DERIVATIVES OF 5-PHENYL- OR 5-HETEROARYLATHIAZOL-2-CARBOXYLIC AMIDE USEFUL FOR THE TREATMENT OF INTER ALIA CYSTIC FIBROSIS

Abstract

The present disclosure is based, in part, on the discovery that disclosed compounds such as those having Formula (IIIa), (III), or (IV) can increase cystic fibrosis transmembrane conductance regulator (CFTR) activity as measured in human bronchial epithelial (hBE) cells.

Claims

1. A compound represented by formula III or IV: ##STR00046## and pharmaceutically acceptable salts, stereoisomers, and prodrugs thereof, wherein: X.sub.3 is selected from the group consisting of O, S, and NR.sub.hh; pp is 1, 2, or 3; R.sub.11 is independently selected for each occurrence from the group consisting of hydrogen, halogen, and C.sub.1-4alkyl (optionally substituted by one, two or three halogens); L.sub.1 is selected from the group consisting of C.sub.1-6alkylene, C.sub.3-6cycloalkylene, C.sub.3-6cycloalkylene-C.sub.1-4alkylene, C.sub.1-3alkylene-NR.sub.hh—S(O).sub.w—, —C.sub.1-3alkylene-S(O).sub.w—NR.sub.hh—, C.sub.3-6cycloalkylene-C.sub.0-2alkylene-S(O).sub.w—NR.sub.hh, and C.sub.3-6cycloalkylene-C.sub.0-2alkylene NR.sub.hh—S(O).sub.w—, wherein L.sub.1 may be optionally substituted by one, two or three substituents selected from the group consisting of halogen, hydroxyl, C.sub.1-3alkyl (optionally substituted by one, two or three substituents each selected independently from R.sub.ff); R.sub.44 is selected from the group consisting of H, halogen, hydroxyl, C.sub.1-3alkoxy, heterocycle, and a 5-6 membered monocyclic or 8-10 membered bicyclic heteroaryl having one, two or three heteroatoms each selected from O, N, and S; wherein the heterocycle and the heteroaryl may be optionally substituted by one or two substituents each selected independently from R.sub.gg; R.sub.ff is selected for each occurrence from group consisting of halogen, hydroxyl, C.sub.1-4alkyl, C.sub.1-4alkyoxy, C.sub.2-4alkenyl, C.sub.3-6cycloalkyl, —NR′R″, —NR′—S(O).sub.w—C.sub.1-3alkyl, S(O).sub.w—NR′R″, and —S(O).sub.w—C.sub.1-3alkyl, where w is 0, 1, or 2, wherein C.sub.1-4alkyl, C.sub.1-4alkyoxy, C.sub.2-4alkenyl and C.sub.3-6cycloalkyl may be optionally substituted by one, two or three substituents each independently selected from the group consisting of halogen, hydroxyl, —NR′R″, —NR′—S(O).sub.w—C.sub.1-3alkyl, S(O).sub.w—NR′R″, and —S(O).sub.w—C.sub.1-3alkyl; R.sub.gg is selected for each occurrence from group consisting of halogen, hydroxyl, C.sub.1-6alkyl, C.sub.1-6alkyoxy, C.sub.2-6alkenyl, C.sub.3-6cycloalkyl, —NR′R″, —NR′—S(O).sub.w—C.sub.1-3alkyl, S(O).sub.w—NR′R″, and —S(O).sub.w—C.sub.1-3alkyl, where w is 0, 1, or 2, wherein C.sub.1-6alkyl, C.sub.1-6alkyoxy, C.sub.2-6alkenyl and C.sub.3-6cycloalkyl may each be optionally substituted by one, two or three substituents each independently selected from the group consisting of halogen, C.sub.1-6alkyl, C.sub.1-6alkoxy, hydroxyl, C(O)OH, —C(O)OC.sub.1-6alkyl, —O—C.sub.3-6cycloalkyl, —O-heterocycle, —O-heteroaryl, —O-phenyl, —NR′R″, —NR′—S(O).sub.w—C.sub.1-3alkyl, S(O).sub.w—NR′R″, and —S(O)w-C.sub.1-3alkyl; w is 0, 1 or 2; and R.sub.hh is selected for each occurrence from the group consisting of H, C.sub.1-6alkyl and C.sub.3-6cycloalkyl.

2. The compound of claim 1, wherein L.sub.1 is C.sub.1-3alkylene or C.sub.3-5cycloalkylene.

3. The compound of claim 1 or 2, represented by: ##STR00047## wherein qq is 0 or 1.

4. The compound of any one of claims 1-4, represented by: ##STR00048##

5. The compound of any one of claims 1-4, wherein R.sub.44 is selected from the group consisting of: pyrrolidinyl, piperidinyl, tetrahydropyranyl, and tetrahydofuranyl.

6. The compound of any one of claims 1-4, wherein R.sub.44 is selected from the group consisting of: ##STR00049## wherein X independently for each occurrence is selected from the group consisting of O, S, NR.sub.hh, C, C(R.sub.88), and C(R.sub.88)(R.sub.99); X.sub.2 independently for each occurrence is selected from the group consisting of O, S and NR.sub.hh; R″ is H or C.sub.1-4alkyl, each R.sub.66, R.sub.77, R.sub.88 and R.sub.99 is independently selected for each occurrence from H and R.sub.gg, and n is 0, 1, 2, or 3.

7. The compound of claim 6, where each R.sub.66, R.sub.77, R.sub.88 and R.sub.99 is independently selected for each occurrence from the group consisting of hydrogen, halogen, hydroxyl, C.sub.1-6 alkyl, C.sub.3-6 cycloalkyl, and heterocycle, wherein C.sub.1-6 alkyl, C.sub.3-6 cycloalkyl, and heterocycle are optionally substituted by one, two or three substituents each independently selected from the group consisting of hydroxyl, C.sub.1-6 alkyl, C.sub.1-6 alkoxy (optionally substituted by C.sub.3-6cycloalkyl, heterocycle, —C.sub.1-2alkyl-heterocycle and C.sub.1-2alkyl-C.sub.3-6cycloalkyl), —S(O).sub.w—C.sub.1-3 alkyl (w is 0, 1, or 2) and —NR′S(O).sub.2C.sub.1-6 alkyl; and R′ is independently selected for each occurrence from H and C.sub.1-4 alkyl.

8. The compound of any one of claims 1-7, wherein pp is 0, 1 or 2, and R.sub.11 is selected from H, F, or methyl.

9. A compound having the Formula (IIIa): ##STR00050## or a pharmaceutically acceptable salt, prodrug or solvate thereof, wherein: R.sub.1 is selected from the group consisting of: ##STR00051## R.sub.2 is selected from the group consisting of optionally substituted aryl and optionally substituted heteroaryl; R.sub.3a is selected from the group consisting of hydrogen, optionally substituted C.sub.1-C.sub.10 alkyl, optionally substituted C.sub.2-C.sub.10 alkenyl, optionally substituted C.sub.2-C.sub.10 alkynyl, optionally substituted C.sub.3-C.sub.12 cycloalkyl, optionally substituted C.sub.3-C.sub.12 cycloalkenyl, optionally substituted aryl, halo, OR.sub.c, NR.sub.dR.sub.d, C(O)OR.sub.c, NO.sub.2, CN, C(O)R.sub.c, C(O)C(O)R.sub.c, C(O)NR.sub.dR.sub.d, NR.sub.dC(O)R.sub.c, NR.sub.dS(O).sub.nR.sub.c, N(R.sub.d)(COOR.sub.c), NR.sub.dC(O)C(O)R.sub.c, NR.sub.dC(O)NR.sub.dR.sub.d, NR.sub.dS(O).sub.nNR.sub.dR.sub.d, NR.sub.dS(O).sub.nR.sub.c, S(O).sub.nR.sub.c, S(O).sub.nNR.sub.dR.sub.d, OC(O)OR.sub.c, (C═NR.sub.d)R.sub.c, optionally substituted heterocyclic and optionally substituted heteroaryl; R.sub.4a is selected from the group consisting of hydrogen, optionally substituted C.sub.1-C.sub.10 alkyl, optionally substituted C.sub.2-C.sub.10 alkenyl, optionally substituted C.sub.2-C.sub.10 alkynyl, optionally substituted C.sub.3-C.sub.12 cycloalkyl, optionally substituted C.sub.3-C.sub.12 cycloalkenyl, optionally substituted aryl, halo, OR.sub.c, S(O).sub.nR.sub.c, NR.sub.dR.sub.d, C(O)OR.sub.c, NO.sub.2, CN, C(O)R.sub.c, C(O)C(O)R.sub.c, C(O)NR.sub.dR.sub.d, NR.sub.dC(O)R.sub.c, NR.sub.dS(O)R.sub.c, N(R.sub.d)(COOR.sub.c), NR.sub.dC(O)C(O)R.sub.c, NR.sub.dC(O)NR.sub.dR.sub.d, NR.sub.dS(O).sub.nR.sub.dR.sub.d, NR.sub.dS(O).sub.nR.sub.c, S(O)NR.sub.dR.sub.d, OC(O)OR.sub.c, (C═NR.sub.d)R.sub.c, optionally substituted heterocyclic and optionally substituted heteroaryl; R.sub.4b is selected from the group consisting of hydrogen, optionally substituted C.sub.1-C.sub.10 alkyl, optionally substituted C.sub.2-C.sub.10 alkenyl, optionally substituted C.sub.2-C.sub.10 alkynyl, optionally substituted C.sub.3-C.sub.12 cycloalkyl, optionally substituted C.sub.3-C.sub.12 cycloalkenyl, optionally substituted aryl, optionally substituted heterocyclic and optionally substituted heteroaryl; R.sub.a is selected from the group consisting of hydrogen, optionally substituted C.sub.1-C.sub.10 alkyl, optionally substituted C.sub.2-C.sub.10 alkenyl, optionally substituted C.sub.2-C.sub.10 alkynyl, optionally substituted C.sub.3-C.sub.12 cycloalkyl, optionally substituted C.sub.3-C.sub.12 cycloalkenyl, optionally substituted heterocyclic, optionally substituted aryl, optionally substituted heteroaryl, C(O)OR.sub.c, C(O)R.sub.c, C(O)C(O)R.sub.c and S(O).sub.nR.sub.c; or alternatively, R.sub.a and the nitrogen atom to which it is attached is taken together with an adjacent C(R.sub.b1)(R.sub.b1) or C(R.sub.b2)(R.sub.b2) to form an optionally substituted, 4- to 12-membered heterocyclic ring containing one or more ring nitrogen atoms, wherein said heterocyclic ring optionally contains one or more ring heteroatoms selected from oxygen and sulfur; each R.sub.b1 and R.sub.b2 is independently selected from the group consisting of hydrogen, optionally substituted C.sub.1-C.sub.10 alkyl, optionally substituted C.sub.2-C.sub.10 alkenyl, optionally substituted C.sub.2-C.sub.10 alkynyl, optionally substituted C.sub.3-C.sub.2 cycloalkyl, optionally substituted C.sub.3-C.sub.12 cycloalkenyl, optionally substituted heterocyclic, optionally substituted aryl, optionally substituted heteroaryl, halo, OR.sub.c, NR.sub.dR.sub.d, C(O)OR.sub.c, NO.sub.2, CN, C(O)R.sub.c, C(O)C(O)R.sub.c, C(O)NR.sub.dR.sub.d, NR.sub.dC(O)R.sub.c, NR.sub.dS(O).sub.nR.sub.c, N(R.sub.d)(COOR.sub.c), NR.sub.dC(O)C(O)R.sub.c, NR.sub.dC(O)NR.sub.dR.sub.d, NR.sub.dS(O).sub.nNR.sub.dR.sub.d, NR.sub.dS(O).sub.nR.sub.c, S(O).sub.nR.sub.c, S(O).sub.nNR.sub.dR.sub.d, OC(O)OR.sub.c and (C═NR.sub.d)R.sub.c; or alternatively, two geminal R.sub.b1 groups or two geminal R.sub.b2 groups and the carbon to which they are attached are taken together to form a C(O) group, or yet alternatively, two geminal R.sub.b1groups or two geminal R.sub.b2 groups are taken together with the carbon atom to which they are attached to form a spiro C.sub.3-C.sub.12 cycloalkyl, a spiro C.sub.3-C.sub.12 cycloalkenyl, a spiro heterocyclic, a spiro aryl or spiro heteroaryl, each optionally substituted; Y is selected from the group consisting of S(O).sub.n, NR.sub.d, NR.sub.dS(O).sub.n, NR.sub.dS(O).sub.nNR.sub.d, NR.sub.dC(O), NR.sub.dC(O)O, NR.sub.dC(O)C(O), NR.sub.dC(O)NR.sub.d, S(O).sub.nNR.sub.d, and O; each R.sub.c is independently selected from the group consisting of hydrogen, optionally substituted C.sub.1-C.sub.10 alkyl, optionally substituted C.sub.2-C.sub.10 alkenyl, optionally substituted C.sub.2-C.sub.10 alkynyl, optionally substituted C.sub.3-C.sub.12 cycloalkyl, optionally substituted C.sub.3-C.sub.12 cycloalkenyl, optionally substituted heterocyclic, optionally substituted aryl and optionally substituted heteroaryl; each R.sub.d is independently selected from the group consisting of hydrogen, optionally substituted C.sub.1-C.sub.10 alkyl, optionally substituted C.sub.2-C.sub.10 alkenyl, optionally substituted C.sub.2-C.sub.10 alkynyl, optionally substituted C.sub.1-C.sub.10 alkoxy, optionally substituted C.sub.3-C.sub.12 cycloalkyl, optionally substituted C.sub.3-C.sub.12 cycloalkenyl, optionally substituted heterocyclic, optionally substituted aryl and optionally substituted heteroaryl; or two geminal R.sub.d groups are taken together with the nitrogen atom to which they are attached to form an optionally substituted heterocyclic or an optionally substituted heteroaryl; k is 0 or 1; m is 0, 1, 2, 3, 4, or 5; each n is independently 0, 1 or 2.

