Substituted piperidines as melanocortin receptor agonists

Abstract

Certain novel substituted piperidine compounds are agonists of the human melanocortin receptor(s) and, in particular, are selective agonists of the human melanocortin-4 receptor (MC-4R). They are therefore useful for the treatment, control, or prevention of diseases and disorders responsive to the activation of MC-4R, such as obesity, diabetes, sexual dysfunction, including erectile dysfunction and female sexual dysfunction.

Description

FIELD OF THE INVENTION

[0002] The present invention relates to piperidine derivatives, their synthesis, and their use as melanocortin receptor (MC-R) agonists. More particularly, the compounds of the present invention are selective agonists of the melanocortin-4 receptor (MC-4R) and are thereby useful for the treatment of disorders responsive to the activation of MC-4R, such as obesity, diabetes, male sexual dysfunction, and female sexual dysfunction.

BACKGROUND OF THE INVENTION

[0003] Pro-opiomelanocortin (POMC) derived peptides are known to affect food intake. Several lines of evidence support the notion that the G-protein coupled receptors (GPCRs) of the melanocortin receptor (MC-R) family, several of which are expressed in the brain, are the targets of POMC derived peptides involved in the control of food intake and metabolism. A specific single MC-R that may be targeted for the control of obesity has not yet been identified, although evidence has been presented that MC-4R signalling is important in mediating feed behavior (S. Q. Giraudo et al., “Feeding effects of hypothalamic injection of melanocortin-4 receptor ligands,” Brain Research, 80: 302-306 (1998)).

[0004] Evidence for the involvement of MC-R's in obesity includes: i) the agouti (Avy) mouse which ectopically expresses an antagonist of the MC-1R, MC-3R and -4R is obese, indicating that blocking the action of these three MC-R's can lead to hyperphagia and metabolic disorders; ii) MC-4R knockout mice (D. Huszar et al., Cell, 88: 131-141 (1997)) recapitulate the phenotype of the agouti mouse and these mice are obese; iii) the cyclic heptapeptide MT-II (a non-selective MC-1R, -3R, -4R, and -5R agonist) injected intracerebroventricularly (ICV) in rodents, reduces food intake in several animal feeding models (NPY, ob/ob, agouti, fasted) while ICV injected SHU-9119 (MC-3R and 4R antagonist; MC-1R and -5R agonist) reverses this effect and can induce hyperphagia; iv) chronic intraperitoneal treatment of Zucker fatty rats with an α-NDP-MSH derivative (BP228) has been reported to activate MC-1R, -3R, -4R, and -5R and to attenuate food intake and body weight gain over a 12-week period (I. Corcos et al., “HP228 is a potent agonist of melanocortin receptor-4 and significantly attenuates obesity and diabetes in Zucker fatty rats,” Society for Neuroscience abstracts, 23: 673 (1997)).

[0005] Five distinct MC-R's have thus far been identified, and these are expressed in different tissues. MC-1R was initially characterized by dominant gain of function mutations at the Extension locus, affecting coat color by controlling phaeomelanin to eumelanin conversion through control of tyrosinase. MC-1R is mainly expressed in melanocytes. MC-2R is expressed in the adrenal gland and represents the ACTH receptor. MC-3R is expressed in the brain, gut, and placenta and may be involved in the control of food intake and thermogenesis. MC-4R is uniquely expressed in the brain, and its inactivation was shown to cause obesity (A. Kask, et al., “Selective antagonist for the melanocortin-4 receptor (HS014) increases food intake in free-feeding rats,” Biochem. Biophys. Res. Commun., 245: 90-93 (1998)). MC-5R is expressed in many tissues, including white fat, placenta and exocrine glands. A low level of expression is also observed in the brain. MC-5R knockout mice reveal reduced sebaceous gland lipid production (Chen et al., Cell, 91: 789-798 (1997)).

[0006] Erectile dysfunction denotes the medical condition of inability to achieve penile erection sufficient for successful sexual intercourse. The term impotence is oftentimes employed to describe this prevalent condition. Approximately 140 million men worldwide, and, according to a National Institutes of Health study, about 30 million American men suffer from impotency or erectile dysfunction. It has been estimated that the latter number could rise to 47 million men by the year 2000. Erectile dysfunction can arise from either organic or psychogenic causes, with about 20% of such cases being purely psychogenic in origin. Erectile dysfunction increases from 40% at age 40, to 67% at age 75, with over 75% occurring in men over the age of 50. In spite of the frequent occurrence of this condition, only a small number of patients have received treatment because existing treatment alternatives, such as injection therapies, penile prosthesis implantation, and vacuum pumps, have been uniformly disagreeable [for a discussion, see “ABC of sexual health-erectile dysfunction,” Brit. Med. J. 318: 387-390 (1999)]. Only more recently have more viable treatment modalities become available, in particular orally active agents, such as sildenafil citrate, marketed by Pfizer under the brand name of Viagra®. Sildenafil is a selective inhibitor of type V phosphodiesterase (PDE-V), a cyclic-GMP-specific phosphodiesterase isozyme [see R. B. Moreland et al., “Sildenafil: A Novel Inhibitor of Phosphodiesterase Type 5 in Human Corpus Cavemosum Smooth Muscle Cells,” Life Sci., 62: 309-318 (1998)]. Prior to the introduction of Viagra on the market, less than 10% of patients suffering from erectile dysfunction received treatment. Sildenafil is also being evaluated in the clinic for the treatment of female sexual dysfunction.

[0007] The regulatory approval of Viagra® for the oral treatment of erectile dysfunction has invigorated efforts to discover even more effective methods to treat erectile dysfunction. Several additional selective PDE-V inhibitors are in clinical trials. UK-114542 is a sildenafil backup from Pfizer with supposedly improved properties. IC-351 (ICOS Corp.) is claimed to have greater selectivity for PDE-V over PDE-VI than sildenafil. Other PDE-V inhibitors include M-54033 and M-54018 from Mochida Pharmaceutical Co. and E-4010 from Eisai Co., Ltd.

[0008] Other pharmacological approaches to the treatment of erectile dysfunction have been described [see, e.g., “Latest Findings on the Diagnosis and Treatment of Erectile Dysfunction,” Drug News & Perspectives, 9: 572-575 (1996); “Oral Pharmacotherapy in Erectile Dysfunction,” Current Opinion in Urology, 7: 349-353 (1997)]. A product under clinical development by Zonagen is an oral formulation of the alpha-adrenoceptor antagonist phentolamine mesylate under the brand name of Vasomax®. Vasomax® is also being evaluated for the treatment of female sexual dysfunction.

[0009] Drugs to treat erectile dysfunction act either peripherally or centrally. They are also classified according to whether they initiate a sexual response or facilitate a sexual response to prior stimulation [for a discussion, see “A Therapeutic Taxonomy of Treatments for Erectile Dysfunction: An Evolutionary Imperative,” Int. J. Impotence Res., 9: 115-121(1997)]. While sildenafil and phentolamine act peripherally and are considered to be “enhancers” or “facilitators” of the sexual response to erotic stimulation, sildenafil appears to be efficacious in both mild organic and psychogenic erectile dysfunction. Sildenafil has an onset of action of 30-60 minutes after an oral dose with the effect lasting about 4 hours, whereas phentolamine requires 5-30 minutes for onset with a duration of 2 hours. Although sildenafil is effective in a majority of patients, it takes a relatively long time for the compound to show the desired effects. The faster-acting phentolamine appears to be less effective and to have a shorter duration of action than sildenafil. Oral sildenafil is effective in about 70% of men who take it, whereas an adequate response with phentolamine is observed in only 35-40% of patients. Both compounds require erotic stimulation for efficacy. Since sildenafil indirectly increases blood flow in the systemic circulation by enhancing the smooth muscle relaxation effects of nitric oxide, it is contraindicated for patients with unstable heart conditions or cardiovascular disease, in particular patients taking nitrates, such as nitroglycerin, to treat angina. Other adverse effects associated with the clinical use of sildenafil include headache, flushing, dyspepsia, and “abnormal vision,” the latter the result of inhibition of the type VI phosphodiesterase isozyme (PDE-VI), a cyclic-GMP-specific phosphodiesterase that is concentrated in the retina. “Abnormal vision” is defined as a mild and transient “bluish” tinge to vision, but also an increased sensitivity to light or blurred vision.

[0010] Synthetic melanocortin receptor agonists (melanotropic peptides) have been found to initiate erections in men with psychogenic erectile dysfunction [See H. Wessells et al., “Synthetic Melanotropic Peptide Initiates Erections in Men With Psychogenic Erectile Dysfunction: Double-Blind, Placebo Controlled Crossover Study,” J. Urol., 160: 389-393 (1998); Fifteenth American Peptide Symposium, Jun. 14-19, 1997 (Nashville, Tenn.)]. Activation of melanocortin receptors of the brain appears to cause normal stimulation of sexual arousal. In the above study, the centrally acting α-melanocyte-stimulating hormone analog, melanotan-II (MT-II), exhibited a 75% response rate, similar to results obtained with apomorphine, when injected intramuscularly or subcutaneously to males with psychogenic erectile dysfunction. MT-II is a synthetic cyclic heptapeptide, Ac-Nle-c[Asp-His-DPhe-Arg-Trp-Lys]-NH2, which contains the 4-10 melanocortin receptor binding region common to α-MSH and adrenocorticotropin, but with a lactam bridge. It is a non-selective MC-1R, -3R, -4R, and -5R agonist (Dorr et al., Life Sciences, Vol. 58, 1777-1784, 1996). MT-II (also referred to as PT-14) (Erectide®) is presently in clinical development by Palatin Technologies, Inc. and TheraTech, Inc. as a non-penile subcutaneous injection formulation. It is considered to be an “initiator” of the sexual response. The time to onset of erection with this drug is relatively short (10-20 minutes) with a duration of action approximately 2.5 hours. Adverse reactions observed with MT-II include nausea, flushing, loss of appetite, stretching, and yawning and may be the result of activation of MC-1R, MC-2R, MC-3R, and/or MC-5R. MT-II must be administered parenterally, such as by subcutaneous, intravenous, or intramuscular route, since it is not absorbed into the systemic circulation when given by the oral route.

[0011] MT-II's erectogenic properties apparently are not limited to cases of psychogenic erectile dysfunction in that men with a variety of organic risk factors developed penile erections upon subcutaneous injection of the compound; moreover, the level of sexual desire was significantly higher after MT-II administration than after placebo [see H. Wessells, “Effect of an Alpha-Melanocyte Stimulating Hormone Analog on Penile Erection and Sexual Desire in Men with Organic Erectile Dysfunction,” Urology, 56: 641-646 (2000)].

[0012] Compositions of melanotropic peptides and methods for the treatment of psychogenic erectile dysfunction are disclosed in U.S. Pat. No. 5,576,290, assigned to Competitive Technologies. Methods of stimulating sexual response in females using melanotropic peptides have been disclosed in U.S. Pat. No. 6,051,555.

[0013] Spiropiperidine and piperidine derivatives have been disclosed in WO 99/64002 (Dec. 16, 1999) and WO 00/74679 (Dec. 14, 2000), respectively, as agonists of the melanocortin receptor(s) and thereby useful for the treatment of diseases and disorders, such as obesity, diabetes, and sexual dysfunction, including erectile dysfunction and female sexual dysfunction.

[0014] Because of the unresolved deficiencies of the various pharmacological agents discussed above, there is a continuing need in the medical arts for improved methods and compositions to treat individuals suffering from psychogenic and/or organic erectile dysfunction. Such methods and compositions should have wider applicability, enhanced convenience and ease of compliance, short onset of action, reasonably long duration of action, and minimal side effects with few contraindications, as compared to agents now available.

[0015] It is therefore an object of the present invention to provide novel piperidine derivatives which are useful as melanocortin receptor agonists and thereby useful to treat obesity, diabetes, and male and female sexual dysfunction.

[0016] It is another object of the present invention to provide novel piperidine derivatives which are selective agonists of the melanocortin-4 (MC-4R) receptor.

[0017] It is another object of the present invention to provide pharmaceutical compositions comprising melanocortin receptor agonists of the present invention with a pharmaceutically acceptable carrier.

[0018] It is another object of the present invention to provide methods for the treatment or prevention of disorders, diseases, or conditions responsive to the activation of the melanocortin receptor in a subject in need thereof by administering the compounds and pharmaceutical compositions of the present invention.

[0019] It is another object of the present invention to provide methods for the treatment or prevention of obesity, diabetes mellitus, male sexual dysfunction, and female sexual dysfunction by administering the compounds and pharmaceutical compositions of the present invention to a subject in need thereof.

[0020] It is another object of the present invention to provide methods for the treatment of erectile dysfunction by administering the compounds and pharmaceutical compositions of the present invention to a subject in need thereof.

[0021] These and other objects will become readily apparent from the detailed description that follows.

SUMMARY OF THE INVENTION

[0022] The present invention relates to novel substituted piperidines of structural formula (I): 1

[0023] These piperidine derivatives are effective as melanocortin receptor agonists and are particularly effective as selective melanocortin-4 receptor (MC-4R) agonists. They are therefore useful for the treatment and/or prevention of disorders responsive to the activation of MC-4R, such as obesity, diabetes as well as male and female sexual dysfunction, in particular, male erectile dysfunction.

[0024] The present invention also relates to pharmaceutical compositions comprising the compounds of the present invention and a pharmaceutically acceptable carrier.

[0025] The present invention also relates to methods for the treatment or prevention of disorders, diseases, or conditions responsive to the activation of the melanocortin receptor in a mammal in need thereof by administering the compounds and pharmaceutical compositions of the present invention.

[0026] The present invention also relates to methods for the treatment or prevention of obesity, diabetes mellitus, male sexual dysfunction, and female sexual dysfunction by administering the compounds and pharmaceutical compositions of the present invention.

[0027] The present invention also relates to methods for treating erectile dysfunction by administering the compounds and pharmaceutical compositions of the present invention.

[0028] The present invention also relates to methods for treating erectile dysfunction by administering the compounds of the present invention in combination with a therapeutically effective amount of another agent known to be useful to treat the condition.

[0029] The present invention also relates to methods for treating or preventing obesity by administering the compounds of the present invention in combination with a therapeutically effective amount of another agent known to be useful to treat the condition.

DETAILED DESCRIPTION OF THE INVENTION

[0030] The present invention relates to substituted piperidines useful as melanocortin receptor agonists. Representative compounds of the present invention are described by structural formula (I): 2

[0031] or a pharmaceutically acceptable salt thereof;

[0032] wherein Q is 3

[0033] Z is O, S, or NR4b;

[0034] each p is independently 1 or 2;

[0035] each n is independently 0, 1, or 2;

[0036] R1 is selected from the group consisting of

[0037] hydrogen,

[0038] C1-8 alkyl,

[0039] (CHR7)n—C3-6 cycloalkyl,

[0040] (CHR7)n—O(CHR7)aryl,

[0041] (CHR7)n-aryl, and

[0042] (CHR7)n-heteroaryl;

[0043]  in which aryl and heteroaryl are unsubstituted or substituted with one to three groups independently selected from R6; and alkyl and cycloalkyl are unsubstituted or substituted with one to three groups independently selected from R6 and oxo;

[0044] R2 is selected from the group consisting of

[0045] hydrogen,

[0046] C1-8 alkyl,

[0047] (CH2)nC3-6 cycloalkyl, and

[0048] (CH2)n-aryl;

[0049] each R3 is independently selected from the group consisting of

[0050] hydrogen,

[0051] C1-8 alkyl,

[0052] (CH2)n-aryl,

[0053] (CH2)nC3-6 cycloalkyl,

[0054] (CH2)n-heteroaryl, and

[0055] (CH2)n-heterocyclyl;

[0056]  in which aryl and heteroaryl are unsubstituted or substituted with one to three groups independently selected from R6; and alkyl, cycloalkyl, and heterocyclyl are unsubstituted or substituted with one to three groups independently selected from R6 and oxo;

[0057] or R3 and R5a and the carbons to which they are attached form a 5- to 7-membered ring optionally containing an additional heteroatom selected from O, S, and NR7;

[0058] R4a and R4b are each independently selected from the group consisting of

[0059] hydrogen,

[0060] C1-8 alkyl,

[0061] (CH2)n-aryl,

[0062] (CH2)nC3-6 cycloalkyl,

[0063] (CH2)n-heteroaryl,

[0064] (CH2)n-heterocyclyl,

[0065] COC(R7)2NH2,

[0066] COR7,

[0067] (CH2)nOR7,

[0068] (CH2)nCO2R7,

[0069] CH2C≡CH,

[0070] CO2R7,

[0071] CH2CHF2,

[0072] CONR7R7, and

[0073] SO2R7;

[0074]  in which aryl and heteroaryl are unsubstituted or substituted with one to three groups independently selected from R6; and alkyl, cycloalkyl, and heterocyclyl are unsubstituted or substituted with one to three groups independently selected from R6 and oxo;

[0075] or R4a and R2 and the carbons to which they are attached form a 5- to 7-membered ring optionally containing an additional heteroatom selected from O, S, and NR7; or

[0076] R4a and R4b and the atoms to which they are attached form a 5- to 7-membered ring;

[0077] R5a and R5b are each independently selected from the group consisting of

[0078] hydrogen,

[0079] C1-8 alkyl,

[0080] (CH2)n-aryl, and

[0081] C3-8 cycloalkyl;

[0082]  wherein alkyl and cycloalkyl are unsubstituted or substituted with one to three groups independently selected from R6 and oxo; aryl is unsubstituted or substituted with one to three groups independently selected from R6;

[0083] or R5a and R5b together with the carbons to which they are attached form a 5- to 7-membered ring;

[0084] R6 is selected from the group consisting of

[0085] hydrogen,

[0086] C1-8 alkyl,

[0087] (CH2)n-aryl,

[0088] (CH2)nC3-7 cycloalkyl,

[0089] (CH2)n-heteroaryl,

[0090] halogen,

[0091] OR7,

[0092] NHSO2R7,

[0093] N(R7)2,

[0094] C≡N,

[0095] CO2R7,

[0096] C(R7)(R7)N(R7)2,

[0097] NO2,

[0098] SO2N(R7)2,

[0099] S(O)0-2R7,

[0100] CF3, and

[0101] OCF3;

[0102]  or two R6 substituents, when on the same carbon atom, can be taken together with the carbon atom to which they are attached to form a cyclopropyl group;

[0103] each R7 is independently selected from the group consisting of

[0104] hydrogen,

[0105] C1-8 alkyl,

[0106] (CH2)n-aryl, and

[0107] (CH2)nC3-7 cycloalkyl;

[0108] each R8 is independently selected from the group consisting of

[0109] hydrogen,

[0110] C1-8 alkyl,

[0111] (CH2)n-aryl,

[0112] (CH2)n-heteroaryl,

[0113] (CH2)n-heterocyclyl, and

[0114] (CH2)nC3-7 cycloalkyl;

[0115]  wherein aryl and heteroaryl are unsubstituted or substituted with one to three groups independently selected from R6; and alkyl, cycloalkyl, heterocyclyl, and (CH2)n are unsubstituted or substituted with one to three groups independently selected from R6 and oxo;

[0116]  or two R8 groups together with the atoms to which they are attached form a 5- to 8-membered mono- or bi-cyclic ring system optionally containing an additional heteroatom selected from O, S, NR7, NBoc, and NCbz;

[0117] each R9 is independently selected from the group consisting of

[0118] hydrogen,

[0119] C1-8 alkyl,

[0120] (CH2)n-aryl,

[0121] (CH2)nC3-6 cycloalkyl,

[0122] (CH2)n-heteroaryl,

[0123] halogen,

[0124] OR7,

[0125] NHSO2R7,

[0126] N(R7)2,

[0127] C≡N,

[0128] CO2R7,

[0129] C(R7)(R7)N(R7)2,

[0130] NO2,

[0131] SO2N(R7)2,

[0132] S(O)0-2R7,

[0133] CF3, and

[0134] OCF3;

[0135] X is selected from the group consisting of

[0136] C1-8 alkyl,

[0137] (CH2)nC3-8 cycloalkyl,

[0138] (CH2)naryl,

[0139] (CH2)nheteroaryl,

[0140] (CH2)nheterocyclyl,

[0141] (CH2)nC≡N,

[0142] (CH2)nCONR8R8,

[0143] (CH2)nCO2R8,

[0144] (CH2)nCOR8

[0145] (CH2)nNR8C(O)R8,

[0146] (CH2)nNR8CO2R8,

[0147] (CH2)nNR8C(O)N(R8)2,

[0148] (CH2)nNR8SO2R8,

[0149] (CH2)nS(O)0-2R8,

[0150] (CH2)nSO2N(R8)(R8),

[0151] (CH2)nOR8,

[0152] (CH2)nOC(O)R8,

[0153] (CH2)nOC(O)OR8,

[0154] (CH2)nOC(O)N(R8)2,

[0155] (CH2)nN(R8)(R8), and

[0156] (CH2)nNR8SO2N(R8)(R8);

[0157]  wherein aryl and heteroaryl are unsubstituted or substituted with one to three groups selected from R6; and alkyl, (CH2)n, cycloalkyl, and heterocyclyl are unsubstituted or substituted with one to three groups independently selected from R6 and oxo;

[0158] Y is selected from the group consisting of

[0159] hydrogen,

[0160] C1-8 alkyl,

[0161] (CH2)nC3-8 cycloalkyl,

[0162] (CH2)naryl,

[0163] (CH2)nheterocyclyl, and

[0164] (CH2)nheteroaryl;

[0165]  wherein aryl and heteroaryl are unsubstituted or substituted with one to three groups selected from R6; and alkyl, (CH2)n, cycloalkyl, and heterocyclyl are optionally substituted with one to three groups selected from R6 and oxo.

[0166] In one embodiment of the compounds of formula I, Q is 4

[0167] wherein Z is O or NR4b; and R2, R3, R4a, R4b, R5a, R5b, and R9 are as defined above.

[0168] In a second embodiment of the compounds of formula I, Q is 5

[0169] wherein R2, R3, R4a, R4b, R5a, and R5b are as defined above.

[0170] In a class of this second embodiment, R4a and R4b are each independently selected from the group consisting of

[0171] hydrogen,

[0172] C1-8 alkyl,

[0173] (CH2)n-aryl,

[0174] (CH2)n-heteroaryl,

[0175] (CH2)n-heterocyclyl,

[0176] (CH2)nC3-6 cycloalkyl,

[0177] (CH2)nCO2R7

[0178] (CH2)nOR7,

[0179] COC(R7)NH2,

[0180] CH2C≡CH, and

[0181] CH2CHF2;

[0182] or R4a and R4b and the atoms to which they are attached form a 6-membered ring;

[0183] R3, R5a, and R5b are each independently hydrogen, C1-4 alkyl, C3-6 cycloalkyl, or aryl; wherein aryl is unsubstituted or substituted with one to three groups independently selected from R6; or R3 and R5a and the carbons to which they are attached form a 6-membered ring optionally containing an additional heteroatom selected from O, S, and NR7.

[0184] In a subclass of this class, R4a and R4b are each independently selected from the group consisting of

[0185] hydrogen,

[0186] C1-4 alkyl,

[0187] CH2-aryl,

[0188] CH2-heteroaryl,

[0189] CH2-heterocyclyl,

[0190] (CH2)0-1 C3-6 cycloalkyl,

[0191] CH2CO2R7

[0192] (CH2)2OR7,

[0193] COC(R7)NH2,

[0194] CH2C≡CH, and

[0195] CH2CHF2;

[0196] or R4a and R4b and the atoms to which they are attached form a 6-membered ring;

[0197] R3, R5a, and R5b are each independently hydrogen, C1-4 alkyl, C3-6 cycloalkyl, or phenyl; wherein phenyl is unsubstituted or substituted with one to three groups independently selected from R6; or R3 and R5a and the carbons to which they are attached form a 6-membered ring optionally containing an additional heteroatom selected from O, S, and NR7.

[0198] In a third embodiment of the compounds of formula I, R1 is CHR7-aryl, CHR7OCHR7-aryl, or CHR7-heteroaryl wherein aryl and heteroaryl are optionally substituted with one or two R6 groups. In a class of this embodiment, R1 is benzyl optionally substituted with one or two groups selected from halogen, C1-4 alkyl, C1-4 alkoxy, CN, CF3, and OCF3. In a subclass of this class, R1 is 4-chlorobenzyl; 4-fluorobenzyl; 3,4-difluorobenzyl; 3,5-difluorobenzyl; 2-cyano-4-fluorobenzyl; or 4-methoxybenzyl.

[0199] In a fourth embodiment of compounds of formula I, R2 is H or CH3.

[0200] In a fifth embodiment of compounds of formula I, X is C1-6 alkyl, (CH2)n-aryl, (CH2)n-heteroaryl, (CH2)n-heterocyclyl, (CH2)nC(O)N(R8)(R8), (CH2)nCO2R8, (CH2)nOR8, (CH2)nS(O)0-2R8, (CH2)nNHC(O)R8, (CH2)nOC(O)NR8R8, or (CH2)nNR8SO2R8; wherein aryl and heteroaryl are optionally substituted with one to three groups selected from R6; heterocyclyl is optionally substituted with one to three groups selected from R6 and oxo; the (CH2)n group is optionally substituted with one to three groups selected from R7, halogen, S(O)0-2R7, N(R7)2, and OR7; and R8 is each independently selected from H, C1-8 alkyl, and C3-6 cycloalkyl optionally substituted with one to three groups selected from R6 and oxo; or two R8 groups together with the atoms to which they are attached form a 5- to 8-membered mono- or bi-cyclic ring system optionally containing an additional heteroatom selected from O, S, NR7, NBoc, and NCbz.

[0201] In a class of this embodiment, X is C1-6 alkyl, (CH2)0-1-heteroaryl, CH2-heterocyclyl, CO2R8, CH2OR8, CH2S(O)0-2R8, NHC(O)R8, CH2NR8SO2R8, CH2OC(O)NR8R8, CH2NR8SO2R8, or C(O)N(R8)(R8); wherein heteroaryl is optionally substituted with one to three groups selected from R6; heterocyclyl is optionally substituted with one to three groups selected from R6 and oxo; and R8 is each independently selected from H, C1-8 alkyl, and C3-6 cycloalkyl optionally substituted with one to three groups selected from R6 and oxo; or two R8 groups together with the atoms to which they are attached form a 5- to 8-membered mono- or bi-cyclic ring system optionally containing an additional heteroatom selected from O, S, NR7, NBoc, and NCbz.

[0202] In a sixth embodiment of compounds of formula I, Y is C1-8 alkyl, (CH2)nC3-7 cycloalkyl, (CH2)n-aryl, (CH2)n-heterocyclyl, or (CH2)n-heteroaryl; wherein aryl and heteroaryl are optionally substituted with one to three groups selected from R6; and (CH2)n, alkyl, cycloalkyl, and heterocyclyl are optionally substituted with one to three groups selected from R6 and oxo. In a class of this embodiment, Y is cyclohexyl, cycloheptyl, cyclopentyl, or C1-6 alkyl, unsubstituted or substituted with one to three groups selected from R6 and oxo. In a subclass of this class, Y is cyclohexyl or C1-6 alkyl, wherein the cyclohexyl and alkyl groups are unsubstituted or substituted with one to three groups selected from R6 and oxo.

