60/495, 757, filed August 14,2003.

BACKGROUND OF THE INVENTION FIELD OF THE INVENTION This invention relates to compounds which are VLA-1 integrin antagonists. This invention also relates to compositions containing such compounds and methods of treatment using such compounds in treating diseases mediated, at least in part, by the VLA-1 integrin.

STATE OF THE ART Integrins are heterodimeric cell surface proteins composed of two noncovalently linked poly- peptide chains, a and. Integrins are the major receptor for cell adhesion to extracellular matrix and play important roles in certain cell-cell and cell-matrix adhesion events. These integrin-medi- ated adhesion events are critical for both normal and pathophysiological processes during cell activa- tion, migration, proliferation and differentiation (for reviews see Hynes (1992) Cell 69 : 11; Springer (1994) Cell 76: 301 ; Hynes (2002) Cell 110 : 673).

VLA-1 (very late antigen-1) is an integrin heterodimer composed of an alpha chain (CD49a, al) and a beta chain (CD29, ßl). VLA-1 is one member of a family of four pi integrin molecules that have been shown to bind to the extracellular matrix pro- teins, collagen and laminin. The ßl integrin colla- gen receptors include alpl (VLA-1), a2ßl (VLA-2), alOßl and allpl. These four collagen receptors share overlapping but distinct expression profiles.

They also appear to have distinct ligand preferences in vitro (Tulla et al. , (2001) J. Biol. Chem.

51: 48206). For example, alpl has been shown to bind more effectively to type IV collagen than type I collagen while a2pl binds to type I collagen better than to type IV collagen (Dickeson et al. , (1999) J.

Biol. Chem. 274: 32182).

VLA-1 is expressed on smooth muscle cells, microvascular endothelial cells, fibroblasts, osteo- blasts and chondrocytes. In addition, VLA-1 is also expressed on activated cells of the immune system including effector T cells, macrophages, and NK cells (de Fougerolles et al. , (2000) J. Clin. In- vest. 105 : 721) ; however, it does not appear to be expressed on B cells or neutrophils. VLA-1 is ex- pressed on T cells in various disease states includ- ing in the joints of arthritis patients (HemLer et al. , (1986) J. Clin. Invest. 78: 696), lesions of giant cell arteritis patients (Schaufelberger et al. , (1993) Clin. Exp. Immunol. 91 : 421), arterio- sclerotic plaques, thyroid infiltrates of patients with autoimmune thyroid disease (Paolieri et al.,

(1992) J. Endocrinol. Invest. 15 : 63) and lungs of chronic bronchitis (Saetta et al. , (1993) Am. Rev.

Respir. Dis. 147 : 301), sarcoidosis (reviewed in HemLer (1990) Annu. Rev. Immunol. 8 : 365) and asthma patients (Corrigan et al. , (1991) Int. Arch. Allergy Appl. Immunol. 94 : 270).

Despite this broad expression profile, al null mice generated by homologous recombination are viable and fertile and have no overt phenotype, demonstrating that the molecule is not required for development (Gardner et al. , (1996) Dev. Biol.

175 : 301). In addition, no increased incidence of infection was noted in the mutant mice. While em- bryonic fibroblasts derived from mutant animals show a striking absence of adhesion to collagen IV, they show no deficit in adhesion to collagen I. Despite the absence of an overt phenotype, no compensatory upregulation of other collagen binding receptors could be identified, suggesting instead that VLA-1 may have redundant roles during development.

Together these data suggest that inhibiting VLA-1 function should be nontoxic.

Inhibiting VLA-1 function using al null mice and/or blocking anti-al antibodies has shown efficacy either prophylactically or therapeutically or both in several animal models of inflammatory disease including 1) delayed-type hypersensitivity as a model of general inflammatory disease (de Fougerolles et al. , (2000) J. Clin. Invest.

105 : 721) ; 2) contact hypersensitivity as a model for skin allergic reactions (A. R. de Fougerolles et al.,

2000, J. Clin. Invest. , 105: 721); 3) anticollagen mAb-induced arthritis as a model of rheumatoid arthritis (de Fougerolles et al., (2000) J. Clin.

Invest. 105 : 721) ; and 4) TNBS-and DSS-induced colitis as models of inflammatory bowel disease (Fiorucci et al. , (2002) Immunity 17: 769; Kriegl- stein et al. , (2002) J. Clin. Invest. 110: 1773).

The mechanism for the reduction in inflam- mation seen with the blocking anti-al mAb and in a1 null mice results from multiple effects. VLA-1 has been shown to mediate adhesion to and migration across collagen matrix. Therefore, VLA-1 expression may be critical for allowing the effector cells to enter the site of inflammation. mAbs against al have also been shown to block collagen-induced cyto- kine release, including release of TNF-a, a key mediator in arthritis (Miyake et al. , (1994) Eur. J.

Immunol. 24: 2000). In addition, VLA-1 also regu- lates matrix metalloproteinase (MMP) expression (Gardner et al. , (1996) Dev. Biol. 175: 301 ; Pozzi et al. (2000) Proc. Natl. Acad. Sci. USA 97 : 2202; Pozzi et al. , (2002) Oncogene 21: 272; Lochter et al., (1999) Mol Biol Cell 10 : 271). Therefore, inhibiting VLA-1 provides a way of reducing inflammation through the synergistic action of a variety of mech- anisms (i. e. , VLA-1 is an upstream regulator of mul- tiple disease promoting factors).

Fibrosis is a common response to chronic injury and represents a paradigm for the cycle of parenchymal wound healing in a variety of tissues (reviewed in Bataller et al. (2001) Semin. Liver

Dis. 21 : 437 ; Bissell. (1998) J. Gastroenterol.

33: 295). When overactive, this wound healing pro- cess can result in pathologic tissue scarring, which results from the progression of several defined steps. First, an infiltrate, consisting of inflam- matory cells and platelets and resident"myofibro- blasts" (identified as hepatic stellate cells in the liver and differentiated mesangial cells in the kidney), accumulates at the site of injury. Second, the local. extracellular matrix (ECM) is altered by de novo production of collagen by the myofibro- blasts. Third, the myofibroblasts migrate and align within the wound site and proliferate. Finally, the myofibroblasts contract the collagen, forming the fibrotic scar which contributes to tissue dysfunc- tion. It is generally believed that a similar pro- cess results in scarring within tissues of the liver, kidney, lung, and skin.

VLA-1 is expressed on myofibroblasts in vitro and in vivo and is believed to regulate their pathologic functions. Alports syndrome is a genetic disorder characterized by progressive glomeruloneph- ritis resulting in fibrosis of the kidneys and ulti- mately kidney failure. Alports syndrome affects approximately 1 in 5000 people and is caused by mutations in the type IV collagen genes. This con- dition has been mimicked in mice by knocking out the gene of the a3 chain of type IV collagen (Alport mouse). Double knockout mice for both type IV collagen and al integrin have a delayed onset and slowed progression of glomerular disease (Cosgrove

et al., (2000) Am. J. Pathol. 157 : 1649). In addi- tion, inhibition of TGF-ffi1 with a soluble receptor construct had a synergistic effect with the inacti- vation of al, slowing the onset and severity of glomerular disease. These results correlated with a dramatic decrease in the accumulation of myofibro- blasts and macrophages in the tubular interstitium of double knockout mice (Sampson et al. , (2001) J.

Biol. Chem. 276: 34182). Another report studying the effects of al expression in transfected glomerular mesangial cells showed that VLA-1 expression levels influenced the cell growth, cell size and collagen matrix remodeling ability of these cells (Kagami et al. , (2000) Kidney Int. 58 : 1088). In addition, al mAb blocks hepatic stellate cell adhesion to colla- gen and endothelin-stimulated hepatic stellate cell- mediated contraction of collagen lattices in vitro, and VLA-1 is the sole integrin utilized by contract- ing hepatic stellate cells in vivo (Racine Sampson et al. , (1997) J. Biol. Chem. 272: 30911). Further- more, blocking anti-al antibody has shown efficacy therapeutically in two independent models of fi- brotic kidney disease (Kagami et al. , (2002) Lab.

Invest. 82 : 1219; Cook et al. , (2002) Am. J. Pathol.

161 : 1265).

VLA-1 may also play a role in regulation of tumor vascularization (angiogenesis) and tumor cell metastasis in many forms of cancer. For exam- ple, VLA-1 may regulate tumor angiogenesis by two distinct mechanisms: 1) by regulating the prolif- eration potential of the vascular endothelial fibro-

blasts (Pozzi et al. , (1998) J. Cell. Biol. 142 : 587 ; Senger et al. , (2002) Am. J. Pathol. 160 : 195), and 2) by regulating the production of matrix metallo- proteinase 9 which in turn regulates the activity of angiostatin, a potent angiogenesis inhibitor (Pozzi et al. , (2000) PNAS 97 : 2202 ; Pozzi et al. , (2002) Oncogene 21: 272). Furthermore, the metastatic po- tential of malignant melanoma cell lines correlates with the expression level of VLA-1 (Schadendorf et al. , (1996) Br. J. Cancer, 74: 194), and blocking VLA-1 binding inhibits tumor cell invasion across reconstituted basement membrane through inhibition of matrix metalloproteinase 3 (stromelysin-1) ex- pression (Lochter et al. , (1999) Mol. Biol. Cell, 10 : 271).

Currently, there are only two descriptions for VLA-1 inhibitors in the patent literature, and both describe large molecular weight polypeptides.

The first is a mAb to VLA-1 (WO 02/083854-A2) and the second is a disintegrin isolated from cobra ven- om (WO 02/22571-A2). Therefore, there still exists a need in the art for low molecular weight antag- onists, specific inhibitors of VLA-1-dependent cell adhesion that have improved pharmacokinetic and pharmacodynamic properties such as oral bioavail- ability and significant duration of action. Such compounds would prove to be useful for the treat- ment, prevention or suppression of various pathol- ogies mediated by VLA-1 binding and cellular adhe- sion, migration, activation or differentiation.

SUMMARY OF THE INVENTION The present invention provides aminopiper- idine amide compounds which are antagonists to the VLA-1 integrin.

In one of its composition aspects, this invention is directed to a compound of Formula I: wherein A and B, together with the nitro- gen atom bound thereto, form a 4-8 membered nitrogen containing heterocyclic group containing 1 to 2 nitrogen atoms, wherein said heterocyclic group may be optionally substituted with 1 to 3 additional substituents each independently selected from the group consisting of alkyl, substituted alkyl, aryl, substituted aryl, cycloalkyl, substituted cyclo- alkyl, heteroaryl, substituted heteroaryl, hetero- cyclic and substituted heterocyclic, hydroxy, alkoxy, thioalkyl, and halo, wherein the one or more alkyl and sub- stituted alkyl substituents, if present, may be attached to either a carbon or a nitrogen atom in said heterocyclic group, wherein the one or more hydroxy, alkoxy, alkylsulfanyl and halo substituents, if present, may not be attached to a nitrogen atom in said hetero- cyclic group,

and wherein the one or more hydroxy, alkoxy, and halo substituents, if present, may not be attached to a carbon atom which is adjacent to a nitrogen atom in said heterocyclic group, and further wherein A together with the nitrogen atom bound thereto form a 4-8 membered heterocyclic group containing two nitrogen atoms, then the two nitrogen atoms are either adjacent to each other, or are separated by at least two carbon atoms, R1 is selected. from the group consisting of alkyl, substituted alkyl, aryl, substituted aryl, cycloalkyl, substituted cycloalkyl, heteroaryl, sub- stituted heteroaryl, heterocyclic, and substituted heterocyclic; R2 and R3 are independently selected from the group consisting of hydrogen, fluoroalkyl, and alkyl ; R4 is selected from the group consisting of hydrogen, alkyl, aryl, heteroaryl, heterocyclic, and cycloalkyl ; R6 and R7 are independently selected from the group consisting of hydrogen, halo, alkyl, sub- stituted alkyl, amino, substituted amino, aminocar- bonyloxy, aminoacyl, aminosulfonyl, sulfonylamino, acylamino, aminoacylamino, heterocyclic, substituted heterocyclic, heteroaryl, substituted heteroaryl, aryl, substituted aryl, ORa, acyloxy, oxycarbonyl- amino, thioalkyl, thioaryl, thioalkylaryl, thioalk- ylheteroaryl, NHS02NRaRa, SC (O) Ra, and SC (O) NRa2,

or R6 and R7, together with the carbon atom bound thereto, form a cycloalkyl, substituted cyclo- alkyl, heterocyclic, or a substituted heterocyclic group ; R8 is selected from the group consisting of CRaRaC (O) ORa, CO2Ra, C (O) NRa2, C (O) NRaORa, C (0) NHSO2Rat CRaRa heteroaryl (e. g., tetrazolyl), CRaRa substituted heteroaryl, CN, and (CRa2) pOH, wherein p is 0,1 or 2 ; R9 and Rl° are independently hydrogen, halo, alkyl, substituted alkyl, amino, substituted amino, aminocarbonyloxy, aminoacyl, acylamino, aminoacylamino, aminosulfonyl, sulfonylamino, het- erocyclic, substituted heterocyclic, heteroaryl, substituted heteroaryl, aryl, substituted aryl,-ORa, acyloxy, oxycarbonylamino, thioalkyl, thioaryl, thioalkylaryl, thioalkylheteroaryl, NHSO2NRa Ra, SC (0) Ra, and-SC (O) NRa2, or R7-C-C-R9 can form a cycloalkylene, cycloalkenylene, heterocyclene, or heterocyclenylene group, or R9 and R10, together with the carbon atom attached thereto, form a >C=O (oxo) group, with the proviso that when R9 and R10 form an oxo group, R8 is not-CN, or R9 and R10, or R5 and R7, together with the carbon atom attached thereto, form a vinyl group of the formula >C=CR11R12 where R11 and R12 are inde- pendently selected from the group consisting of hydrogen, alkyl, substituted alkyl, aryl, and sub- stituted aryl,

or R9-C-R10, together with the carbon atom attached thereto, form a group selected from the group consisting of cycloalkyl, substituted cyclo- alkyl, heterocyclic, and substituted heterocyclic group ; wherein each Ra is independently selected from the group consisting of hydrogen, alkyl, sub- stituted alkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, heterocyclic, and substi- tuted heterocyclic, and, in the case of-NRa2, each Ra, together with the nitrogen atom bound thereto, can form a heterocyclic or substituted heterocyclic group or in the case of CRaRaC (O) ORa, the two Ra can be an oxo group, with the proviso that when the two Ra are an oxo group, then R9 and Rl° are not an oxo group; and pharmaceutically acceptable salts, prodrugs, or tautomers thereof.

In a preferred embodiment are compounds of Formula I exhibiting a biological activity of at least fifty percent inhibition of VLA-1 when tested at a concentration of 50 uM. In a more preferred embodiment are compounds of Formula I exhibiting a biological activity of at least fifty percent inhi- bition VLA-1 when tested at a concentration of 40 pM. In a still more preferred embodiment are com- pounds of Formula I exhibiting a biological activity of at least fifty percent inhibition VLA-1 when tested at a concentration of 20 uM.

In a preferred embodiment of this inven- tion, A and B, together with the nitrogen atom bound

thereto, preferably form a piperidine, pyrrolidine, or azetidine ring.

In another preferred embodiment, R1 is selected from the group consisting of aryl, substi- tuted aryl, heteroaryl, and substituted heteroaryl, and even more preferably R1 is selected from the group consisting of substituted aryl and substituted heteroaryl.

Examples of particularly preferred R1 groups are selected from the group consisting of : 2, 3-dichlorophenyl ; 2, 3-dichloro-4- (2-methoxyphenylthio)- phenyl ; 2, 3-dichloro-4- (4-fluorophenylthio) phenyl ; 4-methylthio-2, 3-dichlorophenyl; and 2,3-dichlorobenzo [b] thiophen-4-yl.

In another preferred embodiment, R2, R3, R4, and R6 are hydrogen.

In yet another preferred embodiment, R is selected from the group consisting of hydrogen, halo, alkyl, substituted alkyl, substituted alkyl- ene, aryl, and heteroaryl. Particularly preferred R groups are selected from the group consisting of: hydrogen ; fluoro ; methyl ; aminomethyl ; phenylamidomethylene ; (2-methoxyphenyl) amidomethylene ; (2-chlorophenyl) amidomethylene; phenyl;

pyridin-3-yl; quinolin-3-yl ; phenylaminbcarbonylaminomethylene ; propylamidomethylene; methoxymethyleneamidomethylene ; t-butyl-amidomethylene ; methylthioethyleneamidomethylene ; <BR> <BR> <BR> <BR> <BR> t-butyl-methyleneamidomethylene ;<BR> <BR> <BR> <BR> <BR> <BR> <BR> <BR> cyclopropylamidomethylene ;<BR> <BR> <BR> <BR> <BR> <BR> <BR> cyclopentylamidomethylene ; cyclohexylamidomethylene; 4-N-acetylpiperidinylamidomethylene ; cyclopentylmethylenamidomethylene; <BR> <BR> <BR> <BR> <BR> piperidinyl-N-ethyleneamidomethylene ;<BR> <BR> <BR> <BR> <BR> <BR> <BR> <BR> 2-methoxyphenylamidomethylene ; i-propylamidomethylene. ; cyclobutylamidomethylene ; <BR> <BR> <BR> <BR> <BR> 2-pyridynylamidomethylene ;<BR> <BR> <BR> <BR> <BR> <BR> <BR> 3-pyridinylamidomethylene ; 4-pyridinylamidomethylene ; 2-thiophenylamidomethylene ; 2-furanoylamidomethylene ; benzo [1, 3] -dioxole-5-amidomethylene; <BR> <BR> <BR> <BR> <BR> 3-methoxyphenylmethyleneamidomethylene ;<BR> <BR> <BR> <BR> <BR> <BR> <BR> <BR> 4-methoxyphenylamidomethylene ; 2-thiophenylmethyleneamidomethylene; cyclopropyl-1-phenyl-l-amidomethylene ; phenylethyleneamidomethylene; phenyloxymethyleneamidomethylene ; 4-chlorophenyloxy-dimethylmethyleneamido- methylene ;

cyclopentyl-1- (4-chlorophenyl)-1- amidomethylene; 2-phenylcyclopropaneamidomethylene; phenylmethyleneoxymethyleneamidomethylene; methylaminoamidomethylene ; morpholinylamidomethylene ; phenylmethyleneaminoamidomethylene; phenylethyleneaminoamidomethylene ; methoxyamidomethylene ; phenylmethyleneoxyamidomethylene ; R-phenylamido ; S-phenylamido ; methylamido ; propylamido ; methoxymethyleneamido ; t-butyl-amido ;' <BR> <BR> <BR> <BR> thiomethylethyleneamido ;<BR> <BR> <BR> <BR> <BR> <BR> t-butylmethyleneamido ; cyclopropylamido ; cyclopentylamido ; cyclohexylamido ; 4-N-acetylipiperidinylamido ; cyclopentylmethylene amido ; piperidynyl-N-ethylene amido ; <BR> <BR> <BR> <BR> 3-methoxyphenylamido ;<BR> <BR> <BR> <BR> <BR> <BR> 4-methoxyphenylamido ;<BR> <BR> <BR> <BR> <BR> <BR> i-propylamido ; 3-chlorophenylamido; 4-chlorophenylamido ; cyclobutylamido; 2-pyridinylamido ;

3-pyridinylamido ; 4-pyridinylamido; 2-thiophenylamido ; 2-furanoylamido; 5-isoxazolylamido; 4-biphenylamido; phenylmethylene amido ; 4-chlorophenylmethylene amido; <BR> <BR> <BR> <BR> 3-methoxyphenylmethyleneamido ;<BR> <BR> <BR> <BR> <BR> 4-methoxyphenylmethyleneamido ;<BR> <BR> <BR> <BR> <BR> <BR> 2-thiophenemethyleneamido ; cyclopropyl-1-phenyl-1-amido ; phenylethyleneamido ; phenyloxymethyleneamido; 4-chlorophenyloxy-1, 1-dimethylmethylene- amido ; phenylethyl-ene-amido ; phenylmethoxyloxymethyleneamido ; methylaminoamido ; dimethylaminoamido; morpholine-N-amido ; phenylmethyleneaminoamido; phenylethyleneaminoamido ; methoxyamido; phenylmethyleneoxyamido; phenylsulfonamido ; 3-methoxyphenylamido ; and 3-chlorophenylamido.

Preferred R8 groups are selected from the groups consisting of CRaRaC (O) ORa,-CO2Ra, where Ra is

as defined above, and, when R9 and R10 form an oxo group, then R8 is preferably hydroxy.

Particularly preferred R8 groups are selec- ted from the group consisting of: carboxyl; -CH(C6H4)COOH; - CH2COOH ; and when R9 and R10 form an oxo group, hydroxy.

Preferred R9 groups are selected from the group consisting of hydrogen, alkyl, aminoacyl, acylamino, and aryl.

Particularly preferred R9 groups are se- lected from the group consisting of : methyl ; hydrogen ; phenyl and - NHC (0) CF3.

Preferred R1° groups are selected from the group consisting of hydrogen and alkyl. Particular- ly preferred R1° groups are selected from the group consisting of methyl and hydrogen.

In other preferred embodiments, R7 and R9, together with the carbon atoms bound thereto form a cycloalkylene or cycloalkenylene group such as, for example, cyclopropylene or cyclohexenylene ; R9 and R10, together with the carbon atom bound thereto form an oxo group [>C (0)] or a vinyl group of the formula >C=CR11R12 where R11 and R12 are as defined above such as, for example, >C=CH2.

Aminopiperidine amide derivatives within the scope of this invention are exemplified by those set forth in Tables I, II, and III as follows: Table 1 S1=H, CH3, CH2Ph, CH2CH2Ph at 2 of 3 position Ex. R1 R7 R9 R8 R10 1 2, 3-dichloro- H-CH3 C (O) OH H 2 2, 3-dichloro-# H =CH2-C (O) OH-- 3 2,3-dichloro-# H H -C(O)OH H 2,3-dichloro-4-(2- R7-C-C-R9=3- 4 -- -C(O)OH H methoxyphenylthio)-# Cyclohexene 2,3-dichloro-4-(2- -CH(#)- 5 H H H methoxyphenylthio)- = COOH 2, 3-dichloro-4- (2- R-C-C-R = methoxyphenylthio)-O cyclopropyl 2,3-dichloro-4-(2- 7 H -CH3 -CH2-COOH H methoxyphenylthio)-# 2,3-dichloro-4-(2- methoxyphenylthio)-O 2,3-dichloro-4-(2- NHC(O)- H methoxyphenylthio)-O CF3 10 2,3-dichloro-4-(2- OH H methoxyphenylthio)-# 11 2,3-dichloro-4-(2- methoxyphenylthio)-# 12 2,3-dichloro-4-(2- OH H methoxyphenylthio)-O 13 2,3-dichloro-4-(2 methoxyphenylthio)-O 2,3-dichloro-4-(4- R9/R10 is H -OH -- fluorophenylthio)-# =O 2,3-dichloro-4-(4- R9/R10 is F -OH -- fluorophenylthio)-@ =O 20 2, 3-dichlorobenzo--CH3 H-C (O) OH H [b] thiophen-4-yl 21 2, 3-dichlorobenzo-H-CH3-C (O) OH H [b] thiophen-4-yl 2,3-dichlorobenzo- OH H [b]thiophen-4-yl 23 2,3-dichlorobenzo- H-C (O) OH H [b]thiophen-4-yl 24 2,3-dichlorobenzo- Phenylamido-H-C (O) OH H [b]thiophen-4-yl methylene Table 1 S1=H, CHg, CH2Ph, CH2CH2Ph at 2 of 3 position Ex. R1 R7 R9 R8 R10 (2-methoxy- 2,3-dichlorobenzo- 25 phenyl)amido H -C(O)OH H [b] thiophen-4-yl methylene (2-Chloro- 2,3-dichlorobenzo- 26 phenyl)amido H -C(O)OH H [b]thiophen-4-yl ethylene 2, 3-dichlorobenzo- CH2NHC (0) NH- [b] thiophen-4-yl. O 31 2,3 dichlorobenzo- [b] thiophen-4-yl 32 2, 3-dichlorobenzo-pyridin-3-yl H-C (O) OH H [b] thiophen-4-yl 2,3-dichlorobenzo- quinolin-3- 33 H -C(O)OH H [b]thiophen--4yl yl

O=phenyl Additional compounds are those having the structure set forth in Table 1, wherein R1 is 2,3- dichlorobenzo [b] thiophen-4-yl ; R9 and R10 are H; R8 is -C(O) OH; and R7 is propylamidomethylene; methoxymethyleneamidomethylene; t-butyl-amidomethylene; methylthioethyleleneamidomethylene ; t-butyl-methyleneamidomethylene; cyclopropylamidomethylene; cyclopentylamidomethylene; cyclohexylamidomethylene ; 4-N-acetylpiperidinylamidomethylene; cyclopentylmethylenamidomethylene;

piperidinyl-N-ethyleneamidomethylene; 2-methoxyphenylamidomethylene; i-propylamidomethylene; cyclobutylamidomethylene; 2-pyridynylamidomethylene; 3-pyridinylamidomethylene; 4-pyridinylamidomethylene ; 2-thiophenylamidomethylene ; 2-furanoylamidomethylene ; benzo [1, 3]-dioxole-5-amidomethylene ;. <BR> <BR> <BR> <BR> <P> 3-methoxyphenylmethyleneamidomethylene ;<BR> <BR> <BR> <BR> <BR> 4-methoxyphenylamidomethylene ;, 2-thiophenylmethyleneamidomethylene ; cyclopropyl-1-phenyl-1-amidomethylene; phenylethyleneamidomethylene ;, phenyloxymethyleneamidomethylene ; 4-chlorophenyloxy-dimethylmethyleneamido- methylene; cyclopentyl-1- (4-chlorophenyl)-1-amido- methylene; 2-phenylcyclopropaneamidomethylene; phenylmethyleneoxymethyleneamidomethylene; methylaminoamidomethylene; morpholinylamidomethylene; phenylmethyleneaminoamidomethylene; phenylethyleneaminoamidomethylene; methoxyamidomethylene; phenylmethyleneoxyamidomethylene ; R-phenylamido; S-phenylamido; methylamido ;

propylamido ; methoxymethyleneamido ; t-butyl-amido ; thiomethylethyleneamido ; t-butylmethyleneamido; cyclopropylamido ; cyclopentylamido ; cyclohexylamido; 4-N-acetylipiper. idinylamido ; cyclopentylmethylene amido; piperidynyl-N-ethylene amido; <BR> <BR> <BR> <BR> 3-methoxyphenylamido ;<BR> <BR> <BR> <BR> <BR> <BR> 4-methoxyphenylamido ; i-propylamido; 3-chlorophenylamido; 4-chlorophenylamido; cyclobutylamido; 2-pyridinylamido; 3-pyridinylamido ; 4-pyridinylamido; 2-thiophenylamido; 2-furanoylamido ;.

5-isoxazolylamido; 4-biphenylamido ; phenylmethylene amido ; 4-chlorophenylmethylene amido; 3-methoxyphenylmethyleneamido; 4-methoxyphenylmethyleneamido; 2-thiophenemethyleneamido; cyclopropyl-1-phenyl-1-amido ; phenylethyleneamido ;

phenyloxymethyleneamido; 4-chlorophenyloxy-1, 1-dimethylmethylene- amido; phenylethyl-ene-amido ; phenylmethoxyloxymethyleneamido; methylaminoamido ; dimethylaminoamido; morpholine-N-amido; phenylmethyleneaminoamido ; phenylethyleneaminoamido ; methoxyamido; phenylmethyleneoxyamido; phenylsulfonamido ; 3-methoxyphenylamido; 3-chlorophenylamido ; phenyl ; phenylmethylene; phenylethylene ; 2-bromo-phenylmethylene; 3-bromo-phenylmethylene; 4-bromo-phenylmethylene ; 2-biphenyl-methylene; 3-biphenyl-methylene ; 4-biphenyl-methylene ; and methoxy.

