WO2010116270A1 - Ep2/4 agonists - Google Patents

Abstract

EP2/4 compounds having improved dual pharmacological activity are described. The uniqueness of using EP2/4 dual agonists resides in their ability to modify both uveoscleral outflow via the ciliary muscle and conventional outflow via trabecular meshwork and Schlemm's canal all in the same treatment paradigm. The compounds can be employed for the treatment of glaucoma and ocular hypertension. Formula (I).

Description

EP2/4 AGONISTS

This application claims the benefit of U. S. Provisional Application No. 61/168,344, filed April 10, 2009, the contents of which are hereby incorporated by reference in its entirety. Background of Invention

Glaucoma is a progressive disease which leads to optic nerve damage, and, ultimately, total loss of vision. The causes of this disease have been the subject of extensive studies for many years, but are still not fully understood. The principal symptom of and/or risk factor for the disease is elevated intraocular pressure or ocular hypertension due to excess aqueous humor in the anterior chamber of the eye. The causes of aqueous humor accumulation in the anterior chamber are not fully understood. It is known that elevated intraocular pressure can be at least partially controlled by administering drugs which reduce either the production of aqueous humor within the eye, such as beta-blockers and carbonic anhydrase inhibitors, or increase the flow of aqueous humor out of the eye, such as miotics and sympathomimetics. Latanoprost, a novel prostaglandin F₂α analogue, is a selective prostanoid FP receptor agonist which reduces the intraocular pressure by increasing the outflow of aqueous humor.

The relationship between EP receptor activation and intraocular pressure lowering effects is well known. There are currently four recognized subtypes of the

EP receptor: EP1 , EP2, EP3, and EP4 (J. Lipid Mediators Cell Signaling, volume 14, pages 83-87 (1996)). Intraocular pressure (lOP) may be lowered by ligands capable of EP2 and EP4 receptor activation, such as PGE2 and certain of its synthetic analogs (Journal of Ocular Pharmacology, volume 4, number 1 , pages 13-18 (1988); Journal of Ocular Pharmacology and Therapeutics, volume 11 , number 3, pages

447-454 (1995) Kharlamb EP4 references - see below).

Numerous publications have suggested the use of prostaglandin agonists for treating bone disorders and/or glaucoma, including: US 4599353; US5296504; W01998/028264; US6288120; US6492412; US6552067; US 6649657; US6747054; US7192979; US7414071 JP2000053566; EP1000619; US6344485; EP1205189;

US2002/0115695; US2002/0161026; US2004/0176423; WO2003/045371 ; US2003/0166631 ; WO1999/019300; US6498172; JP20011163779; EP1108426; US2005/203086; and WO2004/078169; the disclosures of each are hereby incorporated by reference in their entirety for all purposes. There remains, however, a continuing need in this field of art for alternative therapies for the treatment of glaucoma. See, for example: Biswas, S., Bhattacherjee, P. Paterson, CA. , 2004, Prostaglandin e2 receptor subtypes, ep1 , ep2, ep3 and ep4 in human and mouse ocular tissues-a comparative immunohistochemical study. Prostaglandins Leukot

Essent Fatty Acids, 71 , 277-288; Kharlamb, A., 2005, Further studies on the effect of prostanoid ep4 receptor stimulation on aqueous humor dynamics in monkeys, ARVO EAbstract: 3669; Kharlamb, A., 2006. Ciliary muscle relaxation does not explain the profound ocular hypotension produced by a selective prostanoid ep4 receptor agonist 3,7-dithia pge1 in monkeys, ARVO EAbstract: 413; Nilsson, S. F., Drecoll, E.,

Lutjen-Drecoll, E., Toris, C. B., Krauss, A. H., Kharlamb, A., Nieves, A., Guerra, T. Woodward, D. F., 2006. The prostanoid ep2 receptor agonist butaprost increases uveoscleral outflow in the cynomolgus monkey. Invest Ophthalmol Vis Sci. 47, 4042- 4049; Schlotzer-Schrehardt, U., Zenkel, M.Nusing, R.M., 2002. Expression and localization of fp and ep prostanoid receptor subtypes in human ocular tissues.

Invest Ophthalmol Vis Sci. 43, 1475-1487; and Toris, CB. , Camras, CB., Yablonski, M. E. Bm baker, R. F., 1997. Effects of exogenous prostaglandins on aqueous humor dynamics and blood-aqueous barrier function. Surv Ophthalmol. 41 Suppl 2, S69-75. There are currently no known approved treatments which can (as a single pharmacological therapeutic entity) affect both uveoscleral and conventional outflow simultaneously, thereby lowering lOP. The uniqueness of using EP2/4 dual agonists resides in their ability to modify both uveoscleral outflow via the ciliary muscle and conventional outflow via trabecular meshwork (TM) and Schlemm's canal all in the same treatment paradigm. Typically, EP2 agonism with Butaprost has been demonstrated to increase uveoscleral outflow, similar to that observed with FP agonists including latanoprost in preclinical models and patients with glaucoma (Toris et al., 1997; Nilsson et al., 2006). EP4 agonists, on the other hand, have been demonstrated in preclinical models to lower IOP by increasing conventional outflow and causing aqueous humor to exit the anterior chamber via the TM. An increase in conventional outflow facility via TM has been observed in experimental ocular hypertensive primates following treatment with 3,7-dithia PGE1 , an EP4 selective agonist (Kharlamb, 2005; Kharlamb et al., 2006). Both EP2 and EP4 receptors are expressed in both the TM and ciliary muscle tissues of human eyes (Schlotzer- Schrehardt et al., 2002; Biswas et al., 2004). Interestingly, EP4 receptor expression appears to be more abundant in TM cells than in ciliary muscle, suggestive of its role in increasing conventional outflow (Schlotzer-Schrehardt et al., 2002; Biswas et al., 2004). So it is expected that novel EP2/4 compounds with dual pharmacology will likely lead to increased IOP lowering due to the activation of two modalities, uveoscleral and conventional outflow since the 2 EP receptor subtypes are also located in TM and ciliary muscle.

Field of The Invention

The invention relates to EP2/4 dual agonists, methods for their preparation, pharmaceutical compositions containing these compounds, and methods of using these compounds and compositions for lowering intraocular pressure and thereby treating glaucoma.

SUMMARY OF THE INVENTION An object of the present invention is, therefore, a compound of formula (I):

wherein:

A¹, A², A³ and A⁴ are each independently selected from C, N, CR^A, NR^A, C-(O)R^A, N-(O)R^A, C-(R^AR^B) and N-(R^AR^B);

R¹, R², R³, R⁴ and R⁵ are each independently selected from a bond, H, halo, cyano, nitro, oxo, CF₃, OCF₃, Ci_₈ alkyl, C_2-8 alkenyl, C_2-8 alkynyl, C-ι_-8 hydroxyalkyl, Ci_-8 haloalkyl, C_3--_I0 cycloalkyl, C_4--_I0 heterocycloakyl, C_3--_I0 cycloalkoxy, C-ι_-8 alkoxy, Ci_-8 alkoxyalkyl, Cs_-I0 aryl, and Cs_-I0 heteroaryl, wherein each of said Ci_-8 alkyl, C_2-8 alkenyl, C_2-8 alkynyl, Ci_-8 hydroxyalkyl, Ci_-8 haloalkyl, C_{3- 10} cycloalkyl, C_4-I0 heterocycloakyl, C_3-I0 cycloalkoxy, Ci_-8 alkoxy, Ci_-8 alkoxyalkyl, Cs_-I0 aryl, and Cs_-I0 heteroaryl moieties is optionally substituted by one or more of R^A, R^AR^B, CR^A, CR^AR^B, SR^A, SR^AR^B, OR^A, COR^A, S(O)_aR^A, NR^AR^B, CONR^AR^B, N(O)R^AR^B, (CR^AR^B)_b(C_5-io aryl), (CR^AR^B)_b(C_5-io heteroaryl) or (CR^AR^B)_b(C_3-io cycloalkyl);

R^A and R^B are independently selected from a bond, H, halo, cyano, nitro, oxo, CF₃, OCF₃, Ci_-8 alkyl, C_2-8 alkenyl, C_2-8 alkynyl, C_5-io aryl, C₅-₁₀ heteroaryl, arylalkyl, C₃-₁₀ cycloalkyl, C_4--I₀ heterocycloakyl, C3-10 cycloalkoxy, Ci_-8 alkoxy, and C_2-S aI koxyalkyl;

D¹ is C, N, O, S, CR^A, NR^A, OR^A, SR^A, C-(O)R^A, N-(O)R^A, C-(O)OR^A, N-(O)OR^A, C-(R^AR^B), N-(R^AR^B), or S-(R^AR^B); and each a is independently selected from O, 1 , 2, 3, 4 and 5; each b is independently selected from 0, 1 , 2, 3, 4 and 5; or a pharmaceutically acceptable salt or stereoisomer thereof.

In another aspect of the invention, there is provided a compound of formula (I) as herein described wherein at least two of A¹, A², A³ and A⁴ are C.

In a further aspect of the invention, there is provided a compound of formula (I) as herein described wherein R¹ is selected from H, halo, cyano, CF₃, OCF₃, Ci_₈ alkyl, C_2-S alkenyl, C_2-S alkynyl, Ci_-8 hydroxyalkyl, Ci_-8 haloalkyl, C3-10 cycloalkyl, C4-10 heterocycloakyl, C3-10 cycloalkoxy, Ci_-8 alkoxy, Ci_-8 alkoxyalkyl, C_5-io aryl, and C5-10 heteroaryl.

In yet another aspect of the invention, there is provided a compound of formula (I) as herein described wherein R² is selected from H, halo, cyano, CF₃, OCF₃, Ci_-8 alkyl, C_2-8 alkenyl, C_2-8 alkynyl, Ci_-8 hydroxyalkyl, Ci_-8 haloalkyl, C_3-io cycloalkyl, C_4-io heterocycloakyl, C_3-i₀ cycloalkoxy, Ci_-8 alkoxy, Ci_-8 alkoxyalkyl, C_5-io aryl, and C_5-i₀ heteroaryl.

In still another aspect of the invention, there is provided a compound of formula (I) as herein described wherein R³ is C5-10 aryl or C5-10 heteroaryl, and wherein each of said C5-10 aryl or C5-10 heteroaryl moieties is optionally substituted by halo, cyano, Ci_-8 alkyl, C_2-8 alkenyl, C_2-8 alkynyl, Ci_-8 hydroxyalkyl, Ci_-8 haloalkyl, C3-10 cycloalkyl, C4-10 heterocycloakyl, C₃-₁₀ cycloalkoxy, Ci_-8 alkoxy or Ci_-8 alkoxyalkyl.

In another aspect of the invention, there is provided a compound of formula (I) as herein described wherein R⁵ is (CR^AR^B)_b(C_5-io aryl) or (CR^AR^B)_b(C_5-io heteroaryl), and wherein each of said (CR^AR^B)_b(C_5-io aryl) and (CR^AR^B)_b(C_5-io heteroaryl) moieties is independently substituted by at least one H, halo, cyano, nitro, oxo, CF₃, OCF₃, Ci_-8 alkyl, C_2-8 alkenyl, C_2-8 alkynyl, Ci_-8 hydroxyalkyl, Ci_-8 haloalkyl, C_3-i₀ cycloalkyl, C_4-i₀ heterocycloakyl, C3-io cycloalkoxy, Ci_-8 alkoxy, Ci_-8 alkoxyalkyl, C_5-io aryl, or C_5-io heteroaryl moiety.

In a further aspect of the invention, there is provided a compound of formula (I) as herein described wherein R¹ is selected from H, halo, cyano, CF₃, OCF₃, Ci_-8 alkyl, C_2-8 alkenyl, C_2-8 alkynyl, Ci_-8 hydroxyalkyl, Ci_-8 haloalkyl, C_3-i₀ cycloalkyl, C_4--_I0 heterocycloakyl, C3-10 cycloalkoxy, Ci_-8 alkoxy, Ci_-8 alkoxyalkyl, Cs-io aryl, and C5-10 heteroaryl; and R³ is C5-10 aryl or C5-10 heteroaryl, and wherein each of said C5-10 aryl or C_5-io heteroaryl moieties of R³ is optionally substituted by halo, cyano, Ci_-8 alkyl, C_2-8 alkenyl, C_2-8 alkynyl, Ci_-8 hydroxyalkyl, Ci_-8 haloalkyl, C_3-io cycloalkyl, C_4-io heterocycloakyl, C_3-io cycloalkoxy, Ci_-8 alkoxy, Ci_-8 alkoxyalkyl, Cs-io aryl, or C_5-io heteroaryl.

In yet another aspect of the invention, there is provided a compound of formula (I) as herein described wherein R³ is C_5--_I0 aryl or C_5--_I0 heteroaryl, and wherein each of said C_5-I0 aryl or C_5--_I0 heteroaryl moieties is optionally substituted by Ci_-8 alkyl, C_2-8 alkenyl, C_2-8 alkynyl, Ci_-8 hydroxyalkyl, Ci_-8 haloalkyl, C_3-i₀ cycloalkyl, C4-10 heterocycloakyl, C3-10 cycloalkoxy, Ci_-8 alkoxy, Ci_-8 alkoxyalkyl, Cs-io aryl, or C5-10 heteroaryl; and R⁵ is (CR^AR^B)_b(C_5-10 aryl) or (CR^AR^B)_b(C_5-10 heteroaryl), and wherein each of said (CR^AR^B)_b(C_5-10 aryl) and (CR^AR^B)_b(C_5-10 heteroaryl) moieties of R⁴ is independently substituted by at least one H, halo, cyano, nitro, oxo, CF₃, OCF₃, Ci_-8 alkyl, C_2-8 alkenyl, C_2-8 alkynyl, Ci_-8 hydroxyalkyl, Ci_-8 haloalkyl, C_3-i₀ cycloalkyl, C_4-I0 heterocycloakyl, C_3-io cycloalkoxy, Ci_-8 alkoxy, Ci_-8 alkoxyalkyl, C_5-i₀ aryl, or C_5--_I0 heteroaryl moiety.

In still another aspect of the invention, there is provided a compound of formula (II):

wherein: A⁵, A⁶, A⁷ and A⁸ are each independently selected from C, N, CR^C, NR^C, C- (O)R^C, N-(O)R⁰, C-(R^CR^D) and N-(R^CR^D);

R⁶ and R⁷ are each independently selected from a bond, H, halo, cyano, nitro, oxo, CF₃, OCF₃, C-_I-8 alkyl, C_2-8 alkenyl, C₂-₈ alkynyl, Ci_-8 hydroxyalkyl, Ci_-8 haloalkyl, C_3- io cycloalkyl, C4-10 heterocycloakyl, C3-10 cycloalkoxy, Ci_-8 alkoxy, Ci_-8 alkoxyalkyl, C5-10 aryl, and C_5-io heteroaryl, wherein each of said Ci_-8 alkyl, C_2-8 alkenyl, C_2-8 alkynyl, Ci_-8 hydroxyalkyl, Ci_-8 haloalkyl, C3-10 cycloalkyl, C_4- 10 heterocycloakyl, C3-10 cycloalkoxy, Ci_-8 alkoxy, Ci_-8 alkoxyalkyl, C_5-io aryl, and C₅-₁₀ heteroaryl moieties is optionally substituted by one or more of R^c, R^CR^D, CR^C, CR^CR^D, NR^C, OR^C, SR^C, SR^CR^D, COR^C, S(O)_nR⁰, NR^CR^D, CONR^CR^D, N(O)R^CR^D, (CR^cR^D)_m(C_5-io aryl), (CR^cR^D)_m(C_5-io heteroaryl) or (CR^cR^D)_m(C_3-io cycloalkyl);

R⁸ is H, halo, cyano, nitro, oxo, CF₃, OCF₃, Ci_-8 alkyl, C_2-8 alkenyl, C_2-8 alkynyl, Ci- ₈ hydroxyalkyl, Ci_-8 haloalkyl, C_3-i₀ cycloalkyl, C_{4- 10} heterocycloakyl, C_3-i₀ cycloalkoxy, Ci- ₈ alkoxy, Ci_-8 alkoxyalkyl, C_5-i₀ aryl, or C_5-i₀ heteroaryl; R⁹ is H, halo, cyano, nitro, oxo, CF₃, OCF₃, Ci_-8 alkyl, C_2-8 alkenyl, C_2-8 alkynyl, Ci-

₈ hydroxyalkyl, Ci_-8 haloalkyl, C3-10 cycloalkyl, C_4- 10 heterocycloakyl, C3-10 cycloalkoxy, Ci- ₈ alkoxy, Ci_-8 alkoxyalkyl, C_5-io aryl, or C₅-₁₀ heteroaryl, wherein each of said Ci_-8 alkyl, C_2-8 alkenyl, C_2-8 alkynyl, Ci_-8 hydroxyalkyl, Ci_-8 haloalkyl, C₃-₁₀ cycloalkyl, C_4-i₀ heterocycloakyl, C_3-io cycloalkoxy, Ci_-8 alkoxy, Ci_-8 alkoxyalkyl, Cs-io aryl, and Cs-io heteroaryl moieties is optionally substituted by one or more of CR^C, CR^CR^D, SR^C, SR^CR^D, OR^C, COR^C, S(O)_nR⁰, NR^CR^D, CONR^CR^D, N(O)R^CR^D, (CR^cR^D)_m(C_5-i₀ aryl), (CR^cR^D)_m(C_5-io heteroaryl) or (CR^cR^D)_m(C_3-i₀ cycloalkyl);

R¹⁰ is H, halo, cyano, nitro, oxo, CF₃, OCF₃, Ci_-8 alkyl, C_2-8 alkenyl, C_2-8 alkynyl, Ci_-8 hydroxyalkyl, Ci_-8 haloalkyl, C3-10 cycloalkyl, C_4-io heterocycloakyl, C3-10 cycloalkoxy, Ci_-8 alkoxy, Ci_-8 alkoxyalkyl, C5-10 aryl, or C5-10 heteroaryl;

R¹¹ is a bond, oxo, R^c, OR^C, Ci_-8 alkyl, C_2-8 alkenyl, C_2-8 alkynyl, Ci_-8 hydroxyalkyl, Ci_-8 haloalkyl, C3-10 cycloalkyl, C_4-i₀ heterocycloakyl, C3-10 cycloalkoxy, Ci_-8 alkoxy, Ci_-8 alkoxyalkyl, C_5-io aryl, or C₅-₁₀ heteroaryl, wherein each of said Ci_-8 alkyl, C_2-8 alkenyl, C_2- s alkynyl, Ci_-8 hydroxyalkyl, Ci_-8 haloalkyl, C_3-io cycloalkyl, C_4-io heterocycloakyl, C_3-io cycloalkoxy, Ci_-8 alkoxy, Ci_-8 alkoxyalkyl, Cs-io aryl, and Cs-io heteroaryl moieties is optionally substituted by one or more of CR^C, CR^CR^D, SR^C, SR^CR^D, OR^C, COR^C, S(O)_mR^c, NR^CR^D, CONR^CR^D, N(O)R^CR^D, (CR^cR^D)_m(C_5-i₀ aryl), (CR^cR^D)_m(C_5-io heteroaryl) or (CR^cR^D)_m(C_3-i₀ cycloalkyl); R^c and R^D are independently selected from a bond, H, halo, cyano, nitro, oxo, CF₃, OCF₃, C-I_-8 alkyl, C_2-8 alkenyl, C_2-8 alkynyl, C_5-io aryl, C_5-io heteroaryl, arylalkyl, C3-10 cycloalkyl, C_4-i₀ heterocycloakyl, C_3-i₀ cycloalkoxy, Ci_-8 alkoxy, and C_2-S aI koxyalkyl;

D² is C, N, O, S, CR^C, NR^C, OR^C, SR^C, C-(O)R⁰, N-(O)R⁰, C-(O)OR⁰, N-(O)OR⁰, C-(R⁰R⁰), N-(R⁰R⁰), or S-(R⁰R⁰); and each m is independently selected from O, 1 , 2, 3, 4 and 5; and each n is independently selected from O, 1 , 2, 3, 4 and 5; or a pharmaceutically acceptable salt or stereoisomer thereof.

In another aspect of the invention, there is provided a compound of formula (II) as herein described wherein at least two of A⁵, A⁶, A⁷ and A⁸ are C.

In yet another aspect of the invention, there is provided a compound of formula (II) as herein described wherein R⁶ is selected from H, halo, cyano, CF₃, OCF₃, Ci_-8 alkyl, C_2-8 alkenyl, C_2-8 alkynyl, Ci_-8 hydroxyalkyl, Ci_-8 haloalkyl, C_3--_I0 cycloalkyl, C_4--_I0 heterocycloakyl, C_3--_I0 cycloalkoxy, C-ι_-8 alkoxy, C-ι_-8 alkoxyalkyl, C_5--_I0 aryl, and C_5--_I0 heteroaryl.

In still another aspect of the invention, there is provided a compound of formula (II) as herein described wherein R⁷ is selected from H, halo, cyano, CF₃, OCF₃, Ci_-8 alkyl, C_2-8 alkenyl, C_2-8 alkynyl, C-ι_-8 hydroxyalkyl, C-ι_-8 haloalkyl, C_3--_I0 cycloalkyl, C_4--_I0 heterocycloakyl, C_3--_I0 cycloalkoxy, C-ι_-8 alkoxy, C-i_-8 alkoxyalkyl, C_5--_I0 aryl, and C_5--_I0 heteroaryl.

In a further aspect of the invention, there is provided a compound of formula (II) as herein described wherein R⁹ is H, halo, cyano, nitro, oxo, CF₃, OCF₃, Ci_-8 alkyl, C_2-8 alkenyl, C_2-8 alkynyl, Ci_-8 hydroxyalkyl, Ci_-8 haloalkyl, C_3-I0 cycloalkyl, C_4-I0 heterocycloakyl, C_3-I0 cycloalkoxy, Ci_-8 alkoxy, Ci_-8 alkoxyalkyl, C_5-i₀ aryl, or C_5-i₀ heteroaryl; and n is O, 1 , 2 or 3.

In another aspect of the invention, there is provided a compound of formula (II) as herein described wherein R¹⁰ is halo, cyano, Ci_-8 alkyl, C_2-8 alkenyl, C_2-8 alkynyl, Ci_-8 hydroxyalkyl, Ci_-8 haloalkyl, C_3-I0 cycloalkyl, C_4-I0 heterocycloakyl, C_3-I0 cycloalkoxy, Ci_-8 alkoxy, Ci_-8 alkoxyalkyl, C_5-i₀ aryl, or C_5-I0 heteroaryl. In yet another aspect of the invention, there is provided a compound of formula

(II) as herein described wherein R¹¹ is Ci_-8 alkyl, C_2-8 alkenyl, C_2-8 alkynyl, Ci_-8 hydroxyalkyl, Ci_-8 haloalkyl, C_3-I0 cycloalkyl, C_4-I0 heterocycloakyl, C_3-I0 cycloalkoxy, Ci_-8 alkoxy or Ci_-8 alkoxyalkyl. In still another aspect of the invention, there is provided a compound of formula (II) as herein described wherein n is 1 , 2 or 3.

In a further aspect of the invention, there is provided a compound selected from the group consisting of:

eutically acceptable salt or stereoisomer thereof. In another aspect of the invention, there is provided a compound as herein described for use as a medicament.

In a further aspect of the invention, there is provided a use of a compound as herein described for the preparation of a medicament for treating glaucoma and ocular hypertension.

In yet another aspect of the invention, there is provided a pharmaceutical composition comprising a pharmaceutically acceptable carrier and a pharmaceutically effective amount of a compound and/or a salt or stereoisomer thereof as herein described. In yet another aspect of the invention, there is provided a pharmaceutical composition as herein described in a suitable form for topical administration.

In still another aspect of the invention, there is provided a pharmaceutical composition as herein described for the treatment of glaucoma and ocular hypertension.

In another aspect of the invention, there is provided a pharmaceutical composition as herein described, wherein the compound is administered as a solution, suspension or emulsion in an ophthalmically acceptable vehicle.

In a further aspect of the invention, there is provided a method for treating glaucoma or ocular hypertension comprising contacting an effective intraocular pressure reducing amount of a pharmaceutical composition as herein described with the eye in order to reduce eye pressure and to maintain said pressure on a reduced level.

