Deprecated: The each() function is deprecated. This message will be suppressed on further calls in /home/zhenxiangba/zhenxiangba.com/public_html/phproxy-improved-master/index.php on line 456
AU774054B2 - Inhibitors of LFA-1 binding to ICAMs and uses thereof - Google Patents
[go: Go Back, main page]

AU774054B2 - Inhibitors of LFA-1 binding to ICAMs and uses thereof - Google Patents

Inhibitors of LFA-1 binding to ICAMs and uses thereof Download PDF

Info

Publication number
AU774054B2
AU774054B2 AU41919/00A AU4191900A AU774054B2 AU 774054 B2 AU774054 B2 AU 774054B2 AU 41919/00 A AU41919/00 A AU 41919/00A AU 4191900 A AU4191900 A AU 4191900A AU 774054 B2 AU774054 B2 AU 774054B2
Authority
AU
Australia
Prior art keywords
sulfide
chlorophenyl
amino
chloro
phenylamine
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Ceased
Application number
AU41919/00A
Other versions
AU4191900A (en
Inventor
Janet Adolphson
Kerry Fowler
Mark Orme
Donald E. Staunton
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Icos Corp
Original Assignee
Icos Corp
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Icos Corp filed Critical Icos Corp
Publication of AU4191900A publication Critical patent/AU4191900A/en
Application granted granted Critical
Publication of AU774054B2 publication Critical patent/AU774054B2/en
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

Links

Classifications

    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D207/00—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom
    • C07D207/02—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom with only hydrogen or carbon atoms directly attached to the ring nitrogen atom
    • C07D207/30—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom with only hydrogen or carbon atoms directly attached to the ring nitrogen atom having two double bonds between ring members or between ring members and non-ring members
    • C07D207/34—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom with only hydrogen or carbon atoms directly attached to the ring nitrogen atom having two double bonds between ring members or between ring members and non-ring members with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals, directly attached to ring carbon atoms
    • C07D207/36—Oxygen or sulfur atoms
    • C07D207/40—2,5-Pyrrolidine-diones
    • C07D207/416—2,5-Pyrrolidine-diones with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals, directly attached to other ring carbon atoms
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P19/00—Drugs for skeletal disorders
    • A61P19/02—Drugs for skeletal disorders for joint disorders, e.g. arthritis, arthrosis
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A61P35/04—Antineoplastic agents specific for metastasis
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P37/00—Drugs for immunological or allergic disorders
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P37/00—Drugs for immunological or allergic disorders
    • A61P37/02—Immunomodulators
    • A61P37/06—Immunosuppressants, e.g. drugs for graft rejection
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C323/00—Thiols, sulfides, hydropolysulfides or polysulfides substituted by halogen, oxygen or nitrogen atoms, or by sulfur atoms not being part of thio groups
    • C07C323/01—Thiols, sulfides, hydropolysulfides or polysulfides substituted by halogen, oxygen or nitrogen atoms, or by sulfur atoms not being part of thio groups containing thio groups and halogen atoms, or nitro or nitroso groups bound to the same carbon skeleton
    • C07C323/09—Thiols, sulfides, hydropolysulfides or polysulfides substituted by halogen, oxygen or nitrogen atoms, or by sulfur atoms not being part of thio groups containing thio groups and halogen atoms, or nitro or nitroso groups bound to the same carbon skeleton having sulfur atoms of thio groups bound to carbon atoms of six-membered aromatic rings of the carbon skeleton
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C323/00—Thiols, sulfides, hydropolysulfides or polysulfides substituted by halogen, oxygen or nitrogen atoms, or by sulfur atoms not being part of thio groups
    • C07C323/23—Thiols, sulfides, hydropolysulfides or polysulfides substituted by halogen, oxygen or nitrogen atoms, or by sulfur atoms not being part of thio groups containing thio groups and nitrogen atoms, not being part of nitro or nitroso groups, bound to the same carbon skeleton
    • C07C323/31—Thiols, sulfides, hydropolysulfides or polysulfides substituted by halogen, oxygen or nitrogen atoms, or by sulfur atoms not being part of thio groups containing thio groups and nitrogen atoms, not being part of nitro or nitroso groups, bound to the same carbon skeleton having the sulfur atom of at least one of the thio groups bound to a carbon atom of a six-membered aromatic ring of the carbon skeleton
    • C07C323/33—Thiols, sulfides, hydropolysulfides or polysulfides substituted by halogen, oxygen or nitrogen atoms, or by sulfur atoms not being part of thio groups containing thio groups and nitrogen atoms, not being part of nitro or nitroso groups, bound to the same carbon skeleton having the sulfur atom of at least one of the thio groups bound to a carbon atom of a six-membered aromatic ring of the carbon skeleton having at least one of the nitrogen atoms bound to a carbon atom of the same non-condensed six-membered aromatic ring
    • C07C323/35—Thiols, sulfides, hydropolysulfides or polysulfides substituted by halogen, oxygen or nitrogen atoms, or by sulfur atoms not being part of thio groups containing thio groups and nitrogen atoms, not being part of nitro or nitroso groups, bound to the same carbon skeleton having the sulfur atom of at least one of the thio groups bound to a carbon atom of a six-membered aromatic ring of the carbon skeleton having at least one of the nitrogen atoms bound to a carbon atom of the same non-condensed six-membered aromatic ring the thio group being a sulfide group
    • C07C323/37—Thiols, sulfides, hydropolysulfides or polysulfides substituted by halogen, oxygen or nitrogen atoms, or by sulfur atoms not being part of thio groups containing thio groups and nitrogen atoms, not being part of nitro or nitroso groups, bound to the same carbon skeleton having the sulfur atom of at least one of the thio groups bound to a carbon atom of a six-membered aromatic ring of the carbon skeleton having at least one of the nitrogen atoms bound to a carbon atom of the same non-condensed six-membered aromatic ring the thio group being a sulfide group the sulfur atom of the sulfide group being further bound to a carbon atom of a six-membered aromatic ring
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C323/00—Thiols, sulfides, hydropolysulfides or polysulfides substituted by halogen, oxygen or nitrogen atoms, or by sulfur atoms not being part of thio groups
    • C07C323/23—Thiols, sulfides, hydropolysulfides or polysulfides substituted by halogen, oxygen or nitrogen atoms, or by sulfur atoms not being part of thio groups containing thio groups and nitrogen atoms, not being part of nitro or nitroso groups, bound to the same carbon skeleton
    • C07C323/39—Thiols, sulfides, hydropolysulfides or polysulfides substituted by halogen, oxygen or nitrogen atoms, or by sulfur atoms not being part of thio groups containing thio groups and nitrogen atoms, not being part of nitro or nitroso groups, bound to the same carbon skeleton at least one of the nitrogen atoms being part of any of the groups, X being a hetero atom, Y being any atom
    • C07C323/40—Y being a hydrogen or a carbon atom
    • C07C323/41—Y being a hydrogen or an acyclic carbon atom
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D213/00—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members
    • C07D213/02—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members
    • C07D213/04—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members having no bond between the ring nitrogen atom and a non-ring member or having only hydrogen or carbon atoms directly attached to the ring nitrogen atom
    • C07D213/60—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members having no bond between the ring nitrogen atom and a non-ring member or having only hydrogen or carbon atoms directly attached to the ring nitrogen atom with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals, directly attached to ring carbon atoms
    • C07D213/62—Oxygen or sulfur atoms
    • C07D213/70—Sulfur atoms
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D215/00—Heterocyclic compounds containing quinoline or hydrogenated quinoline ring systems
    • C07D215/02—Heterocyclic compounds containing quinoline or hydrogenated quinoline ring systems having no bond between the ring nitrogen atom and a non-ring member or having only hydrogen atoms or carbon atoms directly attached to the ring nitrogen atom
    • C07D215/16—Heterocyclic compounds containing quinoline or hydrogenated quinoline ring systems having no bond between the ring nitrogen atom and a non-ring member or having only hydrogen atoms or carbon atoms directly attached to the ring nitrogen atom with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals, directly attached to ring carbon atoms
    • C07D215/36—Sulfur atoms
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D235/00—Heterocyclic compounds containing 1,3-diazole or hydrogenated 1,3-diazole rings, condensed with other rings
    • C07D235/02—Heterocyclic compounds containing 1,3-diazole or hydrogenated 1,3-diazole rings, condensed with other rings condensed with carbocyclic rings or ring systems
    • C07D235/04—Benzimidazoles; Hydrogenated benzimidazoles
    • C07D235/06—Benzimidazoles; Hydrogenated benzimidazoles with only hydrogen atoms, hydrocarbon or substituted hydrocarbon radicals, directly attached in position 2
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D277/00—Heterocyclic compounds containing 1,3-thiazole or hydrogenated 1,3-thiazole rings
    • C07D277/60—Heterocyclic compounds containing 1,3-thiazole or hydrogenated 1,3-thiazole rings condensed with carbocyclic rings or ring systems
    • C07D277/62—Benzothiazoles
    • C07D277/68—Benzothiazoles with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals, directly attached in position 2
    • C07D277/70—Sulfur atoms
    • C07D277/74—Sulfur atoms substituted by carbon atoms
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D295/00—Heterocyclic compounds containing polymethylene-imine rings with at least five ring members, 3-azabicyclo [3.2.2] nonane, piperazine, morpholine or thiomorpholine rings, having only hydrogen atoms directly attached to the ring carbon atoms
    • C07D295/04—Heterocyclic compounds containing polymethylene-imine rings with at least five ring members, 3-azabicyclo [3.2.2] nonane, piperazine, morpholine or thiomorpholine rings, having only hydrogen atoms directly attached to the ring carbon atoms with substituted hydrocarbon radicals attached to ring nitrogen atoms
    • C07D295/08—Heterocyclic compounds containing polymethylene-imine rings with at least five ring members, 3-azabicyclo [3.2.2] nonane, piperazine, morpholine or thiomorpholine rings, having only hydrogen atoms directly attached to the ring carbon atoms with substituted hydrocarbon radicals attached to ring nitrogen atoms substituted by singly bound oxygen or sulfur atoms
    • C07D295/096—Heterocyclic compounds containing polymethylene-imine rings with at least five ring members, 3-azabicyclo [3.2.2] nonane, piperazine, morpholine or thiomorpholine rings, having only hydrogen atoms directly attached to the ring carbon atoms with substituted hydrocarbon radicals attached to ring nitrogen atoms substituted by singly bound oxygen or sulfur atoms with the ring nitrogen atoms and the oxygen or sulfur atoms separated by carbocyclic rings or by carbon chains interrupted by carbocyclic rings
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D295/00—Heterocyclic compounds containing polymethylene-imine rings with at least five ring members, 3-azabicyclo [3.2.2] nonane, piperazine, morpholine or thiomorpholine rings, having only hydrogen atoms directly attached to the ring carbon atoms
    • C07D295/04—Heterocyclic compounds containing polymethylene-imine rings with at least five ring members, 3-azabicyclo [3.2.2] nonane, piperazine, morpholine or thiomorpholine rings, having only hydrogen atoms directly attached to the ring carbon atoms with substituted hydrocarbon radicals attached to ring nitrogen atoms
    • C07D295/12—Heterocyclic compounds containing polymethylene-imine rings with at least five ring members, 3-azabicyclo [3.2.2] nonane, piperazine, morpholine or thiomorpholine rings, having only hydrogen atoms directly attached to the ring carbon atoms with substituted hydrocarbon radicals attached to ring nitrogen atoms substituted by singly or doubly bound nitrogen atoms
    • C07D295/135—Heterocyclic compounds containing polymethylene-imine rings with at least five ring members, 3-azabicyclo [3.2.2] nonane, piperazine, morpholine or thiomorpholine rings, having only hydrogen atoms directly attached to the ring carbon atoms with substituted hydrocarbon radicals attached to ring nitrogen atoms substituted by singly or doubly bound nitrogen atoms with the ring nitrogen atoms and the substituent nitrogen atoms separated by carbocyclic rings or by carbon chains interrupted by carbocyclic rings
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D307/00—Heterocyclic compounds containing five-membered rings having one oxygen atom as the only ring hetero atom
    • C07D307/77—Heterocyclic compounds containing five-membered rings having one oxygen atom as the only ring hetero atom ortho- or peri-condensed with carbocyclic rings or ring systems
    • C07D307/87—Benzo [c] furans; Hydrogenated benzo [c] furans
    • C07D307/90—Benzo [c] furans; Hydrogenated benzo [c] furans with an oxygen atom in position 1 and a nitrogen atom in position 3, or vice versa

Landscapes

  • Chemical & Material Sciences (AREA)
  • Organic Chemistry (AREA)
  • Health & Medical Sciences (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • General Chemical & Material Sciences (AREA)
  • Medicinal Chemistry (AREA)
  • Veterinary Medicine (AREA)
  • Public Health (AREA)
  • Immunology (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Animal Behavior & Ethology (AREA)
  • General Health & Medical Sciences (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Engineering & Computer Science (AREA)
  • Rheumatology (AREA)
  • Transplantation (AREA)
  • Pain & Pain Management (AREA)
  • Oncology (AREA)
  • Orthopedic Medicine & Surgery (AREA)
  • Physical Education & Sports Medicine (AREA)
  • Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
  • Pyridine Compounds (AREA)
  • Nitrogen And Oxygen Or Sulfur-Condensed Heterocyclic Ring Systems (AREA)
  • Plural Heterocyclic Compounds (AREA)
  • Control Of El Displays (AREA)
  • Organic Low-Molecular-Weight Compounds And Preparation Thereof (AREA)
  • Quinoline Compounds (AREA)
  • Thiazole And Isothizaole Compounds (AREA)
  • Furan Compounds (AREA)
  • Pyrrole Compounds (AREA)

