WO2007134169A2 - Indole, benzimidazole, and benzolactam boronic acid compounds, analogs thereof and methods of use thereof - Google Patents

Abstract

Benzimidazole, indole and benzolactam boronic acid compounds, analogs thereof, and pharmaceutical formulations are described, along with methods of use thereof for inhibiting inflammatory cytokines such as tumor necrosis factor alpha (TNF-α) in a subject in need thereof.

Description

INDOLE, BENZIWIiDAZOLE, AND BENZOLACTAM BORONIC ACID COMPOUNDS, ANALOGS THEREOF AND METHODS OF USE THEREOF

Cross-Reference to Related Applications

[0001] This application claims the benefit under 35 U S C 119(e) of U S S N 60/799 599 filed May 10 2006 hereby incorporated by reference in its entirety

Field of the Invention

[0002] The present dfscfoser provides indoϊe benzimidazole, and benzolactam boronic acid compounds, analogs thereof, pharmaceutical formulations containing the same, and methods of use thereof particularly for inhibiting an inflammatory cytokine such as TNF-α in a subject in need thereof

Background of the Invention

[0003] Tumor necrosis factor α (TNF-α} is an inflammatory cytokine produced by neutrophils, activated lymphocytes macrophages, NK cells, LAK cells, astrocytes, and others TNF-α mediates a variety of cellular activities, including cytotoxic effects against tumor cells, activation of neutrophils, growth proliferation of normal cells, and immunoinflammatory, immunoregulatory, and antiviral responses Unfortunately TNF-α also mediates a variety of pathological activities in diverse number of disease states See generally U S Patent No 5 643,893 to Benson et al , see also PCT Application WO 00/73253 to Palladmo et al Accordingly there is a need for new inhibitors of TNF-α

[0004] Several antibody based TNF-α inhibitors are commercially available For example HUMIRA® (adalimumab) is a recombinant human IgGI monoclonal specific for human TNF and is administered subcutaneousiy ENBREL® (etanercept) is a dimeπc fusson protein consisting of the extracellular tigand-bsnding portion of the human 75 kilodalton (p75) tumor necrosis factor receptor (TNFR) linked to the Fc portion of human IgGI specific for human TNF and is administered by subcutaneous injection REMICAOE® (snflixamab) is a chimeric IgG 1 monoclonal antibody specific for human TNF- α and is administered by intravenous infusion However these antibody based therapeutics have several disadvantages as compared to smaii molecules including immunogemcity cost and are limited to non-oral administration

[0005] Phosphodiesterase inhibitors are potent suppressors of many inflammatory cytokines For example phosphodiesterase 4 inhibitors can inhibit TNF-α release from macrophages monocytes and T cells which suggests that they could be effective in inflammatory diseases including nflammatory bowe^! disease but fay a mechanism that is cMfe^rent from that of the aπftbody based TNF-α rrhibitors (Banner β^f a' Trends in Pharmaceutical Sciences VoI 25 Ho 8 12004} [0006] U S Patent No 5 643 893 to Benson et al describes certain dihydroxyboryl atkyl purine, tndofe and pyrimidine dertvavttes that are useful as inhibitors of inflammatory cytokines tn general such inhibitors are compounds of the formula

[0007] where R₁ and R₂ are both hydrogen atoms or together are a propylene cham bridging the two oxygen atoms n ss 2-6 and P is a purine indole or pyπmidsne base residue bonded via the N^s in the case of a purine base or via the N¹ in the case of an indole or pyπmidme base Certain specific substitutions including 6- and 2 6- substituted puπne derivatives are also described

(0008] PCT Application WO 02/085916 to Ishaq also describes certain dihydroxyboryl atkyl purine inhibitors of inflammatory cytokines of the formula

[0009] where P is a puπne base, and R₁ and R₂ are both hydrogen atoms or together are a 3 to 5 carbon alkyjene chain Certain specific substitutions including δ~ 2 6- and 8- substituted purine derivatives are also described {see e g page 21 lines 6-7}

(OOIOj In spite of the foregoing there remains a need for new compounds particularly for oral administration for the inhibition of inflammatory cytokines such as TNF-α and methods of use thereof

Summary of the invention

[001 1] A first aspect of the present invention is a compound of Formula 1 or Formula

[0012] wherein:

[00Ϊ3] A is N or C₁ subject to the proviso that R⁵ is absent when A is N;

[0014} X is -C(O)-, -S(O)₂-, or a covafent bond;

(0015] Y is linking group such as aikyl, aJkenyl, cycloalkyl, alkyicycioalkyi, alkylcycloaikylalky!, ajkyloxyalkyl, aryl, alkylaryl, aikylarylaikyl. arylalkyi, cycloaikytalkyl, alkylheterocycle, heterocyclealkyl, atkylheterocyclealkyl, heterocycle, aminoalkyi, oxyaikyl aminoaryS, oxyaryl;

[00161 Z is selected from the group consisting of -B(QFOOR², -CON(R^)OR². and ~N(OR¹)COR² or any of the additional alternatives for Z described in greater detail below;

[0017] R¹ and R² are each independently H, loweratky!, or together form G2-C4 aikyiene;

[0018J R³. R⁴, R⁵, R⁶, and R' are each independently selected from the group consisting of H. halo lowerafkyl, haiolowefaikyi, riaSoloweraSkoxy, foweralkoxy^* hydroxy, toweralkoxycarbo. carboxyiic ac?d acy!, aztdo, mercapto, alkylthio. ammo, heterocycleamsno, alkylamino, dialkytamirvo acyiamiπo. aminoacyl, arylamino, aryiaikyl, aryϊalkylamϊno aryioxy, cyano, sulfonamide, aminosulfonyϊ suifone, nrtro, aryialkyloxy, cyctoajkyfoxy cycloalkylalkoxy, cycloalkylammo, urea, cycloaikylalkylamino, cyctoalkyl, alkylcycloalkyl, hydroxyamsno, aϊkoxyacytamsno and arydhio, and 5- or 6- membered organic rings containing 0 to 4 heteroatoms selected from the group consisting of N, O and S. which πngs may be unsubstrtuted or substituted from 1 to 4 times with halo, toweralkyl, hatoloweralkyl, halotoweralkyloxy, (oweralkoxy, hydroxy, lowerafkoxycarbo, carboxylic acid, acyl, azido, mercapto, alkyithio, amino, heterocycleamino. alkylamino, dialkyjamino, acylamtno, aminoacyl, arylamino, aryiaikyl, aryialkylamsno, aryioxy, cyano, sulfonamide, aminosulfonyS, suifone, and nitro; and oxoheterocyclic groups, or a pharmaceutically acceptable salt or prodrug thereof (sometimes referred to as "active compounds" herein).

[0019] Another aspect of the present invention is a compound of Formula III, Formula IV or Formula V

[0020] wherein:

[0021J X is -C(O)-, -S(O)₂-, or a covaieπt bond;

[0022] Y is afkyi, alkeπyl, cycloalkyl, alkylcyclαalkyl, alkylcyctoalkyiaikyi, alkyloxyatkyl, aryl, alkyiaryl, alkylarylalkyi, arylalkyl, cycloalkylalky!, alkylheterocycie, heterocyclealkyl, alkyiheterocycieaikyϊ, heterocycle, aminoalkyt, oxyalkyl, amsnoaryl, or oxyaryl;

[0023J Z is selected from the group consisting of -B(OR¹)OR², -CON{R¹)OR², and -N(OR¹)COR², or any of the aiterrsatives for Z discussed below,

[0024] R¹ and R² are each independently H, loweralkyi, or together form C2-C4 alkylene; and

[0025] R³, R⁴, R⁵, R⁶, R^? and R⁸ are each independently selected from the group consisting of: H, halo, loweraikyl, haloloweraiky!, haloloweralkoxy, loweraikoxy, hydroxy, loweralkoxycarbo, carboxylic acid, acyl, azido, mercapto, atkytthio, amino, heterocycleamino, aikylamino, dialkySamino, acylamino. aminoacyi, arylamiπo, arylalkyl, arylalkylamino, aryloxy, cyano, sulfonamide, aminosulfonyl, sulfone, nitro arylalkyloxy, cycioalkyloxy, cycloalkylalkoxy, cycloalkylamino, urea, cycloalkyialkylamino, cycloalkyl, aikylcycloalkyl, hydroxyamino, alkoxyacylamino, and arylthio;and 5- or 6- membered organic rings containing 0 to 4 heteroatoms selected from the group consisting of N, O and S, which rings may be unsubstituted or substituted from 1 to 4 times with halo, loweralkyl, haloloweralkyl, haloSoweralkySoxy, loweralkoxy, hydroxy, ioweralkoxycarbo, carboxylic acid, acyi, azido, mercapto, alkylthio, amino, heterocycleamino, aikylamino, dϊalkylamtno. acylamino, aminoacyf, arylamino, arylalkyl, arylaikyiamino, aryloxy, cyano, sulfonamide, aminosulfonyi, sulfone, and nitro, and oxoheterocyclic groups;or a pharmaceutically acceptable salt or prodrug thereof (sometimes referred to as "active compounds" herein).

[0026] Another aspect of the present invention is a compound of Formula Vl

[0027] wherein

[0028] A is S. O, SO₂ or NR;

[0029] X is -C(O)-, -S(O)₂-, or a covalent bond;

[0030] Y is aikyl, alkenyl, cycloalkyl, alkylcycloatkyl. alkyjcycloajkyialkyl, alkyloxyalkyl, aryl, alkylaryl. alkylarylaikyl, arylalkyl, cycioalkylalkyi, alkylhetocycSe, heterocyclalkyl, aϊkylheterocyclalkyl, heterocycle, aminoalkyl, oxyalkyl, aminoary!, oxyaryl cycloalkylaikyi, aikylbetocycle, heterocyclalkyl, alkylheterocyclalkyl, heterocycle, aminoalkyl, oxyalkyl, aπrtiπoaryl, oxyaryl;

[0031] Z is selected from the group consisting of "B(OR¹JOR², -CON(R¹)0R², and -N(OR¹)COR² or any of the alternatives for Z described below:

[0032] R¹ and R² are each independently H. Soweralkyl. or together form C2-C4 alkylene: and

[0033] R₁ R³, R⁴, R⁵, R⁶, R⁷ _, R⁸ _, R^s and R¹⁰ are each independently selected from the group consisting of: H₁ halo, ioweralkyl, haloloweralkyl, haloloweralkoxy. loweralkoxy, hydroxy, ioweralkoxycarbo, cycioalkyl, alkylcycloalkyl, carboxyJic acid, acyl, azido, rnercapto, alkylthio, amino, heterocycleamino, alkyiamino, dialkylamino, acylamino, aminoacyl, arylamino, arylalkyl. arylalkylamino. aryloxy, cyano, sulfonamide, aminosulfony!, sulfone, nitro arytalkyloxy, cycloalkyloxy, cycϊoalkylalkoxy, cycloalkylamino. urea, cycloalkylalkyiamino, hytiroxyamino, aikoxyacylamino, and aryithio; and 5- or 6- membered organic rings containing 0 to 4 heteroatoms selected from the group consisting of N, O and S, which rings may be unsυbstituted or substituted from 1 to 4 times with halo, loweralkyi. haloloweralkyl, halcloweralkyioxy, loweralkoxy, hydroxy, ioweralkoxycarbo. carboxySic acid, acyl, azido, mercapto, afkylthto. amino, heterocycieamino, alkylamino, dialkyfamino, acylamino, aminoacyl, aryiamino. arylalkyi. arylalkylam^no, aryioxy, cyano, sulfonamide, aminosulfonyf. sulfone. and mtro^" and oxoheterocyclic groups;

JO034J or R^s and R⁹ together are =O or =S, or a pbarmaceufecalSy acceptable salt or prodrug thereof [0035J Another aspect of the present invention is a compound of Formula VIt:

[00361 wherein:

[0037] A₁ and A₂ are each independently N or C;

[0038] X is -C(O)-. -S(O)₂-, or a covafent bond;

[0039] Y is unking group such as alky!, alkenyl, cycioalkyl, aikyicycloalkyl, aikyicycloalkylalkyl, alkyloxyalkyi, aryj, aSkylaryl, alkylarylalkyl, aryiatkyl. cycioalkylalkyl, alkySheterocycle, heterocyclealkyl, aikylheterocyclealkyl, heterocycle, aminoalkyl. oxyalkyl, aminoaryl, oxyaryi;

[0040] Z is selected from the group consisting of -B(OR¹)OR², -CGN(R¹)0R², and -N(OR¹)COR² or any of the additional alternatives for Z described in greater detail below,

[0041 ] R¹ and R² are each independently H, loweraikyt or together form C2-C4 afkyiene;

[0042J R_n, and R_p are each independently selected from the group consisting of; H, halo, loweraikyl, haloloweralkyl, haloloweralkoxy, loweralkoxy, hydroxy, loweralkoxycarbo, carboxylic acid, acyl, azido, mercapto, alkylthio, amino, heterocycieamino. alkyiamsno, dialkylamino. acyfamino, aminoacyl, arylamino, aryiatkyl, arytalkyiarmno, aryloxy, cyano, sulfonamide, aminosulfonyl. sulfone, nitro; arylalkyloxy, cycloaϊkyϊoxy, cycloafkylalkoxy, cycloalkylamino, urea, cycloalkylalkylamino, eycloalkyi, alkyicycloaikyi, hyclroxyamino. alkoxyacylamϊno, and arylthio; and 5- or δ- membered organic rings containing 0 to 4 heteroatoms selected from the group consisting of N, O and S, which rings may be unsubstituted or substituted from 1 to 4 times with halo, ioweraikyS, haloloweraikyf, hatoloweralkyloxy. loweralkoxy. hydroxy. Soweraikoxycarbo carboxyϊsc acid, acyl. azido, mercapto, alkylthio. amino, heterocycieamino. afkylamino, dialkylamino. acyϊamino, aminoacyl. aryfamino, arylalky!, aryfatkySamino. aryloxy cyano. sulfonamide, aminosulfonyl. suffoπe, and nitro, and oxoheterocyclc groups, subject to the proviso that when A₁ ss C, then π

=1 to 4; when A< is H then to 3 A₂ ts C, then p= 1 to 2, when A₂ is N, then n=1 , or a pharmaceutically acceptable salt or prodrug thereof (sometimes referred to as "active compounds" herein)

[0043] A further aspect of the invention is a method of inhibiting tumor necrosis factor alpha in a subject in need thereof, comprising administering a compound as described above to said subject in an amount effective to inhibit tumor necrosis factor alpha

(0044) A further aspect of the invention ss a method of inhibiting phosphodiesterase in a subject in need thereof, comprising administering a compound or active agent as described herein to the subject in an amount effective to inhibit phosphodiesterase (e g PDE H PDE III, PDE IV PDE V and combinations thereof such as both PDE Il and PDE IV)

[0045] A further aspect of the invention ss a method of treating an inflammatory disease in a subject in need thereof, comprising administering a compound or active agent as described herein to the subject in an amount effective to treat said inflammatory disease

|0046] A further aspect of the invention is a method of treating inflammatory bowel disease in a subject in need thereof, comprising administering a compound or active agent as described herein to the subject in an amount effective to treat inflammatory bowel disease

[0047] A further aspect of the invention is a method of treating rheumatoid arthritis sn a subject in need thereof, composing administering a compound or active agent as described herein to the subject in an amount effective to treat rheumatoid arthritis

[0048] A further aspect of the invention is a method of treating psoriasis in a subject in need thereof comprising administering a compound or active agent as described herein to the subject in an amount effective to treat psoriasis

[0049} A further aspect of the invention is a method of treating ankylosing spondylitis in a subject in need thereof comprising administering a compound or active agent as described herein to the subject in an amount effective to treat ankylosing spondylitis

[0050] A further aspect of the invention is a method of treating psoriatic arthritis in a subject in need thereof, comprising administering a compound or active agent as described herein to the subject in an amount effective to treat psoriatic arthritis

[0051] A further aspect of the invention is a method of treating asthma in a subject in need thereof comprising administering a compound or active agent as described herein to the subject in an amount effective to treat asthma [0052] A further aspect of the invention is a method of treating chronic obstructive pulmonary disease sn a subject in need thereof comprising administering a compound or active agent as described herein to the subject in an amount effective to treat chronic obstructive pulmonary disease

[00S3J A further aspect of the invention is a method of treating Alzheimer's disease sn a subject in need thereof, comprising administering a compound or active agent as described herein to the subject in an amount effective to treat Alzheimer's disease

[0054] A further aspect of the invention is a meϊhod of treating type Il diabetes in a subject sn need thereof comprising administering a compound or active agent as described herein to the subject in an amount effective to treat type Il diabetes

[0055] A further aspect of the invention is a method of treating cancer in a subject sn need thereof comprising administering a compound or active agent as described herein to the subject in an amount effective to treat cancer

[0056) A further aspect of the invention ts a method of treating hypertension in a subject in need thereof comprising administering a compound or active agent as described herein to the subject in an amount effective to treat hypertension

[0057] A further aspect of the invention is a method of treating erectile dysfunction tn a subject in need thereof comprising administering a compound or active agent as described herein to the subject in an amount effective to treat erectile dysfunction

[0058] A further aspect of the invention is the use of a compound or active agent as described herein for the preparation of a medicament for carrying out a method as described herein

(0059) The present invention is explained in greater detail below

BRIEF DESCRIPTION OF THE DRAWINGS

|0060] FiGS 1A-C show the effects of CCI-7155 (50 and 100 mg/kg/day p o ), CC1-7156 (100 mg/kg/day p o ) and sulfasalazine {50 mg/kg/day p o ) on body weight expressed a % change in body weight at Day 0

[0061] FIG 2 shows the effects of CCI-7155 (50 and 100 mg/kg/day p o ), CCl-7156 (100 mg/kg/day p o ) and sulfasalazine (50 mg/kg/day p o ) on macroscopic injury in the colon [0062] FIG 3 shows the effects of CCI-7155 (50 and 100 mg/kg/day p o } CCl-7156 (100 mg/kg/day p o ) and sulfasalazine (50 mg/kg/day p o ) on colon weight Compounds were given in divided doses in a twice a day dosing schedule

[0063] FiG 4 shows the effects of CCI-7155 (50 and 100 mg/kg/day p o ) CCI-7156 (100 mg/kg/day p o ) and sulfasalazine (50 mg/kg/day p o \ on water content in the colon Compounds were given in divided doses in a twice a day dosing schedule

(0064] FiG 5 shows the effects of CCΪ-7155 {50 and 100 rng/kg/day gsven p o m divided doses b i d ) CCS-7156 (100 mg/kg/day given p o m divided doses, b i d ) and sulfasalazine (50 mg/kg/day given p o m divided doses, b i d) on MPO feveis in the colon, expressed as mU/mgprotein

]0065] FiG 6 shows the effects of CCl-7308 (4, 20 and 100 mg/kg/day p o ) or sulfasalazine (50 mg/kg/day p o ) on body weight, expressed a % change in body weight at Day 0

(0066] FIG 7 show the effects of CCl-7308 (4, 20 and 100 mg/kg/day p o) or sulfasalazine (50 mg/kg/day p o } on macroscopic injury in the colon

[0067] FIG 8 shows the effects of CCl-7308 (4 20 and 100 mg/kg/day p o) or sulfasalazine (50 mg/kg/day p o ) on colon weight

[0068] FIG 9 shows the effects of CCl-7308 (4, 20 and 100 mg/kg/day p o) or sulfasalazine (50 mg/kg/day p o ) on TNF-a levels in the colon expressed as pg/mg protein

10069} FiGS 10A- 10C show the effects of CCI-7506 (50 and 100 mg/kg/day p o ) CCI-7507 (25 and 50 mg/kg/day p o ) sulfasalazine (50 mg/kg/day p o ) or infliximab (3 mg/kg i v on Day 1 and 7) on body weight over 14 days expressed a % change of the body weight at Day -1 , prior to TNBS challenge on Day 0

[0070] FIG 11 shows the effects of CCl-7506 (50 and 100 mg/kg/day p o ), CCI-7507 (25 and 50 mg/kg/day p o ), sulfasalazine (SASP 50 mg/kg/day p o ) or infliximab (3 mg/kg i v on Day 1 and 7) on macroscopic injury in the colon, determined 14 days after TNBS challenge as assessed as the colonic lesion area, % of the total area measured

[0071 J FlG 12 shows the effects of CCI-7506 {50 and 100 mg/kg/day p o )» CCI-7507 (25 and 50 mg/kg/day p o } sulfasalazine (50 mg/kg/day p o ) or infliximab (3 mg/kg i v on Day 1 and 7} on macroscopic injury tn the colon, determined 14 days after TNBS challenge as assessed by a Damage Score (0-5 scale) [0072] FfG 13. shows the effects of CCI-7506 (50 and 100 mg/kg/day p o.), CCi-7507 (25 and 50 mg/kg/day p o.), sulfasalazine (50 mg/kg/day p,o.) or infliximab (3 mg/kg i.v on Day 1 and 7) on colon weight, determined 14 days after TNBS challenge.

[0073] FiG 14 shows the effects of CCI-7508 (50 and 100 mg/kg/day p o.), CCI-7507 (25 and 50 mg/kg/day p,o }, sulfasalazine (50 mg/kg/day p.o ) or infliximab (3 mg/kg i v on Day 1 and 7} on MPO levels sn the colon, expressed as mU/mg protein, determined 14 days after TNBS challenge

[00741 FIG. 15 shows the effects of CCt-7506 (50 and 100 mg/kg/day p.o }, CCi-7507 (25 and 50 mg/kg/day p.o.}. sulfasalazine (50 mg/kg/day p.o.) or infliximab (3 mg/kg i.v on Day 1 and 7} on TNF-a levels in the colon, expressed as pg/mg protein, determined 14 days after TNBS challenge.

Detailed Description of the Preferred Embodiments

[0075] A variety of substituent groups are utilized herein, including hydrogen and the groups identified herein. In addition, R groups on adjacent carbons may be joined together to form ring structures, including cycloaiky! and aryl groups. "Haio" as used herein refers to any suitable halogen, including -F, -Cl, -Br, and -I,

{0076] "Mercapto" as used herein refers to an -SH group.

[0077] "Azido" as used herein refers to an -N₃ group.

[0078] "Cyano" as used herein refers to a -CN group.

[0079] "Hydroxyl" as used herein refers to an -OH group

[0080] "Nitro" as used herein refers to an -NO₂ group.

[0081] "Oxy" as used herein refers to a -O- group.

J0082J "Oxo" as used herein refers to a =0 group.

[0083] "Alkyt" as used herein aione or as part of another group, refers to a straight or branched chain hydrocarbon containing from 1 to 10 carbon atoms. Representative examples of alkyl include, but are not limited to, methyl, ethyl, n-propyl, iso-propyl, n-butyl, sec-butyl, iso-butyf, tert-butyl, n-peπtyi. isopentyl. neopentyt. n-hexyl. 3-methyIhexyi, 2,2-dimethylpentyl. 2,3-dimethylpeπtyl, n-heptyl, π-octyl, n-nonyf, n-decyl, and the like "Loweralky!" as used nerem, is a subset of alkyl, in some embodiments preferred, and refers to a straight or branched chain hydrocarbon group containtng from 1 to 4 carbon atoms Representative examples of lower a!κy! include, but are not ϋmrted to methyl, ethyl n-propyϊ, iso-ρrop/1 n-butyl, sso-butyϊ, tert-buty! and the [skβ Alκγl ana ioweratk/f groups may be unsubscrtuted or substituted one or more times with R groups as defined herein including halo, aSkyf, haloalkyl, alkenyl, aikynyl, cycloalkyl, cyctoalkylatkyl, aryl, arylalkyl, heterocyclo, heterocyclcalkyl, hydroxyl, alkoxy, alkenytoxy, aikynyloxy, haloalkoxy, cydoalkoxy. cycloalkylalkyioxy, aryloxy, aryialkyloxy, heterocyciooxy, heterocyclolalkyloxy, mercapto, aikyl-S(O)m, haloalkyi-S(O)m, alkenyl-S(O)m, a)kynyS-S{O)rn. cycloafkyf-S(O)m, cycloaikylalkyl-S(O)m, aryl-SiOJm, arylalkyf-S(O)m, heterocyclo- S(0)m, heterocycloa!kyi-S(G)m. amino, afkylamino. alkeny!amino_: aikynyiamino, haioaikyiamino, cycloalkylamtno, cycloalkylalkylamino, arylamino, arytalkylamino, heterocycloamino. heterocycfoalkylamiπo, dϊsubstituted-amiπo, acylamino, acyloxy, ester, amide, sulfonamide, urea, alkoxyacylamino, amsnøacyloxy, nitro or cyano where m=0, 1 or 2.

[0084] "Alkenyl" as used herein alone or as part of another group, refers to a straight or branched chain hydrocarbon containing from 1 to 10 carbon atoms which include 1 to 4 double bonds in the normal chain. Representative examples of Afkenyl snclude. but are not limited to, vinyl, 2-propenyi, 3- butenyl, 2-butenyl, 4-pentyl, 3-pentyi. 2~hexenyi, 3-hexenyi, 2,4-heptadiene, and the like. These groups may be optionally substituted in like manner as described with alkyl above.

[0085J "Aikynyl" as used herein alone or as part of another group, refers to a straight or branched chain hydrocarbon containing from 1 to 10 carbon atoms which include 1 triple bond in the normal chain. Representative examples of Aikynyl include, but are not limited to, 2-propynyl, 3-butynyl, 2- butynyl, 4-pentenyl, 3-pentenyl, and the like. These groups may be optionally substituted in like manner as described with alkyi above.

[0086] "Alkoxy," as used herein alone or as part of another group, refers to an alkyl group, as defined herein, appended to the parent molecular moiety through an oxy group, as defined herein. Representative examples of alkoxy include, but are not limited to, rnethoxy, ethoxy, propoxy, 2- propoxy, butoxy, tert-butoxy, pentyloxy, hexyloxy and the like. These groups may be optionally substituted in like manner as described with alky! above.

[0087] "Acyl" as used herein aione or as part of another group, refers to a -C(O)R radical, where R is any suitable substituent such as alky!, alkenyl, aikynyl, aryi, alkylaryl, etc. as given herein.