10. The compound of claim 9, wherein m is 0, 1 or 2.

11. The compound of claim 9 or 10, wherein k is 0.

12. The compound of any one of claims 9-11, wherein m is 1.

13. The compound of any one claims 9-12, wherein R.sub.3a is hydrogen.

14. The compound of any one of claims 9-13, wherein R.sub.a is hydrogen or C.sub.1-C.sub.4 alkyl (optionally substituted by 1, 2 or 3 halogens).

15. The compound of any one claims 9-14, wherein each of R.sub.b1 and R.sub.b2 is independently selected from of hydrogen, hydroxyl, C.sub.1-4alkoxy (optionally substituted by one, two or three substituents independently selected from halogen and hydroxyl) and C.sub.1-C.sub.4 alkyl (optionally substituted by one, two or three substituents independently selected from halogen, hydroxyl, and C.sub.1-4alkoxy).

16. The compound of claim 15, wherein each of R.sub.b1 and R.sub.b2 is hydrogen.

17. The compound of any one of claims 9-16, wherein R.sub.2 is selected from the group consisting of phenyl and a 5-6 membered heteroaryl having one or two heteroatoms each selected from N, S, and O, wherein R.sub.2 is optionally substituted by one or two substituents each independently selected from the group consisting of halogen, and C.sub.1-C.sub.4 alkyl (optionally substituted by one, two or three halogens.

18. The compound of any one of claims 9-17, wherein R.sub.2 is phenyl.

19. The compound of any one of claims 9-18, wherein R.sub.2 is phenyl is substituted with one or two R.sub.5, wherein each R.sub.5 is independently selected from the group consisting of optionally substituted C.sub.1-C.sub.10 alkyl, optionally substituted C.sub.2-C.sub.10 alkenyl, optionally substituted C.sub.2-C.sub.10 alkynyl, and halo.

20. The compound of any one of claims 9-17, wherein R.sub.2 is selected from the group consisting of: optionally substituted thienyl, optionally substituted furanyl and optionally substituted pyridinyl.

21. The compound of any one of claims 9-20, wherein R.sub.4a is selected from the group consisting of optionally substituted C.sub.1-C.sub.6 alkyl, optionally substituted C.sub.3-C.sub.7 cycloalkyl, phenyl, OR.sub.c, C(O)OR.sub.c, C(O)R.sub.c, optionally substituted heterocycle and optionally substituted heteroaryl, wherein R.sub.c is selected, independently for each occurrence, from the group consisting of H and C.sub.1-6alkyl.

22. The compound of any one of claims 9-20, wherein R.sub.4a is a heterocycle, or a 5-6 membered monocyclic or a 8-10 membered bicyclic heteroaryl having one, two or three heteroatoms selected from N, S or O, wherein the heterocycle or heteroaryl are optionally substituted by one, two or three substituents independently selected for each occurrence from the group consisting of halogen, C.sub.1-6alkyl (optionally substituted by one, two or three substituents each independently selected from halogen and hydroxyl), C.sub.1-6alkoxy (optionally substituted by one, two or three halogens), hydroxyl, and NR.sub.dR.sub.d wherein R.sub.d is independently for each occurrence selected from H and C.sub.1-4alkyl, or the two R.sub.ds taken together with the N to which they are attached form a heterocyclic ring.

23. The compound of claim 22, wherein R.sub.4a is selected from the group consisting of tetrahydropyranyl, thiadiazolyl, tetrahydrofuranyl, and morpholinyl.

24. The compound of any one of claims 9-20, wherein R.sub.4a is monocyclic heteroaryl containing one, two or three ring nitrogen atoms.

25. The compound of any one of claims 9-20, wherein R.sub.4a is selected from the group consisting of furanyl, pyridinyl, pyrazinyl, pyrazolyl, imidazolyl, isoxazolyl, triazolyl, thiazolyl, oxadiazolyl, thiadiazolyl, thienyl, piperazinyl, and benzimidazolyl, each optionally substituted.

26. The compound of claim 20, wherein R.sub.4a is selected from the group consisting of: ##STR00052## wherein each X is independently O, S or NR.sub.g; each R.sub.g is independently selected from the group consisting of hydrogen, C.sub.1-C.sub.4 alkyl, C.sub.3-C.sub.6 cycloalkyl, and each R.sub.6, R.sub.7 and R.sub.8 is independently selected for each occurrence from the group consisting of hydrogen, C.sub.1-C.sub.6 alkyl, C.sub.2-C.sub.6 alkenyl, C.sub.2-C.sub.16 alkynyl, C.sub.3-C.sub.7 cycloalkyl, C.sub.3-C.sub.7 cycloalkenyl, phenyl, heterocycle, heteroaryl, halo, hydroxyl, carboxyl, OR.sub.c, NR.sub.dR.sub.d, C(O)OR.sub.c, CN, C(O)R.sub.c, wherein the C.sub.1-6alkyl, C.sub.2-C.sub.6 alkenyl, C.sub.2-C.sub.16 alkynyl, C.sub.3-C.sub.7 cycloalkyl, C.sub.3-C.sub.7 cycloalkenyl, phenyl, heterocycle, and heteroaryl of R.sub.6, R.sub.7 and R.sub.8 may each be optionally substituted by one, two or three substituents selected from halo, hydroxyl, C.sub.1-6alkyl and C.sub.1-6alkoxy; R.sub.c is C.sub.1-4alkyl; and R.sub.d is independently for each occurrence selected from the group consisting of H and C.sub.1-4alkyl, or the two R.sub.ds taken together with the N to which they are attached form a heterocyclic ring.

27. The compound of claim 26, wherein R.sub.4a is an optionally substituted C.sub.3-C.sub.7 cycloalkyl.

28. The compound of any one of claims 9-12, wherein R.sub.4a is an optionally substituted cyclopropyl or an optionally substituted cyclobutyl.

29. The compound of claim 27 or 28, wherein R.sub.4a is a C.sub.3-C.sub.7 cycloalkyl substituted with a substituent having the formula: ##STR00053## wherein: each R.sub.h is independently selected from the group consisting of hydrogen, halo, optionally substituted C.sub.1-C.sub.10 alkyl, and optionally substituted C.sub.3-C.sub.6 cycloalkyl, or two geminal R.sub.b groups are independently taken together with the carbon atom to which they are attached to form an optionally substituted heterocyclic or an optionally substituted heteroaryl; R.sub.9 is selected from the group consisting of hydrogen, optionally substituted C.sub.1-C.sub.10 alkyl, optionally substituted C.sub.2-C.sub.10 alkenyl, optionally substituted C.sub.2-C.sub.10 alkynyl, optionally substituted C.sub.3-C.sub.12 cycloalkyl, optionally substituted C.sub.3-C.sub.12 cycloalkenyl, optionally substituted aryl, halo, OR.sub.c, NR.sub.dR.sub.d, C(O)OR.sub.c, NO.sub.2, CN, C(O)R.sub.c, C(O)C(O)R.sub.c, C(O)NR.sub.dR.sub.d, NR.sub.dC(O)R.sub.c, NR.sub.dS(O).sub.nR.sub.c, NR.sub.d(COOR.sub.c), NR.sub.dC(O)C(O)R.sub.c, NR.sub.dC(O)NR.sub.dR.sub.d, NR.sub.dS(O).sub.nNR.sub.dR.sub.d, NR.sub.dS(O).sub.nR.sub.c, S(O).sub.nR.sub.c, S(O).sub.nNR.sub.dR.sub.d, OC(O)OR.sub.c, (C═NR.sub.d)R.sub.c, optionally substituted heterocyclic and optionally substituted heteroaryl; p is 0, 1, or 2; and wherein the C.sub.3-C.sub.7 cycloalkyl is optionally further substituted.

30. The compound of any one of claims 27 to 29, wherein R.sub.4a is selected from the group consisting of: ##STR00054## wherein each R.sub.10 is independently selected from the group consisting of hydrogen, optionally substituted C.sub.1-C.sub.10 alkyl, optionally substituted C.sub.2-C.sub.10 alkenyl, optionally substituted C.sub.2-C.sub.10 alkynyl, optionally substituted C.sub.3-C.sub.12 cycloalkyl, optionally substituted C.sub.3-C.sub.12 cycloalkenyl, optionally substituted aryl, halo, OR.sub.c, NR.sub.dR.sub.d, C(O)OR.sub.c, NO.sub.2, CN, C(O)R.sub.c, C(O)C(O)R.sub.c, C(O)NR.sub.dR.sub.d, NR.sub.dC(O)R.sub.c, NR.sub.dS(O).sub.nR.sub.c, NR.sub.d(COOR.sub.c), NR.sub.dC(O)C(O)R.sub.c, NR.sub.dC(O)NR.sub.dR.sub.d, NR.sub.dS(O).sub.nNR.sub.dR.sub.d, NR.sub.dS(O).sub.nR.sub.c, S(O).sub.nR.sub.c, S(O).sub.nNR.sub.dR.sub.d, OC(O)OR.sub.c, (C═NR.sub.d)R.sub.c, optionally substituted heterocyclic and optionally substituted heteroaryl; alternatively, two geminal R.sub.10 groups are taken together with the carbon atom to which they are attached to form a spiro C.sub.3-C.sub.12 cycloalkyl, a spiro C.sub.3-C.sub.12 cycloalkenyl, a spiro heterocyclic, a spiro aryl or spiro heteroaryl, each optionally substituted; or yet alternatively, two vicinal R.sub.10 groups are taken together with the carbon atoms to which they are attached to form a fused, optionally substituted cyclic group selected from the group consisting of C.sub.4-C.sub.8 cycloalkyl, C.sub.4-C.sub.8 cycloalkenyl, 4- to 8-membered heterocyclic, aryl and heteroaryl, each optionally substituted; or further alternatively, two R.sub.10 groups attached to non-adjacent carbon atoms are taken together with the carbon atoms to which they are attached to form a bridged cyclic group selected from the group consisting of C.sub.4-C.sub.8 cycloalkyl, C.sub.4-C.sub.8 cycloalkenyl, and 4- to 8-membered heterocyclic, each optionally substituted; each R.sub.h is independently selected from the group consisting of hydrogen, halo, optionally substituted C.sub.1-C.sub.10 alkyl, and optionally substituted C.sub.3-C.sub.6 cycloalkyl, or two geminal R.sub.b groups are independently taken together with the carbon atom to which they are attached to form an optionally substituted heterocyclic or an optionally substituted heteroaryl; R.sub.9 is selected from the group consisting of hydrogen, optionally substituted C.sub.1-C.sub.10 alkyl, optionally substituted C.sub.2-C.sub.10 alkenyl, optionally substituted C.sub.2-C.sub.10 alkynyl, optionally substituted C.sub.3-C.sub.12 cycloalkyl, optionally substituted C.sub.3-C.sub.12 cycloalkenyl, optionally substituted aryl, halo, OR.sub.c, NR.sub.dR.sub.d, C(O)OR.sub.c, NO.sub.2, CN, C(O)R.sub.c, C(O)C(O)R.sub.c, C(O)NR.sub.dR.sub.d, NR.sub.dC(O)R.sub.c, NR.sub.dS(O).sub.nR.sub.c, NR.sub.d(COOR.sub.c), NR.sub.dC(O)C(O)R.sub.c, NR.sub.dC(O)NR.sub.dR.sub.d, NR.sub.dS(O).sub.nNR.sub.dR.sub.d, NR.sub.dS(O).sub.nR.sub.c, S(O).sub.nR.sub.c, S(O).sub.nNR.sub.dR.sub.d, OC(O)OR.sub.c, (C═NR.sub.d)R.sub.c, optionally substituted heterocyclic and optionally substituted heteroaryl; p is 0, 1, or 2.