In yet another aspect of the invention, there is provided a pharmaceutical composition comprising a mixture of a compound and/or a salt or stereoisomer thereof as herein described and: (i) a beta-blocker; or (ii) a carbonic anhydrase inhibitor; or (iii) an adrenergic agonist; or a nitrooxy derivative of (i), (ii) or (iii). Definitions

As used herein, the terms "comprising" and "including" are used in their open, non-limiting sense.

As used herein, the term "substituted," means that the specified group or moiety bears one or more substituents. The term "unsubstituted," means that the specified group bears no substituents.

As used herein, the term "optionally substituted" means that the specified group is unsubstituted or is substituted by one or more substituents. As used herein, the terms "treat," "treating" or "treatment" includes preventative (e.g., prophylactic) and palliative treatment.

As used herein, the term "pharmaceutically acceptable" means the carrier, diluent, excipients and/or salt must be compatible with the other ingredients of the formulation and not deleterious to the recipient thereof.

As used herein, the term "alkyl" means a straight or branched chain saturated hydrocarbon. Exemplary alkyl groups include but are not limited to methyl, ethyl, propyl, isopropyl, n-butyl, sec-butyl, tert-butyl, pentyl, isopentyl, neopentyl, tert-pentyl, 1 - methylbutyl, 2-methylbutyl, 3-methylbutyl, hexyl, isohexyl, heptyl, octyl and the like. As used herein, the term "alkenyl" means a straight or branched chain hydrocarbon having at least one double bond, i.e., a C=C. Exemplary alkenyl groups include but are not limited to vinyl, propenyl, butenyl, pentenyl, hexenyl, heptenyl, octenyl and the like.

As used herein, the term "alkynyl" means a straight or branched chain hydrocarbon having at least one triple bond, i.e., a CΞC. Exemplary alkynyl groups include but are not limited to acetylenyl, propargyl, butynyl, pentynyl, hexynyl, heptynyl, octynyl and the like.

As used herein, the term "cycloalkyl" means a cyclic saturated hydrocarbon. Exemplary cycloalkyl groups include but are not limited to cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl and the like.

As used herein, the term "cycloalkenyl" means a cyclic hydrocarbon having at least one double bond, i.e., a C=C. Exemplary cycloalkenyl groups include but are not limited to cyclopropenyl, cyclobutenyl, cyclopentenyl, cyclohexenyl, cycloheptenyl, cyclooctenyl and the like. As used herein, the term "cycloalkynyl" means a cyclic hydrocarbon having at least one triple bond, i.e., a CΞC. Exemplary cycloalkynyl groups include but are not limited to cyclohexynyl, cycloheptynyl, cyclooctynyl and the like.

As used herein, the term "alkoxy" means a straight or branched chain saturated alkyl group bonded through oxygen. Exemplary alkoxy groups include but are not limited to methoxy, ethoxy, propoxy, isopropoxy, butoxy, isobutoxy, tert-butoxy, pentoxy, isopentoxy, neopentoxy, tert-pentoxy, hexoxy, isohexoxy, heptoxy, octoxy and the like.

As used herein, the term "alkylene" means a straight chain or branched chain saturated hydrocarbon wherein a hydrogen atom is removed from each of the terminal carbons. Exemplary alkylene groups include but are not limited to methylene, ethylene, propylene, butylene, pentylene, hexylene, heptylene and the like.

As used herein, the term "halo" or "halogen" means fluoro, chloro, bromo or iodo. As used herein, the terms "heterocyclic" and "heterocyclyl" mean an aromatic or non-aromatic cyclic group containing one to four heteroatoms each independently selected from O, S and N, wherein each group has from 3 to 10 atoms in its ring system. Non-aromatic heterocyclic groups include groups having only 3 atoms in their ring system, whereas aromatic heterocyclic groups have at least 5 atoms in their ring system. Heterocyclic groups include fused ring systems such as benzo-fused rings and the like. An exemplary 3 membered heterocyclic group is aziridine; 4 membered heterocyclic group is azetidinyl (derived from azetidine); 5 membered heterocyclic group is thiazolyl; 7 membered ring heterocyclic group is azepinyl; and a 10 membered heterocyclic group is quinolinyl.

Examples of non-aromatic heterocyclic groups include but are not limited to pyrrolidinyl, tetrahydrofuranyl, dihydrofuranyl, tetrahydrothienyl, tetrahydropyranyl, dihydropyranyl, tetrahydrothiopyranyl, pipehdino, morpholino, thiomorpholino, thioxanyl, piperazinyl, azetidinyl, oxetanyl, thietanyl, homopiperidinyl, oxepanyl, thiepanyl, oxazepinyl, diazepinyl, thiazepinyl, 1 ,2,3,6-tetrahydropyhdinyl, 2-pyrrolinyl, 3-pyrrolinyl, indolinyl, 2H-pyranyl, 4H-pyranyl, dioxanyl, 1 ,3-dioxolanyl, pyrazolinyl, dithianyl, dithiolanyl, dihydropyranyl, dihydrothienyl, dihydrofuranyl, pyrazolidinyl, imidazolinyl, imidazolidinyl, 3-azabicyclo[3.1.0]hexanyl, 3-azabicyclo[4.1.0]heptanyl, 3H-indolyl and quinolizinyl.

Examples of aromatic heterocyclic (heteroaryl) groups include but are not limited to pyridinyl, imidazolyl, pyrimidinyl, pyrazolyl, triazolyl, pyrazinyl, tetrazolyl, furyl, thienyl, isoxazolyl, thiazolyl, oxazolyl, isothiazolyl, pyrrolyl, quinolinyl, isoquinolinyl, indolyl, benzimidazolyl, benzofuranyl, cinnolinyl, indazolyl, indolizinyl, phthalazinyl, pyhdazinyl, triazinyl, isoindolyl, pteridinyl, purinyl, oxadiazolyl, thiadiazolyl, furazanyl, benzofurazanyl, benzothiophenyl, benzothiazolyl, benzoxazolyl, quinazolinyl, quinoxalinyl, naphthyhdinyl, and furopyridinyl. The foregoing groups may be C-attached or N-attached where such is possible.

For instance, a group derived from pyrrole may be pyrrol-1 -yl (N-attached) or pyrrol-3-yl (C-attached). Further, a group derived from imidazole may be imidazol-1 -yl (N-attached) or imidazol-3-yl (C-attached). Heterocyclic groups may be optionally substituted on any ring carbon, sulfur or nitrogen atom(s) by one to two oxygens (oxo), per ring. An example of a heterocyclic group wherein 2 ring carbon atoms are substituted with oxo moieties is 1 ,1-dioxo-thiomorpholinyl.

Exemplary five to six membered heterocyclic aromatic rings having one or two heteroatoms selected independently from oxygen, nitrogen and sulfur include but are not limited to isothiazolyl, pyridinyl, pyridiazinyl, pyrimidinyl, pyrazinyl and the like.

Exemplary partially saturated, fully saturated or fully unsaturated five to eight membered heterocyclic rings having one to four heteroatoms selected independently from oxygen, sulfur and nitrogen include but are not limited to 3H-1 ,2-oxathiolyl, 1 ,2,3- oxadizaolyl, 1 ,2,4-oxadiazolyl, 1 ,2,5-oxadiazolyl and the like. Further exemplary five membered rings are furyl, thienyl, 2H-pyrrolyl, 3H-pyrroyl, pyrrolyl, 2-pyrrolinyl, 3- pyrrolinyl, pyrrolidinyl, 1 ,3-dioxolanyl, oxazolyl, thiazolyl, thiazolyl, imidazolyl, 2H- imidazolyl, 2-imidazolinyl, imidazolidinyl, pyrazolyl, 2-pyrazolinyl, pyrazolinyl, isoxazolyl, isothiazolyl, 1 ,2-dithiolyl, 1 ,3-dithiolyl, 3H-1 ,2-oxathiolyl, 1 ,2,3-oxadizaolyl, 1 ,2,4- oxadiazolyl, 1 ,2,5-oxadiazolyl, 1 ,3,4-oxadiazolyl, 1 ,2,3-triazolyl, 1 ,2,4-trizaolyl, 1 ,3,4- thiadiazolyl, 1 ,2,3,4-oxatriazolyl, 1 ,2,3,5-oxathzaolyl, 3H-1 ,2,3-dioxazolyl, 1 ,2,4- dioxazolyl, 1 ,3,2-dioxazolyl, 1 ,3,4-dioxazolyl, 5H-1 ,2,5-oxathiazolyl and 1 ,3-oxathiolyl. Further exemplary six member rings are 2H-pyranyl, 4H-pyranyl, pyridinyl, piperidinyl, 1 ,2-dioxinyl, 1 ,3-dioxinyl, 1 ,4-dioxanyl, morpholinyl, 1 ,4-dithianyl, thiomorpholinyl, pyridazinyl, pyrimidinyl, pyrazinyl, piperazinyl, 1 ,3,5-thazinyl, 1 ,2,4-triazinyl, 1 ,2,3- trizainyl, 1 ,3,5-trithianyl, 4H-1 ,2-oxazinyl, 2H-1 ,3-oxazinyl, 6H-1 ,3-oxazinyl, 61-1-1 ,2- oxazinyl, 1 ,4-oxazinyl, 2H-1 ,2-oxazinyl, 4H-1 ,4-oxazinyl, 1 ,2,5-oxathiazinyl, 1 ,4-oxazinyl, o-isoxazinyl, p-isoxazinyl, 1 ,2,5-oxathiazinyl, 1 ,2,6-oxathiazinyl, 1 ,4,2-oxadiazinyl and 1 ,3,5,2-oxadiazinyl. Further exemplary seven membered rings are azepinyl, oxepinyl, thiepinyl and 1 ,2,4-diazepinyl. Further exemplary eight membered rings are cyclooctyl, cyclooctenyl and cyclooctadienyl.

Exemplary 3-10 membered heterocyclyl groups include but are not limited to oxetane, azetidine, tetrahydrofuran, pyrrolidine, 2,5-dihydro-1 H-pyrrole, 1 ,3-dioxalane, isoxazolidine, oxazolidine, pyrazolidine, imidazolidine, pyrrolidin-2-one, tetrahydrothiophene-1 ,1 -dioxide, pyrrolidine-2,5-dione, tetrahydro-2H-pyran, piperidine, 1 ,2,3,6-tetrahydropyridine, 1 ,4-dioxane, morpholine, piperazine, thiomorpholine, pipehdin-2-one, pipehdin-4-one, thiomorpholine-1 ,1 -dioxide, 1 ,3-oxazinan-2-one, morpholin-3-one, piperazine-2-one, azepane, 1 ,4-oxazepane, 1 ,4-diazepane, azepan-2- one, 1 ,4-diazepan-5-one, quinuclidine, 2-aza-bicyclo[2.2.1]heptane, 8-aza- bicyclo[3.2.1]octane, 5-oxa-2-aza-bicyclo[2.2.1]heptane, 2-oxa-5-aza- bicyclo[2.2.1]heptan-3-one, 2-oxa-5-aza-bicyclo[2.2.2]octan-3-one, 1 -methyl-5,6- pyrrolyl-7-oxa-bicyclo[2.2.1]heptane, 6-aza-bicyclo[3.2.1]octane, 3,8-diaza- bicyclo[3.2.1]octan-2-one, 2,2-dimethyl-tetrahydro-3aH-[1 ,3]dioxolo[4,5-c]pyrrole, 3,3- cyclohexylpyrrolidine, 1 ,5-diaxo-9-azaspiro[5.5]undecane, octahydro-1 H-isoindole, decahydroquinoline, decahydroisoquinoline, octahydropyrrolo[1 ,2a]pyrazine, octahydro'1 H-pyrido[1 ,2a]pyrazine, octahydropyrrolo[3,4-c]pyridine-3-one, decahydropyrazino[1 ,2-a]azepine, furan, 1 H-pyrrole, isoxazole, oxazole, 1 H-pyrazole, 1 H-imidazole, thiazole, 1 ,2,4-oxadiazole, 1 ,3,4-oxadiazole, 4H-1 ,2,4-triazole, 1 H- tetrazole, pyridine, pyridazine, pyrimidine, pyrazine, pyridine-2(1 H)-one, 1 ,4,5,6- tetrahydrocyclopenta[c]pyrazole, 6,7-dihydro-5H-pyrrolo[2,1 -c][1 ,2,4]triazole, 2,3- dihydroimidazo[2,1 -b]thiazole, imidazo[2,1 -b][1 ,3,4-c]pyridine, 4,5,6,7-tetrahydro-3H- imidazo[4,5-c] pyridine, 5,6,7,8-tetrahydroimidazo[1 ,5-a]pyrazine, 4,5,6,7- tetrahydrothiazole[5,4-c]pyhdine, 5,6,7,8-tetrahydro-[1 ,2,4]triazolo[4,3-a]pyrazine, quinoline, isoquinoline, 2,3-dihydrobenzofuran, 5,6,7,8-tetrahydroquinoline, 3,4-dihydro- 1 H-isochromene, 1 ,2,3,4-tetrahydroisoquinoline, 4H-benzo[d][1 ,3]dioxane, 5,6,7,8- tetrahydropyhdo[3,4-d]pyrimidine, benzofuran, 1 H-indole, benzo[d]oxazole, 1 H- benzo[d]imidazole, H-imidazo[1 ,2-a]pyridine, imidazo[1 ,2-a]pyhmidine, 5,6,7,8- tetrahydroimidazo[1 ,5-a]pyrazine-3(2H)-one, 2,3,4,5-tetrahydro-1 H-benzo[d]azepine, 2,3,4,5-tetrahydrobenzo[f][1 ,4]oxazepine, 5,6,7,8-tetrahydro-4H-isoxazolo[4,3-d]azepine and6,7,8,9-tetrahydro-2H-[1 ,2,4]triazolo[4,3-g][1 ,4]diazepin-3(5H)-one.

It is to be understood that if a carbocyclic or heterocyclic moiety may be bonded or otherwise attached to a designated substrate, through differing ring atoms without denoting a specific point of attachment, then all possible points are intended, whether through a carbon atom or, for example, a thvalent nitrogen atom. For example, the term "pyridyl" means 2-, 3-, or 4-pyridyl, the term "thienyl" means 2-, or 3-thienyl, and so forth.

Pharmaceutically acceptable salts of the compounds of the invention include the acid addition and base salts (including disalts) thereof. Suitable acid addition salts are formed from acids which form non-toxic salts. Examples include the acetate, aspartate, benzoate, besylate, bicarbonate/carbonate, bisulphate/sulphate, borate, camsylate, citrate, edisylate, esylate, formate, fumarate, gluceptate, gluconate, glucuronate, hexafluorophosphate, hibenzate, hydrochloride/chloride, hydrobromide/bromide, hydroiodide/iodide, isethionate, lactate, malate, maleate, malonate, mesylate, methylsulphate, naphthylate, 2-napsylate, nicotinate, nitrate, orotate, oxalate, palmitate, pamoate, phosphate/hydrogen phosphate/dihydrogen phosphate, saccharate, stearate, succinate, tartrate, tosylate and trifluoroacetate salts.

Suitable base salts are formed from bases which form non-toxic salts. Examples include the aluminium, arginine, benzathine, calcium, choline, diethylamine, diolamine, glycine, lysine, magnesium, meglumine, olamine, potassium, sodium, tromethamine and zinc salts. For a review on suitable salts, see "Handbook of Pharmaceutical Salts: Properties, Selection, and Use" by Stahl and Wermuth (Wiley-VCH, Weinheim, Germany, 2002).

A pharmaceutically acceptable salt of a compound of the invention may be readily prepared by mixing together solutions of a compound of the invention and the desired acid or base, as appropriate. The salt may precipitate from solution and be collected by filtration or may be recovered by evaporation of the solvent. The degree of ionization in the salt may vary from completely ionized to almost non-ionized.

The compounds of the invention which are complexes, such as clathrates and drug-host inclusion complexes, are within the scope of the invention. In contrast to the aforementioned solvates, the drug and host are present in stoichiometric or non- stoichiometric amounts. Also included are complexes containing two or more organic and/or inorganic components which may be in stoichiometric or non-stoichiometric amounts. The resulting complexes may be ionized, partially ionized, or non-ionized. For a review of such complexes, see J Pharm Sci, 64 (8), 1269-1288 by Haleblian (August 1975). The compounds of the invention include all polymorphs and isomers thereof, including optical, geometric and tautomeric isomers as hereinafter defined and isotopically-labeled compounds.

The compounds of the invention containing one or more asymmetric carbon atoms may exist as two or more stereoisomers. Where a compound contains an alkenyl or alkenylene group, geometric cis/trans (or Z/E) isomers are possible. Where the compound contains, for example, a keto or oxime group or an aromatic moiety, tautomeric isomerism ('tautomerism') can occur. It follows that a single compound may exhibit more than one type of isomerism. AII stereoisomers, geometric isomers and tautomeric forms of the compounds of the invention are included within the scope of the invention, including compounds exhibiting more than one type of isomerism, and mixtures of one or more thereof. Also included are acid addition or base salts wherein the counterion is optically active, for example, D-lactate or L-lysine, or racemic, for example, DL-tartrate or DL-arginine.

Cis/trans isomers may be separated by conventional techniques well known to those skilled in the art, for example, chromatography and fractional crystallization.

Conventional techniques for the preparation/isolation of individual enantiomers include chiral synthesis from a suitable optically pure precursor or resolution of the racemate (or the racemate of a salt or derivative) using, for example, chiral high pressure liquid chromatography (HPLC).

Alternatively, the racemate (or a racemic precursor) may be reacted with a suitable optically active compound, for example, an alcohol, or, in the case where the compound of the invention contains an acidic or basic moiety, an acid or base such as tartaric acid or 1 -phenylethylamine. The resulting diastereomehc mixture may be separated by chromatography and/or fractional crystallization and one or both of the diastereoisomers converted to the corresponding pure enantiomer(s) by means well known to a skilled person.

Chiral compounds of the invention (and chiral precursors thereof) may be obtained in enantiomehcally-enriched form using chromatography, typically HPLC, on an asymmetric resin with a mobile phase consisting of a hydrocarbon, typically heptane or hexane, containing from 0 to 50% isopropanol, typically from 2 to 20%, and from 0 to 5% of an alkylamine, typically 0.1 % diethylamine. Concentration of the eluate affords the enriched mixture. Mixtures of stereoisomers may be separated by conventional techniques known to those skilled in the art [see, for example, "Stereochemistry of Organic Compounds" by E. L. ENeI (Wiley, New York, 1994)].

The invention includes all pharmaceutically acceptable isotopically-labeled compounds of the invention, wherein one or more atoms are replaced by atoms having the same atomic number, but an atomic mass or mass number different from the atomic mass or mass number usually found in nature.

Examples of isotopes suitable for inclusion in the compounds of the invention include isotopes of hydrogen, such as ²H and ³H, carbon, such as ¹¹C, ¹³C and ¹⁴C, chlorine, such as ³⁶CI, fluorine, such as ¹⁸F, iodine, such as ¹²³I and ¹²⁵I, nitrogen, such as ¹³N and ¹⁵N, oxygen, such as ¹⁵O, ¹⁷O and ¹⁸O, phosphorus, such as ³²P, and sulphur, such as ³⁵S.

Certain isotopically-labelled compounds of the invention, for example those incorporating a radioactive isotope, are useful in drug and/or substrate tissue distribution studies. The radioactive isotopes tritium, i.e., ³H, and carbon-14, i.e., ¹⁴C, are particularly useful for this purpose in view of their ease of incorporation and ready means of detection.

Substitution with heavier isotopes such as deuterium, i.e., ²H, may afford certain therapeutic advantages resulting from greater metabolic stability, for example, increased in vivo half-life or reduced dosage requirements, and hence may be preferred in some circumstances.

Substitution with positron emitting isotopes, such as ¹¹C, ¹⁸F, ¹⁵O and ¹³N, can be useful in Positron Emission Topography (PET) studies for examining substrate receptor occupancy.

Isotopically-labeled compounds of the invention can generally be prepared by conventional techniques known to those skilled in the art or by processes analogous to those described in the accompanying Examples and Preparations using an appropriate isotopically-labeled reagents in place of the non-labeled reagent previously employed. The parenthetical negative or positive sign used herein in the nomenclature denotes the direction plane polarized light is rotated by the particular stereoisomer.

One of ordinary skill will recognize that certain compounds of the invention may contain one or more atoms which may be in a particular stereochemical or geometric configuration, giving rise to stereoisomers and configurational isomers. All such isomers and mixtures thereof are included in the invention. Solvates (hydrates) of the compounds of the invention are also included.

Other features and advantages will be apparent from the specification and claims which describe the invention.

Detailed Description of the Invention The examples and preparations provided below further illustrate and exemplify the compounds of the present invention and methods of preparing such compounds. It is to be understood that the scope of the present invention is not limited in any way by the scope of the following examples and preparations. In the following examples, molecules with a single chiral center, unless otherwise noted, exist as a racemic mixture. Those molecules with two or more chiral centers, unless otherwise noted, exist as a racemic mixture of diastereomers. Single enantiomers/diastereomers may be obtained by methods known to those skilled in the art. In general, the compounds of the invention may be prepared by processes known in the chemical arts, particularly in light of the description contained herein. Certain processes for the manufacture of the compounds of the invention are provided as further features of the invention and are illustrated in the reaction schemes provided below and in the experimental section. The use of various protecting groups in these reactions are also well known and are exemplified in Protective Groups In Organic Synthesis, Second Edition, T.W. Greene and P. G. M. Wuts, John Wiley and Sons, Inc. 1991 , pages 227-229, which is hereby incorporated by reference in its entirety for all purposes.

The utility of the compounds of the invention as medical agents for the reduction of intraocular pressure and accordingly to treat glaucoma is demonstrated by the activity of the compounds in conventional assays, including the in vivo assay and a receptor binding assay. Such assays also provide a means whereby the activities of the compounds can be compared to each other and with the activities of other known compounds. The results of these comparisons are useful for determining dosage levels in mammals, including humans, for the treatment of such diseases.

The compounds of the invention intended for pharmaceutical use may be administered as crystalline or amorphous products. They may be obtained, for example, as solid plugs, powders, or films by methods such as precipitation, crystallization, freeze drying, spray drying, or evaporative drying. Microwave or radio frequency drying may be used for this purpose.

The compounds of the invention intended for pharmaceutical use may be administered alone or in combination with one or more other compounds of the invention or in combination with one or more other drugs (or as any combination thereof). Generally, they will be administered as a formulation in association with one or more pharmaceutically acceptable excipients. The term "excipient" is used herein to describe any ingredient other than the compound(s) of the invention. The choice of excipient will to a large extent depend on factors such as the particular mode of ad ministration, the effect of the excipient on solubility and stability, and the nature of the dosage form.

Pharmaceutical compositions suitable for the delivery of compounds of the present invention and methods for their preparation will be readily apparent to those skilled in the art. Such compositions and methods for their preparation may be found, for example, in 'Remington's Pharmaceutical Sciences', 19th Edition (Mack Publishing Company, 1995).]

The compounds of the invention may be administered directly to the eye, typically in the form of drops of a micronised suspension or solution in isotonic, pH-adjusted, sterile saline. Other formulations suitable for ocular administration include ointments, biodegradable (e.g. absorbable gel sponges, collagen) and non-biodegradable (e.g. silicone) implants, wafers, lenses and particulate or vesicular systems, such as niosomes or liposomes. A polymer such as crossed-l inked polyacrylic acid, polyvinylalcohol, hyaluronic acid; a cellulosic polymer, for example, hydroxypropylmethylcellulose, hydroxyethylcellulose, or methyl cellulose; or a heteropolysacchahde polymer, for example, gelan gum, may be incorporated together with a preservative, such as benzalkonium chloride.

The compounds of the invention can be incorporated into various types of ophthalmic formulations for delivery to the eye. These compounds may be combined with ophthalmologically acceptable preservatives, surfactants, viscosity enhancers, penetration enhancers, buffers, sodium chloride and water to form aqueous, sterile ophthalmic suspensions or solutions. In order to prepare sterile ophthalmic ointment formulations, the active ingredient is combined with a preservative in an appropriate vehicle, such as, mineral oil, liquid lanolin, or white petrolatum. Sterile ophthalmic gel formulations may be prepared by suspending the active ingredient in a hydrophilic base prepared from the combination of, for example, carbopol-940 or the like according to the published formulations for analogous ophthalmic preparations; preservatives and tonicity agents can be incorporated. Ophthalmic solution formulations may be prepared by dissolving the active ingredient in a physiologically acceptable isotonic aqueous buffer. Further, the ophthalmic solution may include an ophthalmologically acceptable surfactant to assist in dissolving the active ingredient. Furthermore, the ophthalmic solution may contain a thickener such as hydroxymethylcellulose, hydroxyethylcellulose, hydroxypropylmethylcellulose, methyl-cellulose, polyvinylpyrrolidone, or the like to improve the retention of the medicament in the conjunctival sac.

