MIC of Ceftazidime (CAZ) alone and CAZ/compound combinations Combination of CAZ and compounds of the invention at 4 μg/mL: MIC (μg/mL) Compounds ECO 190317 CAZ 128 CAZ + 1 1 CAZ + 2 4 CAZ + 3 32 CAZ + 4 <=0.125 CAZ + 5 <0.125 CAZ + 6 8 CAZ + 7 64 CAZ + 8 <0.125 CAZ + 9 64