31. The compound of any one of claims 9-30, wherein Y is S, S(O).sub.2 or S(O).sub.2NR.sub.d.

32. The compound of any one of claims 9-20 wherein R.sub.4b is heterocycle or a 5-6 membered monocyclic or a 8-10 membered bicyclic heteroaryl having one, two or three heteroatoms selected from N, S or O, wherein the heterocycle or heteroaryl are optionally substituted by one, two or three substituents independently selected for each occurrence from the group consisting of halogen, C.sub.1-6alkyl (optionally substituted by one, two or three substituents each independently selected from halogen and hydroxyl), C.sub.1-6alkoxy (optionally substituted by one, two or three halogens), hydroxyl, and NR.sub.dR.sub.d wherein R.sub.d is independently for each occurrence selected from H and C.sub.1-4alkyl, or the two R.sub.ds taken together with the N to which they are attached form a heterocyclic ring.

33. The compound of any one of claims 9-20, wherein R.sub.4b is selected from the group consisting of furanyl, pyridinyl, pyrazinyl, pyrazolyl, imidazolyl, isoxazolyl, triazolyl, thiazolyl, oxadiazolyl, thiadiazolyl, thienyl, piperazinyl, and benzimidazolyl, each optionally substituted.

34. A compound selected from the group consisting of: ##STR00055## and pharmaceutically acceptable salts thereof.

35. A pharmaceutical composition comprising a compound of any one of claims 1-34 and a pharmaceutically acceptable carrier or excipient.

36. The pharmaceutical composition of claim 35, wherein the composition further comprises at least one additional CFTR modulator.

37. The pharmaceutical composition of claim 35, wherein the composition further comprises at least two additional CFTR modulators.

38. A method of enhancing cystic fibrosis transmembrane conductance regulator (CFTR) activity in a subject in need thereof comprising administering to said subject an effective amount of a compound of any one of claims 1-34, or a pharmaceutical composition of any one claims 35-36.

39. The method of claim 38, wherein the activity of a mutant CFTR is enhanced.

40. The method of claim 39, wherein the mutant CFTR is selected from the group consisting ΔF508, S549N, G542X, G551D, R117H, N1303K, W1282X, R553X, 621+1G>T, 1717-1G>A, 3849+10kbC>T, 2789+5G>A, 3120+1G>A, I507del, R1162X, 1898+1G>A, 3659delC, G85E, D1152H, R560T, R347P, 2184insA, A455E, R334W, Q493X, and 2184delA CFTR.

41. The method of claim 40 wherein ΔF508 CFTR activity is enhanced.

42. The method of any one of claims 38-41, wherein the subject is suffering from a disease associated with decreased CFTR activity.

43. The method of claim 42, wherein the disease is selected from the group consisting of cystic fibrosis, congenital bilateral absence of vas deferens (CBAVD), acute, recurrent, or chronic pancreatitis, disseminated bronchiectasis, asthma, allergic pulmonary aspergillosis, chronic obstructive pulmonary disease (COPD), chronic sinusitis, dry eye disease, protein C deficiency, A-β-lipoproteinemia, lysosomal storage disease, type 1 chylomicronemia, mild pulmonary disease, lipid processing deficiencies, type 1 hereditary angioedema, coagulation-fibrinolyis, hereditary hemochromatosis, CFTR-related metabolic syndrome, chronic bronchitis, constipation, pancreatic insufficiency, hereditary emphysema, Sjogren's syndrome, familial hypercholesterolemia, I-cell disease/pseudo-Hurler, mucopolysaccharidoses, Sandhof/Tay-Sachs, Crigler-Najjar type II, polyendocrinopathy/hyperinsulemia, Diabetes mellitus, Laron dwarfism, myleoperoxidase deficiency, primary hypoparathyroidism, melanoma, glycanosis CDG type 1, congenital hyperthyroidism, osteogenesis imperfecta, hereditary hypofibrinogenemia, ACT deficiency, Diabetes insipidus (DI), neurophyseal DI, nephrogenic DI, Charcot-Marie Tooth syndrome, Perlizaeus-Merzbacher disease, Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis, progressive supranuclear palsy, Pick's disease, Huntington's disease, spinocerebellar ataxia type I, spinal and bulbar muscular atrophy, dentatorubral pallidoluysian, myotonic dystrophy, hereditary Creutzfeldt-Jakob disease (due to prion protein processing defect), Fabry disease, and Straussler-Scheinker syndrome.

44. The method of claim 43 wherein the disease is cystic fibrosis.

45. The method of claim 44, wherein the subject is a human patient.

46. The method of claim 45, further comprising administering an additional CFTR modulator.

47. The method of claim 46, wherein at least two additional CFTR modulators are administered.

48. The method of claim 46 or 47, wherein at least one CFTR modulator is a CFTR corrector or potentiator.

49. The method of claim 48, wherein the CFTR corrector is selected from the group consisting of VX-809, VX-661, VX-152, VX-440, GLPG-2222, GLPG2665, and VX-983 and the CFTR potentiator is selected from the group consisting of GLPG-1837, ivacaftor and genistein.

50. The method of claim 49, wherein one of the at least two additional therapeutic agents is a CFTR corrector and the other is a CFTR potentiator.

51. A method of identifying a candidate agent that increases CFTR activity, comprising: a) contacting a cell that expresses a CFTR protein with the candidate agent and a compound of any one of claims 1 to 34; b) measuring the CFTR activity in the cell in the presence of the candidate agent and the compound of any one of claims 1 to 34; and c) comparing the CFTR activity to that in the absence of the test agent, wherein an increase in CFTR activity in the presence of the test agent indicates that the agent increases CFTR activity.

52. The method of claim 51, wherein the cell expresses a mutant CFTR protein.

53. The method of any one of claims 51 and 52, wherein CFTR activity is measured by measuring chloride channel activity of the CFTR, and/or other ion transport activity.

54. The method of claim 53, wherein the method is high-throughput.

55. The method of any one of claims 50-54, wherein the candidate agent is a CFTR corrector or a CFTR potentiator.

56. A method of treating cystic fibrosis in a patient in need thereof, comprising administering an effective amount of compound of any one of claims 1-34 or a pharmaceutical composition of any one of claims 35-37.

Description

EXEMPLIFICATION

[0160] The compounds described herein can be prepared in a number of ways based on the teachings contained herein and synthetic procedures known in the art. In the description of the synthetic methods described below, it is to be understood that all proposed reaction conditions, including choice of solvent, reaction atmosphere, reaction temperature, duration of the experiment and workup procedures, can be chosen to be the conditions standard for that reaction, unless otherwise indicated. It is understood by one skilled in the art of organic synthesis that the functionality present on various portions of the molecule should be compatible with the reagents and reactions proposed. Substituents not compatible with the reaction conditions will be apparent to one skilled in the art, and alternate methods are therefore indicated. The starting materials for the examples are either commercially available or are readily prepared by standard methods from known materials. At least some of the compounds identified as “intermediates” herein are contemplated as compounds of the invention.

Example 1: 5-phenyl-N-((tetrahydrofuran-2-yl)methyl)thiazole-2-carboxamide

[0161] ##STR00023##

[0162] HOBt (0.11 g, 0.8 mmol) and EDC.HCl (0.20 g, 1.1 mmol) were added to a solution 5-phenylthiazole-2-carboxylic acid (0.15 g, 0.73 mmol) in THF (5 mL) followed by the addition of (tetrahydro-furan-2-yl)-methylamine (0.15 g, 1.4 mmol) and stirred at room temperature for 12 h. Volatiles were removed under vacuum and crude was dissolved in DCM. The organic layer was washed with water, brine, dried over anhydrous sodium sulfate and concentrated under vacuum to afford crude product. The crude was purified by combiflash chromatography using EtOAc in hexane to obtain the product (90 mg, 43%) as off white solid. .sup.1H NMR (400 MHz, CDCl.sub.3): δ 7.97 (s, 1H), 7.60-7.59 (m, 2H), 7.53 (brs, 1H), 7.44-7.38 (m, 3H), 4.09-4.06 (m, 1H), 3.94-3.88 (m, 1H), 3.80-3.75 (m, 1H), 3.74-3.68 (m, 1H), 3.45-3.38 (m, 1H), 2.03-1.98 (m, 1H), 1.95-1.89 (m, 2H), 1.64-1.58 (m, 1H); LC-MS: [M+H].sup.+ 289.1; HPLC Purity: 99.85% at 254 nm and 99.85% at 220 nm.

Example 2: N-(2-methoxyethyl)-5-phenylthiazole-2-carboxamide

[0163] ##STR00024##

[0164] The product was prepared using the procedure described in example 1. The crude was purified by combiflash chromatography using EtOAc in hexane to obtain the product (30 mg, 16%) as white solid. .sup.1H NMR (400 MHz, CDCl.sub.3): δ 7.98 (s, 1H), 7.60-7.58 (m, 2H), 7.53 (br, 1H), 7.49-7.38 (m, 3H), 3.67-3.62 (m, 2H), 3.59-3.55 (m, 2H), 3.39 (s, 3H); LC-MS: [M+H].sup.+ 263.0; HPLC Purity: 99.62% at 254 nm and 99.68% at 220 nm.

Example 3: N-(2-morpholinoethyl)-5-phenylthiazole-2-carboxamide

[0165] ##STR00025##

[0166] The product was prepared using the procedure described in example 1. The crude was purified by preparative HPLC obtain the product (85 mg, 37%) as off white solid. .sup.1H NMR (400 MHz, CDCl.sub.3): δ 7.99 (s, 1H), 7.62 (bs, 1H), 7.60-7.58 (m, 2H), 7.45-7.35 (m, 3H), 3.75-3.72 (m, 4H), 3.59-3.54 (m, 2H), 2.61-2.58 (m, 2H), 2.58-2.48 (m, 4H); LC-MS: [M+H].sup.+ 317.8; HPLC Purity: 97.36% at 254 nm and 97.87% at 220 nm.

Example 4: N-(3-(1H-imidazol-1-yl)propyl)-5-phenylthiazole-2-carboxamide

[0167] ##STR00026##

[0168] The product was prepared using the procedure described in example 1. The crude was purified by preparative HPLC obtain the product (92 mg, 40%) as white solid. .sup.1H NMR (400 MHz, CDCl.sub.3): δ 7.98 (s, 1H), 7.60-7.58 (d, J=6.8 Hz, 2H), 7.52 (s, 1H), 7.45-7.36 (m, 3H), 7.34-7.30 (m, 1H), 7.07 (s, 1H), 6.97 (s, 1H), 4.07-4.04 (t, J=7.0 Hz, 2H), 3.51-3.46 (m, 2H), 2.17-2.10 (m, 2H); LC-MS: [M+H].sup.+ 312.9; HPLC Purity: 99.89% at 254 nm and 99.74% at 220 nm.

Example 5:N-(3-(1-methyl-1H-pyrazol-5-yl)propyl)-5-phenylthiazole-2-carboxamide

[0169] ##STR00027##

[0170] The product was prepared using the procedure described in example 1. The crude was purified by column chromatography using flash silica gel eluted with 50% EtOAc in hexane to afford (0.115 g, 36.27%) as off white solid; .sup.1H NMR (400 MHz, CDCl.sub.3) δ 7.97 (s, 1H), 7.60-7.58 (m, 2H), 7.45-7.37 (m, 4H), 7.30 (br, 1H), 6.06 (s, 1H), 3.79 (s, 3H), 3.58-3.53 (q, 2H), 2.72-2.68 (t, 2H), 2.04-1.96 (m, 2H); LCMS [M+H].sup.+ 327.1; HPLC purity: 92.12% at 220 nm and 98.02% at 254 nm.

Example 6: N-(2-(5-(hydroxymethyl)-1,3,4-thiadiazol-2-yl)ethyl)-5-phenylthiazole-2-carboxamide

[0171] ##STR00028##

[0172] Step 1: ethyl 2-(2-(3-((tert-butoxycarbonyl)amino)propanoyl)hydrazinyl)-2-oxoacetate: Et.sub.3N (4.98 g, 49.261 mmol) was added to an ice-cooled solution of tert-butyl (3-hydrazinyl-3-oxopropyl)carbamate (5.0 g, 24.63 mmol) in DCM (100 mL) followed by ethyl oxalyl chloride (4.01 g, 29.55 mmol) and the resulting reaction mixture was stirred at room temperature for 2 h. Then the reaction mixture was diluted with water (50 mL). Organic layer separated off and the aq. layer was further extracted with DCM (50 mL×2). Combined organic layer was dried over anhydrous sodium sulfate and concentrated under reduced pressure to afford the product (7 g, crude) as yellowish sticky mass which was used as such in next step without further purification. As per 1H-NMR, triethyl amine present as an impurity in the compound. LC-MS: [M+H].sup.+=303.9.