Description

WO 00/59878 PCT/USOO/08840 INHIBITORS OF LFA-1 BINDING TO ICAMs AND USES THEREOF
BACKGROUND
The leukocyte function-associated antigen (LFA-1, CD11 a/CD18) is a leukocyte-specific P2 integrin that participates in cell/cell adhesion. Binding activity of LFA-1 is essential to leukocyte extravasation from circulation to a site of injury in an inflammatory response. Three principle ligands are known to bind LFA-1, ICAM- 1, ICAM-2, and ICAM-3, which are intercellular adhesion molecules that play an important role in localizing leukocyte adhesion to endothelial cells at a site of injury.
ICAM-4 and ICAM-5 have also been reported to bind LFA-1. Most leukocytes constitutively express LFA-1, but ligand binding requires activation believed to induce a conformational change and increased avidity ligand binding. For example, ICAM-1 is normally expressed at low levels on the endothelium, however, injuryinduced inflammatory mediators promote enhanced surface expression in cells at the site of the injury which, in turn, promotes localized leukocyte adhesion through binding with activated LFA-1.
The structure of LFA-1 includes distinct intracellular and extracellular domains that are believed to participate and/or regulate ICAM binding. Of particular interest is a region in the cL chain of approximately 200 amino acids, designated the I domain, that is found in all P, integrins, as well as many other proteins. Evidence suggests that the I domain is essential to LFA-1 binding to ICAM-1 and 3. For example, anti-LFA-1 blocking monoclonal antibodies have been mapped to epitopes within the I domain. In addition, recombinant I domain polypeptide fragments have been shown to inhibit integrin-mediated adhesion and bind ICAM-1. Within the I domain of LFA-1 (and other proteins) is a single metal ion dependent adhesion site (MIDAS) that preferentially binds manganese or magnesium ions. Binding of either cation is required for ligand interaction and is believed to induce conformational changes in LFA-1 necessary for binding. Cation binding may therefore be a regulatory mechanism that responds to changes in the extracellular leukocyte environment. This hypothesis is supported by the observation that calcium ion WO 00/59878 PCT/USOO/08840 binding actually inhibits LFA-1 interaction with ICAM-1. Indeed, it has been proposed that an inactive LFA-1 conformation results from calcium binding, and that replacement of the calcium ion with a manganese or magnesium ion is a step required for LFA-1 activation [Griggs, et al., J. Biol. Chem. 273:22113-22119 (1998)]. Other factors have also been shown to induce LFA-1 activation, including T cell receptor engagement, cytokine stimulation, and in vitro PMA stimulation.
In practical terms, the identification of LFA-1/ICAM binding sites provides targets to modulate leukocyte inflammatory responses. Numerous antibodies have been isolated that are capable of inducing LFA-1 activation [see, for example, Landis, et al., J. Cell Biol. 120:1519-1527 (1993)] or, for example, preventing ICAM- 1 interaction [see for example, Randi and Hogg, J. Biol. Chem. 269:12395-12398 (1994)]. The previous identification ofanti-LFA-1 activating antibodies that recognize multiple and distinct extracellular epitopes suggests the existence of more than one regulatory region, presumably independent of cytoplasmic signaling.
Localization of LFA-1 sites that bind ICAM-1 has been investigated through use of chimeric LFA-1 a subunit proteins comprising human and murine components [Huang and Springer, J. Biol. Chem. 270:19008-19016 (1995)]. Studies have indicated that residues that coordinate cation binding and residues proximal to the site are essential for binding ICAM-1 at a relatively flat interface. More precise delineation of the extracellular regulatory region(s) and the contact points for ICAM-1 binding will permit design of efficient modulators.
Thus there exists a need in the art to precisely identify regulatory regions for proteins that participate in inflammatory responses, and in particular LFA- 1 and ICAMs that bind LFA-1. Determining the tertiary (or quartenary) structure of a protein can identify potential regulatory regions to permit the rational design of biologically compatible small molecules for therapeutic and prophylactic intervention for inflammatory disorders. There further exists a need in the art to identify compounds that can inhibit LFA-1 binding to ICAMs that can be used in the treatment of inflammatory disorders.
The discussion of the background to the invention herein is included to explain the context of the invention. This is not to be taken as an admission that any of the material referred to was published, known or part of the common general knowledge in Australia as at the priority date of any of the claims.
Throughout the description and claims of the specification the word "comprise" and variations of the word, such as "comprising" and "comprises", is not intended to exclude other additives, components, integers or steps.
e *o W:\ciska\nki\spedes\4191d.doc WO 00/59878 PCT/US00/08840 SUMMARY OF THE INVENTION The present invention is directed to compounds that bind to a novel regulatory site in the I domain of LFA-1, and thereby inhibit LFA-1 binding to ICAMs that bind LFA-1. The present invention therefore also provides methods to regulate leukocyte adhesion to endothelial cells. Compounds of the invention are useful for the treatment of pathologies, such as those associated inflammatory diseases, autoimmune diseases, tumor metastasis, allograft rejection and reperfusion injury. In particular, the present invention is directed to diaryl sulfides of general structural formula a pharmaceutically acceptable salt, or prodrug thereof, and to the use of diaryl sulfides, and particularly compounds of formula to inhibit LFA-1 binding to an ICAM that binds LFA-1.
1R2-5 A B R R
R
6
(I)
wherein A and B, independently, are aryl groups selected from the group consisting of and 6-membered aromatic rings, including, but not limited to, phenyl, thienyl, furyl, pyrimidinyl, imidazolyl, pyrazolyl, pyridyl, pyrazinyl, pyrrolyl, and pyridazinyl;
R
2 and independently, are selected from the group consisting of hydrogen, wherein R, is hydrogen or an alkyl group containing one to six saturated straight or branched chain carbon atoms 6 alkyl),
-O-RF,
-halo, wherein halo is Cl, F, Br, or I, -NRbRc, where Rb and Rc, independently, are H, C, 6 6 alkyl, or -CH,-aryl, 6 4.
WO 00/59878 PCT/USOO/08840
-NO,
-CN,
-perfluoroR., such as trifluoromethyl, -(CHZ)n-NRbR., wherein n is an integer 1 to 6, a 5- or 6-membered heterocyclic ring, either aliphatic or aromatic, containing one or more of O, N, or S, optionally substituted, such as morpholino, and -S-aryl, wherein aryl is a 5- or 6-membered aromatic ring, optionally substituted; and R 4
R
5 and R 6 independently, are selected from the group consisting of hydrogen,
-R
a
-R
-halo, -NRbR,
-NO
2
-CN,
-perfluoroRa,
-(CH
2 )n-NRbR, and -a 5- or 6-membered heterocyclic ring, aliphatic or aromatic, containing one or more of O, N, or S, and optionally substituted, -S-aryl, or wherein
R
4 and R 5 are taken together to form a 5- or 6-membered aromatic ring, optionally containing one or more of O, N, or S in the ring, optionally substituted.
Examples of novel negative regulators of LFA-1 binding to ICAMs, include, but are not limited to the compounds presented in Table I.
WO 00/59878 WO 0059878PCTIUSOO/08840 Table I Exemplary Negative Regulators 3-Chloro-4-(2-chlorophenylsulfanyl)-phenylamine hydrochloride 4-Nitro- '-chlorophenyl-(2',3'-dichlorophenyl)-sulfide 3-Chloro-4-(2-naphthylsulfanyl)-phenylamine hydrochloride 3-Chloro-4-(2,3-dichlorophenylsulfanyl)-phenylamine hydrochloride 3-Chloro-4-(2,4,5-trichlorophenylsulfanyl)-phenylamine hydrochloride 3-Chloro-4-(2,4-dichlorophenylsulfanyl)-phenylainine hydrochloride 4-(Benzothiazol-2-ylsulfanyl)-3-chloro-phenylamnine 3-Chloro-4-( I-chloro-naphthalen-2-ylsulfanyl)-phenylamine 3-Methoxy-4-(2,3-dichlorophenylsulfanyl)-phenylamine 5-Amino-2-(2,3-dichlorophenylsulfanyl)-acetophenone hydrochloride 4-(2,3-dichlorophenylsulfanyl)-phenylamine 3 -Chloro-4-( 1 -naphthylsulfanyl)-phenylamine hydrochloride 3-Methyl-4-(2,4-dichlorophenylsulfanyl)-phenylamine hydrochloride 1 -Acetamido-3-chloro-4-(2,3-dichlorophenylsulfanyl)-benzene 4-Methylamino-2,2',4'-trichlorodiphenylsulfide 3-Bromo-4-(2,4-dichlorophenylsulfanyl)-phenylaniine hydrochloride 3-Hydroxy-4-(2,3-dichlorophenylsulfanyl)-phenylamine hydrochloride 6-Chloro-5-(2,4-dichlorophenylsulfanyl)-l1H-benzimidazole 4-Amino-2-chlorophenyl-(2'4'-dimethylphenyl)-sulfide hydrochloride 2,5-Dichloro-4-(2,4-dichlorophenylsulfanyl)-phenylamine hydrochloride 4-Aniino-2-chlorophenyl-(2'-methyl-4'-chlorophenyl)-sulfide 4-Amino-2-chlorophenyl-(2',4'-difluorophenyl)-sulfide hydrochloride 4-Amino-2-chlorophenyl-(2',4',6'-trichlorophenyl)-sulfide 4-Amino-2-chlorophenyl-(2'-amino-4'-chlorophenyl)-sulfide 4-Amino-2-chlorophenyl-(2'-chloro-4'-nitrophenyl)-sulfide 4-Amino-2-chlorophenyl-(2'-nitro-4'-chlorophenyl)-sulfide 4-Amino-2-chlorophenyl-(3',4'-dichlorophenyl)-sulfide 4-Amino-2-chlorophenyl-2-(3-chloro-5-trifluoromethylpyridyl)-sulfide Bis-(4,4'-diamino-2,2'-dichlorophenyl)-sulfide WO 00/59878 PTU0/84 PCTIUSOO/08840 4-Amino-2-chlorophenyl-(2',4'-dichlorophenyl)-sulfide 4-Amino-2-chlorophenyl-(4'-acetamido-2'-chlorophenyl)-sulfide 4-Amino-2-chlorophenyl-6-(5-nitroquinolino)-sulfide 4-Amino-2-chlorophenyl-(4'-dimethylamino-2'-chlorophenyl)-sulfide 2-Chloro-4-ainino-5-methylamninophenyl-(2',4'-dichlorophenyl)-sulfide 2-Chloro-4-amino-5 -N-morpholinophenyl-(2',4'-dichlorophenyl)-sulfide 4-Amino-2-trifluoromethylphenyl-(2',4'-dichlorophenyl)-sulfide 4-Amino-2-chlorophenyl-2-(5-nitro-3-bromo)-pyridine sulfide 4-Aminomethyl-2-chlorophenyl-(2', 4'-dichlorophenyl)-sulfide 4,5-Dichloro-2-(2,4-dichlorophenylsulfanyl)-phenylamine 3 ,5-Dichloro-4-(2,4-dichlorophenylsulfanyl)-phenylamine 2,3-Dichloro-4-(2,4-dichlorophenylsulfanyl)-phenylamnine 4-Aniino-2-fluorophenyl-(2',4'-dichlorophenyl)-sulfide 5-Amino-3-chlorophenyl-(2',4'-dichlorophenyl)-sulfide 3-Chloro-4-( I -chloro-naphthalen-2-ylsulfanyl)-phenylamine 1 -Nitro-4-phenylsulfanyl-phenyl)-ethanone 1 -(3-Nitro-4-phenylsulfanyl-phenyl)-ethanone oxime 5-Trifluoromethyl-2-phenylsulfanyl-benzonitrile 1 -(3,5-dichlorophenyl)-3 Bis-2,4,6-Trinitrophenyl-sulfide 2-Methyl-i -(2-o-tolylsulfanyl-phenyl)- IH-pyrrole 3-[2-(4-Chloro-2-nitro-phenylsulfanyl)-phenylamnino-3H-isobenzofuiran- 1 -one 4-(Benzothiazol-2-ylsulfanyl)-3-chloro-phenylamine 2-Nitro-4-chlorophenyl-(2 'aminophenyl)-sulfide 6-Amino-2-chlorophenyl-(4'-methylphenyl)-sulfide 4-Nitrophenyl-(2 '-chlorophenyl)-sulfide 2, 4-Dinitrophenyl-(4'-chlorophenyl)-sulfide 4-Aminophenyl-(2 '-chlorophenyl)-sulfide 2, 4-Diaminophenyl-(4'-isopropylphenyl)-sulfide 4-ir--horpey 2,3 '-dichlorophenyl)-sulfide 4-Amino-2-chlorophenyl-2-( 5-nitro-3-bromo)-pyridine-sulfide WO 00/59878 PCT/USOO/08840 The compounds as represented by structural formula can be prepared by synthetic methods or by metabolic processes. Preparation of the compounds by metabolic processes include both in vivo and in vitro processes. Pharmaceutical compositions comprising compounds of the invention are also contemplated.
The invention also provides methods of inhibiting LFA-1 binding to ICAMs that bind LFA-1 comprising the step of contacting LFA-1 with a diaryl sulfide, and preferably a compound of structural formula I. Likewise, the invention provides methods of inhibiting leukocyte adhesion to endothelial cells comprising the step of contacting leukocytes expressing LFA-1 with a diaryl sulfide, and preferably a compound of the structural formula The invention also comprehends methods for treating an inflammatory disorder comprising the steps of administering to a mammal an amount of a pharmaceutical composition of the invention sufficient to inhibit binding of LFA-1 to a naturlal ligand thereof that competes with ICAM-1 or ICAM-3 for binding to LFA-1. The invention also comprehends methods for treating an inflammatory disorder arising from LFA-1 binding to a natural ligand thereof that competes with ICAM-1 or ICAM-3 for binding to LFA-1, comprising administering to a mammal in need thereof a compound that competes with 3-chloro-4-(1-chloronaphthalen-2-ylsulfanyl)-phenylamine for binding to LFA-1 in an amount sufficient to inhibit binding of the natural ligand to LFA-1. In addition, the invention provides methods of ameliorating a pathological condition associated with LFA-1 binding to an ICAM that binds LFA-1 comprising administering to an individual in need thereof an effective amount of a diaryl sulfide, and preferably a compound of the structural formula to inhibit LFA-1 binding to the ICAM.
Examples of inhibitors of the present invention include, but are not limited to, the compounds set out in Table I.
The invention also provides for use of a compound of the invention in the production of a medicament for the treatment of pathologies associated with LFA- 1 binding to ICAM-1.
The invention also provides methods to identify a negative regulator of LFA-1 binding to a natural ligand thereof that competes with ICAM-1 or ICAM-3 for binding to LFA-1 comprising the steps of: a) contacting LFA-1 with an activator of 4 t WO 00/59878 PCT/USOO/08840 LFA-1 binding; b) measuring LFA-1 binding with the natural ligand in the presence and absence of a test compound; and c) identifying the test compound as an inhibitor when decreased LFA-1 binding to the ligand is detected in the presence of the test compound. In one aspect, the activator is crystal violet.
DETAILED DESCRIPTION OF THE INVENTION An ICs 0 value for a compound is defined as the concentration of the compound required to produce 50% inhibition of a biological activity of interest. As used herein, a negative regulator is defined as a compound characterized by an ICo 0 for inhibition of LFA-1 binding to a natural ligand. Negative regulators of LFA-1 binding are defined to have an IC 5 s of less than about 200 gM, less than about 100 tM, less than about 50 rM, and preferably from about 0.05 M to 40 gM.
The term "pharmaceutically acceptable carrier" as used herein refers to those prodrugs of compounds of the invention which are suitable for use in contact with recipient animals and having undue toxicity, irritation, allergic response commensurate with a reasonable benefit/risk ratio, and effective for their intended use.
The term "prodrug" as used herein refers to compounds which are rapidly transformed in vivo to the parent compound of the above formula, for example, by hydrolysis. A thorough discussion is provided in Higuchi, et al., Prodrugs as Novel Delivery Systems, vol. 14 of the A.C.S.D. Symposium Series, and in Roche Bioreversible Carriers in Drug Design, American Pharmaceutical Association and Pergamon Press, 1987, both of which are incorporated herein by reference. Prodrug design is discussed generally in Hardma, et al., (Eds), Goodman Gilman's The Pharmacological Basis of Therapeutics, Ninth Edition, New York, New York (1996), pp. 11-16. Briefly, administration of a drug is followed by elimination from the body or some biotransformation whereby biological activity of the drug is reduced or eliminated. Alternatively, a biotransformation process may lead to a metabolic by-product which is itself more active or equally active as compared to the drug initially administered. Increased understanding of these biotransformation processes permits the design of so-called "prodrugs" which, 4 C- WO 00/59878 PCT/USOO/08840 following a biotransformation, become more physiologically active in an altered state.
Prodrugs are therefore pharmacologically inactive compounds which are converted to biologically active metabolites. In some forms, prodrugs are rendered pharmacologically active through hydrolysis of, for example, and ester or amide linkage, often times introducing or exposing a functional group on the prodrug. The thus modified drug may also react with an endogenous compound to form a water soluble conjugate which further increases pharmacological properties of the compound, for example, as a result of increased circulatory half-life.
As another alternative, prodrugs can be designed to undergo covalent modification on a functional group with, for example, glucuronic acid, sulfate, glutathione, amino acids, or acetate. The resulting conjugate may be inactivated and excreted in the urine, or rendered more potent than the parent compound. High molecular weight conjugates may also be excreted into the bile, subjected to enzymatic cleavage, and released back into circulation, thereby effectively increasing the biological half life of the originally administered compound.
Compounds of the invention may exist as stereoisomers where asymmetric or chiral centers are present. Stereoisomers are designated by either "S" or depending on arrangement of substituents around a chiral carbon atom.
Mixtures of stereoisomers are contemplated by the invention. Stereoisomers include enantiomers, diastereomers, and mixtures of the two. Individual stereoisomers of compounds of the invention can be prepared synthetically from commercially available starting materials which contain asymmetric or chiral centers or by preparation ofracemic mixtures followed by separation or resolution techniques well known in the art. Methods of resolution include attachment of a mixture of enantiomers to a chiral auxiliary, separation of the resulting mixture by recrystallization or chromatography, and liberation of the optically pure product from the auxiliary; salt formation employing an optically active resolving agent, and (3) direct separation of the mixture of optical enantiomers on chiral chromatographic columns.
Compounds of the present invention include, but are not limited to those embraced by general structural formula above and the compounds set out in WO 00/59878 PCT/US00/08840 Table I.
The invention also provides pharmaceutical compositions comprising one or more compounds of the invention, preferably further comprising a pharmaceutically acceptable carrier or diluent.
The invention further provides methods for inhibiting LFA-1 binding to an ICAM that binds LFA-1 comprising the step of contacting LFA-1, or an ICAMbinding fragment thereof, with a negative regulator compound; said negative regulator binding the LFA-1 L polypeptide, or a fragment thereof, at a site selected from the group consisting of a conformation that binds a diaryl sulfide or a binding site defined by Ile 25 9 Leu 2 98 Ile 2 3 5 Val' 5 7 Leu' 61 and Ile 3 06 of human LFA-1 a, polypeptide and an LFA- domain that binds 3-chloro-4-(1 -chloro-naphthalen-2-ylsulfanyl)-phenylamine having the structure described above. Alternatively, the negative regulator binding site on LFA-1 is defined by amino acid residues Ile 2 59 Leu 2 98 le 235 Val1 57 Leu 61 Ile 3 06 Leu 30 ,Tyr 2 57 Leu 132 Val 233 Val 130 and Tyr 166 In still another alternative, the negative regulator binding site on LFA-1 is defined by amino acid residues Lys 2 87 Leu 29 Ile 259 Leu 302 Ile 235 Val' 57 Tyr 257 Lys 3 0 5 Leu' 61 Leu' 32 Va1 233 Ile 255 Val 130 Tyr 1 66 Ile 3 0 6 Phe 1 34 Phe 1 68 Phe 53 Tyr 3 07 Va 308 Ile 309 Thr 23 Glu 284 Phe 285 Glu 30 1 Met' 54 Ie 237 Ile 1 5 and Leu 295 The LFA-1 regulatory binding site is described in copending U.S. patent application entitled "LFA-1 Regulatory Binding Site and Uses Thereof", filed April 2, 1999, attorney docket number 27866/35375, Serial Number 09/285,477, incorporated herein by reference in its entirety. In one embodiment, methods of the invention include use of cells expressing either LFA-1 or the ICAM.
In methods wherein one of the binding partners is expressed in a cell, the other binding partner is either purified and isolated, in a fluid sample (purified, partially purified, or crude) taken from an individual, or in a cell lysate. The invention also comprehends methods wherein both LFA-1 and the ICAM are expressed in cells. The LFA-1 and ICAM binding partners may be expressed on the same cell type or different cell types. Preferably, the LFA-1 polypeptide is expressed on leukocytes, i.e.
lymphocytes, monocytes, or granulocytes, and the ICAM polypeptide is expressed on endothelial cells.
The invention also provides methods to inhibit leukocyte adhesion to WO 00/59878 PCT/USOO/08840 endothelial cells comprising the step of contacting said leukocyte with a negative regulator of LFA-1 binding to an ICAM that binds LFA-1, said negative regulator binding an LFA-1 regulatory site selected from the group consisting of a conformation that binds a diaryl sulfide or a binding site defined by Ile 2 59 Leu 2 98 Ile 2 35 Val 57 Leu" 1 6 and Ile 3 06 of human LFA-1 ac polypeptide or an LFA-1 domain that binds 3chloro-4-(1-chloro-naphthalen-2-ylsulfanyl)-phenylamine. Alternatively, the diaryl sulfide binding conformation is defined by amino acid residues as described above. In vivo and in vitro methods are contemplated.
The invention also provides methods to ameliorate a pathology arising from LFA-1 binding to an ICAM comprising the step of administering to an individual in need thereof a negative regulator of LFA-1 binding to the ICAM in an amount effective to inhibit LFA-1 binding to the ICAM, said negative regulator binding to an LFA-1 regulatory site selected from the group consisting of a conformation that binds a diaryl sulfide or a site defined by Ile 25 9 Leu 29 8 Ile 2 35 Val' 7 Leu 6 and Ile 3 06 of human LFA-1 or an LFA-1 domain that binds compound 3chloro-4-( 1 -chloro-naphthalen-2-ylsulfanyl)-phenylamine.