[0203] In yet a further embodiment of compounds of formula I, the carbon atom marked with * has the R configuration.

[0204] In yet a further embodiment of compounds of formula I, X is selected from the group consisting of: 67

[0205] —NH—C(O)CH3 —C(O)N(CH3)2 —C(O)NH-t-Bu —NHC(O)tBu; —C(O)NHCH(Et)2; —C(O)NHCH2tBu; —CH2SCH(CH3)2; —CH2S(O)CH(CH3)2; —CH2S(O)2CH(CH3)2; —C(O)NHCH2CH2N(CH3)2; C(O)CH(CH3)2; —CH2NHCOtBu; —CH2OC(O)NMe2; —CH2C(O)NEt2; —CH2OC(Me)2CO2H; —C(O)NHC(Me)2CO2Me; —C(O)NHC(Me)2CO2—H; —CH2N(CH3)COtBu; —CH2N(iPr)COMe; —CH2N(iPr)SO2Me; C(O)NHC(Me)2CH2OMe; C(O)NHC(Me)2CH2OH; —CH2CH2C(Me)2CH; 8

[0206] Representative compounds of the present invention of structural formula Ia or Ib with the indicated stereochemistry at the stereogenic center marked with ** are as follows:

1a
9
Y	X	R6	**	R4a	R4b
10	11	Cl	(R)	H	CH3
12	13	Cl	(S)	H	H
14	15	F	(R)	H	H
16	17	F	(R)	H	H
18	19	F	(S)	H	H
20	21	Cl	(S)	H	H
22	23	Cl	(S)	H	H
24	25	Cl	(R)	H	H
26	27	Cl	(S)	H	H
28	29	Cl	(R)	H	H
30	31	Cl	(R)	H	H
32	33	Cl	(S)	H	H
34	35	Cl	(S)	H	H
36	37	F	(S)	H	H
38	39	F	(S)	H	H
40	41	Cl	(S)	H	H
42	43	Cl	(R)	H	H
44	45	F	(S)	H	H
46	47	F	(R)	H	H
48	49	Cl	(S)	H	H
50	51	Cl	(S)	CH3	H
52	53	F	(R)	CH3	H
54	55	Cl	(S)	CH3	H
56	57	F	(S)	CH3	H
58	59	Cl	(R)	CH3	H
60	61	F	(R)	CH3	H
62	63	Cl	(S)	CH3	H
64	65	Cl	(S)	CH3	H
66	67	Cl	(R)	CH3	H
68	69	Cl	(S)	CH3	H
70	71	Cl	(R)	CH3	H
72	73	Cl	(S)	CH3	H
74	75	Cl	(R)	CH3	H
76	77	Cl	(R)	CH3	H
78	79	Cl	(S)	CH3	H
80	81	Cl	(S)	CH3	H
82	83	F	(S)	CH3	H
84	85	F	(S)	CH3	H
86	87	Cl	(S)	CH3	H
88	89	Cl	(R)	CH3	H
90	91	Cl	(S)	CH3	H
92	93	F	(S)	CH3	H
94	95	Cl	(R)	CH3	H
96	97	F	(R)	CH3	H
98	99	Cl	(S)	CH3	H
100	101	Cl	(S)	CH3	CH3
102	103	Cl	(S)	CH3	CH3
104	105	Cl	(S)	CH3	CH3
106	107	Cl	(R)	CH3	CH3
108	109	Cl	(R)	CH3	CH3
110	111	Cl	(R)	CH3	CH3
112	113	Cl	(S)	CH3	CH3
114	115	Cl	(S)	H	CH3
116	117	Cl	(S)	H	i-Pr
118	119	F	(S)	H	2-hydroxyethyl
120	121	F	(S)	H	2-methoxyethyl
122	123	F	(S)	H	CH2CO2Et
124	125	F	(R)	H	CH3
126	127	F	(R)	H	2-methoxyethyl
128	129	Cl	(S)	H	CH3
130	131	Cl	(S)	H	i-Pr
132	133	Cl	(S)	H	2,2- difluoroethyl
134	135	F	(S)	H	CH3
136	137	F	(S)	H	i-Pr
138	139	F	(S)	H	2,2- difluoroethyl
140	141	Cl	(R)	H	CH3
142	143	Cl	(R)	H	i-Pr
144	145	Cl	(R)	H	cyclopropyl- methyl
146	147	Cl	(R)	H	benzyl
148	149	Cl	(R)	H	2-propynyl
150	151	Cl	(R)	H	cyclobutyl
152	153	Cl	(R)	H	2,2- difluoroethyl
154	155	F	(R)	H	CH3
156	157	F	(R)	H	i-Pr
158	159	F	(R)	H	cyclopropyl- methyl
160	161	F	(R)	H	2,2- difluoroethyl
162	163	Cl	(S)	H	CH3
164	165	Cl	(S)	H	CH3
166	167	Cl	(R)	H	CH3
168	169	Cl	(S)	H	CH3
170	171	Cl	(R)	H	CH3
172	173	Cl	(S)	H	CH3
174	175	Cl	(R)	H	CH3
176	177	Cl	(R)	H	CH3
178	179	Cl	(S)	H	CH3
180	181	F	(S)	H	CH3
182	183	Cl	(S)	H	CH3
184	185	Cl	(R)	H	CH3
186	187	Cl	(S)	H	CH3
188	189	Cl	(R)	H	CH3
190	191	F	(S)	H	CH3
192	193	F	(R)	H	CH3
194	195	F	(R)	H	i-Pr
196	197	F	(S)	H	198
199	200	Cl	(S)	CH3	i-Pr
201	202	Cl	(S)	CH3	i-Pr
203	204	F	(S)	H	Et
205	206	F	(R)	H	H
207	208	F	(S)	H	H
209	210	F	(R)	H	Me
211	212	F	(S)	H	Me
213	214	F	(S)	H	Me
215	216	F	(S)	H	Me
217	218	F	(S)	H	Me
219	220	F	(S)	H	Me
221	222	F	(S)	H	Me
223	224	F	(S)	H	Me
225	226	F	(S)	H	Me
227	228	F	(S)	H	Me
229	230	F	(S)	H	Me
231	232	F	(S)	H	H
233	234	F	(S)	H	Me
235	236	F	(S)	H	Me
237	238	F	(S)	H	H
239	240	F	(S)	H	Me
241	242	F	(S)	H	Me
243	244	F	(S)	H	H
245	246	Cl	(R)	H	Me
247	248	Cl	(S)	H	H
249	250	F	(S)	iPr	Me
251	252	F	(S)	H	Me

[0207]

Ib
253
						Diaste-
R6	**	X	R3	R4a	R4b	reomer
4-fluoro	(R)	254	255	H	H	D1 + D2
4-fluoro	(R)	256	257	H	H	D1
4-fluoro	(R)	258	259	H	H	D2
4-chloro	(R)	260	261	H	H	D1 + D2
4-chloro	(R)	262	263	H	H	D1
4-chloro	(R)	264	265	H	H	D2
4-fluoro	(R)	266	267	H	H	D1 + D2
4-fluoro	(R)	268	269	H	H	D1 + D2
4-fluoro	(R)	270	271	H	H	D1 + D2
4-fluoro	(R)	272	273	H	H
4-chloro	(R)	274	Ph	H	H	D1 + D2
4-fluoro	(R)	275	Ph	H	H	D1 + D2
4-fluoro	(R)	276	Ph	H	H	D1 + D2
4-fluoro	(S)	277	278	H	H	D1 + D2
4-fluoro	(S)	279	280	H	H	D1
4-fluoro	(S)	281	282	H	H	D2
4-fluoro	(S)	283	284	H	H	D1 + D2
4-fluoro	(S)	285	286	H	H	D1
4-fluoro	(S)	287	288	H	H	D2
4-fluoro	(R)	289	290	H	H	D1 + D2
4-fluoro	(R)	291	292	H	H	D1
4-fluoro	(R)	293	294	H	H	D2
4-fluoro	(S)	295	296	H	H	D1 + D2
4-fluoro	(S)	297	298	H	H	D1
4-fluoro	(S)	299	300	H	H	D2
4-fluoro	(R)	301	302	H	H	D1 + D2
4-fluoro	(R)	303	304	H	H	D1
4-fluoro	(R)	305	306	H	H	D2
4-fluoro	(S)	307	308	H	H	D1 + D2
4-fluoro	(S)	309	310	H	H	D1
4-fluoro	(S)	311	312	H	H	D2
4-fluoro	(R)	313	314	H	H	D1 + D2
4-fluoro	(R)	315	316	H	H	D1
4-fluoro	(R)	317	318	H	H	D2
4-chloro	(R)	319	320	H	H	D1 + D2
4-chloro	(R)	321	322	H	H	D1
4-chloro	(R)	323	324	H	H	D2
4-chloro	(S)	325	Ph	H	H	D1 + D2
4-fluoro	(S)	326	Ph	H	H	D1 + D2
4-fluoro	(S)	327	Ph	H	H	D1 + D2
4-chloro	(S)	328	Ph	Me	Me	D1
4-chloro	(S)	329	Ph	Me	Me	D2
4-fluoro	(S)	330	Ph	Me	Me	D1
4-fluoro	(S)	331	Ph	Me	Me	D2
4-fluoro	(S)	332	Ph	Me	Me	D1
4-chloro	(R)	333	Ph	Me	Me	D1
4-chloro	(R)	334	Ph	Me	Me	D2
4-fluoro	(R)	335	Ph	Me	Me	D1
4-fluoro	(R)	336	Ph	Me	Me	D2
4-fluoro	(R)	337	Ph	Me	Me	D1
3,4-difluoro	(S)	338	H	H	H
3,4-difluoro	(S)	339	H	H	Me
3,4-difluoro	(S)	340	iPr	H	H	D1 + D2
3,4-difluoro	(S)	341	iPr	H	H	D1
3,4-difluoro	(S)	342	iPr	H	H	D2
3,5-difluoro	(S)	343	H	H	Me
3,5-difluoro	(S)	344	iPr	H	H
4-fluoro	(S)	345	iPr	Me	H	D1 + D2
2-cyano-4- fluoro	(S)	346	H	H	Me

[0208] Further illustrative but nonlimiting examples of compounds of the present invention that are useful as melanocortin receptor agonists are the following: 347

[0209] or a pharmaceutically acceptable salt thereof.

[0210] Even further illustrative of the compounds of the present invention are those of structural formula Ic with the indicated stereochemistry at the stereogenic center marked with ** selected from the group consisting of:

Ic
348
Y	X	R6	**	R3	R4b
349	350	F	(S)	H	CH3
351	352	F	(S)	H	CH3
353	354	F	(S)	H	CH3
355	356	F	(R)	H	CH3
357	358	F	(S)	H	CH3
359	360	Cl	(R)	H	CH3
361	362	F	(S)	363	H
364	365	F	(R)	366	H
367	368	F	(S)	369	H

[0211] The compounds of structural Formula I are effective as melanocortin receptor agonists and are particularly effective as selective agonists of the MC-4R. They are therefore useful for the treatment and/or prevention of disorders responsive to the activation of MC-4R, such as obesity, diabetes as well as male and/or female sexual dysfunction, in particular, erectile dysfunction, and further in particular, male erectile dysfunction.

[0212] Another aspect of the present invention provides a method for the treatment or prevention of obesity or diabetes in a mammal which comprises administering to said mammal an effective amount of a compound of formula I.

[0213] Another aspect of the present invention provides a method for the treatment or prevention of male or female sexual dysfunction including erectile dysfunction which comprises administering to a patient in need of such treatment or prevention an effective amount of a compound of formula I.

[0214] Yet another aspect of the present invention provides a pharmaceutical composition comprising a compound of formula I and a pharmaceutically acceptable carrier.

[0215] Throughout the instant application, the following terms have the indicated meanings:

[0216] The alkyl groups specified above are intended to include those alkyl groups of the designated length in either a straight or branched configuration. Exemplary of such alkyl groups are methyl, ethyl, propyl, isopropyl, butyl, sec-butyl, tertiary butyl, pentyl, isopentyl, hexyl, isohexyl, and the like.

[0217] The term “halogen” is intended to include the halogen atoms fluorine, chlorine, bromine and iodine.

[0218] The term “aryl” includes phenyl and naphthyl.

[0219] The term “heteroaryl” includes mono- and bicyclic aromatic rings containing from 1 to 4 heteroatoms selected from nitrogen, oxygen and sulfur. “5- or 6-membered heteroaryl” are monocyclic heteroaromatic rings, examples thereof include thiazole, oxazole, thiophene, furan, pyrrole, imidazole, isoxazole, pyrazole, triazole, thiadiazole, tetrazole, oxadiazole, pyridine, pyridazine, pyrimidine, pyrazine, and the like. Bicyclic heteroaromatic rings include, but are not limited to, benzothiadiazole, indole, benzothiophene, benzofuran, benzimidazole, benzisoxazole, benzothiazole, quinoline, benzotriazole, benzoxazole, isoquinoline, purine, furopyridine and thienopyridine.

[0220] The term “5- or 6-membered carbocyclyl” is intended to include non-aromatic rings containing only carbon atoms such as cyclopentyl and cyclohexyl.

[0221] The term “5 and 6-membered heterocyclyl” is intended to include non-aromatic heterocycles containing one to four heteroatoms selected from nitrogen, oxygen and sulfur. Examples of a 5 or 6-membered heterocyclyl include piperidine, morpholine, thiamorpholine, pyrrolidine, imidazolidine, tetrahydrofuran, piperazine, and the like.

[0222] Certain of the above defined terms may occur more than once in the above formula and upon such occurrence each term shall be defined independently of the other; thus for example, NR7R7 may represent NH2, NHCH3, N(CH3)CH2CH3, and the like.

[0223] The term “composition”, as in pharmaceutical composition, is intended to encompass a product comprising the active ingredient(s), and the inert ingredient(s) that make up the carrier, as well as any product which results, directly or indirectly, from combination, complexation or aggregation of any two or more of the ingredients, or from dissociation of one or more of the ingredients, or from other types of reactions or interactions of one or more of the ingredients. Accordingly, the pharmaceutical compositions of the present invention encompass any composition made by admixing a compound of the present invention and a pharmaceutically acceptable carrier.

[0224] “Erectile dysfunction” is a disorder involving the failure of a male mammal to achieve erection, ejaculation, or both. Symptoms of erectile dysfunction include an inability to achieve or maintain an erection, ejaculatory failure, premature ejaculation, or inability to achieve an orgasm. An increase in erectile dysfunction is often associated with age and is generally caused by a physical disease or as a side-effect of drug treatment.

[0225] By a melanocortin receptor “agonist” is meant an endogenous or drug substance or compound that can interact with a melanocortin receptor and initiate a pharmacological response characteristic of the melanocortin receptor. By a melanocortin receptor “antagonist” is meant a drug or a compound that opposes the melanocortin receptor-associated responses normally induced by another bioactive agent. The “agonistic” properties of the compounds of the present invention were measured in the functional assay described below. The functional assay discriminates a melanocortin receptor agonist from a melanocortin receptor antagonist.

[0226] By “binding affinity” is meant the ability of a compound/drug to bind to its biological target, in the the present instance, the ability of a compound of formula I to bind to a melanocortin receptor. Binding affinities for the compounds of the present invention were measured in the binding assay described below and are expressed as IC50's.

[0227] “Efficacy” describes the relative intensity with which agonists vary in the response they produce even when they occupy the same number of receptors and with the same affinity. Efficacy is the property that enables drugs to produce responses. Properties of compounds/drugs can be categorized into two groups, those which cause them to associate with the receptors (binding affinity) and those that produce a stimulus (efficacy). The term “efficacy” is used to characterize the level of maximal responses induced by agonists. Not all agonists of a receptor are capable of inducing identical levels of maximal responses. Maximal response depends on the efficiency of receptor coupling, that is, from the cascade of events, which, from the binding of the drug to the receptor, leads to the desired biological effect.

[0228] The functional activities expressed as EC50's and the “agonist efficacy” for the compounds of the present invention at a particular concentration were measured in the functional assay described below.

[0229] Optical Isomers-Diastereomers-Geometric Isomers-Tautomers

[0230] Compounds of Formula I contain one or more asymmetric centers and can thus occur as racemates and racemic mixtures, single enantiomers, diastereomeric mixtures and individual diastereomers. The present invention is meant to comprehend all such isomeric forms of the compounds of Formula I.

[0231] Some of the compounds described herein contain olefinic double bonds, and unless specified otherwise, are meant to include both E and Z geometric isomers.

[0232] Some of the compounds described herein may exist as tautomers such as keto-enol tautomers. The individual tautomers as well as mixtures thereof are encompassed with compounds of Formula I.

[0233] Compounds of the Formula I may be separated into their individual diastereoisomers by, for example, fractional crystallization from a suitable solvent, for example methanol or ethyl acetate or a mixture thereof, or via chiral chromatography using an optically active stationary phase.

[0234] Alternatively, any diastereomer of a compound of the general Formula I or Ia may be obtained by stereospecific synthesis using optically pure starting materials or reagents of known configuration.

[0235] Salts

[0236] The term “pharmaceutically acceptable salts” refers to salts prepared from pharmaceutically acceptable non-toxic bases or acids including inorganic or organic bases and inorganic or organic acids. Salts derived from inorganic bases include aluminum, ammonium, calcium, copper, ferric, ferrous, lithium, magnesium, manganic salts, manganous, potassium, sodium, zinc, and the like. Particularly preferred are the ammonium, calcium, lithium, magnesium, potassium, and sodium salts. Salts derived from pharmaceutically acceptable organic non-toxic bases include salts of primary, secondary, and tertiary amines, substituted amines including naturally occurring substituted amines, cyclic amines, and basic ion exchange resins, such as arginine, betaine, caffeine, choline, N,N′-dibenzylethylenediamine, diethylamine, 2-diethylaminoethanol, 2-dimethylaminoethanol, ethanolamine, ethylenediamine, N-ethyl-morpholine, N-ethylpiperidine, glucamine, glucosamine, histidine, hydrabamine, isopropylamine, lysine, methylglucamine, morpholine, piperazine, piperidine, polyamine resins, procaine, purines, theobromine, triethylamine, trimethylamine, tripropylamine, tromethamine, and the like.

[0237] When the compound of the present invention is basic, salts may be prepared from pharmaceutically acceptable non-toxic acids, including inorganic and organic acids. Such acids include acetic, benzenesulfonic, benzoic, camphorsulfonic, citric, ethanesulfonic, formic, fumaric, gluconic, glutamic, hydrobromic, hydrochloric, isethionic, lactic, maleic, malic, mandelic, methanesulfonic, malonic, mucic, nitric, pamoic, pantothenic, phosphoric, propionic, succinic, sulfuric, tartaric, p-toluenesulfonic acid, trifluoroacetic acid, and the like. Particularly preferred are citric, fumaric, hydrobromic, hydrochloric, maleic, phosphoric, sulfuric, and tartaric acids.

[0238] It will be understood that, as used herein, references to the compounds of Formula I are meant to also include the pharmaceutically acceptable salts.

[0239] Utility

[0240] Compounds of formula I are melanocortin receptor agonists and as such are useful in the treatment, control or prevention of diseases, disorders or conditions responsive to the activation of one or more of the melanocortin receptors including, but are not limited to, MC-1, MC-2, MC-3, MC-4, or MC-5. Such diseases, disorders or conditions include, but are not limited to, obesity (by reducing appetite, increasing metabolic rate, reducing fat intake or reducing carbohydrate craving), diabetes mellitus (by enhancing glucose tolerance, decreasing insulin resistance), hypertension, hyperlipidemia, osteoarthritis, cancer, gall bladder disease, sleep apnea, depression, anxiety, compulsion, neuroses, insomnia/sleep disorder, substance abuse, pain, male and female sexual dysfunction (including impotence, loss of libido and erectile dysfunction), fever, inflammation, immunemodulation, rheumatoid arthritis, skin tanning, acne and other skin disorders, neuroprotective and cognitive and memory enhancement including the treatment of Alzheimer's disease. Some compounds encompassed by formula I show highly selective affinity for the melanocortin-4 receptor relative to MC-1R, MC-2R, MC-3R, and MC-5R, which makes them especially useful in the prevention and treatment of obesity, as well as male and/or female sexual dysfunction, including erectile dysfunction.

[0241] “Male sexual dysfunction” includes impotence, loss of libido, and erectile dysfunction.

[0242] “Erectile dysfunction” is a disorder involving the failure of a male mammal to achieve erection, ejaculation, or both. Symptoms of erectile dysfunction include an inability to achieve or maintain an erection, ejaculatory failure, premature ejaculation, or inability to achieve an orgasm. An increase in erectile dysfunction and sexual dysfunction can have numerous underlying causes, including but not limited to (1) aging, (b) an underlying physical dysfunction, such as trauma, surgery, and peripheral vascular disease, and (3) side-effects resulting from drug treatment, depression, and other CNS disorders.

[0243] “Female sexual dysfunction” can be seen as resulting from multiple components including dysfunction in desire, sexual arousal, sexual receptivity, and orgasm related to disturbances in the clitoris, vagina, periurethral glans, and other trigger points of sexual function. In particular, anatomic and functional modification of such trigger points may diminish the orgasmic potential in breast cancer and gynecologic cancer patients. Treatment of female sexual dysfunction with an MC-4 receptor agonist can result in improved blood flow, improved lubrication, improved sensation, facilitation of reaching orgasm, reduction in the refractory period between orgasms, and improvements in arousal and desire. In a broader sense, “female sexual dysfunction” also incorporates sexual pain, premature labor, and dysmenorrhea.

[0244] Administration and Dose Ranges

[0245] Any suitable route of administration may be employed for providing a mammal, especially a human with an effective dosage of a compound of the present invention. For example, oral, rectal, topical, parenteral, ocular, pulmonary, nasal, and the like may be employed. Dosage forms include tablets, troches, dispersions, suspensions, solutions, capsules, creams, ointments, aerosols, and the like. Preferably compounds of Formula I are administered orally or topically.

[0246] The effective dosage of active ingredient employed may vary depending on the particular compound employed, the mode of administration, the condition being treated and the severity of the condition being treated. Such dosage may be ascertained readily by a person skilled in the art.

[0247] When treating obesity, in conjunction with diabetes and/or hyperglycemia, or alone, generally satisfactory results are obtained when the compounds of the present invention are administered at a daily dosage of from about 0.001 milligram to about 100 milligrams per kilogram of animal body weight, preferably given in a single dose or in divided doses two to six times a day, or in sustained release form. In the case of a 70 kg adult human, the total daily dose will generally be from about 0.07 milligrams to about 3500 milligrams. This dosage regimen may be adjusted to provide the optimal therapeutic response.

[0248] When treating diabetes mellitus and/or hyperglycemia, as well as other diseases or disorders for which compounds of formula I are useful, generally satisfactory results are obtained when the compounds of the present invention are administered at a daily dosage of from about 0.001 milligram to about 100 milligram per kilogram of animal body weight, preferably given in a single dose or in divided doses two to six times a day, or in sustained release form. In the case of a 70 kg adult human, the total daily dose will generally be from about 0.07 milligrams to about 350 milligrams. This dosage regimen may be adjusted to provide the optimal therapeutic response.

[0249] For the treatment of sexual dysfunction compounds of the present invention are given in a dose range of 0.001 milligram to about 100 milligram per kilogram of body weight, preferably as a single dose orally or as a nasal spray.

[0250] Combination Therapy

[0251] Compounds of Formula I may be used in combination with other drugs that are used in the treatment/prevention/suppression or amelioration of the diseases or conditions for which compounds of Formula I are useful. Such other drugs may be administered, by a route and in an amount commonly used therefor, contemporaneously or sequentially with a compound of Formula I. When a compound of Formula I is used contemporaneously with one or more other drugs, a pharmaceutical composition containing such other drugs in addition to the compound of Formula I is preferred. Accordingly, the pharmaceutical compositions of the present invention include those that also contain one or more other active ingredients, in addition to a compound of Formula I. Examples of other active ingredients that may be combined with a compound of Formula I, either administered separately or in the same pharmaceutical compositions, include, but are not limited to:

[0252] (a) insulin sensitizers including (i) PPARγ agonists such as the glitazones (e.g. troglitazone, pioglitazone, englitazone, MCC-555, BR-49653 and the like), and compounds disclosed in WO97/27857, 97/28115, 97/28137 and 97/27847; (ii) biguanides such as metformin and phenformin;

[0253] (b) insulin or insulin mimetics;

[0254] (c) sulfonylureas, such as tolbutamide and glipizide;

[0255] (d) α-glucosidase inhibitors (such as acarbose),

[0256] (e) cholesterol lowering agents such as (i) HMG-CoA reductase inhibitors (lovastatin, simvastatin, pravastatin, fluvastatin, atorvastatin, and other statins), (ii) sequestrants (cholestyramine, colestipol and a dialkylaminoalkyl derivatives of a cross-linked dextran), (ii) nicotinyl alcohol nicotinic acid or a salt thereof, (iii) proliferator-activater receptor α agonists such as fenofibric acid derivatives (gemfibrozil, clofibrate, fenofibrate and benzafibrate), (iv) inhibitors of cholesterol absorption for example beta-sitosterol and (acyl CoA:cholesterol acyltransferase) inhibitors for example melinamide, (v) probucol, (vi) vitamin E, and (vii) thyromimetics;

[0257] (f) PPARδ agonists, such as those disclosed in WO97/28149;

[0258] (g) antiobesity compounds, such as fenfluramine, dexfenfluramine, phentermine, sibutramine, orlistat, or β3 adrenergic receptor agonists;

[0259] (h) feeding behavior modifying agents, such as neuropeptide Y antagonists (e.g. neuropeptide Y5) such as those disclosed in WO 97/19682, WO 97/20820, WO 97/20821, WO 97/20822 and WO 97/20823;

[0260] (i) PPARα agonists such as described in WO 97/36579 by Glaxo;

[0261] (j) PPARγ antagonists as described in WO97/10813;

[0262] (k) serotonin reuptake inhibitors such as fluoxetine and sertraline;

[0263] (l) growth hormone secretagogues such as MK-0677; and

[0264] (m) agents useful in the treatment of male and/or female sexual dysfunction, such as type V cyclic-GMP-specific phosphodiesterase (PDE-V) inhibitors, including sildenafil and IC-351; alpha-adrenergic receptor antagonists, including phentolamine and yohimbine and pharmaceutically acceptable salts thereof; and dopamine receptor agonists, such as apomorphine.

[0265] In one embodiment of a combination for the treatment of male or female sexual dysfunction, the second ingredient to be combined with a compound of Formula I can be a type V cyclic-GMP-specific phosphodiesterase (PDE-V) inhibitor, such as sildenafil and IC-351 or a pharmaceutically acceptable salt thereof; an alpha-adrenergic receptor antagonist, such as phentolamine and yohimbine or a pharmaceutically acceptable salt thereof; or a dopamine receptor agonist, such as apomorphine or a pharmaceutically acceptable salt thereof.