[0173] Step 2: ethyl 5-(2-((tert-butoxycarbonyl)amino)ethyl)-1,3,4-thiadiazole-2-carboxylate: Lawesson's reagent (9.34 g, 23.10 mmol) was added to a solution of ethyl 2-(2-(3-((tert-butoxycarbonyl)amino)propanoyl)hydrazinyl)-2-oxoacetate (7 g, crude) in 1,4-dioxane (150 mL) and the reaction mixture was refluxed for 2h. Then the reaction mixture was cooled, neutralized with saturated sodium carbonate solution and extracted with EtOAc (100 mL×3). Combined organic layer was dried over anhydrous sodium sulfate and concentrated under reduced pressure to afford crude product. The crude compound thus obtained was purified by silica gel of (100-200 mesh) column chromatography using 20% EtOAc in Hexane as eluent to obtain the product (2.5 g, 33.61% after two steps) as yellow viscous mass. .sup.1H NMR (400 MHz, CDCl.sub.3): δ 5.02 (br, 1H), 4.51 (q, J=7.2 Hz, 2H), 3.61-3.60 (m, 2H), 3.39-3.37 (t, J=6.3 Hz, 2H), 1.44 (t, 3H) 1.41 (s, 9H); LC-MS: [M+H].sup.+=302.0.

[0174] Step 3: ethyl 5-(2-aminoethyl)-1,3,4-thiadiazole-2-carboxylate hydrochloride: 4M HCl in 1,4-dioxane (3 mL) was added to an ice-cooled solution of ethyl 5-(2-((tert-butoxycarbonyl)amino)ethyl)-1,3,4-thiadiazole-2-carboxylate (0.200 g, 0.66 mmol) in dioxane (5 mL) and the resulting reaction mixture was stirred at room temperature for 2 h. Volatiles were removed under reduced pressure to obtain the product (0.150 g, crude) as yellowish solid which was used as such for coupling. LC-MS: [M+H].sup.+=202.2.

[0175] Step 4: ethyl 5-(2-(5-phenylthiazole-2-carboxamido)ethyl)-1,3,4-thiadiazole-2-carboxylate: HATU (0.278 g, 0.73 mmol) and DIPEA (0.188 g, 1.46 mmol) were added sequentially to a solution of 5-phenylthiazole-2-carboxylic acid (0.100 g, 0.48 mmol) in THF (10 mL) and the reaction mixture was stirred for 10 min. Ethyl 5-(2-aminoethyl)-1,3,4-thiadiazole-2-carboxylate hydrochloride (0.147 g, crude) was then added and the reaction mixture was stirred at room temperature for 12 h. Volatiles were removed under vacuum and the crude compound was dissolved in DCM. The organic layer was washed with water, brine, dried over anhydrous sodium sulfate and concentrated under reduced pressure to afford crude product. The crude compound was purified by silica gel (100-200 mesh) column chromatography using 20% EtOAc in Hexane as eluent to obtain the product (0.150 g, 63%) as off white solid. .sup.1H NMR (400 MHz, DMSO): δ 9.13 (t, J=5.9 Hz, 1H), 8.42 (s, 1H), 7.77 (d, J=8.5 Hz 2H), 7.50-7.49 (m, 2H), 7.48-7.44 (m, 1H), 4.40 (q, J=7.1 Hz, 2H), 3.71 (q, J=6.3 Hz, 2H), 3.49 (t, J=6.5 Hz, 2H), 1.32 (t, J=7.12 Hz, 3H); LC-MS: [M+H].sup.+=388.8.

[0176] Step 5: N-(2-(5-(hydroxymethyl)-1,3,4-thiadiazol-2-yl)ethyl)-5-phenylthiazole-2-carboxamide: Sodium borohydride (0.063 g, 1.67 mmol) was added in portions to an ice-cooled solution of ethyl 5-(2-(5-phenylthiazole-2-carboxamido)ethyl)-1,3,4-thiadiazole-2-carboxylate (0.130 g, 0.33 mmol) in ethanol (20 mL) and the reaction mixture was stirred for 2 h at room temperature. Volatiles were removed under reduced pressure and the reaction mixture was diluted with cold water. The aq. layer was extracted with EtOAc (30 mL×3). Combined organic layer was dried over anhydrous sodium sulfate and concentrated under reduced pressure to afford crude product. The crude compound thus obtained was purified by silica gel (100-200 mesh) column chromatography using 70% EtOAc in Hexane as eluent to obtain the product (0.050 g, 43.47%) as white solid. .sup.1H NMR (400 MHz, DMSO-d.sub.6): δ 9.09 (t, J=5.8 Hz, 1H), 8.42 (s, 1H), 7.77 (dd, 2H), 7.50-7.46 (m, 2H), 7.43-7.41 (m, 1H), 6.13 (t, J=5.9 Hz, 1H), 4.79 (d, J=5.9 Hz, 2H), 3.67 (q, J=6.5 Hz, 2H), 3.39-3.35 (m, 2H); LC-MS: [M+H].sup.+=347.0; HPLC purity: 99.27% at 220 nm and 99.85% at 254 nm.

Example 7: N-(2-(5-(hydroxymethyl)-1,3,4-oxadiazol-2-yl)ethyl)-5-phenylthiazole-2-carboxamide

[0177] ##STR00029##

[0178] Step 1: 3-(1,3-dioxoisoindolin-2-yl)-N′-(2-methoxyacetyl)propanehydrazide: EDC.HCl (2.8 g, 15.0 mmol) was added to a solution of 3-(1,3-dioxoisoindolin-2-yl)propanoic acid (2.2 g, 10.0 mmol) in THF (30 mL followed by HOBt (1.84 g, 12.0 mmol) and the reaction mixture was stirred for 20 min at room temperature. 2-methoxyacetohydrazide (1.05 g, 10.0 mmol) was added to the reaction mixture followed by DIPEA (6.4 g, 50.0 mmol) and the reaction mixture was stirred at room temperature for 16 h. Volatiles were removed under reduced pressure and the reaction mixture was diluted with water (30 mL). The aq. layer was extracted with DCM (3×25 mL), combined organic layer was washed with aqueous NaHCO.sub.3 solution (50 mL), dried over anhydrous Na.sub.2SO.sub.4 and concentrated under reduced pressure to obtain the product (2.0 g, 65%) as white solid. .sup.1H NMR (400 MHz, DMSO-d.sub.6): δ 9.91 (s, 1H), 9.72 (s, 1H), 7.88-7.82 (m, 4H), 3.88 (s, 2H), 3.78 (t, J=7.6 Hz, 2H), 3.30 (s, 3H), 2.25 (t, J=7.6 Hz, 2H); LC-MS: [M+H].sup.+=306.3 m/z.

[0179] Step 2: 2-(2-(5-(methoxymethyl)-1,3,4-oxadiazol-2-yl)ethyl)isoindoline-1,3-dione: T.sub.3P (10 mL, 15.7 mmol) was added to a solution of 3-(1,3-dioxoisoindolin-2-yl)-N′-(2-methoxyacetyl)propanehydrazide (2.0 g, 6.55 mmol) in 1, 4-dioxane (60 mL) and the reaction mixture was heated to 100° C. for 18 h. Volatiles were removed under reduced pressure and the crude reaction mixture was diluted with aqueous NaHCO.sub.3 solution (50 mL). The aq. layer was extracted with EtOAc (2×50 mL). Combined organic layer was washed with water (50 mL), brine (50 mL), dried over sodium sulfate, filtered and concentrated under reduced pressure to afford the product (1.4 g, 74%) as off-white solid which was used as such without further purification. .sup.1H NMR (400 MHz, DMSO-d.sub.6): δ 7.89-7.83 (m, 4H), 4.57 (s, 2H), 3.94 (t, J=7.6 Hz, 2H), 3.28 (s, 3H), 3.22 (t, J=7.6 Hz, 2H); LC-MS: [M+H].sup.+=288.2 m/z.

[0180] Step 3: 2-(2-(5-(methoxymethyl)-1,3,4-oxadiazol-2-yl)ethyl)isoindoline-1,3-dione: hydrazine hydrate (0.4 mL) was added to a solution of 2-(2-(5-(methoxymethyl)-1,3,4-oxadiazol-2-yl)ethyl)isoindoline-1,3-dione (0.75 g, 2.6 mmol) in ethanol (10 mL) and the reaction mixture was stirred at room temperature for 16 h. Then the reaction mixture was filtered and concentrated under reduced pressure to obtain the product (0.32 g, crude) which was used as such in next step without further purification. LC-MS: [M+H].sup.+=157.9 m/z.

[0181] Step 4: N-(2-(5-(methoxymethyl)-1,3,4-oxadiazol-2-yl)ethyl)-5-phenylthiazole-2-carboxamide: A solution of 2-(2-(5-(methoxymethyl)-1,3,4-oxadiazol-2-yl)ethyl)isoindoline-1,3-dione (0.314 g, crude) and ethyl 5-phenylthiazole-2-carboxylate (0.29 g, 1.2 mmol) in DMF (5 mL) was heated to 65° C. for 16 h. After completion, the reaction mixture was diluted with water (25 mL) and extracted with DCM (2×25 mL). Combined organic layer was washed with water (30 mL) followed by brine (30 mL) and dried over sodium sulfate. Solvent was removed under reduced pressure to afford crude product. The crude compound was purified by combiflash to obtain the product (0.14 g) as off-white solid. LC-MS: [M+H].sup.+=345.1 m/z. As per 1H-NMR, compound is impure and used as such in next step without further purification.

[0182] Step 5: N-(2-(5-(hydroxymethyl)-1,3,4-oxadiazol-2-yl)ethyl)-5-phenylthiazole-2-carboxamide: BBr.sub.3 (0.15 ml, 1.56 mmol) was added to a −78° C. solution of N-(2-(5-(methoxymethyl)-1,3,4-oxadiazol-2-yl)ethyl)-5-phenylthiazole-2-carboxamide (0.140 g, crude) in DCM (6 mL) and the reaction mixture was stirred at room temperature for 5 h. Then the reaction mixture was poured onto ice, stirred for 10 min. The aq. layer was basified with saturated sodium bicarbonate solution and extracted with DCM (2×20 mL). Combined organic layer was washed with water, dried over sodium sulfate and concentrated under reduced pressure to afford crude product. The crude compound thus obtained was purified by crystallization to obtain the product (35 mg, 4.3% over three steps) as white crystal. .sup.1H NMR (400 MHz, DMSO-d.sub.6): δ 9.07 (t, J=6.0 Hz, 1H), 8.42 (s, 1H), 7.77 (d, J=7.2 Hz, 2H), 7.48 (m, 2H), 7.42 (m, 2H), 5.86 (t, J=6.0 Hz, 1H), 4.59 (d, J=6.0 Hz, 2H), 3.66 (q, J=6.7 Hz, 2H), 3.14 (t, J=7.2 Hz, 2H); LC-MS: [M+H].sup.+=331.1 m/z; HPLC purity 98.02% at 314 nm, 97.60% at 254 nm.

Examples 8 and 9: N-(2-(4-(hydroxymethyl)-5-methyl-1H-1,2,3-triazol-1-yl)ethyl)-5-phenylthiazole-2-carboxamide and N-(2-(5-(hydroxymethyl)-4-methyl-1H-1,2,3-triazol-1-yl)ethyl)-5-phenylthiazole-2-carboxamide

[0183] ##STR00030##

[0184] Step 1: N-(2-(4-(hydroxymethyl)-5-methyl-H-1,2,3-triazol-1-yl)ethyl)-5-phenylthiazole-2-carboxamide and N-(2-(5-(hydroxymethyl)-4-methyl-1H-1,2,3-trazol-1-yl)ethyl)-5-phenylthiazole-2-carboxamide: a solution of 2-(2-azidoethyl)isoindoline-1,3-dione (5.0 g, 23.0 mmol) and 2-butyn-1-ol (2.0 mL, 26.7 mmol) in toluene (50 mL) was heated to 110° C. in a sealed tube and for 48 h. After completion, volatiles were removed under reduced pressure and crude reaction mixture thus obtained was washed with ether (100 mL). Residue was dried to obtain (4.5 g, 68%) as a mixture of two regioisomers which was used as such in the next step without further purification. .sup.1H NMR (400 MHz, DMSO-d.sub.6): δ 7.86-7.82 (m, 8H), 5.36 (t, J=5.2 Hz, 1H), 5.00 (t, J=5.2 Hz, 1H), 4.59 (t, J=5.6 Hz, 2H), 4.56 (t, J=5.2 Hz, 2H), 4.53 (d, J=5.6 Hz, 2H), 4.40 (d, J=5.6 Hz, 2H), 4.02 (t, J=6.0 Hz, 2H), 3.95 (t, J=6.0 Hz, 2H), 2.30 (s, 3H), 2.15 (s, 3H); LC-MS: [M+H].sup.+=287.1 m/z.