[0088J "Haloalkyi," as used herein alone or as part of another group, refers to at least one halogen, as defined herein, appended to the parent molecular moiety through an aikyi group, as defined herein. Representative examples of haloalkyl include, but are not limited to, chloromethyi, 2-ffuoroethyi, triftuoromethyi, pentafluoroethyL 2-chloro-3-fiuoropentyi, and the like.

fO089] "ASkylihio," as used herein alone or as part of another group, refers to an alkyi group, as defined herein, appended to the parent molecuiar moiety through a thso moiety, Representative examples of alkyithio include, but are not limited, methylthro. ethylthio. tert-butylthio, hexylthϊo, and the Nke [0090] "Aryl," as used herein alone or as part of another group, refers to a monocyclic carbocycltc ring system or a bscyciic carbocycisc fused ring system having one or more aromatic rsngs Representative examples of aryl include, azulenyl, incfanyl. indenyl, naphthyl, phenyi, tetrahydronaphthyl, and the like These rings may be optionally substituted with groups selected from hato alkyl, haloalkyl, alkenyi. alkynyl, cycloalkyl, cycloaikylafkyl aryl, arylaikyi, heterocycio, heterocycloalkyl, hydroxyl, alkoxy, aikenyϊoxy, alkynyloxy. hatoalkoxy, cycioalkoxy cycloalkyialkyioxy, aryloxy, aryialkyloxy, heterocyclooxy heterocyclolalkyloxy rnercapto, alkyi-S(O)m, haloalkyI-S(O)m, alkenyl-S{O)m aikynyi-S(O}m, cycloa!kyl-S(O)m, cycloalkylalkyi-S(0)m aryl-S(O)m aryialkyl~S(O)m, heterocyclo-S(O)m heterocycloalkyl-S{O}m, amino, alkylammo alkenyjamino, alkynylamino, haloalkylammo, cycloalkylarrπno, cycloalkylalkylamino, arylamino, arylalkylamino, heterocycloamino, heterocycloatkylamino, disubstituied-ammo, acylamino acyloxy, ester, amide, sulfonamide urea alkoxyacyiamino, aminoacyloxy. πttro or cyano where m=0.1 or 2

[0091 ] "Arylaikyi," as used herein alone or as part of another group, refers to an aryl group, as defined herein, appended to the parent molecular moiety through an alkyl group, as defined herein Representative examples of arylaikyi include, but are not limited to, benzyl, 2-phertylethyl, 3- phenylpropyl, 2-naphth-2-ytethyl, and the like

[0092] "Amino" as used herein means the radical -NH₂

[0093] "Alkyiammo" as used herein alone or as part of another group means the radical -NHR, where R is an alkyl group

J0094] "Arylalkylammo" as used herein alone or as part of another group means the radical -NHR, where R is an arylalkyf group

[0095] "Disubstituted-amino" as used herein alone or as part of another group means the radical - NR_aR_b. where R_a and R_b are independently selected from the groups alky!, haloalkyl, aSkenyi, alkynyl, cycioalkyl, cycloalkylalkyl, ary! aryialkyl, heterocycio, heterocycloalkyl

[0096] "Acylamino" as used herein alone or as part of another group means the radical -NR_aR_b, where R_a is an acyl group as defined herein and R_b is selected from the hydrogen, alkyl haloaikyl, alkenyi, alkynyl, cycSoalkyi, cycloalkylalkyl, aryl, arylaikyi, heterocycio, heterocycloalkyl

[0097] "Acyloxy" as used herein atone or as part of another group means the radical -OR where R is an acyl group as defined herein

10098] "Ester" as used herein alone or as part of another group refers to a -C(O)OR radical, where R is any surtable substituent such as alkyl, ary), atkylaryl, etc [0099] "Amide" as used herein alone or as part of another group refers to a -C(O)NR₃R₆ radical, where R_a and R_b are any suitable substituent such as alkyl. aryl, alkylaryl etc

[00100] "Sulfonamide" as used herein alone or as part of another group refers to a -S{O}₂NR_βR_b radical, where R_a and R_b are any suitable substituent, such as H, alkyl, aryt, alkylaryl, etc,

[00101] "Sulfone" as used herein alone or as part of another group refers to a -S(O)₂R radical, where R is any suitable substituent. such as H, alkyl, aryi, alkylaryl, etc,

[00102] "Aminosulfonyr as used herein alone or as part of another group refers to a -N(R_a)S(O)₂R_b radical, where R_a and R_b are any suitable substituent, such as H, aikyl, aryl, alkylaryl, etc.

[00105] "Urea" as used herein alone or as part of another group refers to an -N(R_c)C(O)NR₃R₁, radical, where R_a, R_b and R_c are any suitable substituent such as H, alkyl, aryl, alkylaryl, etc,

[00104] "Alkoxyacylamino" as used herein alone or as part of another group refers to an - N{R_a)C(O)OR_b radical, where R_a, R_b are any suitable substituent such as H, alkyl, aryl. alkylaryl, etc.

[00105] "Aminoacyl" as used herein alone or as part of another group refers to an -C(O)NR₃Rb radical, where R_a and R_b are any suitable substttuent, such as H, aikyi, aryl. alkylaryl, etc.

[00106] "Amsnoacyloxy" as used herein aione or as part of another group refers to an - OC(O)NR_aR_b radical, where R_a and R_b are any suitable substituent, such as H, alkyl, aryl, alkylaryl, etc.

[00107] "Cycloalkyl," as used herein alone or as part of another group, refers to a saturated or partially unsaturated cyclic hydrocarbon group containing from 3. 4 or 5 to 6, 7 or 8 carbons (which may be replaced in a heterocyclic group as discussed below). Representative examples of cycloalkyl include, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyi. cydoheptyl, and cyciooctyi. These rings may be optionally substituted with halo or loweralky!.

[00108J "Heterocyclic group" or "heterocycle" as used herein alone or as part of another group, refers to a monocyclic- or a bicyclic-ring system. Monocyclic ring systems are exemplified by any 5 or 6 membered ring containing 1, 2, 3, or 4 heteroatoms independently selected from oxygen, nitrogen and suϊfur The 5 membered ring has from 0-2 double bonds and the 6 membered ring has from 0-3 double bonds. Representative examples of monocyclic ring systems include, but are not limited to, azetidine, azepine. azfridiπe, diazepine, 1,3-dtoxolaπe. dioxane. drthiane, furan, imidazole, imidazoline, imidazoline, isothiazole, isothiazotine, isothiazαlidine, isoxazole, isoxazoline, isoxazolϊdiπe, morpholine, oxadiazole. oxadiazoϋne, oxadiazolidine, oxazoie, oxazoline, oxazolidine, piperazine, piperidine. pyran, pyrazine, pyrazole, pyrazoline, pyrazohdine, pyridine, pyrimidine, pyπdazine, pyrrole, pyrroline. pyrrolidine, tetrahydrofuraπ, tetrabydrotbϊophene, tetrazine, tetrazole, thiadiazoie, feadrazofme, thiadsazoiidiπe. thsazoie, ihazolsne, thtazotosne, iNophene thsomorpholine. thsomorphohne sulfone thiopyran tπazme tπazole, tπthiane, and the like Bicyciic ring systems are exemplified by any of the above monocyclic ring systems fused to an aryl group as defined herein a cycloalkyl group as defined herein or another monocyclic ring system as defined herein Representative examples of bicyciic ring systems mcfucie but are not limited to for example benzimidazole benzothiazole benzothiadiazole, benzothiopheπe, benzoxadtazole, benzoxazole benzofuran benzopyran, benzothiopyran, benzodsoxine 1 3-benzodioxole cinnoline tndazole indole indoline indotizine, naphthyπdinβ isobenzofuran isobenzothiophene, isoindole, tsoiπdoline, isoqumoline phthalazine, purine, pyranopyπdsne, qumoline quiπolizme quinoxaline qumazoltne, tetrahydroisoqusnolme, tetrahydroquinolme, thsopyranopyπdine, and the like These rings may be optionally substituted with groups selected from halo, alkyl, haloalkyl, alkenyi, aikynyf, cycloalkyS, cycloalkylalkyl, aryl, arytalkyl, heterocyclo, heterocycSoaiky!, hydroxy!, alkoxy alkenyloxy, alkynyloxy, haloalkoxy, cycioalkoxy, cycloalkylalkyloxy, aryloxy, aryialkytoxy, heterocyclooxy, heterocyclolalkyloxy mercapto, a!ky!-S(O)m, ha!oalky!-S(O)m, alkenyt-S(O)m, alkynyi-S(O)m, cycloalkyϊ-S(G)m, cycloalkylalkyl»S{O)m aryl-S{O)m, aryia!kyl-S(O)m, heterocyclo-S(O)m, heterocycloaikyl-S{O)m amino atkylamiπo alkenyiamino alkynylammo haioalkylammo, cycloalkylamino cycloalkylalkylamino, aryjamino, arylaikyiarmno heterocycioamino, heterocycloalkylamino disubstituted-amino acyfamino, acyloxy ester amide sulfonamide, urea, aJkoxyacylamino amsnoacyloxy nitro or cyano where m=0 1 or 2

[00109J "Oxoheterocydic group" refers to a heterocyclic group such as described above, substituted with one or more oxo groups, such as pyπdιne-N-oxιde

JOOl 10] "Arylthio" as used heresn refers to a group of the formula -S-R₁ where R is aryl as described above

[00111] "Hydroxyamino" as used herein refers to a group of the formula -N(R)OH, where R is any suitable group such as aikyi, aryl, aikyiaryl, etc

[00112} "Treat" as used herein refers to any type of treatment that imparts a benefit to a patient afflicted with a disease, including improvement in the condition of the patient (e g in one or more symptoms}, delay in the progression of the disease, etc

[00113j "Inflammatory bowel disease" as used herein includes both Crohn's disease and ulcerative colitis

(00114} "Cancer" as used herein includes any cancer particularly solid tumors and includes but is not limited to lung cancer colon cancer breast cancer prostate cancer tiver cancer skin cancer ovarian cancer etc [OOllSJ "Pharmaceutically acceptable" as used herein means that the compound or composition is suitable for administration to a subject to achieve the treatments described herein without unduly deleterious side effects in fight of the seventy of the disease and necessity of the treatment

[00116) "Pharmaceutically acceptable prodrugs" as used herein refers to those prodrugs of the compounds of the present invention which are withsn the scope of sound medical judgment suitable for use in contact with the tissues of humans and lower animals without undue toxicity irritation aiiergic response and the like, commensurate with a reasonable risk/benefit ratio and effective for their intended use, as well as the zwitteπonic forms, where possible, of the compounds of the invention The term "prodrug" refers to compounds that are rapidly transformed in vivo to yield the parent compound of the above formuϊae for example by hydrolysis in blood A thorough discussion ss provided in T Higuchi and V Stella, Prodrugs as Novel delivery Systems VoI 14 of the A C S Symposium Series and in Edward B Roche, ed , Bioreversfbfe Carriers in Drug Design, American Pharmaceutical Association and Pergamon Press, 1987, both of which are incorporated by reference herein See a/so US Patent No 6,680,299 Examples include a prodrug that is metabolized in vivo by a subject to an active drug having an activity of active compounds as described herein, wherein the prodrug is an ester of an alcohol or carboxyisc acid group, if such a group is present in the compound, an acetal or ketal of an alcohol group if such a group is present in the compound, an N-Manmch base or an imine of an amine group, if such a group is present in the compound or a Schiff base, oxsme, acetal, enoi ester oxazolidsne, or thiazolidine of a carbonyl group if such a group is present in the compound, such as described in US Patent No 6 680 324 and US Patent No 6 680 322

[00117] Prodrugs of the present invention include esters or compositions as described in US Patent No 6,548,668 to Adams et al US Patent No 6 083,903 to Adams et al , or US Patent No 6 699,835 to Plamondon et al , the disclosures of which are incorporated by reference herein in their entirety

1. Active compounds.

[001 IS] Active compounds of the present invention (this term including pharmaceutically acceptable salts and prodrugs thereof) can be made in accordance with known techniques (see e g , U S Patent Ho 5,643, 893 to Benson et a/ ) or variations thereof which will be apparent to those skilled in the art based on the disclosure provided herein In some embodiments, active compounds of the present disclosure are compounds of Formula I or Formula Il

[00119] wherein:

[00120] A is N or C, subject to the proviso that R is absent when A is N;

[00121] X is, for Formula I, -C(O)-, -S(O)₂-, or a covalent bond more preferably -S(O)₂-, or a covalent bond, and X is, for Formula il, -C(O)-, -S(O)₂-, or a covalent bond;

(00122] Y is a linking group such as alkyt (e.g., -R- where R is C2-C6 alky!), alkenyi (e.g., -R- where R is C2-C6 alkenyl), cycloalkyl (e.g., -R- where R is C3-C6 cyctoalkyl). alkyicycloaikyKe.gr., -R-R'-, where R is C1-C4 alky! and R¹ is C3-C6 cycloalkyl). cylcoalkylalkyl (e.g., -R-R'-, where R is C3-C6 cycloalkyl and R' is C1-C4 aikyl), alkylcycloalkylalkyl (e.g., -R-R'-R"-, wherein R is C1-C4 alkyl, R' is C3-C6 cycloalkyl, and R" is C1-C4 alkyl), alkyioxyalkyl (e.g., -R-O-RS wherein R and R' are C1-C4 alkyl}; aryl (e.g., -R- where R is aryl), alkylaryl (e.g., -R-R'- where R is C1-C4 alkyl and R' is aryl), alkylaryiaSkyl (e.g.. -R-R^r-R"- where R is C1-C4 aikyl, R' is ary!, and R" is C1-C4 aikyl), or aryialkyl (e.g., -R-R'- where R is aryl alkyl and R' is C1-C4 alkyl); cycϊoalkylaikyϊ (e.g. -R-R'-, where R is C3-C6 cycloaikyl and R^* is C1-C4 alkyl), alkylheterocycte (e.g., -R-R', where R is C1-C4 aikyl and R' is a heterocyclic group as described herein), heterocyclealkyl, alkylheterocyclealkyi, heterocycle, amtnoalkyl (e g . -N(R)R'-, where R ss H or C1-C4 alkyl and R¹ ss C1-C4 alky!), oxyalkyl (e g,, -O-R- whβre R is C2-C6 aϊkyϊ), amiπoaryl (e.g., -N(R)R'-, where R is H or C1-C4 alkyl and R¹ is aryl), and oxyaryl (e.g., -O-R-. where R is aryl), and [00123] Z is selected from the group consisting of -B(OR¹)OR^Z, -CON(R¹)OR², and -r^OR^COR² or any of the ackiittonal alternatives for Z described tn greater detail below

[00124] R¹ and R² are each independently H, ioweralkyl, or together form C2-C4 alkylene, and

[00125] R³ R⁴, R⁶ R⁶ and R' are each independently selected from the group consisting of H₁ halo (oweralkyl, halofoweralkyl, haloloweralkoxy ioweralkoxy, hydroxy, lowerafkoxycarbo, carboxylic acid, acyl azido mercapto, alkylthio amino, heterocycleamino, alkylamino, dialkylamiπo acylamino, aminoacyl, aryfamino arylalkyl arylalkytarri!no aryloxy cyaπo, sulfonamide, amtnosulfonyl suifone, nitro arylalkyloxy cycloalkyloxy cycloalkylalkoxy cycloaikylamino urea cycloalkylalkylammo cycloaikyi alkylcycfoalkyl, hydroxyamino alkoxyacylamtno and arylthio and 5- or 6- membered organic rings containing 0 to 4 heteroatoms selected from the group consisting of N O and S which rings may be unsubstitυted or substituted from 1 to 4 times with halo, ioweralkyl haloloweralkyl, haloloweralkyioxy, ioweralkoxy, hydroxy, loweraikoxycarbo, carboxylic acsd acyl, azido, mercapto, aikylthio amino, heterocycleamsno, alkylamino, dfalkylamtno, acyiamino, aminoacyl, arylamino, arylalkyl, arylalkylamino, aryloxy, cyano, sulfonamide, aminosulfonyl, sulfoπe, nitro, and oxoheterocyclic groups, or a pharmaceutically acceptable salt or prodrug thereof

[00126] In some embodiments, R³ is preferably not H Thus in some embodiments R³ is preferably a 5- or 6- membered organic ring containing 0 to 4 heteroatoms selected from the group consisting of N, O and S which ring may be unsubsiitυted or substituted from 1 to 4 times with halo, cycloalkylalkoxy ioweralkyl, haloloweralkyl, haloloweraikyloxy, loweralkoxy, hydroxy, loweraikoxycarbo, carboxylic acsd, acyl, azido, mercapto alkylthto, amino, heterocycleamino, alkylamino, dsatkylamino, acylamino, aminoacyl, arylamino, arylalkyl, arylalkylamino, aryloxy, cyano, sulfonamide, amsnosulfonyl, suifone, nttro and oxoheterocyclic groups

[00127] It will be understood that in Formula Il where R³ is bonded to the ring nitrogen it ss iess preferred for R³ to be halo azido mercapto, ammo, alkylamino, dialkylamino, acylamino cyano and arylalkylamino and more preferred for R³ to be alkyl, Ioweralkyl and haloloweralkyl suifone, amide and aryl

[00128] R^s (S prefeably selected from the group consisting of halo, loweralkyl, haloloweralkyl haioloweralkyloxy, loweralkoxy, hydroxy loweraikoxycarbo carboxyltc acid, acyi, azido, mercapto, alkylthto amino heterocycfeamsno alkylamino dialkylamino, acylamino, amtnoacyi, arylamtno, arylaikyi, arylaikyϊamino aryioxy cyano, sulfonamide amsnosulfonyl , suifone, and nitro R⁵ ts more preferably selected from the group consisting of halo haloioweralkyl, haloloweralkyioxy loweralkoxy, ammo acylarniπo, amtnoacyi, arylafkyl, aryloxy acyl, arylamino, cyano, mtro and heterocycleamino R^s is most preferably cyano, fluoroaSkyl or halo 100129] R⁴ is in some embodiments preferably H In other embodiments R⁴ is preferably selected from the group consisting of halo, Soweralkyl, hateioweralkyl, haioloweralkyloxy, ioweraikoxy hydroxy, loweralkoxycarbo, carboxylic acid acyl, azido, rnercapto, alkyithio, amino, heterocycteamino, alkylamsπo, dialkyiamino, acyiarrnno aminoacy!, arylaminα, arylaikyi, arylalkylamsno. aryloxy, cyano, sulfonamide, aminosulfonyi sulfone, and mtro, more preferably R⁴ is selected from the group consisting of halo haloloweralkyl, haloloweralkytoxy_b loweralkoxy, amtno, acylamtno aminoacyi, arylalky) aryloxy, acyl, arylamtno, cyano, nrtro. and heterocycleamino and sttil more preferably R⁴ is cyano fluoroalkyl or halo

(00130] In some embodiments R⁶ is H in other embodiments R⁶ is preferably selected from the group consisting of halo loweralkyl haloioweralkyl, haloloweralkyϊoxy loweralkoxy hydroxy loweralkoxycarbo carboxylic acid acy!, azido, mercapto alkylthio, amino, heterocycleamino alkylamino, dialkylamino acyiamino, aminoacyf arylamino, arylalkyl arylalkylammo, aryloxy cyano, sulfonamide, ammosulfonyl, sulfone, and mtro, in such other embodiments R^e is more preferably selected from the group consisting of halo, haioloweralkyi, haSoloweralkySoxy, loweraikoxy, ammo, acyiamino aminoacyi, arylalkyl, aryloxy, acy!, arylamsno, cyano, mtro, and heterocycleamino, in such other embodiments R⁶ ss most preferably cyano, fluoroaikyi or halo

[00131] In some embodiments, at least two of R⁴ R⁶, and R⁷ are H In some preferred embodiments R⁶ and R⁷ are both H In some preferred embodiments R⁷ ts H

[00132] Particularly preferred examples of compounds of the present invention are

[00133] 4-{2-(Tπfluoromethyl)-1 H-benzo{d]tmιdazol-1 -yl)butylboromc acid,

[00134] S^^ThiazoW-yO-IH-benzotcOimidazol-i-yOpentylboronic acid,

[00135] 5~(5,6~dιmethyl-1W-benzo[d]imidazol-1-y!)ρeniylboronιc acid

[00136J 5-(1H-ιmιdazo[4,5-c]pyrιd!n-1-yl)pentylboronic acid

[00137] 5-(2-H-Methoxyphenyl)-1 H-benzo[d]ιmidazoM~yl)pentylboronιc acid

[00138] 5-(2"(3~Fiuoro-4-methoxyphenyl)-1H-benzo[d]tmιdazol-1-yl)penty(boronϊC acid,

(00139] 5-{5-c/ano-2-(4-methoxyphenyl)-1 H-benzo[d]ϊmidazoS-1-yl)pentyiboron!c actd,

[00140] 5-(6-cyano-2-(4-methoxyρhenyl)-1 H-benzo[d]inrndazo!~1-yS)pentylboronιc acsd,

[00141] and pharmaceuticaϋy acceptable salts and prodrugs thereof [00142] Another aspect of the present disclosure are compounds of Formula ill. FormuialV or Formula V:

[00143) whereirr

J00144] X is, for Formula II!, -C(O)-, -S(O)₂-, or a covalent bond, more preferably -S(O)₂-. or a cQvalent bond, and X is, for Formulas IV and V, -C(O)-, -S(O)₂-, or a covalent bond:

[00145J Y is a linking group such as alkyl {e g., -R- where R is C2-C6 alky!), alkenyl {e.g , -R- where R is C2-C6 aikenyi), cycloatkyl (e g., -R- where R is C3-C6 cycloalkyl), alkylcycioa!kyl{e.g._: -R-R¹-. where R is C1-C4 alkyl and R¹ is C3-C6 cycloalkyl), cylcoaikyiafkyl (e.g., -R-R¹-, where R is C3-C6 cycioaikyi and R' is CA-CA alkyl). alkyicyciostkyfalkyi (e.g . -R-R'-R"-. wherein R ss C1-C4 afkyi, R^* ^s C3-C6 cycloaikyi, and R" is C1-C4 alky!), aikyioxyaikyl (e.g., -R-O-R'-, wherein R and R' are C1-C4 alky(); aryl (e.g., -R- where R is aryl), atkyiaryi (e.g., -R-R'- where R is C1-C4 alkyl and R' is aryi). alkylarylaikyl (e.g., -R-R'-R"- where R is C1-C4 alky!, R' is aryf, and R" is C1-C4 alkyl), arylalkyl (e.g., - R-R'- where R is aryi aikyl and R¹ is C1-C4 alkyl), cycloaϊkyialkyl (e.g. -R-R'-, where R is C3-C6 cycloaikyi and R' is C1-C4 alkyl}, alkylheterocycle (e.g., -R-R', where R is C1-C4 aikyf and R' is a heterocyclic group as described herein), heterocyciealkyl, aikyiheterocyclealkyi, heterocycle, amϊnoalkyl (e.g., -N(R)R'-, where R is H or C1-C4 alkyl and R' is C1-C4 alkyl), oxyalkyl (e.g., -O-R- where R is C2-C6 alkyl}, amiπoaryl (e.g., -N(R)R'-, where R is H or C1-C4 alky! and R' is aryi), or oxyaryl (e.g., -O-R-, where R is aryi); and

[00146) Z is selected from the group consisting of -B(OR¹ )OR², ~CON(R¹)OR², and -N(OR¹JCOR² or any of the additional alternatives for Z described in greater detail below.

|00147] R¹ and R² are each independently H, loweralkyl, or together form C2-C4 alkylene; and

[00148] R³, R⁴, R⁵, R⁶, and R⁷ are each independently selected from the group consisting of: H, halo, loweralkyl, haloioweralkyl, haloloweralkoxy, loweralkoxy, hydroxy, loweralkoxycarbo, carboxyϋc acid, acyl, azido, mercapto, alkylthio, amino, heterocyc learn mo, alkyiamino, dialkylamino, acylamino, aminoacyl, aryiamino, arylalkyl, arylaikylamino, aryloxy, cyano, sulfonamide, aminosulfonyl, sulfone, nitro arylalkyloxy, cycloalkyloxy, cycloalkylalkoxy, cycioalkyiaminα, urea, cycloalkylaϊkyiamino, cycloaikyi, alkylcycloalkyl, hydroxyamino, alkoxyacylamino, and arylthio; and 5- or 6- membered organic rings containing 0 to 4 heteroatoms selected from the group consisting of N, O and S, which rings may be unsubstituted or substituted from 1 to 4 times with halo, loweraikyt, haloloweralkyl, haioloweralkyloxy, loweralkoxy, hydroxy, loweralkoxycarbo, carboxylic acid, acyl, azido, mercapto, alkylthio, amino, heterocycieamino, alkyiamino, dialkylamino, acylamino, aminoacyl, aryiamino, arylalkyl, arylalkyiamino, aryloxy, cyano, sulfonamide, aminosulføny), sulfone, nitro; and oxoheterocycfic groups; or a pharmaceutically acceptable salt or prodrug thereof.

[00149] In some embodiments of compounds of Formulas IN, IV and V, R^£ is selected from the group consisting of: halo, loweralkyl, haloloweralkyi, haloloweralkyioxy, loweraikoxy, hydroxy, ioweralkoxycarbo, carboxylic acid, acyf, azido. mercapto, aikylthio, amino, heterocycieamino, alkyiamino, dialkyiamino, acyfamino, aminoacyl, arylamino, arylalkyl, arylaikylamino, aryloxy, cyano, sulfonamide, aminosulfonyϊ, sulfone, and nitro, more preferably R⁵ is selected from the group consisting of: halo, nalolαwerafkyi haloloweraikyfoxy, foweralkoxy, amino, acyjamino, aminoacyl, arylalkyl. aryloxy. acyi, aryiamino. cyano, nitro, and heterocycieamino, and most preferably R^s is cyano, fluoroalkyl or halo.

[00150] In some embodiments of compounds of Formulas III, IV and V, R* is H; in other embodiments R⁴ is selected from the group consisting of' halo, loweraikyS, haioloweralkyi, haϊoloweraϊkyioxy, ioweralkoxy, hydroxy, ϊoweraikoxycarbo, carboxyϋc acid, aeyl, aziύo, mercapto, alkylthio. amino, heterocycleamino, alkylam^'tno, dialkylamino_h acylamino, aminoacyl, arylammo, arylatkyt. arylalkylamino, aryloxy, cyano, sulfonamide, aminosulfoπyt. suifone, and nitro. more preferably from the group consisting of' halo, haloloweralky). haiofoweralkyloxy. Soweralkoxy, amino, acyiamiπo, aminoacyi. arylaikyl, aryloxy, acyl. arySamino, cyano, nitro, and heterocycleamino: and most preferably cyano, fluoroaikyl or halo

[00151} In some embodiments of compounds of Formuias III, IV and V, R⁶ is H; in other embodiments R⁶ is selected from the group consisting of: halo, loweralkyi, haloioweralkyS, haloloweralkyloxy, loweralkoxy, hydroxy, loweraikoxycarbo, carboxylic acid, acyl, azido, mercapto, alkylthio, amino, heterocycleamino , alkylamino, dialkylamino, acylamino, aminoacyl, arylamino, arylaikyl, arylalkylamino, aryloxy, cyano. sulfonamide, aminosulfonyl, suifone, and nitro; more preferably halo, haloloweralkyl, haloloweralkyloxy, loweralkoxy, amino, acylamino, aminoacyi, arylalkyl, aryloxy, acyl, arylamino, cyano. nitro, and heterocycleamino; and most preferably cyano, fluoroaikyl or halo.

[00152] In some embodiments of compounds of Formulas III, IV and V, R⁷ is H; in other embodiments R⁷ is selected from the group consisting of : halo, loweralkyi, haloloweralkyl, haloloweralkyloxy, ioweralkoxy. hydroxy, loweralkoxycarbo, carboxyiic acid, acyl, azido, mercapto, aikylthio, amino, heterocycleamino, alkylamino, dialkylamino, acylamino, aminoacyl, arylamino, arylalkyl, arylalkylamino, aryloxy, cyano, sulfonamide, aminosulfonyl, suifone, and nitro, more preferably halo, hatoloweralkyl, haioloweralkyioxy, loweraikoxy, amino, acylamino, aminoacyi, arylalkyl, aryloxy, acyl, arylamino, cyano, nitro, and heterocycleamino, and most preferably cyano, fluoroaikyl or halo.