[0266] Pharmaceutical Compositions

[0267] Another aspect of the present invention provides pharmaceutical compositions which comprises a compound of Formula I and a pharmaceutically acceptable carrier. The pharmaceutical compositions of the present invention comprise a compound of Formula I as an active ingredient or a pharmaceutically acceptable salt thereof, and may also contain a pharmaceutically acceptable carrier and optionally other therapeutic ingredients. The term “pharmaceutically acceptable salts” refers to salts prepared from pharmaceutically acceptable non-toxic bases or acids including inorganic bases or acids and organic bases or acids.

[0268] The compositions include compositions suitable for oral, rectal, topical, parenteral (including subcutaneous, intramuscular, and intravenous), ocular (ophthalmic), pulmonary (nasal or buccal inhalation), or nasal administration, although the most suitable route in any given case will depend on the nature and severity of the conditions being treated and on the nature of the active ingredient. They may be conveniently presented in unit dosage form and prepared by any of the methods well-known in the art of pharmacy.

[0269] In practical use, the compounds of Formula I can be combined as the active ingredient in intimate admixture with a pharmaceutical carrier according to conventional pharmaceutical compounding techniques. The carrier may take a wide variety of forms depending on the form of preparation desired for administration, e.g., oral or parenteral (including intravenous). In preparing the compositions for oral dosage form, any of the usual pharmaceutical media may be employed, such as, for example, water, glycols, oils, alcohols, flavoring agents, preservatives, coloring agents and the like in the case of oral liquid preparations, such as, for example, suspensions, elixirs and solutions; or carriers such as starches, sugars, microcrystalline cellulose, diluents, granulating agents, lubricants, binders, disintegrating agents and the like in the case of oral solid preparations such as, for example, powders, hard and soft capsules and tablets, with the solid oral preparations being preferred over the liquid preparations.

[0270] Because of their ease of administration, tablets and capsules represent the most advantageous oral dosage unit form in which case solid pharmaceutical carriers are obviously employed. If desired, tablets may be coated by standard aqueous or nonaqueous techniques. Such compositions and preparations should contain at least 0.1 percent of active compound. The percentage of active compound in these compositions may, of course, be varied and may conveniently be between about 2 percent to about 60 percent of the weight of the unit. The amount of active compound in such therapeutically useful compositions is such that an effective dosage will be obtained. The active compounds can also be administered intranasally as, for example, liquid drops or spray.

[0271] The tablets, pills, capsules, and the like may also contain a binder such as gum tragacanth, acacia, corn starch or gelatin; excipients such as dicalcium phosphate; a disintegrating agent such as corn starch, potato starch, alginic acid; a lubricant such as magnesium stearate; and a sweetening agent such as sucrose, lactose or saccharin. When a dosage unit form is a capsule, it may contain, in addition to materials of the above type, a liquid carrier such as a fatty oil.

[0272] Various other materials may be present as coatings or to modify the physical form of the dosage unit. For instance, tablets may be coated with shellac, sugar or both. A syrup or elixir may contain, in addition to the active ingredient, sucrose as a sweetening agent, methyl and propylparabens as preservatives, a dye and a flavoring such as cherry or orange flavor.

[0273] Compounds of formula I may also be administered parenterally. Solutions or suspensions of these active compounds can be prepared in water suitably mixed with a surfactant such as hydroxy-propylcellulose. Dispersions can also be prepared in glycerol, liquid polyethylene glycols and mixtures thereof in oils. Under ordinary conditions of storage and use, these preparations contain a preservative to prevent the growth of microorganisms.

[0274] The pharmaceutical forms suitable for injectable use include sterile aqueous solutions or dispersions and sterile powders for the extemporaneous preparation of sterile injectable solutions or dispersions. In all cases, the form must be sterile and must be fluid to the extent that easy syringability exists. It must be stable under the conditions of manufacture and storage and must be preserved against the contaminating action of microorganisms such as bacteria and fungi. The carrier can be a solvent or dispersion medium containing, for example, water, ethanol, polyol (e.g. glycerol, propylene glycol and liquid polyethylene glycol), suitable mixtures thereof, and vegetable oils.

[0275] In the Schemes and Examples below, various reagent symbols and abbreviations have the following meanings:

BOC    t-butyloxycarbonyl
Bu     butyl
calc.  calculated
CBZ    benzyloxycarbonyl
DCM    dichloromethane
DEAD   diethyl azodicarboxylate
DIEA   diisopropylethylamine
DMAP   4-dimethylamino-pyridine
DMF    N,N-dimethylformamide
EDC    1-(3-dimethylaminopropyl)3-ethylcarbodiimide HCl
eq.    equivalent(s)
ESI-MS electron spray ion-mass spectroscopy
Et     ethyl
EtOAc  ethyl acetate
HOAt   1-hydroxy-7-azabenzotriazole
HOBt   1-hydroxybenzotriazole hydrate
HPLC   high performance liquid chromatography
LC-MS  liquid chromatography-mass spectroscopy
LDA    lithium diisopropylamide
MC-xR  melanocortin receptor (x being a number)
Me     methyl
MF     molecular formula
Ms     methanesulfonyl
NMM    N-methylmorpholine
OIC    octahydroindole-2-carboxylic acid
Ph     phenyl
Phe    phenylalanine
Pr     propyl
prep.  prepared
PyBrop bromo-tris-pyrrolidino-phosphonium
       hexafluorophosphate
TFA    trifluoroacetic acid
THF    tetrahydrofuran
Tic    1,2,3,4-tetrahydroisoquinoline-3-carboxylic acid
TLC    thin-layer chromatography

[0276] PREPARATION OF COMPOUNDS OF THE INVENTION

[0277] The novel compounds of the present invention can be prepared according to the procedure of the following schemes and examples, using appropriate starting materials and are further exemplified by the following specific examples. The compounds illustrated in the examples are not, however, to be construed as forming the only genus that is considered as the invention. The following examples further illustrate details for the preparation of the compounds of the present invention. Those skilled in the art will readily understand that known variations of the conditions and processes of the following preparative procedures can be used to prepare these compounds. All temperatures are degrees Celsius unless otherwise noted.

[0278] The following Schemes and Examples describe procedures for making representative compounds of the present invention. Moreover, by utilizing the procedures described in detail in PCT International Applications WO 99/64002 (Dec. 16, 1999) and WO 00/74679 (Dec. 14, 2000), which are incorporated by reference herein in their entirety, in conjunction with the disclosure contained herein, one of ordinary skill in the art can readily prepare additional compounds of the present invention claimed herein.

[0279] The phrase standard peptide coupling reaction conditions means coupling a carboxylic acid with an amine using an acid activating agent such as EDC, DCC, and BOP in a inert solvent such as dichloromethane in the presence of a catalyst such as HOBT. The use of protecting groups for amine and carboxylic acid to facilitate the desired reaction and minimize undesired reactions is well documented. Conditions required to remove protecting groups are found in standard textbooks such as Greene, T, and Wuts, P. G. M., Protective Groups in Organic Synthesis, John Wiley & Sons, Inc., New York, N.Y., 1991. CBZ and BOC are commonly used protecting groups in organic synthesis, and their removal conditions are known to those skilled in the art. For example, CBZ may be removed by catalytic hydrogenation with hydrogen in the presence of a noble metal or its oxide such as palladium on activated carbon in a protic solvent such as ethanol. In cases where catalytic hydrogenation is contraindicated due to the presence of other potentially reactive functionality, removal of CBZ groups can also be achieved by treatment with a solution of hydrogen bromide in acetic acid, or by treatment with a mixture of TFA and dimethylsulfide. Removal of BOC protecting groups is carried out in a solvent such as methylene chloride or methanol or ethyl acetate, with a strong acid, such as trifluoroacetic acid or hydrochloric acid or hydrogen chloride gas.

[0280] It is understood that in some cases the order of carrying out the reaction schemes below may be varied to facilitate the reaction or to avoid unwanted reaction products.

PREPARATION OF 4-SUBSTITUTED PIPERIDINE INTERMEDIATES

[0281] The preparation of 4-substituted piperidine intermediates for coupling with the appropriate carboxylic acid intermediates is disclosed in PCT International Application WO 00/74679 (Dec. 14, 2000), which is incorporated by reference herein in its entirety. The synthesis of additional 4-substituted piperidine intermediates needed to prepare the compounds of the present invention is provided below.

Piperidine Intermediate 1

[0282] 370

[0283] To a solution of 4-cyclohexyl 4-formyl-N-(tertbutyloxycarbonyl)piperidine (2.56 g, 8.68 mmol) in toluene (100 ml) was added acetic acid (2 ml) and 1-amino-1-cyclopentanemethanol (1.0 g, 8.68 mmol). After refluxing by using a Dean-Stark apparatus for 11 hours, the reaction mixture was concentrated. The residue was dissolved in acetic acid (70 ml) and hydrogenated overnight in the presence of platinum oxide (500 mg) under a balloon atmosphere of hydrogen gas. The catalyst was filtered off and solvent was removed to give a colorless oil, which was dissolved in methanol and made basic by addition of NaOH (5N, 4 ml) and concentrated. The residue was partitioned between water and CH2Cl2, the two layers separated, and the aqueous layer extracted with CH2Cl2. The combined organic extracts were washed with brine, dried over MgSO4 and concentrated to give the title compound as a colorless oil (2.1 g).

[0284] MS: calc.for C23H42N2O3: 394.3; Found: 395 (M+1), 417 (M+Na).

Piperidine Intermediate 2

[0285] 371

[0286] To a solution of Intermediate 1 (2.1 g, 5.33 mmol) in CH2Cl2 (70 ml) at 0° was added DMAP (0.65 g, 5.33 mmol), DIEA (3.76 ml, 21.3 mmol) followed by slow addition of phosgene (4.1 ml, 8.0 mmol). After stirring the reaction mixture for one hour at 0° C., the ice-water bath was removed and the reaction mixture was continued to stir at room temperature overnight. The mixture was diluted with CH2Cl2, washed with water and brine, dried over MgSO4 and concentrated to give crude product, which was purified by column chromatography on silica gel (2% EtOAc/CH2Cl2 to 5% EtOAc/CH2Cl2) to give the title compound as a white solid (1.2g).

[0287] MS: calc. for C24H40N2O4: 420.3; Found: (M+1), (M+Na).

Piperidine Intermediate 3

[0288] 372

[0289] To the Intermediate 2 (1.2 g) was added hydrogen chloride (4.0 M in dioxane). The reaction mixture was stirred at room temperature for 30 minutes and the solvent was removed in vacuo to afford the title compound (1.2 g).

[0290] MS: calc.for C19H32N2O2: 320.3; Found: 321.1 (M+H).

Piperidine Intermediate 4

[0291] 373

[0292] Intermediate 4 was prepared from (S)-(+)-2-amino-1-propanol in an analogous manner to the one described for the preparation of Intermediate 1.

[0293] MS: calc.for C20H38N2O3: 354; Found: 355 (M+H).

Piperidine Intermediate 5

[0294] 374

[0295] Intermediate 5 was prepared from Intermediate 4 in an analogous manner to the one described for the preparation of Intermediate 2.

[0296] MS: calc. for C21H36N2O4: 380.3; Found: 381 (M+H).

Piperidine Intermediate 6

[0297] 375

[0298] Intermediate 6 was prepared from Intermediate 5 in an analogous manner to the one described for the preparation of Intermediate 3.

[0299] MS: calc. for C16H28N2O2: 280.3; Found: 281 (M+H).

Piperidine Intermediate 7

[0300] 376

[0301] To a suspension of 1-aminocyclopropane-1-carboxylic acid (2.8 g, 27.7 mmol) in THF (20 ml) was added borane-tetrahydrofuran complex (100 ml, 100 mmol) slowly under nitrogen at room temperature. The reaction mixture was stirred at 70° C. overnight, then cooled to 0° C. After addition of methanol (12.2 ml, 300 mmol), the mixture was allowed to stir for 30 minutes. Then acetic acid (1.6 ml, 27.7 mmol) was added. The reaction mixture was concentrated to provide the title compound as a colorless oil (3.0 g).

Piperidine Intermediate 8

[0302] 377

[0303] Intermediate 8 was prepared from Intermediate 7 in an analogous manner to the one described for the preparation of Intermediate 1.

[0304] MS: calc. for C21H38N2O3: 366.3; Found: 367 (M+H).

Piperidine Intermediate 9

[0305] 378

[0306] To a solution of Intermediate 8 (0.8 g, 2.18 mmol) in CH2Cl2 (40 ml) at 0° was added DMAP (0.266 g, 2.18 mmol), DIEA (1.52 ml, 8.74 mmol) and triphosgene (0.648 g, 2.18 mmol). After stirring the reaction mixture for one hour at 0° C., the ice-water bath was removed and the reaction mixture was allowed to stir at r.t. overnight. The mixture was diluted with CH2Cl2, washed with water and brine, dried over MgSO4 and concentrated to give crude product, which was purified by column chromatography on silica gel (10% CH2Cl2/EtOAc) to give the title compound as a colorless oil (0.13 g).

[0307] ESI-MS: calc. for C22H36N2O4: 392; Found: 393 (M+1).

Piperidine Intermediate 10

[0308] 379

[0309] Intermediate 10 was prepared from Intermediate 9 in an analogous manner to the one described for the preparation of Intermediate 3.

[0310] MS: calc. for C17H28N2O2: 292.2; Found: 293 (M+H).

Piperidine Intermediate 11

[0311] 380

[0312] To a solution of the alcohol (9.41 g, 31.6 mmol) in CH2Cl2 (100 ml) at 0° C. containing molecular sieves (2 g) and 4-methylmorpholine N-oxide (NMMO) (4.449 g, 37.98 mmol) was added TPAP (1.12 g, 3.16 mmol). After stirring the reaction mixture at 0° C. for 0.5 h, the reaction mixture was warmed to room temperature and stirred further for 5 hrs. The reaction mixture was concentrated to half the volume, diluted with hexane (250 ml), filtered through a silica gel pad and concentrated to give pure title compound (9.4 g).

Piperidine Intermediate 12

[0313] 381

[0314] To a solution of the aldehyde (2 g, 6.7 mmol) in toluene (50 ml) was added acetic acid (500 μl). After stirring the reaction mixture at reflux temperature using Dean Stark apparatus for 8 hrs, the mixture was concentrated and dissolved in acetic acid (30 ml). To the mixture was added PtO2 (500 mg) which was stirred under an atmosphere of H2 overnight. The rection mixture was flushed with nitrogen, filtered and concentrated to give the title compound (2 g).

Piperidine Intermediate 13

[0315] 382

[0316] To a solution of the amino alcohol (4.96 g, 13.47 mmol) in CH2Cl2 at 0° C. containing DIEA (6.98 g, 53.9 mmol), DMAP (1.64 g, 13.47 mmol) was added slowly a toluene solution of phosgene (1.93M, 10.47 ml, 20.21 mmol). After stirring the reaction mixture for 1 hr at 0° C., the temperature was raised to room temperarure and stirred further for 2 hrs. The reaction mixture was diluted with CH2Cl2, washed with water, brine, dried and concentrated. The residue was purified by column chromatography over silica gel (5% EtOAc/CH2Cl2) to give pure product (3.95 g).

Piperidine Intermediate 14

[0317] 383

[0318] To a solution of Intermediate 13 (3.95 g) in CH2Cl2 was added 5 ml of a saturated HCl solution of EtOAc. After stirring the reaction mixture for 30 minutes at room temperature, the solvent was removed and the residue lyophilized from a benzene/methanol solution to afford the title compound (3.85 g).

Piperidine Intermediate 15

[0319] 384

[0320] To a suspension of the alcohol (29 g, 97.5 mmol), 4-methylmorpholine-N-oxide (15.8 g, 134.6 mmol), and molecular sieves (15.0 gm) in DCM (500 mL) was added tetrapropylammonium perruthenate (TPAP, 1.03 g, 2.92 mmol) portionwise at room temperature. The mixture was stirred at room temperature for 30 min and TLC showed the reaction was completed. The mixture was filtered through a pad of silica gel, washed with DCM and 2:1 hexane/EtOAc. The mixture was then concentrated to give the aldehyde as a light yellow oil (28.5 g, 99%). 385

[0321] To a solution of methyl diethylphosphonoacetate (24.8 g, 117.8 mmol) in THF (400 mL) was add LDA (2.0 N, 58.9 mL, 117.8 mmol) at 0° C. After 30 min, a solution of the aldehyde from the previous step (28.5 g, 98.2 mmol) in THF (100 mL) was added, and the mixture was stirred at room temperature for two days and was then brought to reflux temperature overnight. The solvent was removed by rotary evaporation. The mixture was quenched with saturated NH4Cl and extracted with EtOAc. The combined organic layers were washed with brine, dried, filtered, concentrated, and purified by medium pressure-liquid chromatography to give the unsaturated ester (31.3 g, 90.7%). 386

[0322] To a solution of the unsaturated ester (20 g, 56.9 mmol) in MeOH (200 mL) was added Pd/C (10%, 6.05 g), and the suspension was placed on a shaker under a hydrogen gas atmosphere (50 psi) overnight. The solid was filtered and washed with MeOH, and solvents were removed to give the product (19.3 g, 96%). 387

[0323] To a solution of the ester (2.9 g, 8.2 mmol) in dry THF (100 mL) was added MeLi (1.4 N in THF, 29.3 mL, 41.0 mmol) at −78° C. The mixture was stirred at −78° C. for 3 h and quenched with HCl (4.0 N in dioxane, 10.0 mL). The solvent was removed and the residue was washed with ether. The ether solution was concentrated to give the product (2.85 g, 98%) as an oil. 388

[0324] To a solution of HCl in dioxane (4 N, 14.1 mL, 56.6 mmol) was added the N-Boc-protected alcohol (2.0 g, 5.66 mmol) at room temperature. The mixture was stirred for 1 h and then the solution was evaporated to give Intermediate 15 (1.34 g, 81.7%) as a white solid.

Piperidine Intermediate 16

[0325] 389

[0326] A dry flask was charged with NaH (60% in oil, 960 mg, 24 mmol) and anhydrous THF (40 mL). Added the alcohol starting material (5.95 g, 20 mmol) in dry THF (20 ml) through a two-ended needle under nitrogen atmosphere. Stirred at room temperature for about 60 min or until bubbling ceased, then added ethyl 2-bromoisopropionate (3.12 ml, 24 mmol). The mixture was stirred at room temperature overnight under nitrogen atmosphere. Quenched the reaction by adding the reaction mixture in portions to EtOAc (200 ml)/ice water (50 ml) with stirring. Transferred the mixture to a separatory funnel and added 1N HCl (30 ml). Extracted the aqueous solution with EtOAc (3×150 ml). Combined the organic phases which were dried over MgSO4. Concentrated in vacuo and purified by flash column chromatography on silica gel using 20% EtOAc in hexane as eluent to give the desired product (1.0 g, 13%). LC-MS: M+1=398.5. 390

[0327] To the stirred solution of the Boc-derivative (1.0 g, 2.5 mmol) in dry THF (50 ml) was added LDA (1.5 M in cyclohexane, 2.0 ml, 3 mmol) dropwise at −78° C. The mixture was stirred at −78° C. for 30 min, then MeI (784 μl, 12.5 mmol) was added. Slowly warmed up to room temperature and stirred at room temperature overnight. Quenched the reaction by adding the reaction mixture in portions to EtOAc (200 ml)/ice water (50 ml) with stirring. Transferred the mixture to a separatory funnel and added 1N HCl (30 ml). Extracted the aqueous solution with EtOAc (3×150 ml). Combined the organic phases and dried over MgSO4. Concentrated in vacuo and purified by flash column chromatography on silica gel using 20% EtOAc in hexane as eluent to give the desired product as a thick oil (681.8 mg). LC-MS: M+1=412.

[0328] 1 H NMR (400 MHz, CDCl3): δ 4.13 (q, J=7.2, 2 H), 3.52 (br, 2H), 3.25 (s, 2H), 3.18-3.12 (m, 2H), 1.75-1.61 (m, 5H), 1.53-1.388 (m, 4H), 1.42 (s, 9H), 1.35 (s, 6H), 1.27 (t, J=7.2, 3H), 1.10 (m, 6H).

[0329] Dissolved the above resulting compound in 4N HCl in dioxane (20 ml). Stirred at room temperature for about 60 min. Evaporated to dryness to give Intermediate 16 as a white solid (541 mg). LC-MS: M+1=312.

Piperidine Intermediate 17

[0330] 391

[0331] To the stirred solution of N-Cbz-4-cyclohexyl-piperidine-4-carboxylic acid (1.0 g, 2.9 mmol) in DCM (20 ml) was added oxalyl chloride (2.0M in DCM, 1.6 ml, 3.19 mmol) dropwise. Then added 3 drops of DMF. Stirred at room temperature for 1 hour and then evaporated to give the desired product. The crude mixture was used in the next step without further purification.

[0332] To the stirred solution of the acid chloride (2.9 mmol) in 1,2-dichloroethane (30 ml) was added α-methylalanine methyl ester (446 mg, 2.9 mmol) and DIEA (1.01 ml, 5.8 mmol). Stirred at 75° C. for 1 hour, and then at 60° C. overnight. Cooled to room temperature and diluted the mixture with DCM. Washed with 1N HCl, satd. NaHCO3 and then satd. NaCl. Dried over Na2SO4 and concentrated in vacuo to give the crude desired product (1.2 g). LC-MS: 445 (M+1).

[0333] Dissolved intermediate from previous step (1.2 g, 2.7 mmol) in ethanol (50 ml). Added Pd-C (10%, 200 mg) and stirred at room temperature in the presence of hydrogen gas for two hours. Filtered off the catalyst, and concentrated in vacuo to give Intermediate 17 (663 mg). LC-MS: 312 (M+1).

Piperidine Intermediate 18

[0334] 392

[0335] To a solution of the enone (6 mmol, 0.7 mL) in MeOH (20 mL) at 0° C. was added NaBH4 (3 mmol, 113 mg). The reaction was stirred at room temp for 1 hr. Volatiles were removed and the residue partitioned between CH2Cl2 and 0.5M HCl. Organic phase was dried over MgSO4 and concentrated to afford a clear colorless oil which was used in the next step without further purification. 393

[0336] A solution of the acid (6 mmol, 1.38 g), EDC (12 mmol, 2.3 g), DMAP (ca.50 mg) and enol (ca. 6 mmol) in CH2Cl2 (25 mL) was stirred at room temp for 72 hours. Reaction mixture was poured into EtOAc (200 mL) and washed successively with 0.5M HCl, 1M NaOH, H2O and brine, dried over Na2SO4 and concentrated. Chromatography over silica gel eluting with 500 mL of 5-10% EtOAc/hexane afforded a clear colorless oil (1.9 g). 394

[0337] To a solution of LDA (2M in THF) (4.38 mmol, 2.2 mL) in THF (10 mL) at −78° C. was added the ester from the previous step (3.98 mmol, 1.3 g) in THF (2 mL) followed 30 min later by TMSCl (4.38 mmol, 0.6 mL). Resultant solution was allowed to warm to room temperature and then heated at reflux for 16 hr. After cooling to room temp, 2M HCl (5 mL) was added and stirring continued for 5 min. Resultant solution was partitioned between Et2O (40 mL) and 2M HCl. The organic phase was washed with H2O and brine, dried over Na2SO4 and concentrated. Chromatography over silica eluting with 20-30% EtOAc/hexane afforded the desired acid as an off-white solid (653 mg). 395

[0338] To a solution of the acid from the previous step (1.46 mmol, 474 mg) in CH2Cl2 (5 mL) at 0° C. was added oxalyl chloride (2M in CH2Cl2) (1.61 mmol, 0.81 mL) and DMF (0.05 mL) and the reaction stirred at 0° C. for 1 hr. Volatiles were removed, azeotroping with toluene and finally under high vacuum for 3 hr to afford the acid chloride. The acid chloride was dissolved in t-butylamine (5 mL) and the resultant cloudy solution was stirred at room temperature overnight. The reaction mixture was concentrated to afford a yellow solid. Chromatography over silica gel eluting with 50 mL of 5% then 100 mL of 10-20% EtOAc/hexane afforded the desired tert-butyl amide as a white solid (282 mg). 396

[0339] A suspension of Pd (10% on activated charcoal) (10 mol %, 79 mg) in a solution of the N-Boc derivative from the previous step (0.75 mmol, 282 mg) in MeOH containing 4M HCl (4M in dioxane) (1.5 mmol, 0.37 mL) was shaken under 45 psi of hydrogen gas for 60 hours. After work-up, the hydrochloride salt was used without further purification in the peptide coupling reaction.

Piperidine Intermediate 19

[0340] 397

[0341] To a solution of the acid (10 mmol, 2.29 g) in CH2Cl2 (40 mL) at room temp was added EDC (20 mmol, 3.8 g) and DMAP (ca.50 mg) followed by 3-methyl-2-buten-1-ol (15 mmol, 1.52 mL). Resultant solution was stirred at room temp overnight. Reaction mixture was poured into EtOAc (200 mL) and washed successively with 0.5M HCl, 1M NaOH, H2O and brine, dried over Na2SO4 and concentrated. Chromatography over silica gel eluting with 500 mL of 5% then 250 mL of 10% EtOAc/hexane afforded the ester as a clear colorless oil (2.97 g). 398

[0342] To a solution of LDA (2M in THF) (7.46 mmol, 3.73 mL) in THF (15 mL) at −78° C. was added the ester from the previous step (6.78 mmol, 2.02 g) in THF (3 mL) followed 30 min later by TMSCl (7.46 mmol, 0.95 mL). Resultant solution was allowed to warm to room temp and heated at reflux for 24 hr. After cooling to room temp, 2M HCl (5 mL) was added and stirring continued for 5 min. Resultant solution was partitioned between Et2O (40 mL) and 2M HCl. The organic phase was washed with H2O and brine, dried over Na2SO4 and concentrated. Chromatography over silica eluting with 10-20% EtOAc/hexane afforded the desired acid as a white solid (1.23 g). 399

[0343] To a solution of the acid from the previous step (4.14 mmol, 1.23 g) in CH2Cl2 (10 mL) at 0° C. was added oxalyl chloride (2M in CH2Cl2) (4.55 mmol, 2.27 mL) and DMF (0.15 mL) and the reaction stirred at 0° C. for 1 hr. Volatiles were removed, azeotroping with toluene and finally under high vacuum for 3 hr to afford the acid chloride. The acid chloride was dissolved in t-butylamine (10 mL) and the resultant cloudy solution was left to stir at room temperature overnight. The reaction mixture was concentrated and partitioned between CH2Cl2 and 2M HCl. Organics were dried over Na2SO4 and concentrated. Chromatography over silica gel eluting with 10-30% EtOAc/hexane afforded a white solid (1.07 g).

[0344] A suspension of Pd (10% on activated charcoal) (10 mol %, 322 mg) in a solution of the compound from the previous step (3.03 mmol, 1.07 g) in MeOH (60 mL) containing 4M HCl in dioxane (6.06 mmol, 1.5 mL) was shaken under 45 psi of hydrogen gas for 5 hr. Reaction was filtered through a short pad of celite and concentrated. Residue was dissolved in EtOAc (20 mL) and HCl (4M in dioxane) (20 mL). Resultant solution was left to stir at room temp for 1 hr. Volatiles were removed and the residue precipitated from a CH2Cl2 solution with Et2O/hexane to afford Intermediate 19 as a white solid.