[0185] Step 2: (1-(2-aminoethyl)-4-methyl-1H-1,2,3-triazol-5-yl)methanol and (1-(2-aminoethyl)-5-methyl-1H-1,2,3-triazol-4-yl)methanol: hydrazine hydrate (0.200 mL, 4.0 mmol) was added to a solution of N-(2-(4-(hydroxymethyl)-5-methyl-1H-1,2,3-triazol-1-yl)ethyl)-5-phenylthiazole-2-carboxamide and N-(2-(5-(hydroxymethyl)-4-methyl-1H-1,2,3-triazol-1-yl)ethyl)-5-phenylthiazole-2-carboxamide (0.575 g, 2.0 mmol) in ethanol (10 mL) and the mixture was heated to 60° C. for 2h. After completion, the mixture was filtered and filtrate was concentrated under reduced pressure to obtain the product as a mixture of isomers (0.305 g, crude) which was used as such in next step without purification. LC-MS: (M+H).sup.+=157.2.

[0186] Step 3: N-(2-(4-(hydroxymethyl)-5-methyl-1H-1,2,3-triazol-1-yl)ethyl)-5-phenylthiazole-2-carboxamide and N-(2-(5-(hydroxymethyl)-4-methyl-1H-1,2,3-triazol-1-yl)ethyl)-5-phenylthiazole-2-carboxamide: HATU (0.685 g, 1.8 mmol) and DIPEA (0.58 g, 4.5 mmol) were added sequentially to a solution of 5-phenylthiazole-2-carboxylic acid (0.305 g, 1.5 mmol) in DMF (5 mL). After 10 min stirring, to the resulting reaction mixture was added (1-(2-aminoethyl)-4-methyl-1H-1,2,3-triazol-5-yl)methanol and (1-(2-aminoethyl)-5-methyl-1H-1,2,3-triazol-4-yl)methanol (0.235 g, crude) and the reaction mixture was stirred at room temperature for 12 h. Reaction mixture was diluted with water (15 mL), extracted with dichloromethane (3×25 mL) and washed with brine (2×50 mL). Combined organic layer was dried over anhydrous Na.sub.2SO.sub.4 and concentrated under reduced pressure to obtain 0.9 g of crude material which was purified by prep HPLC to obtain target compounds. Yield: 25% over two steps.

N-(2-(5-(hydroxymethyl)-4-methyl-1H-1,2,3-triazol-1-yl)ethyl)-5-phenylthiazole-2-carboxamide

[0187] 0.09 g; white solid. .sup.1H NMR (400 MHz, DMSO-d.sub.6): δ 8.98 (t, J=5.6 Hz, 1H), 8.41 (s, 1H), 7.77 (d, J=7.2 Hz, 2H), 7.50-7.40 (m, 3H), 5.32 (t, J=5.2 Hz, 1H), 4.54-4.51 (m, 4H), 3.73-3.69 (m, 2H), 2.20 (s, 3H); LC-MS: [M+H].sup.+=344.0 m/z; HPLC purity 99.68% at 220 nm, 99.63% at 254 nm.

N-(2-(4-(hydroxymethyl)-5-methyl-1H-1,2,3-triazol-1-yl)ethyl)-5-phenylthiazole-2-carboxamide

[0188] 0.085 g; white solid. .sup.1H NMR (400 MHz, DMSO-d.sub.6): δ 9.03 (t, J=5.6 Hz, 1H), 8.42 (s, 1H), 7.78 (d, J=7.2 Hz, 2H) 7.50-7.40 (m, 3H), 5.00 (t, J=5.6 Hz, 1H), 4.47-4.43 (m, 4H), 3.68-3.64 (m, 2H), 2.28 (s, 3H); LC-MS: [M+H].sup.+=344.0 m/z; HPLC purity 99.75% at 220 nm, 99.62% at 254 nm.

Example 10: N-cis-3-(5-(hydroxymethyl)-1,3,4-oxadiazol-2-yl)cyclobutyl)-5-phenylthiazole-2-carboxamide

[0189] ##STR00031##

[0190] Step 1: ethyl 3-oxocyclobutane-1-carboxylate: triethyl orthoacetate (21.31 g, 0.131 mol) was added to a solution of 3-oxocyclobutane-1-carboxylic acid (5.0 g, 0.043 mol) in toluene (100 mL) and the reaction mixture was refluxed for 6 h. The reaction mixture was quenched with a IN HCl solution and the layers were separated off. The organic layer was washed with saturated NaHCO.sub.3 solution (2×50 mL), brine (2×50 mL), dried over anhydrous sodium sulfate and concentrated under reduced pressure to get the product (5.3 g, 85%) as a yellow liquid. .sup.1H NMR (400 MHz, CDCl.sub.3): δ 4.23-4.17 (q, J=7.0 Hz, 2H), 3.44-3.37 (m, 2H), 3.32-3.16 (m, 3H), 1.30-1.26 (t, J=7.0 Hz, 3H).

[0191] Step 2: ethyl cis-3-hydroxycyclobutane-1-carboxylate: sodium borohydride (1.55 g, 0.041 mol) was added to an iced cold solution of ethyl 3-oxocyclobutane-1-carboxylate (5.3 g, 0.037 mol) in methanol (75 mL) and the reaction mixture was stirred for 1 h. The reaction mixture was quenched with acetone (10 mL) and volatiles were removed under reduced pressure. The crude reaction mixture was suspended in NaHCO.sub.3 solution (30 mL) and extracted with DCM (100 mL). The organic layer was washed with brine (30 mL), dried over anhydrous sodium sulfate and concentrated under reduced pressure to get the product (3.2 g, 59%) as a yellow oil. .sup.1H NMR (400 MHz, CDCl.sub.3): δ 4.20-4.09 (overlapped q and m, 3H), 3.68 (d, J=2.3 Hz, 1H), 2.62-2.54 (m, 3H), 2.20-2.10 (m, 2H), 1.26-1.22 (t, J=7.0 Hz, 3H); LC-MS: [M+H].sup.+ 145.1.

[0192] Step 3: ethyl cis-3-((methylsulfonyl)oxy)cyclobutane-1-carboxylate: Et.sub.3N (8.96 mL, 0.0666 mol) was added to a solution of ethyl cis-3-hydroxycyclobutane-1-carboxylate (3.2 g, 0.0222 mol) in DCM (100 mL) followed by MsCl (3.03 g, 0.0266 mol) drop wise and the resulting reaction mixture was stirred at room temperature for 1 h. The reaction mixture was poured onto ice cold water (50 mL) and extracted with DCM. The organic layer was dried over anhydrous sodium sulfate and concentrated under reduced pressure to get the crude product (5.1 g) as a yellow oil. .sup.1H NMR (400 MHz, CDCl.sub.3): δ 4.95-4.88 (m, 1H), 4.17-4.12 (q, J=7.1 Hz, 2H), 3.71 (d, 1H), 2.98 (s, 3H), 2.74-2.66 (m, 3H), 2.60-2.59 (m, 2H), 1.27-1.25 (t, J=7.1 Hz, 3H); LC-MS: [M+H].sup.+ 223.0.

[0193] Step 4: ethyl trans-3-azidocyclobutane-1-carboxylate: a mixture of sodium azide (2.98 g, 0.044 mol) and ethyl cis-3-((methylsulfonyl)oxy)cyclobutane-1-carboxylate (5.1 g, 0.022 mol) in DMF (25 mL) was heated to 90° C. for 16 h. The reaction mixture was poured onto water (70 mL) and extracted with ethyl acetate (2×100 mL). Combined organic layer was dried over anhydrous sodium sulfate and concentrated under reduced pressure to obtain crude product which was chromatographed on 230-400 mesh silica gel using 10% EtOAc in hexane as eluent to afford the product (3.8 g, 100% over two steps) as colorless liquid. .sup.1H NMR (400 MHz, CDCl.sub.3): δ 4.18-4.10 (m, 3H), 3.11-3.04 (m, 1H), 2.60-2.53 (m, 2H), 2.36-2.29 (m, 2H), 1.27-1.22 (t, J=7.1 Hz, 3H); LC-MS: [M+H].sup.+ 171.1.

[0194] Step 5a/b: ethyl trans-3-aminocyclobutane-1-carboxylate hydrochloride: a mixture of ethyl trans-3-azidocyclobutane-1-carboxylate (3.8 g, 0.0221 mol) and 10% Pd/C (1.0 g) in ethanol (50 mL) was hydrogenated (50 psi) for 4 h at room temperature. The reaction mixture was filtered through a celite bed and the filtrate was concentrated under reduced pressure obtain the crude compound. The crude compound was treated with 4 M HCl in dioxane to afford HCl salt (3.8 g, 95%) as colorless viscous oil.

[0195] Step 6: ethyl trans-3-(5-phenylthiazole-2-carboxamido)cyclobutane-1-carboxylate: Et.sub.3N (0.15 mL, 0.0011 mol) and HATU (0.255 g, 0.00066 mol) were added sequentially to a suspension of ethyl trans-3-aminocyclobutane-1-carboxylate hydrochloride (0.100 g, 0.00055 mol) and 5-phenylthiazole-2-carboxylic acid (0.125 g, 0.000612 mol) in THF (10 mL) and the reaction mixture was stirred for 12 h at room temperature. Volatiles were removed under reduced pressure and the crude reaction mixture was diluted with water (10 mL). The aq. phase was extracted with ethyl acetate (2×25 mL). Combined organic layer was dried over anhydrous sodium sulfate and concentrated under reduced pressure to obtain the crude compound which was further purified by (230-400 mesh) silica gel column chromatography using 25% EtOAc in hexane to afford the product (0.130 g, 70%) as off white solid. .sup.1H NMR (400 MHz, DMSO-d.sub.6): δ 7.97 (s, 1H), 7.60-7.57 (m, 2H), 7.45-7.36 (m, 4H), 4.76-4.74 (m, 1H), 4.18 (q, J=7.0 Hz, 2H), 3.11-3.10 (m, 1H), 2.78-2.72 (m, 2H), 2.43-2.35 (m, 2H), 1.28 (t, J=7.0 Hz, 3H); LC-MS: [M+H].sup.+ 330.9 m/z.

[0196] Step 7: trans-3-(5-phenylthiazole-2-carboxamido)cyclobutane-1-carboxylic acid: lithium hydroxide monohydrate (0.153 g, 0.0036 mol) was added to a solution of ethyl trans-3-(5-phenylthiazole-2-carboxamido)cyclobutane-1-carboxylate (1.1 g, 0.0033 mol) in THF:water (45 mL, 2:1) and the resultant reaction mixture was stirred at room temperature for 2 h. The reaction mixture was evaporated under vacuum and partitioned between ethyl acetate (30 mL) and water (10 mL). Separated aqueous layer was acidified with saturated citric acid solution (15 mL), precipitate thus formed was filtered, washed with water and dried to afford the product (0.315 g, 98%) as white solid. .sup.1H NMR (400 MHz, DMSO-d.sub.6): δ 12.21 (s, 1H), 9.25 (d, J=8.1 Hz, 1H), 8.42 (s, 1H), 7.77 (d, J=7.3 Hz, 2H), 7.50-7.40 (m, 3H), 4.60-4.54 (m, 1H), 2.96-2.91 (m, 1H), 2.50-2.39 (m, 4H); LC-MS: [M+H].sup.+ 303.1 m/z.

[0197] Step 8: tert-butyl 2-trans-3-(5-phenylthiazole-2-carboxamido)cyclobutane-1-carbonyl)hydrazine-1-carboxylate: HATU (1.49 g, 0.0039 mol) was added to a solution of trans-3-(5-phenylthiazole-2-carboxamido)cyclobutane-1-carboxylic acid (0.990 g, 0.0032 mol) in THF (90 mL) and stirred at room temperature for 15 min. To the resulting reaction mixture was added N-Boc-hydrazine (0.518 g, 0.0039 mol) followed by Et.sub.3N (0.88 mL, 0.00654 mol) and stirring continued for 2 h at room temperature. The reaction mixture was poured onto water (80 mL) and extracted with ethyl acetate (100 mL). The organic layer was dried over anhydrous sodium sulfate and concentrated under reduced pressure to obtain the crude compound. The crude compound was further purified by silica gel (230-400 mesh) column chromatography using EtOAc as eluent to afford the product (1.8 g, crude) as white solid with polar impurity. LC-MS: [M+H].sup.+ 417.2 m/z.