Table 2 Example Ri R9 RB 14 2, 3-dichloro-4-(2-_ methoxyphenylthio)-# is 2, 3-dichloro-4-(4- methoxyphenylthio)-O 2,3-dichloro-4-(2- 16 -CH3 -C(O)OH methoxyphenylthio)-# O=phenyl

Table 3 Example R1. R7 4-methylthio-2,3- 28 methyl dichlorophenyl 2,3- 29 dichlorobenzo [b] thiophen- methyl 4-yl

Specific compounds within the scope of this invention are exemplified by the following com- pounds: (S)-N- 1- [3- (2, 3-dichlorophenyl)-acryloyl]-piper- idin-4-yl}-2-methyl-succinamic acid; <BR> <BR> N- 1- [3- (2, 3-dichlorophenyl)-acryloyl]-piperidin-4- yl}-2-methylene-succinamic acid;

N- 1- [3- (2, 3-dichlorophenyl)-acryloyl]-piperidin-4- yl}-succinamic acid; Cis-6-(1-{3-[2,3-dichloro-4-(2-methoxy-phenylsul- <BR> <BR> <BR> <BR> fanyl)-phenyl]-acryloyl}-piperidin-4-ylcarbamoyl)- cyclohex-3-enecarboxylic acid; <BR> <BR> <BR> <BR> Cis-6- (1- {3- [2, 3-dichloro-4- (2-methoxy-phenylsul-<BR> <BR> <BR> <BR> <BR> <BR> fanyl)-phenyl]-acryloyl}-piperidin-4-ylcarbamoyl)- cyclohex-3-enecarboxylic acid; <BR> <BR> <BR> <BR> 4- (1- {3- [2, 3-dichloro-4- (2-methoxy-phenylsulfanyl)-<BR> <BR> <BR> <BR> <BR> <BR> phenyl]-acryloyl}-piperidin-4-ylcarbamoyl)-2-phenyl- butyric acid; 2-(1-{3-[2,3-dichloro-4-(2-methoxy-phenylsulfanyl)- phenyl]-acryloyl}-piperidin-4-ylcarbamoyl) cyclo- propanecarboxylic acid; <BR> <BR> <BR> <BR> 4- (1- {3- [2, 3-dichloro-4- (2-methoxy-phenylsulfanyl)-<BR> <BR> <BR> <BR> <BR> <BR> phenyl]-acryloyl}-piperidin-4-ylcarbamoyl)-3-methyl- butyric acid; <BR> <BR> <BR> <BR> 4- (1- {3- [2, 3-dichloro-4- (2-methoxy-phenylsulfanyl)-<BR> <BR> <BR> <BR> <BR> <BR> phenyl]-acryloyl}-piperidin-4-ylcarbamoyl)-butyric acid; N-(1-{3-[2,3-dichloro-4-(2-methoxy-phenylsulfanyl)- phenyl]-acryloyl}-piperidin-4-yl}-(S)-2-(2, 2,2- trifluoro-acetylamino) -succinamic acid; <BR> <BR> <BR> <BR> N- (1- {3- [2, 3-dichloro-4- (2-methoxy-phenylsulfanyl)-<BR> <BR> <BR> <BR> <BR> <BR> phenyl]-acryloyl}-piperidin-4-yl)-2-methyl-succinam- ic aid ; <BR> <BR> <BR> <BR> N-(1-{3-[2, 3-dichloro-4-(2-methoxy-phenylsulfanyl)-<BR> <BR> <BR> <BR> <BR> <BR> phenyl]-acryloyl}-piperidin-4-yl)-2-phenyl-succinam- ic acid ;

N- (1- {3- [2, 3-dichloro-4- (2-methoxy-phenylsulfanyl)- phenyl]-acryloyl}-piperidin-4-yl)-2, 2-dimethyl- succinamic acid; <BR> <BR> <BR> <BR> 4- (1- {3- [2, 3-dichloro-4- (2-methoxy-phenylsulfanyl)-<BR> <BR> <BR> <BR> <BR> <BR> phenyl]-acryloyl}-piperidin-3-ylcarbamoyl)-3-methyl- butyric acid; N-6-(1-{3-[2,3-dichloro-4-(2-methoxy-phenylsulfanyl)- phenyl]-acryloyl}-piperidin-3-yl)-(S)-2-(2, 2,2- trifluoro-acetylamino) -succinamic acid; <BR> <BR> <BR> <BR> N- (1- {3- [2, 3-dichloro-4- (2-methoxy-phenylsulfanyl)-<BR> <BR> <BR> <BR> <BR> <BR> phenyl]-acryloyl}-piperidin-3-yl)-2-methyl-succinam- ic acid; <BR> <BR> <BR> <BR> N-(1-{3-[2, 3-dichloro-4-(4-fluoro-phenylsulfanyl)- phenyl]-acryloyl}-piperidin-4-yl)-malonamic acid; <BR> <BR> <BR> <BR> N- (l- {3- [2, 3-dichloro-4- (4-fluoro-phenylsulfanyl)-<BR> <BR> <BR> <BR> <BR> phenyl]-acryloyl}-piperidin-4-yl)-2-fluoro-malonamic acid ; N-{1-[3-(6, 7-dichloro-benzo [b] thiophen-5-yl)-acryl- oyl]-piperidin-4-yl}-(S)-3-methyl-succinamic acid; N- {1- [3- (6, 7-dichloro-benzo [b] thiophen-5-yl)-acryl- oyl]-piperidin-4-yl}-(S)-2-methyl-succinamic acid; N-11- [3- (6, 7-dichloro-benzo [b] thiophen-5-yl)-acryl- oyl]-piperidin-4-yl}-(S)-2-methyl-succinamic acid; <BR> <BR> <BR> <BR> rac-3-aminamethyl-N-1- [3- (6, 7-dichloro-benzo [b]-<BR> <BR> <BR> <BR> <BR> thiophen-5-yl)-acryloyl]-piperidin-4-yl}-succinamic acid trifluoroacetic acid salt; rac-3-(benzoylamino-methyl)-N-{1-[3-(6, 7-dichloro- benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}- succinamic acid;

rac-N- {1- [3- (6, 7-dichlorobenzo [b] thiophen-5-yl)- acryloyl]-piperidin-4-yl}-3- [ (3-methoxy-benzoyl- amino) -methyl] -succinamic acid; rac-3- [ (3-chlorobenzoylamino)-methyl]-N- 1- [3- (6, 7- dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin- 4-yl}-succinamic acid; rac-N-1- [3- (6, 7-dichlorobenzo [b] thiophen-5-yl)- acryloyl]-piperidin-4-yl}-3-[(3-phenyl-ureido)- methyl] -succinamic acid; N- {1- [3- (2, 3-Dichloro-4-methylsulfanyl-phenyl) - acryloyl]-azetidin-3-yl}-3-methyl-succinamic acid ; N- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)-acryl- oyl]-azetidin-3-yl}-3-methyl-succinamic acid ; N-{1-[3-(6,7-Dichloro-benzo [b] thiophen-5-yl)-acryl- oyl]-piperidin-4-yl}-3-phenyl-succinamic acid; N-{1-[3-(6, 7-dichloro-benzo [b] thiophen-5-yl)-acryl- oyl]-piperidin-4-yl}-3-pyridin-3-yl-succinamic acid ; N-{1-[3-(6, 7-dichloro-benzo [b] thiophen-5-yl)-acryl- oyl]-3-methyl-piperidin-4-yl}-3-methyl-succinamic acid; <BR> <BR> <BR> 3- (Butyrylamino-methyl)-N- {1- [3- (6, 7-dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}-succinamic acid N- (1-[3-(6,7-Dichloro-benzo[b]thiophen-5-yl)-acryloyl]-piperid in-4-yl}-3-[(2-methoxy-acetylamino)-methyl]-succinamic acid N- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}-3-[(2,2-dimethyl-pr opionylamino)-emthyl]- succinamic acid N- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yi)-acryloyl]-piperidin-4-yll-3- [ (3-methylsulfanyl-propionylamino)-methyl]- succinamic acid <BR> <BR> <BR> <BR> <BR> <BR> N- {1= [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}-3- [ (3, 3-dimethyl-butyrylamino)-methyl]-succinamic acid <BR> <BR> <BR> <BR> <BR> <BR> 3- [ (Cyclopropanecarbonyl-amino)-methyl]-N- {1- [3- (6, 7-dichloro-benzo [b] thiophen-5-yl)-acryloyt]-piperidin-4-yl}- succinamic acid

3- [ (Cyclopentanecarbonyl-am ino)-m ethyl]-N- {l- [3- (6, 7-dichloro-benzo [b] thiophen-5-yi)-acryloyl]-piperidin-4-yl}- succinamic acid 3-[(Cyclohexanecarbonyl-amino)-methyl]-N-{1-[3-(6,7-dichloro -benzo[b]thiophen-5-yl)-acryloyl]-piperidin-4-yl}-succinamic acid 3-{[(1-Acetyl-piperidine-4-carbonyl)-amino]-methyl)-N-{1-[3- (6,7-dichloro-benzo[b]thiophen-5-yl)-acryloyl]-piperidin-4-y l}- succinamic acid <BR> <BR> <BR> <BR> <BR> <BR> <BR> 3- [ (2-Cyclopentyl-acetylamino)-methyl]-N- {1- [3- (6, 7-dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}-succinamic acid N-{1-[3-(6,7-Dichloro-benzo[b]thiophen-5-yl)-acryloyl]-piper idin-4-yl}-3-[(3-piperidin-4-yl-propionylamino)-methyl]- succinamic acid <BR> <BR> <BR> <BR> <BR> <BR> <BR> N- {3-(6, 7-Dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperldin-4-yl}-3-[(2-meth oxy-benzoylamino)-methyq-succinamic acid <BR> <BR> <BR> <BR> <BR> <BR> <BR> N- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}-3- [ (3-methyl-butyrylamino)-methyl]-succinamic acid 3-[(Cyclobutanecarbonyl-amino)-methyl]-N-{1-[3-(6,7-dichloro -benzo[b]thiophen-5-yl)-acryloyl]-piperidin-4-yl}-succinamic acid <BR> <BR> <BR> <BR> <BR> <BR> <BR> N- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl-3- { [ (pyridine-2-carbonyl)-amino]-methyl}-succinamic acid /V- {1- [3- (6, 7-Dich ! oro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}-3-{[(pyridine-3-car bonyl)-amino]-methyl}-succinamic acid N-{1-[3(6,7-Dichloro-benzy[b]thiophen-5-yl)-acryloyl]-piperi din-4-yl}-3-{[(pyridine-4-carbonyl)-amino]-methyl}-succinami c acid N-{1-[3-(6,7-Dichloro-benzo[b]thiophen-5-yl)-acryloyl]-piper idin-4-yl}-3-{[(thiophene-2-carbonyl)-amino]-methyl}- succinamic acid N- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}-3-{[(furan-2-carbon yl)-amino]-methyl}-succinamic acid 3- { [ (Benzo [1,3] dioxole-5-carbonyl)-amino]-methyl}-N-{1-[3-(6,7-dichloro-ben zo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}- succinamic acid N-{1-[3-(6,7-Dichloro-benzo[b]thiophen-5-yl)-acryloyl]-piper idin-4-yl}-3-{[2-(3-methoxy-phenyl)-acetylamino]-methyl- succinamic acid <BR> <BR> <BR> <BR> <BR> <BR> <BR> N- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}-3- { [2- (4-methoxy-phenyl)-acetylamino]-methyl}- succinamic acid N- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}-3- [ (2-thiophen-2-yl-acetylamino)-methyl]-succinamic acid

N-{1-[3-(6,7-Dichloro-benzo[b]thiophen-5-yl)-acryloyl]-piper idin-4-yl}-3-{[(1-phenyl-cyclopropanecarbonyl)-amino]-methyl }- succinamic acid <BR> <BR> <BR> <BR> <BR> N- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}-3- [ (3-phenyl-propionylamino)-methyl]-succinamic acid N- {1-[3-(6,7-Dichloro-benzo[b]thiophen-5-yl)-acryloyl]-piperid in-4-yl}-3-[(2-phenoxy-acetylamino)-methyl]-succinamic acid <BR> <BR> <BR> <BR> <BR> 3- { [2- (4-Chloro-phenoxy)-2-methyl-propionylamino]-methyl}-N- {1- [3- (6, 7-dichloro-benzo [b] thiophen-5-yl)-acryloyl]- piperidin-4-yl}-succinamic acid 3-({[1-(4-Chloro-phenyl)-cyclopentanecarbonyl]-amino}-methyl )-N-{1-[3-(6,7-dichloro-benzo[b]thiophen-5-yl)-acryloyl]- . piperidin-4-yl}-succinamicacid<BR> <BR> <BR> <BR> <BR> <BR> N- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}-3- { [ (2-phenyl-cyclopropanecarbonyl)-amino]-methyl}- succinamic acid <BR> <BR> <BR> <BR> <BR> 3- [ (2-Benzyloxy-acetylamino)-methyl]-N- {1- [3- (6, 7-dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}-succinamic. acid N- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}-3- [ (3-methyl-ureido)-methyl]-succinamic acid'.

N- {3-(6, 7-Dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}-3-{[(morpholine-4-c arbonyl)-amino]-methyl}- succinamic acid <BR> <BR> <BR> <BR> <BR> 3- (3-Benzyl-ureidomethyl)-N- {1- [3- (6, 7-dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}-succinamic acid.

N- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}-3- (3-phenethyl-ureidomethyl)-succinamic acid v <BR> <BR> <BR> <BR> <BR> N- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}-3- (methoxycarbonylamino-methyl)-succinamic acid 3-S-Benzoylar'nino-N- {1- [3- (6 ; 7-dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}-succinamic acid 3-R-Benzoylamino-N- {1- [3- (6, 7-dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}-succinamic acid 3-R-Acetylamino-N-{1-[3-(6,7-dichloro-benzo[b]thiophen-5-yl) -acryloyl]-piperidin-4-yl}-succinamic acid 3-R-Acetylamino-N-{1-[3-(6,7-dichloro-benzo[b]thiophen-5-yl) -acryloyl]-piperidin-4-yl}-succinamic acid N-R- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}-3-(2-methoxy-acetyl amino)-succinamic acid <BR> <BR> <BR> <BR> <BR> N-R- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}-3- (2, 2-dimethyl-propionylamino)- succinamic acid N-R-{1-[3-(6,7-Dichloro-benzo[b]thiophen-5-yl)-acryloyl]-pip eridin-4-yl}- 3- (3-methylsulfanyl-propionylamino)-succinamic acid

N-R- {1-[3-(6,7-Dichloro-benzo[b]thiophen-5-yl)-acryloyl]-piperid in-4-yl}- 3- (3, 3-dimethyl-butyrylamino)-succinamic acid 3R-- (Cyclopropanecarbonyl-amino)-N- {1- [3- (6, 7-dichloro-benzo [b] thiophen-5-yl) -acryloyl]-piperidin-4-yl}-succinamic acid<BR> <BR> <BR> 3R-- (Cyclopentanecarbonyl-amino)-N- {1- [3- (6, 7-dichloro-benzo [b] thiophen-5-yl)- acryloyl]-piperidin-4-yl}-succinamic acid <BR> <BR> <BR> 3-R- (Cyclohexanecarbonyl-amino)-N- {1- [3- (6, 7-dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl} - succinamic acid <BR> <BR> <BR> <BR> <BR> 3R-- [ (1-Acetyl-piperidine-4-carbonyl)-amino]-N- {1- [3- (6, 7-dichloro-benzo [b] thiophen-5-yl)-acryloyl]- piperidin-4-yl}-succinamic acid 3-R- (2-Cyclopentyl-acetylamino)-N- {1- [3- (6, 7-dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}- succinamic acid N-R- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}-3- (3-piperidin-1-yl-propionylamino) -succinamic acid <BR> <BR> <BR> NR-- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}-<BR> <BR> <BR> 3- (2-methoxy-benzoylamino)-succinamic acid<BR> <BR> <BR> <BR> N-R- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}-3- (4-methoxy-benzoylamino)-succinamic acid <BR> <BR> <BR> N-R- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}-<BR> <BR> 3- (3-methyl-butyrylamino)-succinamic acid 3-R (2-Chloro-benzoylamino)-N- {1- [3- (6, 7-dichloro-benzo [b] thiophen-5-yl)- acryloyl]-piperidin-4-yl}-succinamic acid<BR> <BR> <BR> <BR> 3-R- (4-Chloro-benzoylamino)-N- {1- [3- (6, 7-dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}- succinamic acid 3R--(Cyclobutanecarbonyl-amino)-N-{1-[3-(6,7-dichloro-benzo[ b]thiophen-5-yl)-acryloyl]-piperidin-4-yl} - succinamic acid N-R-{1-[3-(6,7-Dichloro-benzo[b]thiophen-5-yl)-acryloyl]-pip eridin-4-yl}-3-[(pyridine-2-carbonyl)-amino]- succinamic acid <BR> <BR> <BR> <BR> <BR> N-R- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}-3- [ (pyridine-3-carbonyl)-amino]- succinamic acid N-R-{1-[3-(6,7-Dichloro-benzo[b]thiophen-5-yl)-acryloyl]-pip eridin-4-yl}-3-[(pyridine-4-carbonyl)-amino]- succinamic acid M-R- {1- [3- (6, 7-Dich) oro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}-3-[(thiophene-2-car bonyl)-amino]- succinamic acid N-R-{1-[3-(6,7-Dichloro-benzo[b]thiophen-5-yl)-acryloyl]-pip eridin-4-yl}-3-[(furan-2-carbonyl)-amino]- succinamic acid

N-R- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}-3- [ (isoxazole-5-carbonyl)-amino]- succinamic acid <BR> <BR> <BR> <BR> <BR> <BR> <BR> <BR> 3-R- [ (Biphenyl-4-carbonyl)-amino]-N- {1- [3- (6, 7-dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}- succinamic acid N-R-{1-[3-(6,7-Dichloro-benzo[b]thiophen-5-yl)-acryloyl]-pip eridin-4-yl}-3-phenylacetylamino- succinamic acid 3R- [2- (4-Chloro-phenyl)-acetylamino]-N- {1- [3- (6, 7-dichloro-benzo [b] thiophen-5-yl)-acryloyl]- piperidin-4-yl}- succinamic acid <BR> <BR> <BR> <BR> <BR> <BR> <BR> <BR> N-R- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}-3- [2- (3-methoxy-phenyl)- acetylamino]-succinamic acid <BR> <BR> <BR> <BR> <BR> <BR> <BR> N-R- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}-3- [2- (4-methoxy-phenyl)<BR> <BR> <BR> <BR> -acetylamino]-succinamic acid<BR> <BR> <BR> <BR> <BR> <BR> <BR> <BR> <BR> <BR> <BR> N-R- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}-3-<BR> <BR> <BR> <BR> (2-thiophen-2-yl-acetylamino)-succinamic acid N-R-{1-[3-(6,7-Dichloro-benzo[b]thiophen-5-yl)-acryloyl]-pip eridin-4-yl}-3- [ (1-phenyl-cyclopropanecarbonyl)-amino]-succinamic acid NR-{1-[3-(6,7-Dichloro-benzo[b]thiophen-5-yl)-acryloyl]-pipe ridin-4-yl}- 3- (3-phenyl-propionylamino)-succinamic acid NR-{1-[3-(6,7-Dichloro-benzo[b]thiophen-5-yl)-acryloyl]-pipe ridin-4-yl}- 3- (2-phenoxy-acetylamino)-succinamic. acid <BR> <BR> <BR> 3R-- [2- (4-Chloro-phenoxy)-2-methyl-propionylamino]-N-{1- [3- (6, 7-dichloro-benzo [b] thiophen-5-yl)- acryloyl]-piperidin-4-yl}-succinamic acid 3R- { [1- (4-Chloro-phenyl)-cyclopentanecarbonyl]-amino}-N- {1- [3- (6, 7-dichloro-benzo [b] thiophen-5-yl)- acryloyl]-piperidin-4-yl}-succinamic acid <BR> <BR> <BR> <BR> <BR> <BR> <BR> <BR> NR-- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}-3-<BR> <BR> <BR> <BR> (3-phenyl-acryloylamino)-succinamic acid<BR> <BR> <BR> <BR> <BR> <BR> 3-R- (2-Benzyloxy-acetylamino)-N-{1- [3- (6, 7-dichloro-benzo [b] thiophen-5-yl)-<BR> <BR> <BR> <BR> acryloyl]-piperidin-4-yl}-succinamic acid NR- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl} -3-(3-methyl-ureido)-succinamic acid N-R- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}-3- (3, 3-dimethyl-ureido)-succinamic acid NR-{1-[3-(6,7-Dichloro-benzo[b]thiophen-5-yl)-acryloyl]-pipe ridin-4-yl}-3 -[(morpholine-4-carbonyl)-amino]-succinamic acid 3-R (3-Benzyl-ureido)-N- {1-[3-(6,7-dichloro-benzo [b] thiophen-5-yl)-acryloyl] -piperidin-4-yl}-succinamic acid N-R-{1-[3-(6,7-Dichloro-benzo[b]thiophen-5-yl)-acryloyl]-pip eridin-4-yl}- 3- (3-phenethyl-ureido)-succinamic acid

NR- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}- 3-methoxycarbonylamino-succinamic acid <BR> <BR> <BR> <BR> 3-R-Benzyloxycarbonylamino-N- {1- [3- (6, 7-dichloro-benzo [b] thiophen-5-yl)<BR> <BR> <BR> <BR> <BR> -acryloyl]-piperidin-4-yl}-succinamic acid<BR> <BR> <BR> <BR> <BR> <BR> <BR> N-R- {I- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yi)-acryloyl]-piperidin-4-yl)-3<BR> <BR> <BR> <BR> - (3-methoxy-benzoylamino)-succinamic acid<BR> <BR> <BR> <BR> <BR> <BR> <BR> <BR> 3-R- (3-Chloro-benzoylamino)-N-{1- [3- (6, 7-dichloro-benzo [b] thiophen-5-yl)- acryloyl]-piperidin-4-yl}-succinamic acid 3-R-Benzenesulfonylamino-N- {1- [3- (6, 7-dichloro-benzo [b] thiophen-5-yl)-acryloyl]- piperidin-4-yl}-succinamic acid and N- {1- [3- (6, 7-dichloro-benzo [b] thiophen-5-yl)-acryl- oyl]--piperidin-4-yl}-3-guinolin-3-yl-su. ccinamic acid.

In another aspect, this invention provides pharmaceutical compositions comprising a pharmaceu- tically acceptable carrier and a therapeutically effective amount of a compound defined herein..

In one of the method aspects, this inven- tion is directed to a method for assaying a biolog- ical sample from a mammalian patient suspected of having a, disease, condition or disorder mediated, at least in part, by VLA-1, which method comprises ob- taining a biological sample from said patient and assaying said sample for the presence of VLA-1.

In another aspect, this invention is directed to a method for inhibiting adhesion of mammalian cells to the extracellular matrix medi- ated, at least in part, by VLA-1, which method comprises contacting said cells with a compound or pharmaceutical composition of this invention.

In another one of its method aspects, this invention is directed to a method for treating a disease, condition or disorder whose progression is

regulated, at least in part, by VLA-1 expression or activity in a mammalian patient in need thereof comprising administering to said patient a therapeu- tically effective amount of a compound or composi- tion of this invention.

In a preferred embodiment, said disease, disorder, or condition is selected from the group consisting of asthma, trachoma, Alzheimer's disease, atherosclerosis, AIDS dementia, diabetes, inflamma- tory bowel disease, multiple sclerosis, rheumatoid arthritis, tissue transplantation, tumor metastasis, tumor migration, and/or tumor growth, proliferation of fibroblasts in cancer, solid tumors, meningitis, encephalitis, stroke, cerebral traumas, nephritis, retinitis, atopic dermatitis, psoriasis, myocardial ischemia, acute leukocyte-mediated lung injury, and fibrotic diseases.

In another preferred embodiment, said di- sease, disorder, or condition is a fibrotic disease.

In still another preferred embodiment, said fibrotic disease is selected from the group consisting of systemic sclerosis, mixed connective tissue disease, fibrodysplasia, fibrocystic disease, sarcoidosis, and myositis.

In yet another preferred embodiment, said fibrotic disease has a manifestation of fibrotic vascular intimal hypertrophy, and is selected from the group consisting of vasculitis, polyarteritis nodosa, and temporal arteritis.

In yet another preferred embodiment, said fibrotic disease has a manifestation of fibrotic

hypertrophy of skin and/or muscle tissue, and is selected from the group consisting of scleroderma, eosinophilic fasciitis, discoid lesions associated with lupus or discoid lupus, and surgical adhesions.

In yet another preferred embodiment, said fibrotic disease has a manifestation of fibrotic hypertrophy of nerve tissue, and is selected from the group consisting of cerebrosclerosis, annular sclerosis, diffuse sclerosis, and lobar sclerosis.

In yet another preferred embodiment, said fibrotic disease has a manifestation of fibrotic hypertrophy, or fibrosis'of lung tissue, and is se- lected from the group consisting of pulmonary fibro- sis, idiopathic pulmonary fibrosis, the fibrotic element of pneumoconiosis, pulmonary sarcoidosis, fibrosing alveolitis, the fibrotic or hypertrophic element of cystic fibrosis, chronic obstructive pulmonary disease, adult respiratory distress syn- drome, and emphysema.

In yet another preferred embodiment, said fibrotic disease has a. manifestation of fibrotic hypertrophy, or fibrosis of prostate, liver, the pleura, or pancreas, and is selected from the group consisting of benign prostatic hypertrophy (BPH), nonalcoholic steato hepatitis, and fibrosis of the liver.

In yet another preferred embodiment, said fibrotic disease has a manifestation of fibrotic hypertrophy, or fibrosis of the kidney, and is selected from the group consisting of chronic renal

failure, lupus nephritis, alports syndrome, glomerulonephritis, and diabetic nephritis.

In another preferred embodiment, said disease, disorder, or condition is cancer.

In a preferred embodiment, said cancer is a tumor or a neoplasm selected from the group con- sisting of carcinomas, adenocarcinomas, and sar- comas.

In another preferred embodiment, said can- cer is selected from the group consisting of growth of solid tumors/malignancies, myxoid and round cell carcinoma, locally advanced tumors, human soft tissue carcinoma, cancer. metastases, squamous cell carcinoma, esophageal squamous cell carcinoma, oral carcinoma,. cutaneous T cell lymphoma, Hodgkin's lymphoma, non-Hodgkin's lymphoma, cancer of the adrenal cortex, ACTH-producing tumors, nonsmall cell cancers, breast cancer, gastrointestinal cancers, urological cancers, malignancies of the female gen- ital tract, malignancies of the male genital tract, kidney cancer, brain cancer, bone cancers, skin can- cers, thyroid cancer, retinoblastoma, neuroblastoma, peritoneal effusion, malignant pleural effusion, mesothelioma, Wilms's tumors, gall bladder cancer, trophoblastic neoplasms, hemangiopericytoma, and Kapos's sarcoma.

In still yet another preferred embodiment, said cancer is a cell proliferative disorders and is selected from the group consisting of angiogenesis- mediated diseases, benign tumors, acoustic neuromas, neurofibromas, pyogenic granulomas, biliary tract

cancer, choriocarcinoma, esophageal cancer, gastric cancer, intraepithelial neoplasms, lung cancer, and neuroblastomas.

A compound or composition of this inven- tion may be administered to the mammal by a suitable route, such as orally, intravenously, parenterally, transdermally, topically, rectally, or intranasally.

Mammals include, for example, humans and other primates, pet or companion animals, such as dogs and cats, laboratory animals, such. as rats, mice and rabbits, and farm animals, such as horses, pigs, sheep, and cattle.

DETAILED DESCRIPTION OF THE INVENTION Definitions and Overview As discussed above, the present invention is directed to novel aminopiperidine amide deriva- tives.

It is to be understood that the terminal- ogy used herein is for the purpose of describing particular embodiments only and is not intended to limit the scope of the present invention. It must' be noted that as used herein and in the claims, the singular forms"a, ""and,"and"the"include plural referents unless the context clearly dictates other- wise. In this specification and in the claims which follow, reference will be made to a number of terms which shall be defined to have the following mean- ings:

As used herein,"alkyl"refers to mono- valent alkyl groups having from 1 to 10 carbon atoms and more preferably 1 to 6 carbon atoms. This term is exemplified by groups such as methyl, t-butyl, n- heptyl, octyl, and the like.

"Substituted alkyl"refers to an alkyl group having from 1 to 5 substituents selected from the group consisting of alkoxy, substituted alkoxy, acyl, acylamino, acyloxy, amino, substituted amino, aminoacyl, aminocarbonylamino, aminocarbonyloxy, aryl, substituted aryl, aryloxy, substituted aryl- oxy, cyano, halogen, hydroxyl, nitro, carboxyl, carboxyl esters, cycloalkyl, substituted cycloalkyl, thiol, thioalkyl,-heteroaryl, substituted hetero- aryl, heterocyclic, substituted heterocyclic, and oxycarbonylamino.

"Fluoroalkyl"refers to an alkyl group having from 1 to 4 carbon atoms and from 2 to 7 fluoro atoms.

"Hydroxy"refers to the group-OH.

"Alkylene"refers to divalent alkylene groups having from 1 to 10 carbon atoms, and more preferably 1 to 6 carbon atoms. This term is exemplified by groups such as methylene, n-heptyl- ene, 1,3-octylene, and the like.

"Substituted alkylene"refers to an alkyl- ene group having from 1 to 5 substituents selected from the group consisting of substituents defined for substituted alkyl.

"Alkoxy"refers to the group"alkyl-O-" which includes, by way of example, methoxy, ethoxy,

n-propoxy, iso-propoxy, n-butoxy, tert-butoxy, sec- butoxy, n-pentoxy, n-hexoxy, 1,2-dimethylbutoxy, and the like.

"Substituted alkoxy"refers to the group "substituted alkyl-0-." "Acyl"refers to the groups H-C (O)-, alkyl-C (O) -, substituted alkyl-C (O)-, alkenyl-C (O)-, substituted alkenyl-C(O)-, cycloalkyl-C (O)-, substi- tuted cycloalkyl-C (O)-, aryl-C (O) -, substituted aryl-C (O) -, heteroaryl-C (O) -, substituted hetero- aryl-C (O), heterocyclic-C (O) -, and substituted heterocyclic-C (O)-.

"Acylamino"refers to the group-C (O) NRR where each R is independently selected from the group consisting of hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, aryl, substi- tuted aryl, cycloalkyl, substituted cycloalkyl, het- eroaryl, substituted heteroaryl, heterocyclic, sub- stituted heterocyclic, and where each R is option- ally joined to form together with the nitrogen atom a heterocyclic or substituted heterocyclic ring.

"Acyloxy"refers to the groups alkyl- C (O) O-, substituted alkyl-C (0) 0-, alkenyl-C (O) O-, substituted alkenyl-C (O) O-, aryl-C (O) O-, substituted aryl-C (O)O-, cycloalkyl-C (O) 0-, substituted cyclo- alkyl-C (0) O-, heteroaryl-C (O) 0-, substituted hetero- aryl-C (0) 0-, heterocyclic-C (0) 0-, and substituted heterocyclic-C (0) 0-.

"Alkenyl"refers to monovalent alkenyl groups having from 2 to 10 carbon atoms, and more preferably 2 to 6 carbon atoms, and having at least

1 and preferably from 1-2 sites of alkenyl unsatura- tion.

"Substituted alkenyl"refers to alkenyl groups having from 1 to 5 substituents selected from the group consisting of alkoxy, substituted alkoxy, acyl, acylamino, acyloxy, amino, substituted amino, aminoacyl, aminocarbonylamino, aminocarbonyloxy, aryl, substituted aryl, aryloxy, substituted aryl- oxy, cyano, halogen, hydroxyl, nitro, carboxyl, car- boxyl esters, cycloalkyl, substituted cycloalkyl, thiol, thioalkyl, heteroaryl, substituted hetero- aryl, heterocyclic, substituted heterocyclic, and oxycarbonylamino, provided that the hydroxyl or the thio group is not pendent to an unsaturated carbon atom.

"Amino"refers to the group-NH2.

"Substituted amino"refers to the group -NRR where each R is independently selected from the group consisting of hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, aryl, substi- tuted aryl, cycloalkyl, substituted cycloalkyl, heteroaryl, substituted heteroaryl, heterocyclic, substituted heterocyclic and where each R is option- ally joined to form together with the nitrogen atom a heterocyclic or substituted heterocyclic ring pro- vided that both R's are not hydrogen.

"Aminosulfonyl"refers to the group -SO2NR'R', wherein each R'is independently selected from the group consisting of hydrogen, alkyl, sub- stituted alkyl, alkenyl, substituted alkenyl, aryl, substituted aryl, cycloalkyl, substituted cyclo-

alkyl, heteroaryl, substituted heteroaryl, hetero- cyclic, substituted heterocyclic and where each R is optionally joined to form together with the nitrogen atom a heterocyclic or substituted heterocyclic ring.