In a preferred embodiment, methods of the invention include use of a diaryl sulfide compound to inhibit binding of LFA-1 to an ICAM. A preferred method includes use of a compound of general structural formula a pharmaceutically acceptable salt, or prodrug thereof as described above.
Therapeutic Methods To the extent that leukocyte adhesion to endothelial cells gives rise to a pathological disorder, the invention provides methods to ameliorate pathologies associated with accumulation of leukocytes resulting from LFA-1 binding to an ICAM that binds LFA-1 comprising the step of administering to an individual in need thereof an amount of an inhibitor of LFA-1 binding to the ICAM effective to inhibit LFA-1 binding to the ICAM, said inhibitor binding to LFA-1 at a site presented by amino acid residues Ile 2 59 Leu 2 98 Ile 2 35 Val' 5 7 Leu 1 61 and Ile 3 06 Exemplary medical conditions include, without limitation, inflammatory diseases, autoimmune diseases, reperfusion injury, myocardial infarction, stroke, hemorrhagic shock, organ transplant, M O WO 00/59878 PCT/US00/08840 and the like. Methods of the invention provide for amelioration of a variety of pathologies, including, for example, but not limited to adult respiratory distress syndrome, multiple organ injury syndrome secondary to septicemia, multiple organ injury secondary to trauma; reperfusion injury of tissue, acute glomerulonephritis, reactive arthritis, dermatosis with acute inflammatory components, stroke, thermal injury, Crohn's disease; necrotizing enterocolitis, granulocyte transfusion associated syndrome, and cytokine induced toxicity, and T cell mediated diseases.
Inflammatory cell activation and excessive or unregulated cytokine TNFa and IL-1 production are also implicated in disorders such as rheumatoid arthritis, osteoarthritis, gouty arthritis, spondylitis, thyroid associated ophthalmopathy, Behcet disease, sepsis, septic shock, endotoxic shock, gram negative sepsis, gram positive sepsis, toxic shock syndrome, asthma, chronic bronchitis, allergic respiratory distress syndrome, chronic pulmonary inflammatory disease, such as chronic obstructive pulmonary disease, silicosis, pulmonary sarcoidosis, reperfusion injury of the myocardium, brain, and extremities, fibrosis, cystic fibrosis, keloid formation, scar formation, atherosclerosis, transplant rejection disorders, such as graft vs. host reaction and allograft rejection, chronic glamerulonephritis, lupus, inflammatory bowel disease, such as ulcerative colitis, proliferative lymphocyte diseases, such as leukemia, and inflammatory dermatoses, such as atopic dermatitis, psoriasis, urticaria, uveitis.
Other conditions characterized by elevated cytokine levels include brain injury due to moderate trauma (see J. Neurotrauma, 12, pp. 1035-1043 (1995); J. Clin. Invest., 91, pp. 1421-1428 (1993)), cardiomyopathies, such as congestive heart failure (see Circulation, 97, pp. 1340-1341 (1998)), cachexia, cachexia secondary to infection or malignancy, cachexia secondary to acquired immune deficiency syndrome (AIDS), ARC (AIDS related complex), fever myalgias due to infection, cerebral malaria, osteoporosis and bone resorption diseases, keloid formation, scar tissue formation, and pyrexia.
The ability of the negative regulators of the invention to treat arthritis can be demonstrated in a murine collagen-induced arthritis model [Kakimoto, et al.
Immunol. 142:326-337 (1992)], in a rat collagen-induced arthritis model [Knoerzer, et WO 00/59878 PCT/USOO/08840 al., Toxical Pathol. 25:13-19 (1997)], in a rat adjuvant arthritis model [Halloran, et al., Arthritis Rheum 39:810-819 (1996)], in a rat streptococcal cell wall-induced arthritis model [Schimmer, et al., J. Immunol. 160:1466-1477 (1998)], or in a SCIDmouse human rheumatoid arthritis model [Oppenheimer-Marks, et al., J. Clin. Invest 101:1261-1272 (1998)].
The ability of the negative regulators to treat Lyme arthritis can be demonstrated according to the method of Gross, et al., Science, 218:703-706, (1998).
The ability of the negative regulators to treat asthma can be demonstrated in a murine allergic asthma model according to the method of Wegner, et al., Science, 247:456-459, (1990), or in a murine non-allergic asthma model according to the method of Bloemen, et al., Am. J. Respir. Crit. Care Med. 153:521- 529 (1996).
The ability of the negative regulators to treat inflammatory lung injury can be demonstrated in a murine oxygen-induced lung injury model according to the method of Wegner, et al., Lung, 170:267-279, (1992), in a murine immune complexinduced lung injury model according to the method of Mulligan, et al., J. Immunol., 154:1350-1363, (1995), or in a murine acid-induced lung injury model according to the method of Nagase, et al., Am. J. Respir. Crit. Care Med., 154:504-510, (1996).
The ability of the negative regulators to treat inflammatory bowel disease can be demonstrated in a murine chemical-induced colitis model according to the method of Bennett, et al., J. Pharmacol. Exp. Ther., 280:988-1000, (1997).
The ability of the negative regulators to treat autoimmune diabetes can be demonstrated in an NOD mouse model according to the method of Hasagawa, et al., Int. Immunol. 6:831-838 (1994), or in a murine streptozotocin-induced diabetes model according to the method of Herrold, et al., Cell Immunol. 157:489-500, (1994).
The ability of the negative regulators to treat inflammatory liver injury can be demonstrated in a murine liver injury model according to the method of Tanaka, et al., J. Immunol., 151:5088-5095, (1993).
The ability of the negative regulators to treat inflammatory glomerular injury can be demonstrated in a rat nephrotoxic serum nephritis model according to the method of Kawasaki, et al., J. Immunol., 150:1074-1083 (1993).
WO 00/59878 PCT/US00/08840 The ability of the negative regulators to treat radiation-induced enteritis can be demonstrated in a rat abdominal irradiation model according to the method of Panes, et al., Gastroenterology, 108:1761-1769 (1995).
The ability of the negative regulators to treat radiation pneumonitis can be demonstrated in a murine pulmonary irradiation model according to the method of Hallahan, et al., Proc. Natl. Acad. Sci (USA), 94:6432-6437 (1997).
The ability of the negative regulators to treat reperfusion injury can be demonstrated in the isolated heart according to the method of Tamiya, et al., Immunopharmacology, 29:53-63 (1995), or in the anesthetized dog according to the model of Hartman, et al., Cardiovasc. Res. 30:47-54 (1995).
The ability of the negative regulators to treat pulmonary reperfusion injury can be demonstrated in a rat lung allograft reperfusion injury model according to the method of DeMeester, et al., Transplantation, 62:1477-1485 (1996), or in a rabbit pulmonary edema model according to the method of Horgan, et al., Am. J.
Physiol. 261:H1578-H1584 (1991).
The ability of the negative regulators to treat stroke can be demonstrated in a rabbit cerebral embolism stroke model according to the method of Bowes, et al., Exp. Neurol., 119:215-219 (1993), in a rat middle cerebral artery ischemia-reperfusion model according to the method of Chopp, et al., Stroke, 25:869- 875 (1994), or in a rabbit reversible spinal cord ischemia model according to the method of Clark et al., Neurosurg., 75:623-627 (1991). The ability of the negative regulators to treat cerebral vasospasm can be demonstrated in a rat experimental vasospasm model according to the method of Oshiro, et al., Stroke, 28:2031-2038 (1997).
The ability of the negative regulators to treat peripheral artery occlusion can be demonstrated in a rat skeletal muscle ischemia/reperfusion model according to the method of Gute, et al., Mol. Cell Biochem., 179:169-187 (1998).
The ability of the negative regulators to treat graft rejection can be demonstrated in a murine cardiac allograft rejection model according to the method of Isobe, et al., Science, 255:1125-1127 (1992), in a murine thyroid gland kidney capsule model according to the method of Talento, et al., Transplantation, 55:418-422 (1993), WO 00/59878 PCT/USOO/08840 in a cynomolgus monkey renal allograft model according to the method of Cosimi, et al., J. Immunol., 144:4604-4612 (1990), in a rat nerve allograft model according to the method of Nakao, et al., Muscle Nerne, 18:93-102 (1995), in a murine skin allograft model according to the method of Gorczynski and Wojcik, J. Inmmunol. 152:2011- 2019, (1994), in a murine corneal allograft model according to the method of He, et al., Opthalmol. Vis. Sci., 35:3218-3225 (1994), or in a xenogeneic pancreatic islet cell transplantation model according to the method of Zeng, et al., Transplantation, 58:681-689 (1994).
The ability of the negative regulators to treat graft-vs.-host disease (GVHD) can be demonstrated in a murine lethal GVHD model according to the method of Harning, et al., Transplantation, 52:842-845 (1991).
The ability of the negative regulators to treat cancers can be demonstrated in a human lymphoma metastasis model (in mice) according to the method of Aoudjit, et al., J. Immunol., 161:2333-2338, (1998).
Pharmaceutical Compositions The present invention also provides pharmaceutical compositions which comprise a diaryl sulfide formulated together with one or more pharmaceutically-acceptable carriers.
The pharmaceutical compositions of the invention can be administered to humans and other animals by any suitable route. For example, the compositions can be administered orally, rectally, parenterally, intracistemally, intravaginally, intraperitoneally, topically (as by powders, ointments, or drops), bucally, or nasally.
The term "parenteral" administration as used herein refers to modes of administration which include intravenous, intraarterial, intramuscular, intraperitoneal, intrastemal, intrathecal, subcutaneous and intraarticular injection and infusion.
Pharmaceutical compositions of this invention for parenteral injection comprise pharmaceutically-acceptable sterile aqueous or nonaqueous solutions, dispersions, suspensions or emulsions as well as sterile powders for reconstitution into sterile injectable solutions or dispersions just prior to use. Examples of suitable aqueous and nonaqueous carriers, diluents, solvents or vehicles include water, WO 00/59878 PCT/US00/08840 ethanol, polyols (such as glycerol, propylene, glycol, polyethylene glycol, and the like), and suitable mixtures thereof, vegetable oils (such as olive oils), and injectable organic esters such as ethyl oleate. Proper fluidity can be maintained, for example, by the use of coating materials such as lecithin, by the maintenance of the required particle size in the case of dispersions, and by the use of surfactants.
These compositions may also contain adjuvants such as preservative, wetting agents, emulsifying agents, and dispersing agents. Prevention of the action of microorganisms may be ensured by the inclusion of various antibacterial and antifungal agents, for example, paraben, chlorobutanol, phenol sorbic acid, and the like. It may also be desirable to include isotonic agents such as sugars, sodium chloride, and the like, Prolonged absorption of the injectable pharmaceutical form may be brought about by the inclusion of agents which delay absorption such as aluminum monosterate and gelatin.
In some cases, in order to prolong the effect of the drug, it is desirable to slow the absorption of the drug from subcutaneous or intramuscular injection. This may be accomplished by the use of a liquid suspension of crystalline or amorphous materials with poor water solubility. The rate of absorption of the drug then depends upon its rate of dissolution which, in turn, may depend upon crystal size and crystalline form. Alternatively, delayed absorption of a parenterally administered drug from is accomplished by dissolving or suspending the drug in an oil vehicle.
Injectable depot forms are made by forming microencapsule matrices of the drug in biodegradable polymers such a polylactide-polyglycolide. Depending upon the ratio of drug to polymer and the nature of the particular polymer employed, the rate of drug release can be controlled. Examples of other biodegradable polymers include poly(orthoesters) and poly(anhydrides). Depot injectable formulations are also prepared by entrapping the drug in liposomes or microemulsions which are compatible with body tissue.
The injectable formulations can be sterilized, for example, by filtration through a bacterial- or viral-retaining filter, or by incorporating sterilizing agents in the form of sterile solid compositions which can be dissolved or dispersed in sterile water or other sterile injectable medium just prior to use.
WO 00/59878 PCT/USOO/08840 Solid dosage forms for oral administration include capsules, tablets, pills, powders, and granules. In such solid dosage forms, the active compound is mixed with a least one inert, pharmaceutically-acceptable excipient or carrier such as sodium citrate or dicalcium phosphate and/or fillers or extenders such as starches, lactose, sucrose, glucose, mannitol, and silicic acid, binders such as, for example, carboxymethylcellulose, gums (e.g.alginates, acacia) gelatin, polyvinylpyrrolidone, and sucrose, humectants such as glycerol, disintegrating agents such as agaragar, calcium carbonate, potato or tapioca starch, alginic acid, certain silicates, and sodium carbonate, solution retarding agents such a paraffin, absorption accelerators such as quaternary ammonium compounds, wetting agents such as, for example, cetyl alcohol and glycerol monosterate, absorbents such as kaolin and bentonite clay, and lubricants such as talc, calcium sterate, magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate, and mixtures thereof. In the case of capsules, tablets and pills, the dosage form may also comprise buffering agents.
Solid compositions of a similar type may also be employed as fillers in soft and hard-filled gelatin capsules using such excipients as lactose or milk sugar as well as high molecular weight polyethylene glycols and the like.
The solid dosage forms of tablets, dragees, capsules, pills, and granules can be prepared with coatings and shells such as enteric coatings and other coatings well known in the pharmaceutical formulating art. They may optionally contain opacifying agents and can also be of a composition that they release the active ingredients(s) only, or preferentially, in a part of the intestinal tract, optionally, in a delayed manner. Exemplary materials include polymers having pH sensitive solubility, such as the materials available as Eudragit Examples of embedding compositions which can be used include polymeric substances and waxes.
The active compounds can also be in micro-encapsulated form if appropriate, with one or more of the above-mentioned excipients.
Liquid dosage forms for oral administration include pharmaceuticallyacceptable emulsions, solutions, suspensions, syrups and elixirs. In addition to the active compounds, the liquid dosage forms may contain inert diluents commonly used in the art such as, for example, water or other solvents, solubilizing agents and WO 00/59878 PCT/US00/08840 emulsifiers such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butylene glycol, dimethyl formamide, oils (in particular, cottonseed, groundnut, corn, germ, olive, castor, and sesame oils), glycerol, tetrahydrofurfuryl alcohol, polyethylene glycols and fatty acid esters of sorbitan, and mixtures thereof.
Besides inert diluents, the oral compositions can also include adjuvants such as wetting agents, emulsifying and suspending agents, sweetening, flavoring, and perfuming agents.
Suspensions, in addition to the active compounds, may contain suspending agents as, for example, ethoxylated isostearyl alcohols, polyoxyethylene sorbitol and sorbitan esters, microcrystalline cellulose, aluminum metahydroxide, bentonite, agar-agar, and tragacanth, and mixtures thereof.
Compositions for rectal or vaginal administration are preferably suppositories which can be prepared by mixing the compounds of this invention with suitable non-irritating excipients or carriers such as cocoa butter, polyethylene glycol or suppository wax which are solid at room temperature but liquid at body temperature and therefore melt in the rectum or vaginal cavity and release the active compound.
Compounds of the present invention can also be administered in the form of liposomes. As is known in the art, liposomes are generally derived from phospholipids or other lipid substances. Liposomes are formed by mono- or multilamellar hydrated liquid crystals that are dispersed in an aqueous medium. Any nontoxic, physiologically-acceptable and metabolizable lipid capable of forming liposomes can be used. The present compositions in liposome form can contain, in addition to a compound of the present invention, stabilizers, preservatives, excipients, and the like. The preferred lipids are the phospholipids and the phosphatidyl cholines (lecithins), both natural and synthetic. Methods to form liposomes are known in the art. See, for example, Prescott, Ed., Methods in Cell Biology, Volume XIV, Academic Press, New York, N.Y. (1976), p. 33 et seq.
The compounds of the present invention may be used in the form of pharmaceutically-acceptable salts derived from inorganic or organic acids. By WO 00/59878 PCT[USOO/8840 "pharmaceutically-acceptable salt" is meant those salts which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of humans and lower animals without undue toxicity, irritation, allergic response and the like, and are commensurate with a reasonable benefit/risk ratio. Pharmaceutically-acceptable salts are well known in the art. For example, S. M. Berge, et al., describe pharmaceutically-acceptable salts in detail in J. Pharmaceutical Sciences, 66:1 (1977). The salts may be prepared in situ during the final isolation and purification of the compounds of the invention or separately by reacting a free base function with a suitable acid. Representative acid addition salts include, but are not limited to acetate, adipate, alginate, citrate, aspartate, benzoate, benzenesulfonate, bisulfate, butyrate, camphorate, camphorolsulfonate, digluconate, glycerophosphate, hemisulfate, heptanoate, hexanoate, fumarate hydrochloride, hydrobromide, hydroiodide, 2hydroxyethanesulfonate (isothionate), lactate, maleate, methanesulfonate, nicotinate, 2-naphthalenesulfonate, oxalate, pamoate, pectinate, persulfate, 3-phenylpropionate, picrate, pivalate, propionate, succinate, tartrate, thiocyanate, phosphate, glutamate, bicarbonate, p-toluenesulfonate and undecanoate. Examples of acids which may be employed to form pharmaceutically acceptable acid addition salts include inorganic acids as hydrochloric acid, hydrobromic acid, sulphuric acid and phosphoric acid and such organic acids as oxalic acid, maleic acid, succinic acid and citric acid.
Basic nitrogen-containing groups can be quatemized with such agents as lower alkyl halides such as methyl, ethyl, propyl, and butyl chlorides, bromides and iodides; dialky sulfates like dimethyl, diethyl, dibutyl and diamyl sulfates; long chain halides such as decyl, lauryl, myristyl and stearyl chlorides, bromides and iodides; arylalkyl halides like benzyl and phenethyl bromides and others. Water or oil-soluble or dispersible products are thereby obtained.
Basic addition salts can be prepared in situ during the final isolation and purification of compounds of this invention by reacting a carboxylic acidcontaining moiety with a suitable base such as the hydroxide, carbonate or bicarbonate of a pharmaceutically acceptable metal cation or with ammonia or organic primary, secondary or tertiary amine. Pharmaceutically-acceptable basic addition salts include, but are not limited to, cations based on alkali metals or alkaline earth metals such as WO 00/59878 PCT/USOO/08840 lithium, sodium, potassium, calcium, magnesium and aluminum salts and the like and nontoxic quaternary ammonia and amine cations including ammonium, tetramethylammonium, tetraethylammonium, methylamine, dimethylamine, trimethylamine, triethylamine, diethylamine, ethylamine and the like. Other representative organic amines useful for the formation of base addition salts include ethylenediamine, ethanolamine, diethanolamine, piperidine, piperazine and the like.
Dosage forms for topical administration of a compound of this invention include powders, sprays, ointments and inhalants. The active compound is mixed under sterile conditions with a pharmaceutically-acceptable carrier and any needed preservatives, buffers, or propellants which may be required. Ophthalmic formulations, eye ointments, powders, and solutions are also contemplated as being within the scope of this invention.
Actual dosage levels of active ingredients in the pharmaceutical compositions of this invention may be varied so as to obtain an amount of the active compound(s) that is effective to achieve the desired therapeutic response for a particular patient, compositions, and mode of administration. The selected dosage level will depend upon the activity of the particular compound, the route of administration, the severity of the condition being treated, and the condition and prior medical history of the patient being treated. However, it is within the skill of the art to start doses of the compound at levels lower than required for to achieve the desired therapeutic effort and to gradually increase the dosage until the desired effect is achieved.