[0198] Step 9: N-trans-3-(hydrazinecarbonyl)cyclobutyl)-5-phenylthiazole-2-carboxamide hydrochloride: 4 M HCl in dioxane solution (12 mL) was added to a solution of tert-butyl 2-trans-3-(5-phenylthiazole-2-carboxamido)cyclobutane-1-carbonyl)hydrazine-1-carboxylate (0.800 g, 0.00192 mol) in 1,4-dioxane (5 mL) and the reaction mixture was stirred at room temperature for 6 h. Volatiles were removed under reduced pressure and the reaction mixture was diluted with diethyl ether (100 mL) to afford the product (0.650 g, crude) as off white solid. LC-MS: [M+H].sup.+ 317.2 m/z.

[0199] Step 10: N-trans-3-(2-(2-((tert-butyldimethylsilyl)oxy)acetyl)hydrazine-1-carbonyl)cyclobutyl)-5-phenylthiazole-2-carboxamide: HATU (0.256 g, 0.00067 mol) was added to a solution of N-trans-3-(hydrazinecarbonyl)cyclobutyl)-5-phenylthiazole-2-carboxamide hydrochloride (0.161 g, 0.00085 mol) in THF (20 mL) and stirred at room temperature for 15 minutes. Then added 2-((tert-butyldimethylsilyl)oxy)acetic acid (0.200 g, 0.00056 mol) and Et.sub.3N (0.301 mL, 0.00224 mol) were added and the mixture was stirred at room temperature for 6 h. The reaction mixture was diluted with water (30 mL) and extracted with ethyl acetate (30 mL). The organic layer was dried over anhydrous sodium sulfate and concentrated under reduced pressure obtained crude product. The crude compound was purified by column chromatography on 230-400 mesh silica gel using 90% EtOAc in hexane afforded the product (0.103 g, crude) as off white solid. LC-MS: [M+H].sup.+ 489.1 m/z.

[0200] Step 11: N-trans-3-(5-(((tert-butyldimethylsilyl)oxy)methyl)-1,3,4-oxadiazol-2-yl)cyclobutyl)-5-phenylthiazole-2-carboxamide: Iodine (0.206 g, 0.0008 mol) was added to a solution of PPh.sub.3 (0.212 g, 0.00081 mol) in DCM (30 mL) at room temperature and the reaction mixture was stirred for 15 min. The resulting reaction mixture was cooled in an ice bath and to the reaction mixture was triethyl amine (0.27 mL, 0.00204 mol) followed by N-trans-3-(2-(2-((tert-butyldimethylsilyl)oxy)acetyl)hydrazine-1-carbonyl)cyclobutyl)-5-phenylthiazole-2-carboxamide (0.200 g, 0.00040 mol). The reaction mixture was stirred at room temperature for 3 h. Volatiles were removed under reduced pressure; the crude compound was diluted with ethyl acetate and filtered. The filtrate was concentrated under reduced pressure to afford crude product. The crude compound was purified by (230-400 mesh) silica gel column chromatography using 60% EtOAc in hexane as eluent to obtain the product (0.110 g, 21% over four steps) as off white solid. .sup.1H NMR (400 MHz, DMSO-d.sub.6): δ 9.41 (d, J=8.0 Hz, 1H), 8.44 (s, 1H), 7.79-7.77 (m, 2H), 7.50-7.47 (m, 2H), 7.43-7.41 (m, 1H), 4.87 (s, 2H), 4.73-4.67 (m, 1H), 3.73-3.69 (m, 1H), 2.80-2.72 (m, 2H), 2.60-2.54 (m, 2H), 0.88 (s, 9H), 0.10 (s, 6H); LC-MS: [M+H].sup.+ 471.3 m/z.

[0201] Step 12: N-trans-3-(5-(hydroxymethyl)-1,3,4-oxadiazol-2-yl)cyclobutyl)-5-phenylthiazole-2-carboxamide: tetra butyl ammonium fluoride (0.46 mL, 0.00046 mol, 1 M in THF solution) was added to a cold solution of N-trans-3-(5-(((tert-butyldimethylsilyl)oxy)methyl)-1,3,4-oxadiazol-2-yl)cyclobutyl)-5-phenylthiazole-2-carboxamide (0.110 g, 0.00023 mol) in THF (5 mL) and the reaction mixture was stirred for 1 h. Volatiles were removed under reduced pressure; the crude reaction mixture was suspended in water (10 mL) and extracted with ethyl acetate (15 mL). The organic layer was dried over anhydrous Na.sub.2SO.sub.4 and concentrated under reduced pressure to afford the product (0.070 g, 84%) as off white solid. .sup.1H NMR (400 MHz, DMSO-d.sub.6): δ 9.41 (d, J=8 Hz, 1H), 8.45 (s, 1H), 7.80-7.77 (m, 2H), 7.50-7.42 (m, 3H), 5.89-5.86 (m, 1H), 4.72-4.67 (m, 1H), 4.63 (d, J=4 Hz, 2H), 3.72-3.67 (m, 1H), 2.79-2.71 (m, 2H), 2.61-2.56 (m, 2H); LC-MS: [M+H].sup.+ 356.8 m/z; HPLC purity: 99.39% at 220 nm and 98.86% at 254 nm.

Example 11: N-trans-3-(5-((R)-1-hydroxyethyl)-1,3,4-oxadiazol-2-yl)cyclobutyl)-5-phenylthiazole-2-carboxamide

[0202] ##STR00032##

[0203] Step 1a: methyl (2R)-2-[(tert-butyldimethylsilyl)oxy]propanoate: into a 250-mL round-bottom flask, was placed a solution of methyl (2R)-2-hydroxypropanoate (5 g, 48.03 mmol, 1.00 equiv) and Imidazole (6.5 g, 95.59 mmol, 2.00 equiv) in dichloromethane (100 mL). This was followed by the addition of a solution of tert-butyl(chloro)dimethylsilane (8.7 g, 57.72 mmol, 1.20 equiv) in dichloromethane (50 mL) dropwise with stirring at 0° C. The resulting solution was stirred for 2 hours at room temperature. The reaction was then quenched by the addition of 100 mL of water/ice. The resulting solution was extracted with dichloromethane (3×100 mL) and the organic layers combined. The resulting mixture was washed with brine (3×50 mL), dried over anhydrous sodium sulfate and concentrated under vacuum. This resulted in 7 g (67%) of methyl (2R)-2-[(tert-butyldimethylsilyl)oxy]propanoate as colorless oil.

[0204] Step 1b: (2R)-2-[(tert-butyldimethylsilyl)oxy]propanehydrazide: into a 250-mL round-bottom flask, was placed a solution of methyl (2R)-2-[(tert-butyldimethylsilyl)oxy]propanoate (7 g, 32.06 mmol, 1.00 equiv) in ethanol (100 mL). To the solution was added hydrazine (10 g, 159.81 mmol, 5.00 equiv, 80%). The resulting solution was stirred for 15 hours at 90° C. in an oil bath. The resulting solution was quenched by the addition of water/ice. The resulting solution was extracted with ethyl acetate (3×100 mL) and the organic layers combined. The resulting mixture was washed with brine (2×100 mL), dried over anhydrous sodium sulfate and concentrated under vacuum. This resulted in 6.5 g (93%) of (2R)-2-[(tert-butyldimethylsilyl)oxy]propanehydrazide as colorless oil. LC-MS (ES, m/z): [M+1].sup.+=219.

[0205] Step 1: methyl (trans-3-(1,3-dioxo-2,3-dihydro-1H-isoindol-2-yl)cyclobutane-1-carboxylate: into a 250-mL round-bottom flask, under nitrogen, was placed a solution of methyl 3-cis-hydroxycyclobutane-1-carboxylate (8 g, 61.47 mmol, 1.00 equiv), 2,3-dihydro-1H-isoindole-1,3-dione (18.1 g, 123.02 mmol, 2.00 equiv) and triphenylphosphine (32.3 g, 123.15 mmol, 2.00 equiv) in THF (100 mL). This was followed by the addition of DIAD (24.9 g, 123.14 mmol, 2.00 equiv) dropwise with stirring at 0° C. The resulting solution was stirred for 2.5 hours at room temperature. The resulting mixture was concentrated under vacuum. The residue was applied onto a silica gel column with ethyl acetate/petroleum ether (1:5). The crude product was re-crystallized from petroleum ether/ethyl acetate in the ratio of 10:1. This resulted in 7.2 g (45%) of methyl trans-3-(1,3-dioxo-2,3-dihydro-1H-isoindol-2-yl)cyclobutane-1-carboxylate as a white solid. LC-MS (ES, m/z): [M+1].sup.+=260. .sup.1H NMR (400 MHz, CDCl.sub.3): δ 7.85-7.82 (m, 2H), 7.74-7.71 (m, 2H), 5.08-5.04 (m, 1H), 3.75 (s, 3H), 3.34-3.32 (m, 1H), 3.20-3.12 (m, 2H), 2.66-2.60 (m, 2H).

[0206] Step 2: trans-3-(1,3-dioxo-2,3-dihydro-1H-isoindol-2-yl)cyclobutane-1-carboxylic acid: into a 100-mL round-bottom flask, was placed a solution of methyl trans-3-(1,3-dioxo-2,3-dihydro-1H-isoindol-2-yl)cyclobutane-1-carboxylate (7.2 g, 27.77 mmol, 1.00 equiv) in 1,4-dioxane (100 mL). To the solution was added 5M hydrogen chloride aqueous (10 mL). The resulting solution was stirred for 4 hours at 80° C. in an oil bath. The resulting mixture was concentrated under vacuum. This resulted in 6.2 g (91%) of trans-3-(1,3-dioxo-2,3-dihydro-1H-isoindol-2-yl)cyclobutane-1-carboxylic acid as a white solid. LC-MS (ES, m/z): [M−1].sup.−=244.

[0207] Step 3: (2R)-2-[(tert-butyldimethylsilyl)oxy]-N-[trans-3-(1,3-dioxo-2,3-dihydro-1H-isoindol-2-yl)cyclobutyl]carbonyl]propanehydrazide: into a 250-mL round-bottom flask, was placed a solution of trans-3-(1,3-dioxo-2,3-dihydro-1H-isoindol-2-yl)cyclobutane-1-carboxylic acid (6.2 g, 25.28 mmol, 1.00 equiv), (2R)-2-[(tert-butyldimethylsilyl)oxy]propanehydrazide (6.61 g, 30.27 mmol, 1.20 equiv) and HATU (14.4 g, 37.89 mmol, 1.50 equiv) in THF (100 mL). This was followed by the addition of DIEA (9.81 g, 75.91 mmol, 3.00 equiv) dropwise with stirring at 0° C. The resulting solution was stirred for 1 hour at room temperature. The reaction was then quenched by the addition of 100 mL of water/ice. The resulting solution was extracted with ethyl acetate (3×50 mL) and the organic layers combined. The resulting mixture was washed with brine (2×50 mL), dried over anhydrous sodium sulfate and concentrated under vacuum. The residue was applied onto a silica gel column with ethyl acetate/petroleum ether (1:4). This resulted in 7 g (62%) of (2R)-2-[(tert-butyldimethylsilyl)oxy]-N-[trans-3-(1,3-dioxo-2,3-dihydro-1H-isoindol-2-yl)cyclobutyl]carbonyl]propanehydrazide as colorless oil. LC-MS (ES, m/z): [M+1].sup.+=446.