"Sulfonylamino"refers to the group -NR'R'SO2, wherein each R'is as defined above.

"Aminoacyl"refers to the groups-NRC (O)- alkyl,-NRC (O) substituted alkyl,-NRC (O) alkenyl, -NRC (O) substituted alkenyl,-NRC (O) cycloalkyl,-NRC- (O) substituted cycloalkyl,-NRC (O) aryl, -NRC (O) sub- <BR> <BR> <BR> <BR> stituted aryl,-NRC (O) heteroaryl, -NRC (O) substituted heteroaryl,-NRC (O) heterocyclic, and-NRC (O) substi- tuted heterocyclic, where R is hydrogen or alkyl.

"Aminocarbonyloxy"refers to the groups - NRC (O) 0-alkyl,-NRC (0) 0 substituted alkyl,-NRC (O)- O-Cycloalkyl,-NRC (0) 0 substituted cycloalkyl,-MRC- <BR> <BR> <BR> <BR> (O) O-aryl,-NRC (O) O substituted aryl,-NRC (O) O-het- eroaryl,-NRC (0) O substituted heteroaryl,-NRC (0) O- heterocyclic, and -NRC (0) O substituted heterocyclic, where R is hydrogen or alkyl.

"Oxycarbonylamino"refers to the groups -OC (O) Q where Q is amino or substituted amino.

"Aminocarbonylamino"or"aminoacylamino" refers to the groups-QC (O) Q where each Q is inde- pendently amino or substituted amino.

"Aryl"or"Ar"refers to an unsaturated aromatic carbocyclic group of from 6 to 14 carbon atoms having a single ring (e. g. , phenyl) or multi- ple condensed rings (e. g. , naphthyl or anthryl) which condensed rings may or may not be aromatic

(e. g. , 2-benzoxazolinone, 2H-1, 4-benzoxazin-3 (4H)- one-7-yl, and the like) provided that the point of attachment is on an aromatic carbocyclic group atom.

Preferred aryls include phenyl and naphthyl.

"Substituted aryl"refers to aryl groups which are substituted with from 1 to 3 substituents selected from the group consisting of hydroxy, acyl, acylamino, acyloxy, alkyl, substituted alkyl, alkoxy, substituted alkoxy, alkenyl, substituted alkenyl, amino, substituted amino, aminoacyl,-amino- carbonyloxy, aminocarbonylamino, aryl, substituted aryl, aryloxy, substituted aryloxy, carboxyl, car- boxyl esters, cyano, thiol, thioalkyl, substituted thioalkyl, cycloalkyl, substituted cycloalkyl, halo, nitro, heteroaryl, substituted heteroaryl, hetero- cyclic, and substituted heterocyclic.

"Aryloxy"refers to the group aryl-O- which includes, by way of example, phenoxy, naph- thoxy, and the like.

"Substituted aryloxy"refers to substi- tuted aryl-O-groups.

"Carboxyl"refers to the group-COOH and salts thereof.

"Carboxyl esters"refer to the group-COOR where R is selected from the group consisting of alkyl, substituted alkyl, alkenyl, substituted alkenyl, cycloalkyl, substituted cycloalkyl, aryl, substituted aryl, heteroaryl, substituted hetero- aryl, heterocyclic, substituted heterocyclic.

"Cycloalkyl"refers to monovalent cyclic alkyl groups of from 3 to 8 carbon atoms having a

single cyclic ring including, by way of example, cyclopropyl, cyclobutyl, cyclopentyl, cyclooctyl, and the like.

"Substituted cycloalkyl"refers to a cycloalkyl group, preferably of from 3 to 8 carbon atoms, having from 1 to 5 substituents selected from the same group of substituents as defined for sub- stituted alkyl as well as oxo (=O) and thioxo (=S) groups.

"Cycloalkylene"refers to divalent cyclic alkyl groups of from 3 to 8 carbon atoms having a single cyclic ring including, by way of example, cyclopropylene, cyclobutylene, cyclopentylene, cyclooctylene, and the like.

"Cycloalkenylene"refers to divalent cyclic alkenyl groups of from 4 to 8 carbon atoms having a single cyclic ring and 1-2 sites of unsat- uration including, by way of example, cyclobutenyl- ene, cyclopentenylene, cyclooctenylene, and the like.

"Halo"or"halogen"refers to fluoro, chloro, bromo and iodo and preferably is fluoro, chloro or bromo.

"Heteroaryl"refers to an aromatic group of from 1 to 10 carbon atoms and 1 to 4 heteroatoms selected from the group consisting of oxygen, nitro- gen and sulfur within the ring. Such heteroaryl groups can have a single ring (e. g. , pyridyl or furyl) or multiple condensed rings (e. g. , indolizin- yl or benzothienyl) provided that the point of attachment is to a heteroaryl group atom and further

provided that the heteroaryl'group contains at least five ring atoms. Preferred heteroaryls include pyridyl, pyrrolyl, indolyl and furyl.

"Substituted heteroaryl"refers to heter- oaryl groups which are substituted with from 1 to 3 substituents selected from the group of'substituents defined for, substituted aryl.

"Heterocycle"or"heterocyclic"refers to a monovalent saturated or unsaturated, but not aro- matic, group having a single ring or multiple con- densed rings, from 1 to 10 carbon atoms and from 1 to 4 hetero : atoms selected from the group consisting of nitrogen, sulfur or oxygen within the ring where- in, in fused ring systems, one or more the rings can be aryl or heteroaryl, provided that the hetero- cyclic ring has at least 4 atoms and further pro- vided that the point of attachment is to a hetero- cyclic ring atom.

"Substituted heterocyclic"refers to het- erocycle groups which are substituted with from 1 to 3 substituents selected from the group of substitu- ents defined for substituted cycloalkyl.

Examples of heterocycles and heteroaryls include, but are not limited to, azetidine, pyrrole, imidazole, pyrazole, pyridine, pyrazine, pyrimidine, pyridazine, indolizine, isoindole, indole, dihydro- indole, indazole, purine, quinolizine, isoquinoline, quinoline, phthalazine, naphthylpyridine, quinox- aline, quinazoline, cinnoline, pteridine, carbazole, carboline, phenanthridinei acridine, phenanthroline, isothiazole, phenazine, isoxazole, phenoxazine,

phenothiazine, imidazolidine, imidazoline, piperi- dine, piperazine, indoline, phthalimide, 1,2, 3,4- tetrahydroisoquinoline, 4,5, 6, 7-tetrahydrobenzo [b] - thiophene, thiazole, thiazolidine, thiophene, benzo- [b] thiophene, morpholinyl, thiomorpholinyl (also re- ferred to as thiamorpholinyl), piperidinyl, pyrrol- idine, tetrahydrofuranyl, and the like.

"Heterocyclene"refers to divalent hetero- cyclic groups of from 3 to 8 carbon atoms having a single cyclic ring.

"Heterocyclenylene"refers to divalent heterocyclic groups of from 4 to 8 carbon atoms having a single cyclic ring and 1-2 sites of unsat- uration.

"Thiol"refers to the qroup-SH.

"Thioalkyl"refers to the group-S-alkyl.

"Substituted thioalkyl"refers to the group-S substituted alkyl.

"Thioaryl"refers to the group-S-aryl.

"Thioalkylaryl"refers to the group-S- alkylene-aryl, S substituted alkylene aryl, S alkyl- ene substituted aryl or-S substituted alkylene sub- stituted aryl.

"Thioalkylheteroaryl"refers to the group - S alkylene heteroaryl, S substituted alkylene het- eroaryl, S alkylene substituted heteroaryl or-S substituted alkylene substituted heteroaryl.

"Pharmaceutically acceptable salt"refers to pharmaceutically acceptable salts of a compound of Formula I which salts are derived from a variety of organic and inorganic counter ions well known in

the art and include, by way of example only, sodium, potassium, calcium, magnesium, ammonium, tetraalkyl- ammonium, and the like; and when the molecule con- tains a basic functionality, salts of organic or inorganic acids, such as hydrochloride, hydrobro- mide, tartrate, mesylate, acetate, maleate, oxalate, and the like.

"Prodrugs"as used herein, are compounds which convert (e. g.., hydrolyze, metabolize) in vivo to a compound of the invention. The effectiveness of an. orally administered drug is dependent upon the drug's efficient transport across the mucosal epi- thelium and its stability. in entero-hepatic circula- tion. Drugs that are effective after parenteral administration but less effective orally, or whose plasma half-life is considered too short, may be chemically modified into a prodrug form. The pro- drug should have a pharmacokinetic profile that is different from that of the parent, enabling easier absorption across the mucosal epithelium, better salt formulation and/or solubility, and/or improved systemic stability (for an increase in plasma half- life, for example). Many chemical modifications may be suitable for the creation of the prodrugs accord- ing to the invention, including: (1) Ester or amide derivatives which may be cleaved by, for example, esterases or lipases.

For ester derivatives, the ester is derived from the carboxylic acid moiety of the drug molecule by known means. For amide derivatives, the amide may be de-

rived from the carboxylic acid moiety or the amine moiety of the drug molecule by known means.

(2) Peptides that may be recognized by specific or nonspecific proteinases. A peptide may be coupled to the-drug molecule via amide bond for- mation with the amine or carboxylic acid moiety of the drug molecule by known means.

(3) Derivatives that accumulate at a site of action through membrane selection of a prodrug form or modified prodrug form.

(4) Any combination of 1 to 3.

It will further be appreciated by those skilled in the art that certain moieties known to those skilled in the art as"pro-moieties,"for example as described in"Design of Prodrugs"by H.

Bundgaard (Elsevier) 1985, may be placed on ap- propriate functionalitiesvwhen such functionali. ties are present in compounds of the invention also to form a"prodrug."Further, certain compounds of the invention may act as prodrugs of other compounds of the invention. All protected derivatives, and pro- drugs, of the compounds of the invention are in- cluded within the scope of the invention.

"Biological activity"as used herein re- fers to an inhibition concentration when tested in at least one of the assays outlined in Example A or B..

The terms"substituted"as used with, for example, "substituted alkyl"does not include poly- mers derived therefrom but are limited to a maximum of 3 substituents groups, e. g. , Ar-Ar-Ar.

The term"tautomer"refers to an isomer in which migration of a hydrogen atom results in two or more structures.

Compound Preparation The compounds of this invention can be prepared from readily available starting materials using the following general methods and procedures.

It will be appreciated that where typical or pre- ferred process conditions (i. e. , reaction tempera- tures, times, mole ratios of reactants, solvents, pressures) are given, other process conditions can also be used unless otherwise stated. Optimum re- action conditions may vary with the particular re- actants or solvent used, but such conditions can be determined by one skilled in the art by routine optimization procedures.

Additionally, as will be apparent to those skilled in the art, conventional protecting groups may be necessary to prevent certain functional groups from undergoing undesired reactions. Suit- able protecting groups for various functional groups as well as suitable conditions for protecting and deprotecting particular functional groups are well known in the art. For example, numerous protecting groups are described in T. W. Greene and P. G. M.

Wuts,"Protecting Groups in Organic Synthesis, Second Edition, "Wiley, New York, 1991, and refer- ences cited therein.

Furthermore, the compounds of this inven- tion may contain one or more chiral centers.

Accordingly, if desired, such compounds can be pre- pared or isolated as pure stereoisomers, i. e. , as individual enantiomers or diastereomers, or as stereoisomer-enriched mixtures. All such stereoiso- mers (and enriched mixtures) are included within the scope of this invention, unless otherwise indicated.

Pure stereoisomers (or enriched mixtures) may be prepared using, for example, optically active start- ing materials or stereoselective reagents well known in the art. Alternatively, racemic mixtures of such compounds can be separated using, for example, chi- ral column chromatography, chiral resolving agents, and the like.

Compounds in the present invention may be better understood by the following synthetic Schemes that illustrate methods for the synthesis of com- pounds of the invention.

Scheme 1 0 R3 R2 R3 + R'-X ION yly (optional R2 0 hydrolysis) 1 2 3 each R2 = H, CHg, CF3 R1 and R3 are as defined for Formula I Pd (O) refers to metallic palladium Cinnamic acid intermediates 3 may be pre- pared by a Heck-type, palladium-mediated coupling (e. g. , using tetrakis- (o-tolyl phosphine) palladium-

(0), Pd2 (dba) 3, Pd (OAc) 2, or the like) of halo sub- stituted aromatic derivatives 2 with an appropriate olefinic substrate 1 (e. g. , methyl acrylate, ethyl crotonate, or the like ; S. J. Buchwald, Chem. Eur.

J. 1999, 5, 3107-3112). The intermediate ester may require a separate hydrolysis step to allow for fur- ther elaboration..

Scheme 2 "acetate equivalent" ---------. R'OH ON R' base, solvent (optional hydrolysis) 4 5

6 R3 = H or CH3 Rl is as defined for Formula I An alternative preparation of substituted cinnamic acids 6 is given in Scheme 2. Substituted aromatic aldehydes 5 may be prepared from aromatic carboxylic acids 4 by utilizing a reducing agent (for example, borane-THF, LiAlH4) to reduce the acid to the benzyl alcohol. The resulting alcohols can then undergo partial oxidation by standard methods

(Jones, Swern, Moffat) to the desired aldehydes.

Further elaboration is possible by reacting the aldehyde with an acetate equivalent (using malonic acid, Meldrum's acid, or the like) and a base (such as piperidine,. or the like) in a solvent (e. g., pyridine or 2,6-lutidine) to give the resulting cinnamic acids 6. In an alternate procedure (M.

Mikolajczyk, Synthesis 1976, 6, 396), the aldehyde may be reacted with a Wittig, Horner-Emmons, or Wadsworth-Emmons reagent (e. g., 2- (diethoxyphos- phoryl) -propionic acid ethyl ester, or the like) in a solvent (CH2Cl2, CH2ClCH2C. 1, or like) and a base (NaOH, NaOEt, NaH, or the like) to furnish the de- sired cinnamic ester. The ester may undergo hydrol- ysis to the acid 6 with. the use of a base (LiCS, NaOH, or the like) in a solvent (THF-water, CH2Cl2- water).

Scheme 3 R3 H R4 N-P1 R'OH + coupling m 6 7 H R4 N-Pi R3 R NHZ deprotection R3 Rl*N I go. R N m RlZ1/N I m R2 0

8 9 wherein R1, R2, R3 are as defined for Formula I m is as defined above Cinnamides of amino substituted cyclic amines may be constructed by coupling of a cinnamic acid 6 with an appropriately protected amino substi- tuted cyclic amine 7 (e. g., P1=Boc, Fmoc, or the like ; m=0-4), as shown in Scheme 3. The cinnamic acid is activated (for example, using thionyl chlor- ide, or oxalyl chloride, or 1- (3-dimethylamino- propyl)-3-ethylcarbodiimide and N-hydrox'ysuccin- imide, or the like) and reacted with 7, usually in the presence of a tertiary amine base (e. g. , diiso- propylethylamine, triethylamine, N-methyl morph- oline, or the like) to provide amides 8. The pro- tected amino group of 8 is subsequently deprotected to the amine 9 using appropriate reagents (e. g., anhydrous HCl to remove a Boc group).

Scheme 4 R\. NH R7 O R3 R6 R4 NH > (optional base) + Im R'l n n R2 0 0 9 10

when n=1, then R"=R9 and/or R10 when n=2, then first R"=R9 and/or RIO and second CHR"-COOH=R8 Rl, R2, R3, R6, R7, R9, RIO are as defined for Formula I m is as defined above Scheme 4 illustrates the elaboration of amino cinnamides 9 with various cyclic anhydrides 10 (for example, substituted succinic and glutaric an- hydrides, or the like) to give succinamides (11, n=1) and glutaramides (11, n=2). The use of a tertiary amine base (e. g., diisopropylethylamine, or the like) is optional. In cases where R6 or R7#R", a regioisomeric mixture of amides 11 may be obtained, which can be isolated by conventional techniques as described below.

Scheme 5 RHz p6 p7 0 4 R3 coupling R N n 0 Razz I m 0 R" R O 9 12 R6 R7 0 Ru H R3 N n OiP2 deprotection m R2 o R 0

Rl, R2, R3, R6, R7 are as defined for Formula I R"is as defined above m is as defined above P2 is a suitable protecting, group such as t-butyl and the'like Alternatively, amines 9 may be coupled with a mono-protected diacid 12 (for example, mono- ethyl malonate, mono-ethyl succinate, or the like) to give an amide-ester intermediate 13, as shown in Scheme 5. Amide formation protocols are as de- scribed for Scheme 3. Subsequent deprotection of the ester group of 13, using appropriate reagents (for example, TFA to remove tert-butyl ester groups) yields malonamides (11, n=0), succinamides (11, n=1), or glutaramides (11, n=2).

Scheme 6 R4 NH2 R6 R7 0 HO Op2 coupling pN +'I'n 0 RR" 14 12 R6'R7 0 N) -P2 deprotection O R" X m 15 R3 R3 OH N'in | 6 R7 ° R2 O n 0 1) coupling HO O R 11 i 2) deprotection m 16

P1, P2, n, R", m are as defined above R6, R7, R1, R2, R3 are as defined above In another variation for preparing the compounds of the invention, shown in Scheme 6, the mono-protected diacid 12 is first coupled with an appropriately protected amino substituted cyclic amine 14 (P1=Boc group, or the like; m=0-4) using amide formation protocols described in Scheme 3.

After removal of the protecting group (e. g., an- hydrous HC1 for Boc removal, or the like), amine 16 can be coupled with a cinnamic acid 6 as described for Scheme 3 followed by deprotection of P2 using appropriate reagents (for example, TFA to remove tert-butyl ester groups) to provide amides 11.

Scheme 7 R4 H R3 R4 H X N-P1 4 Cl base + HN m R2 0 solvent 7 17

8 P1, m, Pd (O) are as defined above R1, R2, R3 are as defined for Formula I Scheme 7 illustrates an alternative pro- cedure in which the order of these coupling steps may be reversed. Cyclic amines bearing a protected amine group (7, e. g. , 4-N-Boc-aminopiperidine, or the like) may be coupled with an acryloyl chloride 17 in the presence of an appropriate base (e. g., DIEA, or the like). The acrylamide olefin 18 may be

further elaborated by palladium-mediated coupling with a halo substituted aromatic derivative (using protocols described for Scheme 1) to provide cinnamides 8.

Scheme 8 R R H N OPZ deprotect HN O O I Im 24 R6 ruz H R4 N OP2 6 It3 coupling 0-Rl,), r 0 hydrolysis ; N m 0 0. R2 25

26 each P2 is a suitable carboxy protecting group m is as defined above Ri, R2, R3, R6, R7 are as defined for Formula I Some of the analogues described herein contain a malonamide moiety, which can be prepared as described in Scheme 5, or by direct displacement of the ester group of a dialkyl malonate (e. g. , di- ethyl malonate, or the like) with amines 14 (e. g., 21=benzyl, or the like) leading to malonamides 23.

After deprotection (for example, using H'2 and Pd/C to remove an N-benzyl group, or the like), the result- ing amines 24 can be coupled with a cinnamic acid 6 using amide formation protocols described for Scheme 3, thus providing malonamides 25. Deprotection of the ester group Ra gives desired acids 26.

Scheme 9

30. 31.

R''=substitution on aryl group-as defined above Scheme 9 illustrates the preparation of 5- bromobenzothiophenes, used for the preparation of the corresponding cinnamic acids. A suitable phen- ylsulfanyl acetic acid 28 is prepared from a thio- phenol 27 using bromoacetic acid under standard con- ditions. Sulfide 28 is selectively brominated, e. g. , with bromine in a solvent such as dichloro- methane, glacial acetic acid, or the like, in the presence of iron or iodine as catalysts. The reac- tion is typically performed at room temperature for 1 to 96 hours. Bromide 29 is then converted to an acid halide intermediate by treatment with an in-

organic acid halide (for example, thionyl chloride, phosphorous trichloride, phosphorous pentachloride, phosphrous tribromide, oxalyl chloride, or the like) in an inert solvent (for example dichloromethane or the like), at a temperature in the range of 0°C to 110°C for about 1 to 48 hours. The volatiles are then removed under reduced pressure and the residue is dissolved in an appropriate solvent, typically dichloromethane, and subjected to Friedel-Crafts cyclization by treatment with a Lewis acid such as AlCl3 or polyphosphoric acid. The reaction is gen- erally carried out at-78°C to 25°C. The resulting intermediate ketone product is reduced without isolation by a hydride reducing reagent (for exam- ple, NaBH4 or the like) to-provide alcohol 30.

Alcohol 30 may be used directly in the next step, or first purified by chromatography or recrystalliza- tion as appropriate. The dehydration of 30 to benzothiophene 31'is accomplished by treatment with a Lewis or proti. c acid (for example, boron trifluor- ide etherate.) in an appropriate solvent, (for example glacial acetic acid or the like) at a temperature ranging from ambient temperature to the reflux tem- perature of the solvent. Upon completion of the reaction, the resulting 5-bromo benzothiophene 31 is recovered by conventional means such as neutraliza- tion, precipitation, filtration, recrystallization, and the like.

Scheme 10 coupling separate isomers R6 R7 0/ N n deprotect R2 I I 32 R 0 - 32

11, isomers separates m, n, R"are. as defined above R1, R2, R3, R6, R7 are as defined for Formula I Some of the diamides 11 prepared according to Scheme 4 as regioisomeric mixtures (R6 or R7#R"), were separated chromatographically as 9-fluorenyl- methyl esters 32 as shown in Scheme 10. These fluorenyl esters 32 were synthesized by coupling the carboxylic acid group of the mixture of regioiso- meric compounds 11 with 9-fluorenylmethanol using an appropriate activating reagent (e. g., dicyclohexyl- carbodiimide, or the like). After chromatographic separation, the 9-fluorenylmethyl group was removed without regioisomeric scrambling of R6 or R7 and R" using piperidine, or the like, to generate pure regioisomeric compounds 11.

Scheme 11 0 Brt H O alk P2 34 ON 0 base O base 33 Rx"-, 0 0 O H hydrolysis O N alk P2 o 35 36 'NH ? 2 NHP2 0 ex i Rq N O R3 \--V 9 razz 9 i 1' 0 coupling R'N deprotect razz R O 37

RIO'is a suitable substituent m is as defined above R1, R2, R3 are as defined for Formula I

Scheme 11 illustrates the preparation of succinamides in which an aminomethyl group is ap- pended to the ß-position of the succinamide chain.

Al, kylation of an ester of-glycine (33, P2=Boc group, or the like; alk=methyl, ethyl, or the like) with an a-bromo acetate ester 34 (Rx=tert-butyl, or another group with orthogonal deprotection condi- tions to alk) gives an aminomethyl succinate diester 35. The diester 35 can be selectively mono-depro- tected to give carboxylic acid intermediate 36 (for example, using lithium hydroxide to hydrolyze a methyl ester selectively over a ter-butyl ester).

The monocarboxylic acid 36 is coupled to amines 9 using amide formation protocols described for Scheme 3. Simultaneous deprotection of P2 and R'of amides 37 can be achieved using appropriate reagents (for example, TFA in cases where P2=Boc and RX=tert-butyl, or the like) to provide amino acids 38. The amino group of 37 can be revealed by selective deprotec- tion, and then functionalized by reaction with acti- vated carboxyl-and sulfonyl-containing inputs (e. g. , acid chlorides, sulfonyl chlorides, carbamoyl chlorides, isocyanates, anhydrides, chloroformates,

or the like) in the presence of an appropriate base (e. g. , diisopropylethylamine, or the like), to pro- vide, after ester cleavage, the amine-derivatized compounds 39. Alternatively, the amino group of 38 may be functionalized selectively in the presence of the, carboxylic acid using the activated carboxyl- and sulfonyl-containing inputs described above.

The preparation of the (3-heteroaryl group (Ra is heteroaryl, substituted heteroaryl in Formula I) can be accomplished by well known techniques, an example of. which is provided in Scheme 12 below: Scheme 12 R6 and R7 are as defined for Formula I Het is a suitable heteroaryl or heterocyclic

600

Specifically, in Scheme 12 above, the car- boxyl group of an nitrogen-protected a-amino acid, compound 100, is reduced using conventional tech- niques such as the use of a reducing agent includ- ing, for example, lithium aluminum hydride to pro- vide for the corresponding alcohol, compound 200.

The reaction is preferably conducted in an inert diluent such as tetrahydrofuran, diethyl ether, and the like at a temperature preferably from about -78°C to about 25°C. The reaction is continued until substantial completion which typically occurs from within 0.5 to 18 hours. Upon completion of the reaction, compound 200 can be recovered by conven- tional methods including neutralization, extraction, precipitation, chromatography, filtration, and the like or used in the next step of the reaction with- out purification and/or isolation.

The alcohol, compound 200, is converted to a halo group (e. g. , chloro) again by conventional methods such as contact with a suitable halogenating agent to provide for compound 300. Suitable halo- genating agents include, for example, inorganic acid halides, such as thionyl chloride, phosphorous tri- chloride, phosphorous tribromide or phosphorous pentachloride, under conventional conditions. Gen- erally, this reaction is conducted using about 1 to 5 molar equivalents of the inorganic acid halide, either neat or in an inert solvent, such as dichlor- omethane or carbon tetrachloride, at temperature in the range of about 0°C to about 80°C for about 1 to about 48 hours. A catalyst, such as DMF, may also

be used in this reaction. Upon completion of the reaction, compound 300 can be recovered by conven- tional methods including. neutralization, extraction, precipitation, chromatography, filtration, and the like or used in the next step of the reaction with- out purification and/or isolation.

The conversion-of compound 300 into a Grignard reagent, compound 400,. follows conventional techniques well documented in the literature includ- ing Gutsche et al.,"Fundamentals of Organic Chem- istry, "Prentice Hall (1975), pp. 238 et seq. Spe- cifically, at least an equimolar amount of metallic magnesium is added to compound 3. in a suitable inert diluent such as diethyl ether to form the Grignard reagent, compound 400. This, compound, typically in a single reaction vessel,-is reacted with at least an equimolar equivalent of a heteroaryl or hetero- cyclic halide, i. e., Het-X (compound 500) to provide for compound 600.

Pharmaceutical Formulations When employed as pharmaceuticals, the com- pounds of the subject invention are usually adminis- tered in the form of pharmaceutical compositions.

These compounds can be administered by a variety of routes including oral, parenteral, transdermal, top- ical, rectal, and intranasal. These compounds are effective as both injectable and oral compositions.

Such compositions are prepared in a manner well known in the pharmaceutical art and comprise at least one active compound.

This invention also includes pharmaceu- tical compositions which contain, as the active ingredient, one or more of the compounds of the subject invention above associated with pharmaceu- tically acceptable carriers. In making the composi- tions of this invention, the active ingredient is usually mixed with an excipient, . diluted by an exci. pient or enclosed within such a carrier which can be in the form of a capsule, sachet, paper or other container.. The. excipient. employed is typical- ly an excipient suitable for administration to human subjects or other mammals. When, the excipient serves as a diluent, it can be a solid., semi-solid, or liquid material, which acts as a vehicle, carrier or medium for the active ingredient. Thus, the com- positions can be in the form of tablets, pills, powders, lozenges, sachets, cachets, elixirs, sus- pensions, emulsions, solutions, syrups, aerosols (as. a solid or in a liquid medium), ointments contain- ing, for example, up to 10% by weight of the active compound, soft and hard gelatin capsules, supposi- tories, sterile injectable solutions, and sterile packaged powders.

In preparing a formulation, it may be necessary to mill the active compound to provide the appropriate particle size prior to combining with the other ingredients. If the active compound is substantially insoluble, it ordinarily is milled to a particle size of less than 200 mesh. If the active compound is substantially water soluble, the particle size is normally adjusted by milling to

provide a substantially uniform distribution in the formulation, e. g. , about 40 mesh.

Some examples of suitable excipients in- clude lactose, dextrose, sucrose ; sorbitol, manni- tol, starches, gum acacia, calcium phosphate, al- ginates, tragacanth, gelatin, calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, sterile water, syrup, and methyl cellu- lose. The formulations can additionally include: lubricating agents such as talc, magnesium stearate, and mineral oil ; wetting-agents ;. emulsifying and suspending agents; preserving agents such as methyl and propylhydroxy-benzoates ; sweetening agents ; and flavoring agents. The compositions of. the invention can. be formulated so as to provide quick, sustained or delayed release of the active ingredient after administration to the patient by employing proce- dures known in the art.

The quantity of active component, that is the compound according to the subject invention, in the pharmaceutical composition and unit dosage form thereof may be varied or adjusted widely depending upon the particular application, the potency of the particular compound and the desired concentration.

The compositions are preferably formulated in a unit dosage form, each dosage containing from about 1 to about 500 mg, usually about 5 to about 100 mg, occasionally about 10 to about 30 mg, of the active ingredient. The term"unit dosage forms"re- fers to physically discrete units suitable as uni- tary dosages for human subjects and other mammals,

each unit containing a predetermined quantity of active material calculated to produce the desired therapeutic effect, in association with a suitable pharmaceutical excipient. Preferably, the compound of the subject-invention above is employed at no more than about 20 weight percent of the pharmaceu- tical composition, more preferably no more than about 15 weight percent, with the balance being pharmaceutically inert carrier (s).

The active compound is effective over a wide dosage range (and is generally administered. in a pharmaceutically or therapeutically effective-. amount. It. will be understood, however, that the amount of the compound actuallyadministered will be. determined by a physician, in the light of the rel- : evant circumstances, including the condition to be treated, the severity of the condition being treated, th. e chosen route of administration, the actual compound administered, the age, weight, : and response of the individual patient, the severity of the patient's symptoms, and the like.

In therapeutic use for treating, or com- bating, inflammation in warm-blooded animals, the compounds or pharmaceutical compositions thereof will be administered by any appropriate route, such as orally, topically, transdermally, and/or par- enterally at a dosage to. obtain and maintain a con- centration, that is, an amount, or blood-level of active component in the animal undergoing treatment that will be therapeutically effective. Generally, such therapeutically effective amount of dosage of

active component (i. e. , an effective dosage) will be in the range of about 0.1 to about 100, more prefer- ably about 1 to about 50 mg/kg of body weight./day..

For preparing solid compositions such as tablets, the principal active ingredient is mixed with a pharmaceutical excipient to form a solid pre-. formulation composition containing a homogeneous mixture of a compound of the present invention.

When referring to these. preformulation compositions as homogeneous, it is meant that. the active ingredi- ent is dispersed evenly throughout the composition so that the. composition may be readily subdivided into equally effective unit dosage forms such as tablets, pills and capsules. This solid preformula- tion is then subdivided into unit dosage forms of t. type described above-containing from, for. exam- ple, 0.1 to about 500. mg of the active. ingredient of the present invention.