The compounds of the invention are preferably formulated as topical ophthalmic suspensions or solutions, with a pH of about 4.5 to 7.8. The compounds will normally be contained in these formulations in an amount of 0.1 % to 10% by weight, but preferably in an amount of 0.25% to 5.0% by weight. Thus, for topical presentation, 1 to 3 drops of these formulations would be delivered to the surface of the eye 1 to 4 times a day according to the routine discretion of a skilled clinician.

The compounds of the invention may be combined with soluble macromolecular entities, such as cyclodextrin and suitable derivatives thereof or polyethylene glycol- containing polymers, in order to improve their solubility, dissolution rate, taste-masking, bioavailability and/or stability for use in any of the aforementioned modes of administration.

Drug-cyclodexthn complexes, for example, are found to be generally useful for most dosage forms and administration routes. Both inclusion and non-inclusion complexes may be used. As an alternative to direct complexation with the drug, the cyclodextrin may be used as an auxiliary additive, i.e. as a carrier, diluent, or solubiliser. Most commonly used for these purposes are alpha-, beta- and gamma-cyclodexthns, examples of which may be found in International Patent Applications Nos. WO91 /11172, WO94/02518 and WO98/55148.

Depending on the disease and condition of the patient, the term "treatment" as used herein may include one or more of curative, palliative and prophylactic treatment.

As mentioned above, objects of the present invention are also pharmaceutical compositions containing at least a compound of the present invention of formula (I) together with non-toxic adjuvants and/or carriers typically employed in the pharmaceutical field.

The preferred route of administration is topical. The compounds of the present invention can be administered as solutions, suspensions or emulsions (dispersions) in an ophthalmically acceptable vehicle. The term "ophthalmic acceptable vehicle" as used herein refers to any substance or combination of substances which are non-reactive with the compounds and suitable for administration to patient.

Preferred are aqueous vehicles suitable for topical application to the patient's eyes. Other ingredients which may be desirable to use in the ophthalmic compositions of the present invention include antimicrobials, preservatives, co-solvents, surfactants and viscosity building agents.

The invention also relates to a method for treating glaucoma or ocular hypertension, said method consisting in contacting an effective intraocular pressure reducing amount of a composition with the eye in order to reduce eye pressure and to maintain said pressure on a reduced level.

The doses of prostaglandin nitroderivatives can be determined by standard clinical techniques and are in the same range or less than those described for the corresponding underivatized, commercially available prostaglandin compounds as reported in the: Physician's Desk Reference, Medical Economics Company, Inc., Oradell, N.J., 58^th Ed., 2004; The pharmacological basis of therapeutics, Goodman and Gilman, J. G. Hardman, L. e. Limbird, Tenth Ed.

The compositions contain 0.1 -0.30 μg, especially 1 -10 μg, per application of the active compound.

The treatment may be advantageously carried out whereby one drop of the composition, corresponding to about 30 μl, is administered about 1 to 2 times per day to the patient's eye.

It is further contemplated that the compounds of the present invention can be used with other medicaments known to be useful in the treatment of glaucoma or ocular hypertension, either separately or in combination. For example, the compounds of the present invention can be combined with (i) beta-blockers, such as timolol, betaxolol, levobunolol and the like (see U.S. Patent No. 4,952,581 ); (ii) carbonic anhydrase inhibitors, such as brinzolamide; (iii) adrenergic agonists including clonidine derivatives, such as apraclonidine or brimonidine (see U.S. Patent No. 5,811 ,443). Also contemplated is the combination with nitrooxy derivatives of the above reported compounds, for example nitrooxy derivatives of beta-blockers such as those described in U.S. Patent No. 6,242,432.

The following non-limiting preparations and Examples illustrate the preparation of the compounds of the invention.

Examples

In the examples described below, unless otherwise indicated, all temperatures are set forth in degrees Celsius and all parts and percentages are by weight. Reagents may be purchased from commercial suppliers, such as Sigma-Aldrich Chemical Company, Acros Organics, or Lancaster Synthesis Ltd. and may be used without further purification unless otherwise indicated. Tetrahydrofuran (THF), methylene chloride (CH₂CI₂ Or DCM), N, N-dimethylacetamide (DMA), acetonitrile (MeCN or ACN), and N₁N- dimethylformamide (DMF) may be purchased from Aldrich in Sure-Seal bottles and used as received. All solvents may be purified using standard methods known to those skilled in the art, unless otherwise indicated. Diethyl ether is abbreviated as Et₂O. Ethyl acetate is abbreviated as EtOAc or EA. Thfluoroacetic acid is abbreviated as TFA. Acetic acid is abbreviated as HOAc or AcOH. Trifluoromethanesulfonate, or triflate, is abbreviated as "OTf." te/f-Butoxycarbonyl is abbreviated as BOC. 4-(N₁N-

Dimethylamino)pyridine is abbreviated as DMAP. N-Methyl-morpholine is abbreviated as NMM. acetic anhydride as Ac₂O. N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride is abbreviated as EDAC or EDC.

The reactions set forth below were done generally under a positive pressure of argon or nitrogen or with a drying tube, at ambient temperature (unless otherwise stated), in anhydrous solvents, and the reaction flasks were fitted with rubber septa for the introduction of substrates and reagents via syringe. Glassware was oven dried and/or heat dried. Microwave chemistry was carried out using an EmrysTM Optimizer EXP from Personal Chemistry, Inc. (now Biotage). Analytical thin layer chromatography (TLC) was performed using glass-backed silica gel 60 F 254 pre-coated plates (Merck Art 5719) and eluted with appropriate solvent ratios (v/v). Reactions were assayed by TLC or LCMS and terminated as judged by the consumption of starting material. Visualization of the TLC plates was done with UV light (254 nm wavelength) or with an appropriate TLC visualizing solvent and activated with heat. Analytical HPLC performed with Waters or Agilent instruments. Flash column chromatography (Still et al., J. Org. Chem., 1978, 43, 2923) was performed using silica gel 60 (Merck Art 9385) or various MPLC systems, such as Biotage or ISCO purification systems (i.e., Biotage SP4 model). Preparative HPLC was done with several methods as follows: Prep LC 4000 system from Water with Ultra 120 10 mm C8 column from Peeke Scientific. Mass-directed prep HPLC with an Agilent A2Prep System, with computer-controlled gradients of two mobile phases (100% water with 0.1 % formic acid and 100% acetonitrile with 0.1 % formic acid) through a XBridge C18 column, 250 mm x 30 mm., 5 μmicron particle size, and fraction collection guided by a detector tandem of UV diode array and (ESI) mass spectrometer. Supercritical Fluid Chromatography (SFC) purification was performed on Multigram Il SFC from Berger Instruments using the ProNTo software platform. Chiralpak AS-H 21.2 x 250 mm 5u column was typically used and eluted with 20% MeOH in CO₂ at 140 bar. Flow rate was 60 mL/min. Peaks were collected with UV detection at 260 nm.

The compound structures in the examples below were confirmed by one or more of the following methods: proton magnetic resonance spectroscopy, mass spectroscopy, and elemental microanalysis. Proton magnetic resonance (¹H NMR) spectra were determined using a Bruker spectrometer operating at field strength of 300, 400, or 700 megahertz (MHz). Chemical shifts are reported in parts per million (ppm, δ) downfield from an internal tetramethylsilane standard. Alternatively, ¹H NMR spectra were referenced relative to signals from residual protons in deuterated solvents as follows: CDCI₃ = 7.25 ppm; DMSO-d₆ = 2.49 ppm; CD₃CN = 1.94 ppm, CD₃OD or methanol-c/₄ = 3.30 ppm; C₆D₆ = 7.16 ppm. Peak multiplicities are designated as follows: s, singlet; d, doublet; dd, doublet of doublets; t, triplet; dt, doublet of triplets; q, quartet; br, broadened; m, multiplet. Coupling constants are given in Hertz (Hz). For detailed structural elucidation of selected examples-proton, gCOSY, gHSQC, and gHMBC NMR spectra were acquired using a Bruker Avance 700 MHz NMR spectrometer equipped with a cryo-probe with the proton and carbon coils tuned to 700.13 MHz and 176.07 MHz respectively. A solution of about 10 mg of sample dissolved in 0.75 ml_ dimethylsulfoxide (d-6; 99.8% D) was used to acquire the spectra, which were referenced to the solvent signal (2.50 ppm for proton and 39.51 ppm for carbon).

Mass spectra (MS) data were obtained using Agilent LC mass spectrometer with APCI or (ESI) ionization. High resolution MS (HRMS) were performed on an Agilent G3250AA LCMSD/TOF mass spectrometer. Elemental microanalyses were performed by Atlantic Microlab Inc. and gave results for the elements stated within ±0.4% of the theoretical values.

Preferred compounds in accordance with the invention may be prepared in manners analogous to those specifically described below. The examples and preparations provided below further illustrate and exemplify the compounds of the present invention and methods of preparing such compounds. It is to be understood that the scope of the present invention is not limited in any way by the scope of the following examples and preparations. The skilled artisan will recognize that different acids, amines, alkyl halides, aryl halides, coupling reagents, and heterocycles may be substituted in the following descriptions to suit the preparations of a desired embodiment. The following methods may be scaled upwards or downwards to suit the amount of desired material.

Method A

Example A-1

4-({3-oxo-4-r2-(thfluoromethyl)benzyl1-3,4-dihvdro-2/-/-1 ,4-benzothiazin-2- vDmethvDbenzoic acid

Preparation of A-1 -a 1 -methylethyl 4-aminobenzoate

To a solution of 4-aminobenzoic acid (5.0 g, 36.5 mmol) in 2-propanol (200 ml_) was added thionyl chloride (21 ml, 290 mmol). The mixture was first stirred at room temperature for 3 days and subsequently refluxed for 18 hrs. Carefully, an aqueous sodium bicarbonate solution (150 ml_) was added. It was extracted with ethyl acetate (3 x 80 ml_), dried over sodium sulfate, filtered, and concentrated to obtain a yellow solid in quantitative yield. ¹H-NMR (300 MHz, CDCI₃) δ ppm 7.8 (d, 2, Ar-H), 6.6 (d, 2H, Ar-H), 5.2 (q, 1 H, -CH(CHs)₂), 1.4 (d, 6H, CH(CH₃)₂).

Preparation of A-1 -b 1 -methylethyl 4-(2-bromo-3-ethoxy-3-oxopropvl)benzoate

To a solution of 1 -methylethyl 4-aminobenzoate (4.4 g, 24.6 mmol) in H₂O (30 ml_) was added concentrated aqueous hydrogen bromide solution (7.2 ml 47%, 61.5 mmol). The mixture was cooled to O⁰C and slowly sodium nitrite (1.7 g, 24.6 mmol) in H₂O (30 ml_) was added. The resulting cold mixture was slowly added to a solution of CuBr (0.7 g, 4.9 mmol) and ethyl acrylate (80 ml_, 0.74 mol) in acetone (150 ml_) at 4O⁰C. After 30 min., H₂O (1 L) was added and it was extracted with ethyl acetate (3 * 250 ml_), dried over sodium sulfate, filtered, and concentrated (8.5 g). Based on ¹H-NMR, 60% product was present and it was used in the next step without further purification. ¹H-NMR (300 MHz, CDCI₃) δ ppm 8.0 (d, 2H), 7.3 (d, 2H), 5.2 (q, 1 H), 4.4 (t, 1 H), 4.2 (q, 2H), 3.5 (m, 1 H), 3.3 (m, 1 H), 1.4 (d, 6H), 1.3 (t, 3H).

Preparation of A-1 -c 1 -methylethyl 4-[(3-oxo-3,4-dihydro-2/-/-1 ,4-benzothiazin-2-yl)methyl1benzoate

To a solution of aminothiophenol (0.92 g, 7.4 mmol) in dimethylformamide (30 ml_) was added 1 -methylethyl 4-(2-bromo-3-ethoxy-3-oxopropyl)benzoate (7.4 mmol) in dimethylformamide (30 ml_) and it was heated at 9O⁰C. After 2 hrs the mixture was cooled to room temperature and H₂O (500 ml_) was added. The mixture was extracted with ethyl acetate (5 x 80 ml_), washed with brine, dried over sodium sulfate, filtered, and concentrated to give a crude yield (5.9 g). After purification by automated column chromatography, 1 -methylethyl 4-[(3-oxo-3,4-dihydro-2H-1 ,4-benzothiazin-2- yl)methyl]benzoate was obtained as a yellow solid (1.5 g, 36%). ¹H-NMR (300 MHz, CDCI₃) δ ppm 8.6 (s, 1 H), 8.0 (d, 2H), 7.3 (d, 2H), 7.3 (m, 1 H), 7.2 (m, 1 H), 7.0 (m, 1 H), 6.9 (d, 1 H), 5.2 (q, 1 H), 3.7 (dd, 1 H), 3.4 (dd, 1 H), 2.9 (dd, 1 H), 1.4 (d, 6H).

Preparation of A-1 -d

1 -methylethyl 4-({3-oxo-4-[2-(trifluoromethyl)benzvπ-3,4-dihvdro-2/-/-1 ,4-benzothiazin-2-

To a suspension of sodium hydride (30 mg, 0.74 mmol) in dimethylformamide (2 ml_) was added 1 -methylethyl 4-[(3-oxo-3,4-dihydro-2H-1 ,4-benzothiazin-2- yl)methyl]benzoate (230 mg, 0.67 mmol) in dimethylformamide (5 ml_). After 15 min. stirring, α'-bromo-α,α,α-thfluoro-o-xylene (161 mg, 0.67 mmol). After 1.5 h, an aqueous NH₄CI solution (30 ml_) was added. It was extracted with ethyl acetate (3 * 25 ml_), washed with brine (20 ml_), dried over sodium sulfate, filtered, and concentrated. After purification by column chromatography, using 10% ethyl acetate / heptane, 1 - methylethyl 4-({3-oxo-4-[2-(trifluoromethyl)benzyl]-3,4-dihydro-2H-1 ,4-benzothiazin-2- yl}methyl)benzoate was obtained as a yellow oil (170 mg, 51 %).¹H-NMR (300 MHz,

CDCI₃) δ ppm 8.0 (d, 2H), 7.7 (d, 1 H), 7.0-7.4 (m), 6.9 (dd, 1 H), 5.4 (dd, 2H), 5.2 (q, 1 H), 3.8 (dd, 1 H), 3.4 (dd, 1 H), 2.9 (dd, 1 H), 1.4 (d, 6H) LCMS (M+H)⁺: 500.

Preparation of A-1 4-({3-oxo-4-r2-(thfluoromethyl)benzyl1-3,4-dihvdro-2/-/-1 ,4-benzothiazin-2- vDmethvDbenzoic acid

To solution of 1 -methylethyl 4-({3-oxo-4-[2-(trifluoromethyl)benzyl]-3,4-dihydro-2/-/-1 ,4- benzothiazin-2-yl}methyl)benzoate (188 mg, 0.38 mmol) in dioxane (40 ml_) was added an aqueous sodium hydroxide solution (5 ml, 1 M). After refluxing for 18 h the dioxane was evaporated. An aqueous citric acid solution was added until acidic. It was extracted with ethyl acetate (3 * 30 ml_), dried over sodium sulfate, filtered, and concentrated to yield4-({3-oxo-4-[2-(thfluoromethyl)benzyl]-3,4-dihydro-2/-/-1 ,4-benzothiazin-2- yl}methyl)benzoic acid as a brown solid (50 mg, 29%). ¹H-NMR (300 MHz, CDCI₃) δ ppm 8.1 (d, 2H), 7.7 (d, 1 H), 7.0-7.4 (m), 6.9 (dd, 1 H), 5.4 (dd, 2H), 3.9 (dd, 1 H), 3.5 (dd, 1 H), 3.0 (dd, 1 H). LCMS (M+H)⁺: 458.

The following compounds A-2 to A-48 listed in Table 1 were prepared in a similar manner to method A using the appropriate benzyl bromide in step d. In some cases enantiomers were separated by chiral supercritical fluid chromatography (SFC) performed on Multigram Il SFC from Berger Instruments. The Chiralpak AS-H 21.2 x 250 mm 5u column was typically used and eluted with 20% MeOH in CO₂ at 140 bar. Flow rate was 60 mL/min. Peaks were collected with UV detection at 260 nm.

Method B

Example B-1

4-{r8-chloro-4-(4-cvanobenzyl)-3-oxo-3,4-dihvdro-2/-/-1 ,4-benzothiazin-2- vlimethvDbenzoic acid

Preparation of B-1 -a 2-Ch loro-6-n itrobenzeneth iol

To a solution of 2,3-dichloronitrobenzene (19.2 g, 100 mmol) in DMSO (300 ml_) was added powdered sodium sulfide (24.0 g, 100 mmol). The mixture was stirred at ambient temperature for 18 hours. The reaction mixture was then diluted with 2 L of water. The mixture was clarified by filtration, and the filtrate was acidified to pH=4 with concentrated hydrogen chloride. The mixture was extracted with 3x Et₂O (400 ml_). The organic layers were combined and washed with brine, then dried over magnesium sulfate, and filtered. Removal of the solvent under reduced pressure gave product (18g, 95%) as a yellow solid, which was used without further purification. ¹H NMR (400 MHz, DMSO-c/₆) δ ppm 8.02 (d, J=8.08 Hz, 1 H) 7.94 (dd, J=8.08, 1.26 Hz, 1 H) 7.52 (t, J=7.96 Hz, 1 H) 5.06 (s, 1 H).

Preparation of B-1 -b 1 -Methylethyl 4-(2-Bromo-3-ethoxy-3-oxopropvl)benzoate

Reference: Similar to reaction from Obushak, M. D.; Matiychuk, V. S.; Tsyalkovsky, V. M.; Voloshchuk, R. M. Synthesis of Heterocycles on the Basis of Arylation Products of Unsaturated Compounds. Part 11. 5-R-Benzyl-2- iminoselenazolidin-4-ones from Ethyl 3-Aryl-2-bromopropanoates. Phosphorus, Sulfur, and Silicon and the Related Elements 2004, 179, 107 - 113 (Beilstein ref 5796305 or SciFinder hit: CAN 140:287328 AN 2003:969209. To a solution of 4-amino-benzoic acid isopropyl ester (4.48 g, 25 mmol) in water (30 ml_) was added 67% aq hydrogen bromide solution (4.3 ml_, 62.5 mmol). The mixture was cooled to O C. Then sodium nitrite (1.72 g, 25 mmol) in water (30 ml_) was added. The resulting cold mixture was slowly added to a solution of CuBr (0.71 g, 5 mmol) and ethyl acrylate (81 ml_, 750 mmol) in acetone (150 ml_) at 40^°C. After 30 minutes, H₂O (1 L) was added. The mixture was extracted with ethyl acetate (3x 250 ml_), dried over magnesium sulfate, filtered, and concentrated. Yielded (8.2 g crude, 96%) of brown oil, which was used without further purification. ¹H NMR (400 MHz, CDCI₃) δ ppm 7.99 (d, J=8.34 Hz, 2 H) 7.29 (d, J=8.34 Hz, 2 H) 5.24 (d, J=6.06 Hz, 1 H) 4.40 (t, J=7.83 Hz, 1 H) 4.19 (t, J=7.07 Hz, 2 H) 3.52 (dd, J=14.15, 8.08 Hz, 1 H) 3.30 (dd, J=14.15, 7.33 Hz, 1 H) 1.37 (d, J=6.32 Hz, 6 H) 1.25 (t, J=7.07 Hz, 3H). HRMS Calcd for C₁₅H₁₉BrO₄ (M+Na): 365.0359 Found: 365.0356 Preparation of B-1 -c 1 -Methylethyl 4-{2-[(2-chloro-6-nitrophenyl)sulfanyl]-3-ethoxy-3-oxopropyl}benzoate

2-Chloro-6-nitrobenzenethiol (2.28g, 12 mmol) was dissolved in tetrahydrofuran (40 ml_), and to this solution was added 1 M potassium t-butoxide in t-butanol (24 ml_, 24 mmol). The mixture was stirred under nitrogen for 30 minutes. Bromoacetate (6.86 g, 12 mmol) was added via syringe. The solution stirred at ambient temperature for 6 hours. The mixture was quenched with water and then extracted with ethyl acetate 3x. The combined organic layer was then treated to serial washes with H₂O, 1 N sodium hydroxide, and brine, then dried with magnesium sulfate and concentrated in vacuo. The crude product was purified using column chromatography, eluting with ethyl acetate /heptane 10-45% to give the desired product as light yellow oil (3.7g, 68%). ¹H NMR (400 MHz, CDCI₃) δ ppm 7.96 (d, J=8.34 Hz, 2 H) 7.67 (dd, J=8.08, 1.52 Hz, 1 H) 7.55 (dd, J=7.96, 1.39 Hz, 1 H) 7.44 (t, J=7.96 Hz, 1 H) 7.31 (d, J=8.34 Hz, 2 H) 5.23 (d, J=6.32 Hz, 1 H) 4.09 (dd, J=9.60, 6.06 Hz, 1 H) 3.99 (q, J=7.07 Hz, 2 H) 3.31 (dd,

J=14.02, 9.47 Hz, 1 H) 3.16 (d, J=6.32 Hz, 1 H) 1.36 (d, J=6.32 Hz, 6 H) 1.03 (t, J=7.20 Hz, 3H). HRMS Calcd for C₂IH₂₂CINO₆S (M+Na): 474.0748 Found: 474.0763.

Preparation of B-1 -d 1 -Methylethyl 4-r(8-Chloro-3-oxo-3,4-dihvdro-2/-/-1 ,4-benzothiazin-2-yl)methyl1benzoate

1 -Methylethyl 4-{2-[(2-amino-6-chlorophenyl)sulfanyl]-3-ethoxy-3-oxopropyl}benzoate (2.8 g, 6.3 mmol) was refluxed in 10% hydrogen chloride for 45 min, poured into ice water, and extracted with ethyl acetate (3 x 200 ml_). The combined organic layers were washed with saturated aq sodium bicarbonate, then dried over magnesium sulfate, and the solvent was evaporated under reduced pressure. The product was further purified by column chromatography, eluting with 15-40% ethyl acetate/hexane. Yielded (800 mg, 32%) of brown solid. ¹H NMR (400 MHz, DMSO-Cl₆) δ ppm 10.88 (s, 1 H) 7.85 (d, J=8.34 Hz, 2 H) 7.35 (d, J=8.08 Hz, 2 H) 7.18 (dt, J=15.73, 7.93 Hz, 2 H) 6.96 (dd, J=7.71 , 1.39 Hz, 1 H) 5.11 (quin, J=6.19 Hz, 1 H) 4.02 (dd, J=9.09, 6.06 Hz, 1 H) 3.24 (dd, J=14.02, 5.94 Hz, 1 H) 2.84 (dd, J=14.27, 9.22 Hz, 1 H) 1.31 (d, J=6.32 Hz, 6 H). HRMS Calcd for C₁₉H₁₈CINO₃S (M+Na): 398.0588 Found: 398.0599

Preparation of B-1 -e

1 -methylethyl 4-{[8-chloro-4-(4-cvanobenzyl)-3-oxo-3,4-dihvdro-2/-/-1 ,4-benzothiazin-2- ylimethvDbenzoate

The title compound was made analogous to the method described in Method A using instead 1 -methylethyl 4-[(8-Chloro-3-oxo-3,4-dihydro-2H-1 ,4-benzothiazin-2- yl)methyl]benzoate. ¹H NMR (400 MHz, CDCI₃) δ ppm 8.00 (d, J=8.34 Hz, 2 H) 7.62 (d, J=8.34 Hz, 2 H) 7.28 - 7.31 (m, 2 H) 7.24 - 7.26 (m, 2 H) 7.07 - 7.22 (m, 2 H) 6.87 (d, J= 1.01 Hz, 1 H) 5.23 - 5.35 (m, 2 H) 3.79 (dd, J=9.35, 5.56 Hz, 1 H) 3.38 (dd, J= 13.89, 5.56 Hz, 1 H) 2.92 (dd, J= 13.89, 9.35 Hz, 1 H) 1.38 (d, J=6.06 Hz, 6 H) 0.89 (d, J= 13.64 Hz, 1 H). HRMS Calcd for C₂₇H₂₃CIN₂O₃S (M+H): 491.1190 Found: 491.1187

Preparation of B-1

4-{[8-chloro-4-(4-cvanobenzyl)-3-oxo-3,4-dihvdro-2/-/-1 ,4-benzothiazin-2- ylimethvDbenzoic acid

The title compound was made analogous to the method described in Method A using instead 1 -methylethyl 4-{[8-chloro-4-(4-cyanobenzyl)-3-oxo-3,4-dihydro-2H-1 ,4- benzothiazin-2-yl]methyl}benzoate. ¹H NMR (400 MHz, DMSO-c/₆) δ ppm 7.88 (d, J=8.08 Hz, 2 H) 7.80 (d, J=8.34 Hz, 2 H) 7.37 (t, J=9.09 Hz, 3 H) 7.28 - 7.38 (m, 1 H) 7.25 (dt, J=3.85, 1.99 Hz, 2 H) 7.16 - 7.20 (m, 1 H) 5.34 (br. s., 2 H) 4.27 (td, J=8.78, 6.19 Hz, 1 H) 2.90 (s, 1 H) 1.23 - 1.27 (m, 1 H). HRMS Calcd for C₂₄H₁₇CIN₂O₃S (M+H): 471.0540 Found: 471.0554. The racemic mixture was separated by chiral SFC and the 2 enantiomers were isolated as (-)4-{[8-chloro-4-(4-cyanobenzyl)-3-oxo-3,4-dihydro-2H- 1 ,4-benzothiazin-2-yl]methyl}benzoic acid (B-2, [α]_D° =-37.30; 99% ee) and (+)4-{[8- chloro-4-(4-cyanobenzyl)-3-oxo-3,4-dihydro-2/-/-1 ,4-benzothiazin-2-yl]methyl}benzoic acid (B-3, [α]_D° =30.00; 99% ee).