[0208] Step 4: 2-[trans-3-[5-[(1R)-1-[(tert-butyldimethylsilyl)oxy]ethyl]-1,3,4-oxadiazol-2-yl]cyclobutyl]-2,3-dihydro-1H-isoindole-1,3-dione: into a 250-mL round-bottom flask, was placed a solution of (2R)-2-[(tert-butyldimethylsilyl)oxy]-N-[[trans-3-(1,3-dioxo-2,3-dihydro-1H-isoindol-2-yl)cyclobutyl]carbonyl]propanehydrazide (6.95 g, 15.60 mmol, 1.00 equiv) and TEA (7.89 g, 77.97 mmol, 5.00 equiv) in dichloromethane (100 mL). This was followed by the addition of a solution of 4-methylbenzene-1-sulfonyl chloride (8.92 g, 46.79 mmol, 3.00 equiv) in dichloromethane (50 mL) dropwise with stirring at 0° C. The resulting solution was stirred for 15 hours at room temperature. The reaction was then quenched by the addition of 100 mL of water/ice. The resulting solution was extracted with dichloromethane (2×50 mL) and the organic layers combined. The resulting mixture was washed with brine (2×50 mL), dried over anhydrous sodium sulfate and concentrated under vacuum. The crude product was purified by Flash-Prep-HPLC with the following conditions (IntelFlash-1): Column, C18; mobile phase, H.sub.2O/CH.sub.3CN=100:1 increasing to H.sub.2O/CH.sub.3CN=1:100 within 30 min; Detector, UV 254 nm. This resulted in 3.28 g (49%) of 2-[trans-3-[5-[(1R)-1-[(tert-butyldimethylsilyl)oxy]ethyl]-1,3,4-oxadiazol-2-yl]cyclobutyl]-2,3-dihydro-1H-isoindole-1,3-dione as colorless oil. LC-MS (ES, m/z): [M+1].sup.+=428. .sup.1H NMR (400 MHz, CDCl.sub.3): δ 7.72-7.70 (m, 2H), 7.60-7.58 (m, 2H), 5.04-4.96 (m, 2H), 3.83-3.78 (m, 1H), 3.26-3.24 (m, 2H), 2.67-2.62 (m, 2H), 1.49-1.48 (d, J=6.8 Hz, 3H), 0.76 (s, 9H), 0.01 (s, 3H), 0.00 (s, 3H).

[0209] Step 5: trans-3-[5-[(1R)-1-[(tert-butyldimethylsilyl)oxy]ethyl]-1,3,4-oxadiazol-2-yl]cyclobutan-1-amine: into a 250-mL round-bottom flask, was placed a solution of 2-[trans-3-[5-[(1R)-1-[(tert-butyldimethylsilyl)oxy]ethyl]-1,3,4-oxadiazol-2-yl]cyclobutyl]-2,3-dihydro-1H-isoindole-1,3-dione (1.18 g, 2.76 mmol, 1.00 equiv) in ethanol (100 mL). To the solution was added hydrazine hydrate (3.45 g, 55.13 mmol, 20.00 equiv, 80%). The resulting solution was stirred for 3 hours at room temperature. The solids were filtered out. The resulting mixture was concentrated under vacuum. This resulted in 760 mg (crude) of trans-3-[5-[(1R)-1-[(tert-butyldimethylsilyl)oxy]ethyl]-1,3,4-oxadiazol-2-yl]cyclobutan-1-amine as colorless oil. LC-MS (ES, m/z): [M+1].sup.+=298.

[0210] Step 6: N-[trans-3-[5-[(1R)-1-[(tert-butyldimethylsilyl)oxy]ethyl]-1,3,4-oxadiazol-2-yl]cyclobutyl]-1,3-thiazole-2-carboxamide: into a 100-mL round-bottom flask, was placed a solution of lithio 5-phenyl-1,3-thiazole-2-carboxylate (332 mg, 1.57 mmol, 1.20 equiv), (1r,3r)-3-[5-[(1R)-1-[(tert-butyldimethylsilyl)oxy]ethyl]-1,3,4-oxadiazol-2-yl]cyclobutan-1-amine (390 mg, 1.31 mmol, 1.00 equiv) and HATU (750 mg, 1.97 mmol, 1.50 equiv) in THF (50 mL). This was followed by the addition of DIEA (508 mg, 3.93 mmol, 3.00 equiv) dropwise with stirring at 0° C. The resulting solution was stirred for 1 hour at room temperature. The resulting solution was diluted with 50 mL of water/ice. The resulting solution was extracted with ethyl acetate (3×50 mL) and the organic layers combined. The resulting mixture was washed with brine (2×30 mL), dried and concentrated under vacuum. This resulted in 300 mg (47%) of 5-phenyl-N-[trans-3-[5-[(1R)-1-[(tert-butyldimethylsilyl)oxy]ethyl]-1,3,4-oxadiazol-2-yl]cyclobutyl]-1,3-thiazole-2-carboxamide as a light yellow crude solid. LC-MS (ES, m/z): [M+1].sup.+=485.

[0211] Step 7: 5-phenyl-N-[trans-3-[5-[(1R)-1-hydroxyethyl]-1,3,4-oxadiazol-2-yl]cyclobutyl]-1,3-thiazole-2-carboxamide: into a 50-mL round-bottom flask, was placed a solution of 5-phenyl-N-[(1r,3r)-3-[5-[(1R)-1-[(tert-butyldimethylsilyl)oxy]ethyl]-1,3,4-oxadiazol-2-yl]cyclobutyl]-1,3-thiazole-2-carboxamide (300 mg, 0.62 mmol, 1.00 equiv) in THF (2 mL). To the solution was added TBAF (1 mol/L in tetrahydrofuran, 1 mL). The resulting solution was stirred for 3 hours at room temperature. The resulting solution was diluted with 20 mL of water. The resulting solution was extracted with ethyl acetate (3×30 mL) and the organic layers combined. The resulting mixture was washed with brine (2×10 mL), dried over anhydrous sodium sulfate and concentrated under vacuum. The residue was applied onto a silica gel column with dichloromethane/methanol (20:1). The crude product was purified by Flash-Prep-HPLC with the following conditions (IntelFlash-1): Column, C18; mobile phase, H.sub.2O/CH.sub.3CN=100:1 increasing to H.sub.2O/CH.sub.3CN=1:100 within 30 min; Detector, UV 254 nm. This resulted in 115.6 mg (50%) of 5-phenyl-N-[trans-3-[5-[(1R)-1-hydroxyethyl]-1,3,4-oxadiazol-2-yl]cyclobutyl]-1,3-thiazole-2-carboxamide as a white solid. LC-MS (ES, m/z): [M+1].sup.+=371. .sup.1H NMR (400 MHz, DMSO-d.sub.6): δ 9.42-9.40 (d, J=8.0 Hz, 1H), 8.43 (s, 1H), 7.80-7.78 (m, 2H), 7.51-7.42 (m, 3H), 5.96-5.94 (d, J=6.0 Hz, 1H), 4.95-4.90 (m, 1H), 4.75-4.65 (m, 1H), 3.73-3.67 (m, 1H), 2.80-2.72 (m, 2H), 2.63-2.57 (m, 2H), 1.50-1.48 (d, J=6.4 Hz, 3H).

Example 12: N-trans-3-(5-(hydroxymethyl)-1H-1,2,3-triazol-1-yl)cyclobutyl)-5-phenylthiazole-2-carboxamide

[0212] ##STR00033##

[0213] Step 1: tert-butyl (3-oxocyclobutyl)carbamate: DPPA (4.0 g, 1.1 eq.) is added dropwise to a cold (−5-5° C.) solution of 3-oxocyclobutanecarboxylic acid (1.5 g, 1.0 eq.) and TEA (1.5 g, 1.1 eq.) in toluene (30 mL), and the mixture is stirred at −5˜0° C. for 16 h. The reaction mixture is washed with NaHCO.sub.3 (2*9 mL), water (1*9 mL) and NaCl aq. (1*4.5 mL) at 0˜10° C. The organic phase is dried over Na.sub.2SO.sub.4, filtered and t-BuOH (7.5 mL) is added to the filtrate. The reaction mixture is heated at 90˜100° C. for 16 h. The mixture is concentrated under vacuum at 60˜70° C. and then suspended in TBME (4.5 mL), filtered and the solid is dried over air to give 1.15 g (purity: 98.5%, yield: 47.2%) of product as a white solid.

[0214] Step 2: tert-butyl (cis-3-hydroxycyclobutyl)carbamate: a solution of tert-butyl (3-oxocyclobutyl)carbamate (200 mg, 1.0 eq.) in THF (1 mL) is added dropwise to a cold (below −70° C.) solution of NaBH.sub.4 (20.4 mg, 0.5 eq.) in a solution of THF (1.8 mL) and water (2 mL), maintaining the temperature at −80˜−70° C. (ca. for 2 h for completion of addition). The mixture is stirred at −60˜−50° C. for 3 h, water (2 mL) is added to the reaction mixture and allowed to reach up to 15° C. The reaction mixture is then extracted with ethyl acetate (2 mL, 2*1 mL) and the combined organic layers are washed with brine (1 mL). The organic layer is concentrated under vacuum at 35˜40° C., the solid is dissolved in toluene (1 mL, 80˜90° C.) and then is gradually cooled to 25-30° C. for 2.5 h. The mixture is stirred for 2 h at 25-30° C., filtered, and the solid is dried in the air to give the product (177 mg with ratio of cis:trans (96.4:3.6), yield: 87.6%) as an off-white solid.

[0215] Step 3: tert-butyl (trans-3-azidocyclobutyl)carbamate: a solution of PPh.sub.3 (315 mg) and DIAD (243 mg) in THF (3 mL) is stirred for 20 min at 0-10° C. Then a solution of tert-butyl (cis-3-hydroxycyclobutyl)carbamate (150 mg, 1.0 eq.) and DPPA (265 mg, 1.2 eq.) in THF (1 ml) is added dropwise, the mixture is then warmed to 25-30° C. and it is stirred for 2 h. Brine (3 mL) is added to the reaction mixture, extracted with ethyl acetate (3 mL) and is then concentrated under vacuum to give the crude oil. The mixture is purified by SiO.sub.2 column chromatography and is eluted with ethyl acetate/petroleum ether (0%˜10%) gradually. The product is suspended in n-heptane (0.3 mL) and stirred for 0.5 h at 20˜25° C., the mixture is filtered and the solid is dried in air to give the product in 85% yield and ratio of cis/trans=4:96 checked by .sup.1H NMR.

[0216] Step 4: tert-butyl (trans-3-(5-(hydroxymethyl)-1H-1,2,3-triazol-1-yl)cyclobutyl)carbamate: a solution of tert-butyl (trans-3-azidocyclobutyl)carbamate (246 mg, 1.0 eq.) and prop-2-yn-1-ol (326 mg, 5.0 eq.) in DMF (1.2 mL) is heated at 90˜95° C. for 20 h. The mixture is concentrated under vacuum at 65° C. to give ˜1:1 mixture of 4 and 5 regioisomers (353 mg). The mixture is purified by SFC to give tert-butyl (trans-3-(5-(hydroxymethyl)-1H-1,2,3-triazol-1-yl)cyclobutyl)carbamate (101 mg, P: 99.9% (205 nm), Y: 32%) as a solid.

[0217] Step 5: (1-(trans-3-aminocyclobutyl)-1H-1,2,3-triazol-5-yl)methanol hydrochloride: tert-butyl (trans-3-(5-(hydroxymethyl)-1H-1,2,3-triazol-1-yl)cyclobutyl)carbamate (101 mg, 1.0 eq.) is added slowly (5 portions) to a solution of HCl/dioxane (3.5 mol/L, 2 mL) at 20˜30° C., and then is stirred for 18 h at 20˜30° C. The reaction mixture is concentrated under vacuum at 55° C. to give the product (93.4 mg, assay 67% based on free base, Y: 100%) as a solid.

[0218] Step 6: of 5-phenyl-N-trans-3-[5-(hydroxymethyl)-1H-1,2,3-triazol-1-yl]cyclobutyl]-1,3-thiazole-2-carboxamide as a white solid: DIEA (386 mg, 2.99 mmol, 3.00 equiv) was added dropwise to a cold solution 0° C. of lithio 5-phenyl-1,3-thiazole-2-carboxylate (180 mg, 0.85 mmol, 1.00 equiv), [1-[trans-3-aminocyclobutyl]-1H-1,2,3-triazol-5-yl]methanol hydrochloride (204 mg, 1.00 mmol, 1.00 equiv) and HATU (567 mg, 1.49 mmol, 1.50 equiv) in DMF (5 mL). The resulting solution was stirred for 1 hour at room temperature. The reaction was then quenched by the addition of 50 mL of water/ice. The resulting solution was extracted with ethyl acetate (3×80 mL) and the organic layers combined. The resulting mixture was washed with brine (3×30 mL), dried over anhydrous sodium sulfate and concentrated under vacuum. The crude mixture was purified by Flash-Prep-HPLC with the following conditions (IntelFlash-1): Column, C18; mobile phase, H.sub.2O/CH.sub.3CN=100:1 increasing to H.sub.2O/CH.sub.3CN=1:100 within 35 min; Detector, UV 254 nm. This resulted in 115 mg (38%) of 5-phenyl-N-trans-3-[5-(hydroxymethyl)-1H-1,2,3-triazol-1-yl]cyclobutyl]-1,3-thiazole-2-carboxamide as a white solid. LC-MS (ES, m/z): [M+1].sup.+=356. .sup.1H NMR (400 MHz, DMSO-d.sub.6): δ 9.47-9.45 (d, J=8.0 Hz, 1H), 8.45 (s, 1H), 7.81-7.79 (d, J=7.6 Hz, 2H), 7.65 (s, 1H), 7.52-7.42 (m, 3H), 5.43 (s, 1H), 5.26-5.19 (m, 1H), 4.85-4.76 (m, 1H), 4.56 (s, 2H), 2.90-2.81 (m, 4H).