[00153] In some embodiments of compounds of Formulas IiI₁ IV and V, at least two of R⁴, R⁶, and R⁷ are H. For example, in some embodiments R⁶ and R⁷ are H; in other embodiments R⁴ and R⁶ are H; in other embodiments R^≤ and R⁷ are H; in still other embodiments R⁴ and R⁵ are H.

[00154] In yet another aspect of the present disclosure are compounds of Formu Sa VI:

|00155] wherein

J001S6] A is S, O, SO₂ Or NR

[00157] X ss -C(O)- -S(O)₂- or a covaient bond

[00158] Y is a linking group such as afkyl (e g -R- where R is C2-C6 alky⁾), alkenyl (e g -R- where R is C2-C6 aikenyi) cycloalkyl {e g , -R- where R is C3-C6 cycloalkyl) alkylcycloalkyl(e g , -R-R¹- where R ss C1-C4 a!ky! and R¹ is C3-C6 cycloalkyl) cylcoalkytalkyi (e g , -R-R'- where R is C3-C6 cycloaikyl and R' is C1-C4 alkyl) aikyϊcycloalkylaikyl (e g , -R-R'-R"- wherein R is C1-C4 alkyl, R' is C3-C6 cycϊoaikyf and R" is C1-C4 alkyl) alkyloxyaikyl (e g -R-O-R'-, wherein R and R' are C1-C4 alkyl), aryl (e g , -R- where R is aryl), alkylaryl (e g , -R-R'- where R ts C1-C4 alkyl and R' is aryl), alkylarylalkyl (e g , -R-R'-R"- where R is C1-C4 aikyt, R' ss ary!, and R" ts C1-C4 aSkyi), aryialkyl (e g , - R-R'- where R is aryl alkyl and R' ss C1-C4 alkyl), cycloaikyiaSkyf (e g -R-R'- where R is C3-C6 cycioalkyl and R' is C1-C4 alkyl), alkylheterocycle (e g , -R-R', where R is C1-C4 alkyl and R' is a heterocyclic group as described herein), heterocyclealkyi, alkylheterocyceialkyl, heterocycle, aminoaikyl (e g , -N(R)R'-, where R is H or C1-C4 alkyl and R' ss C1-C4 alky!), oxyaikyi (e g , -O-R- where R is C2-C6 alkyl), aminoary! (e g , -N(R)R'-, where R is H or C1-G4 alkyi and R' is aryl), and oxyaryl (e g , -O-R-, where R is aryl), and

[00159] Z ss selected from the group consisting of -B(0R¹)0R², -CON(R¹)OR², -N(OR¹)COR², or any of the additional alternatives for Z described in greater detail below

(00160} In some embodiments of Formula Vl, at least one of R³, R⁴, R⁵, R⁶, R⁷ or R⁸ is not H

[00161 ] In some embodiments of Formula V! R⁵ is selected from the group consisting of halo, lowerajkyl, haloloweralkyi, haSoloweralkyloxy loweralkoxy, hydroxy, loweralkoxycarbo, carboxylic acid, acyl, azido, mercapto, alkylthio, ammo, heterocycleamsno, alkylamsno, diaikyiamino, acylamino, aminoacyl, arylammo, aryialkyl, arylalkylamino, aryioxy, cyano, sulfonamide, ammosutfonyl sυlfone, nttro and hydroxyamino In more preferred embodiments, R⁵ is selected from the group consisting of halo haloioweralkyl, haloloweralkyloxy, loweralkoxy, ammo, acyiamino, amiπoacyl aryialkyl aryioxy, aaccyyii aarryyllaammmmoo ccyyano, nitro, and heterocycleamtno In still more preferred embodiments, R⁵ is cyano, fluoroalkyl or halo

[00162] in some embodiments of Formula Vl R⁴ is H In other embodiments of Formula Vl, R⁴ is selected from the group consisting of halo joweralkyl, haioioweraikyi halolowerafkytoxy, loweralkoxy hydroxy loweralkoxycarbo, carboxylic acid acyl, aztdo mercapto, alkylthso amino heterocycieamino alkylamirso dsaSkylamsno, acyiamino amsnoacyl, arylammo aryialkyl, aryiatkylamsno aryioxy cyano sulfonamide aminosulfonyl sulfone nsfro and heterocycleamtno more preferably R⁴ is seSected from the group consisting of halo haiofoweralkyl ha⁽oio#era!f<yfoxv loweralkoxy amsno acyiamtπo aminoacyi. arylalkyl, aryloxy, acy!, arylarrϋno, cyano, nitro, and heterocycieamino, and most preferably R⁴ is cyano fSuoroaikyi or halo

[00163] In some embodiments of Formuia Vl, R⁶ is H in other embodiments R⁶ is selected from the group consisting of halo, ioweraikyl, haloloweratkyl, haioloweraikySoxy, ioweralkoxy hydroxy, loweralkoxycarbo, carboxyitc acid acyl, azido mercapto, alkyithio amino heterocycieamino, alkylamtno dialkylamsno, acylamino, aminoacyi, aryiamiπo, arylalkyl, aryialkylarmrto, aryloxy cyano, sulfonamide, aminosulfonyl, suifone, and nitro, more preferably halo haϊoloweralkyl, haløloweraSkyloxy Ioweralkoxy amino, acylamino, aminoacyi, arylatkyl, aryloxy, acyl, arylamino, cyano nttro and heterocycieamino, and most preferably R^e is cyano fluoroalkyi or halo

[00164] In some embodiments of Formuia Vl, R⁷ is H In some preferred embodiments at least two of R⁴ R^e, and R⁷ are H In some stiii more preferred embodiments R⁶ and R⁷ are H

f0016S] In some embodiments R ts selected from the group consisting of H, loweralkyi, haloloweralkyl, haloioweralkyloxy, Ioweralkoxy, loweralkoxycarbo, carboxyhc acid, acyl, acylamino, aminoacyi, arylafkyl, cyano, sulfonamide, aminosulfonyl, and sulfone, more preferably H, loweralkyl, haloloweraikyl, haloioweralkyioxy, Soweralkoxy, SoweraSkoxycarbo, and arylalkyl

[00166] In some embodiments R³ is selected from the group consisting of H, alkyl, aryl, heteraryl, and cycioalkyl

[00167] In some embodiments R⁸ and R⁹ are each independently selected from the group consisting of H and ioweraikyl or R^s and R⁹ are together =0 or =S

[00168 J In some embodiments R⁹ and R¹⁰ are both H

[00169] Examples of particularly preferred compounds of Formula Vl include but are not limited to

[0017Oj 5~(6-fluoro-2>dihydro-3-oxobenzo[b][1 4]oxazιn-4-yi)pentylboronιc acid 5~(2,3-dthydro-3- oxobenzo[b][1,4Jthiaztn-4-y|)penfylboronιc acid, 5-{7-chloro-2 3-dihydro-3-oxobenzo[b][1 ,4]tNazιn-4- y!)perrtylboromc acid, 5-{2 3-dthydro-7-nrtro-3-oxobenzo[b][1,4]oxazin-4~y!)pentylboromc acid, 5-(2 3- d&hydro~3-oxobenzofb3[1 4]oxazιn-4-yl}pentylboron:c acid ethyl 2-(3,4-dιhydro~3~oxo-4-(5- pentyiboronsc acιd)-2H-benzo[b][1 ,4]truazin-2-y!)acetate and pharmaceutically acceptable salts and prodrugs thereof

[00171 j In some embodiments, active compounds of the present disclosure are compounds of Formuia VII

JΘ0172] wherein

[00173] A₁ and A₂ are each independently N or C

[00174] X is -C{0)-, -S(O)₂-, or a covaient bond,

[00175J Y is a isnking group such as alkyl (e.g., -R- where R is C2-C6 alky!), alkeπyl {e.g., -R- where R is C2-C6 alkenyl), cycloalky! (e.g., -R- where R is C3-C6 cycioaikyl), alkylcycloalkyl(e.g., -R-R'-, where R is C1-C4 atkyt and R¹ is C3-C6 cycloalkyl), cylcoalkyialkyl (e.g., -R-R¹-, where R is C3-C6 cycioaikyl and R¹ is C1-C4 alkyi). alkylcycloaikylalkyl (e.g., -R-R'-R"-, wherein R is C1-C4 alkyl, R' is C3-C6 cycioaikyl, and R" is C1-C4 alkyl}, alkyloxyalkyl (e.g., -R-O-R'-, wherein R and R' are C1-C4 alkyl); aryl (e.g., -R- where R is aryl), alkylaryl (e.g., -R-R'- where R ss C1-C4 alkyl and R' is aryl), alkyiarylalky! (e.g., -R-R'-R"- where R is C1-C4 alkyl, R' is aryl, and R" is C1-C4 aikyt), or arylalkyl (e.g., -R-R'- where R is aryl alkyl and R' is C1-C4 alkyl), cycloaikylaikyl (e.g. -R-R'-, where R is C3-C6 cycloalkyl and R' is C1-C4 alkyl), alkyϊheterocycte (e.g , -R-R', where R is C1-C4 alkyi and R' is a heterocyclic group as described herein), heterocycfealkyl, alkySheterocycleaikyi, heterocycie, amsnoaikyl (e.g., -N(R)R¹-, where R is H or C1-C4 alkyl and R' is C1-C4 alkyl), oxyalkyl (e.g., -O-R- where R is C2-C6 alkyl), aminoaryl {e.g.. -N(R)R'-, where R is H or C1-C4 alkyl and R' is aryl), and oxyaryl (e.g , -O-R-. where R is aryl); and

[00176] 2 is selected from the group consisting of -B{OR¹)OR², -CON(R¹JOR², and -N(OR¹JCOR² or any of the additional alternatives for 2 described in greater detail below.

[00177] R¹ and R² are each independently H, loweralkyl, or together form C2-C4 alkylene; and

[00178] R_n, and R_p are each independently selected from the group consisting of: H, halo, loweralkyl, haloloweraSkyl, haloiowerafkoxy, ioweralkoxy. hydroxy, loweralkoxycarbo, carboxylic acid, acyl, azido, mercapfo, aϊkytthio. amino, heterocyclearruno, aikyiammo diaiky'amϊn-o, acyfamtno, amirtoacyl, arylarruna. arylalkyl aryiafkyfammo aryioxy, cyano, sulfonamide, amsnosuffonyϊ. sulfoπe, πftrø, arylalkyloxy, cycioalkyloxy, cycloaikylalkoxy, cycloalkylamino, urea, cycloalkylalkySamino, cycioalkyi. aikylcycloaSkyi, hydroxyamsno, afkoxyacytamino, and arylthio;and 5- or 6- membered organic rings containing 0 to 4 heteroatoms selected from the group consisting of N, O and S, which rings may be unsubstituted or substituted from 1 to 4 times with halo, ioweralkyl, haloloweralkyl, haloloweraikyloxy, loweralkoxy, hydroxy, loweraikoxycarbo, carboxylic acid, acyS, azido, mercapto, aikylthio, amino, heterocycleamino, aikyiamino, dialkylamino, acyiamino, aminoacyS, aryiamino, arylalky!, arylalkylamino, aryloxy, cyano, sulfonamide, amiπosulfbnyl, sulfone, nitro; and oxoheterocyclic groups; subject to the proviso that when A₁ is C₁ then n=1 to 4; when Ai is N, then n=1 to 3; A₂ is C, then p= 1 to 2; when A₂ is N, then n=1; or a pharmaceutically acceptable sait or prodrug thereof.

(OOΪ 79| In addition, compounds of the present invention include compounds of Formulas I₁ 11_« HI. IV. V_h VI₁ and VII, and others above in which substituent -Z is a group of the formula:

O I -NH

HN-N^' H₂N b [0018Oj In addition, compounds of the present invention iπcϊude compounds of Formulas I₁ H, II!, IV. V, VS, and VII, and others herein subststuent -Y-Z ss a group of the formula;

[00181] In addition, compounds of the invention include compounds of Formulas I. II, ill, IV₁ V_: Vl. and VII, and others herein the groups -X-Y-Z are a substituent of the formula:

NH NH CF₃O₂S PhO₂S

[00182] In addition, compounds of the invention include compounds of Formulas I, M₁ III, IV, V, Vl₁ and ViI₁ and others herein, the groups -X-Y-Z represent a substituent of the formula:

[00183] in addition, compounds of the invention include compounds of Formulas 1, II, III, N, V, Vi, and VH₁ and others herein, group -Z is a substituent of the formula

Oxyguanidinc^, cvctic oxyguaπidines

OH

N OH NH,

M

[00184] In addition compounds of the invention includes compounds of the Formulas I Ii, ill, IV V ! and VH and others herein group -Z is a substituent of the formula

85] Examples of active compounds of the present invention include but are not limited to:

(4)

(21)

(23)

(52)

(54)

{71}

(74}

(78)

(83)

(97)

[00186] The active compounds disclosed herein can, as noted above, be prepared m the form of their pharmaceutically acceptable salts Pharmaceutically acceptable salts are salts that retain the desired biological activity of the parent compound and do not impart undessred toxicoiogical effects Examples of such salts are (a) actd addition salts formed with inorganic acids for example hydrochloric acid hydrobromic acsd sulfuric aσd phosphoric acid nrtric acid and the like, and salts formed with organic acids such as, for example acettc actd, oxalic acid, tartaric acid, succinic acid maleic acid fumaric acid, gluconic actd, citric acid mafsc acid ascorbic actd benzoic actd tannic aod, palmitic acid aSgtmc acid, polyglutamsc acid, naphthatenesulfonic acid, methanesuifontc acid, p- ioluenesυifonic actd, πaphthafersedisuifonic actd poiygalacturonic acid and the Mke (b) salts formed from elements! antoπs such as chlorine, bromine and rødfne and (c) sails demed from bases such as ammonium salts alkali metal saSts such as those of sodium and potassium aikaisne earth metai salts such as those of calcium and magnesium and salts with organic bases such as dicyciohexyiamine and N-methyl-D-giucamsne

2. Pharmaceutical formulations,

[00187] The active compounds described above may be formulated for administration in a pharmaceutical earner in accordance with known techniques See, inter alia Remington The Science and Practice of Pharmacy , 21^th Ed Mack Publishing Co , Easton, PA (2006) and Handbook of Pharmaceutical Excspients, 3rd Ed, Kibbe A H ed , Washington DC, American Pharmaceutical Association (2000) hereby incorporated by reference in their entirety In the manufacture of a pharmaceutical formulation according to the invention the active compound (including the physiologically acceptable salts thereof) ss typically admixed with, inter alia, an acceptable carrier The carrier must of course be acceptable in the sense of being compatible with any other ingredients in the formulation and must not be deleterious to the patient The carrier may be a solid or a liquid or both and is preferably formulated with the compound as a unit-dose formulation for example a tablet, which may contain from 0 01 or 0 5% to 95% or 99% by weight of the active compound One or more active compounds may be incorporated in the formulations of the invention which may be prepared by any of the well known techniques of pharmacy consisting essentially of admixing the components optionally including one or more accessory ingredients

[00188} The formulations of the invention include those suitable for oral, rectal, topical, buccal (e g sub-lingual) vaginal, parenteral (e g , subcutaneous intramuscular intradermal or intravenous), topical (/ e both skin and mucosal surfaces, including airway surfaces) and transdermal administration although the most suitable route in any given case wilt depend on the nature and seventy of the condition being treated and on the nature of the particular active compound which is being used

[00189] Formulations suitable for oral administration may be presented tn discrete units, such as capsules cachets, lozenges or tablets each containing a predetermined amount of the active compound, as a powder or granules as a solution or a suspension in an aqueous or non-aqueous liquid or as an oil-in-water or water-ιπ-oιl emulsion Such formulations may be prepared by any suitable method of pharmacy which includes the step of bringing into association the active compound and a suitable earner (which may contain one or more accessory ingredients as noted above! In general the formulations of the invention are prepared by uniformly and intimately admixing the active compound with a liquid or finely divided soifd earner or both and then if necessary shaping the resulting mixture For example a tablet may be prepared by compressing or molding a powder or granules containing the active compound optionally wsth one or more accessory ingredients Compressed tablets mav be prepared by compressing m a suitable machine tπe compound in a free- flowing form, such as a powder or granules optionally mixed with a binder, lubricant, inert diluent, and/or surface active/dispersing agent(s) Molded tablets may be made by molding sn a suitable machine the powdered compound moistened with an inert liquid binder

(00190) Formulations suitable for buccal (sub-lingua!) administration include lozenges comprising the active compound in a flavoured base, usually sucrose and acacia or tragacanth and pastilles comprising the compound sn an inert base such as gefatin and giycerin or sucrose and acacia

[00191] Formulations of the present invention suitable for parenteral administration comprise sterile aqueous and non-aqueous injection solutions of the active compound which preparations are preferably isotonic with the blood of the intended recipient These preparations may contain antioxidants buffers bacteπostats and solutes which render the formulation isotonic with the biood of the intended recipient Aqueous and non-aqueous sterile suspensions may include suspending agents and thickening agents The formulations may be presented sn umfΛdose or multi-dose containers, for example sealed ampoules and vials, and may be stored in a freeze-dned (lyophihzed) condition requiring only the addition of the sterile liquid earner, for example, saline or water-for-mjection immediately prior to use Extemporaneous injection solutions and suspensions may be prepared from sterile powders, granules and tablets of the kind previously described For example, in one aspect of the present invention, there is provided an injectable, stable, sterile composition comprising a compound of Formula I, Il III, IV or V, or a salt thereof, in a unit dosage form in a sealed container The compound or salt is provided in the form of a lyophilizate which is capable of being reconstituted with a suitable pharmaceutically acceptable earner to form a liquid composition suitable for injection thereof into a subject The unit dosage form typscaliy comprises from about 10 mg to about 10 grams of the compound or salt When the compound or salt is substantially water-insoluble a sufficient amount of emulsifying agent which is physiologically acceptable may be employed in sufficient quantity to emulsify the compound or salt in an aqueous carrier One such useful emulsifying agent is phosphatidyl choline

[00192 J Formulations suitable for rectal administration are preferably presented as unit dose suppositoπes These may be prepared by admixing the active compound with one or more conventional solid earners, for example, cocoa butter, and then shaping the resulting mixture

[00193J Formulations suitable for topical application to the skin preferably take the form of an ointment, cream, lotion, paste gel. spray, aerosol, or oil Garners whrch may be used include petroleum jelly, lanoline polyethylene glycols, alcohols, transdermal enhancers and combinations of two or more thereof in some embodiments the compositions described herein can be administered from an inhafer through the mouth or nasai passage for pulmonary delivery

[00194} Formulations suitable for transdermal administration may be presented as discrete patches adapted to remain <n intimate contact wsth the epidermis of the recipient for a prolonged penod of time Formulations suitable for transdermal administration may also be delivered by iontophoresis (see, for example Pharmaceutical Research 3 (6) 318 (1986)) and typically take the form of an optionally buffered aqueous solution of the acttve compound Suitable formulations comprise citrate or btsUris buffer (pH 6) or ethanol/water and contain from 0 1 to 0 2M active ingredient

[00195] Further, the present invention provides liposomal formulations of the compounds disclosed herein and salts thereof The technology for forming liposomal suspensions is well known in the art When the compound or salt thereof is an aqueous-soluble salt, using conventional liposome technology, the same may be incorporated into lipid vesicles In such an instance due to the water solubility of the compound or salt the compound or salt will be substantially entrained within the hydrophilsc center or core of the liposomes The lipid layer employed may be of any conventional composition and may either contain cholesterol or may be cholesterol-free When the compound or salt of interest is water-insoluble, again employing conventional liposome formation technology, the salt may be substantially entrained within the hydrophobic lipid bilayer which forms the structure of the liposome In either instance, the liposomes which are produced may be reduced in size as through the use of standard sonication and homogenizatson techniques Liposomal formulations containing the compounds disclosed herein or salts thereof, may be lyophihzed to produce a iyophilszate which may be reconstituted with a pharmaceutically acceptable carrier, such as water, to regenerate a liposomal suspension

[00196 J Other pharmaceutical compositions may be prepared from the water-insoluble compounds disclosed herein, or salts thereof, such as aqueous base emulsions In such an instance, the composition will contain a sufficient amount of pharmaceutically acceptable emulsifying agent to emulsify the desired amount of the compound or salt thereof Particularly useful emulsifying agents include phosphatidyl cholines, and lecithin

[00197] In addition to the active compounds the pharmaceutical compositions may contain other additives, such as pH-adjusting additives In particular useful pH-adjustsng agents include acids such as hydrochloric acid bases or buffers, such as sodium lactate sodium acetate, sodium phosphate, sodium citrate, sodium borate or sodium gluconate Further the compositions may contain microbial preservatives Useful microbial preservatives include methylparaben, propylparaben and benzyl alcohol The microbial preservative is typically employed when the formulation is placed in a vial designed for multidose use Of course as indicated, the pharmaceutical compositions of the present invention may be lyophiiized using techniques well known tn the art

3. Subjects.

(00198] The present invention is primarily concerned with the treatment of human subjects but the invention may also be earned out on animat subjects particularly mammalian subjects such as truce rats, dogs, cats, livestock and horses for veterinary purposes, and for drug screening and drug development purposes

[00199] Subjects to be treated wtth active compounds or administered active compounds of the present invention are, in general, subjects in which an inflammatory cytokine such as tumor necrosis factor alpha (TNF-α} ss to be inhibited, and/or tn which a phosphodiesterase (PDE) such as phosphodiesterase Il ill, IV₁ and/or V is to be inhibited

|00200] Subjects m need of treatment wrth active agents as described herein include, but are not limited to subjects afflicted wrth invasive diseases infections, and inflammatory diseases or states such as septic shock, cachexia (or weight loss associated wsth chronic diseases such as Alzheimer's disease cancer, or AIDS), rheumatoid arthritis, inflammatory bowel disease (including but not limited to Crohn's disease and ulcerative colitis) multiple sclerosis cogestive or chronic heart failure psoπaεis, asthma, non tnsulsn-dependent diabetes mellitus, cerebral malaria, anemia associated with malaria stroke periodontitis AIDS and Alzheimer's disease Subjects afflicted with such diseases are administered the active compound of the present invention (including salts thereof), alone or in combination with other compounds used to treat the said disease, tn an amount effective to combat or treat the disease

[00201 J A particularly preferred category of diseases for treatment by the methods of the present invention are inflammatory diseases, or inflammations

[00202J While it is presently believed that the aforesaid diseases are treated by the inhibitory effect of the active compounds described herein on TNF-α production (and/or phosphodiesterase 4, kinases implicated in inflammation), applicants do not wish to be bound to any specific theory of the invention and it is intended that the treatment of particular diseases described herein by active compounds described herein be encompassed by the present invention without regard to the underlying physiological mechanism by which such treatment is accomplished

4. Dosage and routes of administration

[00203] As noted above the present invention provides pharmaceutical formulations comprising the active compounds (including the pharmaceutically acceptable salts thereof), m pharmaceutically acceptable carriers for oral rectal, topical, buccal, parenteral, intramuscular, intradermal, or intravenous inhalation and transdermal administration

{00204] The therapeutically effective dosage of any specific compound, the use of which is sn the scope of present invention will vary somewhat from compound to compound, and patient to patient, and will depend upon the condrtson of the patient and the route of delivery In general, a dosage from about 0 05 or 0 1 to about 20 50 or 100 mg/kg subject body weight may be utilized to carry out the present invention For example a dosage from about 0 1 rng_fkg to about 50 or 100 mg'kg may be employed for oral administration, or a dosage of about 0 05 mg/kg to 20 or 50 mg/kg, or more may be employed for intramuscular injection The duration of the treatment may be one or two dosages per day for a peπod of two to three weeks, or until the condition is controlled or treated in some embodiments Sower doses given less frequently can be used prophyiactically to prevent or reduce the incidence of recurrence of the condition being treated

[Θ02Θ5] The present invention is explained in greater detail in the following non-limsting Examples

EXAMPLE 1 4-(2-(THfluoromethyl)-1 H-benzo[d]imidazol-1 -yl)buty lboronic acid

[00206] A 20 mL scintillation viaf was charged with 2-(tπfluorornethyl)benzιmtdazo!e (50 mg, 0 27 mmol, 1 0 equiv) and 95% sodsum hydride (8 mg, 0 32 mmoi, 1 2 equiv} Anhydrous dϊmethylformamide was added, and the reaction mixture was stirred for 10 mm A 1 0 M solution of 4- bromobutylboronic acid {53 mg, 0 30 mmol, 1 1 equiv) in dimethyfformamtde was added The reaction was stirred at ambient temperature After 5 days the reaction mixture was filtered through ceiite and concentrated in vacuo The residue was purified by reverse-phase HPLC to afford 4-(2- {tπfluoromethyl)-1H-benzo[d]ιmidazoi-1-yl)butylboronιc acid (43 mg, 53%) ¹H NMR (300 MHz, CD₃CN) δ 7 93 (d, J = 8 O Hz, 1H), 7 77 {d, J = 8 0 Hz, 1H), 7 59 (t, J = 7 4 Hz, 1 H), 7 50 (m, 1H), 5 61 (s, 2H), 4 47 (t J = 7 7 Hz, 2H), 1 96 {pent, J = 7 8 Hz. 2H), 1 57 (pent, J = 7 8 Hz, 2H), 0 85 (t, J = 7 9 Hz 2H)

EXAMPLES 2-4 5-{2-|ThtazoI-4-yi}-1W-benzo[cflimidazol-1-yi)pentylboronic acid

[00207] Cesium carbonate (486 mg, 1 50 mmo), 3 0 equiv) was added to a solution of thiabendazole (100 mg, 0 50 mmol, 1 0 equiv) in anhydrous dimethylformamide After stirring for 10 mm, a 1 0 M solution of 5-bromopentylboronιc acid {145 mg, 0 75 mmol, 1 5 equiv) was added The reaction mixture was stirred at ambient temperature After 5 h, the reaction mixture was filtered Silica gel diol (1 1 g, 3 equiv) was added to the filtrate and shaken for 30 mm The silica get was washed with 30 mL of acetonitπle followed by 30 ml of 95 5 water-acetomπle with 25 mrnoi trtfluoroacetsc acid The aqueous wash was concentrated in vacuo, and the residue was purified by reverse-phase HPLC to afford 5-(2-(thιazol-4-y!)-1H-benzo[c/]ιmιdazoi-1-yl)pentylboronιc acid (110 mg, 70%)

(00208] A 1 dram vial was charged with thiabendazole (50 mg, 0 25 mmol, 1 0 equiv) and 95% sodium hydride (7 5 rng, 0 30 mmol, 1 2 equiv} Anhydrous dimethylformamide was added, and the reaction mixture was stirred for 10 mm A 1 0 M solution of 5-bromopentylboronιc acid (53 mg, 0 27 mmol, 1 1 equiv) in anhydrous dimethyiformamsde was added, and the reaction mixture was stirred at ambient temperature After 4 days the reactton mixture was filtered and concentrated in vacuo The residue was purified by reverse-phase HPLC to afford 5-(2-(thιazol~4~yl)-1H-benzo[d]fmιdazol-1- yl)pentylborontc ac!d (10 0 mg, 13%) ¹H NMR (300 MHz, CD₃CN) δ 9 39 (br s, 1 H), 8 73 (br s, 1H), 7 88 (m, 1H), 7 72 (m, 1 H), 7 46 (m, 2H), 4 72 <i, J = 7 δ Hz₁ 2H) 1 71 (m 2H), 1 21 (m, 2H), 0 43 (t, J = 6 9 Hz, 2H)