Piperidine Intermediate 20

[0345] 400

[0346] To a solution of CrO3 (15.8 mmol, 1.59 g) in dry CH2Cl2 (20 mL) at −20° C. was added 3,5-dimethylpyrazole (15.8 mmol, 1.52 mg) in one portion. The resultant solution was stirred at −20° C. for 15 min before the addition of the cyclohexene intermediate (0.79 mmol, 289 mg) in CH2Cl2 (2.5 mL) over 3 min. The reaction mixture was warmed to −15° C. and stirred for a further 5 hr. 5N NaOH (51.5 mmol, 10.3 mL) was added and the emulsion stirred at 0° C. for 1 hr then at rt overnight. Aqueous phase was extracted with CH2Cl2, and the combined organics were washed with 1N HCl, water, satd NaHCO3 and brine, dried over Na2SO4 and concentrated. Chromatography over silica gel eluting with 50 mL of 0, 2.5, 5, and 10% EtOAc/hexane afforded the cyclohexenone as a white solid (135 mg). 401

[0347] A suspension of Pd (10% on activated charcoal) (20 mol %, 76 mg) in a solution of the cyclohexenone (0.36 mmol, 135 mg) in MeOH was shaken under 45 psi of hydrogen gas for 60 hr. The reaction mixture was filtered through a short pad of celite and concentrated to afford a clear colorless gum. Chromatography over silica gel eluting with 50 mL of 0, 2.5, 5, 10, and 20% Me2CO/CH2Cl2 afforded the cyclohexanone as a white solid (111 mg).

[0348] To a solution of the cyclohexanone (0.29 mmol, 111 mg) in CH2Cl2 was added (diethylamino)sulfur trifluoride (0.73 mmol, 0.1 mL). Resultant solution was left to stir at room temp for 24 hours. Reaction mixture was poured into saturated NaHCO3. Organic phase was washed with NaHCO3, dried over Na2SO4 and concentrated. Chromatography over silica gel eluting with 10-30% EtOAc/hexane afforded the difluorocyclohexane intermediate as a white solid (84 mg)

[0349] A solution of the difluorocyclohexane intermediate from the previous step (0.2 mmol, 80 mg) in CH2Cl2 and TFA was stirred at room temp for 1 hr. Volatiles were removed and the residue partitioned between NaOH and EtOAc. Organic phase was dried over Na2SO4 and concentrated to give Intermediate 20.

Piperidine Intermediate 21

[0350] 402

[0351] To a suspension of Reike Mg (5 g/200 mL THF) (6 mmol, 6 mL) at 0° C. was added a solution of 4-methyl-1-bromocyclohexane (4 mmol, 708 mg) in THF (4 mL) over a period of about 5 min. The resultant slurry was stirred at room temp for 5 min then cooled to −20° C. A solution of the Cbz-piperidine derivative (1 mmol, 276 mg) in THF (10 mL) was then added. The reaction was stirred at −20° C. for 15 min then poured into ice-cold 50% H2SO4 (25 mL) and stirred for a further 30 min. The emulsion was poured into H2O (100 mL) and extracted with CH2Cl2 (2×25 mL). The combined organic phases were dried over Na2SO4 and concentrated to afford a green oil. To a solution of this oil in DMSO (2 mL) was added NaNO2 (3 mmol, 207 mg) and AcOH (10 mmol, 0.6 mL). The resultant orange solution was stirred at 40° C. for 24 hr. After cooling to room temperature, 1N HCl (2.5 mL) was added and stirring continued for a further 15 min. The mixture was extracted with CH2Cl2 (3×5 mL). The combined organic layers were dried over Na2SO4 and concentrated. Chromatography over silica gel eluting with 100 mL of 10% and 50 mL of 20-30% EtOAc/hexane afforded the desired acid as an off-white solid (100 mg). 403

[0352] To a solution of the acid (0.42 mmol, 151 mg) in CH2Cl2 (2.5 mL) at 0° C. was added oxalyl chloride (2M in CH2Cl2) (0.46 mmol, 0.23 mL) and DMF (4 drops) and the reaction stirred at 0° C. for 1 hr. Volatiles were removed, azeotroping with toluene and finally under high vacuum for 3 hr to afford the acid chloride. The acid chloride was dissolved in CH2Cl2 (2.5 mL) and cooled to 0° C. t-Butylamine (1.26 mmol, 0.13 mL) was added and the resultant cloudy solution was left to stir at room temperature overnight. The reaction mixture was poured into CH2Cl2 (ca. 3 mL) and washed with brine, dried over Na2SO4 and concentrated. Chromatography over silica gel eluting with 250 mL of 20, 25, 30, and 40% EtOAc/hexane afforded the Cbz-protected t-butyl amide as a white foam (174 mg). A mixture of the Cbz-protected t-butyl amide (0.1 mmol, 174 mg) and catalytic Pd (10% on activated C) (20 mg) in methanol was stirred under an atmosphere of hydrogen gas at room temp for 1 hr. The solution was filtered through a short pad of celite and concentrated to give Intermediate 21.

Piperidine Intermediate 22

[0353] 404

[0354] A solution of the amine (400 mg, 1.42 mmol), cyclopropylsulfonyl chloride (600 mg, 4.26 mmol), DIEA (1.47 g, 11.36 mmol) and DMAP (100 mg, 0.8 mmol) in toluene (50 mL) was heated to reflux overnight. A solution of NaOH (5N, 10 mL) was added and allowed the reaction to reflux for an additional 4 h. The reaction mixture was cooled to rt and diluted with EtOAc (200 mL). The combined organics were washed with 0.5N HCl, satd NaHCO3, and brine, dried over Na2SO4 and concentrated. Chromatography over silica gel eluting with 50 mL of 10, 20, 15, 25, 40, and 50% EtOAc/hexane afforded the Boc-protected intermediate as a white solid (615 mg). A solution of this intermediate in CH2Cl2 (4mL) and HCl (4M in dioxane) (4 mL) was stirred at room temp for 1 hr. Volatiles were removed and the product precipitated from a CH2Cl2 solution with Et2O/hexane to give Intermediate 22 (615 mg).

Piperidine Intermediate 23

[0355] 405

[0356] To a solution of the ester (2.36 g, 9.17 mmol) in THF (50 mL) at −78° C. was added LDA (1.5M in THF) (6.72 mL, 10.09 mmol) and followed 45 min later by cyclopropylmethyl bromide (1.49 g, 11.0 mmol) in THE (10 mL). Resultant solution was allowed to warm to room temp overnight. Resultant solution was quenched with sat. NH4Cl and partitioned between EtOAc (40 mL) and 0.5M HCl. The organic phase was washed with H2O and brine, dried over Na2SO4 and concentrated. Chromatography over silica gel eluting with 5% EtOAc/hexane afforded the alkylated product (2.76 g). 406

[0357] A solution of intermediate from the previous step (2.76 g, 8.86 mmol) and LiOH (1.1 g, 44.3 mmol) in MeOH/H2O (70 mL) was heated to reflux overnight. More MeOH was added to the reaction mixture to make the solution homogeneous. The reaction mixture was concentrated to about 10 mL and acidified with 2N HCl to pH about 2. The aqueous solution was extracted with EtOAc (3×100 mL). The organics were washed successively with H2O and brine, dried over Na2SO4 and concentrated. Chromatography over silica gel eluting with 20-70% EtOAc/hexane afforded the Boc-protected acid as a white solid (1.69 g). To a solution of the acid (2.5 g, 8.82 mmol) in CH2Cl2 (5 mL) at 0° C. was added oxalyl chloride (2M in CH2Cl2) (4.85 mL, 9.70 mmol) and DMF (0.05 mL) and the reaction stirred at 0° C. for 1 hr. Volatiles were removed, azeotroping with toluene and finally under high vacuum for 3 hr to afford the acid chloride. The acid chloride was dissolved in t-butylamine (2.8 mL), and the resultant cloudy solution was left to stir at room temp overnight. The reaction mixture was concentrated and partitioned between CH2Cl2 and 2M HCl. Organics were dried over Na2SO4 and concentrated. A solution of the Boc-protected amide in CH2Cl2 (4 mL) and 4.0 M HCl/dioxane (4 mL) was stirred at room temp for 1 hr. Volatiles were removed and Intermediate 23 was precipitated from a CH2Cl2 solution with Et2O/hexane (1.9 g).

Piperidine Intermediate 24

[0358] 407

[0359] This intermediate was prepared in the same manner as Intermediate 23 but using cyclobutylmethyl bromide in place of cyclopropylmethyl bromide in the alkylation step.

Piperidine Intermediate 25

[0360] 408

[0361] To a solution of the acid (600 mg, 2.117 mmol) in THF (5 mL) at 0° C. was added BH3.Me2S (10 M in THF) (0.85 mL, 8.47 mmol) and the solution was left to stir at room temperature for 3 h. The reaction mixture was then cooled to 0° C., and H2O2 (30% aqueous, 2.5 mL) was added dropwise and then 1M NaOH (10 mL). The resultant solution was stirred for 10 min at 0° C. and then a further 30 min at rt. The reaction mixture was poured into EtOAc (100 mL), and washed successively with water, sat. NH4Cl, saturated NaHCO3, and brine, dried over Na2SO4 and concentrated. Chromatography over silica gel eluting with 40% EtOAc/hexane afforded the alcohol intermediate (611 mg). To a solution of the alcohol (611 mg, 2.268 mmol) and Et3N (0.63 mL, 4.5 mmol) in CH2Cl2 (5 mL) at 0° C. was added methanesulfonyl chloride (10 M in THF) (0.35 mL, 4.53 mmol) at 0° C. and the solution was left to stir at room temp 45 min. The reaction was concentrated and was poured into water (100 mL), and extracted with EtOAc (3×100 mL). The organics were dried over Na2SO4 and concentrated. Chromatography over silica gel eluting with 5-30% EtOAc/hexane afforded the mesylate as a solid. To a solution of the mesylate (596 mg, 1.7 mmol) in DMF (5 mL) at rt was added sodium isopropylsulfide (842 mg, 8.57 mmol) and the solution was left to stir at room temp overnight. The reaction was concentrated and was poured into water (100 mL), and extracted with EtOAc (3×100 mL). The organics were dried over Na2SO4 and concentrated. Chromatography over silica gel eluting with 5-30% EtOAc/hexane afforded a solid. A solution of the Boc-protected isopropyl sulfide in CH2Cl2 (4 mL) and 4.0 M HCl/dioxane (4 mL) was stirred at room temp for 1 hr. Volatiles were removed and Intermediate 25 was precipitated from a CH2Cl2 solution with Et2O/hexane (400 mg).

Piperidine Intermediate 26

[0362] 409

[0363] To a solution of the amino alcohol (177 mg, 0.48 mmol), NaOH (192 mg, 4.8 mmol) in CHCl3 (5 mL) and water (2 mL) at 0° C. was added dropwise a solution of BrCH2COBr (263 mg, 1.3 mmol) in CHCl3 (1 mL) over a period of 5 min., and the solution was left to stir at 0° C. for 1 h and then at rt overnight. The reaction was concentrated and was poured into water (100 mL), and extracted with EtOAc (3×100 mL). The organics were washed successively with water, 1N HCl and brine and dried over Na2SO4 and concentrated. Chromatography over silica gel eluting with 25% EtOAc/hexane afforded a solid. A solution of this solid in CH2Cl2 (4 mL) and 4.0 M HCl/dioxane (4 mL) was stirred at room temp for 1 hr. Volatiles were removed and Intermediate 26 precipitated from a CH2Cl2 solution with Et2O/hexane (100 mg). 410

EXAMPLE 1

[0364] Step A

[0365] N-(Benzyloxycarbonyl)-4-cyclohexyl-piperidine-4-carboxylic acid (1-1) (2.5 g, 7.24 mmol) was dissolved in 36 mL of CH2Cl2 and cooled at 0° C. in an ice-H2O bath. Oxalyl chloride (2.0 M solution in CH2Cl2, 3.98 mL, 7.96 mmol) was then added dropwise followed by the addition of 1-2 drops of DMF. This mixture was stirred at 0° C. for 2 h and then concentrated with toluene. The residue was dissolved in CH2Cl2 and cooled at 0° C. in an ice-H2O bath, and then t-butylamine (2.28 mL, 21.72 mmol) was added dropwise. The reaction mixture was then stirred at 0° C. for 2 h, warmed to room temperature, and stirred at room temperature overnight. The resulting mixture was then diluted with CH2Cl2 and washed with brine, dried over MgSO4, filtered, and concentrated to give 1-2 as a solid (2.92 g). Mass spectrum: Calcd for C24H36N2O3: 400.27; Found: 401 (M++1).

[0366] Step B

[0367] Compound 1-2 (7.24 mmol) was dissolved in 30 mL of CH2Cl2 and then 30% HBr in acetic acid (7.2 mL, 36.15 mmol) was added. The mixture was stirred at room temperature for 45 min (reaction monitored by TLC), and then diethyl ether was added. The resulting precipitate was filtered and washed with ether. The solid was dissolved in ethyl acetate and washed with IN NaOH solution, and the aqueous layer was extracted with EtOAc. The combined organic phases were dried over K2CO3, filtered, and concentrated to give 1-3 as a white solid (1.362 g). Mass spectrum: Calcd for C16H30N2O: 266.24; Found: 267 (M++1).

[0368] Step C

[0369] N-Boc-(D)-4-chlorophenylalanine (0.935 g, 3.12 mmol) was dissolved in 14.2 mL of methylene chloride, and then amine 1-3 (0.755 g, 2.84 mmol), NMM (1.20 mL, 11.36 mmol), EDC (0.598 g, 3.12 mmol), and HOBt (0.422 g, 3.12 mmol) were added. The resulting mixture was stirred at room temperature overnight, and then diluted with 20 mL of CH2Cl2 and washed with 20 mL of 1N HCl solution, 20 mL of saturated NaHCO3 solution, 20 mL of H2O, and 20 mL of saturated NaCl solution. The organic phase was dried over MgSO4, filtered, and concentrated to give a white, foamy-solid. The crude product was purified by column chromatography (30:1 to 9:1 methylene chloride-acetone) to give 1-4 as a white solid (1.34 g). Mass spectrum: Calcd for C30H46N3O4Cl: 547.32; Found: 548 (M++1).

[0370] Step D

[0371] Compound 1-4 (1.33 g, 2.43 mmol) was dissolved in 6.1 mL of methylene chloride and 6.1 mL of trifluoroacetic acid, and this solution was stirred at room temperature for 30 min. The mixture was then concentrated with two 8-mL portions of toluene and two 8-mL portions of diethyl ether to give a white solid. The solid was dissolved in EtOAc and washed with 1N NaOH solution, and the aqueous layer was extracted with EtOAc. The combined organic phases were dried over K2CO3, filtered, and concentrated to give 1-5 as a foamy-solid (1.08 g). Mass spectrum: Calcd for C25H38N3O2Cl: 447.27; Found: 448 (M++1).

[0372] Step E

[0373] A 25-mL, round-bottomed flask was purged under nitrogen and charged with (R)-4-(benzyloxycarbonyl)-1-(tert-butoxycarbonyl)-piperazine-2-carboxylic acid (1-6) (0.5 g, 1.37 mmol) (the preparation of this intermediate from commercially available 2(R)-piperazine carboxylic acid was achieved via modifications to the procedures described by Bigge and coworkers in Tetrahedron Lett. 1989, 30, 5193) and 7 mL of DMF. Potassium carbonate (0.228 g, 1.65 mmol) was then added followed by the addition of methyl iodide (0.43 mL, 6.86 mmol), and the resulting mixture was stirred at room temperature overnight. The cloudy yellow mixture was diluted with H2O and EtOAc, and the layers were separated. The aqueous layer was extracted with EtOAc, and the combined organic phases were washed with brine, dried over Na2SO4, filtered, and concentrated. Purification by column chromatography (30% ethyl acetate-hexane) provided 1-7 (0.52 g). Mass spectrum: Calcd for C19H26N2O6: 378.18; Found: 279 (M++1-Boc).

[0374] Step F

[0375] Intermediate 1-7 (0.52 g, 1.37 mmol) was charged with 6.8 mL of EtOH, and 10% Pd/C (0.052 g). A H2 balloon was placed on the top of the flask via a 3-way stopcock and the system was evacuated and purged with H2 three times. The mixture was then stirred at room temperature under H2 overnight. The flask was evacuated and purged with N2 three times, and then the reaction mixture was filtered through a pad of Celite and concentrated to give 1-8 as a clear oil (0.328 g). Mass spectrum: Calcd for C11H20N2O4: 244.14; Found: 245 (M++1).

[0376] Step G

[0377] Compound 1-8 (0.205 g, 0.84 mmol) was dissolved in 4.2 mL of methanol, and then sodium acetate (0.345 g, 4.20 mmol), trifluoroacetic acid (0.065 mL, 0.84 mmol), and 37% aqueous formaldehyde solution (0.30 mL, 4.03 mmol) were added. The mixture was stirred at room temperature for 30 min, and then sodium cyanoborohydride (1.0 M in THF, 2.7 mL, 2.7 mmol) was added. The reaction mixture was stirred at room temperature overnight and then concentrated to give a white sludge. The crude mixture was dissolved in EtOAc and 1N NaOH, and the layers were separated. The organic phase was washed with 1N NaOH solution, H2O, and brine, dried over MgSO4, filtered, and concentrated. Purification by column chromatography (10% methanol-methylene chloride) provided 1-9 (0.14 g). Mass spectrum: Calcd for C12H22N2O4: 258.16; Found: 259 (M++1).

[0378] Step H

[0379] Ester 1-9 (0.14 g, 0.56 mmol) was dissolved in 2.7 mL of methanol, and then 1N NaOH solution (1.12 mL, 1.12 mmol) was added. The mixture was stirred at room temperature overnight, and then concentrated. The residue was dissolved in water and then the pH was adjusted to pH=6. The solution was then concentrated twice with toluene to give 1-10 (0.217 g, 63% purity). Mass spectrum: Calcd for C11H20N2O4: 244.14; Found: 245 (M++1).

[0380] Step I

[0381] (R)-4-Methyl-1-(tert-butoxycarbonyl)piperazine-2-carboxylic acid 1-10 (63% purity, 0.063 g, 0.172 mmol) was dissolved in 0.78 mL of methylene chloride, and then amine intermediate 1-5 (0.07 g, 0.156 mmol), NMM (0.07 mL, 0.624 mmol), EDC (0.033 g, 0.172 mmol), and HOBt (0.023 g, 0.172 mmol) were added. The resulting mixture was stirred at room temperature overnight, and then diluted with 10 mL of CH2Cl2 and washed with 5 mL of 1N HCl solution, 5 mL of saturated NaHCO3 solution, 5 mL of H2O, and 5 mL of saturated NaCl solution. The organic phase was dried over MgSO4, filtered, and concentrated to give a yellow oil. The crude product was purified by column chromatography (9:1 to 1:1 methylene chloride-acetone) to give 1-11 as a white solid (0.080 g). Mass spectrum: Calcd for C36H56N5O5Cl: 673.40; Found: 674 (M++1).

[0382] Step J

[0383] Compound 1-11 (0.078 g, 0.116 mmol) was dissolved in 0.30 mL of methylene chloride and 0.30 mL of trifluoroacetic acid. This solution was stirred at room temperature for 30 min, and then concentrated with two 5-mL portions of toluene and two 5-mL portions diethyl ether to give a white, foamy-solid. The solid was dissolved in EtOAc and washed with 1N NaOH solution, and the aqueous layer was extracted with EtOAc. The combined organic phases were dried over K2CO3, filtered, and concentrated, and the residue was purified by column chromatography (5-10% methanol-methylene chloride) to give a white, foamy-solid. The solid was dissolved in EtOAc and 1.0 M HCl solution in Et2O (0.28 mL, 0.28 mmol) was added. The precipitate was filtered under N2 and dried under vacuum to give 1-12 as a white solid (0.046 g). Mass spectrum: Calcd for C31H48N5O3Cl: 573.34; Found: 574 (M++1).

[0384] 1 H NMR (CD3OD): δ 7.28-7.13 (m, 4 H), 5.02 (m, 1 H), 4.26 (m, 2 H), 3.66-3.50 (m, 2 H), 3.26-3.45 (m, 3 H), 3.00-2.87 (m, 4 H), 2.78 (s, 3 H), 2.43 (m, 2 H), 1.94 (m, 2 H), 1.71 (m, 3 H), 1.58 (d, J=10.7 Hz, 1 H), 1.47 (d, J=11.7 Hz, 1 H), 1.25 (d, J=16.2 Hz, 9 H), 1.25-0.75 (m, 7 H), 0.11 (m, 1 H) 411

EXAMPLE 2

[0385] Step A

[0386] The bis-protected (S)-piperazine-2-carboxamide intermediate (2-1) was prepared by coupling amine 1-5 with (S)-4-(benzyloxycarbonyl)-1-(tert-butoxycarbonyl)-piperazine-2-carboxylic acid in place of (S)-4-methyl-1-(tert-butoxycarbonyl)-piperazine-2-carboxylic acid in Step I of Example 1.

[0387] Step B

[0388] Compound 2-1 (0.164 g, 0.206 mmol) was dissolved in 0.6 mL of methylene chloride and 30% HBr in acetic acid (0.409 mL, 2.06 mmol) was added. The mixture was stirred at room temperature for 45 min (TLC showed no starting material). To this orange solution, diethyl ether (5 mL) was added, and the precipitate was filtered and washed with ether. The solid was dissolved in ethyl acetate and washed with 1N NaOH and the aqueous layer was extracted with EtOAc. The combined organics were dried over K2CO3, filtered and concentrated to give a white solid (0.1096 g). A portion of the white solid (0.050 g, 0.089 mmol) was dissolved in EtOAc and 1.0 M HCl in Et2O (0.22 mL, 0.22 mmol) was added. The precipitate was filtered under N2 and dried under vacuum to give 2-2 as a white solid (0.047 g); mass spectrum: 560 (M+1); 582 (M+Na). 412

[0389] The intermediates shown in Table 1 below having bis-protected piperazine-2-carboxylic acid residues and variable X groups at the 4-position of the piperidine ring were prepared as shown in Scheme 3 above with the indicated stereochemistry at the stereogenic center marked with **.

TABLE 1
			Exact
X	R6	**	Mass	Mass Spec.
413	F	(R)	750.40	751 (M+ + 1)
414	Cl	(S)	793.42	794 (M+ + 1), 816 (M+ + Na)
415	F	(R)	777.55	778 (M+ + 1)
416	F	(S)	777.55	778 (M+ + 1), 800 (M+ + Na)
417	Cl	(R)	793.42	794 (M+ + 1)
418	F	(R)	777.55	778 (M+ + 1)
419	Cl	(S)	779.40	780 (M+ + 1), 802 (M+ + Na)
420	Cl	(S)	791.40	792 (M+ + 1)
421	Cl	(R)	791.40	792 (M+ + 1)
422	Cl	(S)	805.42	806 (M+ + 1)
423	Cl	(R)	805.42	806 (M+ + 1)
424	Cl	(S)	807.43	808 (M+ + 1)
425	Cl	(R)	807.43	808 (M+ + 1)
426	Cl	(R)	807.43	808 (M+ + 1)
427	Cl	(S)	814.38	815 (M+ + 1), 837 (M+ + Na)
428	Cl	(S)	819.36	820 (M+ + 1)
429	F	(S)	924.48	925 (M+ + 1), 947 (M+ + Na)
430	F	(S)	804.46	805 (M+ + 1), 827 (M+ + Na)
431	Cl	(S)	793.42	794 (M+ + 1), 816 (M+ + Na)
432	Cl	(R)	793.42	794 (M+ + 1), 816(M+ + Na)
433	Cl	(S)	821.41	822 (M+ + 1), 844 (M+ + Na), 722 (M+ − Boc)
434	F	(S)	805.44	806 (M+ + 1), 828 (M+ + Na), 706 (M+ − Boc)
435	Cl	(R)	821.41	822 (M+ + 1), 844 (M+ + Na), 722 (M+ − Boc)
436	F	(R)	805.44	806 (M+ + 1), 828 (M+ + Na), 706 (M+ − Boc)
437	Cl	(S)	795.4	796 (M+ + 1), 818 (M+ + Na)

[0390] The following Nα,β-unsubstituted piperazine examples shown in Table 2 below above with the indicated stereochemistry at the stereogenic center marked with ** were prepared from the intermediates in Table 1 by treatment with 30% HBr in acetic acid to afford the hydrobromide salt, which upon neutralization and subsequent treatment with HCl in diethyl ether afforded the hydrochloride salt.

TABLE 2
438
				Exact
Example	X	R6	**	Mass	Mass Spec.
3	439	F	(R)	516.31	517 (M+ + 1), 539 (M+ + Na)
4	440	Cl	(S)	559.23	560 (M+ + 1), 582 (M+ + Na)
5	441	F	(S)	543.36	544 (M+ + 1), 566 (M+ + Na)
6	442	Cl	(S)	545.31	546 (M+ + 1), 568 (M+ + Na)
7	443	Cl	(S)	571.33	572 (M+ + 1), 594 (M+ + Na)
8	444	Cl	(R)	571.33	572 (M+ + 1)
9	445	Cl	(S)	573.34	574 (M+ + 1), 596 (M+ + Na)
10	446	Cl	(R)	573.34	574 (M+ + 1)
11	447	Cl	(R)	573.34	574 (M+ + 1)
12	448	Cl	(S)	580.29	581 (M+ + 1), 603 (M+ + Na)
13	449	Cl	(S)	585.27	586 (M+ + 1), 608 (M+ + Na)
14	450	F	(S)	556.35	557 (M+ + 1), 579 (M+ + Na)
15	451	F	(S)	570.37	571 (M+ + 1), 593 (M+ + Na)
16	452	Cl	(S)	559.33	560 (M+ + 1), 582 (M+ + Na)
17	453	Cl	(R)	559.33	560 (M+ + 1), 582 (M+ + Na)
18	454	F	(S)	571 35	572 (M+ + 1), 594 (M+ + Na)
19	455	F	(R)	571 35	572 (M+ + 1)
20	456	Cl	(S)	561.31	562 (M+ + 1), 584 (M+ + Na)
21	457	F	(R)	543.66	544 (M+ + 1)
22	458	F	(R)		531 (M+ + 1)
23	459	F	(S)		531 (M+ + 1)

[0391] 460

EXAMPLE 24

[0392] Step A

[0393] Compound 2-1 (0.090 g, 0.113 mmol) was dissolved in 0.30 mL of methylene chloride and 0.30 mL of trifluoroacetic acid. This solution was stirred for 30 min at room temperature. The mixture was then concentrated with toluene (3 mL×2) and diethyl ether (3 mL×2) to give a white solid. The solid was dissolved in 0.6 mL of methanol, and then sodium acetate (0.046 g, 0.565 mmol) and 37% aqueous formaldehyde solution (0.041 mL, 0.542 mmol) were added. The reaction mixture was stirred at room temperature for 30 min, and then sodium cyanoborohydride (1.0 M in THF, 0.36 mL, 0.36 mmol) was added. The mixture was stirred at room temperature overnight. The solution was concentrated to give a white sludge, and then dissolved in EtOAc (10 mL) and 1N NaOH (5 mL) and the layers were separated. The organic phase was washed with 1N NaOH (5 mL), H2O (5 mL), and brine (5 mL), dried over MgSO4, filtered, and concentrated to give an oil. Purification by column chromatography using 10% methanol in methylene chloride afforded 4-1 as a white, foamy-solid (0.080 g); mass spectrum: 708 (M+1); 730 (M+Na).