Method C

Example C-1

4-({2-methyl-3-oxo-4-r2-(trifluoromethyl)benzyl1-3,4-dihvdro-2H-1 ,4-benzothiazin-2- vDmethvDbenzoic acid

Preparation of C-1 -a

2-methyl-2H-1.4-benzothiazin-3(4H)-one

A mixture of 2-aminobenzenethiol (2.5g, 20mnnol) and 2-bromopropanoic acid (3.Og, 20mnnol) was heated under nitrogen at 125°C for 16 h. Reaction cooled to ambient temperature and dissolved in dichloromethane washed with saturated sodium bicarbonate solution, dried over anhydrous magnesium sulfate, filtered and concentrated in vacuo. Material purified using preparatory LC/MS to give the titled compound as an off white solid. (1.04g, 29% yield) ¹H NMR (400 MHz, CDCI₃) δ ppm 8.51 (br. s., 1 H) 7.32 (dd, J=7.83, 1.26 Hz, 1 H) 7.16 - 7.22 (m, 1 H) 7.00 - 7.06 (m, 1 H) 6.87 (dd, J=7.83, 1.26 Hz, 1 H) 3.57 (q, J=7.16 Hz, 1 H) 1.51 (d, J=7.07 Hz, 3 H)

Preparation of C-1 -b

2-methyl-4-f2-(trifluoromethyl)benzvn-2H-1.4-benzothiazin-3(4H)-one

2-methyl-2H-1 ,4-benzothiazin-3(4H)-one (0.25g, 1.4mmol) was dissolved in dimethylformamide (3.49ml_, 0.4M). 60% dispersion of sodium hydride in mineral oil (0.056g, 1.4mmol) added and reaction stirred for 10 min. Next, 1 -(bromomethyl)-2-

(trifluoromethyl)benzene (0.33g, 1.4mmol) was added. The reaction mixture was stirred at ambient temperature for 72 h and then charged with 60% sodium hydride dispersion in mineral oil (0.022g, 0.56mmol) followed by 1 -(bromomethyl)-2- (thfluoromethyl)benzene (0.13g, 0.56mmol) and allowed to stir at ambient temperature for five hours. The reaction mixture was worked up by quenching with water, extracted with ethyl acetate, dried over magnesium sulfate, filtered and concentrated. The crude material was purified column chromatography, eluting with 10% ethyl acetate /hexanes to give the titled compound as a thick glass. (0.371 g, 79% yield). ¹H NMR (400 MHz, CDCI₃) δ ppm 7.71 (d, J=7.83 Hz, 1 H) 7.32 - 7.48 (m, 3 H) 7.07 - 7.16 (m, 2 H) 6.98 - 7.05 (m, 1 H) 6.85 (dd, J=8.34, 1.01 Hz, 1 H) 5.36 - 5.52 (m, 2 H) 3.67 (q, J=7.07 Hz, 1 H) 1.56 - 1.59 (m, 3 H).

Preparation of C-1-c

Methyl 4-({2-methyl-3-oxo-4-[2-(trifluoromethyl)benzyl1-3,4-dihvdro-2H-1 ,4-benzothiazin- 2-yl)methyl)benzoate

Anhydrous 1 M LiHMDS (0.35ml_, 0.35mmol) was added to 2-methyl-4-[2- (trifluoromethyl)benzyl]-2H-1 ,4-benzothiazin-3(4H)-one (0.10Og, 0.30mmol)) in tetrahydrofuran (2mL) at -78⁰C under N₂ and stirred for 45 minutes. Methyl A- (bromomethyl)benzoate (0.075g, 0.33mmol) in tetrahydrofuran (31 ml_) was added to the reaction mixture and stirred for 5 minutes. The reaction mixture was quenched with saturated ammonium chloride and extracted with ethyl acetate. The organic later was dried over anhydrous magnesium sulfate, filtered, and concentrated in vacuo. The material was purified by preparatory LC/MS (UV method) to isolate the titled compound as a clear oil, (0.06Og, 42% yield). ¹H NMR (400 MHz, CDCI₃) δ ppm 7.99 (d, J=8.34 Hz, 2 H) 7.72 (d, J=7.58 Hz, 1 H) 7.32 - 7.48 (m, 3 H) 7.24 (d, J=8.34 Hz, 2 H) 7.15 - 7.21 (m, 1 H) 7.06 - 7.11 (m, 2 H) 6.91 (dd, J=8.21 , 1.14 Hz, 1 H) 5.41 - 5.56 (m, 2 H) 3.92 (s, 3 H) 3.02 - 3.17 (m, 2 H) 1.47 (s, 3 H). Preparation of C-1 4-({2-methyl-3-oxo-4-r2-(trifluoromethyl)benzyl1-3,4-dihvdro-2H-1 ,4-benzothiazin-2- vDmethvDbenzoic acid

Methyl 4-({2-methyl-3-oxo-4-[2-(thfluoromethyl)benzyl]-3,4-dihydro-2H-1 ,4-benzothiazin- 2-yl}methyl)benzoate (0.06Og, 0.124mmol) taken up in 1 :1 methanol/tetrahydrofuran (2ml_), to which lithium hydroxide monohydrate ( 0.005g, 0.124mmol) in water (0.2ml_) was added. Reaction stirred for 16 hours at ambient temperature. The reaction mixture was quenched with 1 N hydrochloric acid, extracted with ethyl acetate, washed with brine, dried over anhydrous magnesium sulfate, filtered, and concentrated in vacuo to afford the title compound as an off white solid. (0.054g, 92% yield) ¹H NMR (400 MHz, CDCI₃) δ ppm 8.06 (d, J=8.34 Hz, 2 H) 7.73 (d, J=7.83 Hz, 1 H) 7.33 - 7.48 (m, 3 H) 7.27 - 7.31 (m, 2 H) 7.15 - 7.22 (m, 1 H) 7.04 - 7.13 (m, 2 H) 6.92 (d, J=7.58 Hz, 1 H) 5.41 5.56 (m, 2 H) 3.05 - 3.21 (m, 2 H) 1.48 (s, 3 H).

Method D

Example D-1

4-{r4-(4-cvano-2-nnethylbenzyl)-8-fluoro-3-oxo-3,4-dihvdro-2/-/-1 ,4-benzothiazin-2- vlimethvDbenzoic acid

Preparation of D-1 -a r(2-fluoro-6-nitrophenyl)sulfanyl1acetic acid

To a mixture of 1 -chloro-2-difluoro-6-nitrobenzene (9.83g, 56 mmol) in 63ml_ aq. acetone (1 water : 2 acetone) and thethylamine (79.4ml_, 570mmol), 2-mercaptoacetic acid (5.06 ml_, 72.8 mmol, 70% aq. w/w) was added. The reaction mixture was stirred under nitrogen at 32 C in dark for 1 h. The reaction mixture was diluted with dichloromethane and the washed with water (4x50ml_). The aqueous layer was acidified to pH=4 using 2N hydrogen chloride then was extracted with ethyl acetate (2x50ml_). The combined organics were washed with saturated aqueous sodium chloride, dried (anhydrous magnesium sulfate), filtered, and concentrated to provide the title compound (12.59g, 95% yield). ¹H NMR (400 MHz, DMSO-c/₆) δ ppm 12.82 (br. S., 1 H) 7.83 (d, J=7.07 Hz, 1 H) 7.50 - 7.71 (m, 2 H) 7.64 (dq, J=8.34, 8.08 Hz, 1 H) 3.75 (s, 2H). HRMS Calcd for C₈H₆FNO₄S (M+Na): 253.9894 Found: 253.9898.

Preparation of D-1-b 8-fluoro-2/-/-1 ,4-benzothiazin-3(4/-/)-one

A solution of sodium dithionite (12.3g, 60mmol) in 28 ml_ of water was added slowly to a stirred solution of 2-Fluoro-6-nitro-phenylsulfanyl-acetic acid (4.62 g, l Ommol) in water (4OmL) containing potassium carbonate (11.8 g, 80 mmol) at 3O C for 15 minutes. The reaction mixture was acidified using 6N hydrogen chloride to pH = 3 and the mixture was heated for 1 h at 3O⁰C. The reaction mixture was allowed to cool down and the precipitated solid was filtered out to provide the title compound (2.1 g, 57% yield). ¹H NMR (400 MHz, DMSO-c/₆) δ ppm 10.79 (br. s., 1 H) 7.20 (td, J=8.21 , 6.32 Hz, 1 H) 6.92 (dd, J= 17.68, 1.01 Hz, 1 H) 6.82 (d, J=7.83 Hz, 1 H) 3.52 (s, 2H). HRMS Calcd for C₈H₆FNOS (M+H): 184.0226 Found: 184.0228.

Preparation of D-1 -c 4-r(8-fluoro-3-oxo-2,3-dihvdro-4/-/-1 ,4-benzothiazin-4-yl)methyl1-3-methylbenzonitrile

The title compound was made analogous to the method described in Method A using instead 8-fluoro-2H-1 ,4-benzothiazin-3(4/-/)-one and 4-(bromomethyl)-3- methylbenzonitrile. ¹H NMR (400 MHz, CDCI₃) δ ppm 7.50 (s, 1 H) 7.43 (dd, J=7.96, 1.14 Hz, 1 H) 7.10 (td, J=8.34, 6.06 Hz, 1 H) 7.00 (d, J=7.83 Hz, 1 H) 6.82 - 6.93 (m, 1 H) 6.59 (d, J=8.34 Hz, 1 H) 5.18 (s, 2H) 3.56 (s, 2H) 2.43 (s, 3H). HRMS Calcd for C₁₇H₁₃FN₂OS (M+Na): 335.0625 Found: 335.0631. Preparation of D-1 -d methyl 4-{r4-(4-cvano-2-nnethylbenzyl)-8-fluoro-3-oxo-3,4-clihvdro-2/-/-1 ,4-benzothiazin-

2-vl1methvl)benzoate

The title compound was made analogous to the method described in Method C using instead 4-[(8-fluoro-3-oxo-2,3-dihydro-4H-1 ,4-benzothiazin-4-yl)methyl]-3- methylbenzonitrile. ¹H NMR (400 MHz, DMSO-c/₆) δ ppm 7.91 (m, J=8.34 Hz, 2 H) 7.71 (s, 1 H) 7.58 (dd, J=8.08, 1.52 Hz, 1 H) 7.43 (m, J=8.34 Hz, 2 H) 7.27 (td, J=8.34, 6.32 Hz, 1 H) 7.01 (d, J=8.08 Hz, 2 H) 7.06 (t, J=8.46 Hz, 2 H) 5.12 - 5.33 (m, 2 H) 4.34 (dd, J=9.22, 5.94 Hz, 1 H) 3.85 (s, 3 H) 2.96 (dd, J=14.40, 9.95 Hz, 1 H) 2.40 (s, 3H) 2.25 (s, 1 H). HRMS Calcd for C₂₆H_2IFN₂O₃S (M+Na): 483.1149 Found: 483.1150.

Preparation of D-1

4-{r4-(4-cvano-2-methylbenzyl)-8-fluoro-3-oxo-3,4-dihvdro-2/-/-1 ,4-benzothiazin-2- ylimethvDbenzoic acid

The title compound was made analogous to the method described in Method A using instead methyl 4-{[4-(4-cyano-2-methylbenzyl)-8-fluoro-3-oxo-3,4-dihydro-2H-1 ,4- benzothiazin-2-yl]methyl}benzoate. ¹H NMR (400 MHz, DMSO-c/₆) δ ppm 12.91 (br. s., 1 H) 7.88 (m, J=8.08 Hz, 2 H) 7.70 (s, 1 H) 7.57 (dd, J=8.08, 1.26 Hz, 1 H) 7.38 (m, J=8.08 Hz, 2 H) 7.23 - 7.32 (m, 1 H) 7.05 (t, J=8.46 Hz, 1 H) 7.00 (d, J=8.08 Hz, 1 H) 6.92 (d, J=8.34 Hz, 1 H) 5.23 - 5.35 (m, 1 H) 5.07 - 5.23 (m, 1 H) 4.32 (dd, J=9.35, 6.06 Hz, 1 H) 3.36 (d, J=8.59 Hz, 1 H) 2.94 (dd, J=14.40, 9.35 Hz, 1 H) 2.39 (s, 3H). HRMS Calcd for C₂₅Hi₉FN₂O₃S (M+H): 469.0992 Found: 469.0995. The racemic mixture was separated by chiral SFC and the 2 enantiomers were isolated as (+)4-{[4-(4-cyano-2- methylbenzyl)-8-fluoro-3-oxo-3,4-dihydro-2H-1 ,4-benzothiazin-2-yl]methyl}benzoic acid (D-2, [α]_D° =50.60; 99% ee) and (-)4-{[4-(4-cyano-2-methylbenzyl)-8-fluoro-3-oxo-3,4- dihydro-2H-1 ,4-benzothiazin-2-yl]methyl}benzoic acid (D-3, [α]_D° =-42.86; 97%ee).

The following compounds D-4 to D-7 listed in Table 1 were prepared in a similar manner 5 to Example D-1 starting with 1 -chloro-2-difluoro-6-nitrobenzene and using the appropriate benzyl bromide as the electrophile in Step c. In some cases the enantiomers were separated by chiral SFC to afford the chiral pure product.

Example D-8 o 4-{[4-(4-Cyano-2-methylbenzyl)-8-fluoro-2-methyl-3-oxo-3,4-dihydro-2H-1 ,4- benzothiazin-2-yl]methyl}benzoic Acid

Preparation of D-8-a 5 2-[(2-Fluoro-6-nitrophenyl)sulfanyl]propanoic Acid

The thioacetic acid (4.61 ml, 52 mmol) was added to the reaction mixture of 1 ,2-fluoro-6- nitrobenzene (6.36 g, 40 mmol) in 45 ml_ aq. acetonitrile (15 ml H₂O: 30 ml CH3CN) and Et₃N ( 67 ml, 400 mmol; 10 eq). The reaction was stirred under N₂ at 7O C in the dark for0 48 hour. The reaction mixture was diluted with dichloromethane (100 ml) then washed with water (3 x 100ml) until no product was in the organic layer. The aqueous layer was acidified with (15 ml) 2N hydrogen chloride until pH=4, extracted with dichloromethane twice. The combined organic layer dried over magnesium sulfate, filtered and solvent was evaporated. Crude product was obtained in 92% yield and was carried forward5 without further purification. ¹H NMR (400 MHz, DMSO-c/₆) δ ppm 12.90 (br. s., 1 H)

7.82 (t, J=4.67 Hz, 1 H) 7.62 - 7.73 (m, 2 H) 3.96 (q, J=7.07 Hz, 1 H) 1.32 (d, J=7.07 Hz, 4H). Preparation of D-8-b 8-Fluoro-2-methyl-2/-/-1 ,4-benzothiazin-3(4/-/)-one

The title compound was made analogous to the method described in Method D-1 preparation of D-1 -b using instead 2-[(2-Fluoro-6-nitrophenyl)sulfanyl]propanoic acid and was obtained in 21 % yield and used without further purification. ¹H NMR (400 MHz, DMSO-CZ₆) δ ppm 10.79 (s, 1 H) 7.21 (td, J=8.21 , 6.06 Hz, 1 H) 6.92 (dd, J=17.68, 1.01 Hz, 1 H) 6.83 (d, J=8.08 Hz, 1 H) 3.73 (q, J=7.07 Hz, 1 H) 1.32 (d, J=7.07 Hz, 3H). HRMS Calcd for C₉H₈FNOS (M+Na): 220.0202 Found: 220.0202

Preparation of D-8-c

4-[(8-Fluoro-2-methyl-3-oxo-2,3-dihydro-4/-/-1 ,4-benzothiazin-4-yl)methyl]-3- methylbenzonithle

The title compound was made analogous to the method described in Method A using instead 8-Fluoro-2-methyl-2H-1 ,4-benzothiazin-3(4/-/)-one and 4-(bromomethyl)-3- methylbenzonithle. The title compound was obtained in 82% yield and used without further purification. ¹H NMR (400 MHz, DMSO-c/₆) δ ppm 7.71 (s, 1 H) 7.58 (d, J=8.08 Hz, 1 H) 7.24 (td, J=8.34, 6.57 Hz, 1 H) 7.05 (t, J=8.59 Hz, 1 H) 6.98 (d, J=8.08 Hz, 1 H) 6.86 (d, J=8.34 Hz, 1 H) 5.06 - 5.33 (m, 2 H) 4.02 - 4.11 (m, 1 H) 2.40 (s, 3 H) 1.42 (d, J=6.82 Hz, 3H).

Preparation of D-8-d Methyl 4-{[4-(4-Cyano-2-methylbenzyl)-8-fluoro-2-methyl-3-oxo-3,4-dihydro-2/-/-1 ,4- benzothiazin-2-yl]methyl}benzoate

The title compound was made analogous to the method described in Method C using instead 4-[(8-Fluoro-2-methyl-3-oxo-2,3-dihydro-4H-1 ,4-benzothiazin-4-yl)methyl]-3- methylbenzonithle and was obtained in 59% yield and used without further purification. ¹H NMR (400 MHz, CDCI₃) δ ppm 7.99 (m, J=8.08 Hz, 2 H) 7.52 (s, 1 H) 7.42 (d, J=8.08 Hz, 1 H) 7.23 (m, J=8.34 Hz, 2 H) 7.16 (td, J=8.27, 6.19 Hz, 1 H) 6.90 - 7.01 (m, 2 H) 6.64 (d, J=8.34 Hz, 1 H) 5.21 (s, 2H) 3.93 (s, 3H) 3.09 -3.20 (m, 1 H) 2.95 - 3.09 (m, 1 H) 1.59 (s, 3H) 1.48 (s, 3H).

Preparation of D-8

4-{[4-(4-Cyano-2-methylbenzyl)-8-fluoro-2-methyl-3-oxo-3,4-dihydro-2/-/-1 ,4- benzothiazin-2-yl]methyl}benzoic Acid

The title compound was made analogous to the method described in Method A using instead methyl 4-{[4-(4-Cyano-2-methylbenzyl)-8-fluoro-2-methyl-3-oxo-3,4-dihydro-2H- 1 ,4-benzothiazin-2-yl]methyl}benzoate and was obtained in 75%yield. ¹H NMR (400 MHz, DMSO-c/e) δ ppm 12.84 (br. s., 1 H) 7.87 (m, J=8.34 Hz, 2 H) 7.71 (s, 1 H) 7.57 (d, J=7.83 Hz, 1 H) 7.31 (td, J=8.34, 6.57 Hz, 1 H) 7.25 (m, J=8.08 Hz, 2 H) 7.12 (t, J=8.46 Hz, 1 H) 7.03 (d, J=8.34 Hz, 1 H) 6.90 (d, J=8.08 Hz, 1 H) 5.30 (s, 2H) 2.95 - 3.13 (m, 2H) 2.41 (s, 3H) 1.40 (s, 3H). HRMS Calcd for C₂₆H_2IFN₂O₃S (M+H): 461.1329 Found: 461.1326.

Compound D-9 listed in Table 1 was prepared in a similar manner to Example D-7 starting with 1 ,2-fluoro-6-nitrobenzene and using 2-(thfluoromethyl)-benzylbromide as the electrophile in step c. The following compounds D-10 to D-18 listed in Table 1 were prepared in a similar manner to Example D-1 starting with 2-chloro-3-methyl-nitrobenzene and using the appropriate benzyl bromide as the electrophile in step c.

Compound D-19 to D-21 listed in Table 1 was prepared in a similar manner to Example D-1 starting with 2-chloro-5-fluoronitrobenzene and using 2-(thfluoromethyl)- benzylbromide as the electrophile in step c.

Compound D-22 to D-24 listed in Table 1 was prepared in a similar manner to Example D-1 starting with 2,4-difluoro-nitrobenzene and using 2-(trifluoromethyl)-benzylbromide as the electrophile in step c. The enantiomers were separated by chiral SFC to afford the chiral pure products.

Compound D-25 listed in Table 1 was prepared in a similar manner to Example D-1 starting at step c with 7-methyl-1 ,4-benzothiazin-3(4H)-one and 2-(thfluoromethyl)- benzylbromide.

Compound D-26 to D-28 listed in Table 1 was prepared in a similar manner to Example D-1 starting with 2-fluoro-3-chloronitrobenzene and using 2-(trifluoromethyl)- benzylbromide as the electrophile in step c. The enantiomers were separated by chiral SFC to afford the chiral pure products.

Compound D-29 to D-31 listed in Table 1 was prepared in a similar manner to Example D-1 starting with 2,6-difluoronitrobenzene and using 2-(trifluoromethyl)-benzylbromide as the electrophile in step c. The enantiomers were separated by chiral SFC to afford the chiral pure products. Method E

Example E-1

[3-({3-Oxo-4-[2-(trifluoromethyl)benzyl]-3,4-dihydro-2H-1 ,4-benzothiazin-2- yl}methyl)phenoxy]acetic Acid

Preparation of E-1 -a 4-[2-(Trifluoromethyl)benzyl]-2/-/-1 ,4-benzothiazin-3(4/-/)-one

The title compound was made analogous to Method A using 2H-benzo[b][1 ,4]thiazin- 3(4H)-one and 1 -(bromomethyl)-2-(thfluoromethyl) benzene at ambient temperature and was obtained in 83% yield. ¹H NMR (400 MHz, DMSO-c/₆) δ ppm 7.80 (d, J=7.58 Hz, 1 H) 7.59 (t, J=7.45 Hz, 1 H) 7.47 (dd, J=I. Iλ , 1.39 Hz, 2 H) 7.19 (dd, J=7.96, 1.89 Hz, 1 H) 7.14 - 7.23 (m, 2 H) 7.05 (td, J=7.52, 1.14 Hz, 1 H) 6.87 (dd, J=8.34, 1.01 Hz, 1 H) 5.30 (s, 2H) 3.75 (s, 2H). HRMS Calcd for Ci₆H₁₂F₃NOS (M+Na): 346.0483 Found: 346.0487. Preparation of E-1 -b 2-(3-Methoxybenzyl)-4-[2-(trifluoromethyl)benzyl]-2/-/-1 ,4-benzothiazin-3(4/-/)-one

The title compound was made analogous to Method C using 4-[2- (Trifluoromethyl)benzyl]-2H-1 ,4-benzothiazin-3(4/-/)-one and 1 -(bromomethyl)-3- methoxybenzene instead of ethyl 4-(bromomethyl)benzoate at ambient temperature and was obtained in 40% yield. ¹H NMR (400 MHz, CDCI₃) δ ppm 7.71 (d, J=7.58 Hz, 1 H) 7.33 - 7.47 (m, 4 H) 7.22 - 7.26 (m, 1 H) 7.09 - 7.16 (m, 2 H) 7.00 - 7.07 (m, 1 H) 6.87 - 6.93 (m, 1 H) 6.75 - 6.85 (m, 2 H) 5.36 - 5.59 (m, 2 H) 3.76 - 3.87 (s, 3 H) 3.58 (s, 1 H) 3.39 (dd, J=14.02, 4.93 Hz, 1 H) 2.85 (dd, J=14.02, 10.23 Hz, 1 H). HRMS Calcd for C₂₄H₂₀F₃NO₂S (M+Na): 466.1059 Found: 466.1063.