[00209] Thiabendazole f 10 g 49 75 mmol) was added to a suspension of cesium carbonate (48 5 g 149 mmol 3 0 equtv) in dsmethylformamide After stirring for 30 rmπ a solution of bromopentyiboronic acid { 15 g 77 mrnol ) was added The reaction mixture was stirred for 2 days, then D! water was added until precipitate formed product was filtered then washed with water and filtered agasn White solid was dπed via vacuum (15 g yield 96 %) 1 H NMR (300 MHz, dδ-DMSO) δ 9 39 (br s 1H) 8 73 (br s, 1H) 7 88 sm, 1H) 7 72(m 1H) 7 46 (m 2H), 4 72 (t J=7 δ Hz 2H) 1 71 (m 2H) 1 21(m, 2H) 0 43(t J-6 9 Hz 2H) Elemental analysis C 56 99% H, 5 91% N 13 33% EXAMPLE 5 5-(5,6-dimethyI-1W-benzo[d]imiclazoI-1-yl)pentytboronic acid

[0021 OJ A suspension of 5,6-dιmethyibenzιmιdazole (50 mg 0 34 mmol) and potassium carbonate (70 9 mg, 0 51 mmol) in DlViF (0 3 M) in a 40 mL scintillation vial was stirred for 30 mm A solution of 5-bromopentylboronιc acid, (1 M₁ O O 38 mmol) was added and stsrred at room temperature for 90 h The reaction was filtered through celrte and washed with DMF The filtrate was evaporated and the residue was purified by HPLC to give 5-(5,6-dιmethyl-1W~benzo[d]irnιdazol-1-yl}penty!boromc acid (12 4 mg, 14%) ¹H NMR (CD₃CN 300 MHz) 68 794 (s, 1H), 7 65 (s 1 H), 7 585(s, 1H), 4 333(t, 2 H, J = 7 4 Hz), 2 425 (s 3H) 2 398 (s, 3H), 1 444-1 269 (m, 4H), 0 66{t 2H J = 7 5 Hz)

EXAMPLE 6 5-(1W-imidazo[4,5-cJpyridin-1-yl}pentylboronic acid

N^'

^

OH

OH

[00211 ] A suspension of 5-azabenzιmιdazole (50 mg, 042 mmol) and potassium carbonate (87 01 mg, 0 63 mmol) in DMF (0 3 M) in a 40 m L scintillation vial was stirred for 30 mm A solution of 5- bromopentylboronic acid { 1 M, 0 0 38 mmol) was added and stirred at room temperature for 90 h The reaction was filtered through celite and washed with DMF The filtrate was evaporated and the residue was purified by HPLC to give 5-(1W-ιmιdazo[4 5-e]pyπdsn-1-yi)pentylboromc acid as a mixture of regιoιsomers(14 5 mg 15%) ^"H NMR (CD₃CN)S 9 25{s) 9 194 (s), 8 622(s 1 H) δ 549-8 487 (m 1H) 8 108 (d J = 6 Hz), 8 035(d J = 6 3 Hz) 4 553 (t, J = 7 4) 4 385(p, J = 7 1 Hz}, 1 963-1 871(m 2H) 1 456-1 267ιm 4H) 0 694-0 63Km 2H} EXAMPLE 7 5-(2-(4-Methoxypheny[)-1 H-benzo[d]imidazot-1 -yljpentylboronic acid

[00212] A 20 ml_ scintillation via! was charged with 2~(4-methoxyphenyl)-1H~benzo[c/Jϊmidazole (100 mg, 0.45 mmot, 1.0 eq), tetrabutylammonium iodide (16 mg, 0.04 mmol, 0.1 eq), and 95% sodium hydride (26 mg, 1.04 mmol, 2.3 eq). Tetrahydrofuran was added to the vial, and the reaction mixture was stirred until gas evolution was no longer evident. A 1.0 M solution 5-bromopentySboronic acid (96 mg, 0.49 rnmol, 1.5 eq) was added via syringe. The reaction mixture was stirred on a J-chem shaker at 180 rpm. After 48 h the reaction mixture was concentrated in vacuo. The residue was purified using an ISCO combiflash (12 g SiO₂, 30 mi/min, ethyl acetate to 9:1 ethyl acetate-methanoS). The appropriate fractions were concentrated in vacuo and the resulting oil was lyophilized from 3:1 acetonitrile-water to afford 5-{2-(4-methoxypheny!)-1H-benzo[d]imidazo!-1~yl)pentyJboronιc acid (53 mg, 35%) as a white powder: ¹H NMR (400 MHz, Cf₆-DMSO). δ 7.67 (m, 2H), 7.60 (m, 1 H), 7.36 (s, 2H), 7.22 (m, 1H), 7 1Q (m, 1 H), 7.10 (m, 2H). 4.22 (t, J = 7.3 Hz. 2H)₁ 3.82 (s, 3H), 1.64 (pent, J = 7.4 Hz, 2H), 1.22 (pent, J = 7 6 Hz, 2H), 1 09 (m. 2H), 0.46 (t, J = 7 6 Hz, 2H).

EXAMPLE 8 2-(3-F!uoro-4-methoxyphenyl)-1H-benzo[d]imidazole

(00213) Samples of 3-fiuoro-4-melhoxybenzaldehyde (771 mg, 5 mmol) and 1.2-phenylenediamine {541 mg, 5 mmol) were suspended rn nitrobenzene (2 mL) in a mscrowavable pressure tube (CEM) The mixture was subjected to microwave conditions (CEM Explorer, 200 ⁰C and a hold time of 10 mm). Upon cooling to room temperature, a large amount of a crystalline solid formed. The solid was filtered and tπturated with hexane (3 x 20 ml) and hexane/EtOAc 4:1 (3 x 20 mL) The product was isolated as a tan solid (856 mg 71%) ¹H NMR {400 MHz. CD₃CN) 6 7 84-7 89 (m. 2 H), 7.60 (bs. 2 H), 7 22-7,27 (m, 3 H), 3 96 (s, 3 H) EXAMPLE 9 2-|S-Bromopentyt)-4,4_I5,5-tetramethy⁽-1,3,2-dioxaboroIane

[00214] A solution of 5-bromopentylborontc aod (9 75 g, 50 rnmoi) and ptnacol (5 91 g 50 mmol) in acetonitπle (125 rnL) was stirred at room temperature for 16 hr The reaction mixiufe was concentrated under reduced pressure to give a dark gray residue Puπfication using an !sco purification system (silica column, eluted with hexane/EIOAc 4 1} gave the product as a clear liquid (8 1 g, 58%) Visualization of the product in TLC analysts was achieved using anisaldehyde or KMnO₄ staining followed by heating ¹H NMR (400 MHz, CD₃CN) 53 48 (t, J = 6 8 Hz, 2 H), 1 82- 1 86 (m 2 H), 1 40-1 42 (m, 4 H)₁ 1 23 (s, 12 H) 0 71-0 75 (m, 2 H)

EXAMPLE 10 a-tS-Fiuoro^-methoxyphenylJ-i-fS^ΛS.δ-tetramethyi-I.S^-dioxaborolan^_'ylJpentyO-IH- benzo[d]imidazole

[00215] A suspension of 2-(3-fluoro-4-methoxyphenyl)-1H^»benzo[d]ιmιdazole (300 mg, 1 24 mmol) 2~(5-bromopentyl)-4,4,5,5-tetramethyl-1 _!3 2-dioxaborolan (687 mg, 2 48 mmol) and cesium carbonate (808 mg, 2 48 mmol) sn DMF (2 5 mL) was stirred at room temperature for 22 hr The reaction mixture was diluted with EtOAc (25 mL) and H₂O (25 mL) The organic phase was extracted with aqueous LiCI (10 %, 25 mL) The organic phase was dried (Na₂SO₄) The solvent was removed to afford a brown residue Purification using an lsco purification system (silica column, eluted with hexane/EtOAc 4 1) gave the product as a clear liquid (8 1 g 58%) ¹H NMR (400 MHz₁ CD₃CN) δ 7 53-7 68 (m, 1 Hj 7 50-7 52 (m, 3 H) 7 24-7 30 (m, 3 H) 4 24-4 28 {m 2 H), 3 96 (m, 3 H), 1 71- 1 75 (m, 2 H) 1 10-1 30 (m 16 H) 0 58-062 fm 2 H} EXAMPLE 11 5-<2-(3-Fluoro-4-methoxyphenyl)-1H-benzo[dJimidazol-1-yt)pentyiboronic acid

[00216] Samples of 2-(3-fiuofo-4-methGxyphenyl)-1~(5-(4 4 5,5-tetramethyl-1 3 2-dtoxaborolan-2- yl)pentyl)-1H-benzo[d]irmdazofe (810 mg, 1 85 mmol) and diethanolamme (2 1 g 20 mmol) were combined in a rrucrowavable pressure tube (CEM) The mixture was subjected to microwave conditions (CEM Explorer, 60 "C and a hold time of 10 mm) LC-MS analysis showed some starting material Another portion of diethanolamme (2 1 g, 20 mmol) was added to the viscous mixture The mixture was again subjected to microwave conditions (60 ⁰C and a hold time of 10 mm) LC-MS analysis showed a trace o! the starting materia! remaining Thus, the reaction mixture was diluted with H_ΪO (50 mL) to form an emulsion Extraction was performed sequentially using hexane (50 mL), hexane/EtOAc 4 1 (3x50 mL} and ether (2x 50 mL) To the aqueous phase was added HCI (1M aqueous, 100 mL) followed by CH₂CI₂ (100 mL) The mixture was stirred at room temperature for 20 mm The pH of the aqueous phase was adjusted to 8 using solid K₂CO₃ The organic phase was separated The aqueous phase was extracted with CH₂CI₂ZEtOH 3 1 (3x100 mL) The organic phase was combined and dried (MgSO₄) The solvent was removed under reduced pressure to give an osly residue Acetonitnle/H₂O 1 1 (20 mL) was added to the residue After thorough mixing and solvent removal, an off-white solid was obtained Trituration with hexane/EtOAc 4 1 (3x50 mL) afforded the material slightly contaminated with 2-(3-fluoro-4-methoxyphenyl)-1-(5-(4 4,5 5-tetramethyl-1 3 2- dsoxaboroian-2-yl)pentyl)~1 H-benzo[d]ιmidazote The solid was then dissolved in acetone (5 mL) with heating After cooling, the addition of hexane (30 mL) induced the precipitation of a white solid (250 mg_τ 38%) ! H NMR400 MHz, CD₃CN) δ 7 67-7 69 (m, 1 H) 7 50-7 56 (m, 3 H) 7 23-7 33 (m, 3 H) 4 27 (t, J=S 0 Hz, 2 H) 3 97 (s, 3 H) 1 72-1 80 (m, 2 H)₁ 1 15-1 34 (m, 4 H), 0 60 (t, J=S 0 Hz 2 H)

EXAMPLE 12 ethyl 6-{2-(thiazo!-4-yl)-1H-benzo[d]imidazol-1-yl)hexanoate

[00217] Cesium carbonate { 2425 mg 7 5 mmol, 3 0 equiv) was added to a solution of thiabendazole (500 mg, 2 48 mmol, 1 0 eqsv) in anhydrous dimethylformamide After stirring for 30 mm, a solution of ethyl 5-bromohexanoate (1106 mg 4 98 mmol 2 eqiv) was added The reaction mixture was stirred for 3 hours Then water (8 1) was added and this was extracted with ethyi acetate The ethyl acetate solution was concentrated in vacuo and the residue was puπfted by silica gel column using ethyl acetate/ hexane as an elutmg solvent to afford ethyl 6-{2-(thiazoi-4-yi)-1H-benzo[d]smιdazot-1- yl)hexanoate

[00218] (650 mg 76 %) 1H NMR (300 MHz, d6~DMSO) δ 9 32 (d, J=1 76Hz, 1 H) 8 48(d J=1 76 Hz, 1H), 7 64(t,d J=7 03 Hz, 1 7Hz 2H) 7 25(m 2H) 4 72(t, J=7 3 Hz 2H), 3 99(q, J=7 03 Hz, 2H) 2 19(t, J=7 3 Hz 2H} 1 73(pent, J=7 3 Hz, 2H) 1 476 (pent, J=7 62 Hz, 2H), 1 23 (m. 2H) 1 106 (t, J=7 03 Hz, 3H)

EXAMPLE 13 N-hydroxy-6-{2-(thiazol-4-yi)-1H-benzo[d]imida2ol-1-yl)hexanamtde

[00219] To a neat ethyl 6-(2-(thιazol-4-yl)-1 H-benzoϊd]ιmιdazo!-1-yl)hexanoate {400 mg, 1 Iδrnmo!) N, O-Bιs(trιmethylsιlyl) hydroxylamsne (5,8 mmol, 1 G3g, 5 eq ) was added at room temperature After stirring for 30 mm a solution of 1N NaOH (2 ml) was added followed by the addition of methanol (-7 mi) Then reaction mixture was concentrated via rotovap and then purified on sihca gel column using methylene chloride/ methanol as an elutmg solvent {121 mg, 31 %) 1H NMR (300 MHz, dδ- DMSO) δ 10 27 (s, 1H), 9 32(d J=2 345 Hz, 1 H), 8 537 (s 1 H) 8 48 (d, J«1 759 Hz 1H) 7 637 (t, J=8 793 Hz, 2H) 7 25 (m 2H), 4 70 (t j=7 33 Hz, 2H) 1 862 (t J=7 33 Hz₁ 2H), 1 717 (t, J=7 33 Hz 2H), 1 452 (t J=7 33 Hz, 2H) 1 219(m 2H} EXAMPLE 14 ethyl 5-(2-{thiazol-4-yl)-1H-benzo[d]tmidazol-1-yi}pentanoate

[00220] 1 H NMR (300 MHz₁ d6-DMSO) δ 9 32 (d, J=1 759 Hz₁ 1 H), 8 489 (d, J=2 345 Hz, 1H), 7 643 (t J=6 741 Hz, 2H), 7 25 (m, 2H), 4 748 (t, J=7 034 Hz, 2H) 3 98 (q, J=7 6 Hz, 2H), 3 513 (t, j=6 448 Hz, 2H), 1 610 (pent, J=7 33 Hz, 2H)₁ 1 477 (pent, J=7 622 Hz. 2H)₁ 1 087(t, J=7 034 Hz, 3H)

EXAMPLE 15 N-hydroxy-5-(2-(thiazol-4-yl)-1H-benzo[d]imidazol-1-yl)pentanamide

|00221] 1H NMR (300 MHz, dδ-DMSO) δ 10 34 (broad, 1H), 9 438(s, 1H), 8 754 (S₁ 1 H)₁ 7 88 (d, J= 8 2 Hz, 1 H) 7 7δ(d, J=8 2 Hz₁ 1H) 7 47 (pent, J=5 5 Hz 2H) 4 8 (t, J=7 034 Hz 2H) 1 95(t J=7 3 Hz, 2H), 1 79 (pent, J=7 3 Hz, 2H)₁ 1 52 (pent, J=? 62 Hz, 2H) EXAMPLE 16 ethy! 5-(2-(4-methoxypheny 1J-1 H-benzo[d]imidazol-1 -y l}pentanoate

[00222] 1H NMR (300 MHz₁ c!6-DMSO} δ 7 68(d, J=8 79 Hz, 2H), 7 6(m 2H), 7 2{m, 2H), 7 1{d, J=8 79 Hz, 2H), 4 27{t J=7 3 Hz, 2H), 3 95{q, J=7 034 Hz, 2H), 3 83{s 3H), 2 178{S J=7 3 Hz, 2H), 1 67(m, 2H), 1 37{peπt, J=7 620 Hz, 2H) 1 096{f. J=7 034 Hz₁ 3H)

EXAMPLE 17 N-hydroxy-5-(2-(4-methoxyphenyl)-1 H-benzo[d]imidazol-1-yl)pentanamide

[00223] 1H NMR {300 MHz, d6-DMSO) δ 8 05(d, J=7 62 Hz 1 H), 7 8(d, J=8 79 Hz 4H)₁ 7 6(m, 2H) 7 25(d, J=S 79 Hz, 2H) 4 43(t, J=7 3 Hz, 2H), 3 88{s, 3H) 1 88(t, J=7 034 Hz, 2H), 1 74(m, 2H) 1 44{pent, J=7 62 Hz 2H)

EXAMPLE 18 ethyl 6-(2-{4-methoxypheπyl)-1H-benzo[dJϊmidazol-1-yi)hexanoate

J00224] 1 H NMR (300 MHz, dδ~DMSO) δ 7 68{d, J=8 79 Hz, 2H), 7 6{m 2H), 7 2(m, 2H), 7 1(Cl, J=8 79 Hz. 2H), 4 26{q, J=7 3 Hz₁ 2H), 3 98{m, 2H), 3 83(s, 3H), 2 137 (t, J=7 3 Hz, 2H), 1 67(m, 2H), 1 37{m, 2H)₁ 1 11(m, 5H)

EXAMPLE 19 N-hydroxy-6-(2-(4-methoxyphenyl)-1H-benzo[d]imidazol-1-yl)hexanamide

I00225] 1H NMR (300 MHz, dδ-DMSO) δ 10 311 (broad, 1 H), 7 856 (d, J=7 03 Hz, 1H), 7 76(m 3H), 7 433 (pent, J=5 8 Hz₁ 2H), 7 209 (d, J=8 79 Hz, 2H), 4 322(t, J=7 3 Hz, 2H), 3 865 (s, 3H), 1 842 (t. J=7 3 Hz, 2H), 1 717 (pent J=7 034 Hz, 2H) 1 385 (pent, J=7 3 Hz, 2H), 1 147 (m 2H)

EXAMPLE 20 5-{5-cyano-1W-indol-1-yf)pertty!boronic acid

HO-B

OH

[00226] A 1 dram vial was charged with 5-cyanoιndole (50 mg, 0 35 mmot, 1 O equiv) and 95% sodium hydride {10 6 mg, 0 42 mrnol, 1 2 equiv} Anhydrous dimethylformamide was added, and the reaction mixture was stirred for 10 mm A 1 0 M solution of 5-bromopentylboronic acsd (75 4 mg, 0 39, 1 1 eqiπv) in dimethylformamide was added, and the reaction mixture was stirred at ambient temperature After 4 days the reaction mixture was filtered and concentrated in vacuo The residue was purified by reverse-phase HPLC to afford δ-tδ-cyano-IW-indoi-i-yJJpentylboronic acid (46 5 mg, 52%) ¹ H NMR(SOO MHz, CD₃CN) δ 7 99 (s, 1 H)₁ 7 54 (d, J = 8 9 Hz, 1 H), 7 36-7 45 (m, 2H), 6 58 (d, J = 2 7 Hz, 1H), 4 18 (f, J - 7 16 Hz, 2H), 1 79 (pent J = 7 3 Hz 2H), 1 38 (m, 2H), 1 23 (m, 2H), 0 64 (t J = 7 6 Hz, 2H)

EXAMPLE 21 5-(4-Cyano-1H-mdol-1-yl}pentylboronic acid

[00227} A 1 dram vial was charged with 4-cyaπθindole (51 mg, 0 36 mmol 1 0 equrv) and 95% sodium hydride (20 9 mg 0 83 mmol. 2 3 equrv) Anhydrous dimethylformamide was added, and the reaction mixture was stirred for 10 mm A 1 0 M solution of 5-bromopentylboronιc acid (76 9 mg, Q 39 mmoi 1 1 equiv) in dimethylformamide was added and the reaction mixture was stirred at ambient temperature After 2 days the reaction mixture was filtered and concentrated m vacuo The residue was purified by reverse-phase HPLC to afford 5-(4-cyano-1j4-ιndol-1-yl)pentyfboronιc actd ¹H NMR (300 MHz CD₃CN) o 7 71 (m 1H) 7 45 (m 2H) 7 26 (t J = 8 0 Hz 1 H) 6 61 (d J = 2 1 Hz 1H) 4 19 (t, J = 7 15 Hz, 2H), 1 80 (pent, J = 7.3 Hz, 2H), 1 38 (m, 2H). 1 24 (m, 2H). 0 64 (t, J = 7 4 Hz, 2H)

EXAMPLE 22 5-{2-Methyl-1H-indoU1"yl)pentytboroπic acid

(00228] A 1 dram vsal was charged with 2-methySindole (50 mg, 0 38 mmol, 1 O equiv) and 95% sodium hydride (22 O mg O 87 mmol, 2 3 equpv) Anhydrous dimethylformamide was added, and the reaction mixture was stirred for 10 mm A 1 0 U solution of 5-bromopentyiboromc actd (81 1 mg, 0 42 mmoi, 1 1 equiv) in dimethylformamtde was added, and the reaction mixture was stirred at ambient temperature After 2 days the reaction mixture was filtered and concentrated tn vacuo The residue was purified by reverse-phase HPLC to afford 5-(2-Methyl-1 H-ιndol-1-yl)penfylboronιc acid ¹H NMR {300 MHz, CD₃CN) δ 7 43 (d, J = 7 7 Hz, 1H), 7 31 (d, J = 8 25 Hz, 1 H), 7 07 (m, 1 H), 6 97 (HD, 1 H), 6 18 (S₁ 1H) 4 07 (m, 2H), 2 40 (s, 3H), 1 69 (m, 2H), 1 35 (m, 4H) 066 (m, 2H)

EXAMPLE 23 5-{4-Fluoro-1H-indol-1-yt)pentylboronic acid

[00229] A 1 dram vial was charged with 4-fluoroirtdoie (50 mg, 0 37 mmol, 1 0 equiv) and 95% sodium hydπde (21 5 mg, 0 85 mmol, 2 3 equiv) Anhydrous dimethylformamide was added, and the reaction mixture was stirred for 10 min A 1 0 M solution of 5-bromopentylboronsc acid (79 3 mg, 041 mmol, 1 1 equiv) tn dimethylformamide was added, and the reaction mixture was stirred at ambient temperature After 2 days the reaction mixture was filtered and concentrated in vacuo The residue was purified by reverse-phase HPLC to afford 5-(2-Methyi-i H-ιndol-1-yl)pentylboronic acid ¹H NMR (300 MHz, CD₃CN) 5721 (m, 2H), 7 10 (m, 1H), 6 74 (m 1H), 649 (m, 1H)₁ 4 13 (m, 2H), 1 79 (m, 2H), 1 38 (m, 2H) 1 27 (m, 2H), 0 65 (m, 2H)

EXAMPLE 24 5-|4-Amino-1 H-indoM -yi)penty tboronic acid

[00230) A 1 dram vial was charged with 4-amιπoindole {51 rng, 0 39 mmol, 1 O equiv} and 95% sodium hydride (224 mg O 89 mmol, 2 3 equiv) Anhydrous dimethylformamide was added, and the reaction mixture was stirred for 10 mm A l O M solution of 5-bromopentyiboronιc acid (82 7 mg, 0 42 mmol, 1 1 equfv) in dimethylformamide was added, and the reaction mixture was stirred at ambient temperature After 2 days the reaction mixture was filtered and concentrated in vacuo The residue was purified by reverse-phase HPLC to afford 5-(2-Methyϊ-iH-indoi-1-yl)pentylborontc acid ¹H NMR (300 MHz, CDJCN) δ 7 19 (d, J = 3 3 Hz 1H) 7 08 (m, 2H), 6 63 (d, J = 7 2 Hz 1H) 6 48 (d, J = 2 8 Hz₁ 1H), 4 12 (t J = 7 2 Hz, 2H), 1 78 (pent, J = 7 3 Hz, 2H), 1 38 (m, 2H), 1 25 (m, 2H), 0 64 (t, J = 7 7 Hz, 2H)

EXAMPLE 25 5-{S-Fluoro-2-(3,4-dimethoxyphenyl)-1H-pndoI-1-yl)pentylboronrc acid

[00231] Sodfum hydride (60 wt% dispersion in mineral oil, 81 mg, 2 02, 1 1 equiv) was added to a solution of 5-fluoro-2-{3,4nJiimethoxyphenyl)-1H-indole (500 mg, 1 84 mmol, 1 0 equiv) in 8 2 mL of anhydrous dimethylformamtde The resulting yellow reaction mixture was stirred 10 mm at ambient temperature A solution of 2-(5-bromopentyi)-4,4,5,5-tetramethy!-1 ,3,2-dtoxaborolane (561 mg, 2 02 mmol, 1 1 equtv) in 1 0 mL of anhydrous dimethylformamide was added via syringe After 2 h the reaction mixture was partitioned with 200 mL of 1 1 water-ethyl acetate The layers were separated, and the aqueous layer was extracted with ethyl acetate (2 x 100 mL) The combined organic layers were washed with aqueous lithium chloride and bnne, dried over sodium sulfate, filtered, and concentrated in vacuo The residue was puπfted on an ISCO combiflash (40 g SiO₂, 40 mL/min, 4 1 hexanes-ethyl acetate) -to afford 5-f!uoro-2-(3 4-dιmethoxyphenyl)-1-(5-(4 4,5 δ-tetramethyl-1 ,3 2- dιoxaborolan~2-y!)pentyi)-1/-/-indoie as a clear oil

[§0232] A solution/suspension of 5-fluoro-2-(3,4-dιmethoxyphertyi)-1-(5~(4,4,5 5-tetra methyl -1 ,3,2 - dioxaborolan-2-yl)pentyl}-1 W-indole (114 5 mg. 0 245 mmol, 1 0 equiv) and diethanoiarmne (47 L 0490 mmol, 2 0 equiv) ιn 5 0 mL of diethyl ether was heated at 40 ^CC After 20 h the milky reaction mixture was cooled to ambient temperature, and the precipitate was collected by filtration The solids were washed with diethyl ether The pasty white solid was stirred for 20 mm m 10 mL of 1 1 dιcWoromethane-1 N aqueous hydrochloric acid The layers were separated, and the aqueous layer was extracted wttn dschtormethane (4 x 10 mU The combined organic layers *ere washed with saturated aqueous arnmomum chtonde dried over sodium sulfate filtered and concentrated sn vacuo The residue was taken up in 3 1 acetomtrile-water and lyophtøzed to afford 5-(5-flυoro~2-(3,4- dimethoxyphenyf)-1H-tndol-1-yl)pentyibororac acid as a white powder ¹H NMR (400 MHz, CD₃CN) δ 7.49 <m, 1 H), 7.27 (d, J = 9.8 Hz, 2H), 7 07 (br s, 3H), 6.98 (t, J = 9.6 Hz, 1 H), 4.19 {t. J = 6.1. 2H)₁ 3 89 (s, 3H), 3.87 (s, 3H), 1.65 (m, 2H)₁ 1.24 {m. 2H), 1 ,14 (m, 2H), 0.58 (m, 2H).