Generally dosage levels of about 0.1 to about 1000 mg, about 0.5 to about 500 mg, about 1 to about 250 mg, about 1.5 to about 100, and preferably of about 5 to about 20 mg of active compound per kilogram of body weight per day are administered orally or intravenously to a mammalian patent. If desired, the effective daily dose may be divided into multiple doses for purposes of administration, two to four separate doses per day.
The present invention is illustrated by the following examples.
Example 1 described a high throughput assay to screen for negative regulators of WO 00/59878 PCT/USOO/08840 LFA-1 binding to an ICAM that binds LFA-1. Example 2 relates to binding assays to evaluate the ability of various compounds to inhibit LFA-1 binding to an ICAM.
Example 3 describe synthesis of negative regulators. Example 4 provides results from cell based assays using the negative regulators.
Example 1 High Throughput Screening for LFA-1/ICAM-1 Binding Inhibitors In an effort to identify inhibitors of LFA-1/ICAM-1 binding, a high throughput screening (HTS) assay was designed to efficiently screen large numbers of chemical compounds in a proprietary library as follows.
Preliminary experiments were carried out in order to define the linear range of LFA-1/ICAM-1 interaction. Recombinant ICAM-1/IgGl fusion protein (comprising full length ICAM-1) was prepared as described in U.S. Patent Nos.
5,770,686, 5,837,478, and 5,869,262, each of which is incorporated herein by reference. The fusion protein was biotinylated using a kit obtained from Pierce Chemical (Rockford, IL). Biotinylated protein (BiolgICAM-1) concentration was determined by measuring absorbance at 280 nm, and serial dilutions were prepared to give a final concentration range of 50 gg/ml to 0.008 ig/ml. Titration of BioIgICAM-1 was carried out with the protein first aliquoted into wells on an assay plate. Recombinant LFA-1 was added to each well at the same concentration and the experiment (as described below) was carried to completion. The amount of binding was determined for each well, and from a subsequent plot of the results, a single concentration of BioIgICAM-1 was selected for subsequent experiments. In a similar manner, LFA-1 was titrated using the BioIglCAM-1 concentration selected as described above.
On day 1 of the HTS procedure, the capture antibody, a non-blocking anti-LFA-1 monoclonal antibody (TS2/4.1; ATCC #HB244), was diluted in plate coating buffer (50 mM sodium carbonate/bicarbonate, 0.05% ProClin 300, pH 9.6) to a final concentration of 2 .g/ml. Immulon 0 4 (Dynex Technologies, Chantilly, VA) plate wells were coated with 100 .l diluted antibody solution per well, and incubation was carried out overnight at 4 0 C. On day 2, the WO 00/59878 PCT/US00/08840 plates were warmed to room temperature and washed two times with wash buffer (calcium- and magnesium-free phosphate buffered saline, CMF-PBS) with 0.05% To each well, 200 pl of blocking solution fish skin gelatin in CMF-PBS with 0.05% ProClin 300) was added, and the blocking incubation was carried out at room temperature for 30 min. The blocking solution was removed by aspiration, and the plates were not washed. LFA-1 was diluted to a final concentration of 1 ug/ml in assay buffer fish skin gelatin and 2 mM MgCl1 in CMF-PBS), and 100 ul was added to each well. Incubation was carried out for one hour, and the plates were washed two times with wash buffer.
A 2X stock solution of BiolgICAM-1 was prepared containing 0.1 tg/ml BioIgICAM-1 and 4 pM crystal violet (found to be an activator of LFA-1/ICAM-1 binding) in Assay Buffer (EG&G Wallac, Gaithersburg, MD).
Aliquots (50 ul) of pooled chemicals (22 compounds/pool in 100% DMSO) from the chemical library were added to the wells, followed by addition of 50 pl of the 2X stock of BioIgICAM-1 to provide a final assay volume of 100 pl (containing 2% DMSO). The plates were incubated for one hour at room temperature, and washed once with wash buffer. Europium-labeled streptavidin (Eu-SA; #1244-360, EG&G Wallac) was diluted 1:500 in Assay Buffer, 100 ul of the diluted Eu-SA was added to each well, and the plates were incubated at room temperature for one hour.
Plates were washed eight times with wash buffer, 100 tl of DELFIA enhancement solution (EG&G Wallac) diluted 1:2, was added to each well, and the plates were shaken for five minutes using a Wallac shaker at fast speed. Plates were read using a Wallac DELFIA® fluorescence reader (fluorimeter). Controls included both positive and negative wells and 50% binding wells established using blocking antibodies, anti-LFA-1 monoclonal antibody (TS1/22.1, ATCC #HB202) or anti-ICAM-1 monoclonal antibody. Chemical pools in wells showing 50% or greater inhibition of LFA-1 binding to ICAM-1 were identified and the experiment was repeated using individual chemicals from those pools. Inhibitors of LFA-1/ICAM-1 binding were identified, and a further screen was performed to determine dose dependence of the inhibitory activity. Further study of selected compounds was carried out using biochemical and cellular assay techniques.
WO 00/59878 WO 0059878PCTIUSOO/08840 The compounds were grouped according to common structural features, and it was found that a subset (listed below) of the compounds included a characteristic diaryl sulfide structure.
3-Chloro-4-( I -chloro-naphthalen-2-ylsulfanyl)-phenylamine 1 -(3-Nitro-4-phenylsulfanyl-phenyl)-ethanone 1 -(3-Nitro-4-phenylsulfanyl-phenyl)-ethanone oxime -Trifluoromethyl-2-phenylsulfanyl-benzonitrile 1 -dichlorophenyl)-3-phenylsulfanyl-pyrrolidine-2,5 -dione Bis-2,4,6-Trinitrophenyl-sulfide 2-Methyl-i -(2-o-tolylsulfanyl-phenyl)- 1H-pyrrole 3-[2-(4-Chloro-2-nitro-phenylsulfanyl)-phenylamino-3H-isobenzofuran- I -one 4-(Benzothiazol-2-ylsulfanyl)-3-chloro-phenylamine 2-Nitro-4-chlorophenyl-(2 'aminophenyl)-sulfide 6-Ainino-2-chlorophenyl-(4'-methylphenyl)-sulfide 4-Nitrophenyl-(2 '-chlorophenyl)-sulfide 2, 4-Dinitrophenyl-(4 '-chlorophenyl)-sulfide 4-Aminophenyl-(2 '-chlorophenyl)-sulfide 2, 4-Diaminophenyl-(4'-isopropylphenyl)-sulfide In an effort to optimize the negative regulatory capacity of the identified compounds, various diaryl sulfide derivatives were conducted as described below in Example 3. Additional diaryl sulfide derivatives are described in co-pending provisional patent application entitled "Cell Adhesion-Inhibiting Antiinflammatory and Immune Suppressive Compounds" filed April 2, 1999, attorney docket number 6446.US.Z3, Serial Number 09/2 86,645, incorporated herein by reference in its entirety.
WO 00/59878 PCT/US00/08840 Example 2 Binding Assays A. ICAM-I/LFA-I Biochemical Interaction Assay Compounds that antagonize the interaction between ICAM-1 and LFA-1 can be identified, and their activities quantitated, using both biochemical and cell-based assays. A primary biochemical assay measures the ability of the compound in question to block the interaction between LFA-1 and its adhesion partner ICAM-1, as described below.
In this biochemical assay, 100 pl of anti-LFA-1 antibody at a connection of 5 ug/ml in Dulbecco's phosphate-buffered saline (D-PBS) was used to coat wells of a 96-well microtiter plate overnight at 4°C. The wells were washed twice with wash buffer (CMF-PBS, 0.05% Tween 20) and blocked by addition of 200 pl of D-PBS containing 5% fish skin gelatin. Recombinant LFA-1 (100 gl of 0.7 g/ml) in D-PBS was added to each well. Incubation was continued for one hour at room temperature and the wells were washed twice with wash buffer. Serial dilutions of compounds being assayed as LFA-1/ICAM-1 negative regulators, prepared as mM stock solutions in DMSO, were diluted in D-PBS, 2 mM MgCl 2 1% fish skin gelatin and 500 il of each dilution added to each well, followed by addition of 50 gl of 0.8 jg/ml BioIgICAM-1 to the wells, and the plates were incubated at room temperature for one hour. The wells were then washed twice with wash buffer and 100 tl of Eu-SA (EG&G Wallac) diluted 1:100 in Delfia® assay buffer (EG&G Wallac) were added to the wells. Incubation was carried out for one hour at room temperature. The wells were washed eight times with wash buffer and 100 pl of enhancement solution (EG&G Wallac) was added to each well. Incubation was continued for five minutes with constant mixing. Time-resolved fluorimetry measurements were made using the Victor 1420 Multilabel Counter (EG&G Wallac) and the percent inhibition of each candidate compound was calculated using the following equation: %inhibition 100 average OD w/compound minus background Saverage OD w/o compound minus background] WO 00/59878 PCT/US00/08840 where "background" refers to wells that were not coated with anti-LFA-1 antibody.
B. ICAM-I/JY-8 Cell Adhesion Assay Biologically relevant activity of the compounds in this invention was confirmed using a cell-based adhesion assay that measures the ability of the compounds to block adherence of JY-8 cells (a human EBV-transformed B cell line expressing LFA-I on its surface) to immobilized ICAM-1, as follows. This assay may be performed with or without added IL-8. For IL8 stimulation of the standard JY-8 cells 30 ng/ml IL-8 was added in the 30 minute incubation at 37 0 C with the cells.
For measurement of inhibitory activity in the cell-based adhesion assay, 96-well microtiter plates were coated with 70 pl of recombinant ICAM-1/Ig at a concentration of 5 pg/ml in CMF-PBS overnight at 4 0 C. The wells were washed twice with D-PBS and blocked by addition of 200 pI of D-PBS, 5% fish skin gelatin by incubation for one hour at room temperature. Fluorescent-tagged JY-8 cells (50 pI at 2 x 106 cells/ml in RPMI-1640/1% fetal bovine serum (FBS)) were added to the wells. For fluorescent labeling of JY-8 cells, 5 x 106 cells, washed once in RPMI 1640, were resuspended in 1 ml RPMI-1640 containing 2 pM Calcein AM (Molecular Probes, OR), were incubated at 37 0 C for 30 minutes and washed once with RPMI-1640/1% FBS. Dilutions of compounds to be assayed for LFA-1/ICAM-1 antagonistic activity were prepared in RPMI-1640/1% FBS from 10 mM stock solutions in DMSO and 50 il aliquots were added to duplicate wells. Microtiter plates were incubated for 45 min at room temperature and the wells were washed gently once with RPMI-1640/1% FBS. Fluorescence intensity was measured in a fluorescence plate reader with an excitation wavelength at 495 nm and an emission wavelength at 530 nm. The percent inhibition of a candidate compound at a given concentration was calculated using the following equation: %inhibition 100 X 1 average OD w/compound I average OD w/o compound WO 00/59878 PCT/US00/08840 C. ICAM-3/JY-8 Cell Adhesion Assay Compounds of the present invention have been demonstrated to act via interaction with the integrin LFA-1, specifically by binding to the a L I domain which is known to be critical for the adhesion of LFA-1 to a variety of cell adhesion molecules. As such, it is expected that these compounds should block the interaction of LFA-1 with other CAMs, and this inhibition has been demonstrated for LFA-1 binding to ICAM-3. Compounds of the present invention were evaluated for the ability to block the adhesion of JY-8 cells to immobilized ICAM-3, as follows.
For measurement of inhibitory activity in the cell-based adhesion assay, 96-well microtiter plates were coated with 50 il of recombinant ICAM-3/Ig at a concentration of 10 tg/ml in CMF-PBS overnight at 4 0 C. The wells were washed twice with D-PBS, blocked by addition of 100 ul of D-PBS, 1% bovine serum albumin (BSA) by incubation for one hour at room temperature, and washed once with RPMI-1640/5% heat-inactivated FBS (adhesion buffer). Dilutions of compounds to be assayed for LFA-1/ICAM-3 antagonistic activity were prepared in adhesion buffer from 10 mM stock solutions in DMSO and 100 tl aliquots were added to duplicate wells. JY-8 cells (100 ul at 0.75 x 10 6 cells/ml in adhesion buffer) were then added to the wells (with or without 30 ng/ml IL-8). Microtiter plates were incubated for 30 min at room temperature, the adherent cells were fixed with 50 ul of 14% glutaraldehyde/D-PBS and incubation carried out for an additional 90 min. The wells were washed gently with dH20 and 50 ll ofdH20 was added, followed by 50 ul of 1% crystal violet. After five minutes, the plates were washed twice with dH 2 O and 225 ul ethanol (EtOH) was added to each well to extract the crystal violet from the cells. Absorbance was measured at 570 nm in an ELISA plate reader. The percent inhibition of a candidate compound was calculated using the following equation: %inhibition 100 X 1 average OD w/compound average OD w/o compound WO 00/59878 PCT/US00/08840 Example 3 Synthesis of Negative Regulators Synthesis of various diaryl sulfide compounds according to the invention is described below.
General Procedures and Starting Materials In general, solvents were purchased anhydrous and were not dried or purified further and starting materials were the best commercially available. Thin layer chromatography (TLC) was carried out using E. Merck, silica gel 60 F254, 0.25 mm, glass or aluminum coated plates, or Analtech silica gel uniplates (250 microns of silica). Visualization was achieved by UV. Reported RfS indicate a single spot detected. Flash chromatography was performed on E. Merck silica gel 60, 230-400 mesh. Nuclear magnetic resonance (NMR) spectra were recorded on Bruker DPX 300 or Varian 300 Gemini 2000 spectrometers. Chemical shifts are reported in parts per million (ppm) downfield from tetramethylsilane (TMS). Coupling constants are in Hz. In other cases, NMR spectra were recorded using a Unity XL-200 spectrometer.
Mass spectra were recorded on either a VG 70 SEG instrument at the University of Washington, Department of Medicinal Chemistry, Mass Spectrometry Facility (high resolution) or a Finnigan Mat TSQ 70 spectrometer (low resolution). Only the molecular ions are reported. Elemental analyses were performed by Quantitative Technologies, Inc.
A. General Synthesis Method A: Aminodiarylsulfides 1. General Description of Synthesis Method A One molar equivalent (eq) of a desired thiolphenol and 1 molar equivalent of the respective nitroaryl compound were placed in a dry flask under nitrogen and dissolved in dry acetone. One and a half molar equivalents anhydrous potassium carbonate (K 2 C0 3 was added and the mixture stirred vigorously overnight.
The reaction was then diluted with ether, washed with saturated NaHCO 3 3% NaHSO 4 and saturated NaCl. The organics were dried over Na 2
SO
4 and filtered.
Heptane was then added and the solution was concentrated by boiling until most of .1 1.
WO 00/59878 PCT/USOO/08840 the ether was removed. Upon cooling, the nitrodiaryl sulfide crystallized. The product was collected by filtration, washed with pentane, and dried in vacuo.
One molar equivalent of the diaryl sulfide and 5 molar equivalents tin chloride dihydrate were dissolved in ethanol (10 to 30 volumes). The mixture was heated to 60°C in an oil bath and concentrated HCI was added (10 to 30 volumes).
After three hours, the reaction was allowed to cool and 20 to 60 volumes of ice was added. The mixture was neutralized to pH 10 12 by addition of 5 N NaOH. The mixture was extracted twice with ether and the combined ether extracts were washed with saturated NaHC0 3 and saturated NaCl. The organic layer was dried over NaSO4, filtered, and the solvent stripped by rotary evaporation. The resulting solid or oil was taken up in ether, and approximately 5 molar equivalents of 1 N HC1 in ether was added drop-wise. The resulting solid was collected by filtration, washed with ether, and dried in vacuo.
The general synthesis method A is diagramed schematically below, wherein X is a halogen, chlorine.
R
R
X HS, H Ar K 2 C0 3 0 S Ar O NR Acetone R O O 1) SnC 2 2H 2 0 cone. HC1 EtOH
C
2) HCl/ether R
H
I A r HC1 H 2 N
R
H~l R WO 00/59878 PCT/US00/08840 2. Specific Example of General Synthesis Method A General Synthesis Method A was employed as specifically described below with the exception that tin granules were employed instead of the preferred tin chloride dihydrate shown in the schematic above.
2,4-Dichlorophenyl-(2'-chloro-4'-nitrophenyl)-sulfide 1.54 g (8.60 mmol) 2,4-dichlorothiophenol and 1.65 g (1 eq, 8.60 mmol) 3,4-dichloronitrobenzene were placed in a dry flask under an atmosphere of nitrogen and dissolved in 20 ml dry acetone. 1.78 g (1.5 eq, 12.9 mmol) anhydrous potassium carbonate (K,C0 3 was added and the mixture was stirred magnetically for hrs. The reaction was then diluted with 150 ml ether and then washed with saturated NaHCO 3 (2 x 75 ml), 3% NaHSO 4 (2 x 75 ml) and saturated NaCI (2 x ml). The organic phase was then dried over anhydrous Na 2
SO
4 and filtered. 50 ml heptane was then added, and the solution was concentrated to approximately 50 ml by boiling on a steam bath. Upon cooling high quality crystals formed. These were collected by filtration, rinsed with three portions of pentane, and dried in vacuo 0.05 mm Hg, 2 hrs). 1.43 g (50% yield) of yellow crystals were obtained, mp 145-146 R 0.
37 (Ethyl acetate [EtAc]/Heptane, 1:20). 'H NMR (CDC13) 6.71 (d, J=8.9, 1H), 7.38 (d of d, J 2 1H), 7.59 J=8.2, 1H), 7.63 J=2.2, 1H), 7.94 (d ofd, J 2 1H), 8.26 J=2.5, 1H). MS (EI) m/z 333 98).
Anal. Calcd for C, 2
H
6 C1 3 NO2S: C, 43.08; H, 1.81; N, 4.19. Found: C, 43.06; H, 1.77; N, 4.02.
3-Chloro-4-(2,4-dichlorophenylsulfanyl)-phenylamine Hydrochloride 0.680 g (2.03 mmol) 2,4-Dichlorophenyl-(2'-chloro-4'-nitrophenyl)-sulfide and 0.844 g (3.5 eq, 7.11 mmol) tin granules were slurried in 20 ml concentrated HC1. With vigorous stirring, this was heated to reflux for two days. The mixture was allowed to cool to room temperature and then was diluted with 50 ml ice. While stirring, this mixture was neutralized to pH 10 by the addition of approx. 80 ml 5 N NaOH. This was then extracted with ether (2 x 100 ml). The combined organics were washed with 100 ml saturated WO 00/59878 PTUO/84 PCT/USOO/08840 NaHCO, and 2 x 100 ml saturated NaCi. This was dried over Na,S0 4 and filtered.
This was concentrated on a rotavapor to give a brown oil. The oil was taken up in ml ether, and 4 ml 1 N HCI in ether was then added, drop wise, producing a white solid. This was collected by filtration, rinsed with several portions of ether, and dried (90' 0 C, 3 hrs, 0.05 mm. Hg). 0.548 g (79% yield) of a white solid was obtained, mp 183-185'C (dec), Rf 0.33 (EtAc/Heptane 1:2 w/ 1% TEA). 'H NMR (DMSO-d6) 6.55 J=8.6, lH), 6.66 (d of d, J 1
J
2 IH), 6.89 (br s, 3H), 6.89 I1H), 7.29 J=2.2, I 7.3 2 J=2.2, I1H), 7.62 1=2.2, IlH). MIS (El) mlz 303 M+ [100]. Anal. Calcd for C 1 2
H
8 Cl 3 NS-HCI: C, 42.26; H, 2.66; N, 4.11. Found: C, 42.39; H, 2.57; N, 4.14.
3. Additional Examples of Compounds Prepared by General Synthesis Method A The following compounds were prepared from commercially available reagents using General Synthesis Method A. The starting thiol and nitroaryl compounds are provided, along with the intermediate (thiol nitroaryl intermediate).
3-Chloro-4-(2-chlorophenylsulfanyl)-phenylamine hydrochloride 2-chlorothiophenol 3,4-dichloronitrobenzene 4-nitro-2-chlorophenyl-(2 chlorophenyl) -sulfide Grey solid, mp 197-198 *C (dec), Rf0.1 6 (EtAc/Heptane 1:4 w/ 1 TEA). 'H NMR (DMSO-d6) 6.61 (d of d, J 1=1'9, J273 1 6.70 (d of d, J 1=2.4, J2=8.4, 1H), 6.94 J=2.5, lH), 7.18 (in, 2H), 7.28 J=8.9, 1H), 7.45 (d of d, 11=1.6, J2=7.6, 1H), 7.78 (br s, 3H). MS (El) m/z 269 M+ [100]. Anal. Calcd for C1 2 HIOC1 3 NS-HCI: C, 47.22; H, 3.30; N, 4.59. Found: C, 47.34; H, 3.15; N, 4.45.
3-Chloro-4-(2-naphthylsulfanyl)-phenylamine hydrochloride 2-naphthalenethiol 3,4-dichloronitrobenzene 4-nitro-2-chlorophenyl-2-naphthylh sulfide Off white solid, mp 188'C (dec), RfO0.1 6 (EtAc/Heptane 1:4 w/ 1% TEA). 'H NMR (DMSO-d6) 6.08 (br s, 3H), 6.60 (d of d, JI=2.4, 12=8.4, lH), 6.90 WO 00/59878 WO 0059878PCT/USOO/08840 J=2.5, IH), 7.20 (d of d, J 1=1.9, I 7.30 J=8.6, I1H), 7.45 (in, 2H), 7.53 I1H), 7.76 (mn, IlH), 7.83 (in, 2H). MIS (El) in/z 285 M+ 100]. Anal. Calcd for C, 6 H,2C1NS-HCl: C, 59.64; H, 4.07; N, 4.35. Found: C, 59.59; H, 4.07; N, 4.09.
3-Chloro-4-(2,3-dichlorophenylsulfanyl)-phenylamine hydrochloride 2,3-dichiorothiophenol +3,4-dichloronitrobenzene 4-nitro-chlorophenyl-(2' ,3 dichlorophenyl)-sulfide Off white solid, mp 207-209*C (dec), RfO0.
33 (EtAc/Heptane 1:2 w/ 1% TEA). 'H NMR (DMSO-d6) 6.45 (dof 12=8.1, IH), 6.62 (dof d, J 1=2.2, J 2 1 6.79 (br s, 3H), 6.86 J=2.5, I1H), 7.21 J=7.9, I 7.33 (d, J=8.2, 1H), 7.38 (d of d, J 1
J
2 IH). MIS (El) m/z 303 93). Anal. Calcd for C, 2
H
8 Cl 3 NS-HCl: C, 42.26; H, 2.66; N, 4.11. Found: C, 42.49; H, 2.56; N, 4.10.
3-Chloro-4-(2,4,5-trichlorophenylsulfanyl)-phenylamine hydrochloride 2,3 ,4-trichlorothiophenol 3 ,4-dichloronitrobenzene 4-nitro-2-chlorophenyl- ,4 '-trichlorophenyl)-sulfide Off white solid, mp 192-194*C (dec), RfO0.
33 (EtAc/Heptane 1:2 w/ 1% TEA). 'H NMR (DMSO-d6) 6.46 (br s, 3H), 6.53 1H), 6.65 (d of d, 11=2.3, J2=8.4, 1H), 6.89 J=2.2, 1H), 7.35 J=8.6, 1H), 7.88 IH). MIS (El) mlz 337 73). Anal. Calcd for C, 2
H
7 Cl 4 NS-HCl: C, 38.38; H, 2.15; N, 3.73. Found: C, 38.73; H, 2.07; N, 3.60.
3-Methoxy-4-(2,3-dichlorophenylsulfanyl)-phenylamine 2,3-dichiorothiophenol 2-bromo-5-nitroanisole 4-nitro-2-methoxyphenyl-(2 1,3 1 dichlorophenyl)-sulfide This compound was isolated by crystallization from ether rather than from the formation of the HCl salt. Off white crystals were obtained, mp 166-167 0
C,