Preparation of E-1 -c 2-(3-Hydroxybenzyl)-4-[2-(trifluoromethyl)benzyl]-2H-1 ,4-benzothiazin-3(4H)-one

A solution of 2-(3-methoxybenzyl)-4-[2-(trifluoromethyl)benzyl]-2H-1 ,4-benzothiazin- 3(4H)-one (190 mg, 0.42 mmol) in dichloromethane (4 ml_) was cooled to O C under nitrogen and treated dropwise with 1 M boron thbromide in dichloromethane (1.71 ml_). The solution was left to warm to ambient temperature for 5 hours. The mixture was poured into of ice, extracted with dichloromethane (3x30ml_), and the combined organic layers were concentrated in vacuo. The crude product obtained in 98% yield was directly carried to the next step without further purification. ¹H NMR (400 MHz, CDCI₃) δ ppm 7.71 (d, J=8.08 Hz, 1 H) 7.32 - 7.50 (m, 3 H) 7.00 - 7.25 (m, 4 H) 6.89 (d, J=8.08 Hz, 1 H) 6.62 - 6.84 (m, 3 H) 5.36 - 5.56 (m, 2 H) 3.80 (dd, J=9.98, 5.18 Hz, 1 H) 3.36 (dd, J=14.27, 5.18 Hz, 1 H) 2.84 (dd, J=14.02, 9.98 Hz, 1 H). HRMS Calcd for C₂₃H₁₈F₃NO₂S (M+Na): 452.0953 Found: 452.0956. Preparation E-1 -d

1 -Methylethyl [3-({3-Oxo-4-[2-(trifluoromethyl)benzyl]-3,4-dihydro-2H-1 ,4-benzothiazin-

2-yl}methyl)phenoxy]acetate

2-(3-Hydroxybenzyl)-4-[2-(trifluoromethyl)benzyl]-2/-/-1 ,4-benzothiazin-3(4H)-one (138 mg, 0.320 mmol) was dissolved in dimethylformannide (1 ml_), and to this solution was added potassium t-butoxide (55 mg, 0.32 mmol). The mixture was stirred under nitrogen for 30 minutes, lsopropyl bromoacetate (40 uL, .0.31 mmol) was added via syringe. The solution was then stirred at ambient temperature for 18 hours. The mixture was quenched with water and then extracted with ethyl acetate (3 x 50 ml_). The combined organic layers were then washed sequentially with water, 1 N sodium hydroxide, and brine, dried with magnesium sulfate, and concentrated in vacuo. The crude product was purified using column chromatography, eluting with 0-10% ethyl acetate / heptane. Product was obtained as a yellow solid (111 mg, 69%), which was used without further purification. ¹H NMR (400 MHz, DMSO-c/₆) δ ppm 7.80 (d, J=7.58 Hz, 1 H) 7.60 (t,

J=7.45 Hz, 1 H) 7.49 (t, J=7.33 Hz, 1 H) 7.44 (dd, J=7.83, 1.52 Hz, 1 H) 7.22 (t, J=7.83 Hz, 1 H) 7.17 - 7.31 (m, 1 H) 7.14 (d, J=7.83 Hz, 1 H) 7.07 (t, J=7.07 Hz, 1 H) 6.97 (d, J=8.08 Hz, 1 H) 6.83 - 6.91 (m, 2 H) 6.78 (dd, J=8.97, 1.89 Hz, 1 H) 5.34 - 5.44 (m, 1 H) 5.21 - 5.34 (m, 1 H) 4.98 (quin, J=6.25 Hz, 1 H) 4.71 (s, 2 H) 4.18 (dd, J=9.73, 5.68 Hz, 1 H) 3.21 - 3.31 (m, 1 H) 2.78 (dd, J=14.40, 9.85 Hz, 1 H) 1.20 (dd, J=6.06 Hz, 2.27 Hz, 6H). HRMS Calcd for C₂₈H₂₆F₃NO₄S (M+Na): 552.1427 Found: 552.1437. Preparation of E-1

[3-({3-Oxo-4-[2-(trifluoromethyl)benzyl]-3,4-dihydro-2H-1 ,4-benzothiazin-2- yl}methyl)phenoxy]acetic Acid

To a solution of 1 -methylethyl [3-({3-oxo-4-[2-(trifluoromethyl)benzyl]-3,4-dihydro-2/-/- 1 ,4-benzothiazin-2-yl}methyl)phenoxy]acetate (110 mg, 0.20 mmol) in methanol (4 ml_), 1 N LiOH (0.416 ml_, 0.416 mmol) was added. After stirring at ambient temperature for 18 hours, the mixture was concentrated to remove solvent, then acidified with 1 N hydrogen chloride. The product was extracted with ethyl acetate (3 x 25 ml_), and the combined organic layers were dried with magnesium sulfate, filtered, and concentrated in vacuo. A pale white solid (92 mg, 91 %) was obtained. ¹H NMR (400 MHz, DMSO-c/₆) δ ppm 7.79 (d, J=7.58 Hz, 1 H) 7.59 (t, J=7.58 Hz, 1 H) 7.38 - 7.53 (m, 3 H) 7.10 - 7.33 (m, 3 H) 7.06 (t, J=7.07 Hz, 1 H) 6.96 (d, J=8.08 Hz, 1 H) 6.72 - 6.89 (m, 2 H) 5.24 - 5.45 (m, 2 H) 4.63 (s, 2 H) 4.14 - 4.17 (m, 1 H) 3.28 (d, J=9.09 Hz, 1 H) 2.77 - 2.79 (m,

1 H). LCMS (M+H)⁺: 488.0. The racemic mixture was separated by chiral SFC and the

2 enantiomers were isolated as (-)-[3-({3-oxo-4-[2-(trifluoromethyl)benzyl]-3,4-dihydro- 2H-1 ,4-benzothiazin-2-yl}methyl)phenoxy]acetic acid (E-2, [α]_D° = -55.74; 99% ee) and (+)-[3-({3-oxo-4-[2-(thfluoromethyl)benzyl]-3,4-dihydro-2/-/-1 ,4-benzothiazin-2- yl}methyl)phenoxy]acetic acid (E-3, [α]_D° =61 .62; 95% ee).

Method F

Example F-1

4-({6-Bromo-2-oxo-1-[2-(trifluoromethyl)benzyl]-2,3-dihydro-1 H-pyrido[2,3- b][1 ,4]thiazin-3-yl}methyl)benzoic Acid

Preparation of F-1-a 2,6-Dibromopyridin-3-amine

As suggested in Majumdar, K. C; Mondal, S. Regioselective synthesis of substituted pyrrolopyridines based on Pd(ll)-mediated cross coupling and base induced heteroannulation. Tet. Letters 2007, 48, 6951-6953, to a solution of 3-aminopyridine (4.75 g, 50.5 mmol) in DMSO (100 ml_) and water (2.5 ml_) at 0^°C was added portionwise N-bromosuccinimide (18.9 g, 8.91 mmol). The mixture stirred for 24 hours at ambient temperature. A solid was formed, which was filtered off and dried on a glass frit. The solid was then dissolved in ethyl acetate and dried over sodium sulfate and filtered. Solvent was removed under vacuum to afford a reddish brown solid (11.7g, 92%), which was used without further purification and displayed ¹H NMR that matched previously reported (Parlow, J. J.; Kurumbail, R. G.; Stegeman, R. A.; Stevens, A. M.; Stallings, W. C; South, M. S. Design, Synthesis, and Crystal Structure of Selective 2-Pyridone Tissue Factor Vila Inhibitors. J. Med. Chem. 2003, 46, 4696-4701 ). Preparation of F-1-b 6-Bromo-1 H-pyrido[2,3-b][1 ,4]thiazin-2(3H)-one

Similar to that described in Dunn, A. D.; Norrie, R. The synthesis of pyrido[1 ,4]thiazinones. J. Prakt. Chem. 1990, 332, 444-452: 60% sodium hydride (8.33 g, 347 mmol) was taken up in tetrahydrofuran (1 L), and add methylthioglycolate (20 g, 190 mmol) was added. After stirring for 1 h, 2,6-dibromopyhdin-3-amine (40 g, 160 mmol) was added. The reaction mixture was heat to reflux for 18 hours, allowed to cool, and quenched by adding methanol (80 ml_). Solvent was removed under reduced pressure to afford a brown solid residue. The brown solid was washed with hot ethyl acetate (5 x 200 ml_) and filtered. The ethyl acetate mother liquor was concentrated to 15% of original volume. The solid was filtered, then washed with ethyl acetate to afford the titled product (9.Og, 22%). ¹H NMR (400 MHz, DMSO-c/₆) δ ppm 10.75 (br. s., 1 H) 7.41 (d, J=8.34 Hz, 1 H) 7.18 (d, J=8.34 Hz, 1 H) 3.69 (s, 3 H). Preparation of F-1-c

6-Bromo-1 -[2-(trifluoromethyl)benzyl]-1 H-pyrido[2,3-b][1 ,4]thiazin-2(3H)-one

The title compound was made analogous to the method described in Method A using 1 - (bromomethyl)-2-(thfluoromethyl)benzene at ambient temperature, the title compound was obtained in 44% yield. ¹H NMR (400 MHz, DMSO-c/₆) δ ppm 7.80 (d, J=7.58 Hz, 1 H) 7.61 (t, J=7.58 Hz, 1 H) 7.51 (t, J=7.45 Hz, 1 H) 7.44 (d, J=8.59 Hz, 1 H) 7.20 (d, J=7.83 Hz, 1 H) 7.16 (d, J=8.59 Hz, 1 H) 5.26 (s, 2 H) 3.97 (s, 2 H). Preparation of F-1-d

Methyl 4-({6-Bromo-2-oxo-1 -[2-(trifluoromethyl)benzyl]-2,3-dihydro-1 H-pyrido[2,3- b][1,4]thiazin-3-yl}methyl)benzoate

The title compound was made analogous to the method described in Method C using methyl 4-(bromomethyl)benzoate at ambient temperature, the title compound was obtained in 80% yield as pale white solid. ¹H NMR (400 MHz, DMSO-c/₆) δ ppm 7.91 (d, J=8.08 Hz, 2 H) 7.80 (d, J=7.58 Hz, 1 H) 7.62 (t, J=7.83 Hz, 1 H) 7.48 (t, J=9.22 Hz, 4 H) 7.28 (d, J=8.84 Hz, 1 H) 7.14 (d, J=7.83 Hz, 1 H) 5.16 - 5.33 (m, 2 H) 4.54 (dd, J=8.97, 6.19 Hz, 1 H) 3.85 (s, 3 H) 3.43 (dd, J=14.53, 6.69 Hz, 1 H) 3.06 (dd, J=14.53, 8.97 Hz, 1 H). HRMS Calcd for C₂₄H₁₈BrN₂O₃S (M+H): 551.0246 Found: 551.0266.

Preparation of F- 1

4-({6-Bromo-2-oxo-1 -[2-(thfluoromethyl)benzyl]-2,3-dihydro-1 H-pyrido[2,3-b][1 ,4]thiazin- 3-yl}methyl)benzoic Acid

The title compound was made analogous to the method described in Method A using Methyl 4-({6-Bromo-2-oxo-1 -[2-(thfluoromethyl)benzyl]-2,3-dihydro-1 H-pyrido[2,3- b][1 ,4]thiazin-3-yl}methyl)benzoate at ambient temperature, the title compound was obtained in 20% yield as a white solid. ¹H NMR (400 MHz, DMSO-d₆) δ ppm 12.90 (br. s., 1 H) 7.88 (d, J=8.08 Hz, 2 H) 7.80 (d, J=7.58 Hz, 1 H) 7.58 - 7.65 (m, 1 H) 7.49 (d, J=8.84 Hz, 2 H) 7.41 (d, J=8.08 Hz, 2 H) 7.28 (d, J=8.59 Hz, 1 H) 7.13 (d, J=7.83 Hz, 1 H) 5.18 - 5.41 (m, 2 H) 4.53 (dd, J=9.09, 6.32 Hz, 1 H) 3.41 (d, J=8.34 Hz, 1 H) 3.03 (dd, J=14.65, 9.09 Hz, 1 H). HRMS Calcd TOr C₂₃Hi₆BrN₂O₃S (M+H): 537.0089 Found: 536.9901. Preparation of F-1 -e

Methyl 4-({6-Methyl-2-oxo-1-[2-(trifluoromethyl)benzyl]-2,3-dihydro-1H-pyrido[2,3- ϋ][1,4]thiazin-3-yl}methyl)benzoate

A solution of 2M methylzinc chloride in tetrahydrofuran (0.12 ml_, 0.24 mmol) was added slowly to a stirring solution of methyl 4-({6-bromo-2-oxo-1 -[2-(trifluoromethyl)benzyl]-2,3- dihydro-1 H-pyrido[2,3-ib][1 ,4]thiazin-3-yl}methyl)benzoate (110 mg, 0.2 mmol), tetrakis(triphenylphosphine)-palladium(0) (11 mg, 0.01 mmol), and tetrahydrofuran (0.4 ml_) in the microwave tube. The mixture was microwaved at 8O C for 10 min. The mixture was quenched by addition of saturated ammonium chloride solution (4 ml_), and extracted with ethyl acetate (3x 15 ml_). The combined organic layers were washed with sat. sodium bicarbonate, then brine, dried over magnesium sulfate, and filtered The solvent was removed under reduced pressure. The residue was purified by column chromatography eluted with 15-50% ethyl acetate / heptane. Product was obtained (62 mg, 72%) as a clear oil. ¹H NMR (400 MHz, CDCI₃) δ ppm 8.00 (d, J=8.34 Hz, 2 H) 7.71 (d, J=7.83 Hz, 1 H) 7.44 (t, J=7.45 Hz, 1 H) 7.33 - 7.41 (m, 3 H) 6.98 - 7.09 (m, 2 H) 6.89 (d, J=8.34 Hz, 1 H) 5.39 (s, 2 H) 3.92 (s, 3 H) 3.52 (dd, J=14.15, 5.56 Hz, 1 H) 3.06 (dd, J=14.27, 9.22 Hz, 1 H) 2.47 (s, 3 H) 0.89 (t, J=6.82 Hz, 1 H). HRMS Calcd for C₂₅H₂₁F₃N₂O₃S (M+Na): 509.1117 Found: 509.0884. Preparation of F-2

4-({6-Methyl-2-oxo-1 -[2-(trifluoromethyl)benzyl]-2,3-dihydro-1 H-pyrido[2,3-b][1 ,4]thiazin-

3-yl}methyl)benzoic Acid

The title compound was made analogous to the method described in Method A using Methyl 4-({6-Methyl-2-oxo-1 -[2-(thfluoromethyl)benzyl]-2,3-dihydro-1 H-pyrido[2,3- £>][1 ,4]thiazin-3-yl}methyl)benzoate at ambient temperature, the title compound was obtained in 48% yield as white solid. ¹H NMR (400 MHz, DMSO-c/₆) δ ppm 12.84 (br. s., 1 H) 7.87 (d, J=8.08 Hz, 2 H) 7.78 (d, J=7.83 Hz, 1 H) 7.59 (t, J=7.33 Hz, 1 H) 7.46 - 7.53 (m, 1 H) 7.41 (d, J=8.34 Hz, 2 H) 7.20 - 7.29 (m, 1 H) 7.11 (t, J=3.92 Hz, 2 H) 5.26 - 5.42 (m, 2 H) 4.39 (dd, J=9.22, 6.44 Hz, 1 H) 3.37 - 3.48 (m, 1 H) 2.99 (dd, J=14.40, 9.09 Hz, 1 H) 2.36 (s, 3H). HRMS Calcd for C₂₄H₁₉F₃N₂O₃S (M+H): 473.1141 Found: 473.1244. The racemic mixture was separated by chiral SFC and compound F-3 was isolated at RT=2.747 min (>99%ee) and F-4 was isolated at RT=3.244 min (98%ee).

The following compounds F-5 to F-7 listed in Table 1 were prepared in a similar manner to Example F-2 starting with 3-aminopyridine in step a and using 5-methyl-2- (trifluoromethyl)benzylbromide as the electrophile in step c. The enantiomers were separated by chiral SFC to afford the chiral pure products.

The following compounds F-8 to F-10 listed in Table 1 were prepared in a similar manner to Example F-1 starting with step b using 2-chloro-4-methylpyridin-3-amine. The enantiomers were separated by chiral SFC to afford the chiral pure product. Method G

Example G-1

2-Fluoro-4-r3-oxo-4-(2-tπfluoroιinethyl-benzyl)-3,4-clihvdro-2H-benzori ,41thiazin-2- ylmethyli-benzoic acid

Preparation of G-1 -a 2-Fluoro-4-methyl-benzoic acid methyl ester

To a mixture of 2-fluoro-4-methylbenzoic acid (2.5g, 16mmol) in methyl alcohol (30ml_),thionyl chloride (1.36ml_, 18.9mmol) was added and the mixture was stirred at room temperature for 18h. The solvent was evaporated to provide the title compound (2.6g, 98% yield). ¹H NMR (400 MHz, METHANOL-^) δ ppm 7.79 (t, J=7.83 Hz, 1 H), 7.07 (d, J=8.08 Hz, 1 H), 7.02 (d, J=12.38 Hz, 1 H), 3.88 (s, 3 H), 2.39 (s, 3 H) Preparation of G-1 -b 4-Bromonnethyl-2-fluoro-benzoic acid methyl ester

To a mixture of 2-Fluoro-4-methyl-benzoic acid methyl ester (2.6g, 15.46mmol) in carbontetrachloride (25mL), N-bromo succinimide (2.75g, 15.5mmol) and benzoyl peroxide (0.023g, 0.093mmol) were added and the reaction mixture was stirred at 6O⁰C for 18h. The solution was concentrated to half the volume then the precipitated solid was filtered. The filtrate was concentrated and purified using silica gel column chromatography eluting with 0-10% ethyl acetate in hexanes to provide the title compound (1.6g, 42% yield). ¹H NMR (400 MHz, DMSO-c/₆) δ ppm 7.88 (t, J=7.83 Hz, 1 H), 7.46 (dd, J=11.87, 1.52 Hz, 1 H), 7.41 (dd, J=8.08, 1.52 Hz, 1 H), 4.74 (s, 2 H), 3.85 (s, 3 H).

Preparation of G-1 -c 4-(2-Thfluoromethyl-benzyl)-4H-benzo[1 ,41thiazin-3-one

To the solution of (2H),1 ,4-benzothiazin-3(4H)-one (3.30 g, 20 mmol) in anhydrous dimethylformamide (3OmL) was added the solution of sodium bis(thmethylsilyl)amide in tetrahydrofuran (1.0 M, 21 ml). After stirring for 5 minutes, 2-(thfluoromethyl)benzyl chloride (4.28 g, 22.0 mmol) was added and the mixture was stirred at room temperature for 4 days. After filtration through celite and concentration to remove solvent, the residue was purified on silica gel column chromatography eluting with 0- 20% of ethyl acetate / heptane to provide the title compound (4.4g, 68% yield).¹H NMR (400 MHz, DMSO-c/e) δ ppm 7.80 (d, J=7.83 Hz, 1 H), 7.59 (t, J=7.58 Hz, 1 H), 7.48 (m, 2 H), 7.20 (m, 2 H), 7.06 (td, J=7.58, 1.01 Hz, 1 H), 6.88 (d, J=7.33 Hz, 1 H), 5.31 (s, 2 H), 3.75 (s, 2 H).

Preparation of G-1 -d 2-Fluoro-4-r3-oxo-4-(2-trifluoromethyl-benzyl)-3,4-dihvdro-2H-benzon ,41thiazin-2- ylmethyli-benzoic acid methyl ester

The title compound was made analogous to the method described in Method C using A- (2-Trifluoromethyl-benzyl)-4H-benzo[1 ,4]thiazin-3-one. ¹H NMR (400 MHz, DMSO-c/₆) δ ppm 7.81 (m, 2 H), 7.59 (t, J=7.45 Hz, 1 H), 7.48 (t, J=7.58 Hz, 1 H), 7.44 (dd, J=7.71 , 1.39 Hz, 1 H), 7.33 (dd, J=12.13, 1.26 Hz, 1 H), 7.24 (m, 2 H), 7.14 (d, J=7.83 Hz, 1 H), 7.07 (t, J=7.58 Hz, 1 H), 6.98 (dd, J=8.21 , 0.88 Hz, 1 H), 5.33 (m, 2 H), 4.30 (dd, J=9.47, 5.94 Hz, 1 H), 3.84 (s, 3 H), 3.39 (dd, J=14.40, 6.06 Hz, 1 H), 2.95 (dd, J=14.40, 9.35 Hz, 1 H).

Preparation of G-1

2-Fluoro-4-r3-oxo-4-(2-trifluoromethyl-benzyl)-3,4-dihvdro-2H-benzori ,41thiazin-2- ylmethyli-benzoic acid

The title compound was made analogous to the method described in Method A using 2- Fluoro-4-[3-oxo-4-(2-thfluoromethyl-benzyl)-3,4-dihydro-2H-benzo[1 ,4]thiazin-2- ylmethyl]-benzoic acid methyl ester. ¹H NMR (400 MHz, DMSO-c/₆) δ ppm 13.16 (br. s., 1 H), 7.80 (ddd, J=7.77, 4.17, 3.98 Hz, 2 H), 7.59 (t, J=7.45 Hz, 1 H), 7.50 (m, 1 H), 7.45 (dd, J=7.83, 1.52 Hz, 1 H), 7.28 (m, 2 H), 7.22 (m, 1 H), 7.15 (d, J=7.58 Hz, 1 H), 7.08 (td, J=7.58, 1.01 Hz, 1 H), 6.98 (d, J=8.34 Hz, 1 H), 5.34 (m, 2 H), 4.29 (dd, J=9.35, 6.06 Hz, 1 H), 3.38 (dd, J=14.40, 5.81 Hz, 1 H), 2.94 (dd, J=14.53, 9.47 Hz, 1 H). The enantiomers were separated by chiral SFC to afford the chiral pure products G-2 and G- 3 listed in Table 1. The following compound G-4 listed in Table 1 was prepared in a similar manner to Method G using benzylbromide as the electrophile in step c.

The following compounds G-5 to G-7 listed in Table 1 were prepared in a similar manner to Method G starting with 2-fluoro-4-methylbenzoic acid in step a. The enantiomers were separated by chiral SFC to afford the chiral pure products.

Example H-1

4-({2-oxo-1 -r2-(thfluoromethyl)benzyl1-1 ,2,3,4-tetrahvdroquinolin-3-yl)methyl)benzoic acid

Preparation H-1 -a 1 -r2-(tπfluoronnethyl)benzyl1-3,4-clihvdroquinolin-2(1 H)-one

To a mixture of 3,4-dihydroquinolin-2(1 /-/)-one (0.75 g, 4.8 mmol) in dimethylformannide (15 ml), cooled to O⁰C, was added sodium hydride (0.25 g, 6.3 mmol) in one portion. The reaction mixture was stirred at room temperature for 40 minutes then 1 - (bromomethyl)-4-(thfluoromethyl)benzene (1.3 g, 5.3 mmol) was added and the reaction was stirred at room temperature overnight. The reaction mixture was quenched with aqueous saturated ammonium chloride. The product was extracted out with ethyl acetate (3 X 30 ml_). Combined organics were dried over magnesium sulfate, filtered, and concentrated to dryness. The crude mixture was purified by silica gel chromatography, using 4-40% ethyl acetate / hexanes to provide the title compound (1.4 g, 95 %). LCMS (IvRH)⁺: 306.20.