EXAMPLE 26 ethyl 6-{5-cyarto-1H-indol-1-yi)hexanoate

[00233] Cyanoiπdole ( 500 mg, 3.52 mmol ) was added to a suspension of sodium hydride (1.1 eq. 148 mg of 60% dispersion m mineral oii) in dimethylformamide and the reaction was stirred for 10 min. Then ethyl 6-bromohexanoate (1 5 eq, 1.18 g, 5.28 mmol ) was added dropwsse. The reaction was stirred at ambient temperature for 5 hours. Then water (8 :1) added and this was extracted with ethyl acetate. The ethyl acetate solution was concentrated in vacio and the residue was purified by silica gel column using ethyl acetate/ hexane as an eluting solvent to afford ethyl 6-{5-cyano~1 H-indol-1- yl)bexanoate (850 mg, 85% yield). 1H NMR {300 MHz, d6-DMSO): δ 8.057(d, J=1.172 Hz₁ 1H), 7.65(d. J=8.793 Hz. 1H), 7.6(d, J=2.93 Hz, 1H), 7.5(dd, J=1.759Hz, 8.79Hz, 1H), 6.6(d, J=3,5 Hz, 1H), 4.2(t, J=7.03 Hz, 2H), 3.98{q, J=7.034 Hz, 2H), 2.2(t, J=7.3, 2H)₁ 1.72(pent, J=7.62 Hz, 2H), 1.5(peπt, J=7.62 Hz, 2H), 1.2( m, 2H)₁ 1.1(1, J=7.034 Hz, 3H).

EXAMPLE 27 6-(5-cyano-1H-indol-1-yl)-N-hydroxyhexariamide

[00234] To a neat ethyl 6-{5-cyano-1H-indoH~yi)hexanoafe (850 mg , 2 99 mmol} N, O- Bss4rimethytsriyl) hydroxyiamfπe (14 95 mmol, 2 65g. 5 eq.) was added at room temperature After stirring for 30 mm solution of 1N NaOH (4 ml) was added followed by the addrtron of methanol Then the reaction mixture was concentrated via rotovap and then purified on silica gel column using methylene chlortde methanol as an eiuting solvent to afford 310 mg (38% yield) of 6-(5-cyaπo-1 H- ιndol-1-yl)~N-hydroxyhexanamιde 1H NMR (300 MHz, d6-DMSO) δ 10 298 (broad, 1 H), 8 656(broad, 1H), 8 067(d J=1 172 Hz 1 H), 7 675 (d, J=8 207 Hz, 1 H)₁ 7 581 (d J=3 517 Hz 2H) 7 465(d,d, J=8 79 Hz, 1 172 Hz, 2H) 6 578 (d, J=2 931 Hz, 1 H), 4 197 (t, J=7 034 Hz 2H) 1 88 (t J=7 3 Hz, 2H), 1 717 (pent, J=7 3 Hz, 2H), 1 476 (pent. J=7 620 Hz₁ 2H), 1 167(m 2H)

EXAMPLE 28 Synthesis of 2-substJtuted-2H-benzo[bf[1,4]thiazin~3(4H}-ones

[00235] The thioanshne (1 mmol) and α-brorno-α-sυbstituted acetic acid (0 9 mmol) ss combined in xylenes (5 0 mL) and heated to 100⁰C for six hours After cooling the solvent is removed under reduced pressure, and the target product is purified on an HPLC-MS apparatus (Agilent) by mass directed fractionation

EXAMPLE 29

Synthesis of 5-(#-R-2,3-dihydro-3-oxobenzo[b][1,4] (thia/oxe)zϊn-4-yl)pentylboronic acid

[00236] The parent πng (1 0 mmol), 5-bromo-1-ρentylboronιc acid (2 0 mmol), and cesium carbonate {2 5 mmoi) are combined in 2 0 mL of DMF and shaken at ambient temperature for 48 hours Aiternattveiy the 5~bromo-1-pentylboromc acid is added in 0 5 mmol aliquoϊs every 12 hours for 48 hours This increases both the conversion and yield The reaction mixture is then filtered to remove the cesium carbonate and the solvent is removed under reduced pressure The target product ss purified on an HPLC-MS apparatus (Agilent) by mass directed fractionation EXAMPLE 30 SKβ-fiϋoro-ZjS-dihydro-S-oxobenzofbJti^Joxazin^-yOpentylboronic acid

J00237] ¹H NMR (400 MHz, CD₃CN): 6 98 (1 H, dd), 6.92 (1 H. dd)_h 6.75 (1 H. dt), 4.55 (2H, s), 3. (2H, t), 1.62 (2H, m), 1.43 (2H₁ m), 1.34 (2H, m), OJO (2H, t).

EXAMPLE 31 5-(2,3-dihydro-3-oxobenzo[b][1,4]thiazin-4-y!)pentylboronic acid

OH

B OH

J00238] ¹H NMR (400 MHz, CD₃CN): 7.41 (1H₁ m). 7.28 (2H, m), 7.05 (1H, m), 4.71 (2H, s), 3.37 (2H, m), 1.54 (2H, m), 1 34 (2H, m), 0.91 (2H, m}, 0.70 (2H, t)

EXAMPLE 32 5-{7-ch!oro-2,3-dihydro-3-oxobenzo|;b][1,4]thiazin-4-yl)pentylboronic acid

[00239] ¹H NMR (400 MHz, CD₃CN): 7.44 (1H, d), 7.27 (1 H, d), 7.22 (1 H₁ s), 3.96 (2H, t), 3.39 (2H, s), 1.57 (2H, m), 1.38 (2H, m), 1.28 (2H. m), 0.67 (2H, !).

EXAMPLE 33 5-(2,3-dϊhydro-7-nitro-3-oxobenzo[b][1 ,4]oxazϊn-4~yf)pentylboronic acid

[00240] ¹H NMR (400 MHz. CD₃CN): 7.92 (2H, m), 7.12 (1H, d), 4.73 (2H, s), 3.99 (2H. t), 1.66 (2H₁ m), 1.40 {4H, m}, 0.71 (2H, t).

EXAMPLE 34 S-^S-dihydro-S-oxobenzolbJII ,4]oxazin-4-yi)penty lboronic acid

[00241] ¹H NMR (400 MHz, CD₃CN); 7.13 (1H, d), 7.06 (2H₁ m), 7.00 (1H, m), 4.56 (2H, s), 3.91 (2H, t), 1.62 (2H, t), 1.38 (4H, m), 0.70 (2H, t).

EXAMPLE 35 ethyl 2-(3_r4-dihydro-3-oxo-4-(5-peπty!boronic acid)-2H-benzo [b3[1,4]thiazin-2-yl)acetate

J00242] ¹H NMR (400 MHz, CD₃CN): 7.41 (1 H, dd), 7.31 (2H, m), 7.08 (1 H, dt), 4.13 (2H₁ q), 3.97 (1H, dd). 3.81 (2H, 7), 2.89 (1H₁ dd), 2.54 (1H, dd) 1.57 (2H, m), 1.34 (4H, m), 1.23 (3H, t), 0.67 (2H, t).

EXAMPLE 36 ethyl 6-(7-chIoro-2,3-dihydro-3-oxobeπzo|;b][1,4^'|thiazin-4-yl)hexanoate

[00243J Cesium carbonate ( 2443 mg, 7 5 mmo!, 3.0 eqυiv) was added to a solution of 7-chloro-2H- 1 ,4-benzothiazin-3(4H)-one (500 mg 2.5 mrnoi, 1.0 eqiv) in anhydrous dsmethyiformamkie. After stirring for 30 mm, a solution of ethyl 5-bromohexanoafe (1106 mg, 4 96 mmol 2 eqh/i was added. The reaction mixture was stirred for 3 hours. Then water (8:1) was added and this was extracted with ethyl acetate. The ethyl acetate solution was concentrated in vacuo and the residue was purified by silica gel column using ethyl acetate/hexane as an eiuting solvent to afford ethyl 6-(7-chloro»3~oxo- 2,3'dthydrobenzo[b3[1,4]thiazin-4-y!)hexanoate (545 mg, 64 % yield). 1H NMR (300 MHz. d6- DMSO): 5 7 511 (m, 1H), 7.3(m, 2H), 4.01{m, 2H), 3.91 (t, J=7.3 Hz₁ 2H), 3,49 (s, 2H). 2.2 (t, J=7 3, 2H), 1.48 (m, 4H), 1 ,2(m, 2H), 1 137 (t, J=O.134, 3H)

EXAMPLE 37 e^-chloro^jS-dihydro-S-oxobenzotblfli^lthiazin^-yli-N-hydroxyhexananiicle

[00244| To a neat ethyl 6~(7-ch!oro-3-oxo-2,3-dihydrobenzo[b][1,4Jthiazin-4-yl)hexaπoate (500 mg , 1.45mmol) N, O-Bts(trimethylsilyl) hydroxylamine (7.25 mmol, 1 3g, 5 eq.) was added at room temperature. After stirring for 30 min a solution of 1N NaOH (2 ml) was added followed by the addition of methanol (~7 mi). Then reaction mixture was concentrated via rotovap and then purified on silica gel coiumn using methylene chloride/ methanol as an eluting solvent (62 mg, 13 %) : 1H NMR (300 MHz₁ dδ-DMSO): δ 10.306 (s, 1H), 8.650(s, 1 H). 7.511(m, 1 H), 7 3(m, 2H), 3.91 (t, J=7.3 Hz₁ 2H), 3 49 (s, 2H), 1.883 (t, j=7 3, 2H), 1 45 (m, 4H), 1.2{m, 2H).

EXAMPLE 38:

In vitro receptor binding, enzyme and ADME-Tox assays of the compound of Example 20 (5-{5- cyano-1W-indol-1-yt)perUylborortic acid

[00245] This example shows the effects (5-(5-cyano-1 H-ιndol-1-yl)pentylborortfc acid in various in vitro receptor binding, enzyme and ADME-Tox assays In each experiment, the respective reference compound was tested concurrently with 5-(5-cyano-1W~rndol~1-yl)pentylboronic acid sn order to assess the assay suitability. Reference compound were tested at several concentrations (for IC_5e or EC₅₀ value determination), and the data were compared with historical values previously determined,

f00246] Bind assay. The binding of (S-fS-cyano-IH-indoM-yOpeπtyiboronic acid to the receptors was determined as described in Tabfes 1 and 2. The specific ϋgand binding to receptors is the difference between the total binding and the non-specific binding determined in the presence of an excess of unlabeled ligand. The results are expressed as the percent nhibition of control values percent in the presence of (5-(5-cyaπo-1 W-indoi-1-yl)peπtyfboroπic acid as shown in Table 3 The mean values from two experiments expressed as the percent of control binding was also determined (data not shown) The IG₅₀ values (concentration causing a half-maxsmai inhibition of control specific binding) and Hifl coefficients (n_H) were determined by non-linear regression analysis of the competition curves ussng Hitl equation curve fitting The inhibition constants (K,) were calculated from the Cheng Prusoff equation (K, = SC₅₀Z(I +(LZK₀)) where L = concentration of radioligand in the assay and K_D = affinity of the radioligand for the receptor) see Table 4

[00247]

Table 1

^'~Q44(nf human recombinant clozapine Van Toi et a! (1992;

[00248]

[00249]

[00250]

[00251] Enzyme assays The effect of (5-(5-cyano-1/-/-mdol~1-yl}pentyfboromc acid on the enzymes of Table 5 was determined with the using the experimental conditions described in Table 6

Table 6

[00252] Enzyme Results The mean values for the inhibitory effects of {5-{5-cyano-1H-ιndol-1- yl)pentylborontc acid on the assayed enzymes is summarized m Table 7 The ICso value for each reference compound ss indicated in Table 8 Each is within accepted limrts of the historic average ± 0 5 log units The mean values for the stimulatory effects of f 5-(5-cyano-1H-ιndol-1~y!)pentylboronιc acid summarized Table 9 The EC₆₀ value for each reference compound is indicated in Table 10 Each is within accepted limits of the frsstoπc average ± 0 5 tog units

[00253}

(00254]

_3_ziθdo_L;tyjos_iπe

ATPase(Ha⁺K^") 92E-07 ^"TJ ouabatp [00255]

[00256]

Table 10

EC₅₀

Assay/Reference Compound "H

Adenyiyl cycfase (basalj/forskolin 1.1 E-05 1.0

Guanylyl cyclase (basal)/ sodium nitroprusside 4 2E-06 0.6

[00257] ADME-Tox: In vstro Metabolism. The ADME-Toxiclolgy in vitro metabolism of (5-(5-cyano- 1H-indot-1-yl)pentylboronic acid was determined using the procedures cited in Table 11. The mean values from two experiments of the effects of 1.0E-OS(M) (5-{5-cyano-1H-indol-1-yl)peπty!boronic acid on receptors is summarized in Table 4

[00258]

(00259I

[00260] Abbreviations: BFC: 7-BeπzySoxy-4-(trifluoromethyl)-coumarin: from Discovery Labware, catalog number 451730

CEC: 3-Cyano-7-ethoxycoumarin, from Molecular Probes, catalog number C-684

CHC: 3-Cyano-7-hydroxycoumarin

CYP: Cytochrome P450

G6P: D~G!ucose-6-phosphate, from Sigma, catalog number G,-77J2

GβPDHase: GiucQse-6~phosphate dehydrogenase, from Sigma, catalog number G-4134

HFC: 7-Hydroxy-4-triflυoromethy[coumarin

MFC: 7-Methoxy-4-trifluoromethylcoumarin,fromSigma, catalog number T-3165

NADP: β-Nicotinamide adenine dinucleotide phosphate, from Sigma, catalog number N-0505

[00261] Results ADME-Tox: in Vitro Metabolism. The mean values for the effects of 5-(5-cyano-1 H- indoM-ytJpentyiboronic acid are summarized in Table 13. The data obtained with the reference compounds is shown Table 14

[0Θ262J

i Table 14

^', Assay IC₅₀ Reference Compound (M)

CYP1A2 inhibition (CEC substrate) 5 5E-06 furafylline

⁽ CΫP2C9 Inhibition (7-MFC substrate) 2 5E-07 1 0 I suifaphenazole

CYP2C 19 Inhibition (CEC substrate) 3 OE-06 tranylcypromine

CYP2D6 inhibition (7-MFC substrate) 4 8E-08 0 9 quinidine

CYP3A4 inhibition (BFC substrate) 73E¹OT I 1 4 ketoconazole

[00263] ADME-Tox For QT Prolongation the general procedure is shown in Table 15 and the experimental condition are shown in Table 16 In the event that a negative (< 5 % inhibition) compound was tested, the reference compound was perfused into the bath to ensure blockade of the HERG current, thereby eliminating false negative results For positive (active) compounds, controls with 10 nM E-403 1 were performed in separate cells (same clone) E-4031 from Wako, catalog number 052-06523 For patch-clamp, the incubation conditions were applied until steady-state was achieved

[00264]

Reference

Assay Celts Compound Bibliography

K⁺ channel HEK-293 cell line stably expressing E-4031 Zhou et al

(HERG) HERG (1998)

(patch-clamp)

[00265]

Table 16

Method of

Assay Incubation Conditions (mM) Detection

K^" channel 10-20 mm, 22- Pipette 130 KC1 , 10 NaCi 1 MgCI₂, 10 EGTA, 5 Whole-celi (HERG) 24 ⁰C MgATP, 10 HEPES (pH adjusted to 7 2 with 1 H patch-ciamp (patch- KOH) clamp)

BaIh 137 NaCI, 4 KCI . 1 8 CaCI₂, I MgCIj,

10 DM-GSucose, 10 HEPES

|00266] For HERG (paϊch-ciamp) studies cultured cells (1-3 days) were used for recordings. The ceiis were cultured in DMEM/F 12 + 10 % FBS For recording, cells were plated on collagen-coated coverslips at low density (about 10⁴ cells/mL).The cells were held at -80 mV and depolarized to +20 mV for two seconds, followed by a one second pulse to -40 mV to reveal the tail current. This paradigm was delivered once every eight seconds (0.125 Hz) to monitor the current amplitude. After the current amplitude stabilized, the test compound was delivered to the extracellular medium by bath perfusion. During superfusion, the cell was repetitively stimulated with the protocol described above, and the current amplitude was continuously monitored. Data were acquired and analyzed by using pClamp (Axon Instruments) and Excel (Microsoft), and are reported as mean and individual values. The degree of inhibition {%) was obtained by measuring the tail current amplitude before and after drug perfusion (the difference current was normalized to control and multiplied by 100 to obtain the percent of inhibition).

(00267] Results. ADME-Tox: QT Prolongation Tables 17 contains the mean experimental values for the test compound By adopting a general potency ranking system (Roche et al. ChemBioChem 2002, 3, 455-459} (Low, IC₅₀ > 10 μM; Moderate, 1 μM < IC₆₀ < 10 μM; and High, IC₅₀ < 1 μM), and based on the experimental findings, the test compound can be classified as a moderate-potency HERG- channel blocker,

Table 18

Test Inhibition of Tail

Concentration Current {%) Potency

(μM) MEAN Ranking

1 31.1 Moderate

EXAMPLE 39:

The effect of the compound of Example 7 5-(2-{44fiethoxypheny|}-iH-benzo[dJirnidazoM- yljpentylboronic acid in various in vitro phosphodiesterase and ADME-Tox assays.

[00268] The in vitro pharmacology of 5-(2-(4-Methoxyphenyi)-1H-benzo[d]imidazoi-1- yljpentylboronic acid was determined with several enzymes as described in Table 19 using the experimental conditions of Table 20

Table 20

Reference

Assay Origin Compound Bibliography

Phosphodiesterase 1 bovine brain 8-methDxy-iBfvlX Nichoison et at. Phosphodiesterase differentiated U-937 EHNA Torphy et al (1992)

2(h) cells

Phosphod sesterase human platelets milrinone Weishaar et al m (1986)

Phosphodiesterase U-937 cells rolipram Torphy et al (1992)

4{h)

Phosphodiesterase human platelets dipyridamole Weishaar et ai

5(h) (1986)

Phosphodiesterase 6 bovine retina zapπnast

Table 21

Reaction Method of

Assay Substrate/Stιmulus/T racer Incubation Product Detection

Phosphodiesterase [^JH]cAMP 30 ['H]S¹AMP Scintillation

1 +cAMP (1μM) mm /3O⁰C counting

Phosphodiesterase fHJcAMP 30 [³H]5'AMP Scintillation

2(h) +cAMP (1 μM) mm /30^cC counting

Phosphodiesterase [³H]CAMP 30 [³H]S¹AMP Scintillation

+CAMP (O 1 μM) mm /30⁰C counting

Phosphodiesterase [³HJcAMP 30 [³H]5'AMP Scintillation

A(h) +CAMP (1μM) msn /3O⁰C counting

Phosphodiesterase [³H]CGMP 30 [³H]S¹GMP Scintillation

5(h) +cGMP (1 μM) rmn /3O⁰C counting

Phosphodiesterase 6 [³H]CGMP 30 mm /30°C [³H]S¹GMP Scintillation

+CGMP (2μM) counting

|00269] Results The IC₅₀ values determined for 5-{2-(4-Methoxypheπyl)-1 H-benzo[d]rmιdazol-1~ yOpentyϊboronic acid are indicated in Table 22 The corresponding inhibition curves were also determined (data not shown) The ICso value for each reference compound is indicated in Table 23 Each is wsthin accepted limits of the historic average ± 0 5 log units Table 23 contasn the mean experimental values fo the QT prolongation study as perfored as desribed above By adopting a general potency ranking system {Roche et al ChemBtoChem 2002, 3_T 455-459} (Low, IC₅₀ ^>10 μM, Moderate, 1 μM < IC₅₀ < 10 μM, and High, IC₅₀ < 1 MM), and based on the experimental findings, the test compound can be classified as moderate/Ngh-potency HERG-channei blocker

Table 22 fCso Determination: Summary Results

Assay ICso (M) n_H Flags

Phosphodiesterase 1 N C

Phosphodiesterase 2 (h) 1 1E-07 08

Phosphodiesterase 3 (h) N C

Phosphodiesterase 4 (h) 4 2E-0? 1 0

Phosphodiesterase 5 (h) 9 6E-07 0 7

Phosphodiesterase 6 4 7E-06 09 N. C. Not calculable IC50 vaiue is not calcυiable because of less than 25% inhibition at the highest tested concentration.

Table 23

Test Concentration INHIBITION OF TAIL CURRENT (%) Potency (MM) MEAN Ranking

1 49 7 Moderate/High

EXAMPLE 40: Biological Example Inhibition of TNF-α Production By Peripheral Blood Monocyte Cells (PMSC)

PMBC in RPMl 1640 Cell Culture Medium (containing 1 % Penicillin and 1% Streptomycin) are aliquoteci into 96-well plates at 5 x 10^s cells/well and pre-incubated with test compounds for 30 minutes at 37 ⁰C, After incubation, 1 ug/mL LPS is added to each well to stimulate TNF-α production and the plate is incubated for 24 hours at 37⁰C After incubation, the supernatant is removed and the TNF-α secreted is quantified using EIA detection kits commercially available from R&D Systems (USA), The results from this assay are expressed as percent inhibition of control activity, with the control being stimulated wells with no test compound. Dexamethasone is used as a standard reference compound sn the assay and is tested with each experiment All test compounds are diluted from 10 mM stock solutions in 100% DMSO

nM nM nM

74 nM nM nM

EXAMPLE 41: Effects of several compounds in various in vitro cell biology assays.

[00270J The procedures used in the various assays are shown in Table 25 In each experiment the respective reference compound was tested concurrently with the test compounds in order to assess the assay suitability it was tested at several concentrations (for IC₅₀ value determination)

ASS3/ o- q - ke*e eiee Corf pcα c Refe^e^ce

SFN-ysecreison (h) PBMC dexamethasone Andre et a! (1996)

TNF-α secretion (h) PBMC dexamethasone Schindler et al (1990)

IL-1β secretion (h) PBMC cyclohexsmide Schindler et al (1990)

IL-2 secretion (h) PBMC dexamethasone Konno et al (1994)

IL-4 secretion (h) PBMC dexamethasone Eπdo et al (1993) lL-6 secretion (b) PBMC dexamethasone Schtndler et al (1990)

1L-10 secretion (h) PBMC dexamethasone Rigano et a! (1995)

⁽L-S secretion (h) PBMC dexamethasone Schindler et al (1990)

Ceii viability (rø P8MC erythromycin Mosmann (1983)

HHHBB

[00271] The experimental condition used in the assays are shown in Table 26 The test compounds were assayed at 3 x 10 ^e M

fissay SuDbirafe

IFN-γsecretton (h) PHA (2 μg/m!) 24 h/37°C IFN-/ EiA TNF-α secretionfft) LPS (1 μg/ml) 24 h/37°C TNF-α EIA IL-1β secretion (h) LPS {1 μg/ml} 24 h/37°C iL-lβ EIA IL-2 secretion (h) PHA (20 μg/ml) 48 h/37°C IL-2 EIA SL-4 secretion (h) ConA (20 μg/ml) 48 h /37°C IL-4 EIA I L-6 secretion (h) LPS (1 μg/ml) 24 h /37⁰C IL-6 EIA IL- 10 secretion (h) PHA (3 μg/ml) 48 h /37°C IL-10 EIA tL-8 secretion (Jh) LPS (1 μg/ml) 24 h /37X IL-8 EIA Cell viability (h) IVtTT (0 5 mg/mt) 24 h /37°C formazan Photometry

(00272J Results The mean values for the effects of the test compounds are summarized in tables 27 The results are expressed as a percent of control values and as a percent inhibition of control values obtained in the presence of the test compounds

|00273| The IC₅₀ values (concentration causrng a haif-maxnnai inhibition of control values) and Hill coefficients _\n_H) were determined by non-linear regression analysis of the inhibition curves using HHI equation curve fitting a summary of the data is shown in Table 28 The IC₅₃ value for each reference compound is indicated was determined \data not shown) and was wsthsn accepted limits of the historic average ± 0 5 log units

TABLE 28

Assay Comocuπd iW.,-3

Flags

CCi-7048 2.9E-06 .M

> Cone Above the highest test concentration. IC50 value is above the highest tested concentration. Dose response curve has an inhibitory shape wrth less than 50 % inhibition at the highest tested concentration

N.C. Not calculable. IC50 value is not calculable because of less than 25% inhibition at the highest tested concentration.

EXAMPLE 41 :

Effects of CCI-7155 and CCI-7156, and sulfasalazine, in a rat model of colitis provoked by challenge with tri nitrobenzene sutphonic acid (TNBS).

J00274] The model of colitis that is provoked by intracoionic instillation of trinitrobenzene sulphonic acid (TNBS) as described by Morris and associates and Boughton-Smith and colleagues, and is now widely used and well-characterised (Boughton-Smsth et al, 1988a, 1998b; Morris et al, 1989; Reuter et at, 1996; Kiss et at, 1997; Fries et al, 1998, Galvez et al, 2000; Baiϊinger et al, 2000; Whittle et al, 2003}. The inflammatory response provoked by TNBS ts considered to reproduce many of the macroscopic, histological, and immunological hallmarks of clinical colitis. Thus, open ulceration may be produced, with transmural inflammation and thickening of the bowel wail Histological features include distorted crypt architecture, crypt atrophy, granulomata, giant ceils, basal lymphoid aggregates and the presence of an inflammatory infiltrate (Morris et al, 1989; Yamada et al, 1992; Hoffmann et al, 1997; Torres et al, 1999; Neurath et al, 2000, Villegas et al, 2003). Thus, the model has been used and validated for studying colonic inflammation and therefore to address aspects of the pathogenesis of IBO, as is the industry standard for evaluating potential novel therapeutic agents for this utility (Whittle et al. 2003).

[0027SJ In the present example, the effects of the following compounds were were evaluated in the TNBS model. ™&

CCi-7155

OH

OH

CCI-7156

CH - B OH

[Θ0276J The effects of CCI-7155 and CCI-7156, were evaiuated in the TNBS model at one and two oral dose levels, administered twice a day by gavage, treatment commencing 1 day prior to challenge, A low intracolonic concentration of TNBS (10 mg) was used, known to produce reproducible yet not unduly severe mucosal injury in the colon, determined 3 days after instillation. Colonic macroscopic injury has been assessed, as has colonic weight as a reflection of colonic oedema and wet/dry weight to determine colonic water content, along with determination of myeloperoxidase (MPO) activity {Bradley et al, 1982) as an index of white cell infiltration for the evaluation of tissue injury. In addition to the test compounds [CCi-7155 and CCI-7156], the actions of sulfasalazine, a well-established and currently used treatment has also been evaluated in this model

METHODS AND PROTOCOL

[00277] TNBS Challenge - Maie Wtstar rats (230-28Og) were randomised into groups of 8-10 before commencement of the study. Food was withdrawn 18 h (overnight) before TNBS administration, but the rats were allowed free access to drinking water. On the morning of the day of challenge, Day 0, the rats were transiently anaesthetised with ether and TNBS (10 mg in 0,25 ml of 50% ethaπol) was instilled approximately 6-8 cm into the colon using a soft plastic catheter inserted in the rat rectum. The rats were allowed to recover with free access to food and drinking water. At the end of the experiment. 72 h after TNBS administration (i.e. on the mornsng of Day 3, between θ.00 and 11.00). the distal colon was dissected, and the distal 8 cm photographed and stored appropriately for subsequent analyses.