Rf 0.17 (EtAc/Heptane 1:2 w/ 1% TEA). 'H NMR (DMSO-d6) 3.67 3H), 5.75 (br s, 2H), 6.26 (d of d, J 1=19, I 6.37 J=1.9, I1H), 6.46 J=7.9, I1H), 7.13 J=8.2, IH), 7.16 J=8.1, 1H), 7.31 J=7.9, IH). MIS (El) ni/z 299 M+ [100].
Anal. Calcd for C, 3 H, IC1 2 NOS: C, 52.01; H, 3.69; N, 4.67. Found: C, 51.97; H, 3.59; WO 00/59878 WO 0059878PCTIUSOO/08840 N, 4.67.
5-Amino-2-(2,3-dichlorophenylsulfanyl)-acetophenone hydrochloride 2,3-dichiorothiophenol 2-chioronitroacetophenone 4-nitro-2-acetylphenyl-(2',3 dichlorophenyl)-sulfide Yellow solid, mp 151-153 oC (dec), Rf 0.35 (EtAc/Heptane 1: 1 w/ I% TEA).
I1H NMR (DMSO-d6) 2.51 3H), 7.00-7.09 (in, 311), 7.34 J=7.9, 1 7.41 (br s, 1H), 7.44 (br s, 3H), 7.58 1H). MS (El) m/z 311 M+ [100]. Anal. Calcd for C13HI 1Cl3NOS.HCl: C, 48.23; H, 3.47; N, 4.02. Found: C, 47.78; H, 3.43; N, 3.79.
4-(2,3-dichlorophenylsulfanyl)-phenylamine 2,3-dichlorothiophenol 4-bromonitrophenol 4-nitrophenyl-(2 '-dichlorophenyl)sulfide Off white solid, mp 175'C (dec), Rf 0.31 (EtAc/Heptane 1:2 w/ 1% TEA). 'H NMR (DMSO-d6) 7.13 J=8.2, 2H), 7.26 J=7.8, 1H), 7.42 J=8.6, 2H), 7.47 (d of d, J 2 111), 7.76 (d of d, 111), 8.04 (br s, 3H). MS (El)mnz 269 M+ [100]. Anal. Calcd for C, 2
H
9 Cl 2 NS-HCl: C, 47.00; H, 3.29; N, 4.57. Found: C, 46.92; H, 3.36; N, 4.27.
3-Chloro-4-(l-naphthylsulfanyl)-phenylamine hydrochloride 1 -naphthalenethiol 3,4-dichloronitrobenzene- 4-Nitro-2-chlorophenyl-( 1 -naphthyl)sulfide Off white solid, mp 192-194'C, Rf 0.37 (EtAc/Heptane 1:2 w/ 1% TEA). 'H NMR (DMSO-d6) 6.84 (in, 2H), 7.20 J=1.9, 1H), 7.40 J=7.0, 1H), 7.50 J=7.8, 11H), 7.56-7.61 (in, 2H), 7.94 J=8.2, 1H), 7.99 (mn, 111), 8.12-8.15 (mn, 5H). MS (El) in/z 285 M+ [100]. Anal. Calcd for C, 6
H,
2 ClNS-HCl: C, 59.64; H, 4.07; N, 4.35. Found: C, 59.59; H, 4.19; N, 4.11.
3-Methyl-4-(2,4-dichloroph enylsulfanyl)-phenylamine hydrochloride 2,4-dichlorothiophenol 2-broino-5-nitrotoluene 4-Nitro-2-methylphenyl-(2', 4'dichlorophenyl)-sulfide WO 00/59878 WO 0059878PCTIUSOO/08840 Off white solid, mp 195-197'C, RfO0.
4 1 (EtAc/Heptane 1:2 w/ 1% TEA). 1H NMR (DMSO-d6) 2.22 3H), 6.62 J=8.6, IH), 6.94 J=8.2, 111), 7.04 (br s, IRH), 7.29 I1H), 7.32 I1H), 7.66 J=2.2, I1H), 7.89 (br s, 3H). MS (El) m/z 283 M+ [100]. Anal. Calcd for C, 3
H,
1 C,NSHCI: C, 48.69; H, 3.77; N, 4.37.
Found: C, 48.87; H, 3.73; N, 4.34.
3-Bro mo-4-(2,4-dich loroph enylsulfanyl)-phenyla mine Hydrochloride 2),4-dichiorothiophenol 3 -bromo-4-chloronitrobenzene 4-Nitro-2-bromophenyl- 4' -dichlorophenyl)-sulfide Off white solid, mp 171-173'C (dec), Rf 0.
69 (EtAc/Heptane 1: 1 w/ 1% TEA). 1H NMR (DMSO-d6) 6.66 J=8.6, 1H), 6.81 (d of d, J 1
J
2 =8.4, 1 7.19 J=2.2, I 7.29 J=8.2, I 7.32 (d of d, J 1
J
2 1 7.64 (d, J=2.2, 1H), 7.92 (br s, 3H). MS (El) ni/z 347 60). Anal. Calcd for
C
12
H
8 BrCl 2 NS-HC1: C, 37.39; H, 2.35; N, 3.63. Found: C, 37.78; H, 2.28; N, 3.61.
2,5-Dichloro-4-(2,4-dichlorophenylsulfanyl)-ph enylamine Hydrochloride 2,4-dichlorothiophenol 2,4,5-trichloronitrophenol 4.-Nitro-2, 4 '-dichlorophenyl)-sulfide Off white solid, mp 168-176'C, Rf 0.39 (EtAc/Heptane 1:2 w/ 1% TEA). 1H NMR (DMSO-d6) 6.57 J=8.6, 1H), 6.83 (br s, 3H), 7.06 1H1), 7.30 (d of d, Jj= 2.2, IH), 7.55 IH), 7.63 J=2.2, 114). MS (El) mlz 347 Anal. Calcd for C 12
H
7 Cl 4 NS-0.75HCI: C, 39.34; H, 2.13; N, 3.82. Found: C, 39.34; H, 2.11; N, 3.7 1.
4,5-Dichloro-2-(2,4-dichlorophenylsulfanyl)-pheflylamifle 2,4-dichlorothiophenol 3 ,4-dichloro-2-fluoronitrobenzene 4-Nitro-2-chloro-5fluorophenyl-(2', 4' -dichlorophenyl)-sulfide Starting with, 3,4-dichloro-2-fluoronitrobenzene, the fluoride is displaced to give the 2-sulfanyl compound. This compound was purified by flash chromatography as the free amine, an off white solid, mp 1 I 9-122'C, RfO0.
2 0 (EtAc/Heptane 1:20). 1H NMR (DMSO-d6) 5.93 2H), 6.61 J=8.6, 1H), 7.08 (s, WO 00/59878PCUOI084 PCTIUSOO/08840 1H), 7.31 (d of d, IH), 7.55 111), 7.65 J=1.9, 1H). MIS (El) m/z 337 75). Anal. Calcd for C, 2
H
7 C1 4 NS: C, 42.5 1; H, 2.08; N, 4.13. Found: C, 42.94; H, 1.97; N, 4.08.
3,5-Dich Ioro-4-(2,4-dichlorophenvlsulfanyl)-phenvlamine 2,4-dichiorothiophenol 3,4,5-trichloronitrobenzene 4-Nitro-2, 6-dichiorophenyl- 4' -dichlorophenyl)-sulfide This compound was recrystallized as the free amine to give a white solid, mp 128-131'C, Rf 0.36 (EtAc/Heptane 'H NMR (DMSO-d6) 6.33 (brs, 2H), 6.46 J=8.5, 1H), 6.83 2H), 7.32 (d of d, J 2 IH), 7.65 J=2.2, 111). MIS (El) m/z 337 77). Anal. Calcd for C, 2
H
7 Cl 4 NSt C, 42.51; H, 2.08; N, 4.13. Found: C, 42.15; H, 2.10; N, 4.01.
2,3-Dichloro-4-(2,4-dichlorophenylsulfanyl)-phenylamine 2,4-dichiorothiophenol 2,3,4-trinitrobenzene 2,3-dichloro-4-(2,4dichlorophenylthio)-nitrobenzene This compound was purified by flash chromatography as the free amine to give a white solid, mp 116-119 0 C, RfO0.
3 3 (EtAc/Heptane 1:4 w/ 1% TEA).
'H NMR (DMSO-d6) 6.09 2H), 6.47 J=8.7, 1H), 6.86 J=8.7, IH), 7.32 (d of d, JI2 1H), 7.46 J=8.7, 1H), 7.69 J=2.2, IH). MIS (El) mlz 337 71). Anal. Calcd for C, 2
H
7 Cl 4 NS: C, 42.51; H, 2.08; N, 4.13. Found: C, 42.83; H, 2.02; N, 4.06.
B. General Synthesis Method B A General Synthesis Method B is diagramed schematically below: WO 00/59878 PTUOI84 PCr[USOO/08840 0-X H\ArK)C,C 0- 0 1 Ar R: r Acetone xi)' N+ reflux
R
00 Fe 0 IMeOH aq. NH 4
CI
reflux
R
Ar
H
2 N R0 wherein X is a halogen, preferably chloro.
The following compounds were prepared using General Synthesis Method B: 4-Nitro-2-chlorophenyl-(2',4'-dimethylphenyl)-sulfide 4-Amino-2-chlorophenyl-(2',4'-dimethylphenyl) -sulfide hydrochloride 4-Amino-2-chlorophenyl-(2'-methyl-4'-chlorophenyl)-sulfide 4-Amino-2-chlorophenyl-(2',4'-difluorophenyl)-sulfide hydrochloride 4-amino-2-chlorophenyl-(2',4',6'-trichlorophenyl)-sulfide 4-Amino-2-chlorophenyl-(2'-amino-4'-chlorophenyl)-sulfide 4-Amino-2-chlorophenyl-(3',4'-dichlorophenyl)-sulfide 4-Axnino-2-chlorophenyl-2-(3-chloro-5-trifluoromethylpyridyl)-sulfide Bis-(4,4'-diamino-2,2'-dichlorophenyl)-sulfide 4-Amino-2-chlorophenyl-(2',4'-dichlorophenyl)-sulfide 2-Chloro-4-amino-5-methylaminophenyl-(2',4'-dichlorophenyl)-sulfide 2-Chloro-4-amino-5-morpholinophenyl-(2',4'-dichlorophenyl)-sulfide 4-Amino-2-trifluoromethylphenyl-(2',4'-dichlorophenyl)-sulfide 4-Amino-2- fluorophenyl-(2',4'-dichlorophenyl)-suI fide -Amino-3-chlorophenyl-(2',4'-dichlorophenyl)-sulfide WO 00/59878 PCT/USOO/08840 The initial step in the General Synthesis Method B is illustrated by preparation of the intermediate compound described immediately below.
4-Nitro-2-chlorophenol-(2',4'dimethylphenyl)-sulfide 2,4-Dimethylthiophenol (1.0 g) and 3,4-dichloronitrobenzene (1 eq) were added to 100 ml acetone containing KCO, (5 The mixture was refluxed for 24 hr. After cooling to room temperature, the mixture was filtered and the acetone removed using a rotary vacuum. The resulting residue was dissolved in a small volume ofCH 2 and filtered. The CH,Cl, was then removed using a rotary vacuum.
The resulting residue was treated with methanol (MeOH), which caused the product to precipitate. The yellow solid product was then collected by filtration and dried at 0 C in a vacuum oven for 24 hr. The product yield was 66%, and the melting point was 128-130 0 C. Standard analytical techniques, including proton NMR spectroscopy, elemental analysis and thin layer chromatography, were employed to characterize the product.
The following compounds were prepared using an initial step generally in accordance with the above illustrative intermediate.
4-Amino-2-chlorophenyl-(2',4'-dimethylphenyl)-sulfide Hydrochloride 2-Chloro-4-nitrophenyl-(2',4'-dimethylphenyl)-sulfide (0.85 g) and iron powder in MeOH (300 ml) were added to 20 ml of an aqueous solution of NH 4 C1, in a molar ratio of 1:3:5 for substrate, iron powder and NH 4 CI, respectively. The mixture was stirred mechanically under reflux conditions overnight. The solvent was removed using a rotary vacuum. The resulting residue was dissolved in a small volume of chloroform and filtered. The product was obtained using preparative chromatography by applying the dissolved residue to a small column packed with silica gel (70-230 mesh), followed by elution using a solvent system of chloroform:hexane Fractions containing the product were combined and the solvents removed using a rotary vacuum. The off-white solid product was collected by filtration and dried at 50 0 C in a vacuum oven for 24 hr. The product yield was 57%, and the melting point was 180-185 C (decomposition). Standard analytical techniques, including proton WO 00/59878 PCT/USOO/08840 NMR spectroscopy, elemental analysis and thin layer chromatography, were employed to characterize the product.
4-Amino-2-chlorophenyl-(2'-methyl-4'-chlorophenyl)-sulfide 4-Nitro-2-chlorophenyl-(2'-methyl-4'-chlorophenyl)-sulfide (1.0 g) and iron powder in MeOH (300 ml) were added to 5 ml of an aqueous solution of NH 4 C1, in a molar ratio of 1:3:5 for substrate, iron powder and NHI 4 C, respectively. The mixture was stirred mechanically under reflux conditions overnight. The solvent was removed using a rotary vacuum. The resulting residue was dissolved in a small volume of chloroform:hexanes and filtered. The product was obtained using preparative chromatography by applying the dissolved residue-to a small column packed with silica gel (70-230 mesh), followed by elution using a solvent system of chloroform:hexane Fractions containing the product were combined and the solvents removed using a rotary vacuum. The white solid product was collected by filtration and dried at 60 0 C in a vacuum oven for 24 hr. The product yield was 71%, and the melting point was 91-93 0 C. Standard analytical techniques, including proton NMR spectroscopy, elemental analysis and thin layer chromatography, were employed to characterize the product.
4-Amino-2-chlorophenyl-(2',4'-difluorophenyl)-sulfide Hydrochloride 4-Nitro-2-chlorophenyl-(2',4'-difluorophenyl)-sulfide and iron powder in MeOH (300 ml) were added to 20 ml of an aqueous solution of NH 4 C1, in a molar ratio of 1:3:5 for substrate, iron powder and NH4C1, respectively. The mixture was stirred mechanically under reflux conditions overnight. The solvent was removed using a rotary vacuum. The resulting residue was dissolved in a small volume of chloroform and filtered. The product was obtained using preparative chromatography by applying the dissolved residue to a small column packed with silica gel (70-230 mesh), followed by elution using a solvent system of chloroform:hexane Fractions containing the product were combined and the solvents removed using a rotary vacuum. The colorless solid product was collected by filtration and dried at 0 C in a vacuum oven for 24 hr. The product yield was 88%, and the melting point WO 00/59878 PCT/US00/08840 was 187-190 0 C. Standard analytical techniques, including proton NMR spectroscopy, elemental analysis and thin layer chromatography, were employed to characterize the product.
4-amino-2-chlorophenyl-(2',4',6'-trichlorophenyl)-sulfide 4-Nitro-2-chlorophenyl-(2',4',6;-trichlorophenyl)-sulfide (0.6 g) and iron powder in MeOH (250 ml) were added to 32 ml of an aqueous solution of NH 4 C1, in a molar ratio of 1:3:5 for substrate, iron powder and NH 4 C1, respectively. The mixture was stirred mechanically under reflux conditions overnight. The solvent was removed using a rotary vacuum. The resulting residue was dissolved in a small volume of chloroform and filtered. The product was obtained using preparative chromatography by applying the dissolved residue to a small column packed with silica gel (70-230 mesh), followed by elution using a solvent system of chloroform:hexane Fractions containing the product were combined and the solvents removed using a rotary vacuum. The white solid product was collected by filtration and dried at 40 0 C in a vacuum oven for 24 hr. The product yield was 72%, and the melting point was 109-111 Standard analytical techniques, including proton NMR spectroscopy, elemental analysis and thin layer chromatography, were employed to characterize the product.
4-Amino-2-chlorophenyl-(2'-amino-4'-chlorophenyl)-sulfide 4-Amino-2-chlorophenyl-(2'-nitro-4'-chlorophenyl)-sulfide (1.02 g) and iron powder in MeOH (250 ml) were added to 60 ml of an aqueous solution of
NH
4 C1, in a molar ratio of 1:3:5 for substrate, iron powder and NH 4 C1, respectively.
The mixture was stirred mechanically under reflux conditions overnight. The solvent was removed using a rotary vacuum. The resulting residue was dissolved in a small volume of chloroform and filtered. The product was obtained using preparative chromatography by applying the dissolved residue to a small column packed with silica gel (70-230 mesh), followed by elution using a solvent system of chloroform:hexane Fractions containing the product were combined and the solvents removed using a rotary vacuum. The off-white solid product was collected WO 00/59878 PCT/USOO/08840 by filtration and dried at 50C in a vacuum oven for 24 hr. The product yield was 59%, and the melting point was 86-88 C. Standard analytical techniques, including proton NMR spectroscopy, elemental analysis and thin layer chromatography, were employed to characterize the product.
4-Amino-2-chlorophenyl-(3',4'-dichlorophenyl)-sulfide 4-Nitro-2-chlorophenyl-(3,4'-dichlorophenyl)-sulfide (1.0 g) and iron powder in MeOH (250 ml) were added to 40 ml of an aqueous solution of NH 4 CI, in a molar ratio of 1:3:5 for substrate, iron powder and NIHC1, respectively. The mixture was stirred mechanically under reflux conditions overnight. The solvent was removed using a rotary vacuum. The resulting residue was dissolved in a small volume of chloroform and filtered. The product was obtained using preparative chromatography by applying the dissolved residue to a small column packed with silica gel (70-230 mesh), followed by elution using a solvent system of chloroform:hexane Fractions containing the product were combined and the solvents removed using a rotary vacuum. The colorless solid product was collected by filtration and dried at 0 C in a vacuum oven for 24 hr. The product melting point was 103-105°C.
Standard analytical techniques, including proton NMR spectroscopy, elemental analysis and thin layer chromatography, were employed to characterize the product.
4-Amino-2-chlorophenyl-2-(3-chloro-5-trifluoromethylpyridyl)-sulfide 4-Nitro-2-chlorophenyl-2-(3-chloro-5-trifluoromethylpyridyl)-sulfide g) and iron powder in MeOH (250 ml) were added to 80 ml of an aqueous solution of NH 4 CI, in a molar ratio of 1:3:5 for substrate, iron powder and NH 4 Cl, respectively. The mixture was stirred mechanically under reflux conditions overnight.
The solvent was removed using a rotary vacuum. The resulting residue was dissolved in a small volume of chloroform and filtered. The product was obtained using preparative chromatography by applying the dissolved residue to a small column packed with silica gel (70-230 mesh), followed by elution using a solvent system of chloroform:hexane Fractions containing the product were combined and the solvents removed using a rotary vacuum. The colorless solid product was collected WO 00/59878 PCT/US00/08840 by filtration and dried at 60°C in a vacuum oven for 24 hr. The product yield was 97%, and the melting point was 96-98 0 C. Standard analytical techniques, including proton NMR spectroscopy, elemental analysis and thin layer chromatography, were employed to characterize the product.
Bis-(4,4'-diamino-2,2'-dichlorophenyl)-sulfide 4-Amino-2-chlorophenyl-(2'-chloro-4'-nitrophenyl)-sulfide and iron powder in MeOH (125 ml) were added to 20 ml of an aqueous solution of NH 4 C1, in a molar ratio of 1:3:5 for substrate, iron powder and NH 4 C1, respectively. The mixture was stirred mechanically under reflux conditions overnight. The solvent was removed using a rotary vacuum. The resulting residue was dissolved in a small volume of chloroform and filtered. The product was obtained using preparative chromatography by applying the dissolved residue to a small column packed with silica gel (70-230 mesh), followed by elution using a solvent system of chloroform:hexane Fractions containing the product were combined and the solvents removed using a rotary vacuum. The off-white solid product was collected by filtration and dried at 0 C in a vacuum oven for 24 hr. The product yield was 66%, and the melting point was 113-115 C. Standard analytical techniques, including proton NMR spectroscopy, elemental analysis and thin layer chromatography, were employed to characterize the product.
4-Amino-2-chlorophenyl-(2',4'-dichlorophenyl)-sulfide 2-Chloro-4-nitrophenyl-(2',4'-dichlorophenyl)-sulfide and iron powder in MeOH (50 ml) were added to 50 ml of an aqueous solution of NH 4 C1, in a molar ratio of 1:3:5 for substrate, iron powder and NH4C1 respectively. The mixture was stirred mechanically under reflux conditions overnight. The solvent was removed using a rotary vacuum. The resulting residue was dissolved in a small volume of chloroform and filtered, and the solvent was then removed from the filtrate using a rotary evaporator. The product was obtained by stirring the residue with 75 ml of distilled water (dH20). The colorless solid precipitate was collected by filtration. The product yield was 83%, and the melting point was 105-107°C. Standard analytical WO 00/59878 PCT/USOO/08840 techniques, including proton NMR spectroscopy, elemental analysis and thin layer chromatography, were employed to characterize the product.
4-Amin o-2-chlorophenyl-(4'-acetamido-2'-chlorophenyl)-sulfide 4-Nitro-2-chlorophenyl-(4'-acetamido-2'-chlorophenyl)-sulfide and iron powder in MeOH (250 ml) were added to 20 ml of an aqueous solution of NH 4 C1, in a molar ratio of 1:3:5 for substrate, iron powder and NH 4 Cl, respectively. The mixture was stirred mechanically under reflux conditions overnight. The solvent was removed using a rotary vacuum. The resulting residue was dissolved in a small volume of chloroform and filtered. The product was obtained using preparative chromatography by applying the dissolved residue to a small column packed with silica gel (70-230 mesh), followed by elution using a solvent system of chloroform-hexane Fractions containing the product were combined and the solvents removed using a rotary vacuum. The off-white solid product was collected by filtration and dried at 60 0 C in a vacuum oven for 24 hr. The product yield was 76%, and the melting point was 170-175°C. Standard analytical techniques, including proton NMR spectroscopy, elemental analysis and thin layer chromatography, were employed to characterize the product.
4-Amino-2-chlorophenyl-(4'-dimethylamino-2'-chlorophenyl)-sulfide 4-Nitro-2-chlorophenyl-(4'-dimethylamino-2'-chlorophenyl)-sulfide (0.2 g) and iron powder in MeOH (250 ml) were added to 20 ml of an aqueous solution of NH 4 C1, in a molar ratio of 1:3:5 for substrate, iron powder and NH 4 C1 respectively. The mixture was stirred mechanically under reflux conditions overnight.
The solvent was removed using a rotary vacuum. The resulting residue was dissolved in a small volume of chloroform and filtered. The product was obtained using preparative chromatography by applying the dissolved residue to a small column packed with silica gel (70-230 mesh), followed by elution using a solvent system of chloroform:hexane Fractions containing the product were combined and the solvents removed using a rotary vacuum. The pale yellow solid product was collected by filtration and dried at 60 0 C in a vacuum oven for 24 hr. The product yield was WO 00/59878 PCT/USOO/08840 58%, and the melting point was 133-135 0 C. Standard analytical techniques, including proton NMR spectroscopy, elemental analysis and thin layer chromatography, were employed to characterize the product.
2-Chloro-4-amino-5-methylaminophenyl-(2',4'-dichlorophenyl)-sulfide 2-Chloro-4-nitro-5-methylaminophenyl-(2',4'-dichlorophenyl)-sulfide (0.2 g) and iron powder in methanol (250 ml) were added to 20 ml of an aqueous solution of NH 4 C1, in a molar ratio of 1:3:5 for substrate, iron powder and NH 4 C1 respectively. The mixture was stirred mechanically under reflux conditions overnight.
The solvent was removed using a rotary vacuum. The resulting residue was dissolved in a small volume of chloroform and filtered. The product was obtained using preparative chromatography by applying the dissolved residue to a small column packed with silica gel (70-230 mesh), followed by elution using a solvent system of chloroform:hexane Fractions containing the product were combined and the solvents removed using a rotary vacuum. The brown solid product was collected by filtration and dried at 50 0 C in a vacuum oven for 24 hr. The product yield was 16%, and the melting point was 65-70 0 C. Standard analytical techniques, including proton NMR spectroscopy, elemental analysis and thin layer chromatography, were employed to characterize the product.
2-Chloro-4-amin o-5-N-morpholinophenyl-(2',4'-dichlorophenyl)-sulfide 2-Chloro-4-nitro-5-N-morpholinophenyl-(2',4'-dichlorophenyl)-sulfide (0.9 g) and iron powder in MeOH (250 ml) were added to 20 ml of an aqueous solution of NH 4 C1, in a molar ratio of 1:3:5 for substrate, iron powder and NH 4 C1 respectively. The mixture was stirred mechanically under reflux conditions overnight.