The tablets or pills of the present inven- tion may be coated or. otherwise compounded to pro- vide a dosage form affording the advantage of pro- longed action. For example, the tablet or pill can comprise an. inner dosage and an outer dosage com- ponent, the latter being in the form of an envelope over the former. The two components can be sep- arated by an enteric layer which serves to resist disintegration in the stomach and permit the inner component to pass intact into the duodenum or to be delayed in release. A variety of materials can be used for such enteric layers or coatings, such materials including a number of polymeric acids and

mixtures of polymeric acids with such materials as shellac, cetyl alcohol, and cellulose acetate.

The liquid forms in which the novel com- positions of the present invention may be incorpo- rated for administration orally or by injection include aqueous solutions, suitably flavored syrups, aqueous or oil suspensions, and flavored emulsions with edible oils such as corn oil, cottonseed oil, sesame oil, coconut oil, or peanut oil, as well as elixirs and similar pharmaceutical vehicles.

Compositions for inhalation or insuffla- tion include solutions and suspensions in pharmaceu- tically acceptable, aqueous or organic solvents, or mixtures thereof, and powders. The liquid or solid compositions may contain suitable pharmaceutically acceptable excipients as. described supra. Prefer- ably the compositions are administered by the oral. or nasal respiratory route for local or systemic effect. Compositions in preferably pharmaceutically acceptable solvents may be nebulized by use of inert gases. Nebulized solutions may be inhaled directly from the nebulizing device or the nebulizing device may be attached to a face. mask tent, or intermittent positive pressure breathing machine. Solution, sus- pension, or powder compositions may be administered, preferably orally or nasally, from devices which deliver the formulation in an appropriate manner.

The following formulation examples illus- trate representative pharmaceutical compositions of the present invention.

Formulation Example 1 Hard gelatin capsules containing the following ingredients are prepared:

Ingredient Quantity (mg/capsule) Active ingredient 30.0 Starch 305.0 Magnesium stearate 5.0 The above ingredients are mixed and filled into hard gelatin capsules in 340 mg quantities.

Formulation Example 2 A tablet formula is prepared using the in-. gredients below: Ingredient Quantity (mg/tablet) Active ingredient 25.0 Cellulose, microcrystalline 200. 0' Colloidal silicon dioxide 10.0 Stearic acid 5. 0 The components are blended and compressed to form tablets, each weighing 240 mg.

Formulation Example 3 A dry powder inhaler formulation is pre- pared containing the following components: Ingredient Weight % Active ingredient 5 Lactose 95

The active ingredient is mixed with the lactose and the mixture is added to a dry powder inhaling appliance.

Formulation Example 4 Tablets, each containing 30 mg of active ingredient, are prepared as follows Ingredient Quantity (mg/tablet) Active ingredient 30.0 mg Starch 45. 0 mg Microcrystalline cellulose 35.0 mg Polyvinylpyrrolidone (as 10% solution in sterile water) _ Sodium carboxymethyl starch 4.5 mg Magnesium stearate 0.5 mg Talc 1.0 mg Total 120 mg , The active ingredient, starch and cellu- lose are passed through No. 20 mesh U. S. sieve and mixed thoroughly. The solution'of polyvinylpyrroli- done is mixed with the resultant powders, which are then passed through a 16 mesh U. S. sieve. The granules so produced are dried at 50°C to 60°C and passed through a 16 mesh U. S. sieve. The sodium carboxymethyl starch, magnesium stearate, and talc, previously passed through a No. 30 mesh U. S. si. eve, are then added to the granules which, after mixing, are compressed on a tablet machine to yield tablets each weighing 120 mg.

Formulation Example 5 Capsules, each containing 40 mg of medic- ament are made as follows:

Ingredient Quantity (mg/capsule) Active ingredient 40.0 mg Starch 109.9 mg Magnesium stearate 1.0 mg Total 150.0 mg The active ingredient,-starch and ma. gnesi- um stearate are blended, passed through a No. 20 mesh U. S. sieve, and filled into hard gelatin cap- sules in 15 mg quantities. zu Formulation Example 6 Suppositories, each containing 25 mg of active ingredient are made as follows: Ingredient Amount Active ingredient 25 mg Saturated fatty acid glycerides to 2., 000 mg s The active ingredient is passed through a No. 60 mesh U. S. sieve and suspended in the satur- ated fatty acid glycerides previously melted using the minimum heat necessary. The mixture is then poured into a suppository mold of nominal 2.0 g capacity and allowed to cool.

Formulation Example 7 Suspensions, each containing 50 mg of medicament per 5.0 mL dose are made as follows: Ingredient Amount Active ingredient 50.0 mg Xanthah gum 4. 0 mg Sodium, carboxymethyl cellulose (11%) Microcrystalline cellulose (89%) 50.0 mg Sucrose 1. 75 g Sodium benzoate 10.0 mg Flavor and color q. v. Purified water to 5.0 mL The active ingredient, sucrose and xanthanl. » gum are blended, passed through a No. 10 mesh U. S. sieve, and then mixed with a previously made solu- tion of the microcrystalline cellulose'and sodium carboxymethyl cellulose in water.. The sodium ben- zoate, flavor, and, color are diluted with some of the water and added with stirring. Sufficient watery <BR> <BR> <BR> <BR> is then added to produce the required volume.<BR> <P> .

Formulation Example 8

Ingredient Quantity (mg/capsule) Active ingredient 15.0 mg Starch 407.0 mg Magnesium stearate 3.0 mg Total 425. 0 mg

The active ingredient, starch, and mag- nesium stearate are blended, passed through a No. 20 mesh U. S. sieve, and filled into hard gelatin cap- sules in 425.0 mg quantities.

Formulation Example 9 A subcutaneous formulation may be prepared as follows:

Ingredient Quantity Active ingredient 5.0 mg Corn oil 1. 0 mL Formulation Example 10 A topical formulation may be prepared as follows : Ingredient Quantity Active ingredient 1-10 g Emulsifying wax 30 g Liquid paraffin 20 g. White soft paraffin to 100 g The white. soft paraffin is heated until molten. The liquid paraffin and emulsifying wax are incorporated and stirred until dissolved. The active ingredient is added and stirring is continued until dispersed. The mixture is then cooled until solid.

Formulation Example 11 An intravenous formulation may be prepared as follows: Ingredient Quantity Active ingredient 250 mg Isotonic saline 1000 mL

Another preferred formulation employed in the methods of the present invention employs trans- dermal delivery devices ("patches"). Such trans- dermal patches may be used to provide continuous or discontinuous infusion of the compounds of the pres- ent invention in controlled amounts. The construc- tion and use of transdermal patches for. the delivery of pharmaceutical agents is well known in the art.

See, e. g. , U. S. Patent 5,023, 252, herein incorpo- rated by reference. Such patches may be constructed for continuous, pulsatile, or on demand of pharmaceutical agents.

Frequently, it will be desirable or necessary to introduce the pharmaceutical composi- tion to the brain, either directly or indirectly.

Direct techniques usually involve placement of a drug delivery catheter into the host's ventricular system to bypass the blood brain barrier. One such implantable delivery system used for the transport of biological factors to specific anatomical regions of the body is described in U. S. Patent 5,011, 472 which is herein incorporated by reference.

Indirect techniques, which are generally preferred, involve formulating the composi- tions to provide for drug latentiation by the con- version of hydrophilic drugs into lipid soluble drugs. Latentiation is generally achieved through blocking of the hydroxy, carbonyl, sulfate, and primary amine groups present on the drug to render the drug more lipid soluble and amenable to trans- portation across the blood brain barrier. Alterna-

tively, the delivery of hydrophilic drugs may be enhanced by intra arterial infusion of hypertonic solutions which can transiently open the blood brain barrier.

Other suitable formulations for use in the present invention can be found in Remington's Phar- maceutical Sciences, Mack. Publishing Company, Phila- delphia, PA, 17th ed. (1985).

As noted above, the compounds described herein are suitable for use in a variety of drug delivery systems described above. Additionally, in order to enhance the in vivo serum half life of the administered compound, the compounds may be encap- sulated, introduced into the lumen of liposomes, prepared as a colloid, or other conventional tech- niques may be employed which provide an., extended serum half life of the compounds. A variety of methods are available for'preparing liposomes, as described in, e. g., Szoka et al. , U. S. Patent Nos.

4,235, 871, 4, 501,728 and 4, 837,028, each of which is incorporated herein by reference : Utility The compounds and/or compositions of this invention can be employed to bind VLA-1 in biolog- ical samples, for instance in mammalian patients suspected of having a disease, condition or disorder mediated, at least in part, by VLA-1. Accordingly, these compounds have utility in, for example, assay- ing such samples for VLA-1 mediated adhesion.

In addition, it is contemplated that com- pounds of this invention and/or pharmaceutical com- positions thereof inhibit, in vivo, adhesion of mammalian cells to the extracellular matrix medi- ated, at least in part, by VLA-1 and, accordingly, can be used in the treatment, prevention, or ameli- oration of diseases, conditions, or disorders whose progression or symptoms is regulated, at least in part, by VLA-1 expression or activity. Such di- seases, conditions, or disorders include, but. are not limited to, inflammatory diseases, fibrotic diseases, and cancer.

For example, diseases,-conditions,, and disorders which are expected to be treatable by. the compounds and/or compositions of the present inven- tion'include, but are not limited to, asthma,- trachoma, Alzheimer's disease, atherosclerosis, AIDS dementia, diabetes (including acute juvenile onset diabetes}, inflammatory bowel disease (including ulcerative colitis-and Crohn's disease) ;, multiple sclerosis, rheumatoid arthritis, tissue transplanta- ti. on, tumor metastasis, migration, and/or growth (including angiogenesis), proliferation of fibro- blasts in cancer, solid tumors, meningitis, enceph- alitis, stroke, and other cerebral traumas, neph- ritis, retinitis, atopic dermatitis, psoriasis, myocardial ischemia, acute leukocyte-mediated lung injury such as that which occurs in adult respira- tory distress syndrome, and fibrotic diseases, such as fibrotic diseases of the lung, kidney, liver and vasculature (including idiopathic pulmonary fibro-

sis, systemic sclerosis, glomerulonephritis, chronic hepatitis, chronic renal failure, and nonalcoholic steatohepatitis).

Fibrotic diseases which are expected to be treatable by the compounds and/or compositions of the present invention include systemic sclerosis, mixed connective tissue disease, fibrodysplasia, fibrocystic disease, sarcoidosis, myositis (e. g. , polymyositis, primary idiopathic polymyositis, childhood polymyositis, dermatomyositis, childhood dermatomyositis, primary idiopathic dermatomyositis in adults, inclusion body myositis, polymyositis, or dermatomyositis associated with malignant tumors).

Dermatomyositis can be associated with fibrosing or hypertrophic aspects, including fibrosing alveolitis and pulmonary fibrosis. Treatment using the com- pounds and/or compositions of the present invention is expected to treat, prevent, reduce, or ameliorate such diseases, or hypertrophy, fibrotic hypertrophy, or fibrosis in. such diseases. Amelioration includes reducing the rate of progression of a disease.

Among these fibrotic diseases. are diseases that have as a manifestation fibrotic vascular intimal hypertrophy. These diseases include va, scu- litis (including coronary artery vasculitis), poly- arteritis nodosa or temporal arteritis. Treatment using the compounds and/or compositions of the present invention is expected to treat, prevent, reduce, or ameliorate vascular intimal hypertrophy in such diseases.

These fibrotic diseases further include diseases that have as a manifestation fibrotic hypertrophy of skin and/or muscle tissue. These diseases include scleroderma, eosinophilic fasciit- is, discoid lesions associated with lupus or discoid lupus or surgical adhesions. Treatment using the compounds an. d/or composi. tions of the present inven- tion is expected to treat, prevent, reduce, or ameliorate such indications, or hypertrophy or fibrosis of skin or muscle tissue.

Fibrotic diseases further include diseases that have as a-manifestation fibrotic hypertrophy of nerve tissue. These diseases include cerebroscler- osis, annular sclerosis.. diffuse sclerosis and lobar sclerosis. Treatment using the compounds and/or compositions of the present invention is ex- pected to treat, prevent, reduce, or ameliorate such diseases, or hypertrophy,. fibrotic hypertrophy, or fibrosis of nerve tissue in such diseases.

These fibrotic diseases further include fibrotic lung diseases that have as a manifestation fibrotic hypertrophy, or fibrosis of lung tissue.

These diseases include pulmonary fibrosis (or in- terstitial lung disease or interstitial pulmonary fibrosis), idiopathic pulmonary fibrosis, the fi- brotic element of pneumoconiosis (which is associ- ated with exposure to environmental hazards such as smoking, asbestos, cotton lint, stone dust, mine dust and other particles), pulmonary sarcoidosis, fibrosing alveolitis, the fibrotic or hypertrophic element of cystic fibrosis, chronic obstructive

pulmonary disease, adult respiratory distress syn- drome and emphysema. Treatment using the compounds and/or compositions of the present invention is ex- pected to treat, prevent,. reduce, or ameliorate such diseases, or hypertrophy, fibrotic hypertrophy, or fibrosis in such diseas. es.

Such fibrotic diseases. further include diseases that have as a manifestation fibrotic hypertrophy, or fibrosis of prostate, liver, the pleura (e. g. , pleurisy, pleural.. fibrosis) or pan- creas. These diseases include benign prostatic hypertrophy (BPH) ;. nonalcoholic steato hepatitis and, fibrosis of the liver. Treatment using the com- pounds and/or compositions of the present invention is expected to treat, prevent, reduce, or ameliorate such diseases, or hypertrophy, ibrotic hypertrophy, or fibrosis. in such diseases.

These fibrotic diseases further include diseases. that have. as a manifestation fibrotic hypertrophy, or fib. rosis of the kidney, such as chronic renal failure,. lupus nephritis, alports syndrome, glomerulonephritis and diabetic nephritis.

Treatment using the compound and/or compositions of the present invention is expected to treat, prevent,. reduce, or ameliorate such diseases, or hypertrophy, fibrotic hypertrophy, or fibrosis of the kidney.

Cancers which are expected to be treatable by the compounds and/or compositions of the present invention. typically occur in mammals. Mammals in- clude, for example, humans and other primates, pet or companion animals, such as dogs and cats, labor-

atory animals, such as rats, mice and rabbits, and farm animals, such as horses, pigs, sheep, and cattle.

Tumors or neoplasms include growths of tissue cells in which-the multiplication of the cells is uncontrolled and progressive. Some such growths are benign, but others are termed"mali. g- nant"and can lead to death of the organism.

Malignant neoplasms or"cancers"are distinguished from benign growths in that, in addition to exhib- iting aggressive cellular proliferation, they can invade surrounding tissues and metastasize. More- over, malignant neoplasms-are characterized in that they show a greater loss of differentiation (greater "dedifferentiation") and'organization relative to one another and to surrounding. tissues.. This property is called"anaplasia.".

Tumors or neoplasms which are expected to be treatable by the compounds and/or compositions of the present invention include, but are not limited to., solid tumors, i. e. , carcinomas, adenocarcinomas,' and sarcomas. Carcinomas include those malignant neoplasms derived from epithelial cells which infil- trate (invade) the surrounding tissues give rise to metastases. Adenocarcinomas are carcinomas de- rived from granular tissue, or from tissues which form recognizable glandular structures. Another broad category of cancers includes sarcomas, which are tumors whose cells are embedded in a fibrillar or homogenous substance like embryonic connective tissue.

VLA-1 may be associated with adult and pediatric oncology in various forms of cancer, for example, growth of solid tumors/malignancies, myxoid and round cell carcinoma, locally advanced tumors, human soft tissue carcinoma (including Ewing's sar- coma), cancer metastases (including lymphatic metas- tases), squamous cell carcinoma (particularly of the : head and neck), esophageal squamous cell carcinoma, oral carcinoma, cutaneous T cell lymphoma, Hodgkin's lymphoma, non-Hodgkin's lymphomar cancer of the adrenal cortex, ACTH-producing tumors, nonsmall cell cancers, breast =cancer (including small cell carci- noma and ductal carcinoma), gastrointest. inal. cancers. <BR> <BR> <BR> <BR> <P>(including stomach cancer, colon cancer,. colorectal cancer, polyps associated with colorectal neoplasia, pa. ncreatic cancer and liver cancer), urological can- cers (including bladder cancer, especially primary superficial bladder tumors, invasive transitional cell carcinoma of the bladder, and muscle-invasive bladder cancer-and prostate cancer), malignancies of the. female genital. tract (including ovarian carci- noma, primary peritoneal. epithelial neoplasms, cervical carcinoma, uterine endometrial cancers, vaginal cancer, cancer of the vulva, uterine cancer and solid tumors in the ovarian follicle) malig- nancies of the male genital tract (including testic- ular cancer and-penile cancer), kidney cancer (in- cluding renal cell carcinoma), brain cancer (in- cluding intrinsic brain tumors, neuroblastoma, astrocytic brain tumors, gliomas and metastatic tumor cell invasion in central nervous system), bone

cancers (including osteomas and osteosarcomas), skin cancers (including malignant melanoma, tumor pro- gression of human skin keratinocytes and squamous cell cancer), thyroid cancer, retinoblastoma, neuro- blastoma, peritoneal effusion, malignant pleural effusion, mesothelioma, Wilms's tumors, gall bladder cancer, trophoblastic neoplasms,, hemangiopericytoma, and Kapos's sarcoma. Cancers and other cell pro- liferative disorders treatable by the compounds and/or compositions of the present invention also include angiogenesis-mediated diseases,. benign tumors (e.. g.., hemangiomas)., acoustic neuromas, neurofibromas, pyogenic granulomas, biliary tract cancer, choriocarcinoma, esophageal cancer, gastric cancer, intraepithelial neoplasms, lung cancer, and'. neuroblastomas.

The biological activity of the compounds identified'above may be assayed in a variety of sys- tems. For example, extracellular matrix, such as collagen IV, can be immobilized on a solid surface and adhesion of cells expressing VLA-l can be measured in the presence or absence of compound.

Using such formats, large. numbers of compounds can be screened. Cells suitable for this assay include smooth muscle cells, microvascular endothelial cells, fibroblasts, osteoblasts, chondrocytes, and activated cells of the immune system including effector T cells, macrophages and NK cells. A number of transfected cell lines can also be used, including, for example, CHO, K562, and the like.

The compounds and/or compositions of the invention can also be tested for the ability to inhibit binding between VLA-1 and extracellular matrix such as collagen IV, or between VLA-1 and a labeled compound known to. bind VLA-1 such as a com- pound and/or composition of this invention or anti- bodies to VLA-1. dIn these assays, the extracellular matrix can be soluble or immobilized on a solid surface. VLA-1 may also be expressed as a recombi- nant fusion protein having acidic and basic leucine zipper tails so that binding to extracellular matrix may be detected in an immunoassay.

Many assay formats employ labeled assay components. The labeling systems can be in a vari- ety of forms. The label may be. coupled directly or indirectly to the desired component of the assay according to methods well known in the art. A. wide variety of labels may be. used. The component may be labeled by any one of several methods. The most common method of detection is the use of auto- radiography with 3H, 125I, 35S, 14C, or 32P labeled compounds, and the like. Nonradioactive labels include europium, as well as ligands which bind to labeled antibodies, fluorophores, chemiluminescent agents, enzymes, and antibodies which can serve as specific binding pair members for a labeled ligand.

The choice of label depends on sensitivity required, ease of conjugation with the compound, stability requirements, and available instrumentation.

Appropriate in vivo models for demonstrat- ing efficacy in treating inflammatory responses in-

clude DTH (delayed type hypersensitivity) in mice, rats, guinea pigs, or primates, as well as other inflammatory or fibrotic models dependent upon VLA-1 integrin.

Compounds-having the desired biological activity may be modified as necessary to provide de- sired properties such as improved pharmacological properties (e. g., in vivo stability, bio-availabil- ity), or the ability to be detected in diagnostic applications. Stability can be. assayed in a variety of ways such as by measuring the half-life of the compounds during incubation with peptidases'or'human plasma or serum.

For diagnostic. purposes, a wide variety of labels may be linked to the compounds, which may provide, directly or indirectly, a detectable sig- nal. Thus, the compounds and/or compositions of the subject invention may be modified in a variety, of ways for a variety of end purposes while still re- taining biological activity. In addition, various reactive sites may be introduced for linking to particles, solid substrates, macromolecules, and the like.

Labeled compounds can be used in a variety of in vivo or in vitro applications. A wide variety of labels may be employed, such as radionuclides (e. g. , gamma-emitting radioisotopes such as tech- netium-99 or indium-111), fluorescers (e. g., fluor- escein), enzymes, enzyme substrates, enzyme cofac- tors, enzyme inhibitors, chemiluminescent compounds,. bioluminescent compounds, and the like. Those of

ordinary skill in the art will know of other suit- able labels for binding to the complexes, or will be able to ascertain such using routine experimenta- tion. The binding of these labels is achieved using standard techniques common to those of ordinary skill in the art.

In vitro uses include diagnostic applica- tions such as monitoring inflammatory responses by detecting the presence of cells expressing VLA-1.

The compounds and/or compositions of this invention can also be used for isolating or labeling such cells. In addition, as mentioned above, the com- pounds and/or compositions of the invention can be used to assay for potential inhibitors of VLA- 1/Extracellular matrix interactions.

For in vivo diagnostic, imaging to identi- fy, e. g. , sites of inflammation,, radioisotopes are typically used in accordance with well known tech- niques. The radioisotopes may be bound to the com- pound either directly or indirectly using intermedi- ate functional groups. For instance, cheating agents such as diethylenetriaminepentacetic acid (DTPA) and ethylenediaminetetraacetic acid (EDTA) and similar molecules have been used to bind com- pounds to metallic ion radioisotopes.

The complexes can also be labeled with a paramagnetic isotope for purposes of in. vivo diag- nosis, as in magnetic resonance imaging (MRI) or electron spin resonance (ESR), both of which are well known. In general, any conventional method for visualizing diagnostic images can be used. Usually

gamma-and positron-emitting radioisotopes are used for camera imaging and paramagnetic isotopes are used for MRI. Thus, the compounds can be used to monitor the course of amelioration of an inflamma- tory response in an individual. By measuring the increase or. decrease in cells expressing VLA-1 it is possible to determine whether a particular therapeu- tic regimen aimed at ameliorating the disease is effective.

Pharmaceutical compositions of the inven- tion are suitable for use in a variety of drug de- livery systems.-Suitable formulations for use in the present invention are found in"Remington's Pharmaceutical Sciences,"Mack Publishing Company, Philadelphia, Pa., 17th ed. (1985).

The amount administered to the patient will vary depending upon what is being administered, the purpose of the-administration, such as prophyl- axis or therapy, the state of the patient, the manner of administration, and the like. In thera- peutic applications, compositions are administered to a patient already suffering from a disease in an amount sufficient to cure or at least partially arrest the progression or symptoms of the disease and its complications. An amount adequate to accomplish this is defined as"therapeutically effective dose. "Amounts effective for this use will depend on the disease condition being treated as well as by the judgment of the attending clini- cian depending upon factors such as the severity of

the disease, disorder or condition, the age, weight and general condition of. the patient, and the like.

The compounds administered to a patient are typically in the form of pharmaceutical composi- tions described above ; These compositions may be sterilized by conventional sterilization techniques, or may be sterile filtered. The, resulting aqueous solutions may be packaged for use as is, or lyophil- ized, the lyophilized preparation being combined with a sterile aqueous carrier prior to administra- tion. The pH of the compound preparations typically.. will be between about 3 and 11, more'preferably from about 5 to 9, and most'preferably from about 7 to 8.

It will be understood that use of certain of the foregoing excipients, carriers, or stabilizers will result in the formation of pharmaceutical salts.

The therapeutic dosage of the compounds and/or compositions of the present invention will ; vary according to, for example,-the particular'use. for which the treatment is made, the manner of administration of the compound, the health and condition of the patient, and the judgment of. the prescribing physician. For example, for oral admin- istration, the dose will typically be in the range of about 100 ug to about 50 mg per kilogram body weight per day, preferably about 5 mg to about 20 mg per kilogram body weight per day. In the alterna- tive, for intravenous administration, the dose. will typically be in the range of about 20 ug to about 500 ug per kilogram body weight, preferably about 100 ug to about 300 ug per kilogram body weight.

Alternative routes of administration contemplated include, but are not limited to, intranasal, trans- dermal, inhaled, subcutaneous and intramuscular.

Effective doses can be extrapolated from dose-re- sponse curves derived from in vitro or animal model test systems.

In general, the compounds and/or composi- tions of the subject invention will be administered in a therapeutically effective amount by any of the accepted modes of administration for agents that serve similar utilities. Toxicity and therapeutic efficacy of such compounds can be determined by standard pharmaceutical procedures-in cell cultures or experimental animals,. et. g. , for determining the LD50 (the dose lethal to 50% cf the population)'and the ED50 (the dose therapeutically effective in 50% of the population). The dose ratio between toxic and therapeutic effects is the therapeutic index and it can be expressed as the ratio LD50/ED50. Compounds that exhibit large therapeutic indices are pre-- ferred.

The data obtained from the cell culture assays and animal studies. can be used in formulating a range of dosage for use in humans. The dosage of such compounds lies preferably within a range of circulating concentrations that include the ED50 with little or no toxicity. The dosage may vary within this range depending upon the dosage form employed and the route of administration utilized. For any compound and/or composition used in the method of the invention, the therapeutically effective dose

can be estimated initially from cell culture assays. A dose may be formulated in animal models to achieve a circulating plasma concentration range which in- cludes the IC50 (the concentration of the test com- pound which achieves a half-maximal inhibition, of activity) as determined in cell culture. Such in- formation can be used to more accurately determine useful doses in humans. Levels in plasma may be measured, for example, by high performance liquid chromatography.

The following synthetic and. biological examples are offered to illustrate this. invention and are not to be. construed in any way as limiting the. scope of this invention. Unless otherwise stated, all temperatures are in degrees'Celsius.

EXAMPLES Unless otherwise stated all temperatures are in degrees Celsius. Also, in these examples and elsewhere, have the following mean- ings: 1H NMR _ proton nuclear magnetic resonance 2JHF Geminal HF Coupling Constant A Angstrom ACN acetonitrile APT Attached Proton Test APCI Atmospheric Pressure Chemical Ionization bd-broad doublet Boc = tert-butoxycarbonyl br d broad doublet br s or bs-broad singlet CDC13 deuterated chloroform CDT = N, N'-carbonyldiimidazole' COSY correlated spectroscopy d doublet dba = dibenzylideneacetone DCM = dichloromethane dd = doublet of doublets DIEA diisopropylethylamine DMAP = dimethylaminopyridine DMF dimethylformamide DMSO = dimethyl sulfoxide DMSO-d6 deuterated dimethyl sulfoxide dt = doublet of triplets DTPA = diethylenetriaminepentacetic acid 1-(3-dimethylaminopropyl)-3- EDC (or EDCI) = ethylcarbodimide hydrochloride EDTA = ethylenediaminetetraacetic acid ELSD = Evaporative Light Scattering Detection ESI = electrospray ionization Et ethyl EtOH ethanol Et20 or ether diethyl ether EtOAc ethyl acetate Fmoc = 9-fluorenylmethoxycarbonyl g = grams h = hours HFBA = heptafluorobutyric acid HMBC = heteronuclear multiple bond correlation ~.- heteronuclear multiple quantum correlation HOBt (HOBT) 1-hydroxybenzotriazole HOD = mono deuterated water HPLC high performance liquid chromatography HSQC Heteronuclear Single Quantum Correlation Hz = hertz IPA isopropyl alcohol J = coupling constant in hertz L = liters LC method liquid chromatography method LCMS = liquid chromatography mass spectrometry M _ molar m multiplet m/z = mass/charge ratio Me = methyl mg milligrams MHz = megahertz min = minutes mL = milliliters mm = millimeter mm millimolar mmol = millimole macroporus poly (styrene-Co- divinylbenzene)-p-toluene sulfonic acid MS (EI) = mass spectrometry (electrospray ionization) MTBE tert-butyl methyl ether N normal NaOEt = sodium ethoxide nm nanometers nM. = nanomolar NMM = N-methyl morpholine NMP N-methylpyrrolidone NMR = nuclear-magnetic resonance Pd(OAc)2 = palladium acetate Pd/C-or Pd on C palladium on carbon Pd2 (dba) 3 _ tri (dibenzylideneacetone) dipalladium (0) PS-benzaldeyde = polystyrene benzaldehyde quartet racemic-2,2'-Bisdiphenylphosphino)- rac-BINAP = 1,1'-binaphthyl RPM = rotations per minute RP-HPLC = reverse phase hi. gh pressure liquid chromatography Rt retention time. rt = room temperature s = singlet sat. = t = td triplet of doublets TFA trifluoroacetic acid THF = Tol = toluene TOCSY = total correlation spectroscopy TSP = Thermo Separation Products 5 chemical shift µg = micrograms µL = microliters µM = micromolar BSA = bovine serum albumin CMF-PBS = calcium and magnesium-free Dulbecco's phosphate buffered saline dH20 distilled water D-PBS = Dulbecco's phosphate buffered saline FACS fluorescent-activated cell sorting FBS = fetal bovine serum PMA-phorbol 12-myristate 13-acetate = Roswell Park Memorial Institute media 1640 TBS = tris buffered saline

The following analytical HPLC methods (or LC Methods) are referred to the Examples. The gradient profiles were gradients in- creasing or decreasing linearly over the time period indicated.

Method A: Varian HPLC System: (Pumps : Varian ProStar Solvent Deiivery System, Model 210; Detector: Varian ProStar UV-VIS Detector, Model <BR> <BR> <BR> 345 ; Autosampler : Varian ProStar Autosampler, Model.

430). Analytical column : YMC ODS-AQ, 4. 6x50 mm, S3 u, Maters Corporation. Detection : 220 nm and 254nm. Solvent A: H2O, 0. 1% TFA, 1. 0%, IPA. Solvent B : Acetonitrile, 0. 05% TFA, 1. 0% IPA. Flow Rate: 2. 0 mL/min. Gradient Program : 0. 00min 95% Solvent A, 5% Solvent B; 0.05min. 95% Solvent A, 5% Solvent Bu 4. 12 min 5% Solvent A, 95% Solvent B; 4.24min 5% Solvent A, 95% Solvent B; 4. 30min 95% Solvent A, 5% Solvent B; 5. 00min 95% Solvent A, 5% Solvent B.