J00278] The following primary parameters were measured in the main study: (a) macroscopic scoring of distal 8 cm of colon; φ] myeloperoxidase levels in segments of distal 8 cm of colon in addrtion, the «etghi of the colonic segment was assessed as an indirect and nw-speciftc marker øf oedema, and this was supported by measurement of the wet/dry ratio as an index of water content The body weight of the animals was also determined and expressed as % change from the day of chatienge.

(00279] Treatments. Ail challenged groups were dosed orally twice daily from Day -1 The groups for study were^'

[00280J (a) Vehicle control 0 5% carboxy methyi cellulose (CMC) p.o . twice daily from Day -1

JΘ0281] (b) sulfasalazine 25 mg/kg_: p.o , twice daily from Day -1 (50 mg/kg/day total)

[00282] (C) CCΪ-7155 25 mg/kg_τ p o . twice dairy from Day -1 (50 mg/kg/day total)

[00283] (d) CCI-7155 50 mg/kg, p.o , twice daiiy from Day -1 (100 mg/kg/day total)

[00284] (e) CCI-7156 50 mg/kg, p.o., twice daily from Day -1(100 mg/kg/day total)

[00285] A further group of non-challenged and non-treated animate was used for baseline measurement of colonic MPO.

[00286] The compounds were thus administered orally, twice daily and given on Day -1 before TNBS administration, on Day 0 , the day of TNBS administration and on Day 1 and 2. Tissues were removed 72 h after TNBS administration (on Day3). Dosing was performed once in the morning (Between 9:00 and 11 :00) and once in the late afternoon (between 18^'OO and 20:00).

[00287] Preparation of Compound. CCI-7155 and CCi-7156 were suspended in 0.5% w/v carboxy methyl cellulose in sterile water, to produce a smooth suspension as instructed, and administered in a volume of 2 ml/kg (-0.5 ml per rat per dose),

[00288] Preparation of Sulfasalazine The doses of sulfasalazine used in the present study were derived from previously published work with this agent in the TNBS model (Boughton-Srnith et a!. 1988; Sykes et ai, 1999; Galvez et al, 2000, BobirvDubsgeon et al, 2001). Sulfasalazine was suspended in carboxy methyl cellulose (0.5% w/v in sterile water; Sigma Chemical Co, compound reference M-G262) and administered p.o. in a volume of 0,5 ml. In previous studies in these laboratories, this concentration of carboxy methyi ceiluiose (CMC) had no significant effect on the extent of colitis as determined by macroscopic injury and changes sn inflammatory markers following TNBS challenge.

[00289] Animal Husbandry: Male Wistar rats (270 + 30g body weight) were used throughout

f00290J Rats were maintained in air-conditioned with 20 arr changes per hour and constantly monitored environment with temperature 21 1.2⁰C The rooπrts were illuminated by fluorescent fight on a 12 hour light/dark cycle, fed pelleted rat No 1 maintenance diet RWl(E) and water ad libitum Rats were housed in groups of 3-5 tn polypropylene cages with ammai bedding of graded cellulose wood fibres

[00291} Macrocoptc Injury The distal 8 cm portion of the colon (measured from the rectum) was removed opened longitudinally and gently rinsed with ice-cold phosphate buffer (PBS, pH 7 4) blotted, weighed (Scaftec, Germany) and photographed (Samsung, Digimax 340, digital camera) The tissue was then cut into longitudinal strips, each strip being thus 8 cm long and included the whole of the zone of injury Each tissue was weighed and stored at -3O⁰C for the subsequent determination of myeloperoxidase activity, while a segment was also dried at 120⁰C for 24h for the determination of wet weight/dry weight ratio

|00292) The extent of macroscopscally apparent damage involving regions of haemorrhage necrosis was determined in a randomised manner from the colour images via computerised planimetry (Scion Image B4 02 version, Scion Corp ) Data on the macroscopic measurements are shown m the Appendix I Examples of the macroscopic appearance of the colon following challenge are shown in the Appendix Il The area of macroscopicaliy visible mucosal damage was calculated and expressed as the percentage of the total colonic segment area under study

[00293J Myeloperoxidase Activity The myeloperoxidase activity was determined using the method described by Bradley (Bradley et al, 1982} with msnor modifications The 8 cm longitudinal strips of the colon were weighed, homogenised (Ultra turrax, T25, 2 x 30 sec, 250 mg colon/ 1ml buffer) in ice- cold phosphate buffer (50 mM, pH 6 0), freeze thawed three times and centrifuged (15,000 x g 15 mm at 4 ⁰C) A 12μ! aliquot of the supernatant was mixed with 280μl phosphate buffer (50 mM, pH 6) containing 0 167 mg/ml of O-adenosine (.(hydrochloride and the reaction started with 10 μl 0 03 % hydrogen peroxide and assayed spectrophotometπcally (Benchmark Microplate reader, Bio-Rad Lab , λ = 490 nm) after 90 sec shaking The standards used for preparation of the standard curve were 0, 0 05, 0 1, 0 2, 0 3, 0 4 and 0 5 U peroxidase/ m! phosphate buffer Myeloperoxidase activity (MPO) was expressed as mU/mg protein or wet weight of tissue

[00294] Protein determination The method used is that described by Bradford (Bradford 1976) Thus 20 μl of the diluted samples (25 x or 50 x with distilled water) was msxed with 980 μl distilled water and 200 μ) Bradford reagent added to each sample After mixing and a 10 mm incubation the samples were assayed spectrophotomefπcally (Benchmark Micropiate reader Bio-Rad Lab λ = 595 nm) The standard curve was 0, 2 4, 6, 8 and 10 μg bovine serum aibumsn / mi dsstsiled water Protein was expressed as mg protein/ml

[00295] Reagents and Materials Tnnitrobenzene sulphomc actd was obtained from Fluka (Chemie AG. Buchs, Switzerland) The Bradford protein assay was from BlO-RAD All other assay reagents were from Sigma ChemicaS Company [00296] Statistical Evaluation Results shown in the figures are expressed as mean + S. E. M. from n rats per experimental group For statistical comparisons, the two-tailed Student's t-test and the analysis of variance with the Bonferoni test were used, where appropriate. P<0.05 was taken as significant.

RESULTS

[00297] In FIGS. 1-5 the labels have the following meanings;

TNBS= 2,4,6-Triratrobenzenesuifonic acid solution (1Gmg);

CMC = carbox ymethyi cellulose vehicle;

CMC group = TNBS + 0.5 % CMC (0.5 ml/rat p.o };

Sulfasalazine = TNBS + Sulfasalazine treated group (50 mg/kg/day p.o.)

CCi-7155 50= TNBS + CCI-7155 treated group (50 mg/kg/day p.o.)

CCI-7155 100 = TNBS + CCI-7155 treated group (100 mg/kg/day p.o )

CCI-7156 100 = TNBS + CCI-7156 treated group (100 mg/kg/day p o )

Control = non-treated, non-challenged absolute control.

I00298J Body Weight. FIGS. 1A-C show the effects of CCI-7155 (50 and 100 mg/kg/day p.o.), CCi- 7156 (100 mg/kg/day p,o.) and sulfasalazine (50 mg/kg/day p.o.) on body weight, expressed a % change in body weight at Day 0. Compounds were given in divided doses in a twice a day dosing schedule. Results are expressed as mean ± S.E.M ; n=9~10; significance is shown as aP<0.05 compared with 0.5% CMC group bP<0.05 compared with CCI-7155 50 mg group.

[00299} Following challenge with TNBS₁ there was a fal! in body weight observed in the vehicle- challenge control group over the 3 day period, with the fall in body weight reaching its peak after 2 days (FSGS. 1A-C). In contrast, there was no fall in body weight in the absolute control group that received no treatment nor was challenged with TNBS (data not shown). Treatment with CCI-7155 (50 and 100 mg/kg/day, administered orally in divided doses of 25 and 50 mg/kg b.i.d respectively) caused a dose-dependent attenuation of this fall in body weight, as shown in FIGS. 1A-C. The effects of the higher dose of CCI-7155 were significantly different from the TNBS control group at both Day 2 and Day 3 post-challenge (P<0 05). The effects of CCI-7156 (100 mg/kg/day administered orally in divided doses of 50 mg/kg b.i.d) reached marginal significance (P<Q 056) at Day 3 post-challenge (data not shown). Treatment with sulfasalazine (50 mg/kg/day administered orally in divided doses of 25mg/kg b.i.d} whjie appearing to attenuate the body weight loss (FIGS 1 A-B) did not reach statistical significance for this acton at any of the time points (data not shown)

[00300] Macroscopic Colonic Injury, in this study following intracolonic instillation of TNBS (10mg), the area of colonic injury, determined 72h after challenge in the control group of rats that had only received the 0.5% CMC vehicle p.o. involved 26 + 3% (n=9) of the total colonic area of the segment studied, determined by computerized planimetrtc measurement There was no ctetβctabte macroscopic injury in the colons from the non-chaϋenged group of rats (data not shown) The macroscopic appearance of the colonic mucosa following challenge and with the various treatments was assessed (data not shown) FIG 2 shows the effects of CCI-7155 ^50 and 100 mg/kg/day p o ) CCI-7156 (100 mg/kg/day p o ) and sulfasalazine ξ50 mg/kg/day p o ) on macroscopic injury in the colon Results are expressed as mean ± S E M , n=9-10 ^ap<0 05 compared with 0 5% CMC group ^BP<0 05 compared with CCi-7155 50 mg group ^cP<0 05 compared wsth Sulfasalazine 50 mg group

[00301 J Treatment with CCI-7155 (50 and 100 mg/kg/day administered orally in divided doses) caused a dose-dependent reduction in the area of colonic injury (FtG 2) This reduction in TNBS- induced colonic damage was statistically significant for both doses (P<0 001 and P<0 0001 respectively) as shown in FiG 2

[00302] Treatment with CCi-7156 (100 mg/kg/day administered orally in divided doses) caused a reduction tn the area of colonic injury (FIG 2) This reduction in TNBS-induced colonic damage was statistically significant (P<0 001) as shown in FIG 2

[00303] Treatment with sulfasalazine (50 mg/kg/day administered orally in divided doses) also significantly (P<0 001) reduced the extent of macroscopic injury as shown FIG 2

(00304] Colon Weight As an indirect index of inflammatory oedema in the colonic tissue, the weight of the colonic segments was determined at the end of the study FIG 3 shows the effects of CCi- 7155 (50 and 100 mg/kg/day p o }, CCI-7156 (100 mg/kg/day p o ) and sulfasalazine (50 mg/kg/day p o ) on colon weight Compounds were given in divided doses in a twice a day dosing schedule Results are expressed as mean ± S E IvI , n=9-10, ^aP<0 05 compared with 0 5% CIVIC group ^feP<0 05 compared wsth CCI-7155 50 mg group ^cP<0 05 compared with Sulfasalazine 50 mg group

[00305] As shown in FIG 3, the colonic weight in the groups challenged with TNBS was significantly higher than that of non-challenged colon (absolute control) for a comparabie tissue section Treatment with CCI-7155 caused a dose-dependent reduction m the colon weight (FIG 3) With the higher dose of CCi-7155, the reduction sn the colonic weight of the standard segment was statistically significant whereas that achieved by the lower dose was not (FfG 3) Treatment with CCI-7156 did not cause a significant reduction in the colon weight (FIG 3) A significant reduction in colon weight was also not observed in the sulfasalazine group (FIG 3), despite the reduction in damage seen in those tissues

[00306] Colon Wet/Dry Weight As an index of water content in the colonic tissue the wesght of the colonic segments was determined at the end of the study both wet and after oven-drying FIG 4 shows the effects of CCI-7155 (50 and 100 mg/kg/day p o ) CCI-7156 (100 mg/kg/day p o ) and sulfasalazine (50 mg/kg/day p o j on water content sn the colon Compounds were given in divided doses i" a twice a day dossng schedule Resurts are expressed as mean ± S E M rt=9-10 aP<0 05 compared with 0 5% CMC group bP<0 05 compared wrth CCI-7155 50 mg group cP<0 05 compared with Sulfasalazine 50 mg group

[00307] As shown in FIG 4 the colonic water content in the groups challenged wrth TNBS was significantly higher than that of non-challenged colon (absolute control) for a comparable tissue section As with the colon weight treatment wsth CCI-7155 caused a dose-dependent reduction in the colonic water content (FIG 4) With the higher dose of CCI-7155 the reduction in the colonic water content was significant whereas that achieved by the lower dose was not {FIG 3) Treatment with CCi-7156 did not cause a significant reduction in the colonic water content (FlG 4) Likewise a significant reduction in colon weight was also not observed in the sulfasalazine group (FtG 4} again despite the reduction in damage seen in those tissues

[00308] Colonic MPO Levels FlG 5 shows the effects of CO-7155 (50 and 100 mg/kg/day given p o in divided doses b i d ) CCl-7156 (100 mg/kg/day given p o in divsded doses b i d ) and sulfasalazine (50 mg/kg/day given p o m divided doses b i d) on MPO levels in the colon expressed as mU/mgprotetn Compounds were given in divided doses in a twsce a day dosing schedule Results are expressed as mean ± S E M n=9-10 aP<0 05 compared with 0 5% CMC group bP<0 05 compared with CCI-7155 50 mg cP<0 05 compared with Sulfasalazine 50 mg group

[00309] The level of MPO activity determined in the colonic tissue from rats in the unchallenged control group was significantly increased in the TNBS-challenged group (from 28+ 4 to 254 + 48 mU/mg protein P<0 001) as shown in FiG 5 Treatment with CCI-7155 caused a dose-dependent fall in the elevated MPO levels with a significant (P<0 01) reduction in colonic MPO levels at both doses as shown in FIG 5 Likewise treatment with CCI-7156 caused significant fail in the eievated MPO levels (FIG 5) Treatment with sulfasalazine significantly reduced the elevated colonic levels of MPO as can be seen in FIG 5 The extent of this reduction in MPO levels was in the same range as that brought about by the two experimental compounds (data not shown) The data for MPO has also been expressed as mU/g wet tissue (data not shown) and the relative changes between the groups were identical

[00310] As described above the sntra-cαlomc instillation of TN8S (10mg) caused a subchrortic colitis in the rat Thts macroscopic injury in the colon determined 72h after chaiienge consists of areas of haemorrhagic necrosss with evidence of tissue inflammation and hyperaemia In the present study the degree of macroscopic injury involved a mean of 25% of the measured colonic mucosa Such a moderate degree of injury is useful in the first stage analysis of novel therapeutic compounds for thts utility as it allows sensitive detection of any preventative activity on the lesion development

(00311] Oral administration of novel compound CCI-7155 caused a significant dose-dependent fall tn the extent of rracroscopically assessed THBS-sπduced colonic damage as dtd the single dose level of

CCi 7156 eva!ua!eα [00312] The macroscopic injury provoked by TNBS was accompanied by a substantial increase in the levels of MPO in the colon, which is an index of leukocyte infiltration into the inflamed tissue (Morris et ai, 1989: Reuter et at. 1996; Kiss et al, 1997). As with the macroscopic injury, both CCI- 7155 and CCI-7156 caused significant reduction in colonic MPO activity, and the relative activity of these compound on this parameter appeared comparable to that on macroscopic injury.

(00313] The elevated weight of the colonic segments following TNBS challenge, as an indirect index of oedema, was also dose-dependently reduced by the daily treatment with CCi-7155. a significant effect being observed at the higher dose. Likewise, the wet/dry ratio of the colon segment as an index of water content, was also significantly reduced by the higher dose of CCS-7155. This may reflect actions of this agent on the generalised inflammatory response in the tissue, with white-cell infiltration and oedema being attenuated. Despite the macroscopic injury being attenuated, none of the other treatment groups showed a reduction in these latter indirect parameters.

(00314] The faii in body weight that followed the challenge with TNBS was attenuated by CCI-7155 in a dose-dependent manner, with the higher dose of CC!-7155 preventing the fall in body weight at Day 2 and 3 post challenge. Although the effect of CCI-7156 on body weight change at Day 3 was near-significant, the data tn the other groups was suggestive of an effect at some time-points, but this dsd not reach statistical significance.

fO0315| Although sulfasalazine was introduced in clinical practice in the 1940's, the precise mechanism of its therapeutic action is a continuing discussion point but is, as yet, still unclear, it is known that the compound acts as a pro-drug, arriving essentially unchanged to the colon where it is cleaved by indigenous bacteria into its two constituent products, sulfapyridine and 5-arninosalicylic acid (5-ASA. mesaiamine) by action on its azo linkage. It is considered that 5-ASA is the active moiety, being released in high concentrations locally, and a number of delivery formulations of 5-ASA are in current cfinscal use (Schroeder. 2003). Despite this widespread clinical use, experimental studies with sulfasalazine have produced inconsistent findings of efficacy in a range of IBD models, including TNBS-induced colitis. Thus, this compound has been found to have variable effects on several of the indices of TNBS-colitis, depending on the dose and schedule utilized {Soughton-Smϊth et al, 1988a; Sykes et al, 1999; Galvez et ai, 2000; Bobin-Dubigeon et al, 2001), Studies on the putative active species, 5-ASA are even less clear as to the activity and reproducibility of effects in colitis models (Galvez et al, 2000; Tozaki et al, 2002).

[00316] Regarding the dose of sulfasalazine used m the current study, the clinical dose for the 500 mg tablets of the marketed form. Salazopynn™. is 2-4 tablets x 4 times a day for the treatment of active disease in IBD. Thus, this is a dose range of 4-8 g/day; based on an average body wesght of 75 kg. the lower dose is thus 53 mg/kg/day Indeed, the paed tatnc doses are given as 40-60 mg/kg/day for acute fiare-up. Although pharmacokinetic differences between rat and humans have to be taken into account, the dose !evef used tn the rats is thus dose that that used in therapeutics |00317] In the present work sulfasalztne at the dose of 50 mg/kg/day significantly reduced the degree of colonic injury and reduced the elevated MPO levels in the colonic tissue As can be seen from the data, the activity of CCI-7155 and CCl-7156 on both parameters was comparable to that of sulfasalazine

EXAMPLE 42;

The effects of the CCl-7308 or sulfasalazine in a rat model of colitis provoked by trinitrobenzene sυlphomc acid (TNBS)

[00318J In the present example the effects of the following compound CCl-7308 was evaluated in the TNBS mode!

[00319] In the study a low sntracolonsc concentration of TNBS (10 mg) was used, known to produce reproducible yet not unduly severe mucosal injury in the colon, determined 3 days after instigation In the study, colonic macroscopic injury has been assessed, as has colonic weight as a reflection of colonic oedema, along with determination of MPO activity (Bradley et al, 1982) as an index of white eel! infiltration for the evaluation of tissue injury

[00320] The pathogenesis of the inflammatory bowel diseases, including ulcerative colitis and Crohn's disease, is still not fufly understood but it is likely that pro-inflammatory cytokine release and derangement of the immune response play a role in the inflammatory processes (Kappeler & Muefler, 2000 Papadakis et al 2000) The colonic levels of the cytokine, tumour necrosis factor-α (TNF-α) have been shown to be increased in TNBS-induced colitis (Ameho et al, 1997 Ribbons et al, 1997, Sykes et al, 1999, Sun et al, 2001 , Ten Hove et al 2001, Villegas et ai, 2003) Pharmacological studies using a number of putative inhibitors of the synthesis of TNF-α have suggested efficacy in reducing damage in TNBS-tnduced colitis in the rat (Bobm-Dubigeon et al, 2001) Sn the current study, the colonic levels of TNF-α after TNBS challenge have therefore also been determined

METHODS AND PROTOCOL

(00321] The methods and protocol used were substantially similar to those tn Exampse 39 with some differences described beicw |00322] TNBS Challenge - Male Wistar rats (2?0-330g) were randomised into groups of 10-11 before commencement of the study

[00323] The following primary parameters were measured in the study

[00324] (a) macroscopic scoring of distal 8 cm of colon

[00325] (b) myeloperoxidase levels in segments of distal 8 cm of colon

[00326] (C) TNF-α levels in segments of distal 8 cm of colon.

(00327] in addition, the weight of the colonic segment was assessed as an indirect and non-specific marker of oedema. The body weight of the animals was also determined and expressed as % change from the day of challenge.

[00328] Treatments. Ail challenged groups were dosed orally twice daily from Day -1. The groups for study were:

[00329] (a) Vehicle control 0.5% carboxy methyl cellulose (CMC) p.o., twice daily from Day -1

[00330] (b) sulfasalazine 25 mg/kg, p.o., twice daily from Day -1 (50 mg/kg/day total)

[00331] (c) CCI-7308 2 mg/kg, p.o., twice daily from Day -1 (4 mg/kg/day total)

[00332J (d) CCI-7308 10 mg/kg, p.o., twice daily from Day -1 (20 mg/kg/day total)

[00333] (e) CCI-7308 50 mg/kg, p.o., twice daily from Day -1(100 mg/kg/day total).

[00334] Colon homogenates for cytokine measurements. The colonic tissue samples were thawed, weighed and homogenized (Ultra-turrax, T25. 2 x 30 sec on ice) in 4 volumes (250 mg colon/ml buffer) of a modified a Greenburg buffer (300 mmol/L NaCI, 15 mmol/L Tris, 2 mmoi/L MgCi, 2 mmol/L Triton X-1QQ, 20 ng/ml pepstatin A, 20 ng/m! leupeptin, 20 ng/mi aprotonine. pH: 7.4) Tissue homogenates were lysed for 30 mm. on ice, and then centrifuged (10 mm , 14, 000 x g) The aliquots of the supernatant were stored at -20⁰C until use (Ten Hove et a!., 2001).

[0033SJ Tumour Necrosis Factor-α Activity. The TNF-α levels were determined with quantitative TNF-α solid-phase Enzyme Linked Immunosorbent Assay (ELISA), which is based on the sandwich principle (HyCuIt biotechnology b. V.; Cat number: HK102). The TNF-α standards used were 0, 8.2_h 20.5, 51.2, 128. 320. 800 and 2000 pg/ml. At the end of the EUSA assay, the samples were measured speetrophofometricalty (Benchmark Microplate reader, Bio-Rad Lab; λ = 450 nm). The samples were diluted 2 or 4 times with the sample buffer included in the kit. The TNF-α values were expressed as pg/mg protein This commercsaiiy avertable krt (Hycult Biotechnology b v Utfβn. The Netherlands Catalogue number HKIOk.) used had a range of the standard curve of 0-2000 pg/mi with minimum detection ievei of 10 pg/ml of TNF-α

J00336] RESULTS, in FIGS 6-9 the labels have the following meanings

[0Θ337J TNBS = 2 4,6-Tπnitrobeπzenesulfontc acid solution (10mg)

[00338] CMC = carbox ymethyl cellulose vehicle

[00339] CMC = TNBS + 0 5 % CMC (0 5 ml/rat p o }

[00340] Sulfasalazine = TNBS + Sulfasalazine treated group (50 mg/kg/day p o )

J00341] CCI-7308 4 = TNBS + CCI-7308 treated group (4 0 mg/kg/day p o )

[00342] CCI-7308 20 = TNBS + CCi-7308 treated group (20 mg/kg/day p o )

[00343] CCi-7308 100 = TNBS + CCi-7308 treated group (100 mg/kg/day p o )

[00344] Body Weight Effects of CCI-7308 (4 20 and 100 mg/kg/day p o ) or sulfasalazine (50 mg/kg/day p o } on body weight expressed a % change in body weight at Day 0 Compounds were given m divided doses in a twice a day dosing schedule Results are expressed as mean ± S E M , n=9-11 , ^*P<0 05, ^**P<0 01 , ^**^*P<0 001 compared with CMC group

[00345] Following challenge with TNBS₁ there was a fall in body weight observed in the CMC vehicle-challenge control group over the 3 day period, With the fall in body weight reaching its peak after the first day post-challenge (FiG 6) Treatment with CCI-7308 (4, 20 and 100 mg/kg/day, administered orally in divided doses of 2, 10 and 50 mg/kg b i d respectively) caused a dose- dependent attenuation of this fall in body weight, as shown m FSG 6 The effect of the lower dose of CCI-7308 was significantly different from the challenged CMC control group on Day 1, while the intermediate dose was significantly different from the CMC group on Days 1 2 and 3 (P<0 001) as shown in FIG 6

[00346] The effects of CCI-7156 (100 mg/kg/day administered orally in divided doses of 50 mg/kg b s ύ) also were significant (P<0 001} on Days 1 2 and 3 post-chaiienge (data not shown) Treatment with sulfasalazine (50 mg/kg/day administered orally in divided doses of 25 mg/kg b i d}, attenuated the body weight loss following TNBS challenge (FiG 6), which reached statistical significance at Day 1 2 and 3 (data not shown)

|00347J Macroscopic Colonic Injury FIG 7 shows the effects of CCi-7038 (4 20 and 100 mg/kg/day p ø) or sϋffasaiazrπβ (50 mg> kg/day p o } on macroscopic injury m the colon Compounds were gwen tn divided doses in a twice a day dosing schedule Results are expressed as mean + S E M n=9-11 ***P<Q 001 compared with CMC group

[00348] In this study following intracoionic instillation of TNBS {10mg} the area of colonic injury determined 72h after challenge in the control group of rats that had only received the 0 5% CMC vehicle p o involved 27 + 3% (n=11) of the total colonic area of the segment studied determined by computerized plaπimetnc measurement There was no detectable macroscopic injury in the colons from a non-challenged group of rats (data not shown) The macroscopic appearance of the colonic mucosa following challenge and with the various treatments was determined

[00349] Treatment with CCl-7308 (4 20 and 100 mg/kg/day administered orally sn divided doses) caused a dose-dependent reduction in the area of colonic injury (FIG 7) This reduction in TNBS- induced colonic damage was statistically significant for both the 20 and 100 mg/kg/day doses (P<0 001 for both) whereas that for the lower dose did not reach significance as shown in FIG 7 The data suggests that the maximal effect on this parameter was achieved with the intermediate dose of 20 mg/kg/day (FIG 7) there being no significant effect between the actions of 20 and 100 mg/kg/day (FSG 7) Treatment with sulfasalazine (50 mg/kg/day administered orally in divided doses) also significantly (P<0 001) reduced the extent of macroscopic injury as shown in FlG 7 The degree of inhibition with sulfasalazine was comparable to that achieved with the intermediate dose of CCl- 7308 of 20 mg/kg/day (FIG 7)

[0Θ350J Colon Weight FIG 8 shows the effects of CCI-7038 (4 20 and 100 mg/kg/day p o) or sulfasalazine (50 mg/kg/day p o ) on colon weight Compounds were given in divided doses in a twice a day dosing schedule Results are expressed as mean ± S E M n=9-11

|00351] *P<0 05, ^**P<0 01 compared with CMC group

[00352J As an indirect index of inflammatory oedema in the colonic tissue the weight of the standard colonic segments was determined at the end of the study Treatment with CCl-7308 caused a dose- dependent reduction m the colon weight (FlG 8) With the intermediate and higher dose of CCi-7308 the reduction in the colonic weight of the standard segment was statistically significant whereas that achieved by the lower dose was not (FtG 8) A significant (P<0 05) reduction in colon weight was also observed in the sulfasalazine group (FfG 8)