[0394] Step B

[0395] Compound 4-1 (0.080 g, 0.113 mmol) was dissolved in 0.5 mL of methylene chloride and 30% HBr in acetic acid (0.112 mL, 0.565 mmol) was added. The mixture was stirred at room temperature for 45 min (TLC showed no starting material). To this orange solution, diethyl ether was added and the precipitate was filtered and washed with ether. The solid was dissolved in EtOAc and washed with 1N NaOH, and the aqueous layer was extracted with EtOAc. The combined organics were dried over K2CO3, filtered and concentrated to give a foamy solid. Purification by column chromatography (5% -20% methanol in methylene chloride) provided a solid. The solid was dissolved in EtOAc and 1M HCl in Et2O (0.27 mL, 0.27 mmol) was added whereupon a precipitate formed. The precipitate was filtered under N2 and dried under vacuum to give 4-2 as a white solid (0.044 g). 461

[0396] The following Nα-methyl-Nβ-(Cbz) intermediates shown in Table 3 below above with the indicated stereochemistry at the stereogenic center marked with ** having variable X groups at the 4-position of the piperidine ring were prepared as shown in Scheme 5 above.

TABLE 3
			Exact
X	R6	**	Mass	Mass Spec.
462	F	(R)	750.40	751 (M+ + 1)
463	Cl	(S)	707.37	708 (M+ + 1), 730 (M+ + Na)
464	F	(S)	691.41	692 (M+ + 1), 714 (M+ + Na)
465	Cl	(R)	707.38	708 (M+ + 1)
466	F	(R)	691.41	692 (M+ + 1)
467	Cl	(S)	693.37	694 (M+ + 1), 716 (M+ + Na)
468	Cl	(S)	705.37	706 (M+ + 1)
469	Cl	(R)	705.37	706 (M+ + 1)
470	Cl	(S)	719.38	720 (M+ + 1), 742 (M+ + Na)
471	Cl	(R)	719.38	720 (M+ + 1)
472	Cl	(S)	721.40	722 (M+ + 1), 742 (M+ + Na)
473	Cl	(R)	721.40	722 (M+ + 1)
474	Cl	(R)	721.40	722 (M+ + 1)
475	Cl	(S)	728.35	729 (M+ + 1), 751 (M+ + Na)
476	Cl	(S)	733.32	734 (M+ + 1), 717 (M+ + Na)
477	F	(S)	838.44	839 (M+ + 1), 861 (M+ + Na)
478	F	(S)	718.42	719 (M+ + 1), 741 (M+ + Na)
479	Cl	(S)	707.38	708 (M+ + 1), 730 (M+ + Na)
480	Cl	(R)	707.38	708 (M+ + 1), 730 (M+ + Na)
481	Cl	(S)	735.38	736 (M+ + 1), 758 (M+ + Na)
482	F	(S)	719.41	720 (M+ + 1)
483	Cl	(R)	735.38	736 (M+ + 1), 758 (M+ + Na)
484	F	(R)	719.41	720 (M+ + 1)
485	Cl	(S)	709.36	710 (M+ + 1), 732 (M+ + Na)

[0397] The following Nα-methyl-Nβ-unsubstituted piperazine Examples shown in Table 4 below above with the indicated stereochemistry at the stereogenic center marked with ** were prepared from the intermediates in Table 3 by treatment with 30% HBr in acetic acid to afford the hydrobromide salt, which upon neutralization and subsequent treatment with HCl in diethyl ether afforded the hydrochloride salt.

TABLE 4
486
				Exact
Ex.	X	R6	**	Mass	Mass Spec.
25	487	F	(R)	530.33	531 (M+ + 1), 553 (M+ + Na)
26	488	Cl	(S)	573.34	574 (M+ + 1), 596 (M+ + Na)
27	489	F	(S)	557.37	558 (M+ + 1), 580 (M+ + Na)
28	490	Cl	(R)	573.34	574 (M+ + 1), 596 (M+ + Na)
29	491	F	(R)	557.37	558 (M+ + 1)
30	492	Cl	(S)	559.33	560 (M+ + 1), 682 (M+ + Na)
31	493	Cl	(S)	571.33	572 (M+ + 1)
32	494	Cl	(R)	571.33	572 (M+ + 1)
33	495	Cl	(S)	585.34	586 (M+ + 1), 608 (M+ + Na)
34	496	Cl	(R)	585.34	586 (M+ + 1)
35	497	Cl	(S)	587.36	588 (M+ + 1), 610 (M+ + Na)
36	498	Cl	(R)	587.36	588 (M+ + 1)
37	499	Cl	(R)	587.36	588 (M+ + 1)
38	500	Cl	(S)	594.31	595 (M+ + 1), 617 (M+ + Na)
39	501	Cl	(S)	599.29	600 (M+ + 1)
40	502	F	(S)	570.37	571 (M+ + 1), 593 (M+ + Na)
41	503	F	(S)	584.39	585 (M+ + 1), 607 (M+ + Na)
42	504	Cl	(S)	573.34	574 (M+ + 1), 596 (M+ + Na)
43	505	Cl	(R)	573.34	574 (M+ + 1), 596 (M+ + Na)
44	506	Cl	(S)	601.34	602 (M+ + 1), 624 (M+ + Na)
45	507	F	(S)	585.37	586 (M+ + 1)
46	508	Cl	(R)	601.34	602 (M+ + 1), 624 (M+ + Na)
47	509	F	(R)	585.37	586 (M+ + 1)
48	510	Cl	(S)	575.32	576 (M+ + 1), 598 (M+ + Na)

[0398] 511

EXAMPLE 49

[0399] Step A

[0400] Compound 2-2 (0.059 g, 0.105 mmol) was dissolved in 0.5 mL of methanol, and then sodium acetate (0.046 g, 0.565 mmol), trifluoroacetic acid (0.016 mL, 0.210 mmol), and 37% aqueous formaldehyde solution (0.093 mL, 1.008 mmol) were added. The reaction mixture was stirred at room temperature for 30 min, and then sodium cyanoborohydride (1.0 M in THF, 0.67 mL, 0.67 mmol) was added. The mixture was stirred at room temperature overnight, and then concentrated to give a white sludge. This was dissolved in EtOAc (10 mL) and 1N NaOH (5 mL) and the layers were separated. The organic phase was washed with 1N NaOH (5 mL), H2O (5 mL), and brine (5 mL), dried over MgSO4, filtered, and concentrated to give a white foamy solid. Purification by column chromatography on silica gel using 10% methanol in methylene chloride provided a white foamy solid (0.044 g).

[0401] Step B

[0402] The compound from Step A (0.040 g, 0.067 mmol) was dissolved in EtOAc, and 1.0 M HCl in Et2O (0.16 mL, 0.16 mmol) was added. The precipitate was filtered under N2 and dried under vacuum to give 6-1 as a white solid (0.031 g); mass spectrum: 588 (M+1); 610 (M+Na).

[0403] The following Nα,β-dimethyl-piperazine Examples shown in Table 5 below above with the indicated stereochemistry at the stereogenic center marked with ** were prepared by reductive methylation of the corresponding Nα,β-unsubstituted-piperazine intermediates, neutralization, and subsequent treatment with HCl in diethyl ether.

TABLE 5
512
				Exact
Ex.	X	R6	**	Mass	Mass Spec.
50	513	Cl	(S)	560.31	561 (M+ + 1), 583 (M+ + Na)
51	514	Cl	(S)	587.36	588 (M+ + 1), 610 (M+ + Na)
52	515	Cl	(R)	587.36	588 (M+ + 1)
53	516	Cl	(R)	599.36	600 (M+ + 1)
54	517	Cl	(R)	601.38	602 (M+ + 1)
55	518	Cl	(S)	587.36	602 (M+ + 1), 624 (M+ + Na)

[0404] 519

EXAMPLE 56

[0405] Step A

[0406] Intermediate 7-2 was prepared in a similar fashion as 1-11 but using (S)-4-methyl-1-(tert-butoxycarbonyl)-piperazine-2-carboxylic acid (7-1) in the coupling reaction with amine 1-5. (S)-4-Methyl-1-(tert-butoxycarbonyl)-piperazine-2-carboxylic acid (7-1) (61.0% purity, 0.162 g, 0.405 mmol) was dissolved in 1.8 mL of methylene chloride, and then the amine intermediate 1-5 (0.165 g, 0.368 mmol), NMM (0.16 mL, 1.472 mmol), EDC (0.078 g, 0.405 mmol), and HOBt (0.055 g, 0.405 mmol) were added. The resulting mixture was stirred at room temperature overnight, and then diluted with 10 mL of CH2Cl2 and washed with 5 mL of 1N HCl, 5 mL of saturated NaHCO3, 5 mL of H2O, and 5 mL of saturated NaCl solution, dried over MgSO4, filtered and concentrated to give a yellow oil. The crude product was purified by column chromatography on silica gel (9:1 to 1:1 methylene chloride-acetone) to give 7-2 as a white solid (0.219 g).

[0407] Step B

[0408] Compound 7-2 (0.219 g, 0.324 mmol) was dissolved in 0.80 mL of methylene chloride and 0.80 ml of trifluoroacetic acid. This solution was stirred for 30 min at room temperature, and then concentrated with toluene (5 mL×2) and diethyl ether (5 mL×2) to give a white foamy solid. The solid was dissolved in EtOAc and washed with 1N NaOH, and the aqueous layer was extracted with EtOAc. The combined organics were dried over K2CO3, filtered and concentrated. Purification by column chromatography (5% -10% methanol in methylene chloride) provided a white foamy solid. The solid was dissolved in EtOAc and 1.0 M HCl in Et2O (0.78 mL, 0.78 mmol) was added. The precipitate was filtered under N2 and dried under vacuum to give 7-3 as a white solid (0.083 g); mass spectrum: 574 (M+1); 596 (M+Na).

[0409] The Nα-(tert-butoxycarbonyl)-Nβ-substituted-piperazine-2-carboxylic acid amide intermediates shown in Table 6 below were prepared by reductive alkylation of Nα-(tert-butoxycarbonyl)-piperazine-2-carboxylic acid methyl ester with the appropriate aldehyde or nucleophilic displacement of an alkyl halide with Nα-(tert-butoxycarbonyl)-piperazine-2-carboxylic acid methyl ester, followed by saponification, and coupling with the appropriate substituted N-piperidinyl-Phe amine intermediate. Alternatively, Nα-(tert-butoxycarbonyl)-Nβ-(Cbz)-piperazine-2-carboxylic acid was EDC-coupled with the substituted N-piperidinyl-4-halo-Phe amine intermediate, the Cbz group cleaved by hydrogenolysis (H2, Pd/C), and then either reductive alkylation of the β-piperazine nitrogen with the appropriate aldehyde or nucleophilic displacement of an alkyl halide.

TABLE 6
520
X	R6	**	R4b	Exact Mass	Mass Spec.
521	Cl	(S)	i-Pr	674.38	675 (M+ + 1)
522	F	(S)	2-hydroxyethyl	660.39	661 (M+ + 1)
523	F	(S)	2-methoxyethyl	674.41	675 (M+ + 1), 697(M+ + Na)
524	F	(S)	CH2CO2Et	702.40	703 (M+ + 1), 725 (M+ + Na)
525	F	(R)	Me	630.38	631 (M+ + 1), 653 (M+ + Na)
526	F	(R)	2-methoxyethyl	674.41	675 (M+ + 1), 697 (M+ + Na)
527	Cl	(S)	i-Pr	701.43	702 (M+ + 1)
528	Cl	(S)	2,2-difluoroethyl	723.39	724 (M+ + 1)
529	F	(S)	Me	657.43	658 (M+ + 1)
530	F	(S)	i-Pr	685.46	686 (M+ + 1)
531	F	(S)	2,2-difluoroethyl	707.42	708 (M+ + 1)
532	Cl	(R)	Me	673.40	674 (M+ + 1), 696 (M+ + Na)
533	Cl	(R)	i-Pr	701.43	702 (M+ + 1)
534	Cl	(R)	cyclopropylmethyl	713.43	714 (M+ + 1)
535	Cl	(R)	benzyl	749.43	750 (M+ + 1)
536	Cl	(R)	2-propynyl	697.40	698 (M+ + 1)
537	Cl	(R)	cyclobutyl	713.43	714 (M+ + 1)
538	Cl	(R)	2,2-difluoroethyl	723.39	724 (M+ + 1)
539	F	(R)	Me	657.43	658 (M+ + 1)
540	F	(R)	i-Pr	685.46	686 (M+ + 1)
541	F	(R)	cyclopropylmethyl	697.46	698 (M+ + 1)
542	F	(R)	2,2-difluoroethyl	707.42	708 (M+ + 1)
543	Cl	(S)	Me	659.38	660 (M+ + 1)
544	Cl	(S)	Me	671.38	672 (M+ + 1)
545	Cl	(R)	Me	671.38	672 (M+ + 1)
546	Cl	(S)	Me	685.40	686 (M+ + 1)
547	Cl	(R)	Me	685.40	686 (M+ + 1)
548	Cl	(S)	Me	687.41	688 (M+ + 1)
549	Cl	(R)	Me	687.41	688 (M+ + 1)
550	Cl	(R)	Me	687.41	688 (M+ + 1)
551	Cl	(S)	Me	694.36	695 (M+ + 1), 717 (M+ + Na)
552	F	(S)	Me	684.44	685 (M+ + 1), 707 (M+ + Na)
553	Cl	(S)	Me	673.40	674 (M+ + 1), 696 (M+ + Na)
554	Cl	(R)	Me	673.40	674 (M+ + 1), 696 (M+ + Na)
555	F	(S)	Me	685.42	686 (M+ + 1)
556	F	(R)	Me	685.42	686 (M+ + 1)
557	F	(R)	i-Pr	713.45	714 (M+ + 1)
558	F	(S)	Et		672 (M+ + 1)
559	F	(R)	H		672 (M+ + 1)

[0410] The following Nα-unsubstituted-Nβ-(R4b)-piperazine Examples shown in Table 7 below above with the indicated stereochemistry at the stereogenic center marked with ** were prepared as their bis-hydrochloride salts by cleavage of the Nα-(Boc) group in the corresponding compounds in Table 6 to afford the trifluoroacetate salt, which upon neutralization and subsequent treatment with HCl in diethyl ether afforded the hydrochloride salt.

TABLE 7
560
Ex.	X	R6	**	R4b		Mass Spec.
57	561	Cl	(S)	i-Pr	574.33	575 (M+ + 1), 597 (M+ + Na)
58	562	F	(S)	2-hydroxyethyl	560.34	561 (M+ + 1), 583 (M+ + Na)
59	563	F	(S)	2-methoxyethyl	574.35	575 (M+ + 1), 597 (M+ + Na)
60	564	F	(S)	CH2CO2Et	602.35	603 (M+ + 1), 625 (M+ + Na)
61	565	F	(R)	Me	530.33	531 (M+ + 1), 553 (M+ + Na)
62	566	F	(R)	2-methoxyethyl	574.35	575 (M+ + 1), 597 (M+ + Na)
63	567	Cl	(S)	Me	573.34	574 (M+ + 1), 596 (M+ + Na)
64	568	Cl	(S)	i-Pr	601.38	602 (M+ + 1), 624 (M+ + Na)
65	569	Cl	(S)	2,2-difluoroethyl	623.34	624 (M+ + 1)
66	570	F	(S)	Me	557.37	558 (M+ + 1), 580 (M+ + Na)
67	571	F	(S)	i-Pr	585.41	586 (M+ + 1)
68	572	F	(S)	2,2-difluoroethyl	607.37	608 (M+ + 1)
69	573	Cl	(R)	Me	573.34	574 (M+ + 1), 596 (M+ + Na)
70	574	Cl	(R)	i-Pr	601.38	602 (M+ + 1)
71	575	Cl	(R)	cyclopropylmethyl	613.38	614 (M+ + 1)
72	576	Cl	(R)	benzyl	649.38	650 (M+ + 1)
73	577	Cl	(R)	2-propynyl	597.34	598 (M+ + 1)
74	578	Cl	(R)	cyclobutyl	613.38	614 (M+ + 1)
75	579	Cl	(R)	2,2-difluoroethyl	623.34	624 (M+ + 1)
76	580	F	(R)	Me	557.37	558 (M+ + 1), 580 (M+ + Na)
77	581	F	(R)	i-Pr	585.41	586 (M+ + 1)
78	582	F	(R)	cyclopropylmethyl	597.41	598 (M+ + 1)
79	583	F	(R)	2,2-difluoroethyl	607.37	608 (M+ + 1)
80	584	Cl	(S)	Me	559.33	560 (M+ + 1), 682 (M+ + Na)
81	585	Cl	(S)	Me	571.33	572 (M+ + 1)
82	586	Cl	(R)	Me	571.33	572 (M+ + 1)
83	587	Cl	(S)	Me	585.34	586 (M+ + 1)
84	588	Cl	(R)	Me	585.34	586 (M+ + 1)
85	589	Cl	(S)	Me	587.36	588 (M+ + 1)
86	590	Cl	(R)	Me	587.36	588 (M+ + 1)
87	591	Cl	(R)	Me	587.36	588 (M+ + 1)
88	592	Cl	(S)	Me	594.31	595 (M+ + 1), 617 (M+ + Na)
89	593	F	(S)	Me	584.39	585 (M+ + 1), 607 (M+ + Na)
90	594	Cl	(S)	Me	573.34	574 (M+ + 1), 596 (M+ + Na)
91	595	Cl	(R)	Me	573.34	574 (M+ + 1), 596 (M+ + Na)
92	596	Cl	(S)	Me	601.34	602 (M+ + 1), 624 (M+ + Na)
93	597	Cl	(R)	Me	601.34	602 (M+ + 1), 624 (M+ + Na)
94	598	F	(S)	Me	585.37	586 (M+ + 1)
95	599	F	(R)	Me	585.37	586 (M+ + 1)
96	600	F	(R)	i-Pr	613.40	614 (M+ + 1)
97	601	F	(S)	Et		572 (M+ + 1)
98	602	F	(R)	Me		545 (M+ + 1)
99	603	F	(S)	Me		545 (M+ + 1)

[0411] 604

EXAMPLE 100

[0412] Step A: 4-Phenyl-1,4-piperidinedicarboxylic Acid 1-(1,1-dimethylethyl) Ester (8-2)

[0413] A 12-L, three-necked, round-bottomed flask equipped with a mechanical stirrer was charged with commercially available 4-phenyl-4-piperidinecarboxylic acid p-methylbenzenesulfonate (8-1) (500 g, 1.32 mol), di-tert-butyl dicarbonate (318 g, 1.46 mol), 3000 mL of 1 N NaOH solution (3.0 mol), and 3000 mL of dioxane. After addition, the pH was adjusted to 11-12 using 5 N NaOH solution, and the resulting mixture was stirred at room temperature overnight. The mixture was then concentrated and acidified using 2 N HCl solution to about pH 1. The precipitate was filtered using 2 L of water to wash and dried to give 418 g of title compound 8-2 as a white solid.

[0414] Step B: 4-Cyclohexyl-4-piperidinecarboxylic Acid Hydrochloride (8-3)

[0415] 4-Phenyl-1,4-piperidinedicarboxylic acid 1-(1,1-dimethylethyl) ester (8-2) (202 g, 0.662 mol) was dissolved in 1700 mL of 10% HCl in methanol and rhodium on alumina (25 g) was added. The mixture was placed on a high pressure hydrogenator using 1500 psi of hydrogen at 100° C. for 17 h. The resulting mixture was filtered through Celite using methanol to rinse and concentrated. The resulting solid was triturated with diethyl ether and filtered to afford title compound 8-3 as an off-white solid.

[0416] Step C: 4-Cyclohexyl-1,4-piperidinedicarboxylic Acid 1-(phenylmethyl) Ester (8-4)

[0417] A 2-L, three-necked, round-bottomed flask equipped with a mechanical stirrer and two addition funnels was charged with 4-cyclohexyl-4-piperidinecarboxylic acid hydrochloride (8-3) (157.9 g, 0.637 mol), 1 L of dioxane, and 255 mL of 5 N NaOH solution. The mixture was cooled at about 5° C., and then benzyl chloroformate (92 mL, 0.643 mmol) and 127 mL of 5 N NaOH solution were added dropwise simultaneously via two separate addition funnels while maintaining the temperature at or below 10° C. The reaction was monitored by TLC and, upon completion, the resulting mixture was diluted with 1 L of water and concentrated. The residue was diluted with 2 L of water and the pH was adjusted to about 12 using 5 N NaOH solution. The mixture was then extracted with 1 L of ethyl acetate. The aqueous layer was acidified to pH 1.5-2.0 using 2 N HCl solution, and extracted with three 1-L portions of ethyl acetate. The combined organic layers were washed with 1 L of saturated sodium chloride solution, dried over sodium sulfate, filtered and concentrated to give 165 g of title compound 8-4 as a white solid.

[0418] Step D: 4-Cyclohexyl-4-[[(1,1-dimethylethyl)amino]carbonyl]-1-piperidinecarboxylic Acid Phenylmethyl Ester (8-5)

[0419] 4-Cyclohexyl-1,4-piperidinedicarboxylic acid 1-(phenylmethyl) ester (8-4) (2.50 g, 7.24 mmol) was dissolved in 36 mL of methylene chloride and cooled at 0° C. in an ice-water bath. Oxalyl chloride (2.0 M solution in CH2Cl2, 3.98 mL, 7.96 mmol) was then added dropwise followed by the addition of 1-2 drops of DMF. This mixture was stirred at 0° C. for 2 h and then concentrated with toluene. The residue was dissolved in 36 mL of methylene chloride and cooled at 0° C. in an ice-water bath. tert-Butylamine (2.28 mL, 21.7 mmol) was then added dropwise, and the reaction mixture was stirred at 0° C. for 2 h, warmed to room temperature, and stirred at room temperature overnight. The resulting mixture was diluted with methylene chloride, washed with brine, dried over MgSO4, filtered, and concentrated to give 2.92 g of title compound 8-5 as a solid. LCMS (ESI): m/z 401 (M++1).

[0420] Step E: 4-Cyclohexyl-N-(1,1-dimethylethyl)-4-piperidinecarboxamide (8-6)

[0421] 4-Cyclohexyl-4-[[(1,1-dimethylethyl)amino]carbonyl]-1-piperidinecarboxylic acid phenylmethyl ester (8-5) (10.0 g, 25.0 mmol) was dissolved in 130 mL of ethyl alcohol and 10% palladium on carbon (1 g) was added. The mixture was evacuated and purged with hydrogen three times and then stirred at room temperature overnight. The resulting mixture was filtered through Celite using methylene chloride to rinse and concentrated to afford 6.22 g of title compound 876 as a white solid. This crude product was used in the next reaction without further purification. LCMS (ESI): m/z 267 (M++1). 1H NMR (500 MHz, CDCl3) δ 2.99 (d, J=12.6 Hz, 2H), 2.71 (dd, J=12.3, 11.5 Hz, 2H), 1.93 (d, J=12.8 Hz, 2H), 1.79-1.73 (m, 3H), 1.65, (d, J=12.1 Hz, 1H), 1.51-1.45 (m, 2H), 1.36 (s, 9H), 1.33-0.94 (m, 7H).

[0422] Step F: [(1R)-2-[4-Cyclohexyl-4-[[(1,1-dimethylethyl)amino]carbonyl]-1-piperidinyl]-1-[(4-fluorophenyl)methyl]-2-oxoethyl]carbamic Acid 1,1-dimethylethyl Ester (8-7)

[0423] N-Boc-(D)-4-fluorophenylalanine (7.26 g, 25.63 mmol) was dissolved in 116.5 mL of methylene chloride, and amine 8-6 (6.21 g, 23.3 mmol), DIEA (16.2 mL, 93.2 mmol), EDC.HCl (4.91 g, 25.6 mmol), and HOBt (3.46 g, 25.6 mmol) were added. The resulting mixture was stirred at room temperature overnight and then diluted with 100 mL of methylene chloride. The mixture was washed with 100 mL of 1 N HCl solution, 100 mL of saturated NaHCO3 solution, 100 mL of water, and 100 mL of saturated NaCl solution, dried over MgSO4, filtered, and concentrated to give a white foamy-solid. The crude product was purified by column chromatography (30:1 to 9:1 methylene chloride-acetone) to give 10.1 g (81%) of title compound 8-7 as a white solid. LCMS (ESI): m/z 532 (M++1). 1H NMR (500 MHz, CDCl3) (mixture of two rotamers) δ 7.20-6.90 (m), 5.45 (d, J=8.7 Hz), 5.36 (d, J=8.5 Hz), 5.20 (d, J=5.8 Hz), 4.84-4.77 (m), 4.42 (d, J=12.9 Hz), 3.56 (m), 3.04 (dd, J=12.8, 12.3 Hz), 2.95 (d, J=7.3 Hz), 2.89 (d, J=6.9 Hz), 2.61-2.54 (m), 1.87-1.54, (m), 1.41 (s), 1.35 (s), 1.32 (s), 1.28-0.80 (m), 0.33-0.28 (m).

[0424] Steps G and H: (2S)-2-[[[(1R)-2-[4-cyclohexyl-4-[[(1,1-dimethylethyl)amino]carbonyl]-1-piperidinyl]-1-[(4-fluorophenyl)methyl]-2-oxoethyl]amino]carbonyl]-1,4-piperazinedicarboxylic Acid 1-(1,1-dimethylethyl) 4-(phenylmethyl) Ester (8-8)

[0425] Step G

[0426] Compound 8-7 (5.33 g, 10.0 mmol) was dissolved in 25.1 mL of methylene chloride and 25.1 mL of trifluoroacetic acid. This solution was stirred at room temperature for 30 min. The mixture was then concentrated with four 30-mL portions of methylene chloride to give a white foamy solid. The solid was dissolved in ethyl acetate and washed with 1 N NaOH solution, and the aqueous layer was extracted with ethyl acetate. The combined organics were dried over K2CO3, filtered, and concentrated to give 4.04 g of a white foamy-solid. This crude product was used in the next reaction without further purification. LCMS (ESI): m/z 432 (M++1). 1H NMR (500 MHz, CDCl3) (mixture of two rotamers) δ 7.28-6.24 (m), 5.25 (d, J=8.2 Hz), 4.51 (d, J=13.5 Hz), 3.96-3.90 (m), 3.62 (d, J=13.5 Hz), 3.55 (d, J=13.7 Hz), 3.10-3.05 (m), 2.93-2.81 (m), 2.74-2.69 (m), 2.62-2.57 (m), 2.00 (d, J=13.5 Hz), 1.85-1.47 (m), 1.37 (s), 1.35 (s), 1.33-0.81 (m), 0.41-0.35 (m).