Method B: Varian HPLC System : (Pumps: Varian ProStar Solvent Delivery System, Model 210; Detector: Rainin Dynamax Absorbance Detector, Model UV-DII ; Autosampler: Varian ProStar Autosampler, Model 430). Analytical column: YMC ODS-AQ, 4. 6x50mm, S3m, Waters Corporation. Detection: 220nm and 254nm. Solvent A: H2O, 0. 01% HFBA, 1. 0%

IPA. Solvent B: Acetonitrile, 0. 01% HFBA, 1.0% IPA.

Flow. Rate: 2.0 mL/min. Gradient Program: 0. 00min 95% Solvent A, 5% Solvent B; 0.12min 95% Solvent A, 5% Solvent B; 4.00 min 5% Solvent A, 95% Solvent B; 4. 18min 5% Solvent A, 95% Solvent B; 4.30min 95% Solvent A, 5% Solvent B ; 5. 30min 95% Solvent A, 5% Solvent B.

Method C : Varian HPLC System: (Pumps : Vari. an ProStar Solvent Delivery System, Model 210 ; Detector: Varian ProStar PDA, Model 330 ; Auto- sampler: Varian ProStar Autosampler, Model 430).

Analytical column: YMC ODS-AQ, 4. 6x50mm, S3m, Waters Corporation. Detection: 220nm and 254nm.

Solvent A : dH2O, 001% HFBA, 1. 0% IPA. Solvent B: Acetonitrile, 0. 01% HFBA, 1. 0% IPA. Flow Rate: 2. 0 mL/min. Gradient Program : 0. OOrnin 95% Solvent A, 5% Solvent B ; 0. 12min 95% Solvent A, 5% Solvent B ; 4. 00 min 5% Solvent A, 95% Solvent B; 4.18min 5% Solvent A, 95% Solvent B; 4. 30min 5% Solvent A, 95% Solvent B; 5. 00min 95% Solvent A, 5% Solvent B 5. 30min 95% Solvent A, 5% Solvent B.

Method D: Berger SFC (Berger Dual Pump Fluid Control Module, Model FCM-1200 ; Berger Thermal Control Module, Model TCM-2000 ; Hewlett Packard 1100 Series DAD, Model G1315A ; Alcott Au. tosampler, Model 718AL). Column : Chiralcel OJ-R 150x4. 6 mm. De- tection : 258nm. SFC Conditions : 2 mL/min, 10% MeOH in C02, 35°C, 200 bar.

Method E: Agilent Technologies 1100 HPLC System (Pump: QuatPump Model G1311A; Detector: DAD, Model G1315B ; Column Compartment: Model

G1216A ; Autosampler: ALS, Model G1313A; Degasser: Model G1322A). Analytical column : Chiralcel OD-RH, 150x4. 6 mm. Detection: 220nm. HPLC Conditions : 1.0 mL/min, Isocratic, 70% (H2O, 0. 01% TFA), 30% (ACN, 0. 01% TFA), 35°C.

Method F: Mass Spectrometer: Finnigan LCQTMDuo, APCI. TSP HPLC System: (Pump: TSP SpectrSYSTEM# P4000; Detector: TSP SpectraSYSTEM# UV2000, 220nm and 254m ; Autosampler : TSF Spectra- SYSTEM AS3000 ; Degasser : TSP SpectraSYSTEMG Model SCM1000 Solvent Degasser).. Detection : 220nm and 254nm. Analytical-column : YMC ODS-AQ, 4.6x50mm, S3m, Waters Corporation. Solvent C: H2O, 0.01% HFBA, 1. 0% IPA.. Solvent. D: Acetonitrile, 0. 01% HFBA, 1. 0% IPA. Flow Rate: 2.0 mL/min. Gradient Program : 0. 00min 95% Solvent C,. 5% Solvent D ; 0. 02min 95% Solvent C, 5% Solvent D; 4.00 min 5%' Solvent C, 95%. Solvent D ; 4. 30min 5% Solvent C, 95% Solvent D ;, 4. 50min 95% Solvent C, 5% Solvent D; 5. 50min 95% Solvent C, 5% Solvent D.

Method. G: Mass Spectrometer: Finnigan LCQTMDuo, ESI. TSP HPLC System: (Pump: TSP Spec- traSYSTEM# P4000; Detector: TSP,-SpectraSYSTEMO UV2000, 220nm and 254m; Autosampler: TSP Spectra- SYSTEMs AS3000 ; Degasser: TSP SpectraSYSTEM# Model SCM1000 Solvent Degasser). Detection: 220nm and 254nm. Analytical column: YMC. ODS-AQ 4. 6x50mm S3m, Waters Corporation. Solvent C: 10 mM Ammonium Acetate in H2O. Solvent D: 10 mM Ammonium Acetate in Acetonitrile, 1. 0% IPA. Flow Rate: 2.0 mL/min.' Gradient Program: 0. 00min 75% Solvent C, 25% Solvent

D; 0.02min 75% Solvent C, 25% Solvent D 4. 00 min 5% Solvent C, 95% Solvent D ; 4.30min 5% Solvent C, 95% Solvent D ; 4.50min 75% Solvent C, 25% Solvent D; 5.50min 75% Solvent C, 25% Solvent D.

Method H, :. Mass Spectrometer: Finnigan LCQwDuo, ESI. TSP HPLC System: (Pump : TSP Spec- traSYSTEMs P4000 ; Detector: TSP SpectraSYSTEM'y UV2000, 22Onm and 254m; Autosampler: TSP Spectra- SYSTEMs AS3000; Degasser: TSP SpectraSYSTEM# Model SCM1000 Solvent Degasser). Detection: 220nm and 254nm. Analytical column: Zorbax Extend C18 Rapid Resolution#, 50x4.6mm, 3.5m, 80A, Agilent Technol- ogies. Solvent. C: 10 mM Ammonium Acetate in H2O.

Solvent D: 10 mM Ammonium Acetate in Acetonitrile, 1. 0%. IPA. Flow Rate: 2. 0 mL/min. Gradient Pro- gram: 0. 00min 95% Solvent C, 5% Solvent D; 0. 02min.

95% Solvent C, 5% Solvent D ; 4.00 min 5% Solvent C, 95% Solvent D; 4.30min 5% Solvent C, 95% Solvent D ; 4.50min 95% Solvent C, 5% Solvent D; 5.50min 95% Solvent C, 5% Solvent D.

Method I : Mass Spectrometer:. Finnigan LCQTMDuo, ESI. TSP HPLC System:. (Pump: TSP Spec- traSYSTEMs P2000 ; Detector: TSP SpectraSYSTEM# UV2000,220nm and 254m; Autosampler: TSP Spectra- SYSTEMs AS3000 ; Degasser: TSP SpectraSYSTEM° Model SCM1000 Solvent Degasser). Detection: 220nm and 254nm. Analytical column: Zorbax Extend C18 Rapid Resolution@, 50x4.6mm, 3.5m, 80A, Agilent Technol- ogies. Solvent A: 10 mM Ammonium Acetate in H2O.

Solvent B: 10 mM Ammonium Acetate in Acetonitrile, 1. 0% IPA. Flow Rate: 2.0 mL/min. Gradient Pro-

gram: 0. 00min 95% Solvent A, 5% Solvent B ; 0.02min 95% Solvent A, 5% Solvent B; 4.00 min 5% Solvent A, 95% Solvent B; 4.. 30min 5% Solvent A, 95% Solvent B ; 4.50min 95% Solvent A, 5% Solvent B; 5.50min 95% Solvent A, 5% Solvent B.

Method J: Mass spectrometers: LCQTMDuo and LCQTMDeca. Pump : Series 1100, Quat pump model G131. 1A, Agilent Technologies. Detector: Series 1100, Column model G1216A, Agilent Technologies.

Autosampler :' Series 1100 ALS model G1313A, Agilent Technologies. Degasser: Series 1100, Degasser model G1322A, Agilent Technologies. ELSD : Sedex model 75, Sedere. Analytical column : YMC ODS-AQ 4. 6x50mm S3u, Waters Corporation... Solvent A:. H2O, 0. 01% HFBA, 1. 0% IPA. Solvent. B: Acetonitrile, 0. 01% HFBA, 1. 0% IPA. Flow. Rate:. 2. 0 mL/min.

Gradient Program : 0. 00min 95% Solvent A, 5% Solvent Bu 0. 02min 95% Solvent A, 5% Solvent B; 4. 10 min. 5% Solvent A,. 95% Solvent B ;.. 4.30min 5% Solvent A, 95% Solvent B ;'4. 50min 95% Solvent A, 5% Solvent B; 5.50min 95% Solvent A, 5% Solvent B. Purity based on 220 nm wavelength channel.

Example 1 <BR> <BR> <BR> <BR> (S)-N- {1- [3- (2, 3-Dichloro-phenyl)-acryloyl]-piperldin- 4-yl}-2-methyl-succinamic acid (55 : 45 regioisomeric mixture of 2-methyl and 3-methyl-succinamic acids)

Example 1A 3-(2, 3-Dichloro-phenyl)-acrylic acid To a solution of 2, 3-dichlorobenzaldehyde ('75 g, 429 mmol) in pyridine (220 mL) was added malonic acid (82 g ; 788 mmol) and piperidine (3. 5 g, 41 mmol). The mixture was heated at 110°C until gas evolution had ceased, and then heated for an addi- tional 3 hours at that temperature. The mixture was concentrated in vacuo, dissolved in 3.0 M NaOH (1 L) and H20 (2 L) with heating at 90°C to give complete dissolution, and then allowed to cool overnight.

The resulting white crystalline solid was collected by filtration and washed with water. The aqueous filtrate was extracted with MTBE (2 L) and the crystalline solid was then combined with the basic aqueous layer. This mixture was acidified with 12 M

HC1 to pH 1 with stirring to produce a white solid, which was collected by filtration and washed with water. The solids were dried under high vacuum to give the indicated product as a white powder (82. 7 g, 89%). 1H NMR (acetone-d6,400 MHz) 5 8.06 (d, J=16.0 Hz, 1H), 7.86 (dd, J=7.8, 1.2 Hz, 1H), 7. 66 (dd, J=7.9, 1.3 Hz, 1H), 7. 44 (t, J=8. 0. Hz, 1H), 6.60 (d, J=15. 6 Hz, 1H).

Example 1B <BR> <BR> <BR> <BR> <BR> 1-(4-Amino-piperidin-l-yl)-3-{2, 3-dichloro-<BR> <BR> <BR> <BR> phenyl)-propenone A. solution of Example 1A (5. 43 g, 25. 0 mmol) in DMF (25 mL) was stirred with 4-N-Boc-amino- piperidine (7.51 g, 37.5 mmol), l-hydroxybenzotri- azole hydrate (7.66 g, 0 mmol), N-methylmorph- oline (8.25 mL, 75.0 mmol), and 1- (3-dimethylamino- propyl) -3-ethylcarbodiimide hydrochloride (9.59 g, 50.0 mmol) under N2 for 16 h. The reaction was diluted with methylene chloride (200 mL) and water (200 mL), and the phases were separated. The or- ganic phase was then washed with IN aqueous HC1 (2x100 mL) followed by a sat. solution of NaHCO3 (200 mL). The organic phase was dried (Na2SO4), filtered, and concentrated to a white solid (9.88 g, 99%) which was carried forward without purification. The

Boc group was removed by stirring a suspension of the white solid (9.88 g, 24.7 mmol) with methanol (20 mL), THF (100 mL) and 4N HCl in dioxane (54 mL, 124 mmol) under N2 for 16 h. The suspension was diluted with diethyl ether (200 mL), and the white solid was filtered, washing with diethyl ether (3x50 mL). The solid was partitioned between methylene chloride (100 mL) and 1N aqueous NaOH (100 mL). The phases were separated, and the aqueous phase was re- extracted with methylene chloride (2x100 mL). The combined organic phases were washed with brine (100 mL), dried (Na2SO4), filtered, and concentrated to obtain the product as a waxy, white solid (7.0 g, 95%). 1H NMR (CDC13, 400 MHz) 5 7.95 (d, J=15.2 Hz, 1H), 7.48 (dd, J=7.81, 1.56 Hz,. 1H), 7.44 (dd, J=8. 20, 1. 56 Hz, 1H), 7.20 (m, 1H), 6. 85 (d, J=15. 2 Hz, 1H), 4. 58 (m, 1H), 4.03 (m, 1H), 3., 9 (m, 1H), 2.97 (m, 1H), 2.87 (m, 1H), 1. 91 (m, 2H), 1. 27-1. 55 (m, 4H). LCMS (ESI+ ; Method G) m/z 299, 301 (M+H) +; Rt=2. 00 min (220 nm, 100 area%).

Example 1C (S)-N- {l- [3- (2, 3-Dichloro-phenyl)-acryloyl]-piperidin-4-yl}- 2-methyl-succinamic acid (55: 45 regioisomeric mixture of 2-methyl and 3-methyl-succinamic acids)

A solution of Example 1B (54 mg, 0.18 mmol) in CHC13 (3.25 mL) was shaken (400 RPM) with (S) -methyl succinic anhydride (17 mg, 0.15 mmol; prepared from (S) -2-methyl succinic acid according to the literature :'--Davies, S. G. ; Dixon, D. J. J.

Chem. Soc., Perkin Trans. 1 1998, 2635-2643) for 16 h at room temperature. PS-Benzaldehyde (226 mg, 0.300 mmol) was added (Argoscoop@, Argonaut, Foster City, CA,-USA). The reaction was shaken (400 RPM) for an additional : 6 h, and then filtered, rinsing the resin with methylene chloride (3x2 mL). The filtrate was concentrated. A. solution of 1N aqueous HC1 (5 mL) was added to precipitate the product.

The solid was filtered, washing with water (2x2 mL).

The product was dried (COSY4 dessicator) under high vacuum for 16 h. The product was obtained as a white powder (47 mg, 76%). 1H NMR (10% DMSO-due in CDC13, 400 MHz ; 55: 45 mixture of 2-methyl and 3- methyl succinamide regioisomers, respectively) 5 7. 96 (d, J=15.6 Hz, 2H), 7. 51 (m, 4H), 7.24 (m, 2H), 6. 88 (d, J=15.6 Hz, 2H), 4.53 (m, 2H), 3. 93-4.09 (m, 4H), 3.29 (m, 2H), 2.87-2. 99 (m, 3H), 2.70 (m, 2H), 2.54 (m, 1H), 2.38 (m, 1H), 2.27 (m, 1H), 1.92-2. 07 (m, 4H), 1. 42 (m, 4H), 1.22 (d, J=7.03 Hz, 3H; CH3 of 2-methyl succinamide), 1.18 (d, J=6.25 Hz, 3H; CH3 of 3-methyl succinamide). Regiochemical assignment was based on correlation of COSY, HMQC, HMBC and APT multidimensional NMR experiments. LCMS (APCI+; Method F) m/z 413,415 (M+H) +; Rt=2.63 min (220 nm, 100 area% ; regioisomers not separated). Enantio- meric purity (>98%) was determined by chiral HPLC

(Method E): Example 1C, 2-methyl succinamide: Rt=10. 5 min (60 area%), 3-methyl succinamide regioisomer: Rt=14.3 min (40 area%) ; chiral HPLC (Method E) analysis of a mixture of the enantiomers gave different retention times: Rt=16.2 min (46 area%), Rt=25.2 min (54 area%). Neither of the"R" enantiomers were detected (<2%) by chiral HPLC of Example 1C.

Example 2 N- {1- [3- (2, 3-Dichloro-phenyl)-acryloyl]-piperidin-4-yl}- 2-methylene-succinamic acid (78: 22 regioisomeric mixture of 2-methylene and 3-methylene-succinamic acids) Example 2 was prepared from Example 1B (54 mg, 0.18 mmol) and itaconic anhydride (17 mg, 0.15 mmol) as detailed in Example 1C to provide the title compound (33 mg, 53%) as a white solid. 1H NMR (DMSO-d6,400 MHz, 78: 22 mixture of regioisomers) 5 12.46 (bs, 1H), 8.01 (m, 1H), 7.96 (m, 0.2H, minor regioisomer), 7.91 (d, J=7.80 Hz, 1H), 7.79 (m, 1H), 7.68 (dd, J=7. 80,1. 17 Hz, 1H), 7.35-7. 45 (m, 2H), 6.1 (d, J=1.95 Hz, 1H), 5.78 (m, 0. 2H, minor regio- isomer), 5.65 (d, J=1.56 Hz, 1H), 5.47 (m, 0.2 H, minor regioisomer), 4.23 (m, 2H), 3.82 (m, 1H), 3.18-3. 30 (m, 1H, overlaps with H2O), 3. 08 (s, 2H), 2.90 (m, 1H), 1.78 (m, 2H), 1.30 (m, 2H). Regio-

chemical assignment was based on correlation of COSY, HSQC, and HMBC multidimensional NMR experi- ments. LCMS (APCI- ; Method F) m/z 411,413 (M+H) +; Rt=2.63 min (220 nm, 67.4 area%) and 2.67 (220 nm, 23.9 area%).

Example 3 N- {1- [3- (2, 3-Dichloro-phenyl)-acryloyl]--piperidin- 4-yl}-succinamic acid The title compound was prepared from Example 1B (54 mg, 0. 18 mmol) and succinic anhydride (15 mg, 0.15 mmol) as detailed in Example 1C, with the following purification step added : the crude solid was dissolved in. IN aqueous NaOH (10 mL) and extracted with diethyl ether (3x10 mL). The aqueous phase was then acidified with 6N aqueous HC1, and extracted with methylene chloride (3x20 mL). The organic phase was dried (Na2SO4), filtered, and concentrated to obtain Example 3 as a white solid (22 mg, 37%). lH NMR (DMSO-d6,400 MHz) 5 12.07 (bs, 1H), 8.01 (dd, J=7.80, 1.17 Hz 1H), 7.87 (d, J=7.80, 1H), 7.80 (d, J=15.2 Hz, 1H), 7.67 (dd, J=8. 19,1. 56 Hz, 1H), 7.35-7. 45 (m, 2H), 4.23 (m, 2H), 3.82 (m, 1H), 3.24 (m, 1H), 2.90 (m, 1H), 2.42 (m, 2H), 2.31 (m, 2H), 1.78 (m, 2H), 1.28 (m, 2H). LCMS (APCI+; Method F) m/z 399, 401 (M+H) +; Rt=2.55 min (220 nm, 100 area%).

Various syntheses of Examples 4 through 16. are presented in Table 1 below.

Example 4 Cis-6- » {3-[2, 3-Dichloro-4-(2-methoXy-phenylsulfanyl)- phenyl]-acryloyl}-piperidin-4-ylcarbamoyl)-cyclohex-3- enecarboxylic acid Example 4A <BR> <BR> <BR> <BR> <BR> <BR> (1- {3- [2, 3-Dichloro-4- (2-methoxy-phenylsulfanyl)-phenyl]- acryloyl}-piperidin-4-yl)-carbamic acid tert-butyl ester The title compound was prepared from 3- [2, 3-Dichloro-4- (2-methoxy-phenylsulfanyl)-phenyl]- acrylic acid (1.30 g, 3.66 mmol; WO 00/59880, WO 00/39081) and 4-N-Boc-aminopiperidine (880 mg, 4.39 mmol) according to the amide coupling protocol for Example 1B. The product (1.96 g, 100%) was obtained as a beige powder and carried forward to the next step without purification. 1H NMR (CDCl3,

400 MHz) 5 7. 92 (d, J=15.6 Hz, 1H), 7.47 (m, 2H), 7. 26 (m, 1H, overlaps with CHC13), 7. -02 (m, 2H), 6. 74 (d, J=15.6 Hz, 1H), 6.55 (d, J=8.98 Hz,-lH), 4.58 (m, 1H), 4.45 (m, 1H), 3.95 (mi 1H), 3.83 (s, 3H), 3.69 (m, 1H), 3.19 (m, 1H), 2.85 (m, 1H), 2.02 (m, 2H), 1.44 (s, 9H), 1.33 (m, 2H).

1- (4-Amino-piperidin-1-yl)-3- [2, 3-dichloro-4- (2-<BR> <BR> <BR> <BR> methoxy-phenylsulfanyl)-phenyl]-propeonone The title compound was prepared from Example 4A (1.84 mg, 3. 42 mmol) according to the Boc deprotection protocol described for Example 1B. The product (1. 27 g, 85%) was obtained as a fluffy, white powder after flash chromatographic purifica- tion (3-7% methanol in methylene chloride). 1H NMR (CDCl3, 4Q0 MHz) 5 7.91 (d,'J=15. 6 Hz, 1H), 7.47 (m, 2H), 7.27 (m, 1H, overlaps with CHC13), 7.02 (m, 2H), 6.76 (d, J=14. 5 Hz, 1H), 6.56 (d, J=8. 2 Hz, 1H), 4.57 (m, 1H), 3.99 (m, 1H), 3. 84 (s, 3H), 3.15 (m, 1H), 2.94 (m, 1H), 2.83 (m, 1H), 1.89 (m, 2H), 1.52 (bs, 2H, overlaps with HOD), 1.30 (m, 2H).

Analytical HPLC (Method A) : Rt=2.83 min (94. 5 area%, 220 nm).

Example 4C (Procedure A in Table 1) <BR> <BR> <BR> <BR> <BR> Cis-6- (1- {3- [2, 3-dichloro-4- (2-methoxy-phenylsulfanyl)-<BR> <BR> <BR> phenyl]-acryloyl}-piperidin-4-ylcarbamoyl)-cyclohex-3- enecarboxylic acid A 24-well format, Bohdan block'was charged with a solution of Example 4B in chloroform (0.084 M, 1.0 mL, 0.084 mmol), and a solution of cis- 1, 2,3, 6-tetrahydrophthalic anhydride in chloroform (0.24 M, 0.29 mL, 0.070 mmol, via Packard liquid handler). The block was shaken for 16 h at 500 RPM (Bohdan block shaker). PS-Benzaldehyde (105 mg, 0.14 mmol, via Argoscoop@, Argonaut, Fo, ster City, CA, USA) and chloroform (1. O. mL) were added, and the block was shaken for an additional 16 h at 600 RPM.

The reaction was filtered rinsing the. resins with methylene chloride (10x0. 5 mL). The filtrate was concentrated, and the crude product was purified by partial dissolution (digestion) in hot toluene followed with cooling to room temperature and fil- tration. Example 4C was analyzed by LCMS (results in Table 1).

Example 9 (Proceudre B in Table 1) N- (1- {3- [2, 3-Dd-chloro-4- (2-methoxy-phenylsulfanyl)- phenyl]-acryloyl}-piperidin-4-yl)-(S)-2-(2, 2,2- trifluoro-acetylamino) -succinamic acid After synthesis according'to the procedure described for Example 4C, but omitting the purifica- tion step (digestion in toluene), a solution of Example 9 (0. 140 mmol, prepared on twice the scale) in methylene chloride (10'mL) was shaken with more of the scavenging reagent MP-TsOH (183 mg, 0.28 mmol) for 2 h at 300 RPM.'The r'eaction was filtered rinsing the resins with methylene chloride (10x0. 5 mL). The filtrate was concentrated, and Example 9 was analyzed by LCMS (results in Table 1).

Example 14 <BR> <BR> <BR> <BR> <BR> 4- (1- {3- [2, 3-Dichloro-4- (2-methoxy-phenylsulfanyl)-<BR> <BR> <BR> <BR> phenyl]-acryloyl}-piperidin-3-ylcarboamoyl)-3- methyl-butyric acid

Example 14A (1- {3- [2, 3-Dichlbro-4- (2-methoxy-phenylsulfanyl)- phenyl]-acryloyl}-piperidin-3-yl)-carbamic acid tert-butyl ester The title compound was prepared from 3- [2, 3-Dichloro-4- (2-methoxy-phenylsulfanyl)-phenyl]- acrylic acid (1. 30 g, 3. 66 mmol ; WO 00/59880, WO 00/39081) and 3-N-Bbc-aminopiperidine (880 mg, 4. 39'mmol) according to the amide coupling protocol for Example 1B. The product (1. 89 g, 96%) was obtained as a white powder and carried'forward to the next step without purification. 1H NMR (CDC13, 400 MHz) 6 8. 00 (br d, J=12. 1 Hz, 1H}, 7. 47 (m, 2H),.

7.30 (m, 1H)', 7.02 (m, 2H), 6. 79 (br d, J=12.9 Hz, 1H), 6.55 (d, J=8. 59 Hz, 1H), 4.60 (bs, 1H), 3.89 (m, 1H), 3.83 (s, 3H), 3.66 (m, 3H), 3.45 (m, 1H), 1.91 (m, 1H), 1. 6-1. 8 (m, 3H), 1.3-1. 5 (m, 9H).

Example 14B <BR> <BR> <BR> <BR> <BR> 1- (3-Amino-piperidin-1-yl)-3- [2, 3-dichloro-4-<BR> <BR> <BR> <BR> (2-methoxy-phenylsulfanyl)-phenyl]-propenone The title compound was prepared from Example 14A (1.77 g, 3.29 mmol) according to the Boc deprotection protocol described for Example 1B. The product (1.30 g, 90%) was obtained as a fluffy, white powder after flash chromatographic purifica- tion (3-7% methanol in methylene chloride). 1H NMR (CDC13, 400 MHz) 5 7.93 (br d, J=15. 6 Hz, 1H), 7.47 (m, 2H), 7.26 (m, 1H, overlaps with CHC13), 7.02 (m, 2H), 6.77 (m, 1H), 6.56 (d, J=8.59 Hz, 1H), 4.21- 4.48 (m, 1 H), 3.84 (m, 4H), 2.60-3. 17 (m, 3H), 1.99 (m, 1H), 1.78 (m, 1H), 1.53 (m, 2H), 1.34 (m, 2H).

Analytical HPLC (Method A): Rt=2.86 min (95.9 area%, 220 nm).

Example 14C (Procedure C in Table 1) <BR> <BR> <BR> <BR> 4- (1- {3- [2, 3-Dichloro-4- (2-methoxy-phenylsulfanyl)-<BR> <BR> <BR> phenyl]-acryloyl}-piperidin-3-ylcarbamoyl)-3- methyl-butyric acid The title compound was prepared according to the same procedure described for Example 4C, except using the product in 14B in place of the product from 4B, and the purification step (diges- tion in toluene) was omitted. Example 14C was analyzed by LCMS (results in Table 1).

There is no Example 17.

Table 1: Cyclic Anhydride Acylation Library (Examples 4-16) Starting LC Observed Rt Purity Ex. # Name Procedure Anhydride Method Mass1 (min) (% Area) Cis-6-(1-(3-[2,3-Dichloro-4- cis-3a,4,7,7a- (2-methoxy-phenylsulfanyl)- Tetrahydro- 4 phenyl]-acryloyl}-piperidin-4- isobenzofuran- A F 589,591 3.61 90.4 ylcarbamoyl)-cyclohex-3- 1,3-dione enecarboxylic acid 4-(1-(3-[2,3-Dichloro-4-(2- 3-Phenyl- methoxy-phenylsulfanyl)- dihydro-pyran- 5 phenyl]-acryloyl}-piperidin-4- c J 627,629 3.78 90.1 2,6-dione ylcarbamoyl)-2-phenyl-butyric acid 2-(1-{3-[2,3-Dichloro-4-(2- methoxy-phenylsulfanyl)- 3-Oxa-bicyclo 6 phenyl]-acryloyl}-piperidin-4- [3.1.0]hexane- C J 549,551 3.51 84.8 ylcarbamoyl)- 2,4-dione cyclopropanecarboxylic acid 4-(1-{3-[2,3-Dichloro-4-(2- methoxy-phenylsulfanyl)- 4-Methyl- 7 C J 565,567 3.55 82.1 phenyl]-acrylocyl}-piperidin-4- dihydro-pyran- ylcarbamoyl)-3-methyl-butyric 2,6-dion acid 4-(1-{3-[2,3-Dichloro-4-(2- 8 methoxy-phenylsulfanyl)- Dihydro-pyran- C J 551,553 3.47 91.5 phenyl]-acryloyl}-piperidin-4- 2,6-dione ylcarbamoyl)-butyric acid Table 1: Cyclic Anhydride Acylation Library (Examples 4-16) Starting LC Observed Rt Purity Ex. # Name Procedure Anhydride Method Mass1 (min) (% Area) N-(1-{3-[2,3-Dichloro-4-(2- N-(2,5-Dioxo- methoxy-phenylsulfanyl)- tetrahydro- 9 B F 648,650 3.51 75.5 phenyl]-acryloyl}-piperidin-4- furan-3-yl)- yl)-(S)-2-(2,2,2-trifluoro- 2,2,2-trifluoro- acetyulamino)-succinamic acid acetamide N-(1-{3-[2,3-Dichloro-4-(2- 3-Methyl- 10 methoxy-phenylsulfanyl)- C J 551,553 3.53 92.5 dihydro-furan- phenyl]-acryloyl}-piperidin-4- 2,5-dione yl)-2-methyl-succinamic acid N-(1-{3-[2,3-Dichloro-4-(2- 11 methoxy-phenylsulfanyl)- Dihydro-furan- C J 537,539 3.46 93.2 phenyl]-acryloyl}-piperidin-4- 2,5-dione yl)-succinamic acid N-(1-{3-[2,3-Dichloro-4-(2- 3-Phenyl- 12 methoxy-phenylsulfanyl)- C J 613,615 3.77 91.7 dihydro-furan- phenyl]-acryloyl}-piperidin-4- 2,5-dione yl)-2-phenyl-succinamic acid N-(1-{3-[2,3-Dichloro-4-(2- methoxy-phenylsulfanyl)- 3,3-Dimethyl- 13 C J 565,567 3.63 86.4 phenyl]-acryloyl}-piperidin-4- dihydro-furan- yl)-2,2-dimethyl-succinamic 2,5-dione acid 4-(1-{3-[2,3-Dichloro-4-(2- methoxy-phenylsulfanyl)- 4-Methyl- 14 C J 565,567 3.56 100 phenyl]-acryloyl)-piperidin-3- dihydro-pyran- ylcarbamoyl)-3-methyl-butyric 2,6-dione acid 15 N-(1-{3-[2,3-Dichloro-4-(2- N-(2,5-Dioxo- B F 648,650 3.56 89.1 methoxy-phenylsulfanyl)- tetrahydro- Table 1: Cyclic Anhydride Acylation Library (Examples 4-16) Starting LC Observed Rt Purity Ex. # Name Prodedure Anhydride Method Mass1 (min) (% Area) phenyl]-acryloyl}-piperidin-3- furan-3-yl)- yl)-(S)-2-(2,2,2-trifluoro- 2,2,2-trifluoro- acetylamino)-succinamic acid acetamide N-(1-{3-[2,3-Dichloro-4-(2- 3-Methyl- 16 methoxy-phenylsulfanyl)- C J 551,553 3.55 100 dihydro-furan- phenyl]-acryloyl)-piperidin-3- 2,5-dione yl)-2-methyl-succinamic acid 1 The detailed description of the observed mass is described in the designated LC Method.