[00353J Colonic MPO Levels FIG 8 shows the effects of CCI-7038 (A 20 and 100 mg/kg.day p o) or sulfasalazine (50 mg/kg/day p o ) on MPO activity in the colon expressed as mU/mgprotein Compounds were given in divided doses m a twice a day dossng schedule Results are expressed as mean ± S E M π=9-11 ^***p<0 001 compared with CMC group The level of MPO activity determined tn the coSomc tissue from the TN8S-chafienged group was 273 + 25 mU/mg protein as sho¥/n )fi FiG 8) In a separate eontro* study wth colonic t'ssue from rtoi-freated nαr-chaitenged rats the basal MPO activity was 44 + 12 mli/img protein (n=11), significantly lower than that determined following TNBS challenge

[00354] Treatment with CCI-7308 (4, 20 and 100 mg/kg/day) caused a dose-dependent fall in the elevated MPO activity with a Significant (P<0 001) reduction in colonic MPO levels at all three dose levels as shown in FIG 8 Treatment with sulfasalazine significantly reduced the elevated colonic levels of IvIPO as can be seen in FIG 8 The extent of this reduction in MPO levels by sulfasalazine was, however, significantly less than that brought about by the higher dose of CCI-7308 {data not shown)

|00355] The data for MPO has also been expressed as mU/g wet tissue (data not shown) the relative changes between the groups were identical

[00356] Colonic TNF-α Levels FIG 9 shows the effects of CCl-7038 (4, 20 and 100 mg/kg/day p o) or sulfasalazine (50 mg/kg/day p o ) on TNF-a ievels sn the colon, expressed as pg/mg protein Compounds were given in divided doses sn a twice a day dosing schedule Results are expressed as mean ± S E M , n~9-11 , ^*P<0 05, ^**P<0 01, compared with CMC group

[00357] The level of TNF-α in the colonic tissue from TNBS-challenged rats, determined after 3 days was 445 ± 49 pg/mg protein (FIG 9) In a separate control study with coionic tissue from non-treated, non-challenged rats, the basal TNF-α level was 16 + 4 pg /mg protein (n=11), substantially lower than that determined in colonic tissue following TNBS challenge

(00358] Treatment with CCf-7308 dose-dependently reduced the level of TNF-α in the coionic tissues, with the effects of the intermediate and higher dose of achieving significance (FiG 9), while those of the lower dose did not (data not shown)

[00359] A very similar pattern was observed when the data was expressed as TNF-α, pg/g wet tissue with the reduction in levels, with the low doses of not reaching significance while those with the intermediate and higher doses did (data not shown)

[00360] Treatment with sulfasalazine also significantly reduced the elevated colonic levels of TNF-α as can be seen in FIG 9 The extent of this reduction in TNF-α levels by sulfasalazine was not significantly different from that brought about by the intermediate or higher dose of CCI-7308 (data not shown}

[00361 ] As in prevsous studies ihe intra-colonsc instillation of TNBS {10mgl caused a subchronic colitis in the rat This macroscopic injury in the colon determined 72h after challenge, consists of areas of haemorrhagic necrosis with evidence of tissue inflammation and hyperaenrua In the present study the degree of macroscopic injury jnvoived a mean of 27% of the measured coionsc mucosa and a!fo#s sensitive detection of any preventative actwfy on the lesson development [00362] Oral administration of the compound CCl-7308 in a twice a day regimen commencing one day prsor to TNBS challenge, caused a significant dose-dependent fall in the extent of macroscopically assessed TNBS-induced colonic damage The findings suggest that the intermediate dose of CCl- 7308 of 20 mg/kg/day in divided doses is probably close to the maximal effect, with the higher dose of 100 mg/kg/day producing only a comparable degree of inhibition of lesion area There was no evidence of a bell-shaped dose response curve wrthsn the dose range studied with this compound Thus this agent may provide a broad therapeutic window for its effective dose-range

[00363] The macroscopic injury provoked by TNBS was accompanied by a substantial increase in the levels of MPO in the colon, which is an index of leukocyte infiltration into the inflamed tissue (Morns et at, 1989 Reuter et al, 1996, Kiss et al 1997) and reached levels comparable to those reported in the previous study for Nuada (Whittle and Varga, 2004) As with the macroscopic injury, CCl-7308 caused significant and dose-dependent reduction in colonic MPO activity Interestingly a significant reduction tn MPO was also observed with the lower dose of CCi-7308 that did not significantly reduce the macroscopic lesions This could reflect the differences of the statistical variances within the data for each of the parameters from these two groups Whether this finding could also indicate a primary action of CCI-7308 at these lower doses on acute neutrophils influx into the inflammatory site is unknown and would require further investigation

[00364] The elevated weight of the colonic segments following TNBS challenge as an indirect index of oedema was also dose-depeπdently reduced by the dasly treatment with CCi-7308, a significant effect besng observed at the intermediate and higher dose This may reflect actions of this agent at such doses on the generalised inflammatory response in the tissue wrth both white-cell infiltration and oedema being attenuated at these doses

[00365] The fail in body weight that followed the challenge with TNBS was attenuated by CCI-7308 in a dose-dependent manner, wtth the intermediate and higher doses preventing the fall in body weight on all 3 days post-challenge

[00366] In the present study sulfasalazine at the dose of 50 mg/kg/day significantly reduced the degree of colonic injury and reduced the elevated MPO levels &n the colonic tissue As can be seen from the data in general, the profile of activity of CCI-7038 on both parameters was similar to that of sulfasalazine, although lower doses of CCI-7308 were effective Preliminary indication of relative potency from a comparison of the respective molecular weights would suggest that CCI-7308 is some 2 5 times as active as sulfasalazine in reducing macroscopic injury

[00367J The clirsica! dose for the 500 mg tablets of the marketed form Saiazopyπn^ is 2-4 tablets x 4 times a day for the treatment of active disease in IBD Based on an average body weight of 75 kg and the dose range of 4-8 g<day the lower dose ts thus 53 mg/kg,day while the paediairsc doses are given as 40-60 mg kg/day fo^r acute ^flare-up A'tnough pharmacokinetic cWereiees oetween rat aid humans would have to take into account the effective dose level of sulfasalazine used in the rat in the current study of 50 mg/kg/day, is thus within the range used in the therapeutic control of IBD This suggests that this model can be predictive of the therapeutic effect of novel agents in colitis

[00368] The elevated colonic leveis of THF-α following challenge with TNBS, a known endogenous mediator of colrtts and a good biomarker of disease activity was significantly reduced by sulfasalazine, as reported previously by others (Ameho et al, 1997 Ribbons et al 1997 Sykes et al 1999 Sun et al, 2001 Ten Hove et al 2001 , VHiegas et al, 2003) Moreover in the present work CCI-7308 significantly reduced the TNF-α levels in a dose-dependent manner with the intermediate and higher doses reaching significance The degree of inhibition of the TNF-α levels by CCI-7308 was comparable to that produced by sulfasalazine

[00369] The pre-treatment of rats with a single intravenous dose of infliximab (Remicade™), a therapeutic protein targeting TNF-α, was found to reduce the macroscopic injury, colonic MPO and TNF-α levels observed 8 days after TNBS challenge (Woodruff et at, 2003) The degree of inhibition with infliximab may be comparable to the range to that seen with CCI-7308 at the intermediate and higher doses in the current work

[00370] This study indicates that CCI-7038, given by oral gavage twice daily commencing 1 day prior to challenge dose-dependently reduces the degree of tissue injury therapeutic activity in this 3-day rat model of colitis, reducing macroscopic colonic injury at both the intermediate and higher doses employed (20 and 100 rng/kg/day) The biomarkers of colonic inflammation, MPO and also TNF-α, the latter being a known inflammatory mediator involved in colitis, were also dose-dependently reduced in the inflamed tissue by these doses of CCi-7308 Overall, the findings suggest that a dose of CCi-7308 of 20 mg/kg/day in divided doses is close to the maximal effective dose in this model This profile of actions of CCI-7038 were comparable to those seen with sulfasalazine an agent used widely in the clinsc in the therapy of IBD and estimates of relative potency suggest a 2 5-fold greater activity with CCI-7308 on macroscopic injury and probably the other biomarkers

EXAMPLE 43

Comparison of the effects of CCI-7506, CCI-7507, sulfasalazine or infliximab in a rat model of chronic coϊitis provoked by trirtϊtrobenzene sulphonic acid (TNBS) over 14 days

[00371 J In the present example, the effects of the following compounds were were evaluated in the TNBS model CCl-7506

OH

B^'

OH

[00372] In the chronic model of colitis, assessment of the colonic inflammation is made 14 clays or longer, after the intracolonic challenge with TNBS {Boughton-Smith et al, 1988a, 1988b; Wallace et a!, 1989, Rachmilewitz et al, 1989; Wallace and Keenan, 1990; Ameho ef al, 1987; Sans et al, 1999, Sun et a!, 2001; Marie et al, 2003; Moreels et al, 2004; Gonzalez et al, 2004). This model ailows treatment with experimental agents to commence following the estabiishment of the colonic injury, typically 24 hours after the TNBS challenge (Galvez et al, 2000, Villegas, 2003, Gonzalez et al, 2004). The model should therefore identify the ability of the experimental compounds to accelerate the diminution of the inflammatory response and to promote healing of the colonic lesions. This model thus has relevance additional to the acute model, as the clinical correlate is the therapeutic intervention in patients with existing IBD not m remission or with flare-up, to reduce the crisis. This contrasts with the acute TNBS model where the compounds are administered one or two days prior to challenge, the clinical correlate being the use of prophylactic therapy to prevent flare-up and maintain remission in IBD patients.

J00373) In this current study, the low intracofonic challenge concentration of TNBS used in the acute studies was also used for the chronic study over a 14 day period. This concentration and timing was based on the findings from pilot studies where a range of concentrations of TNBS and treatment conditions were evaluated over a 14 day period. The dose of TNBS {10 rng) in rats starved for 12 hours, proved to yield significant colonic injury after 14 days, not dissimilar from that with the high dose of 30 mg, yet substantially reduced the high incidence of mortality and diarrhoea observed with the higher dose in the model and as reported by others with this higher dose over sub-chronic periods (Woodruff et al, 2003)

|00374] The methods and protocol used were substantially similar to those in Example 39-40. with some differences described below. [00375] TN8S Challenge Male Wistar rats {average body weight 21Og) were randomised into groups before commencement of the study in alt groups includtng the non-challenged and non- treated absolute control group food was withdrawn for 12 h before TNBS administration (ι e overnight on Day ~1), but the rats were allowed free access to drinking water

(00376) On the morning of the day of challenge (Day Q, between 9 00 and 11 GO a m ) the rats were transiently anaesthetised with ether and the TNBS solution (10 mg in 0 25 ml of 50% ethanol) was instilled approximately 6-8 cm into the colon using a soft plastic catheter inserted in the rat rectum The rats were allowed to recover with free access to food and drinking water At the end of the experiment, on the morning of Day 14, between 9 00 and 11 00), the colon was dissected, and the distal 8 cm photographed and immediately processed or stored appropriately for subsequent analyses

|00377] The following primary parameters were measured sn the study macroscopic scoring of dtstai 8 cm of colon, myeloperoxidase levels in segments of distal 8 cm of colon TNF-α levels in segments of distal 8 cm of colon

f00378| in addition, the weight of the standard colonic segment was assessed as an indirect and non-specific marker of oedema The body weight of the animals was also determined each evening of the study, starting on Day-1 , and also on the morning of Day 14 The data is shown graphically as the % change from the weight on Day~-1, prior to challenge

[00379J Treatments The TNBS challenged groups for study were {a) Vehicle control 0 5% carboxy methyl cellulose (CMC) p o , twice daily from Day (b) CCI-7506 25 mg/kg, p o , twice daily from Day 1 {50 mg/kg/day total), (c) CCI-7506 50 mg/kg, p o , twice daily from Day 1 (100 mg/kg/day total), (d) CCi-7507 12 5 mg/kg, p o twice dasly from Day 1 (25 mg/kg/day total), (e) CCi- 7507 25 mg/kg, p o , twice daily from Day 1(50 mg/kg/day total), (f) Sulfasalazine 25 mg/kg, p o twice daily from Day 1 (50 mg/kg/day total) (g) Infliximab 3 mg/kg, single slow i v injection, on

Day 1 and Day 7

[00380J The experimental compounds that were administered orally were gsven twice a day from Day 1 , i e 24 h following TNBS administration for the remainder of the 14 day experimental period infliximab was administered by slow i v injection of Day 1 and on Day 7 This fatter group of rats were also administered 0 5% w/v CMC (0 5 ml p o ) twice a day from Day 1 Dosing was performed once m the morning (between 9 00 and 11 00) and once in the late afternoon (between 18 00 and 21 00) In addition a group of rats that were non-treated and non-challenged were also evaluated for base-line measurements

[00381} Preparation and Dose of Infliximab The dose of infliximab (Remicade Centecor-Schβπng

Plough) of 3 mg/kg as a slow intravenous ϊrsjection used m this protocol, iS comparable Io that used m the clinical studies on IBD This does has also been used in the experimental setting in vivo to attenuate the response to TNF-α in acute or chronic inflammatory conditions in the rat (Kulmatycki et al 2001 Woodruff et al 2003) and in our own in-house studies in the acute TNBS model Infliximab was dissolved in the supplied diluent sterile saline for injection immediately prior to use as indicated in the technical documents supplied with the material

[00382] Macrocopsc Injury Macrosopic injury was performnewd as described above In addition to the quantitative measurement of area of damage the degree of colonic damage was aiso assessed in a randomised blinded fashion using a Damage Score, utilizing a 1-5 scale than has been adapted from that used previously (Boughton-Smith et al, 1988a) 0 = No Damage 1 = One region of localized inflammation or thickening (No ulcers), 2 = Linear ulceration, but no significant inflammation 3 = Linear ulceration with inflammation at one site, 4= Two or more sites of ulceration and/or inflammation (Ulcers present in at least one site), 5 = Two or more sites of uiceration and inflammation or one major site of ulceration and inflammation extending > 1 cm aiong the length of the colon

(00383] Results In the following figures the labels have the following menaing TNBS= 2 4,6- Tπnitrobenzenesulfonic acid solution (10mg) CIvIC = carboxy methylceffulose Abs control = non-challenged and non-treated

[00384] CMC = TNBS + 0 5 % CMC (b i d 0 5 ml/rat p o )

J00385] CCi-7506-50 mg = TNBS + CCi-7506 treated group (50 mg/kg/day p o total dose}

[00386] CCI-7506-100 mg = TNBS + CCl-7506 treated group (100 mg/kg/day p o total dose)

[00387] CCI'7507-25 mg= TNBS + CCI-7507 treated group (25 mg/kg/day p o total dose}

100388] CCi-7507-50 mg= TNBS + CCI-7507 treated group (50 mg/kg/day p o total dose)

[00389] SASP= TNBS + Sulfasalazine treated group (50 mg/kg/day p o total dose)

(00390] Infliximab^ T N BS+ Infliximab (3 mg/kg s v on Day 1 and Day 7 } + 0 5 % CMC (b i d 0 5 ml/rat p o )

[00391 J Body Weight FIGS 1GA-1GC show the effects of CCi-7506 (50 and 100 mg/kg/day p o > CCI-7507 (25 and 50 mg/kg/day p o ), sulfasalazine fSO mg/kg/day p o } or infliximab (3 mg'kg s v on Day 1 and 7) on body weight over 14 days expressed a % change of the body weight at Day -1 prior to TNBS challenge on Day 0 The oraily administered compounds were given in divided doses in a twice a day dosing schedule, commencing in the morning of Day 1 i e 24 h after TNBS challenge ASl groups including the non-chailenged non-treated absolute coπtol group were starved for 12h overnight on Day -1 Results are expressed as meap + S E M n=11-15 for the test groups and r~δ for the absolute control group, statistical significance is shown as *P<0 05 ^**P<0 01 ^***P<0 001 compared with the chaitenged control CMC group

[00392] In the non-challenged and non-treated absolute control group there was an apparent small fall in body weight of the rats on Day 0 from sis value on Day-1 presumably as a consequence of the 12h period of food removal overnight From Day 1 onwards the body weight in this group progressively increased and was then significantly different for the CMC challenged group at ail time points up to 14 days {FIGS 1A-1C) As in the absolute control group, there was an apparent snitiaS small transient fall in body weight on Day 0 compared with Day-1 in all of the TNBS-challenged groups (FIGS 1A-1C) There was no statistically significant difference between any of the groups on Day 0 (data not shown)

|00393] Following challenge with TNBS there was a further fail in body weight observed in the CMC- chaiienged control group on Day 1 (P<0 001), reaching its peak on Day 2 post-challenge The body weight then recovered progressively during the 14 day period, and by Day 4 was no longer significantly different from the value at Day-1 There was no significant difference in the % change in body weights between any of the challenged groups on Day 1 (data not shown)

[00394] Treatment with CCI-7506 (50 and 100 mg/kg/day administered orally m divided doses of 25 and 50 mg/kg b i d respectively) commencing 24h after TNBS challenge on Day 1 caused an attenuation of thts fall ;n body weight, as shown in FIG 10A The change in body weights of the lower dose of CCi-7506 was significantly different from those m the challenged CMC control group on Days 2 to 10 and also on Days 13 and 14 With the higher dose the changes in the fall in body weight were likewise attenuated, and were significantly different from the CMC group on Days 2, 3 and 4 (P<0 05) as shown in FIG 1C

[003951 CCI-7507 (25 and 50 mg/kg/day, administered orally in divided doses of 12 5 and 25 mg/kg b I d) also attenuated the TNBS-iπduced fail in body weight (FIG 1 B) With the lower dose, the change in body weight was significantly different from that in the CMC challenged group (P<0 05) on Days 2, 3 4, 8, 9 and 10 post-chailenge With the higher dose of CCi-7507 (50 mg/kg/day), the change in body weight was significantly different form the CMC group on all days from Day 2 to 10

[00396] Treatment with sulfasalazine (50 mg/kg/day administered orally in divided doses of 25 mg/kg b i d), also attenuated the body weight loss following TNBS challenge (FiG 10C) which reached statistical significance compared wrth the CMC challenged group on at Day 2 3 6 and 7

[00397J Intravenous injection of infliximab (3 mg/kg on Day 1 and on Day 7; attenuated the fall in body weight following TNBS wrth the change compared to the CMC challenged group being significant on Day 2 and 3 (FlG 10C) [00398] Macroscopic Colonic Injury Area of Damage FIG 11 shows the effects of CCI-7506 (50 and 100 mg/kg/day p o ), CCi-750? {25 and 50 mg/kg/day p o ) sulfasalazine (50 rπg/kg/day p o } or infliximab (3 mg/kg i v on Day 1 and 7) on macroscopic injury in the coton determined 14 days after TNBS challenge as assessed as the colonic lesson area % of the total area measured The orafϊy administered compounds were given in divided doses in a twice a day dossng schedule commenctng on Day 1 i e 24 h after TNBS challenge Results are expressed as mean ± S E M n=11-15 for the test groups and n=6 for the absolute control group, statistical significance is shown as ^*P<0 05 ^**P<0 01 , ^***p<0 001 compared with the challenged control CMC group

[00399] In the group challenged with TNSS (10 mg) on Day 0 followed by the vehicle 0 5% CMC vehicle p o twice a day, commencing from Day 1 i e 24h after challenge the area of injury determined after 14 days involved 44 + 4% (n=i4) and of the total colonic area of the segment studied, determined by computerized pianimetπc measurement (FIG 11) In a pilot study, administration of the vehicle 0 5% CMC vehicle p o , twice a day for Day 1 , had no significant effect on the area of colonic injury induced by TMBS (10 mg) after 14 days There was no detectable macroscopic injury in the colons in the non-challenged, non-treated absolute control group of rats (FIG 11)

[00400] Treatment with CCI-7506 (50 and 100 mg/kg/day administered orally in divided doses), commencing 24h after the TNBS challenge caused a dose-dependent reduction in the area of colonic injury observed on Day 14 (FiG 11) This reduction m TNBS-induced colonic damage was statistically significant for both doses (P<0 01 see Appendix I Table 10 for full tabular data) Treatment with CCI-7507 (25 and 50 mg/kg/day administered orally in divided doses) commencing 24h after challenge likewise caused a dose-dependent reduction in the area of colonic injury observed at Day 14 (FIG 11) This reduction in TNBS-induced colonic damage was statistically significant for both doses (P<0 001 , data not shown) The effect of CCi-7507 at the dose of 50 mg/kg/day was significantly (P<0 05) greater than that observed with CCI-7506 at that same dose (data not shown) Treatment with sulfasalazine (50 mg/kg/day administered orally in divided doses) commenctng 24h after challenge, significantly (P<0 001) reduced the extent of macroscopic injury seen at Day 14 after challenge as shown in FIG 11 This effect was not significantly different from that observed with any of the other active treatment groups Intravenous injection of infliximab (3 mg/kg on Day 1 and on Day 7 after challenge) significantly attenuated the area of injury following TNBS observed on Day 14 (FIG 11) This effect was not significantly greater than that observed with CCI-7506 CCI-7507 or sulfasalazine (FIG 12)

[0040Ϊ] Macroscopic Damage Score In addition to the area of visible injury the degree of macroscopic colonic injury was assessed using as Damage Score (_.scale 1-5) as shown in FIG 12 Results are expressed as mean ± S E M , n=11-15 for the test groups and n=δ for the absolute control group statistical significance is shown as ^**P<0 01 ^***p<0 001 compared wsth the challenged control CMC g^oup [00402] As can be seen, the scores in the treatment groups closely followed the profile of that determined by the quantitative measurement of area of damage. Thus, CCI-7506 (50 and 100 mg/kg/day) and CCI-7507( 25 and 50 mg/kg/day) at both doses reduced the damage score observed in the colons at Day 14, as did both sulfasalazine (50 mg/kg/day) and infliximab ( 3 mg/kg on Day 1 and 7), as shown in FIG. 12.

[00403] Colon Weight. As an indirect index of inflammatory oedema in the colonic tissue, the weight of the standard colonic segments was determined at the end of the study. The colonic weight in the CMC challenged group was significant higher at Day 14 than thai in the non-chaitenged, non-treated group (FIG. 13). Results are expressed as mean ± S E M , n=11-15 for the test groups and n=6 for the absolute control group; statistical significance is shown as ^*P<0.05, ^**P<Q 01 ^***P<0 001 compared with the challenged control CMC group.

[00404] Treatment with CCi-7506 at both 50 and 100 mg/kg/day caused a significant reduction in the colon weight determined on Day 14 (FiG.13) CCI-7507 (25 and 50 mg/kg/day) also significantly reduced the colonic weight of the standard segment at both doses compared with that in the CMC group (FIG.13). A significant (P<0.05) reduction in colon weight was also observed in the sulfasalazine group and in the infliximab group (FtG.13).

[00405J Colonic MPO Levels. The level of MPO activity in the colonic tissue from the TNBS- challenged group determined after 14 days was 521 + 56 mU/mg protein, being significantly different from that determined in the absolute control group, as shown in FIG, 14, Results are expressed as mean ± S EM., n=11-15 for the test groups and n=6 for the absolute control group; statisticaS significance is shown as ^**P<0.01 , ^***P<O.OO1 compared with the challenged control CMC group

[00406| Treatment with CCi-7506 (50 and 100 mg/kg/day) caused a significant fall in the elevated MPO activity deterrmned at 14 days, at both dose levels, as shown in FlG. 14. Treatment with CCI- 7507 (25 and 50 mg/kg/day) also caused a significant and dose-dependent fall in the elevated MPO activity at both dose levels, as shown in FlG. 14. Treatment with sulfasalazine (50 mg/kg/day) significantly (P<0.G01) reduced the elevated colonic levels of MPO as can be seen in FtG. 14, This effect was not significantly different from that observed with any of the other active treatment groups (data not shown). Intravenous injection of infliximab (3 mg/kg on Day 1 and on Day 7 after challenge) significantly attenuated the increase in MPO, following TNBS, observed on Day 14 (FIG. 14). This effect was not significantly greater than that observed with the lower or higher doses of either CCI- 7506 or CCI-7507, or that with sulfasalazine (FIG. 14)

[00407] Coϊonsc TNF-α Levels Challenge with TN8S significantly eievated the levels of TNF-α in the colonic tissue determined after 14 days compared with the absolute control groupCFIG. 15) Results are expressed as mean ± S E M., n=11-i5 for the test groups and n=6 for the absolute control group, statistical significance is shown as ^*P<0 05, ^*"P<O.G01 compared with the challenged control CMC group The level of TNF-α in the colonic tissue from TNBS-challenged rats, determined after 14 days after challenge was 589 + 66 pg/mg protein (FIG 15)

[00408] Treatment with CCI-7506 dose-dependently reduced the Sevel of TNF-α in the colonic tissues (FSG 15) Thus, whereas CCI-7506 (50 mg/kg/day) had no significant effect on the colonic TNF-α levels the higher dose of 100 mg/kg/day did achieve significance (FIG 15) Administration of either dose of CCI-7507 (25 and 50 rng/kg/dayj caused a Significant reduction in the elevated levels of TNF-α determined at Day 14 as shown in FIG 15 Treatment with sulfasalazine also significantly reduced the elevated colonsc levels of TNF-α observed on Day 14 following TNBS challenge, as can be seen m FiG 15 Intravenous injection of infliximab (3 mg/kg on Day 1 and on Day 7 after challenge) significantly attenuated the increase in TNF-α levels FIG 15 This effect was not significantly greater than that observed with either of the doses of CCI-7507, but was significantly greater than that achieved with the lower dose of CCI-7506 (50 mg/kg/day) or with sulfasalazine

[004091 This Example indicates that both CCI-7506 (50 and 100 mg/kg/day) and CCI-7507 ( 25 and 50 mg/kg/day) given by oral gavage twice daily commencing 1 day following challenge, dose- dependently reduce the degree of tissue injury a 14 day rat model of colitis, reducing macroscopic colonic injury at the doses of both agents employed The biomarkers of colonsc inflammation, colon weight and colonic MPO levels, along with TNF-α, the latter being an inflammatory mediator involved in colics, were also reduced in the inflamed tissue by these doses of CCI-7506 and CCI-7507 This profile of pharmacological actions of CCI-7506 and CCI-7507 were comparable to those seen with sulfasalazine, an agent used widely in the clinic in the therapy of IBD, and preliminary estimates could suggest a greater activity of CCI-7507 on macroscopic injury, and the other key btomarkers sn the chronic study Moreover, the profile of activity with the compounds was also comparable to those of infliximab

[00410] The foregoing is illustrative of the present invention, and is not to be construed as limiting thereof The invention is defined by the following claims with equivalents of the claims to be included therein

Claims

THAT WHICH IS CLAIMED IS:

1. A compound of Formula I or Formula II:

wherein:

A is N or C, subject to the proviso that R⁵ is absent when A is N;

X is -C(O)-, -S(O)₂-. or a covalent bond:

Y is alky!, alkenyl, cycloaikyl, alkyicycloalkyt, aikylcycloatkylalkyi, alkyloxyaSkyl, aryl, aikylaryl, alkylarytaikyi, aryfalkyl , cycioalkylaikyi, alkylheterocycie. heterocyclealkyl, alkyfheterocyclealkyl, heterocycle, aminoafkyl, oxyalkyl, aminoaryi, oxyaryl;