Preparation H-1 -b methyl 4-({2-oxo-1 -[2-(trifluoromethyl)benzvπ-1 ,2,3,4-tetrahvdroquinolin-3- vDmethvDbenzoate

To a mixture of the diisopropylamine (0.74 ml_, 5.1 mmol) in tetrahydrofuran (20 ml_ ) was added 2.5 M butyllithium (2.2 ml_, 5.4 mmol)) dropwise. After stirring for 15 minutes, the mixture was cooled to -78 ⁰C. and a mixture of 1 -[2-(thfluoromethyl)benzyl]- 3,4-dihydroquinolin-2(1H)-one (1.4 g, 4.7 mmol) in tetrahydrofuran (15 ml_) was added. After stirring for 45 minutes, a mixture of methyl 4-(bromomethyl)benzoate (1.3 g, 5.6 mmol) in tetrahydrofuran (10 ml_) was added and was allowed to stir for 1 h at -78⁰C continued to stir at room temperature overnight. The reaction was quenched with sat. aqueous ammonium chloride. The product was extracted out with ethyl acetate (3 X 50 mL). Combined organics were dried over magnesium sulfate, filtered, and concentrated to dryness. The crude mixture was purified by silica gel chromatography, using 5-40% ethyl acetate in hexanes to provide the title compound (0.10 g, 4.8 %). ¹H NMR (300 MHz, DMSO-c/e) δ ppm 7.95-7.90 (2H, m), 7.84-7.80 (1 H, m), 7.62-7.56 (1 H, m), 7.53- 7.48 (1 H, m), 7.47-7.42 (2 H, m), 7.23-7.19 (1 H, m), 7.18-7.12(2 H, m), 7.02-6.96 (1 H, m), 6.68-6.63 (1 H, m), 5.45-5.07 (2 H, m), 5.86 (3 H, s), 3.37-3.29 (1 H, m), 3.22-3.12 (1 H, m), 2.88-2.74 (3 H, m).

Preparation of H-1

4-({2-oxo-1 -[2-(trifluoromethyl)benzyl1-1 ,2,3,4-tetrahvdroquinolin-3-yl)methyl)benzoic acid

To a mixture of methyl 4-({2-oxo-1 -[2-(trifluoromethyl)benzyl]-1 ,2,3,4-tetrahydroquinolin- 3-yl}methyl)benzoate (0.10 g, 0.22 mmol) in 1.5 mL of 1 :1 :1 MeOH: tetrahydrofuran : water, was added 1.5 mL of aqueous 2 N sodium hydroxide. The mixture was stirred at room temperature for 1 hour then concentrated to dryness. The crude mixture was purified on the prep HPLC to give the title compound as a white solid (7.8 mg, 7.8%). ¹H NMR (300 MHz, CH₃OD) δ ppm 7.97-7.85 (2H, m), 7.70-7.65 (1 H, m), 7.45-7.38 (1 H, m), 7.37-7.30 (1 H, m), 7.27-7.22 (2 H, m), 7.09-6.99 (3 H, m), 6.95-6.88(1 H, m), 6.64-6.58 (1 H, m), 5.36-5.17 (2 H, m), 3.28-3.22 (1 H, m), 3.05-2.95 (1 H, m), 2.90-2.82 (1 H, m), 2.75-2.62 (2 H, m).

Method 1-1

Example 1-1

4-[5-Fluoro-2-oxo-1 -(2-trifluoronnet hyl-benzyl)-1 ,2,3,4-tetrahydro-quinolin-3-ylnnethyl1- benzoic acid

Preparation 1-1 -a

2-(2-Fluoro-6-nitro-benzyl)-nnalonic acid diethyl ester

Sodium hydride (1.08g, 26.9mmol) was suspended in dimethylformamide (2OmL) at O⁰C. Diethyl malonate (3.9ml_, 24.9mmol) was added in three portions, and the resulting suspension was stirred at O⁰C for 10 min. A solution of 2-bromomethyl-1 -fluoro-3- nitrobenzene (5.Og, 21 mmol) in dimethylformamide (2OmL) was added dropwise. The reaction mixture was stirred for 45 min at O⁰C and then diluted with saturated ammonium chloride and extracted with ethyl acetate (2x50mL). The combined organics were dried over anhydrous magnesium sulfate, filtered, and concentrated to provide the title compound (6.Og, 90%). LCMS (IvRH)⁺: 314 Preparation 1-1 -b

2-(2-Amino-6-fluoro-benzyl)-malonic acid diethyl ester

2-(2-Fluoro-6-nitro-benzyl)-malonic acid diethyl ester (2g, δ.Ommol) was dissolved in methanol (5OmL), ammonium chloride (1.38g, 25.5mmol) and zinc dust (2.04g, 30.6mmol) were added and the mixture was stirred at room temperature for15min. Zinc was filtered out on a celite pad and the pad was washed with methanol (5OmL). Solvent was evaporated to provide the title compound (1g, 70%). LCMS (M+H)⁺: 284. Preparation 1-1 -c

5-Fluoro-3,4-dihvdro-1 H-quinolin-2-one

To a solution of 2-(2-amino-6-fluoro-benzyl)-malonic acid diethyl ester (1.Og) in acetic acid (1 OmL) was added hydrochloric acid (1 OmL), and the mixture was stirred at 9O⁰C for 1 h. The reaction mixture was poured in water and extracted with ethyl acetate (2x30mL). The combined organics were dried over magnesium sulfate and the solvent was evaporated. The residue was purified using silica gel column, eluting with 0-100% ethyl acetate in heptane to provide the title compound (0.2g, 34%). ¹H NMR (400 MHz, DMSO-c/e) δ ppm 10.26 (br. s., 1 H), 7.17 (m, 1 H), 6.78 (t, J=8.46 Hz, 1 H), 6.70 (d, J=7.83 Hz, 1 H), 2.88 (t, J=7.58 Hz, 2 H), 2.47 (m, 2 H) Preparation 1-1 -d

5-Fluoro-1 -(2-trifluoromethyl-benzyl)-3,4-dihvdro-1 H-quinolin-2-one

The title compound was made analogous to the method described in Method A using 5- fluoro-3,4-dihydro-1 H-quinolin-2-one and 2-(trifluoromethyl)benzylbromide. ¹H NMR (400 MHz, DMSO-c/e) δ ppm 7.80 (d, J=7.58 Hz, 1 H) 7.57 (t, J=7.45 Hz, 1 H) 7.48 (t, J=7.58 Hz, 1 H) 7.17 (m, 2 H) 6.91 (t, J=8.59 Hz, 1 H) 6.47 (d, J=8.34 Hz, 1 H) 5.23 (s, 2 H) 3.02 (t, J=7.45 Hz, 2 H) 2.80 (m, 2 H).

Preparation of 1-1 -e

4-[5-Fluoro-2-oxo-1 -(2-trifluoromethyl-benzyl)-1 ,2,3,4-tetrahvdro-quinolin-3-ylmethyl1- benzoic acid methyl ester

The title compound was made analogous to the method described in Method C using 5- fluoro-1 -(2-trifluoromethyl-benzyl)-3,4-dihydro-1 H-quinolin-2-one. ¹H NMR (400 MHz, DMSO-CZ₆) δ ppm 7.92 (d, J=8.08 Hz, 2 H) 7.81 (d, J=7.58 Hz, 1 H) 7.59 (m, 1 H) 7.50 (t, J=7.33 Hz, 1 H) 7.45 (d, J=8.08 Hz, 2 H) 7.17 (m, 2 H) 6.91 (t, J=8.72 Hz, 1 H) 6.51 (d, J=8.34 Hz, 1 H) 5.41 (m, 1 H) 5.10 (d, J=17.43 Hz, 1 H) 3.85 (s, 3 H) 3.33 (m, 1 H) 3.21 (m, 1 H) 2.92 (dd, J=15.66, 5.81 Hz, 1 H) 2.82 (dd, J=13.64, 9.35 Hz, 1 H) 2.69 (dd, J=15.92, 11.37 Hz, 1 H). LCMS (M+H)⁺: 472.

Preparation of 1-1

4-[5-Fluoro-2-oxo-1 -(2-trifluoromet hyl-benzyl)-1 ,2,3,4-tetrahydro-quinolin-3-ylmethyl1- benzoic acid

The title compound was made analogous to the method described in Method A using 4- [5-fluoro-2-oxo-1 -(2-trifluoromethyl-benzyl)-1 ,2,3,4-tetrahydro-quinolin-3-ylmethyl]- benzoic acid methyl ester. ¹H NMR (400 MHz, DMSO-c/₆) δ ppm 12.90 (br. s., 1 H) 7.90 (d, J=8.34 Hz, 2 H) 7.81 (d, J=7.58 Hz, 1 H) 7.59 (t, J=7.45 Hz, 1 H) 7.50 (t, J=7.58 Hz, 1 H) 7.41 (d, J=8.34 Hz, 2 H) 7.17 (m, 2 H) 6.91 (t, J=8.59 Hz, 1 H) 6.51 (d, J=8.34 Hz, 1 H) 5.41 (d, J=17.68 Hz, 1 H) 5.11 (d, J=17.68 Hz, 1 H) 3.33 (m, 1 H) 3.20 (m, 1 H) 2.93 (dd, J=15.92, 6.06 Hz, 1 H) 2.80 (dd, J=13.52, 9.22 Hz, 1 H) 2.69 (dd, J=15.92, 11.37 Hz, 1 H). LCMS (M+H)⁺: 458. The enantiomers were separated by chiral SFC to afford the chiral pure products I-2 and I-3 listed in Table 1.

The following compounds I-4 to I-6 listed in Table 1 were prepared in a similar manner to the preparation of 1-1 using 4-(bromomethyl)benzonitrile as the electrophile in step d. The enantiomers were separated by chiral SFC to afford the chiral pure products.

Example I-7

4-({5-chloro-2-oxo-1 -[2-(trifluoromethyl)benzyl1-1 ,2,3,4-tetrahvdroquinolin-3- vDmethvDbenzoic acid

Preparation of 2-(bromomethyl)-1 -chloro-3-nitrobenzene

To a mixture of 1 -chloro-2-methyl-3-nitrobenzene (4.0 g, 23 mmol) in 25 ml_ of tetrachloromethane was added 1 -bromopyrrolidine-2,5-dione (4.6 g, 26 mmol) and 2,2'- (E)-diazene-1 ,2-diylbis(2-methylpropanenitrile) (0.70 g, 4.7 mmol). The mixture was stirred at reflux for 16h. The mixture was filtered and concentrated to dryness. The crude mixture was purified by silica gel chromatography, using 5-40% ethyl acetate in hexanes to provide the title compound (4.9 g, 84 %). ¹H NMR (300 MHz, CDCI3) δ ppm 7.86 (1 H, dd, J =1.2, 8.19 Hz) 7.69 (1 H, dd, J= 1.3, 8.1 Hz) 7.43 (1 H, t, J=8.2 Hz) 4.89 (2 H, s).

Preparation of l-7-a diethyl (2-chloro-6-nitrobenzyl)propanedioate

The title compound was made analogous to the method described in Example 1-1 using 2-(bromomethyl)-1 -chloro-3-nitrobenzene. LCMS (M+H)⁺: 330.0.

Preparation of l-7-b diethyl (2-amino-6-chlorobenzyl)propanedioate

The title compound was made analogous to the method described in Example 1-1 using diethyl (2-chloro-6-nitrobenzyl)propanedioate. LCMS (M+H)⁺: 300.0. Preparation of l-7-c

5-chloro-3,4-dihvdroquinolin-2(1 /-/)-one

The title compound was made analogous to the method described in Example 1-1 using diethyl (2-amino-6-chlorobenzyl)propanedioate. LCMS (M+H)⁺: 182.0. Preparation of l-7-d

5-chloro-1 -r2-(trifluoromethyl)benzyl1-3,4-dihvdroquinolin-2(1 /-/)-one

The title compound was made analogous to the method described in Method A using 5- chloro-3,4-dihydroquinolin-2(1 H)-one and 2-(thfluoromethyl)benzylbromide. LCMS (M+H)⁺: 340.0.

Preparation of l-7-e methyl 4-({5-chloro-2-oxo-1 -[2-(trifluoromethyl)benzvπ-1 ,2,3,4-tetrahydroquinolin-3- vDmethvDbenzoate

The title compound was made analogous to the method described in Method C using 5- chloro-1 -[2-(trifluoromethyl)benzyl]-3,4-dihydroquinolin-2(1 /-/)-one. LCMS (M+H)⁺: 488.0.

Preparation of I-7

4-({5-chloro-2-oxo-1 -r2-(trifluoromethyl)benzyl1-1 ,2,3,4-tetrahvdroquinolin-3- vDmethvDbenzoic acid

The title compound was made analogous to the method described in Method A using methyl 4-({5-chloro-2-oxo-1 -[2-(trifluoromethyl)benzyl]-1 ,2,3,4-tetrahydroquinolin-3- yl}methyl)benzoate and obtained in 27 % yield. ¹H NMR (300 MHz, CDCI₃) δ ppm 8.14- 8.03 (2H, m), 7.76-7.69 (1 H, m), 7.48-7.31 (4 H, m), 7.12-6.98 (3 H, m), 6.64-6.57 (1 H, m), 5.49-5.26 (2 H, m), 3.48-3.37 (1 H, m), 3.19-3.00 (2 H, m), 2.92-2.76 (2 H, m). The enantiomers were separated by chiral SFC to afford the chiral pure products I-8 and I-9 listed in Table 1.

The following compounds 1-10 to 1-12 listed in Table 1 were prepared in a similar manner to the preparation of I-7 using 4-(bromomethyl)benzonitrile as the electrophile in step d. The enantiomers were separated by chiral SFC to afford the chiral pure products.

The following compounds 1-13 to 1-15 listed in Table 1 was prepared in a similar manner to Method I starting 2-(bromomethyl)-1 -methyl-3-nitrobenzene in step a and using 2- (trifluoromethyl)benzylbromide as the electrophile in step c. The enantiomers were separated by chiral SFC to afford the chiral pure products.

The following compound 1-16 listed in Table 1 was prepared in a similar manner to Method I starting 2-(bromomethyl)-1 -methyl-3-nitrobenzene in step a and using 4- (bromomethyl)benzonithle as the electrophile in step c.

Method J

Example J-1

4-({2-Oxo-1 -[2-(trifluoromethyl)benzyl]-1 ,2-dihydroquinolin-3-yl}methyl)benzoic Acid

Preparation of J-1 -a Diethyl (4-Cyanobenzyl)propanedioate

Prepared essentially as described in literature (Morphy, J. R.; et al J. Chem. Soc, Perkin Trans. 2 1990, 573-585), except without dimethylformamide: Sodium (4.6 g, 200 mmol) was carefully dissolved in ethanol (100 ml_) under nitrogen atmosphere. To this solution at ambient temperature was added diethyl malonate (32.0 g, 200 mmol). After 1 h, to the resultant slurry was added a solution of 4-cyanobenzyl bromide (39.2g, 200 mmol) in ethanol (60 ml_). Immediate formation of a solid-gel was observed. The mixture was stirred at reflux for 24h. The resultant heterogeneous mix was concentrated in vacuo, diluted with water (500 ml_) and ethyl acetate (150 ml_), and solid at interface filtered off. The aqueous layer was separated and extracted with ethyl acetate (2x150 ml_). The combined organic layers were washed with water (3x150 ml_), dried over magnesium sulfate, filtered, and concentrated to yield a gummy solid, which was purified via column chromatography with silica gel and eluted with a stepwise gradient of hexanes: ethyl acetate (10:1 to 4:1 ) to afford 21.2 g (39%) of oil, which matched the reported ¹H NMR (Morphy, J. R.; et al J. Chem. Soc, Perkin Trans. 2 1990, 573-585) and was used without further purification.

Preparation of J-1 -c Ethyl 2-(4-Cvanobenzyl)prop-2-enoate

To a stirring solution of diethyl (4-cyanobenzyl)propanedioate (21.2 g, 77.0 mmol) in ethanol (110 ml_) was added a solution of KOH (4.3 g, 77 mmol) in ethanol (100 ml_). A precipitate formed immediately, prompting shaking of the resultant viscous mixture, which eventually stirred at ambient temperature overnight. The cloudy mixture was concentrated in vacuo to a solid, which was taken up into water (100 ml_) and extracted with ether (3x4 ml_). The aqueous layer was acidified with 38% aq hydrogen chloride and extracted with diethyl ether (2x80 ml_), ethyl acetate (80 ml_). The extracts were combined, washed with water (2x80 ml_), dried over anhydrous magnesium sulfate, filtered, concentrated in vacuo, and azeotroped with dichloromethane (2x5 ml_) to yield presumed malonate monoacid J-1-b as an oil, which was used without further characterization or purification. A mixture of crude malonate J-1 -b ( 16.0 g, 65.0 mmol), pyridine (25 ml_), pipehdine (0.89 ml_), paraformaldehyde (3.0 g) stirred under nitrogen at 7O⁰C overnight. The resultant mix was poured onto ice and acidified with 38% aq hydrogen chloride and extracted with ether (3x50 ml_) and ethyl acetate (50 ml_). The combined extracts were washed with sat aq sodium bicarbonate (2x5 ml_), dried over anhydrous magnesium sulfate, filtered, and concentrated down to afford an oil, which was purified via chromatography with silica gel and elution with ethyl acetate :hexanes (1 :100 to 4:100) to provide 11.06 g (79.1 %) of exo-methylene product as an oil, which was used without further purification. ¹H NMR (400 MHz, DMSO-c/₆) δ ppm 7.77 (d, J=8.34 Hz, 3 H) 7.40 (d, J=8.34 Hz, 2 H) 6.20 (s, 1 H) 5.73 (d, J= 1.26 Hz, 1 H) 4.09 (q, J=7.07 Hz, 2 H) 3.69 (s, 2 H) 1.16 (t, J=7.07 Hz, 3H).

Preparation of J-1 -d 4-[(2-Oxo-1 ,2-dihydroquinolin-3-yl)methyl]benzonitrile

To a 50 ml_ pressure vessel were added 2-iodoaniline (110 mg, 0.5 mmol), ethyl 2-(4- cyanobenzyl)but-3-enoate (215 mg, 1.00 mmol), Pd(OAc)₂ (5.6 mg, 0.025 mmol), PPh₃ (13 mg, 0.05 mmol), NaOAc (408 mg, 3.00 mmol), and dimethylformamide (5 ml_). After the system flushed with argon, the reaction of mixture was allowed to heat at 100 C for 18 hours. The reaction of mixture was passed through a short silica gel column to eliminate black precipitate. Remove of the solvent, the crude product was purified by column chromatography eluted with 15 to 75% ethyl acetate/ heptane, the title compound was obtained (45 mg, 30%) as a brown solid. ¹H NMR (400 MHz, DMSO-c/₆) d ppm 11.83 (s, 1 H) 7.76 (d, J=8.34 Hz, 2 H) 7.79 (s, 1 H) 7.60 (d, J=7.83 Hz, 1 H) 7.41 - 7.54 (m, 3 H) 7.29 (d, J=7.83 Hz, 1 H) 7.15 (t, J=7.45 Hz, 1 H) 3.92 (s, 2H). HRMS Calcd for Ci₇H₂₂N₂OiS (M+H): 261.1022 Found: 261.1027.

Preparation of J-1 -e 4-({2-Oxo-1 -r2-(trifluoromethyl)benzyl1-1 ,2-dihvdroquinolin-3-yl)methyl)benzonitrile

The title compound was made analogous to the method described in Method A using 4- [(2-0X0-1 ,2-dihydroquinolin-3-yl)methyl]benzonitrile and 2-(thfluoromethyl)benzylbromide at ambient temperature. ¹H NMR (400 MHz, CDCI₃) δ ppm 7.86 (d, J=8.34 Hz, 1 H) 7.78 (s, 1 H) 7.70 (t, J=7.20 Hz, 2 H) 7.63 (dt, J=8.40, 7.17 Hz, 2 H) 7.52 (d, J=8.34 Hz, 2 H) 7.44 (d, J= 1.26 Hz, 1 H) 7.42 (d, J=3.03 Hz, 1 H) 7.34 - 7.38 (m, 1 H) 7.28 - 7.30 (m, 2 H) 5.72 (s, 2 H) 4.13 (s, 2 H). HRMS Calcd for C₂₅Hi₇F₃N₂O (M+H): 419.1365 Found: 419.1432.

Preparation of J-1 4-({2-Oxo-1 -[2-(trifluoromethyl)benzvπ-1 ,2-dihydroquinolin-3-yl)methyl)benzoic Acid

To a solution of 4-({2-oxo-1 -[2-(trifluoromethyl)benzyl]-1 ,2-dihydroquinolin-3- yl}methyl)benzoic acid (25 mg, 0.06 mmol) in ethanol (0.25 ml_) was added 8M sodium hydroxide (0.225 ml_, 1.8 mmol). The mixture was refluxed for 18. Acidified with 2N hydrogen chloride to pH = 4, then extracted with ethyl acetate (3x 15 ml_). The combined organics were dried over magnesium sulfate, and solvent evaporated. The crude product was purified by chromatography, eluted with 10 to 50% ethyl acetate / heptane to obtain the product (20 mg, 76%) as white solid. ¹H NMR (400 MHz, DMSO- Cf₆) δ ppm 12.83 (br. s, 1 H) 8.13 (s, 1 H) 7.80 - 7.88 (m, 3 H) 7.73 - 7.80 (m, 2 H) 7.64 (td, J=7.64, 1.39 Hz, 1 H) 7.56 (dt, J=5.24, 2.56 Hz, 2 H) 7.41 - 7.48 (m, 2 H) 7.31 (d, J=8.34 Hz, 2H) 5.66 (s, 2H) 4.15 (s, 2H). HRMS Calcd for C₂₅Hi₈F₃NO₃ (M+H): 438.1311 Found: 438.1310. Method K-1

Example K-1 4-({(3S,R4S,f?)-4-methyl-2-oxo-1 -r2-(tπfluoromethyl)benzyl1-1 ,2,3,4-tetrahvdroquinolin-

3-vl)methyl)benzoic acid

Preparation of K-1 -a 4-methyl-3,4-dihvdroquinolin-2(1 /-/)-one

To 130 g of polyphosphoric acid was added 3-methylindan-1 -one (9.82g, 67.2 mmol) and stirred with mechanical stirrer for 10 minutes. Sodium azide (4.58 g, 70.5 mmol) was added in portions while stirring for 20 minutes. The mixture was heated in 50 ⁰C oil bath while stirring overnight. The reaction mixture was cooled down and mixed with 600 ml of ice water and sonicated until all the polyphosphoric acid dissolves. This mixture was extracted by CH₂CI₂ 3 times, totally -700 ml. The CH₂CI₂ phase was washed with brine once, dried with sodium sulfate. After silica gel chromatography eluting with 0 - 50% hexane/EtOAc gradient 4.66 g of product was obtained (43% yield). ¹H NMR (400 MHz, DMSO-c/₆) δ ppm 1.17 (d, J=6.8Hz, 3 H) 2.22(dd, J=6.8 Hz, 16.2Hz, 1 H) 2.58(dd, J=5.8 Hz, 15.9Hz, 1 H) 2.95 - 3.13 (m, 1 H) 6.81 -6.88 (m, 1 H) 6.89-6.98 (m, 1 H) 7.08-7.16 (m, 1 H) 7.18 (d, J=7.3 Hz, 1 H) 10.07 (s, 1 H). LRMS (M+H)⁺: 162.2.