Example 18 <BR> <BR> N- {3-[2, 3-Dichloro-4-(4-fluoro-phenylsulfanyl)- phenyl]-acryloyl}-piperidin-4-yl)-malonamic acid

Example 18A N- (l-Benzyl-piperidin-4-yl)- malonamic acid ethyl ester A, stirred mixture of 4-amino-l-benzyl- piperidine (1.00 g, 5.25 mmol) and diethyl malonate (7.90 mL, 52.0 mmol) was heated to 100°C under N2 for 7 h. The reaction was allowed to cool'to room tem- perature and stirring was continued for 16 h. The crude product was purified by flash chromatography, using methylene chloride to elute the excess diethyl malonate, followed by 5% methanol in methylene chloride to elute the product. Example 18A (1.00 g, 63%) was obtained as a yellow oil which solidified upon standing. 1H NMR (CDC13, 400 MHz) 5 7.29-7. 32 (m, 4H), 7.26 (m, 1H), 7.07 (bd, J=6.64 Hz, 1H), 4.20 (q, J=7.03 Hz, 2H), 3.84 (m, 1H), 3.49 (s, 2H), 3.28 (s, 2H), 2.78 (m, 2H), 2.15 (m, 2H), 1.91 (m, 2H), 1.53 (m, 2H), 1.29 (t, J=7. 03 Hz, 3H).

Example 18B N-Piperidin-4-yl-malonamic acid ethyl ester A solution of Example 18A (1.00 g, 3.29 mmol) in absolute EtOH (13 mL) was stirred with Pd on C (348 mg, 0.164 mmol; 10% Pd, 50% water, Degussa type) under an H2 atmosphere for 16 h. The suspen- sion was filtered through celite,'rinsing with EtOH.

The filtrate was concentrated. The resulting resi- due was triturated with diethyl ether and filtered.

The resulting sticky solid was dissolved in methyl- ene chloride and concentrated to give Example 18B as a foamy, yellow solid. 1H NMR (CDCl3, 400 MHz) 5 7. 62 (bd, J=7.42 Hz, lH}, 4. 19 (q, J=7.16 Hz, 2H), 4.06 (m, 1H), 3.43 (m, 2H), 3.29-3. 34 (m, 2H), 2.98 (m, 2H), 2.14 (m, 2H), 1.87 (m, 2H), 1. 29 (t, J=7.22 Hz, 3H).

Example 18C N- (l- {3- [2, 3-Dichloro-4- (4-fluoro-phenylsulfanyl)- phenyl]-acryloyl}-piperidin-4-yl)-malonamic acid ethyl ester The title compound was prepared from 3- [2, 3-dichloro-4- (4-fluoro-phenylsulfanyl)-phenyl]- acrylic acid (415 mg, 1.21 mmol; synthesized in a similar manner to the acrylic acid used in Example 4A by methodology described in WO 00/59880, WO 00/39081 except that 4-fluorothiophenol was substituted for 2-methoxythiophenol) and Example 18B (390 mg, 1. 8 mmol) according to amide coupling protocol described for Example 1B. The product (650 mg, 100%) was obtained as an off-white solid. 1H NMR (DMSO-d6, 400 MHz) 5 8. 31 (bd, J=7.81 Hz, 1H), 7.90 (d, J=8.59 Hz, 1H), 7. 73 (d, J=15.2 Hz, 1H), 7.66 (m, 2H), 7.41 (m, 2H), 7.29 (d, J=15.6 Hz, 1H), 6.67 (d, J=8.59 Hz, 1H), 4.24 (m, 1H), 4.12 (m, 1H), 4.06 (q, J=7. 16 Hz, 2H), 3.83 (m, 1H), 3.22 (s, 2H), 2. 96 (m, 2H), 1.77 (m, 2H), 1.31 (m, 2H), 1.17 (t, J=7.03 Hz, 3H). LCMS (APCI+ ; Method F) m/z 539,541 (M+H) +; Rt=3.77 min (220 nm, 86.6 area%).

Example 18D N-(1-{3-[2,3-Dichloro-4-(4-fluoro-phenylsulfanyl)- phenyl]-acryloyl}-piperidin-4-yl)-malonamic acid A solution of Example 18C (650 mg, 1.20 mmol) in 1: 1 methanol/THF (6 mL) was stirred with 2.4 M aqueous LiOH (1. 0 mL, 2.4 mmol) for 2. 5. h.

The solvent was removed-in vacuo ; and the residue was suspended in H2O (10 mL) and extracted with ethyl' acetate (10 mL). The aqueous phase was acidified by dropwise addition of 1M aqueous HC1, and then ex- tracted with ethyl acetate (10 mL), dried (Na2SO4), filtered, and concentrated to give Example 18D as an off-white solid (200 mg, 33%). 1H NMR'(DMSO-d6, 400 MHz) 5 12.43 (bs, 1H), 8.05 (d, J=7. 42 Hz, 1H), 7.87 (d, J=8. 59 Hz, 1H), 7.73 (d, J=15. 2 Hz, 1H), 7.65 (m, 2H), 7.4 (m, 2H), 7.26 (d, J=15.6 Hz, 1H), 6.66 (d, J=8.59 Hz, 1H), 4. 25 (m, 1H), 4.09 (m, 1H), 3. 82 (m,'lH), 3.23 (m, 1H), 3.10 (s, 2H), 2. 92 (m, 1H), 1.79 (m, 2H), 1.28 (m, 2H). LCMS (APCI+; Method F) m/z 511,513 (M+H) +; Rt=3.44 min (220 nm, 95.4 area%).

Example 19 N-(1-{3-[2,3-Dichloro-4-(4-fluoro-phenylsulfanyl)- phenyl]-acryloyl}-piperidin-4-yl)-2-fluoro- malonamic acid

Example 19A N-(1-Benzyl-piperidin-4-yl)-2-flùoro-malonamic acid ethyl ester The title compound was prepared according to the procedure for Example 18As except using di- ethyl fluoromalonate in place of diethyl malonate.

1H NMR (CDCl3, 400 MHz) 5 7. 23-7. 34 (m, 5H), 6.24 (bd, J=5.86 Hz, 1H), 5.22 (d, 2JHF=49. 2 Hz, 1H), 4.33 (m, 2H), 3.84 (m, 1H), 3.49 (s, 2H), 2. 82 (m, 2H), 2. 13 (m, 2H), 1.92 (m, 2H), 1.53 (m, 2H), 1.33 (m, 3H). LCMS (APCI+; Method F) m/z 323 (M+H) +; Rt=1.83 min (220 nm, 100 area%).

Example 19B 2-Fluoro-N-piperidin-4-yl-malonamic acid ethyl ester The title compound was prepared according to the procedure for Example 18B, except using the product of Example 19A in place of the product from 18A and the diethyl ether trituration was not per- formed. 1H NMR (CDC13, 400 MHz) 5 6. 235 (m, 1H), 5.23 (d, 2JHF =48.8 Hz, 1H), 4. 34 (m, 2H), 3.92 (m, 1H), 3.09 (m, 2H), 2.69 (m, 2H), 1.94 (m, 2H), 1.32-1. 42 (m, 5H). MS (ESI+) m/z 233 (M+H) +.

Example 19C N-(1-{3-[2,3-Dichloro-4-(4-fluoro-phenylsulfanyl)- phenyl]-acryloyl}-piperidin-4-yl)-2-fluoro- malonamic acid ethyl ester The title compound was prepared from 3- [2, 3-dichloro-4-(4-fluoro-phenylsulfanyl)-phenyl]- acrylic acid (98 mg, 0.287 mmol; synthesized in a similar manner to the acrylic acid used in Example 4A by methodology described in WO 00/59880, WO 00/39081 except that 4-fluorothiophenol was

substituted for 2-methoxythiophenol) and Example 19B (100 mg, 0.431 mmol) according to amide coupling protocol described for Example 1B, with the follow- ing purification step added. The product was puri- fied by flash chromatography eluting with 3% meth- anol in methylene chloride. Example 19C (87 mg, 85%) was obtained as a white foam. 1H NMR (DMSO-d6, 400 MHz} 5 8.56 (bd, J=7. 81 Hz, 1H), 7'. 87 (d, J=8. 59 Hz, 1H), 7. 73 (d, J=15.6 Hz, 1H), 7. 65' (m, 2H), 7. 38 (m, 2H), 7.27 (d, J=15.2 Hz, 1H), 6.66 (d, J=8. 20 Hz, 1H), 5.45 (d, 2JHF =47.6 Hz, 1H), 4@33 (m, 1H), 4.11-4. 25 (m, 3H), 3. 89 (m, 1H), 3. 20 (m, 1H), 2.85 (m, 1H), 1.72-1. 81 (m, 2H), 1.37 (m, 2H), 1.20 (t, J=7.03 Hz, 3H). LCMS (APCI+; Method F) m/z 557,559 (M+H) + ; Rt=3.88 min (220 nm, 86.1 area%).

Example 19D <BR> <BR> <BR> <BR> <BR> <BR> N- (l- {3- [2, 3-Dichloro-4- (4-fluoro-phenylsulfanyl)-<BR> <BR> <BR> <BR> phenyl]-acryloyl}-piperidin-4-yl)-2-fluoro- malonamic acid The title compound was prepared from Exam- ple 19C (75 mg, 0.14 mmol) according to saponifica- tion conditions described for Example 18D, except that the aqueous phase was not extracted with ethyl acetate before acidification with 1 M aqueous HC1.

The resulting precipitate was filtered, washed with

CH3CN (5 mL), and dried under vacuum to obtain Example 19D (50 mg, 70%) as a white solid. 1H NMR (DMSO-d6,400 MHz) 5 8.48 (d, J=7. 81 Hz, 1H), 7.87 (d, J=8.59 Hz, 1H), 7.73 (d, J=15.2 Hz, 1H), 7.66 (m, 2H), 7.40 (m,-2H) j 7. 27 (d, J=15.2 Hz, 1H), 6.66 (d, J=8. 98 Hz, 1H), 5.27 (d, 2JHF=48. 8 Hz, 1H), 4.32 (m, 1H), 4.14 (m, 1H), 3.88 (m, 1H), 3. 2-3.5 (m, 1H, buried under H20 peak), 2. 85 (m, 1H), 1.75 (m,. 2H), 1.36 (m, 2H). LCMS (APCI+ ; Method F with an alter- native gradient program : 0. 00min 75% Solvent C, 25% Solvent D ; 0.02min 75% Solvent C, 25% Solvent D ; 4.00 min 5% Solvent C ; 95% Solvent D; 4.30min 5% Solvent C, 95% Solvent D; 4.50min 75% Solvent C, 25% Solvent D ; 5. 50min 75% Solvent C, 25% Solvent D) m/z 529,532 (M+H) + ; Rt=3.01 min (220 nm, 100 area%).

Example 20 N-{1-[3-(6, 7-Dichloro-benzo [b] thiophen-5-yl)- acryloyl]-piperidin-4-yl}-(S)-3-methyl- succinamic acid

Example 20A (2, 3-Dichloro-phenylsulfanyl)-acetic acid A 2 L, 3-neck flask, with a mechanical stir shaft, was charged with 2, 3-dichlorothiophenol (50.0 g, 279 mmol) and bromoacetic acid (40.7 g, 293 mmol). The mixture was suspended in water (300 mL) and 3 M aqueous NaOH (300 mL, 900 mmol) was added.

A reflux condenser was attached and a thermocouple was placed in the remaining port. The mixture was heated to 95°C internal temperature. The mixture was refluxed 15 h and then cooled to room tempera- ture. Once the mixture had cooled, 4 M aqueous HC1 (400 mL, 1.6 mol) was added and the mixture extrac- ted with ethyl acetate (2x1. 25 L). The combined organic layers were dried (MgS04), filtered and con- centrated to an off-white solid. To the solids was added hexanes (150 mL) and the mixture heated to reflux with stirring for 15 min. The mixture was cooled to room temperature and collected by filtra- tion and washed with additional hexanes (100 mL).

The solid was dried under vacuum affording Example 20A (55.6 g, 84%) as a white solid. 1H NMR (acetone d6,400 MHz) 5 7.39-7. 43 (m, 2H), 7.31-7. 37 (m, 1H), 3.96 (s, 2H).

Example 20B (4-Bromo-2, 3-dichloro-phenylsulfanyl)- acetic acid To a solution of Example 20A (58. 0 g, 245 mmol) in acetic acid (250 mL) was added iron chips (3.5 g, 63 mmol). The mixture was gently heated to. dissolve Example 20A. The mixture was. cooled to room temperature and bromine (25 mL, 490, mmol) was added over 20 min by addition funnel. The reaction was stirred for 4 days, then more bromine (10 mL) was added in one portion and stirring continued for 5 h. Next, a third bolus of bromine (10 mL) was added and the mixture stirred 12 h. The, mixture was then quenched by careful addition of a saturated solution of NaHS04 and stirred for 45 min. A white precipitate was collected-by filtration and washed with water. The solid was dried under vacuum for two days in a desiccator followed by six hours, in a vacuum oven at 70°C, affording Example 20B (76.7 g, 99%) containing 90% bromination at the 4-position.

H NMR (acetone d6,400 MHz) 5 7.70 (d, J=8. 98 Hz, 1H), 7.35 (d, J=8.59 Hz, 1H), 4.00 (s, 2H).

Example 20C 5-Bromo-6,7-dichloro-2, 3-dihydro-benzo [b] thiophen-3-ol To a suspension'of Example 20B (50.0 g, 158 mmol) in THF (160 mL) was added 3 drops of DMF followed by oxalyl chloride (21. 1 g, 166.1 mmol) over a 15 min period. The mixture was heated to 50°C for 3 h. Next, the mixture was cooled to room temperature and concentrated in vacuo to obtain an intermediate acid chloride (51. 3'g) as a dark yellow oil that was used in the next step without purifica- tion. A 1 L flask was charged with A1C13 (179-'g, 1.35 mol) and methylene chloride (250 mL), and then cooled to-78°C (dry ice/acetone). A solution of the acid chloride (prepared above) in methylene chloride (150 mL) was added dropwise over 20 min.

When addition was complete the dry ice bath was removed and'the reaction was allowed to warm to room temperature and stir for an additional 1 h. The mixture was quenched by slowly pouring onto ice (2 L). The mixture was extracted with ethyl acetate (2x1 L), and the combined organic phases were washed with brine (500 mL). The red organic layer was dried (MgSO4), filtered and concentrated to an orange solid. The solid was dried under high vacuum for 14 h to give an intermediate ketone, which was used in the next step without purification. The ketone was

resuspended in ethanol (500 mL, absolute) and NaBH4 (12 g, 316 mmol) was added portionwise while cooling on an ice bath. The reaction was warmed to room temperature after addition was complete. After 2 h, the reaction was transferred to a 4 L Erlenmeyer flask and quenched with 4 M aqueous HC1 (caution, vigorous gas evolution). The mixture was concen- trated in vacuo, diluted with water (200 mL), and extracted with ethyl acetate (2x500 mL). The combined organic phases were washed with brine (250 mL), dried (MgSO4), filtered and concentrated to give a dark brown oil (45 g). Purification by chromatog- raphy on silica gel (Biotage Flash 75 L, 10% ethyl acetate in hexanes followed by 20% ethyl acetate in hexanes) afforded Example 20C (23. 0g, 48% for 3 steps) as a red oil that solidified upon standing.

H NMR (CDC13, 400 MHz) 5 7.53 (d, J=0.78 Hz, 1H), 5.44 (m, 1H), 3.66 (dd, J=12.1, 6.63 Hz, 1H), 3.35 (dd, J=12.1, 4.68 Hz, 1H), 2. 23 (d, J=8.19 Hz, 1H).

Example 20D 5-Bromo-6, 7-dichloro-benzo [b] thiophene To a 1 L flask was added Example 20C (23. 0g, 76.7 mmol) and acetic acid (230 mL). The mixture was sonicated for 30 min. BF3-Et2O (15 mL, 115 mmol) was added dropwise over 1 min at room

temperature. After 15 min the homogeneous mixture was diluted with water (500 mL) and a white precipi- tate formed. After stirring for an additional 15 min, the solid was collected by filtration and washed with water (200 mL). The solid was dissolved in ethyl acetate (500 mL) and washed with a satur- ated solution of NaHCO3 (200 mL) followed by brine (200 mL). The organic phase was dried (MgSO4), filtered and concentrated. The resulting pink solid was recrystallized from boiling hexanes (200 mL).

The crystals (14.3 g) that formed had a dark red color. The mother liquor ; was concentrated to give a tan solid (6.6 g) which was combined with the first crop to yield Example 20D (20. 9 g, 97%). 1H NMR (CDC13,400 MHz) 5. 7.97 (s, 1H), 7.51 (d, J=5. 47 Hz,.

1H), 7.24 (d, J=5. 47 Hz, 1H).

Example 20E 3- (6, 7-Dichloro-benzo [b] thiophen- 5-yl) -acrylic acid methyl ester A 250 mL round bottom flask was charged with Example 20D (9.80 g, 35.0 mmol), Pd2 (dba) 3 (796 mg, 0.869 mmol), and o- (Tol) 3P (809 mg, 2.66 mmol).

The flask was purged with N2 (3x),. and then charged with anhydrous DMF (70 mL), methyl acrylate (8.98 g, 104 mmol) and triethylamine (14.5 mL, 104 mmol).

The reaction was purged with N2 (2x) again. The mixture was stirred under N2 at 100°C for 16h. The reaction was cooled to room temperature and a thick precipitate formed. The suspension was partitioned between methylene chloride (300 mL) and water (100 mL). The phases were and the organic phase was washed with 0.5 N aqueous HC1 (4x100 mL).

The organic phase was dried (Na2SO4), filtered and concentrated to obtain a beige fluffy solid (11.2 g). The crude product was purified by partial dissolution (digestion) in hot heptanes followed by cooling to room temperature and. filtration to give Example 20E (8.22g, 82%). 1H NMR (CDC13, 400 MHz) 5 8.18 (d, J=16.0 Hz, 1H), 7.97 (s, 1H), 7.53 (d, J=5.47 Hz, 1H), 7.36 (d, J=5.47 Hz, 1H), 6. 48. (d, J=15. 6 Hz, 1H), 3.85 (s, 3H).

Example 20F 3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)- acrylic acid The title compound was prepared from Example 20E (8.22 g, 28. 6 mmol) according to the procedure described for Example 18D. Example 20F was obtained as an off-white powder (6.60 g, 84%) after recrystallization from hot toluene. 1H NMR (10% DMSO-d6 in CDC13, 400 MHz) 5 8.12 (d, J=16. 4 Hz, 1H), 8.06 (s, 1H), 7.60 (d, J=5.47 Hz, 1H), 7.41 (d, J=5. 47 Hz, 1H), 6. 49 (d, J=16. 0 Hz, 1H).

Example 20G {1- [3- (6, 7-Dichloro-benzo [b] thiophen- 5-yl)-acryloyl]-piperidin-4-yl}-carbamic acid tert-butyl ester Example 20F (2.00 g, 7.32 mmol) was re- acted with (2.20 g, 11.0 mmol) according to the amide coupling protocol de- scribed for Example 1B. Example 20G was obtained as an orange-colored solid (3.34 g, 100%) which was carried forward without purification. 1H NMR (CDCl3, 400 MHz) 5 8.08 (d, J=15. 6 Hz, 1H), 7.94 (s, 1H), 7.52 (d, J=5.47 Hz, 1H), 7.35 (d, J=5. Hz, 1H), 6.90 (d, J=15.2 Hz, 1H), 4.65 (m, 1H), 4. 49 (m, 1H), 4. 06 (m, 1H), 3.73 (m, 1H), 3.26 (m, 1H), 2.89 (m, 1H), 2.05 (m, 2H), 1.46 (s, 9H), 1.40 (m, 2H). LCMS (APCI+; Method, F) m/z 455, 457 (M+H) + ; Rt=3.77 min (220 nm, 100 area%).

Example 20H 1-(4-Amino-piperidin-1-yl)-3-(6, 7-dichloro- benzo [b] thiophen-5-yl)-propenone The Boc group was removed from Example 20G (4.52 g, 9.93 mmol) according to.. the procedure de- scribed for Example 1B. Example : 20H was obtained as a waxy, crème-colored solid (3. 10 g, 88%). 1H NMR (DMSO-d6, 400 MHz) õ 8.55 (s, 1H), 7.98 (d, J=5.46 Hz, 1H), 7.89 (d, J=15. 2 Hz, 1H), 7.56 (d, J=5.46 Hz, 1H), 7.42 (d, J=15.2 Hz, 1H), 4.26 (dd, J=40.9, 13.3 Hz, 2H), 3. 20 (t, J=11. 1 Hz, 1H), 2. 86 (m, 2H), 2.15 (bs, 2H), 1. 78 (m, 2H), 1.18 (m, 2H). LCMS (ESI+ ; Method H) m/z 355, 357 (M+H) + ; Rt=2.42 min (220 nm, 100 area%).

Example 20I <BR> <BR> <BR> <BR> N- {1- [3- (6, 7-Dichloro-benzo [b] thiophen- 5-yl)-acryloyl]-piperidin-4-yl}-3- methyl-succinamic acid (1: 1 regioisomeric mixture of 2-methyl and 3-methyl- succinamic acids)

A solution of Example 20H (312 mg, 0.88 mmol) in methylene chloride (10 mL) was stirred with DIEA (230 pL, 1.32 mmol) and (S) -methyl succinic anhydride (100 mg, 0.88 mmol ; prepared from (S)-2- methyl succinic acid according to the literature: Davies, S. G. ; Dixon, D. J. J. Chem. Soc., Perkin Trans. 1 1998,2635-2643) under N2 for 1 h. The reaction was diluted with methylene chloride (20 mL) and methanol (5 mL) and washed with 1N aqueous HC1 (10 mL). The aqueous phase was back-extracted with 10% methanol in methylene chloride (2x20 mL). The combined organic phases were washed with brine (20 mL), dried (Na2SO4), filtered, and concentrated to obtain a waxy solid (412 mg, 100%) which contained a 1: 1 mixture of Example 20N and its regioisomer Example 21 as analyzed by chiral HPLC (Method D): Example 20N: Rt=14.2 min (50.2 area%) and Example 21: Rt=19.8 min (49.8 area%). LCMS (ESI- ; Method H) m/z 467,469 (M-H)- ; Rt=2.40 min (220 nm, 100 area% ; regioisomers not separated).

Example 20J <BR> <BR> N- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl]-piperidin- 4-yl}- (S)-3-methyl-succinamic acid 9H-fluoren-9- ylmethyl ester

A solution of the 1: 1 regioisomeric mix- ture of carboxylic acids from Example 20I (412 mg, 0.88 mmol) in methylene chloride (20 mL) was stirred with DMAP (21 mg, 0.18 mmol) and (9H-fluoren-9-yl)- methanol (258 mg, 1.32 mmol). The reaction was cooled to 0°C under N2 and dicyclohexylcarbodiimide (272 mg, 1.32 mmol) was added. The reaction was stirred for 16 h at rt under N2. The resulting sus- pension was filtered, rinsing the solid. with methyl- ene chloride. The filtrate was concentrated and the. regioisomers. were separated by Biotage Flash 40 using the following solvent gradient: 3: 1 EtOAc/- hexanes (500 mL), 9 : 1 EtOAc/hexanes (500 mL), and neat EtOAc (1 L). Mixed fractions were rechromato- graphed using a similar elution'gradient. Example 20J (152 mg, 53% of theoretical recovery) was ob- tained as a white powder. 1H NMR (DMSO-d6, 400 MHz) 5 8.55 (d, J=3. 12 Hz, 1H), 7. 98 (d, J=5. 46 Hz, 1H), 7.90 (m, 4H), 7.67 (d, =7. 02 Hz, 2H), 7.55 (m, 1H), <BR> <BR> <BR> <BR> 7. (m, 3HJa 7. 34 (m, 2H), 4.32 (m, 3H), 4.24 (m, 2H), 3.83 (m, 1H), 3.26 (m, 1H), 2.89 (m, 1H), 2.67 (m, 2H), 2.36 (m, 1H), 1.77 (m, 2H), 1.29 (m, 2H), 1.02 (d, J=4.68 Hz, 3H).

Example 20K 4-Benzyl-3-propionyl-oxazolidin-2-one

Adapted from Levy, D. E. et al. , J. Med.

Chem. 1998, 41, 199-223. To an oven-dried 2L flask under N2 was added (R)-4-benzyl-oxazolidin-2-one (30.0 g, 169. 3 mmol) and THF (300mL, 99. 9% anhy- drous) via canula.-The flask was cooled to-78°C (dry ice/acetone). n-BuLi (70 mL, 169 mmol, 2.41 M in hexanes) was added dropwise over a 30 min period.

After the addition was complete, the'mixture was stirred for 30 min and propionyl chloride (17.2 g, 186 mmol) was added dropwise. Stirring was con- tinued for lsh at-78°C. The reaction was allowed to warm to room temperature over 1 h and quenched by pouring into'a 1 L separatory funnel containing sat- urated NH4Cl' (300 mL). The mixture was'extracted with ethyl acetate (600 mL). The organic phase was washed with saturated NaHCO3 (200 mL) and brine (200 mL). The organic phase was dried (Na2S04), filtered and concentrated to give Example 20K (42.2 g, 107%) as a light-colored oil. The crude product was' carried forward without purification.' /o I.. CN I /O a 0 Example 20L (S)-4- (4-Benzyl-2-oxo-oxazolidin-3-yl)- 3-methyl-4-oxo-butyric acid tert-butyl ester To an oven-dried 1 L flask under N2 was added THF (250 mL, 99. 9% anhydrous) and diisopropyl-

amine (14.3 g, 141 mmol). The flask was cooled to 0°C (ice bath) and n-BuLi (56 mL, 141 mmol, 2.54 M in hexanes) was added dropwise. The flask was cooled to-78°C (dry ice/acetone bath). Example 20K (30.0 g, 128.6 mmol, dissolved in 40 mL anhydrous THF) was added dropwise over 30 min. The yellow solution was, stirred at-78°C for 2 h. Bromo-acetic acid tert-butyl ester (67.7 g, 347 mmol) was added dropwise over 20 min. Stirring was continued for 30min at-78°C. The mixture was warmed to 0°C by placing the flask in an ice water bath for 20 min.

The-mixture was quenched by pouring into a 1 L separatory funnel containing saturated NH4Cl (200 mL). The mixture was extracted with EtOAc (300 mL), the phases were separated and the organic was washed. with 1N aqueous HCl. (100 mL), saturated NaHCO3 (100 mL), and brine (100 ; mL). The organic was dried (Na2SO4), filtered, and concentrated. The crude product was purified using a Biotage Flash 65 column. The column was wetted with solvent (9: 1 hexane/MTBE ; 1 L) and wet-mounted with the crude (dissolved in 50 mL DCM ; two columns were performed to purify all of the product). The title compound was obtained as a white, crystalline solid (29.1 g, 65%), and was pure by HPLC, LCMS and 1H NMR.

Example 20M (S) -2-Methyl-succinic acid 4-tert-butyl ester To a 1 L flask was added Example 20L (29.0 g, 83. 5 mmol) and a solution of 4: 1 THF/water (300 mL). The mixture was cooled to 0°C (ice water bath)' and 30% aqueous H2O2 (17. 0 g, 501 mmol) was added dropwise over 10 min while stirring. An aqueous solution'of'LiOH (2"M, 4.73 g, 117 mmol) was added, and the reaction was stirred for'4 h in'the ice bath. The reaction was quenched by careful addition of Na2SO3 (2.73 M, 14. 7 g, 117 mmol). The THF was removed in vacuo and the resulting suspension was extracted with DCM (400 mL). The aqueous phase was reserved (contains product) and the DCM phase was washed with 0.1 N aqueous NaOH (100 DCM was dried (Na2SO4), filtered and concentrated ín vacuo to afford the chiral auxiliary (reusable).

The aqueous phase was cooled in an ice bath and acidified with 2 N'aqueous HCl to pH 4. The cloudy solution was extracted with EtOAc (300mL), dried (Na2SO4), filtered and concentrated to give the product as a colorless oil, which solidified to a white solid upon standing. To remove trace amounts of the auxiliary (which alters the optical rotation) the product was purified using a Biotage Flash 40 column, eluting with 30% EtOAc in hexanes. The title compound (13.4 g, 85%) is less polar than the

auxiliary. [a] D=-6. 56° (c=0.904, CHC13) ; literature: [a] D=-7. 0° (c=0.86, CHC13) ; Davies, S. J. Chem. Soc., Perkin Trans 1 : Org. and Biorg. Chem. 1998, 17, 2635-2644. 1H NMR (CDC13, 400 MHz) 5 11.73 (bs, 1H), 2. 91 (m, 1H), 2.64 (dd, J=16. 4,8. 91 Hz, 1H), 2.36 (dd, J=16.4, 5.85 Hz, 1H), 1.44 (s, 9H), 1.24 (d, J=7.4 Hz, 3H).

Example 20N N- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5- yl)-acryloyl]-piperidin-4-yl}-(S)-3- methyl-succinamic acid A solution of Example 20J (150 mg, 0.23 mmol) in methylene chloride (5 was shaken'with piperidine (115 uL, 1. 16 mmol} for 16 at RPM.