The solvent was removed using a rotary vacuum. The resulting residue was dissolved in a small volume of chloroform and filtered. The product was obtained using preparative chromatography by applying the dissolved mixture to a small column packed with silica gel (70-230 mesh), followed by elution using a solvent system of chloroform:hexane Fractions containing the product were combined and the solvents removed using a rotary vacuum. The off-white solid product was collected I. WO 00/59878 PCT/USOO/08840 by filtration and dried at 50 0 C in a vacuum oven for 24 hr. The product melting point was 153-155 0 C. Standard analytical techniques, including proton NMR spectroscopy, elemental analysis and thin layer chromatography, were employed to characterize the product.
4-Amino-2-trifluoromethylph enyl-(2',4'-dichlorophenyl)-sulfide 4-Nitro-2-trifluoromethylphenyl-(2',4'-dichlorophenyl)-sulfide (0.6 g) and iron powder in MeOH (250 ml) were added to 20 ml of an aqueous solution of
NH
4 CI, in a molar ratio of 1:3:5 for substrate, iron powder and NH 4 C1, respectively.
The mixture was stirred mechanically under reflux conditions overnight. The solvent was removed using a rotary vacuum. The resulting residue was dissolved in a small volume of chloroform and filtered. The product was obtained using preparative chromatography by applying the dissolved residue to a small column packed with silica gel (70-230 mesh), followed by elution using a solvent system of chloroform:hexane Fractions containing the product were combined and the solvents removed using a rotary vacuum. The product was collected by filtration and dried at 50 0 C in a vacuum oven for 24 hr. The product melting point was not determined because it was a colorless oil. Standard analytical techniques, including proton NMR spectroscopy, elemental analysis and thin layer chromatography, were employed to characterize the product.
4-Amino-2-fluorophenyl-(2',4'-dichlorophenyl)-sulfide 4-Nitro-2-fluorophenyl-(2',4'-dichlorophenyl)-sulfide and iron powder in MeOH (250 ml) were added to 20 ml of an aqueous solution of NH 4 C1, in a molar ratio of 1:3:5 for substrate, iron powder and NH 4 C1, respectively. The mixture was stirred mechanically under reflux conditions overnight. The solvent was removed using a rotary vacuum. The resulting residue was dissolved in a small volume of chloroform and filtered. The product was obtained using preparative chromatography by applying the dissolved residue to a small column packed with silica gel (70-230 mesh), followed by elution using a solvent system of chloroform:hexane Fractions containing the product were combined and the solvents removed using a WO 00/59878 PCT/USOO/08840 rotary vacuum. The pale yellow solid product was collected by filtration and dried at 0 C in a vacuum oven for 24 hr. The product yield was 55%, and the melting point was 101-102 C. Standard analytical techniques, including proton NMR spectroscopy, elemental analysis and thin layer chromatography, were employed to characterize the product.
5-Amino-3-chlorophenyl-(2',4'-dichlorophenyl)-sulfide 3-Chloro-5-nitrophenyl-(2',4'-dichlorophenyl)-sulfide (1.0 g) and iron powder in MeOH (250 ml) were added to 20 ml of an aqueous solution of NH 4 C1, in a molar ratio of 1:3:5 for substrate, iron powder and NH 4 C1, respectively. The mixture was stirred mechanically under reflux conditions overnight. The solvent was removed using a rotary vacuum. The resulting residue was dissolved in a small volume of chloroform and filtered. The product was obtained using preparative chromatography by applying the dissolved residue to a small column packed with silica gel (70-230 mesh), followed by elution using a solvent system of chloroform:hexane Fractions containing the product were combined and the solvents removed using a rotary vacuum. The product was collected by filtration and dried at 250C in a vacuum oven for 24 hr. The product yield was 16%, and the melting point was not determined because it was a brown oil. Standard analytical techniques, including proton NMR spectroscopy, elemental analysis and thin layer chromatography, were employed to characterize the product.
C. General Synthesis Method C A General Synthesis Method C is diagramed schematically below: Cl 0 SH 0 .O-0 K2CO 3 SA-N
O-
1I Acetone
H
2 N Ar reflux H 2 WO 00/59878 PCT/US00/08840 The following compounds were prepared by General Synthesis Method C: 4-Amino-2-chlorophenyl-(2'-chloro-4'-nitrophenyl)-sulfide 4-Amino-2-chlorophenyl-(2'-nitro-4'-chlorophenyl)-sulfide 4-Amino-2-chlorophenyl-6-(5-nitroquinolino)-sulfide 4-Amino-2-chlorophenyl-(2'-chloro-4'-nitrophenyl)-sulfide 2-Chloro-4-aminothiophenol (2.0 g) and 3,4-dichloronitrobenzene (1 eq.) were added to 300 ml of acetone containing K,C0 3 (20 The mixture was refluxed for 24 hr at 60 0 C. After cooling to room temperature, the mixture was filtered and the acetone removed using a rotary vacuum. The resulting residue was dissolved in a small volume of chloroform and filtered. The chloroform was then removed using a rotary vacuum. The resulting residue was triturated with MeOH, which caused the product to precipitate. The precipitate was collected by filtration and the yellow solid product dried at 80 0 C in a vacuum oven for 24 hr. The product yield was 59%, and the melting point was 135-136°C. Standard analytical techniques, including proton NMR spectroscopy, elemental analysis and thin layer chromatography, were employed to characterize the product.
4-Amino-2-chlorophenyl-(2'-nitro-4'-chlorophenyl)-sulfide 2-Chloro-4-aminothiophenol (2.0 g) and 2,5-dichloronitrobenzene (1 eq.) were added to 300 ml of acetone containing K,CO (20 The mixture was refluxed for 24 hr at 60 0 C. After cooling to room temperature, the mixture was filtered and the acetone removed using a rotary vacuum. The resulting residue was dissolved in a small volume ofCH 2 C1l and filtered. The CH 2 Cl 2 was then removed using a rotary vacuum. The resulting residue was triturated with MeOH, which caused the product to precipitate. The precipitate was collected by filtration and the yellow solid product dried at 60°C in a vacuum oven for 24 hr. The product yield was 57%, and the melting point was 191-193°C. Standard analytical techniques, including proton NMR spectroscopy, elemental analysis and thin layer chromatography, were employed to characterize the product.
WO 00/59878 PCT/US00/08840 4-Amino-2-chlorophenyl-6-(5-nitroquinolino)-sulfide 2-Chloro-4-aminothiophenol (0.474 g) and (1 eq.) were added to 250 ml of acetone containing KCO 3 (20 The mixture was refluxed for 24 hr at 60 0 C. After cooling to room temperature, the mixture was filtered and the acetone removed using a rotary vacuum. The resulting residue was dissolved in a small volume of chloroform and filtered. The chloroform was then removed using a rotary vacuum. The resulting residue was triturated with MeOH, which caused the product to precipitate. The product was obtained using preparative chromatography by applying the dissolved residue to a small column packed with silica gel (70-230 mesh), followed by elution using a solvent system of chloroform:hexane Fractions containing the product were combined and the solvents removed using a rotary vacuum. The yellow solid product was collected by filtration and dried at 50C in a vacuum oven for 24 hr. The product yield was 62%, and the melting point was 129-131 Standard analytical techniques, including proton NMR spectroscopy, elemental analysis and thin layer chromatography, were employed to characterize the product.
D. Specific Synthesis Procedures Full preparative methods are provided for the following compounds: 1-Acetamido-3-chloro-4-(2,3-dichlorophenylsulfanyl)-benzene 3-Hydroxy-4-(2,3-dichlorophenylsulfanyl)-phenylamine Hydrochloride 6-Chloro-5-(2,4-dichlorophenylsulfanyl)- H-benzimidazole 1-Acetamido-3-chloro-4-(2,3-dichlorophenylsulfanyl)-benzene 0.165 g (2.1 eq, 1.35 mmol) 4-dimethylaminopyridine (4-DMAP) was placed into a dry flask under an atmosphere of nitrogen and dissolved in 5 ml anhydrous tetrahydrofuran (THF). 2 ml of acetic anhydride was added followed by 0.220 g (1 eq, 0.643 mmol) 3-chloro-4-(2,3-dichlorophenylsulfanyl)-phenylamine hydrochloride. This was stirred for 18 hrs. The mixture was then diluted with 75 ml ether and washed with sat. NaHCO 3 (3 x 50 ml), 0.3 N HC1 (3 x 30 ml) and saturated NaCl (2 x 30 ml), dried over Na 2
SO
4 filtered, and the solvent stripped on a rotavapor.
WO 00/59878 PCT/USOO/08840 Flash chromatography (1.9 x 27 cm, EtAc/Heptane afforded 0.135 g (61% yield) of a white solid, mp 167-169 0 C, Rf 0.25 (EtAc/Heptane 'H NMR (DMSO-d6) 2.07 3H), 6.59 (d of d, J 2 1H), 7.24 J=8.1, 1H), 7.46 (d of d, J 1
J
2 1H), 7.55 2H), 8.04 J=1.9, 1H), 10.35 1H). MS (El) m/z 345 82). Anal. Calcd for C, 4 HICl 3 NOS: C, 48.51; H, 2.91; N, 4.04.
Found: C, 48.29; H, 2.88; N, 3.92.
3-Hydroxy-4-(2,3-dichlorophenylsulfanyl)-phenylamine Hydrochloride 0.19 g (0.67 mmol) 3-Methoxy-4-(2,3-dichlorophenylsulfanyl) phenylmine was placed into a dry flask under an atmosphere of nitrogen, dissolved in 10 ml dry CH 2 Cl1, and cooled to 0.32 ml (5 eq., 3.3 mmol) boron tribromide was added drop-wise, with stirring. The cold bath was removed and this was left to react for 20 hrs. The solution was again cooled to and then 10 ml MeOH was added drop-wise. This was allowed to warm to room temperature and was stirred for 1 hr.
The solvent was stripped on a rotavapor and the resulting oil was twice taken up in ml MeOH and again stripped. The material was again taken up in 10 ml MeOH and then diluted with 100 ml EtAc. The white precipitate that formed was removed by filtration, and then the filtrate was washed with saturated NaCI (3 x 50 ml), dried over Na2SO 4 filtered, and the solvent stripped by rotary evaporation. The resulting oil was purified by flash chromatography (2.9 x 28 cm, EtAc/Heptane and was then dissolved in 25 ml of ether and precipitated as the HCI salt by the addition of 5 ml 1 N HC1 in ether. This was collected by filtration, washed with ether, and dried (90 oC, 3 hr, 0.3 mm Hg) to gave 0.17 g (80% yield) of an off-white solid, mp 222 0 C (dec), R, 0.49 (EtAc/Heptane 'H NMR (DMSO-d6) 6.56-6.61 2H), 6.79 J=1.9, 1H), 6.90 (br hump), 7.19 J=8.1, 1H), 7.27 J=8.2, 1H), 7.38 (d of d, J=1.4,
J
2 1H), 10.37 (br s, 1H). MS (EI) m/z 285 M+ [100]. Anal. Calcd for
C,
2 HgC1 2 NOSHCl: C, 44.67; H, 3.12; N, 4.34. Found: C, 44.53; H, 2.91; N, 4.17.
6-Chloro-5-(2,4-dichlorophenylsulfanyl)- H-benzimidazole 4-Chloro-2-nitro-5-(2,4-dichlorophenylsulfanyl)-phenylamine was prepared by the general procedure with the exception that it was purified as the free WO 00/59878 PCT/USOO/08840 amine by flash chromatography. 1.37 g (3.93 mmol) of the free amine was added to ml DMF and 10 ml of EtOH. 4.43 g (5 eq, 19.7 mmol) tin chloride dihydrate was added followed by 10 ml concentrated HC. This was heated to 60 0 C for 20 hrs. The reaction was allowed to cool to ambient temperature, and was then diluted with 30 ml water, and brought to pH 12 by the addition of 30 ml 5 N NaOH. This mixture was twice extracted with 150 ml ether. The combined organics were washed with 100 ml sat. NaHCO 3 and 2 x 100 ml sat. NaCI, dried over Na 2
SO
4 and filtered. 40 ml heptane was then added, and this was concentrated on a rotavapor and dried in vacuo (100°C, 2 hr, 0.3 mm Hg) to yield 1.06 g of an analytically pure white solid, mp 206-208 0 C, Rf0.51 (CH 2 Cl/MeOH w/ 1% TEA). 'H NMR (DMSO-d6) 6.63 J=8.6, 1H), 7.28 (d of d, J J 2 1H), 7.69 J=2.2, 1H), 7.86 (br s, 1H), 7.93 1H), 8.38 1H), 12.80 1H). MS (EI) m/z 328 97). Anal. Calcd for
C,
3
H
7 Cl 3
N
2 S: C, 47.37; H, 2.14; N, 8.50. Found: C, 47.40; H, 2.04; N, 8.32.
E. Specific Synthesis Protocols The following compounds were prepared by methods as described below.
4-Methylamino-2,2',4'-trichlorodiphenylsulfide 4-Amino-2-chlorophenyl-(4'-acetamido-2'-chlorophenyl)-sulfide 4-Amino-2-chlorophenyl-(4'-dimethylamino-2'-chlorophenyl)-sulfide 4-Aminomethyl-2-chlorophenyl-(2',4'-dichlorophenyl)-sulfide 4-Methylamino-2,2',4'-trichlorodiphenylsulfide 4-Amino-2,2',4'-trichlorodiphenyl-sulfide (0.305 g, 1.0 mmol) was added to 15 ml of formic acid. The mixture was stirred for 8 hr, after which 0.23 ml of 37% formaldehyde was added and the mixture refluxed for 8 hr. The solvent was then removed using a rotary evaporator. The resulting residue was then applied to a small column containing silica gel (70-230 mesh). The product was then eluted from the column using chloroform. The solvent was removed from the eluate using a rotary evaporator and the pale yellow solid product collected. The product yield was 44%, and the melting point was 211 Standard analytical techniques, including proton NMR spectroscopy, elemental analysis and thin layer chromatography, were WO 00/59878 PCT/USOO/08840 employed to characterize the product.
4-Nitro-2-chlorophenyl-(4'-acetamido-2'-chlorophenyl)-sulfide 4-Amino-2,2'-dichloro-4'-nitrodiphenylsulfide (1.0 g, 3.17 mmol) was warmed in 15 ml of acetic anhydride containing a trace ofp-toluenesulfonic acid. The mixture was allowed to stand for 1 hr, after which the solvent was removed using a rotary evaporator. The residue was dissolved in EtAc and poured over a small column of silica gel (70-230 mesh). The filtrate was collected, evaporated to dryness, and recrystallized from acetonitrile to give a yellow solid product. The product yield was 97%, and the melting point was 163-165°C. Standard analytical techniques, including proton NMR spectroscopy, elemental analysis and thin layer chromatography, were employed to characterize the product.
4-Nitro-2-chlorophenyl-(4'-dimethylamino-2'-chlorophenyl)-sulfide 4-Amino-2-chlorophenyl-(2'-chloro-4'-nitrophenyl)-sulfide (1.0 g, 3.17 mmol) was added to a suspension of 60% sodium hydride (1.43 g) in 250 ml of THF at 0°C. Iodomethane (0.2 ml, in 20 ml of THF) was then added, and the mixture stirred at room temperature for 48 hr. The mixture was then applied to Analtech silica gel plates. Each plate had a thickness of 1000 microns of silica. The two products of the reaction, 4-nitro-2-chlorophenyl-(4-methylamino-2'-chlorophenyl)-sulfide and 4-nitro-2-chlorophenyl-(4'-dimethylamino-2'-chlorophenyl)-sulfide, were eluted using a 1:1 mixture of CHC1 3 and hexane. The slower eluting band corresponded to the former and the faster eluting band corresponded to the latter. After collecting the bands, the compounds were dissolved in CHC1 3 The solvent was removed by rotary evaporation to provide a yellow solid product. The yield of the desired product was 36%, and the melting point was 138-139 0 C. Standard analytical techniques, including proton NMR spectroscopy, elemental analysis and thin layer chromatography, were employed to characterize the product.
4-Aminomethyl-2-chlorophenyl-(2',4'-dichlorophenyl)-sulfide 2-Chloro-4-cyanopheny-(2',4'-dichlorophenyl)-sulfide (1.0 g, 3.18 WO 00/59878 PCT/USOO/08840 mmol) was added to 200 ml of THF under nitrogen at OoC. Lithium aluminum hydride (0.24 g) was then added to the solution in portions. The mixture was then allowed to stand for 2 hr at OoC, after which 1 ml of 20% NaOH was added to the mixture, followed by 1 ml of dH,O. The solution was then filtered, and solvents were removed using a rotary evaporator. The resulting residue was then applied to a small column containing silica gel (70-230 mesh). The column was washed with a 1:1 mixture of chloroform and hexane, after which the product was eluted using a rotary evaporator, and the product was obtained as a colorless oil. The product yield was and the melting point was not determined. Standard analytical techniques, including proton NMR spectroscopy, elemental analysis and thin layer chromatography, were employed to characterize the product.
F. Other compounds Other compounds synthesized in accordance with the general synthetic methods include the following.
Table II 4-Nitro-2-chlorophenyl-(2',4'-dimethylphenyl)-sulfide 4-Nitro-2-chlorophenyl-(2'-methyl-4'-chlorophenyl)-sulfide 4-Nitro-2-chlorophenyl-(2', 4'-difluorophenyl)-sulfide 4-Nitro-2-chlorophenyl-(2', 6'-trichlorophenyl)-sulfide 4-Nitro-2-chlorophenyl-(3', 4'-dichlorophenyl)-sulfide 4-Nitro-2-chlorophenyl-2-(3-chloro-5-trifluoromethylpyridyl)-sulfide 2-Chloro-4-nitrophenyl-(2', 4'-dichlorophenyl)-sulfide 4-Nitro-2-chlorophenyl-(4'-acetamido-2'-chlorophenyl)-sulfide 4-Nitro-2-chlorophenyl-(4'-dimethylamino-2'-chlorophenyl)-sulfide 2-Chloro-4-nitro-5-methylaminophenyl-(2', 4'-dichlorophenyl)-sulfide 2-Chloro-4-nitro-5-morpholinophenyl-(2', 4'-dichlorophenyl)-sulfide 4-Nitro-2-trifluoromethylphenyl-(2', 4'-dichlorophenyl)-sulfide 4-Nitro-2-fluorophenyl-(2', 4'-dichlorophenyl)-sulfide 3-Chloro-5-nitrophenyl-(2', 4'-dichlorophenyl)-sulfide WO 00/59878 WO 0059878PCTIUSOO/08840 4-Nitro-2-chlorophenyl-( 1 -naphthyl)- sulfide 4-Nitro-2-methylphenyl-(2', 4'-dich lorophenyl)-sul fide 4-Nitro-2-bromophenyl-(2', 4' -di chlorophenyl)-sul fide 4-Nitro-2, 5 -dichlorophenyl-(2', 4' -dichlorophenyl)-sulfide 4-Nitro-2, 6-dichlorophenyl-(2', 4' -dichlorophenyl)-sulfide 4-Nitro-2-chloro-5-fluorophenyl-(2', 4'-dichlorophenyl)-sulfide 4-Nitro-2, 3-dichlorophenyl-(2', 4' -dichlorophenyl)-sulfide 4-Nitro-2-chlorophenyl-(4' -chloro-2 '-aminophenyl)-sulfide 4-Nitro-5-acetamido-2-chlorophenyl-(2', 4'-dichlorophenyl)-sulfide 4-Nitro-2-chlorophenyl-(4' -methylamino-2 '-chlorophenyl)-sulfide 4-Nitro-2-chlorophenyl-(4' -benzylamino-2' -chlorophenyl)-sulfide 4-Nitro-2-chlorophenyl-(4 '-dibenzyiamino-2 '-chlorophenyl)-sulfide 4-Nitro-5-phenylsulfo-2-chlorophenyl-(2', 4 '-dichlorophenyl)-sulfide 3-Nitro-5-chlorophenyl-(2', 4' -dichlorophenyl)-sulfide Example 4 Cell-Based Assay Results Compounds of the invention displayed activity in the cell-based assays described above as set out in Table Ml below which provides RiM 'C 50 values for inhibition of LEA- I binding to ICAM-1I and ICAM-3 where tested. Paired values indicate inhibition in the absence and presence of IL-8. Multiple paired values Y/Z) indicate repeated experiemnents. Dashes indicate the experiment was not performed. "NT" indicates that the compound was not tested in a particular assay.
While the present invention has been described in terms of specific embodiments, it is understood that variations and modifications will occur to those skilled in the art. Accordingly, only such limitations as appear in the appended claims should be placed on the invention.
TABLE III LFA1/ICANI-I IC50 IL-8 LFA-I /ICANI-3 IC50 I L-8 Compound 3 -Chiloro-4-(2-chlorophenylsulfanyl)-phenylarnine hydrochloride 3-Chloro-4-(2-naphthylsul fanyl)-phenylamine hydrochloride 3-Chloro-4-(2 ,3-dichlorophenylsulfanyl)-phenylamine hydrochloride 3 -Chloro-4-(2,4,5-trichlorophenylsul fanyl)-phenylamine hydrochloride 3-Chloro-4-(2 ,4-dichlorophenylsulfanyl)-phenylaniine Hydrochloride 3-Metlioxy-4-(2,3-dichlorophenylsulfanyl)-phenylamine S -Amino-2-(2 ,3-dichlorophenylsulfanyl)-acetophenone hydrochloride 4-(2,3-dichlorophenylsulfanyl)-phenylamine 3 -Ch loro-4-( 1-naphthylsulfanyl)-phenylamine hydrochloride 3 -Methyl-4-(2,4-dichlorophenylsul fanyl)-phenylamine hydrochloride I -Acetamido-3-chloro-4-(2,3-dichlorophenylsul fanyl)-benzene 4-Methylamino-2,2',4'-trichlorodiphenyl sulfide 3-Bromo-4-(2,4-dichlorophenylsul fanyl)-phenylamine Hydrochloride 3 -Hydroxy-4-(2,3 -dichiorophenylsul fanyl)-phenylamine Hydrochloride 6-Chloro-5-(2,4-dichlorophenylsulfanyl)-l H-benzimidazole 4-Ami no-2 -ch lorophenyl-(2',4'-dimethylphenyl)-sulIfide Hydrochloride 2,5-Dichloro-4-(2,4-dichlorophenylsul fanyl)-phenylamine Hydrochloride 4-Amino-2-chlorophenyl-(2'-m ethyl-4'-chloropleyl)- sulfide 4-Amino-2-chlorophenyl fl uorophenyl)- sulIfide Hydrochloride 4-Am ino-2 -ch Iorophenyl-(2',4',6'-tri chlorophelyl)-sul fide 4-Amino-2 -chilorophenyl-(2'-amino-4'-chlorophenyl)-sul fide 4-Ami no-2-chlorophenyl-(2'-chloro-4'-ni trophelyl) -sulfide 4-Ami no-2-chlorophenyl-(2'-ni tro-4'-ch lorophenyl)-sul fide 4-Ami no-2-chlorophenyl-(3',4'-dichlorophenyl)-sulIfide 4-Amino-2-chlorophenyl-2-(3 -ch loro-5 -tri fluoromethylpyridyl)- sulfide Bis-(4,4'-diarnino-2,2'-dichlorophenyl)-sulfide 4-Amino-2-chlorophenyl-(2',4'-dich lorophenyl)-sulIfide 4-Amino-2-chlorophenyl-(4'-acetamido-2'-chlorophelyl)-suI fide 7.6/4.7; 17.7/16.3 7.8/7.2; 10.0/9.3 1. 1/0.7; 3.4/3.9 7.5/12.5; 18.9/22.6 1.6/1.9; 3.0/4.1; 5.0/6.0 8.7/7.0 /20 4-8.0; 8.5/7.6 /31 7.9/8.2; 7.0/6.5 /29.5 34.0/>40 4.0/7.5; 2.6/3.1 14.2/14.2 27.8/>41I 12 .6/2 3.0 3.3/3.9; 3.6/3.1; 2.2/4.0 13.4/22.6 16.5/17.8 8.7/12.4 29.0/36.0 10.0/12.0 6.7/7.8; 6.0/5.6 17.4/14.5 9.3/13.5; 7.6/7.3 35 .5/30. 5 6.0/6/7; 5.9/4.2 17.0/29.0
NT
NT
1.8/1.0
NT
1.2/1.4
NT
NT
NT
NT
NT
NT
NT
NT
NT
NT
NT
1.1/1.1
NT
NT
NT
NT
NT
2.5/2.3;
NT
NT
NT
2.1/2.0
NT
1.1/3.0 4-Amino-2-ch lorophenyl-6-(5 -nitroquinolino)-sulIfide 4-Amino-2-chlorophenyl-(4'-dimethylamino-2'-ch lorophenyl)-sul fide 2-Chloro-4-arnino-5-methylaminophenyl-(2',4'-dichlorophenyl)-sulfide 2-Chloro-4-amino-5-N-rnorpholinophenyl-(2',4'-dichlorophenyl)-sulfide 4-Amino-2-tri fl uoromethylphenyl-(2',4'-dich lorophenyl)-sulIfide 4-Aminomethyl-2-chlorophenyl-(2',4'-dich lorophenyl)-sul fide -Dichloro-2-(2,4-dichlorophenylsulfanyl)-phenylamine 3 ,5-Dichloro-4-(2,4-dich lorophenylsul fanyl)-phenylamine 2,3 -Dichloro-4-(2,4-dichlorophenylsulfanyi)-plienylamine 4-Amino-2 -fl uorophenyl-(2',4'-dichloropheniyl)-sulIfide 5-Amino-3 -clilorophenyl-(2',4'-dichlorophenyl)-sulIfide 4-(Benzothiazol-2-ylsulfanyl)-3-chloro-phenylamnme 3-Chloro-4-( 1 -chloro-naphthalen-2-ylsulfanyl)-phenylamine 4-Nitro-2-chlorophenyl-(2 '-dichloroplienyl)-sul fide 4-Am ino-2-ch forophenyl-2-(5 -nitro-3 -bromo)-pyri dine-sulIfide 6.0/7.3; 3.8/7 9.7/9.2 17.7/31.6 3 5.7/24.3 7.5/6.6 >40/ 18.7 31.5/35.0; 33.7/36 3.9/5.0 27.5/2 7.5 4.8/5.9 3 1.8/32.9 10.0/8.3; 7.8/8.8 1.0/1.5 29. 0/40 .0 >40/3 1.0 2.5/3.8
NT
NT
NT
NT
NT
NT
NT
NT
NT
NT
3.0/3.5 0.5/0.3
NT
NT