[0427] Step H

[0428] (2S)-1,2,4-piperazinetricarboxylic acid 1-(1,1-dimethylethyl) 4-(phenylmethyl) ester (the preparation of this intermediate from commercially available 2-(S)-piperazine carboxylic acid was achieved via modifications to the procedures described by Bigge and coworkers in Tetrahedron Lett. 1989, 30, 5193) (3.73 g, 10.3 mmol) was dissolved in 47 mL of methylene chloride, and the crude amine intermediate (4.02 g, 9.31 mmol), DIEA (6.49 mL, 37.2 mmol), EDC.HCl (1.97 g, 10.3 mmol), and HOBt (1.385 g, 10.3 mmol) were added. The resulting mixture was stirred at room temperature overnight and then diluted with 100 mL of methylene chloride. The mixture was washed with 100 mL of 1 N HCl solution, 100 mL of saturated NaHCO3 solution, 100 mL of water, and 100 mL of saturated NaCl solution, dried over MgSO4, filtered, and concentrated to give an off-white, foamy solid. The crude product was purified by column chromatography (30:1 to 3:1 methylene chloride-acetone) to give 5.77 g (80%) of title compound 8-8 as a white solid. LCMS (ESI): m/z 778 (M++1). 1H NMR (500 MHz, CDCl3) (mixture of two rotamers) δ 7.37-6.79 (m), 5.30-5.00 (m), 4.71-4.50 (m), 4.40 (d, J=10.5 Hz), 4.04-3.79 (br s), 3.53 (d, J=12.6 Hz), 3.13-2.79 (m), 2.60-2.55 (m), 1.88-1.57 (m), 1.47 (s), 1.35 (s), 1.33 (s), 1.17-0.81 (m), 0.41-0.23 (m).

[0429] Steps I and J: (2S)-2-[[[(R)-2-[4-Cyclohexyl-4-[[(1,1-dimethylethyl)amino]carbonyl]-1-piperidinyl]-1-[(4-fluorophenyl)methyl]-2-oxoethyl]amino]carbonyl]-4-methyl-1-piperazinecarboxylic Acid 1,1-dimethylethyl Ester (8-9)

[0430] Step I

[0431] Compound 8-8 (5.77 g, 7.41 mmol) was dissolved in 37 mL of ethyl alcohol and 10% palladium on carbon (0.577 g) was added. The mixture was then evacuated and purged with hydrogen three times and stirred at room temperature overnight. The resulting mixture was filtered through Celite using methylene chloride to rinse and concentrated to give 4.28 g of a white solid. This crude product was used in the next reaction without further purification. LCMS (ESI): m/z 644 (M++1). 1H NMR (500 MHz, CDCl3) (mixture of two rotamers) δ 7.19-6.92 (m), 5.21 (d, J=5.1 Hz), 5.17-5.10 (m), 4.60-4.46 (m), 4.41 (d, J=9.4 Hz), 3.98-3.85 (m), 3.60-3.46 (m), 3.06 (app t, J=13.2 Hz), 2.97-2.91 (m), 2.78 (d, J=12.8 Hz), 2.68-2.57 (m), 1.90-1.58 (m), 1.48 (s), 1.42-1.33 (m), 1.2-0.82 (m), 0.44-0.38 (m).

[0432] Step J

[0433] The crude amine intermediate from Step I (3.28 g, 5.10 mmol) was dissolved in 25.5 mL of methanol, and sodium acetate (2.09 g, 25.5 mmol), trifluoroacetic acid (0.39 mL, 5.10 mmol), and 37% aqueous formaldehyde solution (1.83 mL, 24.5 mmol) were added. The reaction mixture was stirred at room temperature for 20 min and then sodium cyanoborohydride (1.0 M in THF, 16.3 mL, 16.3 mmol) was added. The resulting mixture was stirred at room temperature overnight and then concentrated to give a white sludge. The crude mixture was dissolved in 30 mL of ethyl acetate and 15 mL of 1 N NaOH solution, and the layers were separated. The organic phase was washed with 15 mL of 1 N NaOH solution, 15 mL of water, and 15 mL of brine, dried over MgSO4, filtered, and concentrated to give a solid. Purification by column chromatography (3% methanol in methylene chloride) afforded 2.84 g of title compound 8-9 as a white solid. LCMS (ESI): m/z 658 (M++1). 1H NMR (500 MHz, CDCl3) (mixture of two rotamers) δ 7.18-6.91 m), 5.21 (d, J=7.3 Hz), 5.18-5.12 (m), 4.73-4.35 (m), 4.05-3.92 (m), 3.61-3.29 (m), 3.08-2.99 (m), 2.96-2.91 (m), 2.80-2.56 (m), 2.26 (s), 2.25 (s), 2.05-2.02 (m), 1.93-1.53 (m), 1.48 (s), 1.44-1.32 (m), 1.21-0.83 (m), 0.49-0.38 (m).

[0434] Steps K and L: (2S)-N-[(1R)-2-[4-Cyclohexyl-4-[[(1,1-dimethylethyl)amino]carbonyl]-1-piperidinyl]-1-[(4-fluorophenyl)methyl]-2-oxoethyl]-4-methyl-2-piperazinecarboxamide Dihydrochloride (8-11)

[0435] Step K

[0436] Compound 8-9 (2.82 g, 4.29 mmol) was dissolved in 10.7 mL of methylene chloride and 10.7 mL of trifluoroacetic acid. This solution was stirred at room temperature for 30 min. The mixture was then concentrated with four 30-mL portions of methylene chloride to give a white foamy solid. The solid was dissolved in ethyl acetate and washed with 1 N NaOH solution, and the aqueous layer was extracted with ethyl acetate. The combined organics were dried over K2CO3, filtered, and concentrated to give 2.18 g of 8-10 as a white foamy-solid. This crude product was used in the next reaction without further purification. LCMS (ESI): m/z 558 (M++1); 1H NMR (600 MHz, CD3CN) (mixture of two rotamers) δ 2.53; 2.51 (H-2ax); 4.28; 4.26 (H-2eq); 1.28; 1.11 (H-3ax); 1.93; 1.27 (H-3eq); 1.31; 0.68 (H-5ax); 1.90; 1.83 (H-5eq); 2.75; 2.97 (H-6ax); 3.72; 3.66 (H-6eq); 5.74; 5.72 (7NH); 1.30 (H-9); 4.99 (H-2′); 7.55; 7.63 (2′NH); 2.91 (H-3′a); 2.84 (H-3′b); 7.18; 7.12 (H-5′); 7.01; 6.96 (H-6′); 3.22 (H-2″); 2.50 (H-3″a); 2.08 (H-3″b); 2.32 (H-5″a); 2.00 (H-5″b); 2.80 (H-6″a); 2.69 (H-6″b); 2.13 (NMe); 1.20 (H-1′″); 1.61 (H-2′″a); 0.90 (H-2′″b); 1.72 (H-3′″, H-4′″a); 1.14 (H-3′″, H-4′″b); 1.04 (H-4′″, H-3′″a); and 0.91 (H-4′″; H-3′″b).

[0437] Step L

[0438] The crude amine from Step K (1.89 g, 3.39 mmol) was dissolved in 8 mL of methylene chloride and 1.0 M HCl in ethyl ether (8.13 mL, 8.13 mmol) was added. The precipitate was filtered and dried under vacuum to give 2.09 g as a white solid. LCMS (ESI): m/z 558 (M++1). 605

EXAMPLE 101

[0439] Step A

[0440] Compound 9-1 was hydrogenated to remove the Cbz group. N-tert-Butoxycarbonyl-(D)-alanine (0.050 g, 0.263 mmol) was dissolved in 1.2 mL of methylene chloride, and then the piperazine intermediate obtained after cleavage of the Cbz group above (0.154 g, 0.239 mmol), NMM (0.11 mL, 0.956 mmol), EDC (0.050 g, 0.263 mmol), and HOBt (0.036 g, 0.263 mmol) were added. The resulting mixture was stirred at room temperature overnight, and then diluted with 10 mL of CH2Cl2 and washed with 5 mL of 1N HCl, 5 mL of saturated NaHCO3, 5 mL of H2O, and 5 mL of saturated NaCl solution, dried over MgSO4, filtered and concentrated to give a yellow oil. The crude product was purified by column chromatography (30:1 to 9:1 methylene chloride-acetone) to give a white solid (0.18 g). Mass spectrum: Calcd for C43H67N6O8F: 814.5; Found: 815 (M++1), 715 (M+-Boc).

[0441] Step B

[0442] The bis-Boc intermediate was dissolved in 0.54 mL of methylene chloride and 0.54 mL of trifluoroacetic acid. This solution was stirred for 30 min at room temperature, and then concentrated with toluene (3 mL×2) and diethyl ether (3 mL×2) to give 9-2 as a TFA salt (0.22 g). Mass spectrum: Calcd for C33H51N6O4F: 614.40; Found: 615 (M++1). 606

EXAMPLE 102

[0443] Intermediate 10-1 (0.066 g, 0.093 mmol) was dissolved in 0.24 mL of methylene chloride and 0.24 ml of trifluoroacetic acid. This solution was stirred for 30 min at room temperature, and then concentrated with toluene (5 mL×2) and diethyl ether (5 mL×2). The resulting foam was dissolved in 0.5 mL of methanol, and then sodium acetate (0.038 g, 0.465 mmol) and 37% aqueous formaldehyde solution (0.033 mL, 0.446 mmol) were added. The mixture was stirred at room temperature for 30 min, and then sodium cyanoborohydride (1.0 M in THF, 0.30 mL, 0.30 mmol) was added. The reaction mixture was stirred at room temperature overnight, and then concentrated to give a white sludge. The crude mixture was dissolved in EtOAc (10 mL) and 1N NaOH (5 mL), and the layers were separated. The organic phase was washed with 1N NaOH (5 mL), H2O (5 mL), and brine (5 mL), dried over MgSO4, filtered, and concentrated. Purification by column chromatography using 10% methanol in methylene chloride provided a white foamy solid. The solid was dissolved in EtOAc, and 1.0 M HCl in Et2O (0.23 mL, 0.23 mmol) was added. The precipitate was filtered under N2 and dried under vacuum to give 10-2 as a white solid (0.043 g); mass spectrum: Calcd for C33H54N5O3Cl: 615.39; Found: 616 (M++1).

EXAMPLE 103

[0444] 607

[0445] Example 103 was prepared in a similar fashion as Example 102, but using the 4-cyclohexyl-4-(ethoxycarbonyl)-piperidine intermediate instead; mass spectrum: Calcd for C32H49N4O4Cl: 588.34; Found: 589 (M++1). 608

EXAMPLE 104

[0446] Step A

[0447] A solution of methyl (R)-1-(tert-butoxycarbonyl) piperazine-2-carboxylate (11-1) (0.37 g, 1.52 mmol) in 5 mL of anhydrous THF was treated with a solution of LDA (1.5 M in cyclohexane, 2.02 mL, 3.04 mmol) at −78° C. under N2. After 0.5 h, a solution of methyl iodide (0.28 mL, 4.56 mmol) in THF was added. The mixture was stirred at −78° C. for 2.5 h, warmed up to room temperature, and stirred for 3 days. The resulting mixture was diluted with EtOAc and washed with sat. NH4Cl solution and brine. The organic phase was dried over MgSO4, filtered and concentrated. Purification by column chromatography (30% ethyl acetate-hexane) provided 11-2 (0.024 g). Mass spectrum: Calcd for C13H24N2O4: 272.17; Found: 273 (M++1).

[0448] Step B

[0449] Ester 11-2 (0.022 g, 0.081 mmol) was dissolved in 0.5 mL of methanol and lithium hydroxide in 0.5 mL of water was added. The mixture was stirred at 50° C. overnight, and then concentrated. The residue was dissolved in water and then the pH was adjusted to about 6. The solution was then concentrated twice with toluene to give 11-3 as a solid. Mass spectrum: Calcd for C12H22N2O4: 258.16; Found: 259 (M++1).

[0450] Step C

[0451] The crude acid 11-3 (0.081 mmol) was dissolved in 0.4 mL of methylene chloride, and then the amine intermediate 1-5 (0.033 g, 0.074 mmol), NMM (0.033 mL, 0.296 mmol), EDC (0.016 g, 0.074 mmol), and HOBt (0.011 g, 0.074 mmol) were added. The resulting mixture was stirred at room temperature overnight, and then diluted with 5 mL of CH2Cl2 and washed with 2 mL of 1N HCl, 2 mL of saturated NaHCO3, 2 mL of H2O, and 2 mL of saturated NaCl solution, dried over MgSO4, filtered and concentrated to give a yellow oil. The crude product was purified by column chromatography (30:1 to 9:1 methylene chloride-acetone) to give 11-4 as a white solid (0.012 g). Mass spectrum: Calcd for C37H58N5O5Cl: 687.41; Found: 688 (M++1), 710 (M++Na).

[0452] Step D

[0453] Intermediate 11-4 was dissolved in 0.05 mL of methylene chloride and 0.05 mL of trifluoroacetic acid, and the solution was stirred for 30 min at room temperature. The resulting mixture was then concentrated with toluene (2 mL×2) and diethyl ether (2 mL×2) to give 11-5 as a bis-TFA salt (0.012 g). Mass spectrum: Calcd for C32H50N5O3Cl: 587.36; Found: 588 (M++1), 610 (M++Na). 609

EXAMPLES 105-111

[0454] Examples 105-111 were prepared according to Scheme 12 above. The required bridged piperazine intermediate 12-3 was prepared as follows:

[0455] Step A

[0456] Piperazine (1.0 g, 11.61 mmol) was charged with 50 mL of toluene, triethylamine (3.24 mL, 23.22 mmol), and ethyl dibromopropionate (1.69 mL,11.61 mmol). The mixture was then heated at 80° C. and stirred overnight. The resulting white precipitate was filtered and the filtrate was concentrated to give an oil. Purification by column chromatography (3-10% methanol in methylene chloride) provided the bridged piperazine ester 12-2 (1.20 g). Mass spectrum: Calcd for C9H16N2O2: 184.12; Found: 185 (M++1).

[0457] Step B

[0458] Ester 12-2 was dissolved in methanol and 1N NaOH solution was added. The mixture was stirred at room temperature overnight, and then concentrated. The residue was dissolved in water and then the pH was adjusted to 6. The solution was then concentrated twice with toluene to give the bridged piperazine acid 12-3 (1.81 g, 50.2% purity). Mass spectrum: Calcd for C7H12N2O2: 156.09; Found: 157 (M++1).

[0459] Step C

[0460] Bridged piperazine acid 12-3 was coupled with the appropriate amine, and the isolated product was treated with HCl in diethyl ether to afford Examples 105-111 as their bis-hydrochloride salts.

TABLE 8
610
Example	X	R6	Exact mass	Mass Spec
105	611	Cl	558.30	559 (M+ + 1)
106	612	F	542.33	543 (M+ + 1)
107	613	Cl	585.34	586 (M+ + 1)
108	614	F	569.37	570 (M+ + 1)
109	615	Cl	613.34	614 (M+ + 1)
110	616	F	597.37	598 (M+ + 1)
111	617	Cl	567.31	568 (M+ + 1), 590 (M+ + Na)

[0461] 618

[0462] Example 112 was prepared in a similar fashion as Examples 105-111 but using the D-3-(2-naphthyl)alanine-derived amine intermediate for the coupling reaction. Mass spectrum: Calcd for C34H46N4O4: 574.35; Found: 575 (M++1). 619

EXAMPLES 113-114

[0463] Examples 113 and 114 were prepared according to Scheme 12 above. The required saturated quinoxaline intermediate 13-4 was prepared as follows:

[0464] Step A

[0465] Cis-1,2-diaminocyclohexane (13-1) (2.95 g, 25.87 mmol) was charged with 45 mL of THF, benzaldehyde (5.78 mL, 56.91 mmol) and MgSO4 (1.61 g), and stirred at room temperature for 2 h. The mixture was filtered and the solution was concentrated. The residue was dissolved in methanol (129.35 mL) and sodium acetate was added. After 20 min, sodium cyanoborohydride (1.0 M in THF, 77.6 mL, 77.6 mmol) was added, and the reaction mixture was stirred at room temperature overnight. The mixture was then concentrated, diluted with ethyl acetate, and washed with 1N aq. NaOH solution and brine, dried over MgSO4, filtered and concentrated. Purification by column chromatography (9:1 CH2Cl2/acetone) afforded 13-2 as a yellow oil (0.51 g). Mass spectrum: Calcd for C20H26N2: 294.21; Found: 295 (M++1).

[0466] Step B

[0467] Dibenzylamine 13-2 (0.498 g, 1.693 mmol) was charged with 8.5 mL of toluene and triethylamine (0.47 mL, 3.386 mmol). The mixture was heated at 40° C. and then ethyl 2,3-dibromopropionate (0.25 mL, 1.693 mmol) was added. The reaction mixture was then stirred at 80° C. overnight, filtered, and concentrated. Purification by column chromatography (30:1 CH2Cl2/acetone) provided 13-3 as a yellow solid (0.215 g). Mass spectrum: Calcd for C25H32N2O2: 392.25; Found: 393 (M++1).

[0468] Step C

[0469] Ester 13-3 (0.215 g, 0.55 mmol) was charged with 2.75 mL of methanol and a solution of lithium hydroxide (0.026 g, 1.10 mmol) in 0.5 mL of water. The mixture was stirred at 50° C. for 2 days and concentrated. The residue was dissolved in water and the pH was adjusted to 6. The mixture was then concentrated with toluene twice to give 13-4 as a solid (0.351 g, 57% purity). Mass spectrum: Calcd for C23H28N2O2: 364.22; Found: 365 (M++1).

TABLE 9
620
Example	X	Exact mass	Mass Spec
113	621	777.50	778 (M+ + 1)
114	622	750.45	751 (M+ + 1)

EXAMPLE 115

[0470] 623

[0471] Example 113 (0.021 g, 0.027 mmol) was charged with 0.14 mL of EtOH and 10% Pd/C (0.0063 g). A H2 balloon was placed on the top of the condenser via a 3-way stopcock, and the system was evacuated and purged with H2 three times. The mixture was then stirred at 60° C. under H2 overnight. The oil bath was removed and the flask was evacuated and purged with N2 three times. The reaction mixture was filtered through a pad of Celite and concentrated to give a foamy solid. The solid was dissolved in CH2Cl2 and 1.0 M HCl in diethyl ether was added. The mixture was then concentrated to give Example 115 as a solid (0.017 g). Mass spectrum: Calcd for C36H56N5O3F: 625.44; Found: 626 (M++1).

EXAMPLE 116

[0472] 624

[0473] Example 114 (0.021 g, 0.027 mmol) was charged with 0.14 mL of EtOH, 3.0 M HCl (0.12 mL, 0.122 mmol), and 10% Pd/C (0.0063 g). A H2 balloon was placed on the top of the condenser via 3-way stop-cock and the system was evacuated and purged with H2 three times. The mixture was then stirred at room temperature under H2 for 5 h. The flask was evacuated and purged with N2 three times, and then the reaction mixture was filtered through a pad of Celite and concentrated to give Example 116 as a solid (0.010 g). Mass spectrum: Calcd for C32H7N4O3F: 570.36; Found: 571 (M++1). 625

EXAMPLE 117

[0474] Step A

[0475] A 100-mL, three-necked, round-bottomed flask equipped with a condenser was charged with 3-methyl-2-butanone (14-1) (4.20 mL, 39.26 mmol) and 24 mL of methanol. The solution was stirred and cooled in an ice-H2O bath at 0-5° C., and bromine (2.02 mL, 39.26 mmol) was added in a rapid, steady stream by syringe. The reaction temperature was maintained at 10° C. during the reaction time. The red color of the solution faded gradually in about 45 min and then 12 mL of H2O was added, and the mixture was stirred at RT overnight. The resulting solution was diluted with 36 mL of H2O and extracted with Et2O (2×30 mL). The combined organic layers were washed with 40 mL of 10% potassium carbonate solution and H2O, dried over MgSO4, filtered, and concentrated to give 14-2 as a clear oil (5.23 g, 80%). 1H NMR (500 MHz, CDCl3): δ 3.99 (s, 2H), 2.99 (m, 1H), 1.16 (d, 6H).

[0476] Step B

[0477] N-α-Cbz-N-β-Boc-(D)-diaminopropionic acid (14-3) (5.04 g, 14.91 mmol) was dissolved in 75 mL of DMF, and then K2CO3 (2.47 g, 17.89 mmol) and methyl iodide (4.64 mL, 74.55 mmol) were added. This mixture was stirred at RT overnight. The cloudy solution was diluted with EtOAc-H2O, and the aqueous layer was extracted with EtOAc. The combined organics were washed with brine, dried over MgSO4, filtered, and concentrated. Purification by chromatography on silica gel (30% EtOAc/hexane) gave 14-4 as a clear oil (5.21 g, 99%). LCMS (ESI): m/z 253 (M++1-Boc).

[0478] Step C

[0479] Compound 14-4 (5.20 g, 14.78 mmol) was dissolved in 74 mL of EtOH and 10% palladium on carbon (0.52 g) was added. The reaction mixture was stirred at RT under H2 overnight. The reaction mixture was then filtered through Celite using CH2Cl2 and concentrated to give 14-5 as a clear oil (2.96 g, 91%). LCMS (ESI): m/z 219 (M++1).

[0480] Step D

[0481] Compound 14-5 (1.50 g, 6.87 mmol) was dissolved in 34.4 mL of DMF, and then DIEA (1.20 mL, 6.87 mmol) and compound 14-2 (1.13 g, 6.87 mmol) were added. Then the mixture was stirred at RT overnight. The reaction mixture was diluted with EtOAc and washed with brine and H2O. The organic layer was dried over MgSO4, filtered, and concentrated. Purification by chromatography on silica gel (30% and 50% EtOAc/hexane) gave 14-6 as a yellow oil (0.995 g, 48%). LCMS (ESI): m/z 303 (M++1), 247 (M+−55).

[0482] Step E

[0483] Compound 14-6 (0.78 g, 2.59 mmol) was dissolved in 13.0 mL of 1:1 TFA-CH2Cl2 and stirred at RT for 30 min. Then the mixture was concentrated with CH2Cl2 two times. This TFA salt was dissolved in 32 mL of 1,2-dichloroethane, and sodium triacetoxyborohydride (0.77 g, 3.63 mmol) and acetic acid (0.15 mL, 2.59 mmol) were added. The reaction mixture was stirred at RT under N2 overnight. To this reaction mixture, 16 mL of saturated aqueous NaHCO3 solution was added, and the mixture was concentrated with toluene to give a yellow solid. The crude compound was dissolved in 13 mL of CH2Cl2 and then Boc-anhydride (1.24 g, 5.70 mmol) and TEA (1.08 mL, 7.77 mmol) were added. The resulting mixture was stirred at RT overnight, and then diluted with CH2Cl2. The organic phase was washed with 1N HCl and brine, dried over MgSO4, filtered and concentrated. Purification by chromatography on silica gel (10% EtOAc/hexane) gave 14-7 as a white solid (0.51 g, 50%). LCMS (ESI): m/z 187 (M++1-2Boc).

[0484] Step F

[0485] Compound 14-7 (0.51 g, 1.31 mmol) was dissolved in 4 mL of MeOH and 1N aqueous NaOH solution (2.62 mL, 2.62 mmol) was added. This mixture was stirred at RT overnight, and then concentrated. Water was then added and the pH was adjusted to about 2 using 1N HCl solution. The acidic solution was then extracted with EtOAc, and the combined organics were washed with brine, dried over MgSO4, filtered, and concentrated to give 14-8 as a white foamy-solid (0.40 g, 82%). LCMS (ESI): m/z 173 (M++1-2Boc).

[0486] Step G

[0487] N,N-Di-Boc-5-isopropyl-(R)-piperazine-2-carboxylic acid (14-8) (0.105 g, 0.281 mmol) was dissolved in 1.30 mL of methylene chloride, and then amine intermediate 14-9 (0.110 g, 0.255 mmol), DIEA (0.18 mL, 1.02 mmol), EDC (0.054 g, 0.255 mmol), and HOBt (0.038 g, 0.281 mmol) were added. The resulting mixture was stirred at room temperature overnight, and then diluted with 10 mL of CH2Cl2 and washed with 5 mL of 1N HCl solution, 5 mL of saturated NaHCO3 solution, 5 mL of H2O, and 5 mL of saturated NaCl solution. The organic phase was dried over MgSO4, filtered, and concentrated to give a yellow oil. The crude product was purified by column chromatography on silica gel (30:1 to 9:1 methylene chloride-acetone) to give 14-10 as a white solid (0.148 g, 74%).

[0488] LCMS (ESI): m/z 786 (M++1).

[0489] Step H

[0490] Compound 14-10 (0.146 g, 0.185 mmol) was dissolved in 0.46 mL of methylene chloride and 0.46 mL of trifluoroacetic acid. This solution was stirred at room temperature for 1 h, and then concentrated with four, 2-mL portions of CH2Cl2 to give 14-11 as a white foamy-solid (0.142 g, 94%). LCMS (ESI): m/z 586 (M++1). This mixture of two diastereoisomers was separated into each diastereoisomer (D1 and D2) by preparative HPLC.

[0491] The following 5-substituted-piperazine Examples shown in Table 10 with the indicated stereochemistry at the stereogenic cneter marked with an ** were prepared in a similar manner as Example 117, but using the appropriate N,N-di-Boc-5-substituted-(R)-piperazine-2-carboxylic acids in place of 14-8 which are prepared in a similar manner as 14-8 from the ketone precursors corresponding to 14-1.