The solution was washed with 1N aqueous HC1 (2x10 mL), dried (Na2SO4), filtered, and concentrated. The crude product was purified by Biotage Flash 40 elut- ng with 10% methanol in methylene chloride (200 mL) to elute the 9H-fluoren-9-yl by-products, followed by 40% methanol in methylene chloride (800 mL) to elute the title compound 20N (69 mg, 63%), which was obtained as a white powder in 98% regioisomeric pur- ity. Alternatively the title compound can be made from Example 20H and Example 20M as described in Example 1B followed by deprotection of the tert-

butyl ester with TFA according to the method de- scribed in Example 23E. 1H NMR (DMSO-d6, 400 MHz) 5 8.56 (s, 1H), 8.12 (d, J=6.63 Hz, 1H), 7.98 (d, J=5.07 Hz, 1H), 7.89 (d, J=15.2 Hz, 1H), 7.55 (d, J=5.46 Hz, 1H), 7.42 (d, J=15. 2 Hz, 1H), 4.24 (dd, J=28.9, 12. 9 Hz, 2H), 3. 82 (m, 1H), 3.29 (m, 1H), 2.95 (m, 1H), 2. 63. (m, 1H), 2.33 (dd, J=15.6, 7.41 Hz, 1H), 2.07 (m, 1H), 1.78 (m, 2H), 1.33 (m, 2H), 1.00 (d, J=7.02 Hz, 3H). Regiochemical assignment was based on correlation of COSY, HMBC, HSQC and TOCSY multidimensional NMR experiments. LCMS (ESI- ; Method H) m/z 467, 469 (M-H)- ; Rt=2. 40 min (220 nm, 100% area). Enantiomeric purity (>98%) and regio- isomeric purity (98%) were determined by chiral HPLC (Method D): Example 20N: Rt=14. 0 min 1 area%) and the regioisomer, Example 21 :. Rt=19. 7 min (1.9 area%) ; chiral HPLC analysis (Method D) of a mixture of the enantiomers, Example 22, gave distinguishable retention times: Rt=12. 6'min (50.6 area%), Rt=16.1 min (49.4 area%). The"R"enantiomer was not de- tected (<2%) by chiral HPLC of Example 20K.

Example 21 N- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)- acryloyl]-piperidin-4-yl}-(S)-2-methyl- succinamic acid

Example 21 (regioisomer of Example 20N) was prepared in greater than 90% regioisomeric purity according to the same reaction protocol described for Example 20N. lH NMR (DMSO-d6) 5 8.52 (s, 1H), 7.98 (d, J=7. 41 Hz, 1H), 7.92 (d, J=15.2 Hz, 1H), 7.92 (d, J=5. 46 Hz, 1H), 7.53 (d, J=5. 07 Hz, 1H), 7.39 (d, J=15. 2 Hz, 1H), 4.26 (dd, J=45.6, 12.5 Hz, 2H), 3.87 (m, 1H), 3.3 (m, 1H ; overlaps with water), 2.94 (m, 1H), 2.68 (m, 1H), 2.44 (dd, J-14. 8,7. 02 Hz, 1H), 2.12 (dd, J=14. 4, 7.41 Hz, 1H), 1.82 (m, 2H), 1.35 (m, 2H),'1. 06 (d J=7. 02 Hz, 3H). Regiochemical assignment was based on correla-. tion of COSY, HMBC, HSQC and TOCSY multidimensional NMR experiments. LCMS (APCI+ ; Method F). m/z 469, 471 (M+H) + ; Rt=2.98 min (220 nm ; 100 area%).

Enantiomeric purity (>98%) and regioisomeric purity (91%) were determined by chiral HPLC (Method D) : Example 20N: Rt=13.9 min (8. 9 area%) and Example 21: Rt=19.2 min (91.1 area%) ; chiral HPLC analysis (Method D) of a mixture of the enantiomers, Example 22, gave distinguishable retention times: Rt=12. 6 min (50.6 area%), Rt=16. 1 min (49.4 area%). The"R" enantiomer of Example 21 was not detected (<2%) by chiral HPLC.

Example 22 N- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5- yl)-acryloyl]-piperidin-4-yl}-(R)-2-methyl- succinamic acid (1: 1 regioisomeric mixture of 2-methyl and 3-methyl-succinamic acids) The title compound was prepared as a 1: 1 regioisomeric mixture from Example 20H (89 mg, 0.25 mmol) and (R) -methyl succinic anhydride (29 mg, 0.25 mmol; prepared from (R) -2-methyl succinic acid according to the literature: Davies, G. ; Dixon, D. J. J. Chem. Soc., Perkin Trans. 1 1998, 2635- 2643) as described in the synthesis of Example 20I.

Example 22 (105 mg, 89%) was obtained as a beige solid. 1H NMR (DMSO-d6, 400 MHz, 1: 1 mixture of regioisomers) 5 12.1 (bs, 2H), 8.56 (s, 2H), 7.98 (d, J=5.46 Hz, 2H), 7.82-7. 92 (m, 4H), 7.56 (d, J=5.46 Hz, 2H), 7.43 (d, J=15.2 Hz, 2H), 4.26 (m, 4H), 3.84 (m, 2H), 3.28 (m, 2H), 2.92 (m, 2H), 2.60- 2.74 (m, 2H), 2.39-2. 48 (m, 2H), 2.11-2. 23 (m, 2H), 1.79 (m, 4H), 1.31 (m, 4H), 1.06 (d, J=7.02 Hz, 3H; 2-methyl succinamic acid regioisomer), 1.02 (d, J=7.02 Hz, 3H; 3-methyl succinamic acid regioiso- mer). Regiochemical assignment was based on cor- relation of COSY, HMBC, HSQC and TOCSY multidimen- sional NMR experiments. Enantiomeric purity (>98%) was verified by chiral HPLC (Method D): Example 22: Rt=12.6 min (50.6 area%), Rt=16. 1 min (49.4 area%).

The enantiomers gave distinguishable retention times, Example 20N: Rt=14.2 min (50.2 area%) and Example 21: Rt=19.8 min (49.8 area%). LCMS (APCI+; Method F) m/z 469,471 (M+H) + ; Rt=2.99 min (220 nm, 100 area% ; regioisomers not separated).

TFA Example 23 rac-3-Aminomethyl-N-{1-[3-(6, 7-dichloro- benzo [b] thiophen-5-yl)-acryloyl]-piperidin- 4-yl} -succinamic acid trifluoroacetic acid salt

Example 23A 3-tert-Butoxycarbonylamino- propionic acid methyl ester To a stirred solution of (3-alanine methyl ester hydrochloride (14.0 g, 100 mmol) and diiso- propylethylamine (44 mL, 250 mmol) in methylene chloride (500 mL) at 0°C (ice bath) was added di- tert-butyl dicarbonate (21.8 mL, 95 mmol) dropwise.

The reaction was removed from the ice bath, and allowed to warm to room temperature (gas evolution

observed, vent adequately). The'reaction was stirred for 16 h. The mixture was washed with water (200 mL), 1N aqueous HC1 (2x150 mL; much gas evo- lution, vent adequately), and a saturated solution of NaHCO3 (200 mL)-. The organic phase was dried (Na2SO4), filtered, and concentrated to obtain Exam- ple 23A as a pale yellow oil (20.0 g, 98. 5%). Exam- ple 23A has also been prepared by esterification of Boc-ß-alanine with methanol (Hayashida, Osamu, et al, J. Org. Chem.'2002, 67, 8291-8298) ; 1H NMR (CDC13, 400 MHz) 5 5. 06 (m, 1H), 3. 70 (s, 3H), 3. 40 (m, 2H), 2.53 (m, 2H), 1.44 (s, 9H).

Example 23B rac-2-(ter-Butoxycarbonylamino- methyl) -succinic acid 4-tert-butyl ester 1-methyl ester To a stirred solution of diisopropylamine (16 mL, 115 mmol) in THF (500 mL, 99. 9% anhydrous) at 0°C (ice bath). under N2 was added n-butyllithium (2.56 M in hexanes, 45 mL, 115 mmol) dropwise. The solution was cooled to-78°C (dry ice/acetone) and Example 23A (10.2 g, 50. 0 mmol) was added dropwise.

The yellow suspension was stirred at-78°C for 3 h. tert-Butyl bromoacetate (12.7 g, 65.0 mmol) was then added dropwise. The reaction was stirred for 5 min,

and then removed from the-78°C bath and immediately submerged in an ice water bath. The mixture was stirred for 10 min (note: leaving the reaction for longer periods of time results in poor yields), and then poured into-a saturated solution of NH4C1 (300 mL). The mixture was shaken, and then diluted with ethyl acetate (700 mL). The phases were shaken again, and then separated. The organic phase was washed with 1N aqueous HC1 (2x100 mL), a. saturated solution of NaHCO3 (200 mL), and brine (-200 mL). The organic phase was dried (Na2SO4), filtered, and con- centrated. The crude product (14.0 g) was purified using a Biotage Flash 75 chromatography system, eluting with the following gradient: 10% ethyl ace- tate in hexanes followed by 15% ethyl acetate in hexanes. Example 23B was obtained as a colorless oil (6.67 g, 42%). 1H NMR'(CDC13, 400 MHz) 5 4.93 (m, 1H), 3.72 (s, 3H), 3.38 (m, 2H), 2.96 (m, 1H), 2.63 (dd, J=16.8, 7. 80-Hz, 1H), 2.50 (dd, J=16.8, 5.85 Hz, 1H), 1. 39-1. 54 (m, 18H). LCMS (ESI+ ; Method H) m/z 340 (M+Na) + ; Rt=3.18 min (220 nm, 100 area%).

Example 23C rac-2- (tert-Butoxycarbonylamino- methyl) -succinic acid 4-tert-butyl ester

The title compound was prepared from Exam- ple 23B (7.56 g, 23.8 mmol) according to the proce- dure described for Example 18D, except the amount of lithium hydroxide hydrate was reduced (1. 50 g, 35.7 mmol) to 1.5 equivalents. Example 23C (6. 78 g, 94%) was obtained as a pale yellow oil, which solidified upon standing. 1H NMR (CDCl3, 400 MHz) # 10.61 (bs, 1H), 5.04 (m, 1H), 3.41 (m, 2H), 2.98 (m, 1H), 2.65 (dd, J=16.8, 7. 41 Hz, 1H), 2. 38-2. 57 (m, 1H), 1. 37- 1.54 (m, 18H). LCMS (ESI ;. Method H) m/z 302 (M-H)-; Rt=2.07 min.

Example 23D rac-3- (tert-Butoxycarbonylamino-methyl)-N- {1- [3-(6, 7-dichloro-benzo [b] thiophen-5-yl)- acryloyl]-piperidin-4-yl}-succinamic acid tert-butyl ester The title compound was prepared from Exam- ple 23C (910 mg, 3.00 mmol) and Example 20H (888 mg, 2.50 mmol) according to the amide coupling procedure described for Example 1B. Example 23D (1.03 g, 64%) was obtained as a white powder after purification using a Biotage Flash 40 chromatography system, eluting with the following gradient: 1: 1 ethyl acetate/hexanes, 3: 1 ethyl acetate/hexanes, then 9: 1

ethyl acetate/hexanes. IH NMR (CDC13, 400 MHz) 5 8.56 (bs, 1H), 7.98 (d, J=5. 46 Hz, 1H), 7.86-7. 94 (m, 2H), 7.55 (d, J=5. 07 Hz, 1H), 7.44 (d, J=15.2 Hz, 1H), 6.77 (m, 1H), 4.29 (m, 2H), 3.85 (m, 1H), 3.28 (m, 1H), 3.10 (m, 1H), 2.89-2. 98 (m, 2H),. 2.68 (m, 1H), 2.43 (dd, J=16. 0, 9.75 Hz, 1H) ; 2.27 (dd, J=16. 0,4. 68 Hz, lH), 1.78 (m, 2H), 1. 27-1. 45 (m, 20H). LCMS (ESI+ ; Method I) m/z 662, 664 (M+Na) +; Rt=4.16 min (220 nm, 100 area%).

TFA Example 23E rac-3-Aminomethyl-N- 1- [3- (6, 7-dichloro-benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}- succinamic acid trifluoroacetic acid salt A solution of Example 23D (1.00 g, 1.56 mmol) in methylene chloride (20 mL) was stirred with trifluoroacetic acid (2.4 mL, 31 mmol) under N2 for 16 h at room temperature. The reaction was then concentrated in vacuo. The residue was resuspended in methylene chloride (10 mL), and the solvent was concentrated again. This process was repeated (3x) in order to remove residual trifluoroacetic acid.

The crude product was triturated with diethyl ether (10 mL), and the resulting white solid was filtered.

Example 23E (890 mg, 95%) was obtained as a white powder. 1H NMR (DMSO-d6, 400 MHz) 5 12.5 (bs, 1H), 8.56 (s, 1H), 8.15 (d, J=7.80 Hz, 1H), 7.99 (d, J=5.07 Hz, 1H), 7.77-7. 95 (m, 4H), 7. 56^ (d, J=5.46 Hz, 1H), 7.44 (d, J=15. 2 Hz., 1H), 4.25 (m, 2H), 3.89 (m, 1H), 3.32 (m, 1H), 2.84-3. 11 (m, 4H), 2.58 (m, 1H, overlaps DMSO), 1. 82 (m, 2H), 1.37 (m, 2H).

LCMS (ESI- ; Method I) m/z 482,484 (M-H)- ; Rt=2.64 min (220 nm, 100 area%).

Example 24 rac-3- (Benzoylamino-methyL)-N- {1- [3- (6, 7-dichloro- benzo [b] thiophen-5-yl)-acryloyl]-piperidin-4-yl}- succinamic acid To a stirred suspension of Example 23E (300 mg, 0.50 mmol) and diisopropylethylamine (0.35 mL, 2.0 mmol) in methylene chloride (10 mL) at 0°C (ice bath} was added benzoyl chloride (0.24 M solu- tion in chloroform, 2.1 mL, 0.50 mmol) dropwise.

After addition was complete, the reaction was stirred for 30 min at 0°C. The reaction was then quenched by addition of 1 N aqueous HC1 (5 mL). The heterogeneous mixture was extracted with 20% meth-

anol in methylene chloride (30 mL). The organic phase was washed with 1N aqueous HC1 (3x20 mL).

Before each aqueous extraction, methanol (5 mL) had to be added to maintain two homogeneous phases. The organic phase was dried (Na2SO4), filtered, and con- centrated. crude product was purified by par- tial dissolution (or digestion) in hot acetonitrile (10 mL) followed by cooling to room temperature.

The resulting white solid. was filtered, washing with acetonitrile (3x2 mL). A second digestion, in ethyl acetate, was performed to raise the purity above 95%. The powder was dried (CaS04 dessicator) under high vacuum for 2 days. Example 24 (135 mg, 46%) was obtained as a white powder. 1H NMR (DMSO-d6, 400 MHz) 5 12.1 (bs, 1H), 8.55 (s, 1H), 8.43 (m, 1H), 7. 93-8. 00 (m, 2H), 7.89 (d, J=15.2 Hz, 1H), 7.80- 7.85 (m, 2H), 7.56 (d, J=5.07 Hz, 1H), 7.37-7. 53 (m, 4H), 4.14-4. 33 (m, 2H), 3.86 (m, 1H), 3.21-3. 45 (m, 3H, overlaps with water), 2.90 (m, 2H), 2.55 (m, 1H, overlaps with DMSO), 2. 40 (dd, J=16.4, 5.07 Hz, 1H), 1.77 (m, 2H), 1. 31 (m, 2H). LCMS (ESI+; Method I) m/z 588,590 (M+H) +, 610,612 (M+Na) +; Rt=2.80 min (220 nm, 100 area% ; 254 nm, 96.9 area%).

Example 25 rac-N-{1-[3-(6,7-Dichloro-benzo[b]thiophen-5-yl)- <BR> <BR> acryloyl]-piperidin-4-yl}-3- [ (3-methoxy-<BR> <BR> <BR> benzoylamino)-methyl]-succinamic acid The title compound was prepared from : Example 23E (150 mg, 0.25 mmol) and 3-methoxybenzoyl chloride (0.24 M solution in chloroform, 1. 0 mL, 0.24 mmol) according to the procedure described by Example 24. Example 25 (99 mg, 64%) was obtained as a white powder after purification. by partial dis- solution (or digestion) in hot acetonitrile (10 mL) followed by cooling to room temperature and filtra- tion. 1H NMR, (DMSO-d6,400 MHz} 5 12.1 (bs, 1H), 8. 55 (s, 1H), 8.40 (m, 1H), 7. 98 (d, J=5. 46 Hz, 1H), 7.94 (d, J=7.80 Hz, 1H), 7.89 (d, J=15.2 Hz, 1H), 7.56 (d, J=5.46 Hz, 1H), 7.30-7. 46 (m, 4H), 7.07 (m, 1H), 4.13-4. 30 (m, 2H), 3.86 (m, 1H), 3.78 (m, 3H, 3-methoxy group split into two singlets due to hindered rotation), 3. 31-3. 32 (m, 3H, overlaps water)', 2.89 (m, 2H), 2.55 (m, 1H, overlaps DMSO), 2.38 (dd, J=16.4 Hz, 5.07 Hz, 1H),'1. 77 (m, 2H), 1.32 (m, 2H). LCMS (ESI- ; Method I) m/z 616,618 (M-H)- ; Rt=2.86 min (220 nm, 96.4 area%).

Example 26 rac-3-[(3-Chloro-benzoylamino)-methyl]-N-{1- [3- (6, 7-dichloro-benzo [b] thiophen-5-yl)- acryloyl]-piperidin-4-yl}-succinamic acid The title compound was prepared from Example 23E (150 mg, 0.25 mmol) and 3-Chlorobenzoyl chloride (0. 24 M solution in chloroform, 1.0 mL, 0.24 mmol) according to the procedure described'by Example 24. Example 26 (93 mg, 60%) was obtained as a white powder after purification by partial dis- solution (or digestion) in hot acetonitrile (10 mL) followed by cooling to room temperature and filtra- tion. 1H NMR (DMSO-d6, 400 MHz) 5 8.96 (m, 1H), 8.52 (s, 1H), 8.13 (m, 1H), 7.75-8. 00 (m, 4H), 7.33-7. 57 (m, 4H), 4.12-4. 29 (m, 2H), 3.85 (m, 1H), 3.20-3. 47 (m, 3H, overlaps water), 2.92 (m, 2H), 2.39 (m, 1H), 2.26 (m, 1H), 1.76 (m, 2H), 1.31 (m, 2H)'. LCMS (ESI- ; Method I) m/z 622,624 (M-H)- ; Rt=2.95 min (220 nm, 1GO area% ; 254 nm, 93.9 area%).

Example 27 rac-N- {l- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)- acryloyl]-piperidin-4-yl}-3- [ (3-plienyl-ureido)- methyl] -succinamic acid The title compound was prepared from Exam- ple 23E (150 mg, 0.25 mmol) and phenyl isocyanate (0.24 M solution in chloroform, 1. 0 0.24 mmol) according to the procedure described by Example 24, except the amount of diisopropylethylamine (0.11 mL, 0.63 mmol) was reduced to 2.5 equivalents. Example 27 (89 mg, 59%) was obtained as a white powder after purification by partial dissolution (or digestion) in hot acetonitrile (10 mL) followed by cooling to room temperature and filtration. 1H NMR (DMSO-d6, 400 MHz) 5 12.1 (bs, 1H), 8.50-8. 62 (m, 2H), 7.96- 8.06 (m, 2H), 7.90 (d, J=15.2 Hz, 1H), 7.56 (d, J=5. 46 Hz, 1H), 7.35-7. 47 (m, 3H), 7.15-7. 26 (m, 2H), 6.87 (m, 1H), 6.11 (m, 1H), 4.27 (m, 2H), 3.87

(m, 1H), 3.18-3. 32 (m, 3H, overlaps water), 2.94 (m, 1H), 2.77 (m, 1H), 2.45-2. 56 (m, 1H, overlaps DMSO), 2.33 (dd, J=16.4, 5.85 Hz, 1H), 1.80 (m, 2H), 1. 36 (m, 2H). LCMS (ESI- ; Method I) m/z 601,603 (M-H)- ; Rt=2.87 min (220-nm,-100 area%).

Example 28 N- {1- [3- (2, 3-Dichloro-4-methylsulfanyl- phenyl)-acryloyl]-azetidin-3-yl}-3- methyl-succinacmic acid

Example 28A l-Bromo-2, 3-dichloro-4-methylsulfanyl-benzene Dimethyl sulfate (2.90 mL, 30.7 mmol) was added dropwise to a stirred solution of NaOH (1.28 g, 32.1 mmol) and 2,3-dichlorobenzenethiol (5.00 g, 27.9 mmol) in water (50 mL) at 0°C. The reaction mixture was heated at 110°C for 2 h and allowed to cool to ambient temperature. The reaction mixture was extracted with ether and the organic layer was washed with water and brine, dried (MgS04), and filtered. Evaporation of the solvent followed by

recrystallization from hexane gave the title com- pound (3.89 g, 72%) as a white solid. 1H NMR (CDC13, 400 MHz) 5 7.24 (dd, J=8.2 Hz, J=1.6 Hz, 1H), 7.18 (t, J=8. 2 Hz, 1H), 7. 04 (dd, J=8.2 Hz, J=1. 6 Hz, 1H), 2.48 (s, 3H).

Example 28B l-Bromo-2, 3-dichloro-4-methylsulfanyl-benzene Bromine (0.79 mL, 15 mmol) was added drop- wise to a stirred solution of Example 28A (2. 97 g, 15.4 mmol) in CH2C12 (50 mL) at 0°C. The resulting mixture was stirred at room temperature overnight.

Additional bromine was added to the reaction mixture in 0.05 mL portions until the reaction was com- pleted, as indicated by HPLC. The reaction mixture was diluted with CH2Cl2 and washed with 10% NaHS03.

The organic layer was dried (MgS04) and concentrated under reduced pressure. The residue was purified by flash chromatography on silica gel eluting with 1% EtOAc/hexane) to provide the title compound (3.68 g, 88%) as a white solid. 1H NMR (CDC13, 400 MHz) 5 7.50 (d, J=8. 4 Hz, 1H), 6.91 (d, J=8.4 Hz, 1H), 2.47 (s, 3H).

Example 28C 3- (2, 3-Dichloro-4-methylsulfanyl- phenyl) -acrylic acid methyl ester The title compound was prepared from Example 28B (1.60 g, 5.916 mmol) and methylacrylate (1.6 mL, 17.75 mmol) by the procedure described in Example 20E. 1H NMR (400 MHz, DMSO-d6) 5 7.92 (d, J=8.59 Hz, 1H), 7. 87 (d, J=16. 01 Hz, 1H), 7.29 (d, J=8.59 Hz, 1H), 6.72 (d, J=16.01 Hz, 1H), 3.72 (s, 3H), 2. 55 (s, 3H).

Example 28D 3- (2, 3-Dichloro-4-methylsulfanyl- phenyl) -acrylic acid The title compound 28D (774mg, 79%) was prepared from Example 28C by the method described in Example 20F. 1H NMR (400 MHz, DMSO-d6) 5 12.63 (s, 1H), 7.89 (d, J=8.59 Hz, 1H), 7.82 (d, J=16.01 Hz, 1H), 7.27 (d, J=8.59 Hz, 1H), 6.60 (d, J=15.62 Hz, 1H), 2.54 (s, 3H). Analytical HPLC (Method C) Rt=3.165 min (220 nm, 96 area%).

Example 28E <BR> <BR> <BR> <BR> <BR> <BR> {1- [3- (2, 3-Dichlo. ro-4-methylsulfanyl-<BR> <BR> <BR> phenyl)-acryloyl]-azetidin-3-yl}- carbamic acid tert-butyl ester A solution of Example 28D (100 mg, 0.380 mmol), HOBt. H20 (87 mg, 0.570 mmol), NMM (0.105 mL, 0.950 mmol) and azetidin-3-yl-carbamic acid tert- butyl ester (79 mg, 0.456 mmol), in dry DMF (2 mL) was treated with EDCI (109 mg, 0.570 mmol) at 0°C and allowed to stir at ambient temperature under N2 atmosphere for 18 h. The reaction mixture was diluted with warm 2% methanol/EtOAc (40 mL), washed with water (2x10 mL) and brine (lx10 mL). The organic phase was separated (while keeping warm, otherwise product precipitates out), dried (Na2SO4), filtered, and evaporated to dryness. The residue obtained was crystallized from methanol to provide the desired product (121 mg, 76. 26%) as a white solid. 1H NMR (400 MHz, DMSO) 5 7. 93 (d, J=8.59 Hz, 1H), 7.70 (d, J=15.62 Hz, 1H), 7.60 (br d, J=7. 81 Hz, 1H), 7.30 (d, J=8.59 Hz, 1H), 6. 78 (d, J=15.62 Hz, 1H), 4.54-4. 50 (m, 1H), 4.35-4. 27 (m, 1H), 4.18- 4.13 (m, 1H), 4.09-4. 06 (m, 1H), 3.82-3. 78 (m, 1H), 2.57 (s, 3H), 1.40 (s, 9H). LCMS (APCI+ ; Method F) m/z 417,419 (M+H) + ; Analytical HPLC (Method C) Rt=3.487 min (220 nm, 100 area %).

Example 28F 1-(3-Amino-azetidin-1-yl)-(2, 3- dichloro-4-methylsulfanyl-phenyl)- propenone HC1 salt A suspension of Example 28E (120 mg, 0.288 mmol) in CH2Cl2 (1 mL) was treated with 4N HC1 in dioxane (4 mL) at 0°C under N2 atmosphere for 30 min and allowed to stir at ambient temperature for 2 h.

The reaction was. monitored by HPLC at 1 h intervals.

After 2 hours the solvent was removed under reduced pressure and dried under vacuum for 5 h to provide the HC1 salt of the amine (110 mg, 108. 6%) as a white solid. 1H NMR (DMSO, 400 MHz) 5 8. 64 (br s, 2H), 7.99 (d, J=8.59 Hz, 1H), 7.74 (d, J=15.62 Hz, 1H), 7.30 (d, J=8.59 Hz, 1H), 6.85 (d, J=15. 62 Hz, 1H), 4.59-4. 55 (m, 1H), 4.33-4. 28 (m, 1H), 4.22-4. 17 (m, 1H), 4. ao-3. 96 (m, 1H). LCMS (APCI+ ; Method F) m/z 317,318 (M+H) +. Analytical HPLC (Method C) Rt=2.066 min (220 nm, 100 area %, 254 nm 100 area t)-

Example 28G N- {1- [3- (2, 3-Dichloro-4-methylsulfanyl-phenyl]- azetidin-3-yl} -3-methyl-succinamic acid tert-butyl ester The title compound 28G (75 mg, 44. 37%) was prepared from Example 28F and Example 20M as de- cribed in Example 28E. 1H NMR (DMSO, 400 MHz) 5 8.55 (d, J=6.64 Hz, 1H), 7. 94 (d, J=8. 59 Hz, 1H), 7.71 (d, J=15.62 Hz, 1H), 7. 30 (d, J=8. 59 Hz, 1H), 6 81 (d, J=15.62 Hz, 1H), 4.61-4. 53 (m, 1H), 4.50-4. 43 (m, 1H), 4.19 (m, 1H), 4. 09-4. 01 (m, 1H), 3. 83-3. 78 (m, 1H), 2. 67-2.59 (m, 1H), 2.57 (s, 3H), 2. 21 (m, 1H), 1.37 (s, 9H}, 1. 03 (d, J=7.03 Hz, 3H). LCMS (APCI+; Method F) m/z 487,489 (M+H) +. Analytical HPLC (Method C) Rt=3. 201 min (220 nm, 100 area %, 254 nm. 100 area %).

Example 28H N-{1-[3-(2, 3-Dichloro-4-methylsulfanyl-phenyl) - acryloyl]-azetidin-3-yl}-3-methyl-succinamic acid

The title compound (51 mg, 82%) was pre- ared from Example 28G as described in Example 23E.

H NMR (DMSO, 400 MHz) 5 12.08 (bs, 1H), 8.55 (d, J=6. 64 Hz, 1H), 7.92 (d, J=9.37 Hz, 1H), 7.72 (d, J=15. 62 Hz, 1H), 7.30 (d, J=8.59 Hz, 1H), 6.80 (d, J=15. 23 Hz, 1H), 4.58-4. 53 (m, 1H), 4.48-4. 42 (m, 1H), 4. 17 (m, 1H), 4.09-4. 03 (m, 1H), 3. 83-3. 77 (m, 1H), 2.67-2. 62 (m, 1H), 2. 57 (s, 3Hj, 2.25-2. 19 (m, 1H), 1.04 (d, J= 7.03 Hz, 3H). LCMS (APCI+; Method F) m/z 431, 433 (M+H) +. Analytical HPLC (Method C) Rt=2.556 min (220 nm, 85 area %).

Example 29 N- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)- acryloyl]-azetidin-3-yl}-3-methyl-succinamic acid Example 29A {l- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)- acryloyl]-acetidin-3-yl}-carbamic acid tert-butyl ester

The title compound (139 mg, 89%) was pre- ared from Example 20F (100 mg, 0.366 mmol) and azetidin-3-yl-carbamic acid tert-butyl ester (76 mg, 0.439 mmol) as described in Example 28E. Example 29A was obtained as a white powder. 1H NMR (400 MHz, DMSO) # 8.48 (s, 1H), 7. 98 (d, J=5.47 Hz, 1H), 7.84 (d, J=15.62 Hz, 1H), 7.62 (br d, J=7.03 Hz, 1H), 7.58 (d, J=5.47 Hz, 1H), 6.78 (d, J=15.62 Hz, 1H), 4.55 (t, J=8.59 Hz,'1H), 4.38-4. 30 (m, 1H), 4.20- 4.15 (m, 1H), 4.13-4. 09 (m, 1H), 3, 84-3. 80 (m, 1H),' 1.40 (s, 9H). LCMS (APCI+; Method F) m/z 427, 429 (M+H) +i Analytical HPLC (Method C) Rt=3.473 min (220 nm, 100 area %).

- Example 29B 1-(3-Amino-azetidin-1-yl)-3-(6, 7-dichloro- benzo [b] thiophen-5-yl)-propenone HCl salt The title compound was prepared from Example 29A as described in Example 28F. 1H NMR (DMSO, 400 MHz) 5 8. 79 (s, 2H), 8.54 (s, 1H), 7.99 (d, J=5. 47 Hz, 1H), 7.87 (d, J=15.62 Hz, 1H), 7.58 (d, J=547 Hz, 1H), 6.94 (d, J=15.23 Hz, 1H), 4.65- 4.59 (m, 1H), 4.38-4. 35 (m, 1H), 4.24-4. 19 (m, 1H), 4.11-4. 06 (m, 1H), 4.04-4. 00 (m, 1H). LCMS (APCI+ ; Method F) m/z 327,329 (M+H) +. Analytical HPLC (Method C) Rt=2.147 min (220 nm, 100 area %, 254 nm 97.64 area %).