Preparation of K-1 -b 4-methyl-1 -r2-(trifluoromethyl)benzyl1-3,4-dihvdroquinolin-2(1 /-/)-one

To the solution of 4-methyl-3,4-dihydroquinolin-2(1 /-/)-one (806 mg, 5.00 mmol) in 15 ml of DMF, was added 5.5 ml of 1.0 M sodium bis(trimethylsilyl)amide (5.50 mmol) in tetrahydrofuran. After stirring for 10 minutes, 1 -(bromomethyl)-2-

(thfluoromethyl)benzene (1.26 g, 5.25 mmol) in 5 ml of DMF was added and the mixture was stirred at room temperature for 25 minutes. The mixture was then concentrated to remove solvent and the residue was purified on silica gel, eluting with 0 to 20% hexane/EtOAc gradient. 1.31 g of product was obtained (82% yield). ¹H NMR (400 MHz, DMSO-CZ₆) δ ppm 1.28 (d, J=7.1 Hz, 3 H) 2.57(dd, J=7.1 Hz, 15.7Hz, 1 H) 2.92(dd,

J=10.1 Hz, 15.7Hz, 1 H) 3.15 - 3.26 (m, 1 H) 5.13 (d, 17.2Hz, 1 H) 5.35 (d, 17.4Hz, 1 H) 6.61 (d, 7.4Hz, 1 H) 6.98-7.09 (m, 1 H) 7.09-7.21 (m, 2H) 7.32 (d, 7.3Hz, 1 H) 7.49 (t, 7.6Hz, 1 H) 7.58 (t, 7.6Hz, 1 H) 7.82 (d, J=7.6 Hz, 1 H) LRMS (M+H)⁺: 320.2

Preparation of K-1 -c methyl 4-({(3S,R4S,f?)-4-methyl-2-oxo-1 -r2-(trifluoromethyl)benzyll-1 ,2,3,4- tetrahvdroquinolin-3-vl)methyl)benzoate

At -78 ⁰C, to the solution of 4-methyl-1 -[2-(trifluoromethyl)benzyl]-3,4-dihydroquinolin- 2(1 H)-one (0.32 g, 1.0 mmol) in 15 ml of THF was added 1.1 ml of lithium bis(trimethylsilyl)amide (1.0 M in THF, 1.00 mmol) and stirred for 15 minutes. Then the solution of 4- (bromomethyl)benzoic acid methyl ester (0.23 g, 1.0 mmol) in 5 ml of THF was added slowly and reaction mixtures was stirred at cooling bath for 30 minutes. It was then warmed up to rt and concentrated to remove solvent. The residue was purified on silica gel column (0-20% heptane/EtOAc) 0.44 g of crude product was obtained and used directly to next step. LRMS (M+H)⁺: 468.2

Preparation of K-1 4-({(3S,R4S,f?)-4-methyl-2-oxo-1 -r2-(thfluoromethyl)benzyl1-1 ,2,3,4-tetrahvdroquinolin-

3-yl)methyl)benzoic acid

The title compound was made analogous to the method described in Method A using methyl 4-({(3S,R4S,R)-4-methyl-2-oxo-1 -[2-(thfluoromethyl)benzyl]-1 ,2,3,4- tetrahydroquinolin-3-yl}methyl)benzoate. ¹H NMR (400 MHz, DMSO-c/₆) δ ppm 1.18 (d, J=7.3Hz, 3 H) 2.69 - 2.80 (m, 1 H) 2.81 -3.00 (m, 3H) 3.00 - 3.13 (m, 1 H) 4.94 (d, J=17.4Hz, 1 H) 5.54 (d, J=17.2 Hz, 1 H) 6.70 (d, J=8.1 Hz, 1 H) 7.04-7.13 (m, 2H) 7.18- 7.25 (m, 1 H) 7.29 (d, J=8.4 Hz, 2 H) 7.50 (t, J=7.6 Hz, 1 H) 7.63 (t, J=7.5, 1 H) 7.82 (d, J=7.6 Hz, 1 H) 7.86 (d, J=8.3 Hz, 2 H) 12.85 (br. s., 1 H); LRMS (M+H)⁺: 454.2.

The following compounds K-2 to K-3 listed in Table 1 are enantiomers of compound K-1 separated by chiral SFC.

Method L-1

Example L-1

4-({7-chloro-1 -nnethyl-3-oxo-4-r2-(trifluoroιinethyl)benzyl1-1 ,2,3,4-tetrahvdroquinoxalin-2- vDmethvDbenzoic acid

Preparation of L-1 -a /V-(5-chloro-2-nitrophenyl)-/V-methylqlvcine

To a mixture of 2,4-dichloro-1 -nitrobenzene and sarcosine in 35 mL of EtOH and 15 mL of water was added sodium bicarbonate. The reaction mixture stirred at 80 ⁰C for 2 days. The mixture was diluted with 50 mL of water and acidified to a pH of 4 with 1 N hydrogen chloride. The product was extracted out with dichloromethane (3X50 mL) dried over magnesium sulfate, filtered, and concentrated to dryness to give the titled compound (2.8 g, 75 %). LCMS (IvRH)⁺: 245.0.

Preparation of L-1 -b 6-chloro-4-methyl-3,4-dihvdroquinoxalin-2(1 /-/)-one

A solution of sodium dithionite (14 g, 69 mmol) in 30 ml_ of water was added slowly to a stirred solution of Λ/-(5-chloro-2-nitrophenyl)-Λ/-methylglycine (2.8 g, 11 mmol) in 3OmL water containing potassium carbonate (12 g, 86 mmol). After stirring for 8 hours, the mixture was brought to a pH of 4 using 6N hydrogen chloride. The product was extracted out with ethyl acetate (3X50 ml_). Combined organics were dried over magnesium sulfate, filtered, and concentrated to dryness to give the titled compound (1.5 g, 67 %). LCMS (IvRH)⁺: 197.00.

Preparation of L-1 -c 6-chloro-4-methyl-1 -r2-(trifluoromethyl)benzyl1-3,4-dihvdroquinoxalin-2(1 /-/)-one

The title compound was made analogous to the method described in Method A using 6- chloro-4-methyl-3,4-dihydroquinoxalin-2(1 H)-one. LCMS (M+H)⁺: 355.0.

Preparation of L-1 -d methyl 4-({7-chloro-1 -methyl-3-oxo-4-[2-(trifluoromethyl)benzvH-1 ,2,3,4- tetrahydroquinoxalin-2-yl)methyl)benzoate

The title compound was made analogous to the method described in Method C using 6- chloro-4-methyl-1 -[2-(trifluoromethyl)benzyl]-3,4-dihydroquinoxalin-2(1 H)-one. LCMS (M+H)⁺: 503.0. Preparation of L-1

4-({7-chloro-1 -nnethyl-3-oxo-4-r2-(trifluoroιinethyl)benzyl1-1 ,2,3,4-tetrahvdroquinoxalin-2- vDmethvDbenzoic acid

The title compound was made analogous to the method described in Method A using methyl 4-({7-chloro-1 -methyl-3-oxo-4-[2-(trifluoromethyl)benzyl]-1 ,2,3,4- tetrahydroquinoxalin-2-yl}methyl)benzoate and obtained in 57 % yield. ¹H NMR (400 MHz, DMSO-c/e) δ ppm 7.84-7.74 (3H, m), 7.64-7.46 (2 H, m), 7.33-7.25 (2 H, m), 6.91 - 6.84 (1 H, m), 6.79-6.69 (2 H, m), 6.59-6.52 (1 H, m), 5.40-5.00 (2 H, m), 4.59-4.52 (1 H, m), 3.13-3.03 (1 H, m), 2.98-2.91 (4 H, m).

The following compounds L -2 to L-3 listed in Table 1 are enantiomers of compound L-1 separated by chiral SFC.

Example L-4

4-({8-chloro-1 -methyl-3-oxo-4-[2-(trifluoromethyl)benzyl1-1 ,2,3,4-tetrahvdroquinoxalin-2- vDmethvDbenzoic acid

Preparation of L-4-a r(2-chloro-6-nitrophenyl)(methyl)amino1acetate

The title compound was made analogous to the method described in Example L-1 preparation of L-1 -a using 2,4-dichloro-1 -nitrobenzene. LCMS (M+H)⁺: 245.0.

Preparation of L-4-b 5-chloro-4-methyl-3,4-dihvdroquinoxalin-2(1 H)-one

The title compound was made analogous to the method described in Example L-1 preparation L-1-b using [(2-chloro-6-nitrophenyl)(methyl)amino]acetate. LCMS (M+H)⁺: 197.0.

Preparation of L-4-c 5-chloro-4-methyl-1 -r2-(trifluoromethyl)benzyl1-3,4-dihvdroquinoxalin-2(1 /-/)-one

The title compound was made analogous to the method described in Method A using 5- chloro-4-methyl-3,4-dihydroquinoxalin-2(1 H)-one and 2-(trifluoromethyl)benzylbromide. LCMS (M+H)⁺: 355.0. Preparation of L-4-d methyl 4-({8-chloro-1 -methyl-3-oxo-4-r2-(trifluoromethyl)benzyl1-1 ,2,3,4- tetrahydroquinoxalin-2-yl)methyl)benzoate

The title compound was made analogous to the method described in Method C using 5- chloro-4-methyl-1 -[2-(trifluoromethyl)benzyl]-3,4-dihydroquinoxalin-2(1 H)-one. LCMS (M+H)⁺: 503.0. Preparation of L-4

4-({8-chloro-1 -nnethyl-3-oxo-4-r2-(trifluoroιinethyl)benzyl1-1 ,2,3,4-tetrahvdroquinoxalin-2- vDmethvDbenzoic acid

The title compound was made analogous to the method described in Method A using 5- chloro-4-methyl-3,4-dihydroquinoxalin-2(1 /-/)-one in place of 6-chloro-4-methyl-3,4- dihydroquinoxalin-2(1 H)-one and was obtained in 25 % yield. ¹H NMR (300 MHz, DMSO-CZ₆) δ ppm 7.89-7.80 (3H, m), 7.70-7.60 (1 H, m), 7.57-7.49 (1 H, m), 7.38-7.31 (2 H, m), 7.28-7.22 (1 H, m), 7.17-7.06 (2 H, m), 6.80-6.73 (1 H, m), 5.55-5.06 (2 H, m), 4.14-4.05 (1 H, m), 2.99-2.88 (1 H, m), 2.77 (3 h, s), 2.76-2.68 (1 H, m). The following compounds L-5 to L-6 listed in Table 1 are enantiomers of compound L-4 separated by chiral SFC.

The following compounds L -7 to L-9 listed in Table 1 were prepared in a similar manner to compound L-4 5-fluoro-2-(trifluoromethyl)benzyl bromide as the electrophile in step c. The enantiomers were separated by chiral SFC to afford the chiral pure products.

Example L-10 4-({7-fluoro-1 -methyl-3-oxo-4-[2-(trifluoromethyl)benzyl1-1 ,2,3,4-tetrahydroquinoxalin-2- vDmethvDbenzoic acid

Preparation of L-10-a Λ/-(5-fluoro-2-nitrophenyl)-Λ/-nnethylqlvcine

Following the procedure described in Example L-1 preparation of L-1-a using 2-chloro- 4-fluoro-1 -nitrobenzene to provide the title compound. ¹H NMR (400 MHz, DMSO-c/₆) δ ppm 12.87 (br. s., 1 H), 7.87 (dd, J=9.09, 6.32 Hz, 1 H), 6.88 (dd, J=12.25, 2.65 Hz, 1 H), 6.73 (ddd, J=9.28, 7.14, 2.53 Hz, 1 H), 4.00 (s, 2 H), 2.83 (s, 3 H). LCMS (M+H)⁺: 227.

Preparation of L-10-b 6-fluoro-4-methyl-3,4-dihvdroquinoxalin-2(1 /-/)-one

Following the procedure described in Example L-1 preparation of L-1-b using Λ/-(5- fluoro-2-nitrophenyl)-Λ/-methylglycine and2-(trifluoromethyl)benzylbromide to provide the title compound. ¹H NMR (400 MHz, DMSO-c/₆) δ ppm 10.42 (s, 1 H), 6.74 (dd, J=8.59, 5.81 Hz, 1 H), 6.54 (dd, J=11.12, 2.53 Hz, 1 H), 6.47 (m, 1 H), 3.67 (s, 2 H), 2.76 (s, 3 H). LCMS (M+H)⁺: 181

Preparation of L-10-c 6-fluoro-4-methyl-1 -r2-(trifluoromethyl)benzyl1-3,4-dihvdroquinoxalin-2(1 /-/)-one

Following the procedure described in Method A using 6-fluoro-4-methyl-3,4- dihydroquinoxalin-2(1 H)-one to provide the title compound. ¹H NMR (400 MHz, DMSO- Cf₆) δ ppm 7.81 (d, J=7.58 Hz, 1 H), 7.58 (t, J=7.45 Hz, 1 H), 7.49 (t, J=7.58 Hz, 1 H), 7.10 (d, J=7.83 Hz, 1 H), 6.68 (dd, J=10.99, 2.40 Hz, 1 H), 6.52 (m, 2 H), 5.24 (s, 2 H), 3.97 (s, 2 H), 2.85 (s, 3 H). LCMS (M+H)⁺: 339.

Preparation L-10-d 4-({7-fluoro-1 -nnethyl-3-oxo-4-r2-(tπfluoroιinethyl)benzyl1-1 ,2,3,4-tetrahvdroquinoxalin-2- vDmethvDbenzoate

Following the procedure described in Method C using 6-fluoro-4-methyl-1 -[2- (trifluoromethyl)benzyl]-3,4-dihydroquinoxalin-2(1 /-/)-one to provide the title compound. ¹H NMR (400 MHz, DMSO-c/₆) δ ppm 7.79 (m, 3 H), 7.58 (t, J=7.96 Hz, 1 H), 7.49 (t, J=7.58 Hz, 1 H), 7.32 (d, J=8.34 Hz, 2 H), 6.89 (d, J=7.83 Hz, 1 H), 6.60 (dd, J=10.99, 2.65 Hz, 1 H), 6.49 (m, 2 H), 5.31 (d, J=17.68 Hz, 1 H), 5.04 (d, J=17.68 Hz, 1 H), 4.53 (t, J=6.69 Hz, 1 H), 3.83 (s, 3 H), 3.07 (m, 1 H), 2.94 (m, 1 H), 2.90 (s, 3 H).

Preparation of L-10

4-({7-fluoro-1 -methyl-3-oxo-4-r2-(trifluoromethyl)benzyl1-1 ,2,3,4-tetrahvdroquinoxalin-2- vDmethvDbenzoic acid

To a solution of methyl 4-({7-fluoro-1-methyl-3-oxo-4-[2-(trifluoromethyl)benzyl]-1 ,2,3,4- tetrahydroquinoxalin-2-yl}methyl)benzoate (0.20Og, 0.41 mmol) in ethyl alcohol (1 OmL) was added 20% aqueous lithium hydroxide (3mL). The mixture was stirred at room temperature for 18h then was acidified to pH=5 with 2N aqueous hydrochloric acid and extracted with ethyl acetate (2x15mL). The combined organics were washed with saturated aqueous sodium chloride, dried (anhydrous magnesium sulfate), filtered and concentrated to provide the title compound (0.18g, 90%). ¹H NMR (400 MHz, DMSO-c/₆) δ ppm 12.84 (br. s., 1 H), 7.78 (m, 3 H), 7.58 (m, 1 H), 7.49 (t, J=8.34 Hz, 1 H), 7.29 (d, J=8.34 Hz, 2 H), 6.88 (d, J=8.08 Hz, 1 H), 6.60 (dd, J=10.99, 2.65 Hz, 1 H), 6.48 (m, 2 H), 5.32 (d, J=17.68 Hz, 1 H), 5.03 (m, 1 H), 4.53 (t, J=6.44 Hz, 1 H), 3.06 (m, 1 H), 2.94 (m, 1 H), 2.91 (s, 3 H).

The following compounds L-11 to L-12 listed in Table 1 are enantiomers of compound L-10 separated by chiral SFC.

The following compounds L-13 to L-21 listed in Table 1 were prepared in a similar manner to Method L starting with 2-chloro-1 -fluoro-3-nitrobenzene in step a and the appropriate benzylbromide as the electrophile in step c. The enantiomers were separated by chiral SFC to afford the chiral pure products.

Method M

Example M-1

4-r3-Oxo-4-(2-trifluoromethyl-benzyl)-3,4-dihvdro-2H-benzoπ ,41oxazin-2-ylmethyl1- benzoic acid

Preparation M-1 -a 4-(2-Thfluoromethyl-benzyl)-4H-benzo[1 ,41oxazin-3-one

To a solution of 2H-1 ,4-benzoxazine-3-(4H)-one (1.0 g, 6.6 mmol) in N₁N- dimethylfornnannide (10 mL) at O⁰C, was added sodium hydride (0.32g, 7.97 mmol) The mixture was then stirred at room temperature for 40min. 2-thfluoromethylbenzyl bromide (1.74g, 7.30mmol) was added and the mixture was stirred for 1 h. The reaction was quenched with saturated ammonium chloride solution and extracted with ethyl acetate (2x20 mL). The combined organics were washed with saturated aqueous sodium chloride, dried (anhydrous magnesium sulfate), filtered and concentrated. The residue was purified by silica gel chromatography using 0-25% ethyl acetate in hexanes to provide the title compound (1.5 g, 74%). ¹H NMR (400 MHz, DMSO-Cl₆) δ ppm 7.82 (d, J=7.83 Hz, 1 H), 7.59 (t, J=7.45 Hz, 1 H), 7.51 (t, J=7.45 Hz, 1 H), 7.23 (d, J=7.83 Hz, 1 H), 7.08 (m, 1 H), 7.01 (td, J=7.64, 1.39 Hz, 1 H), 6.95 (td, J=7.64, 1.64 Hz, 1 H), 6.68 (dd, J=7.96, 1.39 Hz, 1 H), 5.25 (s, 2 H), 4.87 (s, 2 H).

Preparation M-1 -b

4-r3-Oxo-4-(2-trifluoromethyl-benzyl)-3,4-dihvdro-2H-benzoπ ,41oxazin-2-ylmethyl1- benzoic acid methyl ester

To a solution of 4-(2-Thfluoromethyl-benzyl)-4H-benzo[1 ,4]oxazin-3-one (1.2 g, 3.3 mmol) in tetrahydrofuran (10 mL) at -78⁰C was added lithium-bis-(trimethylsilyl)-amid (4.98 mL, 4.98mmol).The resulting mixture was stirred at -78⁰C for 45min. 4- (bromomethyl)benzoic acid methyl (0.855g, 3.65mmol) in tetrahydrofuran (5 mL) was added to the mixture and was stirred at -78⁰C for 1 h then allowed to stir at ambient temperature for 18h. Saturated aqueous ammonium chloride was added and the mixture extracted with ethyl acetate (2X20mL). The combined organics were washed with saturated aqueous sodium chloride, dried (anhydrous magnesium sulfate), filtered, and concentrated. The residue was purified by silica gel chromatography using 10-50% ethyl acetate in hexanes to provide the title compound.

Preparation of M-1

4-r3-Oxo-4-(2-trifluoromethyl-benzyl)-3,4-dihvdro-2H-benzori ,41oxazin-2-ylmethyl1- benzoic acid

To a solution of 4-[3-oxo-4-(2-trifluoromethyl-benzyl)-3,4-dihydro-2H-benzo[1 ,4]oxazin- 2-ylmethyl]-benzoic acid methyl ester (0.08g, 0.18mmol) in ethyl alcohol (1OmL) was added 1 N aqueous sodium hydroxide (5ml_). The mixture was stirred at room temperature for 2hours then was acidified to pH=5 with 2N aqueous hydrochloric acid and extracted with ethyl acetate (2x15ml_). The combined organics were washed with saturated aqueous sodium chloride, dried (anhydrous magnesium sulfate), filtered and concentrated. The residue was purified by reverse phase HPLC to provide the title compound (0.013g, 17%). ¹H NMR (400 MHz, DMSO-Cl₆) δ ppm 12.91 (br. s., 1 H), 7.89 (d, J=8.08 Hz, 2 H), 7.82 (d, J=7.33 Hz, 1 H), 7.57 (t, J=7.45 Hz, 1 H), 7.50 (t, J=7.33 Hz, 1 H), 7.45 (d, J=8.08 Hz, 2 H), 7.09 (d, J=7.83 Hz, 1 H), 6.97 (m, 3 H), 6.71 (d, J=7.83 Hz, 1 H), 5.23 (m, 3 H), 3.39 (d, J=4.04 Hz, 1 H), 3.24 (m, 1 H) Method N

Example N-1

4-f3-Oxo-4-(2-trifluorOmethyl-benzyl)-3.4-dihvdro-2H-pyridof3.2-b1f1.41oxazin-2- ylmethyli-benzoic acid

Preparation N-1 -a 4-(2-Trifluoromethyl-benzyl)-4H-pyridor3,2-biri ,41oxazin-3-one

Following the procedure described in Method A using 2H-pyrido[3,2,B]-1 ,4-oxazin- 3(4H)-one in place of 2H-1 ,4-benzoxazine-3-(4H)-one to provide the title compound in 58% yield. ¹H NMR (400 MHz, METHANOL-d₄) δ ppm 7.90 (dd, J=4.80, 1.52 Hz, 1 H), 7.72 (d, J=7.58 Hz, 1 H), 7.45 (d, J=7.58 Hz, 1 H), 7.39 (m, 2 H), 7.14 (d, J=7.83 Hz, 1 H), 7.02 (dd, J=8.08, 4.80 Hz, 1 H), 5.54 (s, 2 H), 4.86 (s, 2 H)

Preparation N-1 -b 4-f3-Oxo-4-(2-trifluorOmethyl-benzyl)-3.4-dihvdro-2H-pyridof3.2-b1f1.41oxazin-2- ylmethyli-benzonitrile

Following the procedure described in Method C using 4-(2-Trifluoromethyl-benzyl)-4H- pyrido[3,2-b][1 ,4]oxazin-3-one and α-bromo-p-tolunitrile to provide the title compound in 30% yield. ¹H NMR (400 MHz, DMSO-Cl₆) δ ppm 7.93 (dd, J=4.80, 1.26 Hz, 1 H), 7.78 (m, 3 H), 7.54 (d, J=8.34 Hz, 2 H), 7.48 (m, 2 H), 7.42 (dd, J=7.96, 1.39 Hz, 1 H), 7.07 (dd, J=7.83, 4.80 Hz, 1 H), 7.01 (d, J=7.33 Hz, 1 H), 5.39 (m, 3 H), 3.43 (m, 1 H), 3.36 (d, J=8.08 Hz, 1 H)

Preparation of N-1

4-r3-Oxo-4-(2-trifluoromethyl-benzyl)-3,4-dihvdro-2H-pyridor3,2-biπ ,41oxazin-2- ylmethyli-benzoic acid

A mixture of 4-[3-Oxo-4-(2-trifluoromethyl-benzyl)-3,4-dihydro-2H-pyrido[3,2- b][1 ,4]oxazin-2-ylmethyl]-benzonitnle (0.36g, 0.85mnnol) in hydrochloric acid (12.5ml_) and ethyl alcohol (2.5ml_) was heated at reflux for 4h. The solvent was evaporated and the residue was purified by silica gel chromatography using 30-100% ethyl acetate in hexanes to provide the title compound and 4-[3-Oxo-4-(2-thfluoromethyl-benzyl)-3,4- dihydro-2H-pyrido[3,2-b][1 ,4]oxazin-2-ylmethyl]-benzoic acid ethyl ester. ¹H NMR (400 MHz, DMSO-dβ) δ ppm 7.92 (m, 1 H), 7.86 (d, J=8.34 Hz, 2 H), 7.77 (d, J=6.82 Hz, 1 H), 7.44 (m, 5 H), 7.07 (dd, J=7.83, 4.80 Hz, 1 H), 6.96 (d, J=7.33 Hz, 1 H), 5.39 (d, J=7.83 Hz, 2 H), 5.35 (dd, J=8.21 , 4.17 Hz, 1 H), 3.40 (d, J=4.29 Hz, 1 H), 3.27 (d, J=8.34 Hz, 1 H).