Claims (7)

1. A compound selected from the group consisting of:
3-Chloro-4-(2-chlorophenylsulfanyl)-phenylamine hydrochloride 3-Chloro-4-(2-naphthylsulfanyl)-phenylamine hydrochloride 3-Chloro-4-(2,3-dichlorophenylsulfanyl)-phenylamine hydrochloride. 3-Chloro-4-(2,4,5-trichlorophenylsulfanyl)-phenylamine hydrochloride 3-Methoxy-4-(2 ,3-dichlorophenylsulfanyl)-phenylamine
5-Amino-2-(2,3-dichlorophenylsulfanyl)-acetophenone hydrochloride 4-(2,3-dichlorophenylsulfanyl)-phenylamine 3-Chloro-4-(1 -naphthylsulfanyl)-phenylamine hydrochloride 3-Methyl-4-(2 ,4-dichlorophenylsulfanyl)-phenylamine hydrochloride 1 -Acetamido-3-chloro-4-(2 ,3-d ichlorophenylsulfanyl)-benzene 4-M ethylamino-2,2',4'-trichlo rod iphenylsu Ifid e 3-Bromo-4-(2,4-dichlorophenylsulfanyl)-phenylamine hydrochloride 3-Hydroxy-4-(2,3-dichlorophenylsulfanyl)-phenylamine hydrochloride
6-Chloro-5-(2,4-dichlorophenysulfanyl)-1 H-benzimidazole 4-Amino-2-chlorophenyl-(2',4'-dimethylphenyl)-sulfide hydrochloride 2,5-Dichloro-4-(2,4-dichlorophenylsulfanyl)-phenylamine hydrochloride 4-Amino-2-chlorophenyl-(2'-methyl-4'-chlorophenyl)-su Ifide 4-Amino-2-chlorophenyl-(2',4'-difluorophenyl)-sulfide hydrochloride 4-amino-2-chlorophenyl-(2',4',6'-trichlorophenyl)-sulfide 4-Amino-2-chlorophenyl-(2'-amino-4'-chlorophenyl)-sulfide 4-Amino-2-chlorophenyl-(2'-nitro-4'-chlorophenyl )-sulfide 25 4-Amino-2-chlorophenyl-2-(3-chloro-5-trifluoromethylpyridyl)-sulfide 4-Amino-2-chlorophenyl-(4'-acetamido-2'-chlorophenyl )-sulfide 4-Amino-2-chlorophenyl-6-(5-nitroqu inol ino)-su Ifide 4-Amino-2-chlorophenyl-(4'-d imethylamino-2'-chlorophenyl)-sulfide 2-Chloro-4-amino-5-methylaminophenyl-(2',4'-dichlorophenyl) sulfide 2-Chloro-4-amino-5-morpholinophenyl-(2',4'-dichlorophenyl)-sulfide 4-Amino-2-trifluoromethylphenyl-(2',4'-d ichiorophenyl )-sulfide :4,5-Dichloro-2-(2,4-dichlorophenylsulfanyl)-phenylamine 2,3-Dichloro-4-(2,4-dichlorophenylsulfanyl)-phenylamine 4-Amino-2-fluorophenyl-(2',4'-dichlorophenyl)-sulfide Y:\Mw\NKI NO DELETE MA1919-OO.doo 5-Amino-3-chlorophenyl-(2',4'-d ichiorophenyl )-su Ifide 4-N itro-2-chlorophenyl-(2',4'-dimethylphenyl )-sulfide 4-Nitro-2-chlorophenyl- (2'-methyl-4'-chlorophenyl)-su Ifide 4-N itro-2-chlorophenyl-(2',4'-d ifluorophenyl)-sulfide 4-N itro-2-chlorophenyl-(2',4',6'-trichlorophenyl )-sulfide 4-N itro-2-chlorophenyl-2-(3-chloro-5-trifluoromethylpyridyl)-sulfide 2-Chloro-4-nitrophenyl-(2', 4'-dichlorophenyl)-sulfide 4-Nitro-2-chlorophenyl-(4'-acetamido-2'-chlorophenyl) -sulfide 4-N itro-2-chlorophenyl-(4'-d imethylamino-2'-chlorophenyl )-sulfide 2-Chloro-4-nitro-5-methylaminophenyl-(2',4'-dichlorophenyl)-sulfide 2-Chloro-4-nitro-5-morpholinophenyl-(2',4'-dichlorophenyl)-sulfide 4-Nitro-2-trifluoromethylphenyl-(2',4'-d ichlorophenyl)-su Ifide 4-Nitro-2-fluorophenyl-(2',4'-dichlorophenyl)-sulfide 3-Chloro-5-nitrophenyl-(2',4'-dichlorophenyl)-sulfide 4-N itro-2-methylphenyl-(2',4'-d ichlorophenyl)-sulfide 4-Nitro-2-bromophenyl-(2',4'-dichlorophenyl)-sulfide 4-N itro-2, 5-d ichlorophenyl-(2',4'-d ichiorophenyl )-sulfide 4-Nitro-2, 6-dichlorophenyl-(2', 4-dichlorophenyl)-sulfide 4-N itro-2-chloro-5-fluorophenyl-(2',4'-d ichlorophenyl)-su [fide 4-N itro-2, 3-dichlorophenyl-(2,4'-dichlorophenyl)-sulfide 4-Nitro-2-chlorophenyl-(4'-chloro-2'-aminophenyl)-sulfide 4-N itro-5-acetamido-2-chlorophenyl-(2',4'-dichlorophenyl )-sulfide 4-N itro-2-chlorophenyl-(4'-methylamino-2'-chlorophenyl)-sulfide 4-Nitro-2-chlorophenyl-(4'-benzylamino-2'-chlorophenyl)-sulfide 25 4-N itro-2-chlorophenyl-(4'-d ibenzylamino-2'-chlorophenyl)-sulfide 4-Nitro-5-phenylsu Ifo-2-chlorophenyl-(2' ichlorophenyl)-sulfide 3-Nitro-5-chlorophenyl-(2',4'-d ichiorophenyl )-sulfide. 2. A pharmaceutical composition comprising the compound of claim 1 and a pharmaceutically acceptable carrier. 3. A pharmaceutical composition comprising a compound selected from the **group consisting of: 3-Chloro-4-(I -chloro-naphthalen-2-ylsulfanyl)-phenylamine Y'\M~y\NKI NO DELETE MR\41919O0doo 56 I -(3-Nitro-4-phenylsulfanyl-phenyl )-ethanone I -(3-Nitro-4-phenylsulfanyl-phenyl )-ethanone oxime 5-Trifluoromethyl-2-phenylsulfanyl-benzonitrile 1 -(3,5-dichlorophenyl)-3-phenylsulfanyl-pyrrolidine-2,5-dione Bis-2 ,4,6-Trinitrophenyl-sulfide 2-Methyl-i -(2-o-tolylsulfanyl-phenyl I H-pyrrole 3-[2-(4-Chloro-2-nitro-phenylsulfanyl )-phenylamino-3H-isobenzofu ran-I1-one 4-(Benzoth iazol-2-ylsulIfanyl)-3-ch loro-phenyla mine 2-Nitro-4-chlorophenyl-(2'-aminophenyl)-sulfide 6-Amino-2-chlorophenyl-(4'-methylphenyl)-su Ifide 4-Nitrophenyl-(2'-chlorophenyl)-sulfide. 2, 4-Dinitrophenyl-(4'-chlorophenyl) sulfide 4-Aminophenyl-(2'-chlorophenyl)-su Ifide 2,4-Diaminophenyl-(4'-isopropylphenyl)-sulfide 4-Nitro-2-chlorophenyl-(2',3'-dichlorophenyl)-sulfide 4-Amino-2-chlorophenyl-2-(5-nitro-3-bromo)-pyridine sulfide 4-Amino-2-chlorophenyl-(2'-nitro-4'-chlorophenyl )-sulfide. 4. A method for treating an inflammatory disorder, comprising a step of administering to a mammal an amount of the pharmaceutical composition of claim 2 or 3 sufficient to inhibit binding of LFA-1 to a natural ligand thereof that competes with lOAM-I1 or ICAM-3 for binding to LFA-I. A method for inhibiting LFA-1 binding to an lOAM that binds LFA-1 25 comprising the step of contacting LFA-I with a diaryl sulfide, 0 wherein the diaryl sulfide is a compound represented by the general structural 0*4: formula as:R A s B R 2 wherein A and B, independently, are aryl groups selected from the group 30 consisting of 5- and 6-membered aromatic rings, including, but not limited to, phenyl, thienyl, furyl, pyrimidinyl, imidazolyl, pyrazolyl, pyridyl, pyrazinyl, pyrrolyl, and pyridazinyl; Y:\Mary\NKI NO DELETE MR\41919.O0,do 57 R 1 R 2 and R 3 independently, is selected from the group consisting of hydrogen, -Ra, wherein Ra is hydrogen or an alkyl group containing one to six saturated straight or branched chain carbon atoms (C 1 -6 alkyl), -O-Ra, -halo, wherein halo is CI, F, Br, or I, -NRbRc, where Rb and Rc, independently, are H, C 1 -6 alkyl, or -CH 2 -aryl, -NO 2 -C(=O)Ra, -CN, -perfluoroRa, such as trifluoromethyl, -(CH 2 )n-NRbRc, wherein n is an integer from 1 to 6, a 5- or 6-membered heterocyclic ring, either aliphatic or aromatic, containing one or more of O, N, or S, optionally substituted, such as morpholino, and -S-aryl, wherein aryl is a 5- or 6-membered aromatic ring, optionally substituted; and R 4 R 5 and R 6 independently, are selected from the group consisting of hydrogen, -Ra, -O-Ra, -halo, -NRbRc, -N02, S 25 -C(=O)Ra, -CN oi. -perfluoroRa, -N-C(=O)Ra, S: -(CH2)n-NRbRc, and S 30 a 5-or 6- membered heterocyclic ring, aliphatic or aromatic, containing one or more of O, N, or S, optionally substituted, -S-aryl, or wherein R 4 and R 5 are taken together to form a 5- or 6-membered aromatic ring, optionally S' containing one or more of O, N, or S in the ring, optionally substituted. Y:\May\NKI NO DELETE MR\41919-00.doc 58 6. The method according to claim 5 wherein the diaryl sulfide is selected from the group consisting of: 3-Chloro-4-(2-chlorophenylsulfanyl )-phenylamine hydrochloride 4-Nitro-2-chlorophenyl-(2',3'-d ichlorophenyl)-sulfide 3-Chloro-4-(2-naphthylsulfanyl)-phenylamine hydrochloride 3-Chloro-4-(2,3-dichlorophenylsulfanyl)-phenylamine hydrochloride 3-Chloro-4-(2 ,4,5-trichlorophenylsu Ifanyl )-phenylamine hydrochloride 3-Chloro-4-(2,4-dichlorophenylsulfanyl)-phenylamine hydrochloride 4-(Benzothiazol-2-ylsulfanyl)-3-chloro-phenylamine 3-Chloro-4-(1 -chloro-naphthalen-2-ylsulfanyl)-phenylamine 3-Methoxy-4-(2,3-d ichlorophenylsu Ifa nyl)-phenyla mine 5-Amino-2-(2,3-dichlorophenylsulfanyl)-acetophenone hydrochloride 4-(2 ,3-d ich lo rophenylsu Ifanyl )-phenylam ine 3-Chloro-4-(1-naphthylsulfanyl)-phenylamine hydrochloride 3-Methyl-4-(2,4-dichlorophenylsulfanyl)-phenylamine hydrochloride 1 -Acetamido-3-chloro-4-(2,3-dichlorophenylsulfanyl)-benzene 4-Methylamino-2,2',4'-trichlorodiphenylsulfide 3-Bromo-4-(2,4-dichlorophenylsulfanyl)-phenylamine hydrochloride 3-Hyd roxy-4-(2 ,3-d ichlorophenylsulfanyl )-phenylamine hydrochloride 6-Chloro-5-(2 ,4-dichlorophenylsu lfanyl)-1 H-benzimidazole 4-Amino-2-chlorophenyl-(2'4'-d imethylphenyl)-su Ifide hydrochloride 2,5-Dichloro-4-(2,4-dichiorophenylsulfanyl)-phenylamine hydrochloride 4-Amino-2-chlorophenyl-(2'-methyl-4'-chlIorophenyl )-sulfide 0 4-Amino-2-chlorophenyl-(2',4'-difluorophenyl)-sulfide hydrochloride 0. 25 4-Amino-2-chlorophenyl-(2',4',6'-trichlorophenyl)-sulfide 4-mn--hoohnl(2-mn-'clrpey )-ulid 4-Amino-2-chlorophenyl-(2'-chloro-4'-nitrophenyl)-sulfide 0 4-Amino-2-chlorophenyl-(2'-nitro-4'-hlorophenyl)-sulfide 4-Amino-2-chlorophenyl-(3','- iochlorophenyl)-su fide 0000004-Amino-2-chlorophenyl-(2',4'-d ichlorophenyl)-sulfide 3 4-Amino-2-chlorophenyl-2('-cetamio--cloropehnpyl)-sulfide 0 ."*4-Amino-2-chlorophenyl-6(5'4-ihro inl)-sulfide Y:\M~ry\NKI NO DELETE MR\41919-OO.doe 59 4-Amino-2-chlorophenyl-(4'-dimethylamino-2'-chlorophenyl)-sulfide 2-Chloro-4-amino-5-methylaminophenyl-(2',4'-dichlorophenyl)-su Ifide 2-Chloro-4-amino-5-N-morpholinophenyl-(2',4'-dichlorophenyl)-sulfide 4-Amino-2-trifluoromethylphenyl-(2',4'-d ichlorophenyl)-sulfide 4-Amino-2-chlorophenyl-2-(5-nitro-3-bromo)-pyridime sulfide 4-Aminomethyl-2-chlorophenyl-(2', 4'-d ichlorophenyl)-sulfide 4,5-Dichloro-2-(2,4-dichlorophenylsulfanyl)-phenylamine 3,5-Dichloro-4-(2,4-dichlorophenylsulfanyl)-phenylamine 2,3-Dichloro-4-(2,4-dichlorophenylsulfanyl)-phenylamine 4-Amino-2-fluorophenyl-(2',4'-dichlorophenyl)-sulfide 5-Amino-3-chlorophenyl-(2',4'-dichlorophenyl)-sulfide 3-Chloro-4-(i -ch loro-naphthalen-2-ylsulIfa nyl)-phe nyla mine 1 itro-4-phenylsulIfa nyl-phenyl)-etha none 1-(3-N itro-4-phenylsulIfa nyl-phenyl)-etha none oxime 5-Trifluoromethyl-2-phenylsulfanyl-benzonitrile 1 ichiorophenyl )-3-phenylsu Bis-2 ,4 ,6-Trin itrophenyl-su Ifide 2-Methyl-I -(2-o-tolylsulfanyl-phenyl)-I H-pyrrole 3-[2-(4-Ch loro-2-n itro-phenyl sulfa nyl)-phenylam ino-3H-isobenzofu ran-i1 -one 4-(Benzothiazol-2-ylsulfanyl)-3-chloro-phenylamine 2-N itro-4-chlorophenyl-(2'aminophenyl)-sulfide 6-Amino-2-chlorophenyl-(4'-methylphenyl)-sulfide 4-Nitrophenyl-(2'-chlorophenyl)-sulfide 2, 4-Dinitrophenyl-(4'-chlorophenyl)-sulfide 4-Aminophenyl-(2'-chlorophenyl)-sulfide 2, 4-Diaminophenyl-(4'-isopropylphenyl)-sulfide 4-Nitro-2-chlorophenyl-(2',3'-dichlorophenyl)-sulfide 4-Amino-2-chlorophenyl-2-(5-n itro-3-bromo)-pyridime-sulfide. 30 7. A method for treating an inflammatory disorder arising from LEA-i binding to a natural ligand thereof that competes with ICM1or ICAM-3 for binding to LEA-i, comprising administering to a mammal in need thereof a compound that competes with 3-chloro-4-(i -chloro-naphthalen-2-ylsulfanyl)-phenylamine for Y:\M~y\NKI NO DELETE MR\41919-OOdoe binding to LFA-1 in an amount sufficient to inhibit binding of the natural ligand to LFA-1, wherein said compound is a diaryl sulfide represented by the general structural formula R, R4 R2 A R 3 R (I) wherein A and B, independently, are aryl groups selected from the group consisting of 5- and 6-membered aromatic rings, including, but not limited to, phenyl, thienyl, furyl, pyrimidinyl, imidazolyl, pyrazolyl, pyridyl, pyrazinyl, pyrrolyl, and pyridazinyl; R 1 R 2 and R 3 independently, is selected from the group consisting of hydrogen, -Ra, wherein Ra is hydrogen or an alkyl group containing one to six saturated straight or branched chain carbon atoms (C 1 -6 alkyl), -O-Ra, -halo, wherein halo is CI, F, Br, or I, -NRbRc, where Rb and Rc, independently, are H, C1-6 alkyl, or -CH 2 -aryl, -NO 2 -C(=O)Ra, -CN, 20 -perfluoroRa, such as trifluoromethyl, -N-C(=O)Ra, -(CH 2 )n-NRbRc, wherein n is an integer from 1 to 6, a 5- or 6-membered heterocyclic ring, either aliphatic or aromatic, containing one or more of O, N, or S, optionally substituted, such as morpholino, and -S-aryl, wherein aryl is a 5- or 6-membered aromatic ring, optionally substituted; and R 4 R 5 and R 6 independently, are selected from the group consisting of S.hydrogen, -Ra, 30 -0-Ra, -halo, -NRbRc, Y:\May\NKI NO DELETE MR\41919-00dod -NO 2 -ON -perfi uoroR, -NC(=0)Ra, -(CH 2 )rn-NRbRc, and -a 5- or 6-membered heterocyclic ring, aliphatic or aromatic, containing one or more of 0, N, or S, optionally substituted, -S-aryl, or wherein R 4 and R 5 are taken together to form a 5- or 6-membered aromatic ring, optionally containing one or more of 0, N, or S in the ring, optionally substituted.
8. The method according to claim 7 wherein the diaryl sulfide is selected from the group consisting of: 3-Chloro-4-(2-chlorophenylsulfanyl)-phenylamine hydrochloride 4-Nitro-2-chlorophenyl-(2',3'-dichlorophenyl)-sulfide 3-Chloro-4-(2-naphthylsulfanyl)-phenylamine hydrochloride 3-Chloro-4-(2,3-dichlorophenylsulfanyl)-phenylamine hydrochloride 3-Chloro-4-(2,4,5-trichlorophenylsulfanyl)-phenylamine hydrochloride 3-Chloro-4-(2,4-dichlorophenylsulfanyl)-phenylamine hydrochloride 4-(Benzothiazol-2-ylsulfanyl)-3-chloro-phenylamine 3-Chloro-4-(1 -chloro-naphthalen-2-ylsulfanyl)-phenylamine 3-Methoxy-4-(2,3-dichlorophenylsulfanyl)-phenylamine 5-Amino-2-(2,3-dichlorophenylsulfanyl)-acetophenone hydrochloride 25 4-(2,3-dichlorophenylsulfanyl)-phenylamine 3-Chloro-4-(1-naphthylsulfanyl)-phenylamine hydrochloride 3-Methyl-4-(2,4-dichlorophenylsulfanyl)-phenylamine hydrochloride I1 -Acetamido-3-chloro-4-(2 ,3-d ichlorophenylsulfanyl)-benzene 4-Methylamino-2,2',4'-trichlorodiphenylsulfide 3-Bromo-4-(2,4-d ich lorophe nylsulfa nyl)-phenyla mine hydrochloride 3-Hydroxy-4-(2,3-dichlorophenylsulfanyl)-phenylamine hydrochloride 6-Chloro-5-(2,4-dichlorophenylsulfanyl)-1 H-benzimidazole :4-Amino-2-chlorophenyl-(2'4'-dimethyl phenyl)-sulfide hydrochloride loro-4-(2,4-d ichlorophenylsu Ifa nyl)-phenyla mine hydrochloride Y:\M.,y\NKI NO DELETE MR\41919-OO.doo 62 4-Amino-2-chlorophenyl-(2'-methyl-4'-ch lorophenyl )-sulfide 4-Amino-2-chlorophenyl-(2',4'-difluorophenyl)-su Ifide hydrochloride 4-Amino-2-chlorophenyl-(2',4',6'-trichlorophenyl)-sulfide 4-Amino-2-chlorophenyl-(2'-amino-4'-chlorophenyl)-su Ifide 4-Amino-2-chlorophenyl-(2'-chloro-4'-nitrophenyl)-sulfide 4-Amino-2-chlorophenyl-(2'-nitro-4'-chlorophenyl)-sulfide 4-Amino-2-chlorophenyl-(3',4'-d ichiorophenyl )-su Ifide 4-Amino-2-chlorophenyl-2-(3-chloro-5-trifluoromethyl pyridyl)-sulfide Bis-(4,4'-d iamino-2 ichiorophenyl )-sulfide 4-Amino-2-chlorophenyl-(2',4'-dichlorophenyl)-sulfide 4-Amino-2-chlorophenyl-(4'-acetamido-2'-chlorophenyl)-sulfide 4-Amino-2-chlorophenyl-6-(5-nitroquinolino)-sulfide 4-Amino-2-chlorophenyl-(4'-dimethylamino-2'-chlorophenyl)-sulfide 2-Chloro-4-amino-5-methylaminophenyl-(2',4'-d ichiorophenyl )-su Ifide 2-Chloro-4-amino-5-N-morpholinophenyl-(2',4'-dichlorophenyl)-sulfide 4-Amino-2-trifluoromethylphenyl-(2' ,4'-dichlorophenyl)-sulfide 4-Amino-2-chlorophenyl-2-(5-nitro-3-bromo)-pyridime sulfide 4-Aminomethyl-2-chlorophenyl-(2', 4'-dichlorophenyl)-sulfide loro-2-(2,4-d ichlorophenylsu lfanyl)-phenyla mine 3, 5-Dichloro-4-(2 ,4-d ichlorophenylsu Ifanyl )-phenylamine 2,3-Dichloro-4-(2,4-dichlorophenylsulfanyl)-phenylamine 4-Amino-2-fluorophenyl-(2',4'-dichlorophenyl)-sulfide 5-Amino-3-chlorophenyl-(2',4'-dichlorophenyl)-sulfide 3-Chloro-4-(1 -ch loro-naphthalen-2-ylsulIfa nyl)-phenyla mine 1 itro-4-phenylsulIfa nyl-phenyl)-etha none 1 1-(3-N itro-4-phenylsulIfa nyl-phenyl)-etha none oxime 5-Trifluoromethyl-2-phenylsu Ifanyl-benzonitrile 1 -(3,5-dichlorophenyl)-3-phenylsulfanyl-pyrrolidine-2,5-dione *'.xBis-2 ,4,6-Trinitrophenyl-sulfide 2-Methyl-i -(2-o-tolylsulfanyl-phenyl)-1 H-pyrrole 3-[2-(4-Chloro-2-n itro-phenylsulfanyl )-phenylamino-3H-isobenzofuran-1 -one 4-(Benzothiazol-2-ylsulfanyl)-3-chloro-phenylamine 2-Nitro-4-chlorophenyl-(2'aminophenyl)-sulfide 6-Amino-2-chlorophenyl-(4'-methyl phenyl )-sulfide Y:%Ma~y\NYI NO DELETE MR\4199-00AMO 4-Nitrophenyl-(2'-chlorophenyl)-sulfide 2, 4-Dinitrophenyl-(4'-chlorophenyl)-sulfide 4-Aminophenyl-(2'-chlorophenyl)-sulfide 2, 4-Diaminophenyl-(4'-isopropylphenyl)-sulfide 4-Nitro-2-chlorophenyl-(2',3'-dichlorophenyl)-sulfide 4-Amino-2-chlorophenyl-2-(5-nitro-3-bromo)-pyridine-sulfide.
9. A method of inhibiting binding of LFA-1 to a natural ligand thereof that competes with ICAM-1 or ICAM-3 for binding to LFA-1, comprising contacting a 0o cell that expresses LFA-1 on its surface with a compound that competes with 3- chloro-4-(1-chloro-naphthalen-2-ylsulfanyl)-phenylamine for binding to LFA-1 in an amount that is sufficient to inhibit binding to LFA-1 to the natural ligand, wherein said compound is a diaryl sulfide represented by the general structural formula R, R 4 R 2 A S B R3 R 6 (I) wherein A and B, independently, are aryl groups selected from the group consisting of 5- and 6-membered aromatic rings, including, but not limited to, phenyl, thienyl, furyl, pyrimidinyl, imidazolyl, pyrazolyl, pyridyl, pyrazinyl, pyrrolyl, and pyridazinyl; 20 R 1 R 2 and R 3 independently, is selected from the group consisting of hydrogen, S-Ra, wherein Ra is hydrogen or an alkyl group containing one to six saturated straight or branched chain carbon atoms (C1-6 alkyl), -O-Ra, -halo, wherein halo is CI, F, Br, or I, S. -NRbRc, where Rb and Rc, independently, are H, C1- 6 alkyl, or -CH 2 -aryl, -NO 2 -C(=O)Ra, -CN, 30 -perfluoroRa, such as trifluoromethyl, -N-C(=0)Ra, -(CH 2 )n-NRbRc, wherein n is an integer from 1 to 6, Y:\Mary\NKI NO DELETE MR\41919-00.doc 64 a 5- or 6-membered heterocyclic ring, either aliphatic or aromatic, containing one or more of O, N, or S, optionally substituted, such as morpholino, and -S-aryl, wherein aryl is a 5- or 6-membered aromatic ring, optionally substituted; and R 4 R 5 and R 6 independently, are selected from the group consisting of hydrogen, -Ra, -O-Ra, -halo, -NRbRc, -NO 2 -C(=O)Ra, -CN -perfluoroRa, -N-C(=O)Ra, -(CH 2 )n-NRbRc, and -a 5- or 6-membered heterocyclic ring, aliphatic or aromatic, containing one or more of O, N, or S, optionally substituted, -S-aryl, or wherein R 4 and R 5 are taken together to form a 5- or 6-membered aromatic ring, optionally containing one or more of O, N, or S in the ring, optionally substituted.
10. The method according to claim 7 or 9, wherein the ligand is ICAM-1 or ICAM-3. DATED: 26 April 2004 PHILLIPS ORMONDE FITZPATRICK Attorneys for: 30 ICOS CORPORATION l *oooo *o* Y:\Mary\NKI NO DELETE MR\41919-00.doc
AU41919/00A 1999-04-02 2000-04-03 Inhibitors of LFA-1 binding to ICAMs and uses thereof Ceased AU774054B2 (en)