2 is selected from the group consisting of -B(OR¹JOR², ~C0N(R¹)0R², and -N(OR¹)COR²;

R¹ and R² are each independently H, loweralkyl, or together form C2-C4 alkylene, and

R³, R⁴, R^s, R⁶, and R⁷ are each independently selected from the group consisting of^' H, halo, loweralkyl, hatoloweralkyi, haloloweraikoxy, ioweralkoxy, hydroxy, toweralkoxycarbo, carboxylfc aαd, acyl, azido, mercapto, aikylthio, amino, heterocycleamino, aikylamsno, dtalkylamino, acyiamino, aminoacy), arylamino. arylalkyS, arylalkylamsno arySoxy, cyano. sulfonamide, aminosulfonyt, suifone. nitro; arylalkyloxy. cycloalkyfoxy, cycloafkylaikoxy, cycloalkylamino. urea, cycloalkylalkylamino. cycloaikyl, alkylcycloalkyl, hydroxyamϊno, alkoxyacylamtno, and aryithra; and 5- or 6- membered organic rings containing O to 4 heteroatoms selected from the group consisting of N, O and S₁ which πngs may be unsubstituted or substituted from 1 to 4 times with halo, loweraikyi haioloweralky!, haloloweraikytoxy, foweralkoxy, hydroxy, towerafkøxycarbσ, carboxylic acid acyl, azido, mercapto, alkylthso, amino, heterocycteammo. alkytamsno, dtalkyiamsno, acyiamino. aminoacyi aryiamino, arylalkyl, arylaSkyiamiπo aryloxy, cyano, sulfonamide aminosulfonyl suSfone, nrtro, and oxoheterocycltc groups or a pharrnaceuttcally acceptable salt or prodrug thereof

2 The compound of claim 1 , wherein R⁵ is selected from the group consisting of halo loweralkyl haioloweralkyl haioioweralkyloxy, loweralkoxy, hydroxy loweralkoxycarbo, carboxylic acid acyl, azido, mercapto, atkylthio ammo, heterocycleamino, alkyiamino, dialkylamtno, acylamtno, aminoacyl, aryiarmno, arylaiky!, arylaSkylamino, aryloxy, cyano, sulfonamide, aminosulfonyl, sulfone nitro, and hydroxyamino

3 The compound of clasm 1 , wherein R⁵ is selected from the group consisting of halo haiofoweralkyl, haloloweraikyioxy, loweralkoxy, ammo acylamino, aminoacyl arylalkyl, aryloxy, acyi, aryiamino, cyano nitro, and heterocycleamino

4 The compound of claim 1 wherein R⁵ is cyano, fluoroalky! or halo

5 The compound of claim 1 wherein R"¹ is H

6 The compound of claim 1 , wherein R⁴ is selected from the group consisting of hato, loweralkyl, haloloweralkyl haioloweralkyloxy, loweralkoxy, hydroxy, Soweralkoxycarbo, carboxyiic acid, acyl, azido, mercapto, aikylthio, amino, heterocycleamino, aikylamtno dialkylammo, acylamino, aminoacyl, aryiamino, arylalkyl, arylalkylamino, aryloxy, cyano, sulfonamide, amtnosulfonyl, sulfone, and mtro

7 The compound of claim 1 , wherein R⁴ is selected from the group consisting of halo, haloloweralkyl, haioioweralkyloxy, loweralkoxy, amino, acylamino, aminoacyl, arylaiky! aryloxy, acyi arylamino, cyano nttro, and heterocycleamino

8 The compound of ciatm 1 , wherein R⁴ is cyano fluoroalkyl or halo

9 The compound of claim 1 wherein R^e is H

10 The compound of claim 1 wherein R^β is selected from the group consisting of halo loweralkyi haloloweralkyi halαioweralkytoxy loweralkoxy hydroxy, lowerafkoxycarbo carboxylic acid, acyl, azido, mercapto aSkylthio amino, heterocycleamino, alkyiammo dialkylammo acyiamino, aminoacy! arylarmno arylalkyS arylalkylamino aryloxy cyano sulfonamide aminosulfonyl, sulfone, and nitro

11 The compound of claim 1 , wherein R^s is selected from the group consisting of halo haloloweralkyf hatoloweralkyloxy foweralkoxy amino acylamino aminoacyi arylalkyf aryloxy, acyl arylamsno cyano nitro and heterocycleamino

12 The compound of ciatm 1 wherein R⁶ rs cyano fluoroalkyl or halo

13 The compound of claim 1 , wheresn R⁷ is H

14 The compound of claim 1, wherein at least two of R⁴, R⁶ and R⁷ are H

15 The compound of ciaim 1, wherein R⁶ and R⁷ are H

16 The compound of claim 1, wherein A is N

17 The compound of ciaim 1, wherein A is C

18 The compound of claim 1 wherein R^J is a 5- or 6- rnembered organic ring containing 0 to 4 heteroatoms selected from the group consisting of N, O and S, which ring may be unsubstituted or substituted from 1 to 4 times with halo loweralkyl haloloweralkyi hatoloweralkyloxy loweraikoxy hydroxy toweraikoxycarbo, carboxylic acid, acyl azido, mercapto alkylthto amino, heterocycleamino, alkylamino dsalkylamino, acylamsno, amtnoacyl, arylamsno, arylalkyl, arylalkylamino, aryloxy cyano, sulfonamide aminosulfonyl sulfone, nitro, and oxoheterocyclic groups

19 The compound of claim 1, wherein said compound is selected from the group consisting of

4-(2-{Tπfluoromethyl)-1 H-benzo[d]ιmιdazol-1 ~yl)butylboronιc acid 5-{2-(Thιazol-4-yl)-1/-/-benzoJdjιmιdazol-1-y!)pentylboronsc acid, 5-(5,6-dιmethyl-1H-benzo[d]ιmsdazol~1~yl)pentylboromc acid, 5-(1 H-ιmιdazø[4,5-c]pyπdm-1-yl)peniylbofonϊc acid 5-(2-(4-Methoxyphenyl)-1H-benzo[d]ιmιdazoM-yl)pentylboronsc acid, 5-{2-{3-Ffuoro-4-methoxyphenyl)-1 H-benzo[d]ιm!dazo!-1-yf)penty!boronιc acid 5-(5-cyano-2-{4'methoxyphenyl)-1H-benzo[d|!mιdazol-1-yl)pentylboronιc acid 5-{6-cyano-2-(4-methoxyphenyl)-1 H-benzo|d]ιmιdazo1~1 -yi)pentylboronic acid and pharmaceutically acceptable salts and prodrugs thereof

20 The compound of claim 1 wherein said compound is 5-(2"(Tπιazo!-4-yl)-1 H- benzo[c/]ifnidazol-1-yl)pentylboronιc acsd and pharmaceutically acceptable salts and prodrugs thereof

21 The compound of claim 1 wherein said compound ts 5~{2^4-Methoxyphenylj-1H- beπzQ{djsmtøazαM-y!}pentyibøronιc acid and pharmaceutically acceptable safe ana prodrugs thereof

22 The compound of claim 1, wherein sad compound ts 5~(2-{3-Fluoro-4-methoxyphenyl)- 1H-benzo[d]imidazot-1-yl}pentylboronic acsd and pharmaceutically acceptable salts and prodrugs thereof

23 The compound of claim 1 , wherein said compound is selected from the group consisting of

5-{5"Cyano-2-(4-methoxyphenyl}-1 H-benzo[dJimidazol-1-yl)pentylborontc acsd. 5-(6-cyano-2-{4-methoxypheny!)-1 H-benzoEd]ιmιdazol-1-yl)pentylboronιc acid, and pharmaceutically acceptable salts and prodrugs thereof

24 A compound of Formula 111, Formula IV, or Formula V

wherein

X is -C(O)-, -S(O)₂-, or a covalent bond,

Y ss alky!, alkenyl, cycloalkyl, afkylcycloalkyi, alkylcycloaikylalkyS, aikyloxyalkyt, aryl, alkylaryl alkylarylalkyl, arylalkyi cycloaikylalkyl, alkylheterocycle, heterocyclealkyl, aikytheterocyclraikyl, heterocycle amiπoalkyl, oxyaikyf, amsnoaryl, or oxyaryl

Z is selected from the group consisting of -B(0R¹)0R² -C0N(R¹)0R^z and ™N(0R¹)C0R²,

R¹ and R² are each independently H loweraikyl or together form C2-C4 alkylene and

R³ R⁴ R⁶ R⁶ R' and R⁸ are each independently selected from the group consisting of H, halo loweralkyϊ, haloloweraSkyl haloloweralkoxy, loweraϊkoxy hydroxy loweralkoxycarbo carboxyhc acid acyl azido mercapto alkylthto amino heterocycleamino, aikylammo, dialkylarntno, acyiamino, amsnoacyl, aryiamino, arylaSkyl arylalkylamino aryloxy, cyano sulfonamide, aminosulfonyl, suifone nttro, aryfaikyloxy cycloaikyloxy, cycioalkylaikσxy. cycloalkylamino, urea, cycloalkySalkyiamtno, cycloalkyl, alkylcycloalkyl, hydroxyamino, alkoxyacylamino, and arylthio, and 5- or 6- membered organic rings containing O to 4 heteroatoms selected from the group consisting o! N, O and S₁ whsch rings may be unsubstituted or substituted from 1 to 4 times With halo, loweralkyl, haloloweralkyt, haloloweraikyloxy, ioweralkoxy, hydroxy, loweralkoxycarbo, carboxylsc acid, acyl, azido, mercapto, aikylthio, amino, heterocycleamino, aikylammo dsaikylamino, acylamino, ammoacyl, aryiamino arylalkyi, aryJaJkyiarmno, aryloxy cyano, sulfonamide, aminosulfonyl, suifone, and nitro, and oxoheterocycltc groups, or a pharmaceutically acceptable salt or prodrug thereof

25 The compound of claim 24, wherein R⁵ is selected from the group consisting of halo, loweralkyl, haloloweralkyl haloloweraikyloxy, Ioweralkoxy, hydroxy, loweralkoxycarbo, carboxylfc acfd, acyl azido mercapto, atkylthio amino heterocycleamino, aikyiammo, dialkylammo acyfamino amsnoacyl, aryiamino, arylalkyi, arylalkylamino aryloxy cyano sulfonamide, aminosulfonyl, suffone and mtro

26 The compound of claim 24, wherein R^ε is selected from the group consisting of halo, haioioweralkyl, haloloweraikyioxy, Soweralkoxy, amino, acytamtno, aminoacyl, arylalkyi, aryloxy, acyl, aryiamino, cyano, nrtro, and heterocycleamino

27 The compound of claim 24, wherein R⁵ is cyano, fluoroalkyl or haϊo

28 The compound of claim 24 wherein R⁴ is H

29 The compound of claim 24 wherein R⁴ is selected from the group consisting of halo loweralkyl haioioweralkyl, haloloweraikyloxy Ioweralkoxy hydroxy loweralkoxycarbo carboxyte aαd acyl azido mercapto aikylthio ammo heterocycfeamino sikylamsno diatkylamsno acylarnsπo aminoacyt. arylamino, arylalkyl, arylalkylamino, aryloxy. cyano, sulfonamide, aminosulfony!, sυifone, and nrtro

30 The compound of ciatm 24, wherein R⁴ is selected from the group consisting of haio, haloloweralkyl, haloloweraikyloxy, ioweratkoxy, amino, acylanrnno, amsπoacyl aryialkyl, aryioxy, acyl arylarnino, cyano. nrtro, and heterocycteamino

31 The compound of claim 24, wherein R⁴ is cyano, fluoroalky! or halo

32 The compound of claim 24, wherein R^s is H

33 The compound of claim 24 wherein R^e is selected from the group consisting of halo, toweralkyl, halofoweralkyl halofoweralkyloxy, loweralkoxy, hydroxy, loweralkoxycarbo. carboxylic acid, acyl, azido, mercapto, alkylthso, amino, heterocycleamino, alkyiamino, dialkylamtno, acylamino, aminoacyl, arylamino, arylaikyi, arySalkylamino, aryloxy, cyano, sulfonamide, aminosulfonyl, suifone, and πitro

34 The compound of ciairn 24, wherein R⁶ is selected from the group consisting of halo, haloloweralkyl, haloloweraikyloxy, loweralkoxy, ammo, acylamino, aminoacyl, arylalkyl, aryloxy, acyl, arylammo, cyano, nitro, and heterocycleamino

35 The compound of claim 24, wherein R⁶ is cyano, fiuoroalkyi or halo

36 The compound of claim 24, wherein R⁷ is H

37 The compound of claim 24, wherein R⁷ is selected from the group consisting of halo loweralkyl, haloloweraikyi haloloweraikyloxy, loweralkoxy, hydroxy, loweralkoxycarbo, carboxyjic acid, acyl, azido mercapto, alkyithio, ammo heterocycleamtno, alkylamino, dtalkylamino acylamino, aminoacyl, aryiamino, arylalkyl arylalkylammo, aryfoxy, cyano, sulfonamide arruπosuifonyl, sulfone and nrtro

38 The compound of claim 24. wherein R⁷ is selected from the group consisting of halo, haioloweralkyϊ, hafoloweraikyloxy loweraikoxy ammo, acylamino, ammoacyl, arylaikyi, aryioxy, acyi, arylamtno, cyano, nitro and heterocycleamsπo

39 The compound of ciasm 24 wheretn R⁷ is cyano, fluoroalkyl or halo

40 The compound of clasm 24 wherein at least two of R⁴ R⁶ and R⁷ are H

41. The compound of claim 24, wherein R⁶ and R⁷ are H.

42. The compound of claim 24, wherein R⁴ and R⁶ are H.

43. The compound of claim 24, wherein R^E and R^f are H.

44. The compound of claim 24, wherein R⁴ and R⁵ are H.

45. The compound of claim 24, wherein said compound is selected from the group consisting of:

5-(5-cyano-1H-ιndol-1~y!)pentylboronιc acid; and pharmaceutically acceptable salts and prodrugs thereof.

46. A compound of Formula Vl:

wherein,

A ss S. O₁ SO₂ or NR₁ X is -C(O)-, -S(O)₂-, or a covaient bond;

Y is aikyi, alkenyl, cycloalkyl, alkylcycloalkyl, alkylcycloalkylalkyl, alkyloxyaikyS, aryl, alkylaryi, alkylarylalkyl, arylalkyl, cycloalkylalkyS alkylheterocycSe, heterocyclealkyl, alkySheterocyctealkyl, heterocycle, aminoalkyl, oxyalkyl, aminoaryl oxyary!.

Z is selected from the group consisting of -8(OR¹)OR² -CON(R¹)OR². and -N(OR¹)COR², R¹ and R' are each independently H. loweraiky!, or together form C2-C4 afkylene; and R, R³, R*. R⁵, R⁶, R⁷ R^a _. R⁹ and R^1C are each independently selected from the group consisting of. H. haio, loweraiky!, hafoloweraϊkyl, haloloweraikoxy, ϊoweralkoxy, hydroxy, foweralkoxycarbo, cycloalkyl, alkyicycloalkyi, carboxylic acid, acyl, azido, mercapto, alkySihio, amino, heterocycleamino, alkySamino, dialkylamino, acylamino, aminoacyl, arylamiπo, arylalkyi, arylalkylamino, aryloxy, cyano, sulfonamide, aminosulfonyl. sulfone, nitro, arylalkyloxy, cycioatkyloxy, cycloaikylafkoxy, cycloalkyiamifio. urea cydoalkyiβlkytamfnø. cycloatk/l, aikylcycloafkyE, hydroxyamtπo, alkoxyacyiamino and aryfthio. and 5- or 6- membered organic rings containing 0 to 4 heteroatoms selected from the group consisting of N O and S, which rings may be υnsubstituted or substituted from 1 to 4 times with halo loweralky! haiotoweralkyl, haloioweralkyloxy, loweralkoxy hydroxy, (oweraikoxycarbo, carboxyiic acid, acyl, azido mercapto alkylthio amino, heterocyclearmnα, alkylamino dialkylamino acylamino, aminoacyl, arylamsno, arylaSkyi, arylalkylamtno, aryloxy, cyano, sulfonamide, amiπosulfonyl sulfone, nitro, and oxoheterocyclic groups, or R^s and R⁹ together are =0 or =S, or a pharmaceutically acceptable salt or prodrug thereof

47 The compound of claim 46, wherein at least one of R³, R⁴, R⁵ R⁶, R⁷ or R^B ts not H

48 The compound of claim 46, wherein R⁵ is selected from the group consisting of halo, loweralkyl haloϊowerafkyi, haloloweralkyloxy, loweralkoxy, hydroxy, loweralkoxycarbo, carboxyiic acid, acyS, azido, mercapto, alkylthio, amino, heterocycSeamino, alkylamino, dialkylamino, acylamino, aminoacyl arylamino, arylalky! arylalkySamino aryloxy, cyano, sulfonamide, aminosulfonyl, sulfone, nitro and hydroxyamino

49 The compound of claim 46, wherein R⁵ is selected from the group consisting of halo, haioloweralkyl, haloloweralkyloxy, loweralkoxy, amino, acylamino, aminoacyl, arylaikyl, aryloxy, acyl, arylamino, cyano, nitro, and heterocycleamino

50 The compound of claim 46, wherein R⁵ is cyano, fϊuoroaikyl or halo

51 The compound of claim 46, wherein R⁴ is H

52 The compound of claim 46, wherein R⁴ is selected from the group consisting of halo, loweraikyl, haioloweralkyl, haloioweralkyloxy loweralkoxy hydroxy, loweralkoxycarbo, carboxyiic acid acyl, azido, mercapto, afkyllhio, amino heterocycleamino alkytamino, dialkylamino, acylamino aminoacyl, aryiamino, arylalkyS, arylaikyiamsrso aryfoxy, cyano sulfonamide amtnosulfonyl, sulfone nitro and heterocycleamino

53 The compound of claim 46 wherein R⁴ is selected from the group consisting of halo haioioweralkyl, haloloweralkyloxy loweralkoxy, ammo acylamtπo, aminoacyl aryiaikyl arySoxy acyl, aryfamino cyano nitro and heterocycleamino

54 The compound of claim 46 wherein R⁴ ss cyano fluoroalkyl or halo

55 The compound of claim 46 wherein R⁶ is H

56 The compound of claim 46. wherein R⁶ is selected from the group consisting of halo, loweraikyl haioloweralkyl, haloloweralkyloxy, ioweralkoxy, hydroxy loweraikoxycarbo, carboxyiic acid, acyl, azido, mercapto, alkylthio, ammo, heterocycleammo. alkylamino dialkylamino, acylamino, amiπoacyl, arylamino, arylalkyl. arytalkylamino. aryloxy, cyano. sulfonamide, aminosuifonyi, sulfone, and nitro.

57. The compound of claim 46, wherein R⁶ is selected from the group consisting of: halo, haloloweralkyi, hafoloweralkyloxy, Ioweralkoxy, amino, acylamino, aminoacyl, arylalkyi, aryloxy, acyl. aryjamino, cyano, nitro, and heterocycleamiπo.

58. The compound of claim 46, wherein R⁶ is cyano, fluoroaikyl or halo

59. The compound of claim 46, wherein R⁷ is H.

60. The compound of claim 46, wherein at least two of R⁴, R⁶, and R⁷ are H.

61. The compound of claim 46, wherein R⁶ and R⁷ are H.

62. The compound of claim 46, wherein R is selected from the group consisting of H, ioweralkyl, halotoweralkyl, halofoweralkyioxy, ioweralkoxy, loweraikoxycarbo. carboxyiic acid, acyl, acylamino, aminoacyl, arylalkyl, cyano, sulfonamide, aminosulfonyi, and sulfone.

63. The compound of claim 46, wherein R is selected from the group consisting of H, loweralkyl, haloloweralkyi, haloloweraikyloxy, ioweralkoxy, !owera!koxycarbo. and arylalkyi.

64 The compound of claim 46, wherein R³ is selected from the group consisting of H, alkyl, aryl, heteraryl, and cycloaikyl,

65. The compound of claim 46, wherein R⁸ and R⁹ are each independently selected from the group consisting of H and loweralkyl, or R⁸ and R⁹ are together =0 or =S

66 The compound of claim 46, wherein R^s and R¹⁰ are both H

6? The compound of clatm 46, wherein said compound ϊs seiected from the group consisting of:

5-(6-fluoro-2,3-dihydro-3-oxobenzo[b][1.4joxazm-4-yl)pentylboronιc acid; 5-(2,3-dihydro-3-oxobenzo[bJ[1 4]thϊazin-4-yl)ρenty!boronic acid; 5-{7-chioro-2.3-dihydro-3-oxoben2θ{b][1,4]th)azιn-4-yl)peπtylboronic acid; 5-(2.3-dEhydro-?-nrtro-3-oxobenzo|bl1.4joxaz!π-4-y!)per}tyiboronic acid; 5-(2,3-dιhydro3-oxobenzo[b][1 ,4]oxazin-4-yl)penty!boromc acid; ethyl 2-(3,4-dihydro-3-oxo-4-(5-pentylboronic acid)-2H-benzo[bJ[1.4]thιazιn-2-yl)acetate; and pharmaceutically acceptable salts and prodrugs thereof,

68. The compound of claim 46, wherein said compound is selected from the group consisting of:

5-{6-fiuoro-2_¥3-d!hydro-3-oxobenzo[bJ[1.4]oxazin-4-yi)penty!boroπic acid; 5-{7~ch!oro-2.3-dihydro-3-oxobeπzo[b]t1,4]thiazm-4-yI)pentylboronic acsd: and pharmaceutically acceptable salts and prodrugs thereof.

69. A pharmaceutical composition comprising a compound according to any one of claims Claims 1 , 24 and 46 in a pharmaceutically acceptable carrier.

70. The composition of claim 69, wherein said carrier is an aqueous carrier.

71 A pharmaceutical composition comprising a compound according to any one of Claims 1, 24 and 46 in a pharmaceutically acceptable carrier

72 The composition of claim 69, wherein said carrier is an aqueous carrier.

73. A compound of Formula VII

wherein:

A₁ and A₂ are each independently N or C;

X is -C(O)- -S(O)₂-, or a covalent bond,

Y is alkyl alkenyl cycloalkyl. alkylcycloalkyl. alkylcycloalkylalkyl, alkyloxyafky! ary!, alkytaryl. a.kylarylalkyl, aryfalkyt , cycjoalkylalkyl, alkyjheterocycle, heterocycleafkyl, alkylheterocycteatkyf, heterocycie, armnoalkyl, oxyafkyl aminoaryt oxyaryl.

Z is selected from the group consisting of -BsOR¹JOR², ~CQNtR')OR^z, and -N(OPJ)COR² R¹ and R² are each independently H, loweraikyl, or together form C2-G4 alkylene, and R_n and R_p are each independently selected from the group consisting of H, halo, loweraikyl, hatoloweralkyl haloloweralkoxy loweralkoxy, hydroxy, loweralkoxycarbo, carboxyϊic acid acyi, azfdo, mercapto, alkylthso, amino, heSerocycleamino, alkyianmno, dialkylamino acylamino aminoacyl arylammo, arylalkyl arylalkylammo, aryloxy, cyano, sulfonamide, amtnosulfoπyl sulfone nitro, arylaikyloxy, cycloalkyloxy, cycloalkyialkoxy, cycioalkylamino, urea, cycloafkylalkylamsno cycloalkyl, alkylcycloalkyl hydroxyamino. alkoxyacylamsno, and arylthio and 5- or 6- mernbered organic rings containing 0 to 4 heteroatorns selected from the group consisting of N O and S which rings may be unsubstttuted or substituted from 1 to 4 times with halo loweraikyl, haloloweraikyl, haloloweralkyloxy Soweralkoxy hydroxy, loweralkoxycarbo, carboxylsc acsd, acyl aztdo, mercapto, alkySthio, amino, heterocycteamiπo, alkylamino dialkylamino, acylamino, aminoacyl, arylammo, arylalkyl, arylalkylamino, aryioxy, cyano, sulfonamide, aminosulfonyl, sulfone, niiro, and oxoheterocychc groups, subject to the proviso that when A₁ ss C, then n=1 to 4. when A₁ is N, then n=1 to 3, A₂ is C, then p= 1 to 2, when A₂ is N₁ then π=1 or a pharmaceutically acceptable salt or prodrug thereof

74 A method of inhibiting tumor necrosis factor alpha in a subject in need thereof, comprising administering a compound according to any one of Ciaims 1 , 24 46, and 74 to said subject in an amount effective to inhibit tumor necrosis factor alpha

75 A method of inhibiting phosphodiesterase in a subject in need thereof, comprising administering a compound according to any one of Claims 1, 24, 46, and 74 to said subject in an amount effective to inhibit phosphodiesterase

76 A method of claim 754, wherein said phosphodiesterase (PDE) is selected from the group consisting of PDE II, PDE III, PDE IV, PDE V and combinations thereof

77 A method of treating an inflammatory disease in a subject in need thereof, comprising administering a compound according to any one of Claims 1 , 24, 46, and 74 to said subject in an amount effective to treat sasd inflammatory disease

78 A method of treating inflammatory bowel disease in a subject in need thereof comprising administering a compound according to any one of Claims 1 , 24 46, and 74 to said subject in an amount effective to treat inflammatory bowei disease

79 A method of treating rheumatoid arthritis in a subject in need thereof comprising administering a compound according to any one of Claims 1 , 24, 46 and 74 to sasd subject m an amount effective to treat rheumatoid arthritis 80 A method of treating psoriasis in a subject tn need thereof, comprising administering a compound according to any one of Claims 1 , 24, 46 and 74 to said subject in an amount effective to treat psoriasis

81 A method of treating ankylosing spondylitis in a subject in need thereof, comprising administering a compound according to any one of Claims 1, 24, 46, and 74 to sasd subject tn an amount effective to treat ankylosing spondylitis

82 A method of treating psoriatic arthπtis in a subject in need thereof comprising administering a compound according to any one of Claims 1 24 46 and 74 to said subject in an amount effective to treat psoriatic arthritis

83 A method of treating asthma tn a subject in need thereof comprising administering a compound according to any one of Claims 1, 24 46, and 74 to said subject in an amount effective to treat asthma

84 A method of treating chronic obstructive pulmonary disease m a subject in need thereof comprising administering a compound according to any one of 1 , 24 46, and 74 to said subject sn an amount effective to treat chronic obstructive pulmonary disease

85 A method of treating Alzheimer's disease in a subject in need thereof, comprising administering a compound according to any one of Claims 1 , 24, 46, and 74 to said subject in an amount effective to treat Alzheimer's disease

86 A method of treating type Il diabetes in a subject in need thereof comprising administering a compound of according to any one of Claims 1, 24, 46, and 74 to said subject in an amount effective to treat type Il diabetes

87 A method of treating cancer tn a subject tn need thereof, comprising administering a compound accordrng to any one of Claims 1 24, 46 and 74 to said subject in an amount effective to treat cancer

88 A method of treating hypertension in a subject in need thereof, comprising administering a compound of according to any one of Claims 1 24 46 and 74 to satd subject in an amount effective to treat hypertension

89 A method of treating erectile dysfunction in a subject in need thereof comprising adminssterfng a compound according to any one of 1 24 46 and 74 to sasd subject in an amount effective to treat erectile dysfunetσn