TABLE 10
626
					R10,	Diaste-	LCMS
Ex.	R6	**	X	R3	R11	reomer	(ESI): m/z
118	F	(R)	627	628	H	D1 + D2	586 (M+ + 1)
119	F	(R)	629	630	H	D1	586 (M+ + 1)
120	F	(R)	631	632	H	D2	586 (M+ + 1)
121	Cl	(R)	633	634	H	D1 + D2	602 (M+ + 1)
122	Cl	(R)	635	636	H	D1	602 (M+ + 1)
123	Cl	(R)	637	638	H	D2	602 (M+ + 1)
124	F	(R)	639	640	Me	D1 + D2	662 (M+ + 1)
125	F	(R)	641	642	H	D1 + D2	614 (M+ + 1)
126	F	(R)	643	644	H	D1 + D2	559 (M+ + 1)
127	F	(R)	645	646	H	D1 + D2	575 (M+ + 1)
128	F	(R)	647	648	H		591 (M+ + 1)
129	Cl	(R)	649	Ph	H	D1 + D2	636 (M+ + 1)
130	F	(R)	650	Ph	H	D1 + D2	620 (M+ + 1)
131	F	(R)	651	Ph	H	D1 + D2	648 (M+ + 1)
132	F	(S)	652	653	H	D1 + D2	586 (M+ + 1)
133	F	(S)	654	655	H	D1	586 (M+ + 1)
134	F	(S)	656	657	H	D2	586 (M+ + 1)
135	F	(S)	658	659	H	D1 + D2	584 (M+ + 1)
136	F	(S)	660	661	H	D1	584 (M+ + 1)
137	F	(S)	662	663	H	D2	584 (M+ + 1)
138	F	(R)	664	665	H	D1 + D2	584 (M+ + 1)
139	F	(R)	666	667	H	D1	584 (M+ + 1)
140	F	(R)	668	669	H	D1 + D2	584 (M+ + 1)
141	F	(S)	670	671	H	D1 + D2	600 (M+ + 1)
142	F	(S)	672	673	H	D1	600 (M+ + 1)
143	F	(S)	674	675	H	D2	600 (M+ + 1)
144	F	(R)	676	677	H	D1 + D2	600 (M+ + 1)
145	F	(R)	678	679	H	D1	600 (M+ + 1)
146	F	(R)	680	681	H	D2	600 (M+ + 1)
147	F	(S)	682	683	H	D1 + D2	598 (M+ + 1)
148	F	(S)	684	685	H	D1	598 (M+ + 1)
149	F	(S)	686	687	H	D2	598 (M+ + 1)
150	F	(R)	688	689	H	D1 + D2	598 (M+ + 1)
151	F	(R)	690	691	H	D1	598 (M+ + 1)
152	F	(R)	692	693	H	D2	598 (M+ + 1)
153	Cl	(R)	694	695	H	D1 + D2	814 (M+ + 1)
154	Cl	(R)	696	697	H	D1	814 (M+ + 1)
155	Cl	(R)	698	699	H	D2	814 (M+ + 1)
156	Cl	(S)	700	Ph	H	D1 + D2	636 (M+ + 1)
157	F	(S)	701	Ph	H	D1 + D2	620 (M+ + 1)
158	F	(S)	702	Ph	H	D1 + D2	648 (M+ + 1)

[0492] 703

EXAMPLE 159

[0493] Intermediate 15-1 (0.0558 g, 0.068 mmol) was dissolved in 0.34 mL of 1:1 TFA-CH2Cl2 and stirred at RT for 1 h, and then the mixture was concentrated with CH2Cl2. The resulting TFA salt was dissolved in 0.34 mL of MeOH, and then sodium acetate (0.056 g, 0.68 mmol) and 37% aqueous formaldehyde solution (0.05 mL, 0.653 mmol) were added. After 20 min, sodium cyanoborohydride (1.0 M in THF, 0.44 mL, 0.44 mmol) was added. The reaction mixture was stirred at room temperature overnight and then concentrated. The crude mixture was dissolved in EtOAc and 1N NaOH, and the layers were separated. The organic phase was washed with 1N NaOH solution, H2O, and brine, dried over MgSO4, filtered, and concentrated. Purification by column chromatography (50-75% EtOAc/hexane, and then 1% TEA in EtOAc) provided two diastereoisomers as a white foamy solid [0.014 g (D1) and 0.009 g (D2)]. Each of these diastereoisomers [0.012 g, 0.019 mmol (D1); and 0.007 g, 0.011 mmol (D2)] was separately dissolved in CH2Cl2, and 1.0 M HCl solution in Et2O (0.28 mL, 0.28 mmol; and 0.03 mL, 0.03 mmol, respectively) was added. The precipitates were filtered under N2 and dried under vacuum to give white solids [0.012 g (D1) and 0.0072 g (D2)]. LCMS (ESI): m/z 664 (M++1).

[0494] The following Nα-methyl-Nβ-methyl-5-phenyl-piperazine Examples shown in Table 11 with the indicated stereochemistry at the stereogenic cneter marked with an ** were prepared in a similar manner as Example 159, but using the appropriate N,N-di-Boc-5-phenyl-(D)-piperazine-2-carboxamide intermediate in place of 15-1.

TABLE 11
704
			Diastereo-		LCMS
Ex.	R6	**	mer	X	(ESI): m/z
160	Cl	(S)	D1	705	664 (M+ + 1)
161	Cl	(S)	D2	706	664 (M+ + 1)
162	F	(S)	D1	707	648 (M+ + 1)
163	F	(S)	D2	708	648 (M+ + 1)
164	F	(S)	D1	709	676 (M+ + 1)
165	Cl	(R)	D1	710	664 (M+ + 1)
166	Cl	(R)	D2	711	664 (M+ + 1)
167	F	(R)	D1	712	648 (M+ + 1)
168	F	(R)	D2	713	648 (M+ + 1)
169	F	(R)	D1	714	676 (M+ + 1)

[0495] 715

EXAMPLE 170

[0496] Step A

[0497] 2-Chloro-4-fluorobenzyl bromide (16-2) (1.0 g, 4.47 mmol), N-(diphenylmethylene)glycine ethyl ester (1-1) (1.067 g, 3.99 mmol) and benzyl triethylammonium chloride (0.922 g, 4.05 mmol) were dissolved in CH2Cl2, and 10% aqueous NaOH solution was added. The resulting two-phase mixture was stirred at RT overnight. The organic layer was separated and concentrated. The residue was dissolved in ether, washed with 1120, dried over MgSO4, filtered, and concentrated to give 16-3 as a clear oil (1.445 g, 88%).

[0498] LCMS (ESI): m/z 410 (M++1).

[0499] Step B

[0500] Compound 16-3 was dissolved in THF and 5% aqueous HCl solution was added. The mixture was stirred at RT for 1 h. Saturated NaHCO3 solution was then added slowly and the mixture was extracted with EtOAc. The combined organics were washed with 1120 and brine, dried over MgSO4, filtered, and concentrated. Purification by chromatography on silica gel (3-30% EtOAc/hexane) gave 16-4 as a clear oil (0.626 g, 75%). LCMS (ESI): m/z 246 (M++1).

[0501] Step C

[0502] Compound 16-4 (0.6238 g, 2.54 mmol) was dissolved in 12.7 mL of CH2Cl2, and then Boc-anhydride (0.61 g, 2.79 mmol) and TEA (0.53 mL, 3.81 mmol) were added. The resulting mixture was stirred at RT overnight and then diluted with CH2Cl2. The organic phase was washed with 1N HCl and brine, dried over MgSO4, filtered and concentrated to give 16-5 as an oil (0.88 g).

[0503] LCMS (ESI): m/z 246 (M++1-Boc).

[0504] Step D

[0505] Compound 16-5 (0.88 g, 2.54 mmol) was dissolved in 8 mL of MeOH and 1N aqueous NaOH solution (5.08 mL, 5.08 mmol) was added. The resulting mixture was stirred at RT overnight and then concentrated. Water was added and the pH was adjusted to about 2 using 1N HCl solution. The acidic solution was extracted with EtOAc, and the combined organic layers were washed with brine, dried over MgSO4, filtered, and concentrated to give intermediate 16-6 as a white solid (0.65 g, 81%).

[0506] LCMS (ESI): m/z 218 (M++1-Boc).

[0507] Step E

[0508] The following Examples shown in Table 12 were prepared following the procedures shown in Scheme 1 and detailed in Example 1 but using the appropriately substituted Boc-phenylalanine intermediate as prepared in Steps A-D above or commercially available Boc-phenylalanines and the appropriately protected piperazine-2-carboxylic acid intermediate.

TABLE 12
716
					LCMS
Ex.	R6	*	R3	R4b	(ESI): m/z
171	3,4-difluoro-	(R)	H	H	562 (M+ + 1)
172	3,4-difluoro-	(R)	H	Me	576 (M+ + 1)
173	3,4-difluoro-	(R)	i-Pr(D1 +D2)	H	604 (M+ + 1)
174	3,4-difluoro-	(R)	i-Pr(D1)	H	604 (M+ + 1)
175	3,4-difluoro-	(R)	i-Pr(D2)	H	604 (M+ + 1)
176	3,5-difluoro-	(R)	H	Me	576 (M+ + 1)
177	3,5-difluoro-	(R)	i-Pr	H	604 (M+ + 1)
178	2,4-difluoro	(RS)	H	H	562 (M+ + 1)
179	2,4-difluoro	(RS)	H	Me	576 (M+ + 1)
180	2-chloro-4- fluoro	(RS)	H	H	578 (M+ + 1)
181	2-chloro-4- fluoro	(RS)	H	Me	592 (M+ + 1)

[0509] 717

EXAMPLE 182

[0510] Step A

[0511] Intermediate 17-1 (0.28 g, 0.926 mmol) was dissolved in 4.6 mL of DMF, and then DIEA (0.16 mL, 0.926 mmol) and methyl iodide (0.12 mL, 1.852 mmol) were added. The resulting mixture was stirred at RT overnight. The reaction mixture was diluted with EtOAc and washed with brine and H2O, and the organic layer was dried over MgSO4, filtered, and concentrated to give a red oil which was treated with TFA in methylene chloride to remove the Boc-protecting group. After the usual work-up, the crude compound was dissolved in 18 mL of 1,2-dichloroethane, and then sodium triacetoxyborohydride (0.275 g, 1.30 mmol) and acetic acid (0.053 mL, 0.926 mmol) were added. The reaction mixture was stirred at RT under N2 overnight. The reaction mixture was then diluted with 9 mL of saturated aqueous NaHCO3 solution and then concentrated with toluene to give a yellow solid. The crude compound was dissolved in 4.6 mL of CH2Cl2, and then Boc-anhydride (0.22 g, 1.02 mmol) and TEA (0.19 mL, 1.39 mmol) were added. The reaction mixture was stirred at room temperature overnight and then diluted with CH2Cl2. The solution was washed with H2O and brine, dried over MgSO4, filtered and concentrated. Purification by chromatography on silica gel (10% EtOAc/hexane) gave 17-2 as a white solid (0.02 g, 7%). LCMS (ESI): m/z 301 (M++1), 245 (M+−55).

[0512] Step B

[0513] Compound 17-2 (0.020 g, 0.067 mmol) was dissolved in 0.2 mL of MeOH and 1N aqueous NaOH solution (0.14 mL, 0.14 mmol) was added. The mixture was stirred at RT overnight and then concentrated. Water was added and the pH was adjusted to about 7 using 1N HCl solution. The aqueous solution was then extracted with EtOAc, and the combined organic layers were washed with brine, dried over MgSO4, filtered, and concentrated to give a white foamy solid. This acid was dissolved in 0.5 mL of methylene chloride, and then the amine intermediate 14-9 (0.027 g, 0.061 mmol), DIEA (0.04 mL, 0.244 mmol), EDC (0.013 g, 0.067 mmol), and HOBt (0.009 g, 0.067 mmol) were added. The resulting mixture was stirred at room temperature overnight. The resulting mixture was diluted with CH2Cl2 and washed with H2O, and brine, and the organic phase was dried over MgSO4, filtered, and concentrated. The crude product was purified by column chromatography on silica gel (9:1 methylene chloride-acetone, and then 3-10% MeOH in CH2Cl2) to give 17-3 as a white solid (0.006 g, 14%).

[0514] LCMS (ESI): m/z 700 (M++1).

[0515] Step C

[0516] Compound 17-3 (0.0058 g, 0.0083 mmol) was dissolved in 0.1 mL of methylene chloride and 0.1 mL of trifluoroacetic acid. This solution was stirred at room temperature for 30 min, and then concentrated with four, 2-mL portions of CH2Cl2 to give 17-4 as a white, foamy-solid (0.006 g, 92%). LCMS (ESI): m/z 600 (M++1). 718

EXAMPLE 183

[0517] Step A

[0518] N-α-Cbz-(L)-Diaminopropionic acid (18-1) (4.0 g, 16.79 mmol) was suspended in 32 mL of MeOH and the flask was cooled at 0° C. in an ice-water bath. Chlorotrimethylsilane (4.7 mL, 36.94 mmol) was then added dropwise and the reaction mixture was stirred at 0° C. for 1 h, and then warmed to RT and stirred at RT overnight. This mixture was concentrated and dried under vacuum to give 18-2 as a white solid (4.77 g, 98%). LCMS (ESI): m/z 253 (M++1).

[0519] Step B

[0520] Compound 18-2 (2.50 g, 8.66 mmol) was dissolved in 34.4 mL of DMF, and then DIEA (3.02 mL, 17.32 mmol) and compound 14-2 (1.43 g, 8.66 mmol) were added. The resulting mixture was then stirred at RT overnight. The reaction mixture was diluted with EtOAc and washed with water and brine, and the organic layer was dried over MgSO4, filtered, and concentrated . Purification by chromatography (30-40% EtOAc/hexane) gave 18-3 as a yellow oil (1.41 g, 49%).

[0521] LCMS (ESI): m/z 337 (M++1).

[0522] Step C

[0523] Compound 18-3 (1.26 g, 3.73 mmol) was dissolved in 70 mL of EtOH and 10% palladium on carbon (0.40 g) was then added. The resulting mixture was stirred at RT under H2 overnight. This mixture was then filtered through Celite using CH2Cl2, and the filtrate was concentrated to give 18-4 as an orange oil (0.65 g, 94%).

[0524] LCMS (ESI): m/z 187 (M++1).

[0525] Step D

[0526] Compound 18-4 (0.643 g, 3.45 mmol) was dissolved in 17.3 mL of CH2Cl2, and then Boc-anhydride (1.66 g, 7.59 mmol) and TEA (1.44 mL, 10.35 mmol) were added. The resulting mixture was stirred at RT overnight. The reaction mixture was diluted with CH2Cl2 and washed with 1N HCl and brine, dried over MgSO4, filtered, and concentrated. Purification by chromatography (10% EtOAc/hexane) gave 18-5 as a white solid (0.82 g, 62%). LCMS (ESI): m/z 231 (M+-Boc-55).

[0527] Step E

[0528] Compound 18-5 (0.796 g, 0.16 mmol) was dissolved in 6.0 mL of MeOH and 1N aqueous NaOH solution (4.12 mL, 4.12 mmol) was added. This mixture was stirred at RT overnight. The solution was concentrated and resulting residue was diluted with water and the pH was adjusted to about 2 using 1N HCl solution. The aqueous solution was extracted with EtOAc, and the combined organic layers were washed with brine, dried over MgSO4, filtered, and concentrated to give 18-6 as a white, foamy-solid (0.412 g, 54%). LCMS (ESI): m/z 217 (M+-Boc-55).

[0529] Step F

[0530] N,N-Di-Boc-6-isopropyl-(S)-piperazine-2-carboxylic acid (18-6) (0.08 g, 0.215 mmol) was dissolved in 1.0 mL of methylene chloride, and then amine intermediate 14-9 (0.0843 g, 0.195 mmol), DIEA (0.14 mL, 0.78 mmol), EDC (0.041 g, 0.215 mmol), and HOBt (0.029 g, 0.215 mmol) were added. The resulting mixture was stirred at room temperature overnight, and then diluted with 10 mL of CH2Cl2 and washed with 5 mL of 1N HCl solution, 5 mL of saturated NaHCO3 solution, 5 mL of H2O, and 5 mL of saturated NaCl solution. The organic phase was dried over MgSO4, filtered, and concentrated to give a yellow oil. The crude product was purified by column chromatography (30:1 to 3:1 methylene chloride-acetone) to give 18-7 as a white solid (0.0743 g, 49%). LCMS (ESI): m/z 786 (M++1).

[0531] Step G

[0532] Compound 18-7 (0.0729 g, 0.093 mmol) was dissolved in 0.23 mL of methylene chloride and 0.23 mL of trifluoroacetic acid. This solution was stirred at room temperature for 1 h, and then concentrated with four, 2-mL portions of CH2Cl2 to give 18-8 as a white foamy-solid (0.0732 g, 97%). LCMS (ESI): m/z 586 (M++1). This mixture of two diastereoisomers was separated into each diastereoisomer (D1 and D2) by preparative HPLC.

EXAMPLE 184

[0533] 719

[0534] This example was prepared in the same manner as Example 183, but using N,N-di-Boc-6-isopropyl-(R)-piperazine-2-carboxylic acid in place of the corresponding (S)-isomer for the coupling reaction in Step F of Example 183. LCMS

[0535] (ESI) m/z 586 (M++1). 720

EXAMPLE 185

[0536] Step A

[0537] 1,2-Diamino-2-methylpropane (19-1) (0.50 mL, 4.77 mmol) was dissolved in 20 ml of DMF, and then ethyl 2,3-dibromopropionate (0.70 mL, 4.77 mmol) and TEA (1.33 mL, 9.54 mmol) were added. This mixture was stirred at RT overnight, and then concentrated to give an oil. Purification by chromatography on silica gel (9:1 CH2Cl2/acetone, and then 3-10% MeOH in CH2Cl2) gave 19-2 as a yellow foamy-solid (0.76 g). LCMS (ESI): m/z 187 (M++1).

[0538] Step B

[0539] Compound 19-2 (0.76 g, 4.06 mmol) was dissolved in 15.0 mL of CH2Cl2, and then Boc-anhydride (1.77 g, 8.12 mmol) and TEA (1.70 mL, 12.18 mmol) were added. The resulting mixture was stirred at RT overnight. The reaction mixture was then diluted with CH2Cl2 and washed with 1N HCl and brine, dried over MgSO4, filtered, and concentrated. Purification by chromatography on silica gel (10% EtOAc/hexane) gave 19-3 as a white solid (0.065 g, 4%).

[0540] LCMS (ESI): m/z 187 (M+-2Boc).

[0541] Step C

[0542] Compound 19-3 (0.062 g, 0.16 mmol) was dissolved in 0.5 mL of MeOH and 1N aqueous NaOH solution (0.32 mL, 0.32 mmol) was added. This mixture was stirred at RT overnight. The solution was concentrated and resulting residue was diluted with water and the pH was adjusted to about 2 using 1N HCl solution. The aqueous solution was extracted with EtOAc and the combined organic layers were washed with brine, dried over MgSO4, filtered, and concentrated to give acid 19-4 as a white, foamy-solid (0.053 g, 93%). LCMS (ESI): m/z 159 (M++1-2Boc), 203 (M+-Boc-55), 381 (M++Na).

[0543] Step D

[0544] Acid 19-4 (0.51 g, 0.142 mmol) was dissolved in 0.65 mL of methylene chloride, and then amine intermediate 14-9 (0.058 g, 0.129 mmol), DIEA (0.09 mL, 0.516 mmol), EDC (0.027 g, 0.142 mmol), and HOBt (0.027 g, 0.142 mmol) were added. The resulting mixture was stirred at room temperature overnight, and then diluted with 10 mL of CH2Cl2 and washed with 5 mL of 1N HCl solution, 5 mL of saturated NaHCO3 solution, 5 mL of H2O, and 5 mL of saturated NaCl solution. The organic phase was dried over MgSO4, filtered, and concentrated to give a yellow oil. The crude product was purified by column chromatography on silica gel (30:1 to 9:1 methylene chloride-acetone) to give 19-5 as a white solid (0.0724 g, 73%). LCMS (ESI): m/z 772 (M++1), 672 (M+-Boc).

[0545] Step E

[0546] Compound 19-5 (0.0692 g, 0.090 mmol) was dissolved in 0.22 mL of methylene chloride and 0.22 mL of trifluoroacetic acid. This solution was stirred at room temperature for 1 h, and then concentrated with four, 2-mL portions of CH2Cl2 to give 19-6 as a white foamy-solid (0.069 g, 96%). LCMS (ESI): m/z 572 (M++1). 721

EXAMPLE 186

[0547] Intermediate 20-1 (0.14 g, 0.204 mmol) was dissolved in 0.51 mL of methylene chloride and 0.51 mL of trifluoroacetic acid. This solution was stirred for 30 min at room temperature. The mixture was then concentrated with methylene chloride (3mL×2) to give a white solid. The solid was dissolved in 1.0 mL of methanol, and then sodium acetate (0.084 g, 1.02 mmol) and acetone (0.072 mL, 0.98 mmol) were added. The reaction mixture was stirred at room temperature for 30 min, and then sodium cyanoborohydride (1.0 M in THF, 0.65 mL, 0.65 mmol) was added. The mixture was stirred at room temperature overnight. The solution was concentrated, and the residue was taken up in EtOAc (10 mL) and 1N NaOH (5 mL) and the layers were separated. The organic phase was washed with 1N NaOH (5 mL), H2O (5 mL), and brine (5 mL), dried over MgSO4, filtered, and concentrated. Purification by column chromatography on silica gel (3% to 10% methanol in methylene chloride) gave 20-2 as a white foamy solid (0.088 g, 69%); mass spectrum: 628 (M+1). 722

EXAMPLE 187

[0548] Step A

[0549] Intermediate 14-4 (2.19 g, 6.22 mmol) was dissolved in 16 mL of methylene chloride and 16 mL of trifluoroacetic acid. This solution was stirred at room temperature for 45 min, and then concentrated with two, 6-mL portions of CH2Cl2 to give an oil. The acid was then dissolved in ethyl acetate and washed twice with aqueous 1 N NaOH solution, dried over K2CO3, filtered and concentrated to give 21-1 as a clear oil (0.999 g, 64%). LCMS (ESI): m/z 253 (M++1).

[0550] Step B

[0551] Compound 21-1 (0.999 g, 3.96 mmol) was dissolved in 20 mL of DMF, and then DIEA (0.92 mL, 5.28 mmol) and compound 14-2 (0.436 g, 2.64 mmol) were added. The resulting mixture was then stirred at room temperature for 4.5 days. The reaction mixture was diluted with EtOAc and water, and the layers were separated. The aqueous phase was extracted with ethyl acetate, and the combined organic layers were washed with water and brine, dried over Na2SO4, filtered, and concentrated to give 21-2 as a golden yellow oil. Purification by chromatography on silica gel (20-50% EtOAc-hexane) gave a yellow oil (0.665 g, 75%). LCMS (ESI): m/z 337 (M++1)

[0552] Step C

[0553] Compound 21-2 (0.250 g, 0.743 mmol) was dissolved in 7 mL of 1,2-dichloroethane, and then 37% aqueous formaldehyde (0.36 mL, 4.46 mmol) and sodium triacetoxyborohydride (0.636 g, 2.97 mmol) were added. The reaction mixture was stirred at RT under N2 overnight. The reaction mixture was then diluted with 6 mL of saturated aqueous NaHCO3 solution and stirred at room temperature for 15 minutes. The layers were then separated and the aqueous phase was extracted with methylene chloride. The combined organic layers were washed with brine, dried over Na2SO4, filtered and concentrated to give a light yellow oil. Purification by chromatography on silica gel (20-50% EtOAc/hexane) gave 21-3 as a clear oil (0.144 g, 55%). LCMS (ESI): m/z 351 (M++1).

[0554] Step D

[0555] Compound 21-3 (0.144 g, 0.411 mmol) was dissolved in 8.5 mL of EtOH, and 10% palladium on carbon (0.044 g) was added. The resulting mixture was stirred at RT under H2 overnight. This mixture was then diluted with MeOH and filtered through Celite using MeOH to wash the filter. The filtrate and washings were concentrated to give 21-4 as a yellow oil (0.077 g, 94%). LCMS (ESI): m/z 201 (M++1).

[0556] Step E

[0557] Compound 21-4 (0.077 g, 0.385 mmol) was dissolved in 2 mL of CH2Cl2, and then Boc-anhydride (0.092 g, 0.423 mmol) and TEA (0.08 mL, 0.578 mmol) were added. The resulting mixture was stirred at RT for 3.5 days. The reaction mixture was concentrated to give a yellow-brown oil. Purification by silica gel chromatography (20-50% EtOAc/hexane) gave 21-5 as a yellow oil (0.048 g, 41%). LCMS (ESI): m/z 301 (M++1).

[0558] Step F

[0559] Compound 21-5 (0.048 g, 0.160 mmol) was dissolved in 0.64 mL of THF, and then 0.16 mL of water and LiOH-H2O (0.020 g, 0.479 mmol) were added. This mixture was stirred at RT overnight and then heated at 45° C. for 4 h. The solution was then diluted with 0.5 mL of 1N HCl solution (pH=4-5) and concentrated to give 21-6 as an oily solid. LCMS (ESI): ml/z 287 (M++1).

[0560] Step G

[0561] Acid 21-6 (0.160 mmol) was dissolved in 1.0 mL of methylene chloride, and then amine intermediate 14-9 (0.065 g, 0.145 mmol), DIEA (0.10 mL, 0.580 mmol), EDC (0.031 g, 0.160 mmol), and HOBt (0.022 g, 0.160 mmol) were added. The resulting mixture was stirred at room temperature overnight, and then diluted with 10 mL of CH2Cl2 and washed with 5 mL of saturated NaHCO3 solution, 5 mL of H2O, and 5 mL of saturated NaCl solution. The organic phase was dried over Na2SO4, filtered, and concentrated to give a yellow oil. The crude product was purified by column chromatography on silica gel (3:1 methylene chloride-acetone) to give 21-7 as a white solid (0.017 g, 16%). LCMS (ESI): m/z 716 (M++1).

[0562] Step H

[0563] Compound 21-7 (0.017 g, 0.024 mmol) was dissolved in 0.10 mL of methylene chloride and 0.10 mL of trifluoroacetic acid. This solution was stirred at room temperature for 1 h, and then concentrated with four, 2-mL portions of CH2Cl2 to give 21-8 as an off-white solid (0.020 g). LCMS (ESI): m/z 616 (M++1).

EXAMPLE 188

[0564] 723

[0565] This Example was prepared in a similar fashion as Example 187, but using the protected D-piperazine-2-carboxylic acid for coupling with 14-9, in place of the L-piperazine 21-6. 724

EXAMPLE 188

[0566] Step A

[0567] To a mixture of 2-isopropylpyrazine (22-1) (163.7 mmol, 20 mg) and DMF (188 mmol, 14.58 mL) was added sulfuryl chloride (163.7 mmol, 13.15 mL) slowly over 2 hr (syringe pump) maintaining the temperature below 40° C. Reaction was stirred at room temperature overnight. The mixture was cooled to 0° C. and H2O (40 mL) was added cautiously followed by 5N NaOH (ca. 60 mL) to neutralize the solution. Water (600 mL) was added and the emulsion was distilled until no further oil condensed (about half the original volume). The emulsion was extracted with CH2Cl2 (4×200 mL). The combined organic layers were dried over Na2SO4 and concentrated to afford a clear colourless oil containing by NMR analysis a 1:1 mixture of starting material and desired product 22-2. This product was used without further purification in Step B.

[0568] Step B

[0569] A mixture of a 1:1 mix of isopropylpyrazine and 2-chloro-3-isopropylpyrazine from Step A (36 mmol), DPPE (0.9 mmol, 371 mg), Pd(OAc)2 (0.9 mmol, 202 mg) and triethylamine (45 mmol, 6.3 mL) in DMF/MeOH (1:2) (18 mL) was stirred at 40 psi CO(g) at 60° C. overnight. The mixture was filtered through a short pad of celite, concentrated and partitioned between EtOAc/H2O. Organic phase was washed with brine, dried over Na2SO4 and concentrated. Chromatography over silica gel eluting with 20% EtOAc/hexane afforded 22-3 as a clear colorless oil (1.38 g).

[0570] Step C

[0571] A suspension of PtO2 (20 mol %, 45 mg) in a solution of 22-3 (1 mmol, 180 mg) in AcOH (20 mL) was shaken under 45 psi of hydrogen gas for 16 hr. Reaction mixture was filtered through a short pad of celite and concentrated to afford 22-4 as a white foam.