Applications Claiming Priority (3)

Application Number Priority Date Filing Date Title
US28532599A 1999-04-02 1999-04-02
US09/285325 1999-04-02
PCT/US2000/008840 WO2000059878A2 (en) 1999-04-02 2000-04-03 INHIBITORS OF LFA-1 BINDING TO ICAMs AND USES THEREOF

Related Child Applications (1)

Application Number Title Priority Date Filing Date
AU2004205210A Division AU2004205210A1 (en) 1999-04-02 2004-08-24 Inhibitors of LFA-1 binding to ICAMs and uses thereof

Publications (2)

Publication Number Publication Date
AU4191900A AU4191900A (en) 2000-10-23
AU774054B2 true AU774054B2 (en) 2004-06-17

Family

ID=23093752

Family Applications (2)

Application Number Title Priority Date Filing Date
AU41919/00A Ceased AU774054B2 (en) 1999-04-02 2000-04-03 Inhibitors of LFA-1 binding to ICAMs and uses thereof
AU2004205210A Abandoned AU2004205210A1 (en) 1999-04-02 2004-08-24 Inhibitors of LFA-1 binding to ICAMs and uses thereof

Family Applications After (1)

Application Number Title Priority Date Filing Date
AU2004205210A Abandoned AU2004205210A1 (en) 1999-04-02 2004-08-24 Inhibitors of LFA-1 binding to ICAMs and uses thereof

Country Status (21)

Country Link
EP (1) EP1165504A2 (en)
JP (1) JP2002541141A (en)
KR (1) KR20020003559A (en)
CN (2) CN1351588A (en)
AU (2) AU774054B2 (en)
BG (1) BG106019A (en)
BR (1) BR0009421A (en)
CA (1) CA2369005A1 (en)
CZ (1) CZ20013524A3 (en)
EA (1) EA200101017A1 (en)
EE (1) EE200100514A (en)
HR (1) HRP20010777A2 (en)
HU (1) HUP0201904A3 (en)
IL (1) IL145528A0 (en)
IS (1) IS6096A (en)
MX (1) MXPA01009915A (en)
NO (1) NO20014768L (en)
NZ (1) NZ515238A (en)
PL (1) PL351323A1 (en)
SK (1) SK14022001A3 (en)
WO (1) WO2000059878A2 (en)

Families Citing this family (19)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6515124B2 (en) * 2000-02-09 2003-02-04 Hoffman-La Roche Inc. Dehydroamino acids
EP1294704A1 (en) 2000-06-29 2003-03-26 Abbott Laboratories Aryl phenylheterocyclyl sulfide derivatives and their use as cell adhesion-inhibiting anti-inflammatory and immune-suppressive agents
US7153944B2 (en) * 2000-07-31 2006-12-26 The General Hospital Corporation High affinity integrin polypeptides and uses thereof
AU2001286403A1 (en) * 2000-07-31 2002-02-13 The General Hospital Corporation High affinity integrin polypeptides and uses thereof
US7064180B2 (en) 2000-07-31 2006-06-20 The General Hospital Corporation High affinity integrin polypeptides and uses thereof
US7323552B2 (en) 2000-07-31 2008-01-29 The General Hospital Corporation Variant integrin polypeptides and uses thereof
UA81749C2 (en) 2001-10-04 2008-02-11 Х. Луннбек А/С Derivated of phenylpiperazine as serotonin reuptake inhibitorS
EP1539744A4 (en) 2002-07-11 2007-06-06 Vicuron Pharm Inc N-hydroxyamide derivatives possessing antibacterial activity
WO2004087156A1 (en) 2003-04-04 2004-10-14 H. Lundbeck A/S 4-(2-phenylsulfanyl-phenyl)-piperidine derivatives as serotonin reuptake inhibitors
PL1701940T3 (en) 2003-12-23 2008-11-28 H Lundbeck As 2-(1h-indolylsulfanyl)-benzyl amine derivatives as ssri
CA2557794A1 (en) 2004-03-15 2005-10-06 Eli Lilly And Company Opioid receptor antagonists
AR052308A1 (en) 2004-07-16 2007-03-14 Lundbeck & Co As H DERIVATIVES OF 2- (1H-INDOLILSULFANIL) -ARILAMINE AND A PHARMACEUTICAL COMPOSITION CONTAINING THE COMPOUND
US7629473B2 (en) 2005-06-17 2009-12-08 H. Lundbeck A/S 2-(1H-indolylsulfanyl)-aryl amine derivatives
AR054393A1 (en) 2005-06-17 2007-06-20 Lundbeck & Co As H DERIVATIVES OF BENZO (B) FURANO AND BENZO (B) THIOPHEN, PHARMACEUTICAL COMPOSITIONS CONTAINING THEM AND THEIR USE IN THE MANUFACTURE OF A MEDICINAL PRODUCT FOR THE TREATMENT OF DISEASES MEDIATED BY THE INHIBITION OF THE REINFORCEMENT OF AMINA BOSS NEUTRANTS.
JP2007008877A (en) * 2005-06-30 2007-01-18 Sato Pharmaceutical Co Ltd Pharmaceutical composition with 2-pyridone derivative as active ingredient
EA200801120A1 (en) * 2005-10-18 2008-08-29 Сигма-Тау Индустрие Фармасьютике Риуните С.П.А. NAPHTILE DERIVATIVES AS AN INHIBITORS AGTAINING BETA-AMYLOID
US8436025B2 (en) * 2008-09-19 2013-05-07 CompleGen Partners, Inc. Compounds and methods for PKC theta inhibition
US9884046B1 (en) * 2017-06-26 2018-02-06 Macau University Of Science And Technology Method of treating lung cancer
NL2026714B1 (en) 2020-10-20 2022-06-16 Lumicks Ca Holding B V Improved detection of lymphocyte - target cell interaction

Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
GB1156005A (en) * 1965-07-20 1969-06-25 Bayer Ag Process for the production of Aminophenyl Thioethers
US3948893A (en) * 1973-03-20 1976-04-06 Bayer Aktiengesellschaft 1-Phenyl-substituted 1,3,5-triazines
US4275077A (en) * 1979-01-15 1981-06-23 Celamerck Gmbh & Co. Kg Insecticidal acyl urea derivatives

Family Cites Families (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US3576872A (en) * 1968-05-03 1971-04-27 Exxon Research Engineering Co Herbicidal s-aryl arylamides
US4973599A (en) * 1989-03-14 1990-11-27 Hoffman-La Roche Inc. Phenylthioheterocyclic derivatives
GB9403408D0 (en) * 1994-02-23 1994-04-13 Wellcome Found Therapeutic benzonitriles
TW530047B (en) 1994-06-08 2003-05-01 Pfizer Corticotropin releasing factor antagonists
US6110922A (en) * 1998-12-29 2000-08-29 Abbott Laboratories Cell adhesion-inhibiting antiinflammatory and immune-suppressive compounds

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
GB1156005A (en) * 1965-07-20 1969-06-25 Bayer Ag Process for the production of Aminophenyl Thioethers
US3948893A (en) * 1973-03-20 1976-04-06 Bayer Aktiengesellschaft 1-Phenyl-substituted 1,3,5-triazines
US4275077A (en) * 1979-01-15 1981-06-23 Celamerck Gmbh & Co. Kg Insecticidal acyl urea derivatives

Also Published As

Publication number Publication date
HUP0201904A2 (en) 2002-09-28
NO20014768D0 (en) 2001-10-01
CZ20013524A3 (en) 2002-06-12
EE200100514A (en) 2002-12-16
HRP20010777A2 (en) 2002-12-31
IL145528A0 (en) 2002-06-30
HUP0201904A3 (en) 2003-04-28
NO20014768L (en) 2001-11-29
WO2000059878A3 (en) 2001-08-09
MXPA01009915A (en) 2003-08-20
EP1165504A2 (en) 2002-01-02
PL351323A1 (en) 2003-04-07
AU2004205210A1 (en) 2004-09-23
AU4191900A (en) 2000-10-23
BR0009421A (en) 2002-03-26
CN1351588A (en) 2002-05-29
BG106019A (en) 2002-06-28
EA200101017A1 (en) 2002-04-25
SK14022001A3 (en) 2002-03-05
CA2369005A1 (en) 2000-10-12
WO2000059878A2 (en) 2000-10-12
KR20020003559A (en) 2002-01-12
CN1721401A (en) 2006-01-18
JP2002541141A (en) 2002-12-03
IS6096A (en) 2001-09-28
NZ515238A (en) 2005-02-25

Similar Documents

Publication Publication Date Title
AU774054B2 (en) Inhibitors of LFA-1 binding to ICAMs and uses thereof
US6787542B2 (en) Aryl phenylheterocyclyl sulfide derivatives and their use as cell adhesion-inhibiting anti-inflammatory and immune-suppressive agents
EP1165505B1 (en) Cell adhesion-inhibiting antiinflammatory and immune-suppressive compounds
US6110922A (en) Cell adhesion-inhibiting antiinflammatory and immune-suppressive compounds
DE69627377T2 (en) INHIBITORS OF PROTEIN ISOPRENYL TRANSFERASE
JPH06501706A (en) Aminosulfonyl carbamate
CZ225495A3 (en) Derivative of 4-aminopyridine, process of its preparation and pharmaceutical composition containing thereof
EP0955296A2 (en) Tertiary alkyl functionalized piperazine derivatives
US5977110A (en) Substituted cyclohexylamines as central nervous systems agents
AU2002346724A1 (en) Human adam-10 inhibitors
US6166050A (en) 3,4-diamino-3-cyclobutene-1,2-dione derivatives which inhibit leukocyte adhesion mediated by VLA-4
AU747815B2 (en) Cyclic amide compounds
EP0749971B1 (en) Thiopyran derivatives
EP1771432B1 (en) Non-imidazole heterocyclic compounds as histamine h3 receptor modulators
US4665173A (en) 2-acetyl- and 2-propionylpyridine selenosemicarbazones
DE60011241T2 (en) NEW DIAZOL DERIVATIVES AND THEIR USE AS SEROTONERGIC ACTIVE SUBSTANCES
US5475012A (en) N-cyanoamidine derivatives as anti-influenza agents
US4014884A (en) Basically substituted 3,4-dihydro-2H-isoquinolin-1-thiones
US5166157A (en) Naphthyl piperazines useful as 5-HT1A receptor ligands
US4558044A (en) 1,2,4-Benzothiadiazines
EP0105732A2 (en) Histamine H2 receptor antagonists of the benzisothiazole amino and benzthiadiazine amino series
CA1312869C (en) Pyridine derivatives
HK1040985B (en) Cell adhesion-inhibiting antiinflammatory and immune-suppressive compounds

Legal Events

Date Code Title Description
FGA Letters patent sealed or granted (standard patent)