Example 29C N- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)-acryloyl]- acid tert-butyl ester The title compound 29C (42 mg, 25%) was prepared from Example 29B as described in Example 28G. 1H NMR (DMSO, 400 MHz) 5 8.57 (d, J=6.64 Hz, 1H), 8.49 (s, 1H), 7.98 (d, J=5.47 Hz, 1H), 7.85 (d, J=15.62 Hz, 1H), 7.57 (d, J=5.47 Hz, 1H), 6.89 (d, d, J1=1. 56 Hz, J2=15. 62 Hz, 1H), 4.59 (m, 1H), 4.48 (m, 1H), 4.21 (m, 1H), 4.11 (m, 1H), 3.83 (m, 1H), 2.64 (m, 1H), 2.22 (m, lH), 1.38 (s, 9H), 1.04 (d, J=7. 03 Hz, 3H). LCMS (APCI+; Method F) m/z 497,498 (M+H) +. Analytical HPLC (Method C) Rt=3.376 min (220 nm, 100 area %).

Example 29D N-11- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl) -acryloyl]- azetidin-3-yl}-3-methyl-succinamic acid

The title compound 29D was prepared from Example 29C as described in Example 28H. LCMS (APCI+; Method F) m/z 441,443 (M+H) +. Analytical HPLC (Method C) Rt=2.370 min (220 nm, 85 area %).

There is no. Example 30.

Example 31 N- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)- acryloyl]-piperidin-4-yl}-3-phenyl-succinamic acid

Example 31A 2-Phenyl-succinic acid 4-tert-butyl ester 1-methyl ester The title compound was prepared by the procedures described in Example 23B, substituting Example 23A with phenyl-acetic acid methyl ester. 1H NMR (CDC13, 400 MHz) 5 7.30 (m, 5H), 4.02 (dd, J=10.0 Hz, J=5.6 Hz, 1H), 3.68 (s, 3H), 3.10 (dd, J=16.4 Hz, J=10.0 Hz, 1H), 2.60 (dd, J=16. 4 Hz, J=5.6 Hz, 1H), 1. 40 (s, 9H).

Example 31B 2-Phenyl-succinic acid 4-tert-butyl ester The title compound was prepared by the procedures described in Example 23C, substituting Example 23B with Example 31A. 1H NMR (DMSO-d6, 400 MHz) 5 7.29 (m, 5H), 3. 86 (dd, J=10. 2 Hz, J=5. 9 Hz, 1H), 2.90 (dd, J=16. 4 Hz, J=10. 2 Hz, 1H), 2.53 (dd, J=16.4 Hz, J=5.5 Hz, 1H), 1.34 (s, 9H). LCMS (ESI- ; Method H) m/z 249 (M-H)-; Rt=2. 37 min (220 nm, 100 area%).

N-{1-[3-(6, 7-Dichloro-benzo [b] thiophen-5-yl)-acryloyl]- piperidin-4-yl}-3-phenyl-succinamic acid tert-butyl ester

The title compound was prepared by the procedures described in Example 23D, substituting Example 23C with Example 31B. 1H NMR (CDC13, 400 MHz) 5 8.04 (d, J=15.2 Hz, 1H), 7.91 (s, 1H), 7.52 (d, J=5.5 Hz, 1H), 7.31 (m, 7H), 6.85 (br d), 5.42 (d, J=7.6 Hz, 1H), 4.55 (m, 1H),. 4. 01 (m, 1H), 3.83 (dd, J=8. 6, J=5.9 Hz, 1H), 3.24 (m, 1H), 3.17 (dd, J=16. 4, J=9.0 Hz, 1H), 2.86 (m, 1H), 2.56 (dd, J=16.4, J=5.9 Hz, 1H), 1.98 (m, 2H), 1.38 (s, 9H), 1.24 (m, 2H). LCMS (APCI+ ; Method I) m/z 587,589 (M+H) + ; Rt=4.28 min (220 nm, 100 area%).

Example 31D N- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)- acryloyl]-piperidin-4-yl}-3-phenyl-succinamic acid The title compound was prepared by the procedures described in Example 23E, substituting Example 23D with Example 31C. H NMR (DMSO-d6, 400 MHz) 5 12.12 (s, 1H), 8.54 (d, J=9.4 Hz, 1H), 8.06 (d, J=7.6 Hz, 1H), 7.98 (s, 1H), 7. 88 (d, J=14.8 Hz, 1H), 7.55 (m, 1H), 7.40 (m, 1H), 7.31 (m, 4H), 7.22 (m, 1H), 4.20 (m, 2H), 3.87 (m, 1H), 3.80 (m, 1H), 3.24 (m, 1H), 2.93 (m, 2H), 2.53 (m, 1H), 1.83 (m, 1H), 1.64 (m, 1H), 1.35 (m, 1H), 1. 20 (m, 1H). LCMS (APCI+; Method I) m/z 531,533 (M+H) +; Rt=3.29 min . (220 nm, 100 area%)..

Example 32 N- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)- acryloyl]-piperidin-4-yl}-3-pyridin-3-yl- succinamic acid

Example 32A 2-Pyridin-3-yl-succinic acid 4-tert-butyl ester 1-ethyl ester The title compound was prepared by the procedures described, in Example 23B, substituting . Example 23A with pyridin-3-yl-acetic acid ethyl ester. 1H NMR (CDC13, 400 MHz) 5 8.57 (d, J=2.4 Hz, 1H), 8.54 (dd, J=4.8 Hz, J=1.6 Hz, 1H), 7.67 (dt, J=8. 0 Hz, J=1.6 Hz, 1H), 7.29 (dd, J=8.0 Hz, J=4. 8 Hz, 1H), 4.16 (m, 2H), 4.05 (dd, J=9.2 Hz, J=6.4 Hz, 1H), 3.11 (dd, J=16.4 Hz, J=9. 2 Hz, 1H), 2.63 (dd, J=16.4 Hz, J=6. 4 Hz, 1H), 1.40 (s, 9H), 1. 21 (t, J=7.2 Hz, 3H). LCMS (APCI+; Method I) m/z 280 (M+H) + ; Rt=3.19 min (220 nm, 100 area%).

Example 32B 2-Pyridin-3-yl-succihic acid 4-tert-butyl ester The title compound was prepared by the procedures described in Example 23C, substituting Example 23B with Example 32A. 1H NMR (DMSO-d6, 400 MHz) 5 8.52 (d, J=2.0 Hz, 1H), 8.47 (dd, J=4.8 Hz, J=1. 6 Hz, 1H), 7.72 (dt, J=7.8 Hz, J=2.0 Hz, 1H), 7.36 (dd, J=7.8 Hz, J=4. 8 Hz, 1H), 3.95 (dd, J=9.4 Hz, J=6.2 Hz, 1H), 2.96 (dd, J=16. 4 Hz, J=9.8 Hz, 1H), 2.62 (dd, J=16. 4 Hz, J=6.2 Hz, 1H), 1.34 (s, 9H). LCMS (APCI+; Method I) m/z 252 (M+H) + ; Rt=1.80 min (220 nm, 100 area%).

Example 32C N- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)-acryloyl]- piperidin-4-yl}-3-pyridin-3-yl-succinamic acid tert- butyl ester The title compound was prepared by the . procedures described in Example 23D, substituting Example 23C with Example 32B. 1H NMR (CDC13, 400 MHz) 5 8.51 (m, 2H),. 8.04 (d, J=15'. 6 Hz, 1H), 7.91 (s, 1H), 7.71 (dt, J=8.2 Hz, J=2.0 Hz, 1H), 7. (d, J=5.6 Hz, 1H), 7.33 (d, J=4.2 Hz, 1H), 7.26 (m, 1H), 6.85 (d, J=15.2 Hz, 1H), 5.91 (d, J=7. 8 Hz, 1H), 4.60 (m, 1H), 4.02 (m, 2H), 3.81 (dd, J=9.4 Hz, J=5.8 Hz, 1H), 3.23 (m, 1H), 3.15 (dd, J=16.8 Hz, J=9. 4 Hz, 1H), 2.85 (m, H), 2.57 (dd, J=16.8 Hz, J=5.5 Hz, 1H), 1.99 (m, 2H), 1. 39 (s, 9H), 1.30 (m, 2H). LCMS (APCI+; Method I) m/z 588, 590 (M+H) +; Rt=3.73 min (220 nm, 100 area%).

Example 32D N- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)-, acryloyl]-piperidin-4-yl}-3-pyridin-3-yl- succinamic acid The title compound was prepared by the procedures described in Example 23E, substituting Example 23D with Example 32C. 1H NMR (DMSO-d6,400 MHz) 5 12.27 (s, 1H), 8.55 (m, 3H), 8.18 (d, J=7.6 Hz, 1H), 7.97 (s, 1H), 7.88 (m, 2H), 7.48 (m, 3H), 4.21 (m, 2H), 3.97 (m, 1H), 3. 80 (m, 1H), 3.26 (m, 1H), 3.00 (dd, J=16.4 Hz, J=9. 0 Hz, 1H),, 2.91 (m, 1H), 2.63 (dd, J=16.4 Hz, J=5. 9 Hz, 1H), 1.84 (m, 1H), 1.66 (m, 1H), 1.35 (m, 1H), 1. 21 (m, 1H). LCMS (APCI+; Method I) m/z 532,534 (M+H) +; Rt=2.67 min (220 nm, 100 area%).

Example. 33 N- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl) - acryloyl]-piperidin-4-yl}-3-quinolin-3- yl-succinamic acid

Example 33A Quinolin-3-yl-acetic acid tert-butyl ester To a solution of 3-bromoquinoline (0.208 g, 1. 00 mmol) in THF (5 mL) was added (1-tert- butoxy-vinyloxy) -tert-butyl-dimethyl-silane (0.472 g, 2.05 mmol), Pd (Tol3P) 2Cl2 (0.039 g, 0.05 mmol) and KOAc (0.196 g, 2.00 mmol). The mixture was heated at reflux for 48 h under nitrogen. After cooling to room temperature, the reaction was quenched with sat. NH4C1, diluted with Et2O and stirred for 20 min. Phases were separated. The aqueous phase was extracted with Et20. The combined organic layers were washed with water, brine, dried and concen- rated. The residue was purified by flash-chromatog-

aphy (silica gel, gradient elution from 10: 1 to 6: 1 hexanes/EtOAc) to give the title compound (0.202 g, 83%) as a pale yellow oil. 1H NMR (CDC13, 400 MHz) õ 8. 84 (d, J=2. 0 Hz, 1H), 8.09 (d, J=8.4 Hz, 1H), 8.06 (s, 1H), 7.80 (d, J=8.0 Hz, 1H), 7.69 (t, J=8.0 Hz, 1H), 7.54 (t, J=8. 0 Hz, 1H), 3.72 (s, 2H), 1.45 (s, 9H). LCMS (APCI+ ; Method I) m/z 244 (M+H) + ; Rt=3.36 min (220 nm, 100 area%).

Example 33B Quinolin-3-yl-acetic acid methyl ester To a stirred solution of Example 33A (0.153 g, 0.629 mmol) in MeOH (2 mL) was added hydrochloride in ether (2 M, 2 mL). After stirring for 48 h, the reaction mixture was concentrated in vacuo. The resulting solid was suspended in IPA- ether (1: 5) and stirred for 30 min. The mixture was filtered to give the compound (0.105 g, 83%) as a white solid. 1H NMR (CDC13, 400 MHz) 5 8.84 (d, J=2.0 Hz, 1H), 8. 11 (d, J=8.4 Hz, 1H), 8.08 (s, 1H), 7.80 (d,. J=9.6 Hz, 1H), 7.70 (t, J=8.0 Hz, 1H), 7.55 (t, J=8.0 Hz, 1H), 3.83 (s, 2H), 3.73 (s, 3H). LCMS (APCI+ ; Method I) m/z 202 (M+H) + ; Rt=2.54 min (220 nm, 100 area%).

Example 33C 2-Quinolin-3-yl-succinic acid 4-tert-butyl ester 1-methyl ester The title compound was prepared by the procedures described in Example 23B, substituting Example 23A with Example 33B. 1H NMR (CDCl3, 400 MHz) 5 8. 87 (d, J=2.4 Hz, 1H), 8.10 (d, J=8. 8 Hz, 1H), 8.07 (d, J=2.0 Hz, 1H), 7.80 (d, J=8.0 Hz, 1H), 7.71 (t, J=8.0 Hz, 1H), 7. 56 (t, J=8. 0 Hz, 1H), 4.26 (dd, J=9.6 Hz, J=6.0 Hz, 1H), 3.70 (s, 3H), 3.24 (dd, J=16. 8 Hz, J=9. 6 Hz, 1H), 2.74 (dd, J=16.8 Hz, <BR> <BR> <BR> J=6. 0 Hz, 1H), 1.40 (s, 9H). LCMS (APCI+ ; Method I) m/z 316 (M+H) + ; Rt=3.15 min (220 nm, 100 area%).

Example 33D 2-Quinoli. n-3-yl-succinic acid 4-tert-butyl ester The title compound was prepared by the procedures described in Example 23C, substituting

Example 23B with Example 33D. 1H NMR (DMSO-d6,400 MHz) 5 12.75 (s, 1H), 8.88 (d, J=2.4 Hz, 1H), 8.28 (d, J=2.0 Hz, 1H), 8.01 (d, J=8.2 Hz, 1H>, 7.97 (d, J=8.2 Hz, 1H), 7.75 (t, J=8.0 Hz, 1H), 7. 61 (t, J=8.0 Hz, 1H), 4.17 (dd, J=9.4 Hz, J=6.2 Hz, 1H), 3.10 (dd, J=16.4 Hz, J=9.4 Hz, 1H), 2.76 (dd, J=16.4 Hz, J=6.2 Hz, 1H), 1. 34 (s, 9H). LCMS (APCI+; Method I) m/z 302 (M+H) + ; Rt=2.13 min (220 nm, 100 area%).

Example 33E N- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)- acryloyl]-piperidin-4-yl}-3-quinolin-3-yl- succinamic acid tert-butyl ester The title compound was prepared by the procedures described in Example 23D, substituting Example 23C with Example 33D. 1H NMR (CDC13, 400 MHz) 1H NMR (CDC13, 400 MHz) 5 8.83 (d, J=2.0 Hz, 1H), 8.15 (d, J=2.0 Hz, 1H), 8.06 (d, J=8.4 Hz, 1H), 8.01 (d, J=15.6 Hz, 1H), 7.89 (br s, 1H), 7.79 (d, J=8.4 Hz, 1H), 7.69 (t, J=6.8 Hz, 1H), 7.55 (d, J=6.8 Hz, 1H), 7.51 (t, J=8.4 Hz, 1H), 7.33 (br s, 1H), 6.84 (m, 1H), 5.87 (d,. J=7.6 Hz, 1H), 4.61 (m,

1H), 4.03 (m, 2H), 4.16 (m, 2H), 3.27 (dd, J=16.8 Hz, J=9.6 Hz, 1H), 3.20 (m, 1H), 2.86 (m, 1H), 2.68 (dd, J=16.8 Hz, J=5.2 Hz, 1H), 2.09 (m, 1H), 1.93 (m, 1H), 1.43 (m, 2H), 1.39 (s, 9H). LCMS (APCI+ ; Method I) m/z 638,-640 (M+H) + ; Rt=4.04 min (220 nm, 100 area%).

Example 33F N- {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)- acryloyl]-piperidin-4-yl}-3-quinolin-3-yl- succinamic acid The title compound was prepared by the procedures described in Example 23E, substituting Example 23D with Example 33E. 1H NMR (DMSO-d6, 400 MHz) 5 8. 98 (s, 1H), 8.53 (br d, 1H), 8. 38 (s, 1H), 8. 26 (d, J=7. 2 Hz, 1H), 8.04 (m, 2H), 7.97 (m, 1H), 7. 87 (d, J=15.2 Hz, 1H), 7.80 (m, 1H), 7.67 (m, 1H), 7.54 (m, 1H), 7.38 (m, 1H), 4.16 (m, 3H), 3.83 (m, 1H), 3.25 (m, 1H), 3.13 (dd, J=16. 4. Hz, J=8.8 Hz, 1H), 2.90 (m, 1H), 2.76 (dd, J=16.4 Hz, J=6. 4 Hz, 1H), 1.86 (m, 1H), 1.65 (m, 1H), 1.37 (m, 1H), 1.18

(m, 1H). LCMS (APCI+ ; Method I) m/z 582,584 (M+H) +; Rt=2.88 min (220 nm, 100 area%).

The following examples 34-118 were pre- pared by procedures described in Examples 25 or 26.

Alternatively, these compounds can be made via solution phase method in solvents such as chloroform or tetrahydrofuran, with the following reagents: PS-DIEA, PS-DMAP, and corresponding acyl chlorides or isocyanates, by shaking in a 24-well format Bohdan mini-block at 550 rpm for 16 hrs. The work- up consisted of rinsing with DMF and/or'THF. The crude solid was purified by preparative HPLC (Methods F or J). LCMS (Method I).

TABLE II 0 HO 0 ru O NH HN /O N O CI CL ci 68 ; s ho 0 O NH HN 0 _ _ 69 CI Cl S-/ Cil STRUCTURE

Example 119 <BR> <BR> N [1- (3-Isoquinolin-4-yl-acryloyl)-piperidin-4-yl]-3-methyl-succi namic acid

Example 119A 3-Isoquinolin-4-yl-acrylic acid methyl ester To a solution of 4-bromoisoquinoline (1 g, 4.8 mmol) in THF (20 mL) were added Pd (OAc) 2 (432 mg, 1.9 mmol), P (o-tol) 3 (2.9 g, 9.6 mmol), NaI (360 mg, 2.4 mmol), and Et3N (2 mL, 14.4 mmol) under a N2 atmosphere. After stirring 30 min, methyl acrylate (0.9 mL, 9.6 mmol) was added and the reaction mix- ture heated to 60°C overnight. The reaction mixture was filtered, concentrated, then purified by flash column chromatography (1: 1 EtOAc/Hexanes) to give the title compound as an orange solid. LCMS (Method I) m/z 214 (M+H) + ; Rt = 2.58 min.

Example 119B <BR> <BR> <BR> N [1- (3-Isoquinolin-4-yl-acryloyl)-piperidin-4-yl]-3-methyl-succi namic acid The title compounds were prepared by pro- cedures described in Examples 20F, 20G, 20H and 20I.

The final product was purified by preparative'HPLC (Method F). LCMS (Method I) m/z 396 (M+H) +

Example 120 N [1- (3-Benzo [b] thiophen-3-yl-acryloyl)-piperidin-4-yl]-3-methyl-succinamic acid

Example 120A 3-Benzo [b] thiophen-3-yl-acrylic acid methyl ester The title compound was prepared by the procedures described in Example 119A, substituting 4-bromoisoquinoline with 3-bromothianaphthalene.

The crude product was purified by flash column chromatography (1: 4 EtOAc/Hexanes) to give the title compound as a brown oil.

Example 120B <BR> <BR> <BR> <BR> <BR> N- [l- (3-Benzo [b] thiophen-3-yl-acryloyl)-piperidin-4-yl]-3-methyl-succinamic acid The title compounds were prepared by pro- cedures described in Examples 20F, 20G, 20H, and 20I. The final product was purified by preparative HPLC (Method F). LCMS (Method I) m/z 401 (M+H) + Example 121 N {1- [3- (6, 7-Dichloro-benzo [b] thiophen-5-yl)-acryloyl]-3-methyl-piperidin-4-yl}-3-methyl-s uccinamic acid.

Example 121A.

4-Amino-3-methyl-piperidine-1-carboxylic acid benzyl ester

To 1-benzyl-3-methyl-piperidin-1-one (3.2 g, 16 mmol) in benzene (100 ml) was added benzyl chloroformate (7.25 g, 42.5 mmol), and the solution refluxed for 15 hours. After concentration and purification by chromatography (dichloromethane as eluant) the product was as clear oil (3.80 g, 98%). This material was carried to the next step and diluted in methanol (100ml), then treated with ammonium acetate (11 g, 138 mmol) and stirred at room temperature for 3 hrs. The solution was cooled to 0°C for 10 min, and sodium cyanoborohydride (0.97

g) was added. The resultant suspension was stirred at 0°C for 10 min and then at room temperature for 4 hrs. The reaction was concentrated, diluted with water and brought to pH 3-4 with 1 N HC1. The aqueous phase was washed with ethyl acetate, brought to pH 10 with 1 N NaOH and extracted with dichloro- methane, dried over MgS04 and concentrated to yield the final product as aclear oil (2.6 g, 69%). LCMS (Method G) m/z 249 (M+H) + Example 121B N-{1-l3-(6, 7-Dichloro-benzolb] thiophen-5-yl)-acryloyll-3-methyl-piperidin-4-yl}-3-methyl-s uccinamicacid

The title compounds were prepared by pro- cedures leading to Example 20I. The final product was purified by preparative HPLC (Method F). LCMS (Method G) m/z 483 (M+H) + From the foregoing description, various modifications and changes in composition and methods will occur to those skilled in the art. All such modifications coming within the scope of the appended claims are intended to be included therein.

The following Methods may be used to test compounds of this invention. Unless otherwise indi- ated, the reagents used in the following examples are commercially available and may be purchased from

Sigma-Aldrich Company, Inc. (Milwaukee, WI, USA) or Alfa Aesar (Ward Hill, MA, USA).

Example A Inhibition of soluble VLA-1 binding to Collagen IV Generation of sVLA-1-LZ A soluble form of the alpl heterodimer (sVLA-1-LZ) was generated by truncating each chain of the heterodimer at the beginning of the transmem- rane region and adding an acidic and basic leucine zipper sequence to the al and pi chains, respective- y. The al chain (described in U. S. Patent Applica- ion Publication No. 2003/0088061) was truncated after residue P1141 of the pro-peptide and the 47 amino acid acidic leucine zipper cassette was added.

The PI chain (Genbank accession no. P05556) was truncated after residue D728 of the pro-peptide and the 47 amino acid basic leucine cassette was added. Both constructs were then expressed in CHO DG44 cells, and the heterodimer was purified from the culture supernatant using an antibody affinity column specific for the leucine zipper sequences, followed by size-exclusion chromatography on an S200 sizing column (Pharmacia, now Pfizer Inc. , New York, NY USA) using methods standard the art. (See also, U. S. Patent Application Publication No.

2003/0088061). The purified protein was stored at - 70°C.

Briefly, an al-LZ construct was generated which has the extracellular domain of al fused to a

C-terminal leucine zipper sequence. The extra- cellular domain of al was amplified using standard PCR methods and reagents in order to add restriction sites for the subcloning. The sequences of the primers used in the PCR reaction were: 5'primer : a1-01 : ATT CTC GAG ACC GCC ACC ATG GTC CCC AGG CGT CC (SEQ. ID. 1) 3'primer : a1-04 : ATT ACG CGT TGG CAC TCT GCC CGG TAG (SEQ. ID. 2) The primers above were used in a PCR reac- tion with an al cDNA clone (described previously, U. S. Patent Application Publication No.

2003/0088061). The resulting PCR product was sub- cloned 5'to the acidic leucine zipper sequence in the mammalian expression vector pDEF38 (described in U. S. Patent Application Publication No.

2003/0088061). The resulting plasmid was verified by sequencing.

The domain of pi likewise was amplified using standard PCR methods and re- agents in order to add restriction sites for the subcloning. The sequences of the primers used in the PCR reaction were: 5'primer : al-02 : ATT CTC GAG ACC GCC ACC ATG AAT TTA CCA ATT TTC TGG (SEQ. ID. 3) 3'primer : al-03 : GTT CCA TTC ACC CCG TTC TTG C (SEQ. ID. 4) In order to generate a pi leucine zipper soluble molecule, the 5'end of the ß-1 insert was generated by PCR from a ßl cDNA and subcloned 5'to

the basic leucine zipper sequence in the mammalian expression vector pNEF38 (described in U. S. Patent Application Publication No. 2003/00.88061). The resulting plasmid was verified by sequencing.

Alpha 1 Leucine Zipper (Acidic) construct protein sequence:

Beta 1 Leucine Zipper (basic) construct protein sequence: Signal Sequences are shown in italics Mature polypetides are shown in plain text

Acidic and Basic Leucine Zippers are underlined and bold Screening Assay for Measuring sVLA-1-LZ Binding to Collagen IV A biochemical assay for measuring binding of sVLA-1-LZ to Collagen IV using time-resolved fluorescence, suitable for high-throughput screen- ing, was developed. Briefly, 96-well Immulon 4 ELISA plates were coated with sVLA-1-LZ at 2 pg/mL in 50mM NaHCO3, pH 9.2, and incubated overnight at 4°. The plates were then washed two times with 300 pL/well Wash Buffer (TBS ; 0.1% Tween-20 ; 2mM Mol2), and blocked for l hour with 200 uL/well Blocking Buffer (2% BSA; CMF-PBS; 2mM MgCl2) at room tempera- ture. Compounds of this invention were prepared in DMSO at 200X the desired final assay concentration.

Final concentrations were selected from a range be- tween 0. 01 nM-100pM. DMSO inhibitor stocks were then diluted to 2X final'concentration in Dilution Buffer (TBS ;. 0. 01% BSA; 2mM MgCl). Fifty pL/well Dilution Buffer, alone or in combination with, anti- al mAb (Immunodiagnostic, #8149a, 2. 5 ug/mL), 10 mM EDTA, DMSO or DMSO+2X inhibitor was added to the wells of the plate, followed by 50 pL/well Collagen IV-biotin at 2 pg/mL in Dilution buffer. Collagen IV-biotin was generated by biotinylating human Collagen IV (Sigma-Aldrich, Milwaukee, WI, USA) using a biotin labeling kit (Pierce Biotechnology, Rockford, IL, USA) following the manufacturer's protocol. The plates were incubated for 1 hour at

room temperature, and washed four times with Wash Buffer (300 pL/well). The plates were then incu- bated with 100 pL/well of 1: 1000 diluted (with H2O) Strepavidin-Europium (PerkinElmer, Boston, MA, USA) for 30 min. at room temperature. The plates were then washed four times with Wash Buffer (300 pL/- well). One hundred uL/well of Delphia Enhancement Solution (PerkinElmer ; diluted 1: 1 with dH2O) was added, and the plates were shaken for 5 minutes.

Binding was'then analyzed. by time resolve fluor- escence (TRF) using a Victor Plate-reader (Perkin- Elmer). Results were analyzed using the equations below. The percent of inhibition was plotted versus the log concentration of. inhibitor across a twelve point titration, and a linear regression trend-line was drawn.

Specific binding = TRF signal (Collagen-IV-biotin in Dilution Buffer plus Ab, DMSO only or DMSO + inhbitor)-TRF signal (Collagen-IV-biotin with EDTA) % inhibition= 1-specific binding of Collagen-tV-biotin with inhibitor in DMSO x 100 specific binding of Collagen-IV-biotin with DMSO only

As a variation in the assay, the effect of serum protein in the assay was determined by substi- tuted Collagen IV-biotin at 2ug/mL in FBS, instead

of Dilution buffer, resulting in a final serum con- centration of 50% in the assay. The percent inhibi- tion in the presence and absence of 50% FBS was then compared.

Example B Inhibition of VLA-1 dependent K562-al cell adhesion to Collagen IV Generation of K562-al cells The al cDNA was subcloned into the mammal- ian expression vector pMH-Neo (Hahn, W. C. et al., 1993. Gene 127: 267). The'resulting clone a1/- pMHneo/40 was verified by sequencing.'The al/- pMHneo/40 construct was introduced into'K562 cells by electroporation. The transfected cells were initially maintained in complete RPMI with 10% FBS.

Two days after the transfection, the cells were spun down and resuspended in complete RPMI with 10% FBS and 0.5mg/mL G418 (Sigma-Aldrich) to select for the neomycin resistance conferred by the pMHneo plasmid.

Transfectants with functional al expression were selected by panning the cells for binding to Type IV collagen (an al ligand). In panning experiments, K562 transfectants were stimulated with 20ng/mL of PMA and allowed to adhere to plate-bound Type IV collagen. After a 30-60 min incubation at 37°C, the unbound cells were washed away and the adherent cells were recovered with versene. Cellular expres- sion of VLA-1 was verified by FACS analysis with an al mAb.

K562-al cell adhesion assay to Collagen IV The day prior to the assay, the K562-a1 cells were split 1: 2 into fresh culture media (RPMI- 1640, 10% FBS) and cultured at 37°C in CO2. Ninety- six well Immulon-4 plates were coated with either 1. 25 ug/mL collagen IV (Sigma) in CMF-PBS or the 21 mAb at 5pg/mL in Coating-Buffer (50 mM sodium car- bonate, 50 mM sodium bicarbonate, pH 9.6), and incubated overnight at 4°C. On the day of the assay, the plates were washed with D-PBS and then blocked with 1% BSA/D-PBS for 1.5-2. 0 hours at room temperature. Compounds of this invention were pre- pared in DMSO at 66.67X the desired final assay concentration. Final were selected from a range between 1 nM-100 uM. Deep-well blocks (96-square well titer plates, Beckman) were prepared with 700pL RPMI/well. DMSO solutions of diluted compound were then added to the deep-well block (10. 5pL diluted compound/700uL RPMI) to generate 1.5X solutions. After blocking, the Immulon-4 plates were aspirated and 200, uL RPMI only, RPMI + 1.5X DMSO/RPMI Solution, or RPMI + 1.5X compound in DMSO/RPMI solution was added to each well. The plates were then incubated at 37°C in 5% C02 for 10- 30 min. K562-a1 cells were counted, spun down, washed once with RPMI and resuspended in RPMI+60ng/- mL PMA at a density of 1. OX106 cells/mL. One hundred pL cell suspension was then added to each well and incubated at 37°C in 5% C02 for 30 min. Adherent cells were then fixed to the plate with 14% glutar- aldehyde/D-PBS for 1.5 hours. Plates were washed

with water and the cells were stained with 5% crystal violet for 5 min. at room temperature. The plates were washed with water again, and the crystal violet dye was extracted from the cells with 200 pL/well 70% ethanol. The plates were read on a plate reader at A570nm and A410nm. Results were analyzed using the equations below. The percent inhibition was plotted versus the log concentration of inhibitor across an eight point titration, and a linear regression trend-line was drawn. As a varia- tion in the assay, the effect of serum protein in the assay was determined by substituting RPMI+50% FBS instead of The percent inhibition in the presence and absence of 50% FBS was then compared. Percent cells binding = A570-A410 (binding to Collagen IV) x 100 A570-A410 (binding to Alphal antibody) Percent Inhibition = 0 cells binding with inhibitor x 100 % cells binding with DMSO It is contemplated that all of this inven- tion will exhibit at least fifty percent inhibition in one or of the above assays when tested at a concentration of 50 uM.