[0572] Step D

[0573] To a solution of 22-4 (1 mmol, 186 mg) in 10% Et3N/MeOH was added Boc2O (2.4 mmol, 765 mg) and the resultant solution was stirred at room temperature overnight to give a mixture of 2 products. Additional Boc2O was added and the reaction heated to 50° C. for 5 hr. The volatiles were removed and the residue was partitioned between 0.5M HCl and EtOAc. Organic phase was washed with saturated NaHCO3 and brine, dried over Na2SO4 and concentrated. Chromatography over silica gel eluting with 50 mL of 0, 2.5, 5, 10, 20, and 30% Me2CO/CH2Cl2 afforded 22-5 which was hydrolyzed under basic conditions to give acid 22-6.

[0574] Step E

[0575] To a solution of 14-9 (0.1 mmol, 47 mg) in CH2Cl2 at room temperature was added the acid 22-6 (0.1 mmol, 46 mg) followed by HOBt (0.12 mmol, 16 mg), EDC (0.12 mmol, 23 mg) and NMM (0.45 mmol, 0.05 mL). Resultant solution was stirred at room temperature overnight. Reaction mixture was poured into EtOAc (20 mL) and washed successively with 0.5M HCl, saturated NaHCO3, water and brine, dried over Na2SO4 and concentrated. Chromatography over silica gel eluting with 100 mL of 5 and 10%, then 50 mL of 20 and 30% Me2CO/CH2Cl2 afforded two diastereoisomeric products D1 and D2 as white solids. Each diastereoisomer was deprotected with TFA in methylene chloride. Yield D1: 20 mg; D2: 16 mg. 725

EXAMPLE 189

[0576] Step A

[0577] A suspension of PtO2 (40 mol %, 330 mg) in a solution of 23-1 (3.62 mmol, 500 mg) in AcOH (36 mL) was shaken under 45 psi of hydrogen gas for 72 hours. Reaction mixture was filtered through a short pad of celite and concentrated to afford a white foam. To a solution of the resulting amino acid in 10% Et3N/MeOH (15 mL) was added Boc2O (8.69 mmol, 2.77 g), and the resultant solution was stirred at room temperature overnight. Volatiles were removed and the residue was partitioned between 0.5M HCl and EtOAc. Organic phase was washed with saturated NaHCO3 and brine, dried over Na2SO4 and concentrated. Chromatography over silica gel eluting with 500 mL of 20-30% Me2CO/CH2Cl2 afforded 23-2 (191 mg).

[0578] Step B

[0579] To a solution of amine 14-9 (0.11 mmol, 50 mg) in CH2Cl2 at room temperature was added the acid 23-2 (0.11 mmol, 37 mg) followed by HOBt (0.13 mmol, 17 mg), EDC (0.13 mmol, 25 mg) and NMM (4.5 mmol, 49 mg). Resultant solution was stirred at room temperature overnight. Reaction mixture was poured into EtOAc (20 mL) and washed successively with 0.5M HCl, saturated NaHCO3, H2O and brine, dried over Na2SO4 and concentrated. Chromatography over silica gel eluting with 50 mL of 0, 2.5, 5, 10, and 20% Me2CO/CH2Cl2 afforded 23-3 as a white solid. A solution of 23-3 in CH2Cl2 and TFA was stirred at room temp for 1 hr. Volatiles were removed and the residue precipitated from a CH2Cl2 solution with Et2O/hexane to give the desired compound 23-4. 726

EXAMPLE 190

[0580] Step A

[0581] A mixture of 3-chloro-2,5-dimethylpyrazine (24-1) (100 mmol, 14.26 g), DPPE (5 mmol, 2.06 g), Pd(OAc)2 (5 mmol, 1.22 g) and sodium acetate (100 mmol, 8.2 g) in DMF/H2O (3:1) (80 mL) was stirred at 40 psi CO at 60° C. overnight. Mixture was filtered through a short pad of celite, concentrated and partitioned between CH2Cl2/1N NaOH. Aqueous phase was washed with CH2Cl2, acidified with 2N HCl and extracted wih EtOAc. Organic extracts were washed with brine, dried over Na2SO4 and evaporated to give 24-2 (1.6 g).

[0582] Step B

[0583] To a solution of amine 14-9 (0.1 mmol, 47 mg) in CH2Cl2 at room temp was added 24-2 (0.1 mmol, 15 mg) followed by HOBt (0.12 mmol, 16 mg), EDC (0.12 mmol, 23 mg) and NMM (0.45 mmol, 0.05 mL). Resultant solution was stirred at room temperature overnight. Reaction mixture was poured into EtOAc (10 mL) and washed successively with 0.5M HCl, saturated NaHCO3, H2O and brine, dried over Na2SO4 and concentrated. Chromatography over silica eluting with 50 mL of 0, 2.5, 5, 10, and 20% of Me2CO/CH2Cl2 afforded 24-3 as a white solid (50 mg).

[0584] Step C

[0585] A suspension of PtO2 (40 mol %, 73 mg) in a solution of 24-3 (0.81 mmol, 456 mg) in AcOH (25 mL) was stirred for 1 hr under a balloon atmosphere of hydrogen gas. Reaction mixture was filtered through a short pad of celite and concentrated to afford the desired product 24-4 as a white foam. 727

EXAMPLE 191

[0586] Step A

[0587] To a solution of amine 14-9 (0.1 mmol, 47 mg) in CH2Cl2 at room temperature was added 25-1 (0.11 mmol, 14 mg) followed by HOBt (0.12 mmol, 16 mg), EDC (0.12 mmol, 23 mg) and NMM (0.45 mmol, 0.05 mL). Resultant solution was stirred at room temperature overnight. Reaction mixture was poured into EtOAc (10 mL) and washed successively with 0.5M HCl, saturated NaHCO3, H2O and brine, dried over Na2SO4 and concentrated. Chromatography over silica gel eluting with 50 mL of 0, 2.5, 5, 10, and 20% Me2CO/CH2Cl2 afforded 25-2 as a white solid (43 mg).

[0588] Step B

[0589] A suspension of PtO2 (40 mol %, 12 mg) in a solution of 25-2 (0.14 mmol, 75 mg) in AcOH (5 mL) was stirred for 1 hr under a balloon atmosphere of hydrogen gas. Reaction mixture was filtered through a short pad of celite and concentrated. The crude product was purified by prep HPLC to afford the title compound 25-3 (5 mg). 728

EXAMPLE 192

[0590] Step A

[0591] A solution of 26-1 (216 mg, 0.39 mmol) in H2SO4 (3 mL) was stirred at rt for 10 min. The resultant solution was placed in an ice-bath and fuming HNO3 (0.03 mL) was added dropwise. After the addition was complete, the reaction was allowed to stir for 15 min at 0° C. The mixture was then poured into stirring ice-water (30 mL) and then allowed to warm to rt. Concentrated ammonia was added until pH 9 was obtained, and the resultant emulsion was extracted with EtOAc (3×100 mL). The organics were washed successively with 1N ammonia, water, and brine and dried over Na2SO4 and concentrated to give 26-2 (230 mg).

[0592] Step B

[0593] To an ice-cold solution of 26-2 (171 mg, 0.249 mmol) in water (2 mL) was added a solution of NaNO2 (20 mg, 0.28 mmol) until the resultant solution tested positive for excess nitrous acid with KI-starch paper. The solution was neutralized with Na2CO3 and was added to a suspension of KCN and CuCN in water (4 mL) at rt. The mixture was gradually heated to 50° C. After being heated for 2 h, the mixture was extracted with CHCl3 (4×25 mL). Organic phase was dried over Na2SO4 and concentrated. Chromatography over silica gel eluting with 5% MeOH/CH2Cl2 afforded the desired nitrile 26-3 (120 mg).

BIOLOGICAL ASSAYS

[0594] A. Binding Assay. The membrane binding assay was used to identify competitive inhibitors of 25I-NDP-alpha-MSH binding to cloned human MCRs expressed in L- or CHO- cells.

[0595] Cell lines expressing melanocortin receptors were grown in T-180 flasks containing selective medium of the composition: 1 L Dulbecco's modified Eagles Medium (DMEM) with 4.5 g L-glucose, 25 mM Hepes, without sodium pyruvate, (Gibco/BRl); 100 ml 10% heat-inactivated fetal bovine serum (Sigma); 10 ml 10,000 unit/ml penicillin & 10,000 ug/ml streptomycin (Gibco/BRl); 10 ml 200 mM L-glutamine (Gibco/BRl); 1 mg/ml Geneticin (G418) (Gibco/BRl). The cells were grown at 37° C. with CO2 and humidity control until the desired cell density and cell number was obtained.

[0596] The medium was poured off and 10 mls/monolayer of enzyme-free dissociation media (Specialty Media Inc.) was added. The cells were incubated at 37° C. for 10 minutes or until cells sloughed off when flask was banged against hand.

[0597] The cells were harvested into 200 ml centrifuge tubes and spun at 1000 rpm, 4° C, for 10 min. The supernatant was discarded and the cells were resuspended in 5 mls/monolayer membrane preparation buffer having the composition: 10 mM Tris pH 7.2-7.4; 4 ug/ml Leupeptin (Sigma); 10 uM Phosphoramidon (Boehringer Mannheim); 40 ug/ml Bacitracin (Sigma); 5 ug/ml Aprotinin (Sigma); 10 mM Pefabloc (Boehringer Mannheim). The cells were homogenized with motor-driven dounce (Talboy setting 40), using 10 strokes and the homogenate centrifuged at 6,000 rpm, 4° C., for 15 minutes.

[0598] The pellets were resuspended in 0.2 mls/monolayer membrane prep buffer and aliquots were placed in tubes (500-1000 ul/tube) and quick frozen in liquid nitrogen and then stored at −80° C.

[0599] Test compounds or unlabelled NDP-α-MSH was added to 100 μL of membrane binding buffer to a final concentration of 1 μM. The membrane binding buffer had the composition: 50 mM Tris pH 7.2; 2 mM CaCl2; 1 mM MgCl2; 5 mM KCl; 0.2% BSA; 4 ug/ml Leupeptin (SIGMA); 10 uM Phosphoramidon (Boehringer Mannheim); 40 ug/ml Bacitracin (SIGMA); 5 ug/ml Aprotinin (SIGMA); and 10 mM Pefabloc (Boehringer Mannheim). One hundred μl of membrane binding buffer containing 10-40 ug membrane protein was added, followed by 100 μM 125I-NDP-α-MSH to final concentration of 100 pM. The resulting mixture was vortexed briefly and incubated for 90-120 min at room temp while shaking.

[0600] The mixture was filtered with Packard Microplate 196 filter apparatus using Packard Unifilter 96-well GF/C filter with 0.1% polyethyleneimine (Sigma). The filter was washed (5 times with a total of 10 ml per well) with room temperature of filter wash having the composition: 50 mM Tris-HCl pH 7.2 and 20 mM NaCl. The filter was dried, and the bottom sealed and 50 ul of Packard Microscint-20 was added to each well. The top was sealed and the radioactivity quantitated in a Packard Topcount Microplate Scintillation counter.

[0601] B. Functional assay. Functional cell based assays were developed to discriminate melanocortin receptor agonists from antagonists.

[0602] Cells (for example, CHO- or L-cells or other eukaryotic cells) expressing a human melanocortin receptor (see e.g. Yang-YK; Ollmann-MM; Wilson-BD; Dickinson-C; Yamada-T; Barsh-GS; Gantz-I; Mol-Endocrinol. 1997 Mar; 11(3): 274-80) were dissociated from tissue culture flasks by rinsing with Ca and Mg free phosphate buffered saline (14190-136, Life Technologies, Gaithersburg, Md.) and detached following 5 minutes incubation at 37° C. with enzyme free dissociation buffer (S-014-B, Specialty Media, Lavellette, N.J.). Cells were collected by centrifugation and resuspended in Earles Balanced Salt Solution (14015-069, Life Technologies, Gaithersburg, Md.) with additions of 10 mM HEPES pH 7.5, 5 mM MgCl2, 1 mM glutamine and 1 mg/ml bovine serum albumin. Cells were counted and diluted to 1 to 5×106/ml. The phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine was added to cells to 0.6 mM.

[0603] Test compounds were diluted in dimethylsulfoxide (DMSO) (10−5 to 10−10 M) and 0.1 volume of compound solution was added to 0.9 volumes of cell suspension; the final DMSO concentration was 1%. After room temperature incubation for 45 min., cells were lysed by incubation at 100° C. for 5 min. to release accumulated cAMP.

[0604] cAMP was measured in an aliquot of the cell lysate with the Amersham (Arlington Heights, Ill.) cAMP detection assay (RPA556). The amount of cAMP production which resulted from an unknown compound was compared to that amount of cAMP produced in response to alpha-MSH which was defined as a 100% agonist. The EC50 is defined as the compound concentration which results in half maximal stimulation, when compared to its own maximal level of stimulation.

[0605] Antagonist assay: Antagonist activity was defined as the ability of a compound to block cAMP production in response to alpha-MSH. Solution of test compounds and suspension of receptor containing cells were prepared and mixed as described above; the mixture was incubated for 15 min., and an EC50 dose (approximately 10 nM alpha-MSH) was added to the cells. The assay was terminated at 45 min. and cAMP quantitated as above. Percent inhibition was determined by comparing the amount of cAMP produced in the presence to that produced in the absence of test compound.

[0606] C. In Vivo Food Intake Models

[0607] 1) Overnight food intake. Sprague Dawley rats are injected intracerebroventricularly with a test compound in 400 mL of 50% propylene glycol/artificial cerebrospinal fluid one hour prior to onset of dark cycle (12 hours). Food intake is determined using a computerized system in which each rats food is placed on a computer monitored balance. Cumulative food intake for 16 hours post compound administration is measured.

[0608] 2) Food intake in diet induced obese mice. Male C57/B16J mice maintained on a high fat diet (60% fat calories) for 6.5 months from 4 weeks of age are are dosed intraperitoneally with test compound. Food intake and body weight are measured over an eight day period. Biochemical parameters relating to obesity, including leptin, insulin, triglyceride, free fatty acid, cholesterol and serum glucose levels are determined.

[0609] D. Rat Ex Copula Assay

[0610] Sexually mature male Caesarian Derived Sprague Dawley (CD) rats (over 60 days old) are used with the suspensory ligament surgically removed to prevent retraction of the penis back into the penile sheath during the ex copula evaluations. Animals receive food and water ad lib and are kept on a normal light/dark cycle. Studies are conducted during the light cycle.

[0611] 1) Conditioning to Supine Restraint for Ex Copula Reflex Tests. This conditioning takes ˜4 days. Day 1, the animals are placed in a darkened restrainer and left for 15-30 minutes. Day 2, the animals are restrained in a supine position in the restrainer for 15-30 minutes. Day 3, the animals are restrained in the supine position with the penile sheath retracted for 15-30 minutes. Day 4, the animals are restrained in the supine position with the penile sheath retracted until penile responses are observed. Some animals require additional days of conditioning before they are completely acclimated to the procedures; non-responders are removed from further evaluation. After any handling or evaluation animals are given a treat to ensure positive reinforcement.

[0612] 2) Ex Copula Reflex Tests. Rats are gently restrained in a supine position with their anterior torso placed inside a cylinder of adequate size to allow for normal head and paw grooming. For a 400-500 gram rat, the diameter of the cylinder is approximately 8 cm. The lower torso and hind limbs are restrained with a non-adhesive material (vetrap). An additional piece of vetrap with a hole in it, through which the glans penis will be passed, is fastened over the animal to maintain the preputial sheath in a retracted position. Penile responses will be observed, typically termed ex copula genital reflex tests. Typically, a series of penile erections will occur spontaneously within a few minutes after sheath retraction. The types of normal reflexogenic erectile responses include elongation, engorgement, cup and flip. An elongation is classified as an extension of the penile body. Engorgement is a dilation of the glans penis. A cup is defined as an intense erection where the distal margin of the glans penis momentarily flares open to form a cup. A flip is a dorsiflexion of the penile body.

[0613] Baseline and or vehicle evaluations are conducted to determine how and if an animal will respond. Some animals have a long duration until the first response while others are non-responders altogether. During this baseline evaluation latency to first response, number and type of responses are recorded. The testing time frame is 15 minutes after the first response.

[0614] After a minimum of 1 day between evaluations, these same animals are administered the test compound at 20 mg/kg and evaluated for penile reflexes. All evaluations are videotaped and scored later. Data are collected and analyzed using paired 2 tailed t-tests to compared baseline and/or vehicle evaluations to drug treated evaluations for individual animals. Groups of a minimum of 4 animals are utilized to reduce variability.

[0615] Positive reference controls are included in each study to assure the validity of the study. Animals can be dosed by a number of routes of administration depending on the nature of the study to be performed. The routes of administration includes intravenous (IV), intraperitoneal (IP), subcutaneous (SC) and intracerebral ventricular (ICV).

[0616] E. Models of Female Sexual Dysfunction

[0617] Rodent assays relevant to female sexual receptivity include the behavioral model of lordosis and direct observations of copulatory activity. There is also a urethrogenital reflex model in anesthetized spinally transected rats for measuring orgasm in both male and female rats. These and other established animal models of female sexual dysfunction are described in McKenna K E et al, A Model For The Study of Sexual Function In Anesthetized Male And Female Rats, Am. J. Physiol. (Regulatory Integrative Comp. Physiol 30): R1276-R1285, 1991; McKenna K E et al, Modulation By Peripheral Serotonin of The Threshold For Sexual Reflexes In Female Rats, Pharm. Bioch. Behav., 40:151-156, 1991; and Takahashi L K et al, Dual Estradiol Action In The Diencephalon And The Regulation Of Sociosexual Behavior In Female Golden Hamsters, Brain Res., 359:194-207, 1985.

EXAMPLES OF A PHARMACEUTICAL COMPOSITION

[0618] As a specific embodiment of an oral composition of a composition of the present invention, 5 mg of Example 2 is formulated with sufficient finely divided lactose to provide a total amount of 580 to 590 mg to fill a size O hard gelatin capsule.

[0619] As another specific embodiment of an oral composition of a compound of the present invention, 2.5 mg of Example 2 is formulated with sufficient finely divided lactose to provide a total amount of 580 to 590 mg to fill a size O hard gelatin capsule.

[0620] Representative compounds of the present invention were tested and found to bind to the melanocortin-4 receptor. These compounds were generally found to have IC50 values less than 2 μM. Representative compounds of the present invention were also tested in the functional assay and found generally to activate the melanocortin-4 receptor with EC50 values less than 1 μM.

[0621] While the invention has been described and illustrated in reference to certain preferred embodiments thereof, those skilled in the art will appreciate that various changes, modifications and substitutions can be made therein without departing from the spirit and scope of the invention. For example, effective dosages other than the preferred doses as set forth hereinabove may be applicable as a consequence of variations in the responsiveness of the mammal being treated for obesity, diabetes, or sexual dysfunction or for other indications for the compounds of the invention indicated above. Likewise, the specific pharmacological responses observed may vary according to and depending upon the particular active compound selected or whether there are present pharmaceutical carriers, as well as the type of formulation and mode of administration employed, and such expected variations or differences in the results are contemplated in accordance with the objects and practices of the present invention. It is intended, therefore, that the invention be limited only by the scope of the claims which follow and that such claims be interpreted as broadly as is reasonable.

Claims

1. A compound of structural formula I:

2. The compound of claim 1 wherein Q is

3. The compound of claim 2 wherein Q is

4. The compound of claim 3 wherein R4a and R4b are each independently selected from the group consisting of hydrogen, C1-8 alkyl, (CH2)n-aryl, (CH2)n-heteroaryl, (CH2)n-heterocyclyl, (CH2)nC3-6 cycloalkyl, (CH2)nCO2R7 (CH2)nOR7, COC(R7)2NH2, CH2C≡CH, and CH2CHF2; or R4a and R4b and the atoms to which they are attached form a 6-membered ring; R3, R5a, and R5b are each independently hydrogen, C1-4 alkyl, C3-6 cycloalkyl, or aryl; wherein aryl is unsubstituted or substituted with one to three groups independently selected from R6; or R3 and R5a and the carbons to which they are attached form a 5- to 7-membered ring optionally containing an additional heteroatom selected from O, S, and NR7.

5. The compound of claim 4 wherein R4a and R4b are each independently selected from the group consisting of hydrogen, C1-4 alkyl, CH2-aryl, CH2-heteroaryl, CH2-heterocyclyl, (CH2)0-1C3-6 cycloalkyl, CH2CO2R7 (CH2)2OR7, COC(R7)2NH2, CH2C≡CH, and CH2CHF2; or R4a and R4b and the atoms to which they are attached form a 6-membered ring; R3, R5a, and R5b are each independently hydrogen, C1-4 alkyl, C3-6 cycloalkyl, or phenyl; wherein phenyl is unsubstituted or substituted with one to three groups independently selected from R6; or R3 and R5a and the carbons to which they are attached form a 6-membered ring optionally containing an additional heteroatom selected from O, S, and NR7.

6. The compound of claim 1 wherein R1 is CHR7-aryl, CHR7OCHR7-aryl, or CHR7-heteroaryl wherein aryl and heteroaryl are optionally substituted with one or two groups independently selected from R6.

7. The compound of claim 6 wherein R1 is benzyl optionally substituted with one or two groups selected from halogen, C1-4 alkyl, C1-4 alkoxy, CN, CF3, and OCF3.

8. The compound of claim 7 wherein R1 is 4-chlorobenzyl; 4-fluorobenzyl; 3,4-difluorobenzyl; 3,5-difluorobenzyl; 2-cyano-4-fluorobenzyl; or 4-methoxybenzyl.

9. The compound of claim 1 wherein R2 is H or CH3.

10. The compound of claim 1 wherein X is C1-6 alkyl, (CH2)n-aryl, (CH2)n-heteroaryl, (CH2)n-heterocyclyl, (CH2)nC(O)N(R8)(R8), (CH2)nCO2R8, (CH2)nOR8, (CH2)nS(O)0-2R8, (CH2)nNHC(O)R8, (CH2)nOC(O)NR8R8, or (CH2)nNR8SO2R8; wherein aryl and heteroaryl are optionally substituted with one to three groups selected from R6; heterocyclyl is optionally substituted with one to three groups selected from R6 and oxo; the (CH2)n group is optionally substituted with one to three groups selected from R7, halogen, S(O)0-2R7, N(R7)2, and OR7; and R8 is each independently selected from H, C1-8 alkyl, and C3-6 cycloalkyl optionally substituted with one to three groups selected from R6 and oxo; or two R8 groups together with the atoms to which they are attached form a 5- to 8-membered mono- or bi-cyclic ring system optionally containing an additional heteroatom selected from O, S, NR7, NBoc, and NCbz.

11. The compound of claim 10 wherein X is C 1-6 alkyl, (CH2)0-1-heteroaryl, CH2-heterocyclyl, CO2R8, CH2OR8, CH2S(O)0-2R8, NHC(O)R8, CH2NR8SO2R8, CH2OC(O)NR8R8, CH2NR8SO2R8, or C(O)N(R8)(R8); wherein heteroaryl is optionally substituted with one to three groups selected from R6; heterocyclyl is optionally substituted with one to three groups selected from R6 and oxo; and R8 is each independently selected from H, C1-8 alkyl, and C3-6 cycloalkyl optionally substituted with one to three groups selected from R6 and oxo; or two R8 groups together with the atoms to which they are attached form a 5- to 8-membered mono- or bi-cyclic ring system optionally containing an additional heteroatom selected from O, S, NR7, NBoc, and NCbz.

12. The compound of claim 1 wherein Y is C1-8 alkyl, (CH2)nC3-7 cycloalkyl, (CH2)n-aryl, (CH2)n-heterocyclyl, or (CH2)n-heteroaryl; wherein aryl and heteroaryl are optionally substituted with one to three groups selected from R6; and (CH2)n, alkyl, cycloalkyl, and heterocyclyl are optionally substituted with one to three groups selected from R6 and oxo.

13. The compound of claim 12 wherein Y is cyclohexyl, cycloheptyl, cyclopentyl, or C1-6 alkyl; wherein alkyl and cycloalkyl are unsubstituted or substituted with one to three groups selected from R6 and oxo.

14. The compound of claim 13 wherein Y is cyclohexyl or C1-6 alkyl, wherein the cyclohexyl and alkyl groups are unsubstituted or substituted with one to three groups selected from R6 and oxo.

15. The compound of claim 1 wherein the carbon atom marked with * has the R configuration.

16. The compound of claim 1 wherein X is selected from the group consisting of:

17. The compound of claim 16 of structural formula Ia selected from the group consisting of

18. The compound of claim 16 of structural formula Ib selected from the group consisting of

19. The compound of claim 16 selected from the group consisting

20. The compound of claim 16 of structural formula Ic selected from the group consisting of

21. A method for the treatment or prevention of disorders, diseases or conditions responsive to the activation of the melanocortin receptor in a mammal in need thereof which comprises administering to the mammal a therapeutically effective amount of a compound according to claim 1.

22. The method of claim 21 wherein the melanocortin receptor is the melanocortin-4 receptor.

23. A method for the treatment or prevention of obesity in a mammal in need thereof which comprising administering to a mammal a therapeutically effective amount of a compound according to claim 1.

24. A method for the treatment or prevention of diabetes mellitus in a mammal in need thereof comprising administering to a mammal a therapeutically effective amount of a compound according to claim 1.

25. A method for the treatment or prevention of male or female sexual dysfunction in a mammal in need thereof comprising administering to a mammal a therapeutically effective amount of a compound according to claim 1.

26. A method for the treatment or prevention of erectile dysfunction in a mammal in need thereof comprising administering to a mammal a therapeutically effective amount of a compound according to claim 1.

27. A pharmaceutical composition which comprises a compound of claim 1 and a pharmaceutically acceptable carrier.

28. The pharmaceutical composition of claim 27 further comprising a second active ingredient selected from the group consisting of an insulin sensitizer, an insulin mimetic, a sulfonylurea, an α-glucosidase inhibitor, an HMG-CoA reductase inhibitor, a sequestrant cholesterol lowering agent, a β3 adrenergic receptor agonist, a neuropeptide Y antagonist, a type V cyclic-GMP-selective phosphodiesterase inhibitor, an α2-adrenergic receptor antagonist, and a dopamine receptor agonist.

29. The pharmaceutical composition of claim 28 wherein the second active ingredient is selected from the group consisting of a type V cyclic-GMP-selective phosphodiesterase inhibitor, an α2-adrenergic receptor antagonist, and a dopamine receptor agonist.

30. The pharmaceutical composition of claim 29 wherein the type V cyclic-GMP-selective phosphodiesterase inhibitor is sildenafil citrate or IC-351.

31. A method of treating male or female sexual dysfunction in a mammal in need thereof, comprising administering to the mammal a therapeutically effective amount of the composition of claim 27.

32. A method of treating male or female sexual dysfunction in a mammal in need thereof, comprising administering to the mammal a therapeutically effective amount of the composition of claim 29.

33. The method of claim 31 wherein the male sexual dysfunction is erectile dysfunction.

34. The method of claim 32 wherein the male sexual dysfunction is erectile dysfunction.