Example 13: CFTR Activity Assays

[0219] i. Ussing Measurements

[0220] As discussed above, Ussing measurements are used to measure CFTR activity. In this method, primary lung epithelial cells (hBEs) homozygous for the Cystic Fibrosis-causing ΔF508 mutation are differentiated for a minimum of 4 weeks in an air-liquid interface on SnapWell filter plates prior to the Ussing measurements. Cells are apically mucus-washed for 30 minutes prior to treatment with compounds. The basolateral media is removed and replaced with media containing the compound of interest diluted to its final concentration from DMSO stocks. Treated cells are incubated at 37° C. and 5% CO.sub.2 for 24 hours. At the end of the treatment period, the cells on filters are transferred to the Ussing chamber and equilibrated for 30 minutes. The short-circuit current is measured in voltage clamp-mode (V.sub.hold=0 mV), and the entire assay is conducted at a temperature of 36° C. −36.5° C. Once the voltages stabilized, the chambers are clamped, and data is recorded by pulse readings every 5 seconds. Following baseline current stabilization, the following additions can be applied and the changes in current and resistance of the cells can be monitored:

[0221] 1. Benzamil to the apical chamber to inhibit ENaC sodium channel.

[0222] 2. Forskolin to both chambers to activate ΔF508-CFTR by phosphorylation.

[0223] 3. Genistein to both chambers to potentiate ΔF508-CFTR channel opening.

[0224] 4. CFTRinh-172 to the apical chamber to inhibit ΔF508-CFTR Cl-conductance.

[0225] The inhibitable current (that current that is blocked by CFTRinh-172) is measured as the specific activity of the ΔF508-CFTR channel, and increases in response to compound in this activity over that observed in vehicle-treated samples are identified as the correction of ΔF508-CFTR function imparted by the compound tested.

[0226] ii. hBE Equivalent Current (Ieq) Assay

[0227] Primary lung epithelial cells homozygous for the Cystic Fibrosis-causing ΔF508 mutation were differentiated for a minimum of 4 weeks in an air-liquid interface on Costar 24 well HTS filter plates prior to the equivalent current (Ieq) measurements. Cells were apically mucus-washed for 30 minutes 24h prior to treatment with compounds. The basolateral media was removed and replaced with media containing the compound of interest diluted to its final concentration from DMSO stocks. Treated cells were incubated at 37° C. and 5% CO.sub.2 for 24 hours. At the end of the treatment period, the media was changed to the Ieq experimental solution for 30 minutes before the experiment and plates are maintained in a CO.sub.2-free incubator during this period. The plates containing the cells were then placed in pre-warmed heating blocks at 36° C.±0.5 for 15 minutes before measurements are taken. The transepithelial voltage (V.sub.T) and conductance (G.sub.T) were measured using a custom 24 channel current clamp (TECC-24) with 24 well electrode manifold. The Ieq assay measurements were made following additions with standardized time periods: [0228] 1. The baseline V.sub.T and G.sub.T values were measured for approximately 20 minutes. [0229] 2. Benzamil was added to block ENaC for 15 minutes. [0230] 3. Forskolin plus VX-770 were added to maximally activate ΔF508-CFTR for 27 minutes. [0231] 4. Bumetanide was added to inhibit the NaK.sub.2Cl cotransporter and shut-off secretion of chloride.

[0232] The activity data captured was the area under the curve (AUC) for the traces of the equivalent chloride current. The AUC was collected from the time of the forskolin/VX-770 addition until the inhibition by bumetanide addition. Correction in response to compound treatment was scored as the increase in the AUC for compound-treated samples over that of vehicle-treated samples. The results are shown below in Table 2. (** indicates activity ≧200% of VX-809 (1 uM) with compound at 10 uM and VX-809 at 1 uM; * indicates activity 100-200% of VX-809 (1 uM) with compound at 10 uM and VX-809 at 1 uM. .sup.##indicates activity ≧200% of VX-809 (3 uM) with compound at 10 uM and VX-809 at 3 uM; .sup.#indicates activity 100-200% of VX-809 (3 uM) with compound at 10 uM and VX-809 at 3 uM

TABLE-US-00004 TABLE 2 # Structure leq Ussing 1 [00034] ** 2 [00035] ** 3 [00036] ** 4 [00037] ** 5 [00038] * 6 [00039] ** ## 7 [00040] ** ## 8 [00041] ** ## 9 [00042] ** 10 [00043] ## 11 [00044] ## 12 [00045] ##

Example 14

[0233] i. Ussing Measurements

[0234] As discussed above, Ussing measurements can be used to measure CFTR activity. In this method, primary lung epithelial cells (hBEs) with a Cystic fibrosis causing class I mutation are differentiated for a minimum of 4 weeks in an air-liquid interface on SnapWell™ filter plates prior to the Ussing measurements. Cells are apically mucus-washed for 30 minutes prior to treatment with compounds. The basolateral media is removed and replaced with media containing the compound of interest diluted to its final concentration from DMSO or aqueous stocks. Treated cells are incubated at 37° C. and 5% CO.sub.2 for 24 hours. At the end of the treatment period, the cells on filters are transferred to the Ussing chamber and equilibrated for 30 minutes. The short-circuit current is measured in voltage clamp-mode (V.sub.hold=0 mV), and the entire assay is conducted at a temperature of 36° C.-36.5° C. Once the voltages stabilize, the chambers are clamped, and data are recorded by pulse readings every 5 seconds. Following baseline current stabilization, the following additions are applied and the changes in current and resistance of the cells are monitored:

[0235] 1. Benzamil to the apical chamber to inhibit ENaC sodium channel.

[0236] 2. Forskolin to both chambers to activate ΔF508-CFTR by phosphorylation.

[0237] 3. Ivacaftor or Genistein to the apical chamber to potentiate ΔF508-CFTR channel opening.

[0238] 4. CFTRinh-172 to the apical chamber to inhibit ΔF508-CFTR Cl-conductance.

[0239] The forskolin-sensitive current and inhibitable current (that potentiated current that is blocked by CFTRinh-172) are measured as the specific activity of the ΔF508-CFTR channel, and increase in response to compound in this activity over that observed in vehicle-treated samples are identified as the correction of ΔF508-CFTR function imparted by the compound tested.

Example 15

[0240] i. Ussing Measurements

[0241] As discussed above, Ussing measurements can be used to measure CFTR activity. In this method, primary lung epithelial cells (hBEs) with a Cystic Fibrosis-causing class III mutation are differentiated for a minimum of 4 weeks in an air-liquid interface on SnapWell™ filter plates prior to the Ussing measurements. Cells are apically mucus-washed for 30 minutes prior to treatment with compounds. The basolateral media is removed and replaced with media containing the compound of interest diluted to its final concentration from DMSO stocks. Treated cells are incubated at 37° C. and 5% CO.sub.2 for 24 hours. At the end of the treatment period, the cells on filters are transferred to the Ussing chamber and equilibrated for 30 minutes. The short-circuit current is measured in voltage clamp-mode (V.sub.hold=0 mV), and the entire assay is conducted at a temperature of 36° C.-36.5° C. Once the voltages stabilize, the chambers are clamped, and data is recorded by pulse readings every 5 seconds. Following baseline current stabilization, the following additions are applied and the changes in current and resistance of the cells is monitored:

[0242] 1. Benzamil to the apical chamber to inhibit ENaC sodium channel.

[0243] 2. Forskolin to both chambers to activate ΔF508-CFTR by phosphorylation.

[0244] 3. VX-770 or Genistein to the apical chamber to potentiate ΔF508-CFTR channel opening.

[0245] 4. CFTRinh-172 to the apical chamber to inhibit ΔF508-CFTR Cl-conductance.

[0246] The forskolin-sensitive current and inhibitable current (that potentiated current that is blocked by CFTRinh-172) are measured as the specific activity of the ΔF508-CFTR channel, and increase in response to compound in this activity over that observed in vehicle-treated samples are identified as the correction of ΔF508-CFTR function imparted by the compound tested.

Example 16

[0247] i. Ussing Measurements

[0248] As discussed above, Ussing measurements can be used to measure CFTR activity. In this method, primary lung epithelial cells (hBEs) with a Cystic Fibrosis-causing class V mutation are differentiated for a minimum of 4 weeks in an air-liquid interface on SnapWell™ filter plates prior to the Ussing measurements. Cells are apically mucus-washed for 30 minutes prior to treatment with compounds. The basolateral media is removed and replaced with media containing the compound of interest diluted to its final concentration from DMSO stocks. Treated cells are incubated at 37° C. and 5% CO.sub.2 for 24 hours. At the end of the treatment period, the cells on filters are transferred to the Ussing chamber and equilibrated for 30 minutes. The short-circuit current is measured in voltage clamp-mode (V.sub.hold=0 mV), and the entire assay is conducted at a temperature of 36° C.-36.5° C. Once the voltages stabilize, the chambers are clamped, and data is recorded by pulse readings every 5 seconds. Following baseline current stabilization, the following additions are applied and the changes in current and resistance of the cells is monitored:

[0249] 1. Benzamil to the apical chamber to inhibit ENaC sodium channel.

[0250] 2. Forskolin to both chambers to activate ΔF508-CFTR by phosphorylation.

[0251] 3. VX-770 or Genistiein to the apical chamber to potentiate ΔF508-CFTR channel opening.

[0252] 4. CFTRinh-172 to the apical chamber to inhibit ΔF508-CFTR Cl-conductance.

[0253] The forskolin-sensitive current and inhibitable current (that potentiated current that is blocked by CFTRinh-172) are measured as the specific activity of the ΔF508-CFTR channel, and increases in response to compound in this activity over that observed in vehicle-treated samples are identified as the correction of ΔF508-CFTR function imparted by the compound tested.

[0254] ii. hBE Equivalent Current (Ieq) Assay

[0255] Primary lung epithelial cells homozygous for the Cystic Fibrosis-causing ΔF508 mutation are differentiated for a minimum of 4 weeks in an air-liquid interface on Costar 24 well HTS filter plates prior to the equivalent current (Ieq) measurements. Cells are apically mucus-washed for 30 minutes 24h prior to treatment with compounds. The basolateral media is removed and replaced with media containing the compound of interest diluted to its final concentration from DMSO stocks. Treated cells are incubated at 37° C. and 5% CO.sub.2 for 24 hours. At the end of the treatment period, the media is changed to the Ieq experimental solution for 30 minutes before the experiment and plates are maintained in a CO.sub.2-free incubator during this period. The plates containing the cells are then placed in pre-warmed heating blocks at 36° C.±0.5 for 15 minutes before measurements are taken. The transepithelial voltage (V.sub.T) and conductance (G.sub.T) are measured using a custom 24 channel current clamp (TECC-24) with 24 well electrode manifold. The Ieq assay measurements are made following additions with standardized time periods: [0256] 1. The baseline V.sub.T and G.sub.T values are measured for approximately 20 minutes. [0257] 2. Benzamil is added to block ENaC for 15 minutes. [0258] 3. Forskolin plus VX-770 (ivacaftor) are added to maximally activate ΔF508-CFTR for 27 minutes. [0259] 4. Bumetanide is added to inhibit the NaK.sub.2Cl cotransporter and shut-off secretion of chloride.

[0260] The activity data captured is the area under the curve (AUC) for the traces of the equivalent chloride current. The AUC is collected from the time of the forskolin/VX-770 addition until the inhibition by bumetanide addition. Correction in response to compound treatment is scored as the increase in the AUC for compound-treated samples over that of vehicle-treated samples.

[0261] While this invention has been particularly shown and described with references to preferred embodiments thereof, it will be understood by those skilled in the art that various changes in form and details may be made therein without departing from the scope of the invention encompassed by the appended claims.

INCORPORATION BY REFERENCE

[0262] All publications and patents mentioned herein, including those items listed below, are hereby incorporated by reference in their entirety for all purposes as if each individual publication or patent was specifically and individually incorporated by reference. In case of conflict, the present application, including any definitions herein, will control.

EQUIVALENTS

[0263] While specific embodiments of the subject invention have been discussed, the above specification is illustrative and not restrictive. Many variations of the invention will become apparent to those skilled in the art upon review of this specification. The full scope of the invention should be determined by reference to the claims, along with their full scope of equivalents, and the specification, along with such variations.

[0264] Unless otherwise indicated, all numbers expressing quantities of ingredients, reaction conditions, and so forth used in the specification and claims are to be understood as being modified in all instances by the term “about.” Accordingly, unless indicated to the contrary, the numerical parameters set forth in this specification and attached claims are approximations that may vary depending upon the desired properties sought to be obtained by the present invention.