Method O

Example 0-1

4-[2-Oxo-1 -(2-trifluoromethyl-benzyl)-2,3-dihvdro-1 H-pyridor2,3-biπ ,41oxazin-3- ylmethyli-benzoic acid

Preparation O-1 -a 1 H-Pyridor2,3-biπ ,41oxazin-2-one

To a mixture of glycolic acid methyl ester (5g, 50mmol) in tetrahydrofuran (2OmL), 1 M lithium-bis-(trimethylsilyl)-amid in tetrahydrofuran (55ml_) was added slowly. After 5 min, 2-chloro-3-nitro-pyhdine (8.8g, 54.4mmol) in N,N-dimethylformamide (15mL) was added to the previous mixture and was stirred at room temperature for 18h. The reaction was quenched with saturated ammonium chloride solution and extracted with ethyl acetate (2x50 ml_). The combined organics were washed with saturated aqueous sodium chloride, dried (anhydrous magnesium sulfate), filtered and concentrated. The residue was purified by silica gel chromatography using 0-50% ethyl acetate in hexanes to provide (3-Nitro-pyridin-2-yloxy)-acetic acid methyl ester which was dissolved in methanol and raney nickel was added and the reaction mixture was stirred under hydrogen balloon for 2h. The catalyst was removed using celite pad and the solvent was evaporated to yield (3-Amino-pyhdin-2-yloxy)-acetic acid methyl ester in 50% yield. A mixture of (3-Amino-pyridin-2-yloxy)-acetic acid methyl ester in acetic acid was heated at 6O⁰C for 10min. the mixture was concentrated to provide the title compound which was used with no further purification.

Preparation O-1 -b 1 -(2-Trifluoronnethyl-benzyl)-1 H-pyrido[2,3-b1H ,41oxazin-2-one

Following the procedure described in Method A using 1 H-Pyhdo[2,3-b][1 ,4]oxazin-2-one in place of 2H-1 ,4-benzoxazine-3-(4H)-one to provide the title compound. ¹H NMR (400 MHz, DMSO-Cl₆) δ ppm 7.86 (dd, J=4.55, 1.77 Hz, 1 H), 7.82 (d, J=7.58 Hz, 1 H), 7.59 (s, 1 H), 7.51 (s, 1 H), 7.31 (d, J=7.58 Hz, 1 H), 7.04 (m, 2 H), 5.23 (s, 2 H), 5.07 (s, 2 H).

Preparation O-1 -c

4-[2-Oxo-1 -(2-trifluoromethyl-benzyl)-2,3-dihvdro-1 H-pyridor2,3-b1H ,41oxazin-3- ylmethyli-benzonitrile

Following the procedure described in Method C using 1 -(2-Trifluoromethyl-benzyl)-1 H- pyrido[2,3-b][1 ,4]oxazin-2-one and α-bromo-p-tolunithle to provide the title compound. LCMS (M+H)⁺: 424.

Preparation of 0-1

4-[2-Oxo-1 -(2-trifluoromethyl-benzyl)-2,3-dihvdro-1 H-pyridor2,3-b1H ,41oxazin-3- ylmethyli-benzoic acid

Following the procedure described in Method A using 4-[2-Oxo-1 -(2-thfluoromethyl- benzyl)-2,3-dihydro-1 H-pyhdo[2,3-b][1 ,4]oxazin-3-ylmethyl]-benzoic acid ethyl ester obtained the title compound. ¹H NMR (400 MHz, DMSO-Cl₆) δ ppm 12.99 (br. s., 1 H), 7.80 (d, J=8.34 Hz, 3 H), 7.75 (d, J=8.84 Hz, 1 H), 7.46 (m, 2 H), 7.37 (m, 2 H), 6.96 (m, 3 H), 5.43 (dd, J=8.08, 4.29 Hz, 1 H), 5.15 (m, 2 H), 3.37 (m, 1 H), 3.21 (m, 1 H).

Assay for Binding to Prostaglandin E2 Receptors

Membrane Preparation Transfected HEK-293 cells stably expressing prostaglandin E2 type 1 receptors (EP1 ), type 2 (EP2), type 3 (EP3) or type 4 (EP4) receptors are harvested. All operations here forth are performed at 4 ⁰C. Cells are pelleted in PBS with Eppendorf centrifuge (model 5804R with A-4-44 rotor) and resuspended in Stock Buffer A plus Protease Inhibitors [10 mM MES-NaOH (pH 6.0), 10 mM MgCl.sub.2, 1 mM EDTA, 1 mM Pefabloc peptide, (Sigma, St. Louis, Mo.), 10 uM Phosporamidon peptide, (Sigma, St. Louis, Mo.), 1 uM Pepstatin A peptide, (Sigma, St. Louis, Mo.), 10 uM Elastatinal peptide, (Sigma, St. Louis, Mo.), 100 uM Antipain peptide, (Sigma, St. Louis, Mo.)]. Cells are lysed by sonification with a Branson Sonifier (Model #450) or by hand-held homogenizer in 20 seconds bursts for 4-5 cycles. Unlysed cells and debris are separated by centrifugation at 25O.times.g for 10 min. Membranes are then harvested by ultra-centhfugation at 70,000.times.g for 30 minutes. Pelleted membranes are resuspended in Stock Buffer A plus Protease Inhibitors and 10% glycerol to 2-10 mg protein per ml, with protein concentration being determined according to BCA method. Resuspended membranes are then stored frozen at -80 ⁰C until use. Binding Assay Frozen membranes as prepared are thawed and diluted to 0.25 mg protein per ml in Stock Buffer A plus Protease Inhibitors. One volume of membrane preparation is combined with 0.05 volume test compound or buffer and one volume of 1.5 nM 3H- prostaglandin E2 (#TRK 431 , Amersham, Arlington Heights, III.) in Stock Buffer A plus Protease Inhibitors. The mixture (205 ul total volume) is incubated for 60 min at 25 ⁰C. The membranes are then recovered by filtration through Milllipore filter plates, MultiScreen-HTS-FB (cat# MSFCN6B50) by vacuum with MultiScreenHTS Vacuum Manifold (cat# MSVMHTS00). The membranes with bound 3H-prostaglandin E2 are trapped by the filter, the buffer and unbound 3H-prostaglandin E2 pass through the filter into waste. Each sample is then washed 5 times with 100 ul/well of [10 mM MES-NaOH (pH 6.0), 10 mM MgCl.sub.2, 1 mM EDTA]. The filters are then dried by heating in a microwave oven. To determine the amount of 3H-prostaglandin bound to the membranes, the dried filter plates are secured with Millipore plate adapter (cat# NC9292307), scintillant cocktail added and counted on the TopCount (PerkinElmer). IC50s are determined from the concentration of test compound required to displace 50% of the specifically bound 3H-prostaglandin E2.

Assay for Binding to Prostaglandin EP4 Receptors Membrane Preparation

Stably transfected HEK-293 cells expressing prostaglandin E2 type 1 receptors (EP1 ), type 2 (EP2), type 3 (EP3) or type 4 (EP4) receptors are grown to 70% confluency. All operations here forth are performed at 4⁰C. Cells are harvested with PBS buffer and centhfuged in Eppendorf centrifuge (model 5804R with A-4-44 rotor) to obtain pellets. Cell pellets are re-suspended in Stock Buffer A plus Protease Inhibitors consisting of [10 mM MES-NaOH (pH 6.0), 1 mM EDTA, 10 mM MgCI₂, 1 mM Pefabloc, (Sigma-Aldhch), 100 uM Antipain, (Sigma-Aldrich, 10 uM Phosporamidon, (Sigma-Aldrich), 10 uM Elastatinal, (Sigma-Aldrich), 1 uM Pepstatin A peptide, (Sigma-Aldrich)]. These are lysed by sonification with a Branson Sonifier (Model #450) or by hand-held homogenizer in 20 seconds bursts for 4-5 cycles. Lysate is then centhfuged at 250 times g for 10 min. Supernatant and pellet fractions are both placed on high speed centhfugation in Beckman Ultra Centrifuge with Ti 45 rotor at 70,000 times g, 4⁰C for 30 minutes. Membrane receptors are re-suspended to 2-10 mg protein per ml in Stock Buffer A plus Protease Inhibitors and 10% glycerol. Protein concentration is determined via BCA Method. Aliquots of membrane receptors are then stored frozen in at -8O⁰C until use. Binding Assay

Prepare test compounds to desired concentrations in 100% DSMO. Frozen membranes as prepared are thawed and diluted to 0.25 mg protein per ml in Stock Buffer A plus Protease Inhibitors. In pre-wetted 96-well filter plates, add one volume of 1.5 nM 3H- prostaglandin E2 (#TRK 431 , Amersham, Arlington Heights, III.) in Stock Buffer A plus Protease Inhibitors, followed by 0.05 volume of test compound. Reaction starts with the addition of one volume of diluted membranes (205 ul total volume). Incubation is for 40 min at room temperature. Membrane products are then captured by filtration through Milllipore filter plates, MultiScreen-HTS-FB (cat# MSFCN6B50) by vacuum with MultiScreenHTS Vacuum Manifold (cat# MSVMHTSOO). The membranes with bound 3H-prostaglandin E2 are trapped by the filter, the buffer and unbound 3H-prostaglandin E2 pass through the filter into waste. Each sample is then washed 5 times with 100 ul per well of [10 mM MES-NaOH (pH 6.0), 1 mM EDTA, 10 mM MgCI₂,]. The filter plates are then dried by heating in an oven. To determine the amount of 3H-prostaglandin bound to the membranes, dried filter plates are secured with Millipore plate adapter (cat# NC9292307), scintillant cocktail added and counted on the TopCount (PerkinElmer).

IC50s are determined from the concentration of test compound required to displace 50% of the specifically bound 3H-prostaglandin E2.

Table 1

Claims

What is claimed is:

1 ) A compound of formula (I):

wherein:

A¹, A², A³ and A⁴ are each independently selected from C, N, CR^A, NR^A, C-(O)R^A, N-(O)R^A, C-(R^AR^B) and N-(R^AR^B);

R¹, R², R³, R⁴ and R⁵ are each independently selected from a bond, H, halo, cyano, nitro, oxo, CF₃, OCF₃, Ci-s alkyl, C2-8 alkenyl, C2-8 alkynyl, Ci_-8 hydroxyalkyl, Ci_-8 haloalkyl, C_3-i₀ cycloalkyl, C_4-i₀ heterocycloakyl, C_3-i₀ cycloalkoxy, Ci_-8 alkoxy, Ci_-8 alkoxyalkyl, C_5-io aryl, and C_5--_I0 heteroaryl, wherein each of said Ci_-8 alkyl, C_2-8 alkenyl, C_2-8 alkynyl, Ci_-8 hydroxyalkyl, Ci_-8 haloalkyl, C_3-i₀ cycloalkyl, C_4-i₀ heterocycloakyl, C₃-₁₀ cycloalkoxy, Ci_-8 alkoxy, Ci_-8 alkoxyalkyl, C_5-io aryl, and C5-10 heteroaryl moieties is optionally substituted by one or more of R^A, R^AR^B, CR^A, CR^AR^B, SR^A, SR^AR^B, OR^A, COR^A, S(O)_aR^A, NR^AR^B, CONR^AR^B, N(O)R^AR^B, (CR^AR^B)_b(C_5-io aryl), (CR^AR^B)_b(C_5-io heteroaryl) or (CR^AR^B)_b(C_3-i₀ cycloalkyl);

R^A and R^B are independently selected from a bond, H, halo, cyano, nitro, oxo, CF₃, OCF₃, Ci_-8 alkyl, C_2-8 alkenyl, C_2-8 alkynyl, C_5-io aryl, C₅-₁₀ heteroaryl, arylalkyl, C₃-₁₀ cycloalkyl, C_4-i₀ heterocycloakyl, C_3-i₀ cycloalkoxy, Ci_-8 alkoxy, and C_2-8 alkoxyalkyl;

D¹ is C, N, O, S, CR^A, NR^A, OR^A, SR^A, C-(O)R^A, N-(O)R^A, C-(O)OR^A, N-(O)OR^A, C-(R^AR^B), N-(R^AR^B), or S-(R^AR^B); and each a is independently selected from 0, 1 , 2, 3, 4 and 5; each b is independently selected from 0, 1 , 2, 3, 4 and 5; or a pharmaceutically acceptable salt or stereoisomer thereof.

2) A compound of formula (I) according to claim 1 wherein: R¹ is selected from H, halo, cyano, CF₃, OCF₃, Ci-s alkyl, C2-8 alkenyl, C2-8 alkynyl, Ci_-8 hydroxyalkyl, Ci_-8 haloalkyl, Cs-io cycloalkyl, C_4-io heterocycloakyl, C_3-Io cycloalkoxy, C-_I-8 alkoxy, Ci_-8 alkoxyalkyl, C_5-io aryl, and C_5-i₀ heteroaryl; and

R³ is C_5-io aryl or C_5--_I0 heteroaryl, and wherein each of said C_5--_I0 aryl or C_5--_I0 heteroaryl moieties of R³ is optionally substituted by halo, cyano, Ci_-8 alkyl, C_2-8 alkenyl, C_2-8 alkynyl, Ci_-8 hydroxyalkyl, Ci_-8 haloalkyl, C_3-i₀ cycloalkyl, C_4-i₀ heterocycloakyl, C₃-₁₀ cycloalkoxy, Ci_-8 alkoxy, Ci_-8 alkoxyalkyl, C_5-i₀ aryl, or C₅-₁₀ heteroaryl.

3) A compound of formula (I) according to claim 1 wherein:

R³ is C_5-I0 aryl or C_5-I0 heteroaryl, and wherein each of said C_5-I0 aryl or C_5-I0 heteroaryl moieties is optionally substituted by Ci_-8 alkyl, C_2-8 alkenyl, C_2-8 alkynyl, Ci_-8 hydroxyalkyl, Ci_-8 haloalkyl, C_3-i₀ cycloalkyl, C_4-i₀ heterocycloakyl, C_3-I0 cycloalkoxy, Ci_-8 alkoxy, Ci_-8 alkoxyalkyl, C_5-i₀ aryl, or C_5-I0 heteroaryl; and

R⁵ is (CR^AR^B)_b(C_5-i₀ aryl) or (CR^AR^B)_b(C_5-i₀ heteroaryl), and wherein each of said (CR^AR^B)_b(C_5-i₀ aryl) and (CR^AR^B)_b(C_5-i₀ heteroaryl) moieties of R⁴ is independently substituted by at least one H, halo, cyano, nitro, oxo, CF₃, OCF₃, Ci_-8 alkyl, C_2-8 alkenyl, C_2-8 alkynyl, Ci_-8 hydroxyalkyl, Ci_-8 haloalkyl, C_3-i₀ cycloalkyl, C_4-i₀ heterocycloakyl, C_3-I0 cycloalkoxy, Ci_-8 alkoxy, Ci_-8 alkoxyalkyl, C_5-i₀ aryl, or C_5-I0 heteroaryl moiety.

4) A compound of Formula (II):

wherein:

A⁵, A⁶, A⁷ and A⁸ are each independently selected from C, N, CR^C, NR^C, C-

(O)R^C, N-(O)R⁰, C-(R⁰R⁰) and N-(R⁰R⁰); R⁶ and R⁷ are each independently selected from a bond, H, halo, cyano, nitro, oxo, CF₃, OCF₃, Ci-₈ alkyl, C₂-₈ alkenyl, C₂-₈ alkynyl, Ci_-8 hydroxyalkyl, Ci_-8 haloalkyl, C_{3- l}o cycloalkyl, C_4-i₀ heterocycloakyl, C_3--_I0 cycloalkoxy, Ci_-8 alkoxy, Ci_-8 alkoxyalkyl, C_5-i₀ aryl, and C_5-io heteroaryl, wherein each of said Ci_-8 alkyl, C_2-8 alkenyl, C_2-8 alkynyl, Ci_-8 hydroxyalkyl, Ci_-8 haloalkyl, C_3-i₀ cycloalkyl, C_4-i₀ heterocycloakyl, C_3-i₀ cycloalkoxy, Ci_-8 alkoxy, Ci_-8 alkoxyalkyl, C_5-io aryl, and C5-10 heteroaryl moieties is optionally substituted by one or more of R^c, R^CR^D, CR^C, CR^CR^D, NR^C, OR^C, SR^C, SR^CR^D, COR^C, S(O)_nR⁰, NR^CR^D, CONR⁰R⁰, N(O)R⁰R⁰, (CR⁰R⁰)_m(C_5-io aryl), (CR⁰R⁰)_m(C_5-io heteroaryl) or (CR^cR°)_m(C_3-io cycloalkyl); R⁸ is H, halo, cyano, nitro, oxo, CF₃, OCF₃, Ci_-8 alkyl, C_2-8 alkenyl, C_2-8 alkynyl, Ci- s hydroxyalkyl, Ci_-8 haloalkyl, C_3-io cycloalkyl, C_4-io heterocycloakyl, C_3-io cycloalkoxy, Ci- s alkoxy, Ci_-8 alkoxyalkyl, Cs-io aryl, or Cs-io heteroaryl;

R⁹ is H, halo, cyano, nitro, oxo, CF₃, OCF₃, Ci_-8 alkyl, C_2-8 alkenyl, C_2-8 alkynyl, Ci- ₈ hydroxyalkyl, Ci_-8 haloalkyl, C_3-i₀ cycloalkyl, C_4-i₀ heterocycloakyl, C_3-i₀ cycloalkoxy, Ci- ₈ alkoxy, Ci_-8 alkoxyalkyl, C_5-io aryl, or C5-10 heteroaryl, wherein each of said Ci_-8 alkyl, C_2-8 alkenyl, C_2-8 alkynyl, Ci_-8 hydroxyalkyl, Ci_-8 haloalkyl, C_3-i₀ cycloalkyl, C_4-i₀ heterocycloakyl, C_3-i₀ cycloalkoxy, Ci_-8 alkoxy, Ci_-8 alkoxyalkyl, C_5-io aryl, and C_5-io heteroaryl moieties is optionally substituted by one or more of CR⁰, CR⁰R⁰, SR⁰, SR⁰R⁰, OR⁰, COR⁰, S(O)_nR⁰, NR⁰R⁰, CONR⁰R⁰, N(O)R⁰R⁰, (CR^cR⁰)_m(C₅-io aryl), (CR^cR⁰)_m(C₅-io heteroaryl) or (CR^cR°)_m(C_3-io cycloalkyl);

R¹⁰ is H, halo, cyano, nitro, oxo, CF₃, OCF₃, Ci_-8 alkyl, C_2-8 alkenyl, C_2-8 alkynyl, Ci_-8 hydroxyalkyl, Ci_-8 haloalkyl, C_3-i₀ cycloalkyl, C_4-i₀ heterocycloakyl, C_3-i₀ cycloalkoxy, Ci_-8 alkoxy, Ci_-8 alkoxyalkyl, C_5-io aryl, or C5-10 heteroaryl;

R¹¹ is a bond, oxo, R°, OR⁰, Ci_-8 alkyl, C_2-8 alkenyl, C_2-8 alkynyl, Ci_-8 hydroxyalkyl, Ci_-8 haloalkyl, C_3-i₀ cycloalkyl, C_4-i₀ heterocycloakyl, C_3-i₀ cycloalkoxy, Ci_-8 alkoxy, Ci_-8 alkoxyalkyl, C_5-io aryl, or C₅-₁₀ heteroaryl, wherein each of said Ci_-8 alkyl, C_2-8 alkenyl, C_{2- 8} alkynyl, Ci_-8 hydroxyalkyl, Ci_-8 haloalkyl, C_3-i₀ cycloalkyl, C_4-i₀ heterocycloakyl, C_3-i₀ cycloalkoxy, Ci_-8 alkoxy, Ci_-8 alkoxyalkyl, C_5-io aryl, and C_5-io heteroaryl moieties is optionally substituted by one or more of CR⁰, CR⁰R⁰, SR⁰, SR⁰R⁰, OR⁰, COR⁰, S(O)_mR°, NR⁰R⁰, CONR⁰R⁰, N(O)R⁰R⁰, (CR^cR⁰)_m(C₅-io aryl), (CR^cR⁰)_m(C₅-io heteroaryl) or (CR°R⁰)_m(C_3-i₀ cycloalkyl); R^c and R^D are independently selected from a bond, H, halo, cyano, nitro, oxo, CF₃, OCF₃, Ci_-8 alkyl, C_2-8 alkenyl, C_2-8 alkynyl, C_5-io aryl, C_5-io heteroaryl, arylalkyl, C_3-i₀ cycloalkyl, C_4-i₀ heterocycloakyl, C_3-i₀ cycloalkoxy, Ci_-8 alkoxy, and C_2-S aI koxyalkyl;

D² is C, N, O, S, CR^C, NR^C, OR^C, SR^C, C-(O)R⁰, N-(O)R⁰, C-(O)OR⁰, N-(O)OR⁰, C-(R⁰R⁰), N-(R⁰R⁰), or S-(R⁰R⁰); and each m is independently selected from O, 1 , 2, 3, 4 and 5; and each n is independently selected from O, 1 , 2, 3, 4 and 5; or a pharmaceutically acceptable salt or stereoisomer thereof.

5) A compound of formula (II) according to claim 4 wherein R⁶ is selected from H, halo, cyano, CF₃, OCF₃, Ci-s alkyl, C_2-8 alkenyl, C_2-8 alkynyl, d-s hydroxyalkyl, Ci_-8 haloalkyl, C_3-io cycloalkyl, C_4-io heterocycloakyl, C_3-io cycloalkoxy, Ci_-8 alkoxy, Ci_-8 alkoxyalkyl, C_5--_I0 aryl, and C_5--_I0 heteroaryl.

6) A compound of formula (II) according to claim 4 wherein R⁷ is selected from H, halo, cyano, CF₃, OCF₃, Ci_-8 alkyl, C_2-8 alkenyl, C_2-8 alkynyl, Ci_-8 hydroxyalkyl, Ci_-8 haloalkyl, C_3-I0 cycloalkyl, C_4-i₀ heterocycloakyl, C_3-I0 cycloalkoxy, Ci_-8 alkoxy, Ci_-8 alkoxyalkyl, C_5-i₀ aryl, and C_5-I0 heteroaryl.

7) A compound of formula (II) according to claim 4 wherein:

R⁹ is H, halo, cyano, nitro, oxo, CF₃, OCF₃, Ci_-8 alkyl, C_2-8 alkenyl, C_2-8 alkynyl, Ci- s hydroxyalkyl, Ci_-8 haloalkyl, C_3-I0 cycloalkyl, C_4-i₀ heterocycloakyl, C_3-I0 cycloalkoxy, Ci- ₈ alkoxy, Ci_-8 alkoxyalkyl, C_5-i₀ aryl, or C_5-I0 heteroaryl; and n is O, 1 , 2 or 3.

8) A compound of formula (II) according to claim 4 wherein R¹⁰ is halo, cyano, Ci_-8 alkyl, C_2-8 alkenyl, C_2-8 alkynyl, Ci_-8 hydroxyalkyl, Ci_-8 haloalkyl, C_3-I0 cycloalkyl, C_4-I0 heterocycloakyl, C_3-I0 cycloalkoxy, Ci_-8 alkoxy, Ci_-8 alkoxyalkyl, C_5-i₀ aryl, Or C_5-I0 heteroaryl.

9) A compound of formula (II) according to claim 4 wherein R¹¹ is Ci_-8 alkyl, C_2-8 alkenyl, C_2-8 alkynyl, Ci_-8 hydroxyalkyl, Ci_-8 haloalkyl, C_3-I0 cycloalkyl, C_4-I0 heterocycloakyl, C_3-I0 cycloalkoxy, Ci_-8 alkoxy or Ci_-8 alkoxyalkyl. 10) A compound selected from the group consisting of:

or a pharmaceutically acceptable salt or stereoisomer thereof.

11 ) A compound of formula (I) or (II) according to any one of claims 1 to 10 for use as a medicament.

12) Use of a compound according to any one of claims 1 to 10 for the preparation of a medicament for treating glaucoma and ocular hypertension. 13) A pharmaceutical composition comprising a pharmaceutically acceptable carrier and a pharmaceutically effective amount of a compound of formula (I) or (II) and/or a salt or stereoisomer thereof as defined in any one of claims 1 to 10.

14) A method for treating glaucoma or ocular hypertension comprising contacting an effective intraocular pressure reducing amount of a pharmaceutical composition according to claim 13 with the eye in order to reduce eye pressure and to maintain said pressure on a reduced level.

15) A pharmaceutical composition comprising a mixture of a compound of formula (I) or (II) and/or a salt or stereoisomer thereof as defined in any one of claims 1 to 10 and:

(i) a beta-blocker; or

(ii) a carbonic anhydrase inhibitor; or

(iii) an adrenergic agonist; or a nitrooxy derivative of (i), (ii) or (iii).