Title of Invention	"HETEROARYL COMPOUNDS WHICH INHIBIT LEUKOCYTE ADHESION MEDIATED BY ALPHA-4 INTEGRINS"
Abstract	Disclosed are compounds which bind 4 integrins, where the 4integrin is preferably VLA-4. Certain of these compounds also inhibit leukocyte adhesion and, in particular, leukocyte adhesion mediated by 4integrins, where the 4 integrin is preferably VLA-4. Such compounds are useful in the treatment of inflammatory diseases in a mammalian patient, e.g., human, such as asthma, Alzheimer's disease, atherosclerosis, AIDS dementia, diabetes, inflammatory bowel disease, rheumatoid arthritis, tissue transplantation, tumor metastasis and myocardial ischemia. The compounds can also be administered for the treatment of inflammatory brain diseases such as multiple sclerosis. (FIG). nil

Title of Invention

"HETEROARYL COMPOUNDS WHICH INHIBIT LEUKOCYTE ADHESION MEDIATED BY ALPHA-4 INTEGRINS"

Abstract

Disclosed are compounds which bind 4 integrins, where the 4integrin is preferably VLA-4. Certain of these compounds also inhibit leukocyte adhesion and, in particular, leukocyte adhesion mediated by 4integrins, where the 4 integrin is preferably VLA-4. Such compounds are useful in the treatment of inflammatory diseases in a mammalian patient, e.g., human, such as asthma, Alzheimer's disease, atherosclerosis, AIDS dementia, diabetes, inflammatory bowel disease, rheumatoid arthritis, tissue transplantation, tumor metastasis and myocardial ischemia. The compounds can also be administered for the treatment of inflammatory brain diseases such as multiple sclerosis. (FIG). nil

Full Text	HETEROARYL COMPOUNDS WHICH INHIBIT LEUKOCYTE ADHESION MEDIATED BY a4 INTEGRINS BACKGROUND OF THE INVENTION Field of the Invention [0001] This invention relates to compounds which inhibit leukocyte adhesion and, in particular, leukocyte adhesion mediated by a4 integrins, where the a4 integrin is preferably VLA-4. References [0002] The following publications, patents and patent applications are cited in this application as superscript numbers: [0003] 1 Hemler and Takada, European Patent Application Publication No. 330,506, published August 30, 1989 [0004] 2 Elices, et al., Cell, 60.577-584 (1990) [0005]3 Springer, Nature, 246:425-434 (1990) [0006] 4 Osborn, Cell, 62:3-6 (1990) [0007] 5 Vedder, et al., Surgery, 106:509 (1989) [0008] 6 Pretolani, et al., J. Exp. Med., 180:795 (1994) [0009] 7 Abraham, et al., J. Clin. Invest., 21:776 (1994) [0010] 8 Mulligan, et al., J. Immunology, 150_:2407 (1993) [0011] 9 Cybulsky, et al., Science, 25.1:788 (1991) [0012] l0 Li, et al., Arterioscler. Thromb., 12:197 (1993) [0013] 11 Sasseville, et al., Am. J. Path., 144:27 (1994) [0014] l2 Yang, et al., Proc. Nat. Acad. Science (USA), 90: 10494 (1993) [0015] 13 Burkly, et al., Diabetes, 42:529 (1994) [0016] H Baron, et al., J. Clin. Invest., 92.: 1700 (1994) [0017] 15 Hamann, et al., /. Immunology, 152:3238 (1994) [0018] l6 Yednock, et al.. Nature, 256:63 (1992) [0019] 17 Baron, et al., J. Exp. Med., 121:57 (1993) [0020] 18 van Dinther-Janssen, et al., J. Immunology, 142:4207 (1991) [0021] 19 van Dinther-Janssen, et al., Annals. RheumaticDis., 52:672 (1993) [0022]20 Elices, et al., /. Clin. Invest., 22:405 (1994) [0023] n Postigo, et al., /. Clin. Invest., 89_: 1445 (1991) [0024] 22 Paul, et al., Transpl. Proceed., 25:813 (1993) [0025] B Okarhara, et al., Can. Res., 54:3233 (1994) [0026]24 Paavonen, et al., Int. J. Can., 58:298 (1994) [0027] M Schadendorf, et al., /. Path., 120:429 (1993) [0028] 2S Bao, et al., Diff., 52:239 (1993) [0029]27 Lauri, et al., British J. Cancer, 68:862 (1993) [0030] M Kawaguchi, et al., Japanese J. Cancer Res., 82:1304 (1992) [0031]29 Konradi, et al., PCT/US00/01686, filed, January 21, 2000. [0032] All of the above publications are herein incorporated by reference in their entirety to the same extent as if each individual publication was specifically and individually indicated to be incorporated by reference in its entirety. State of the Art [0033] VLA-4 (also referred to as a4ß1 integrin and CD49d/CD29), first identified by Hemler and Takada1, is a member of the ß1 integrin family of cell surface receptors, each of which comprises two subunits, an a chain and a ß chain. VLA-4 contains an a4 chain and a ß1 chain. There are at least nine ß1 integrins, all sharing the same ß1 chain and each having a distinct a chain. These nine receptors all bind a different complement of the various cell matrix molecules, such as fibronectin, laminin, and collagen. VLA-4, for example, binds to fibronectin. VLA-4 also binds non-matrix molecules that are expressed by endothelial and other cells. These non-matrix molecules include VCAM-1, which is expressed on cytokine-activated human umbilical vein endothelial cells in culture. Distinct eß1topes of VLA- 4 are responsible for the fibronectin and VCAM-1 binding activities and each activity has been shown to be inhibited independently.2 [0034] Intercellular adhesion mediated by VLA-4 and other cell surface receptors is associated with a number of inflammatory responses. At the site of an injury or other inflammatory stimulus, activated vascular endothelial cells express molecules that are adhesive for leukocytes. The mechanics of leukocyte adhesion to endothelial cells involves, in part, the recognition and binding of cell surface receptors on leukocytes to the corresponding cell surface molecules on endothelial cells. Once bound, the leukocytes migrate across the blood vessel wall to enter the injured site and release chemical mediators to combat infection. For reviews of adhesion receptors of the immune system, see, for example, Springer3 and Osborn.4 [0035] Inflammatory brain disorders, such as experimental autoimmune encephalomyelitis (EAE), multiple sclerosis (MS) and meningitis, are examples of central nervous system disorders in which the endothelium/leukocyte adhesion mechanism results in destruction to otherwise healthy brain tissue. Large numbers of leukocytes migrate across the blood brain barrier (BBB) in subjects with these inflammatory diseases. The leukocytes release toxic mediators that cause extensive tissue damage resulting in impaired nerve conduction and paralysis. [0036] In other organ systems, tissue damage also occurs via an adhesion mechanism resulting in migration or activation of leukocytes. For example, it has been shown that the initial insult following myocardial ischemia to heart tissue can be further complicated by leukocyte entry to the injured tissue causing still further insult (Vedder et al.).5 Other inflammatory or medical conditions mediated by an adhesion mechanism include, by way of example, asthma,6-8 Alzheimer"s disease, atherosclerosis,9-10 AIDS dementia,11 diabetes12-14 (including acute juvenile onset diabetes), inflammatory bowel disease15 (including ulcerative colitis and Crohn"s disease), multiple sclerosis,16-17 rheumatoid arthritis,18-21 tissue transplantation,22 tumor metastasis,23-28 meningitis, encephalitis, stroke, and other cerebral traumas, nephritis, retinitis, atoß1c dermatitis, psoriasis, myocardial ischemia and acute leukocyte-mediated lung injury such as that which occurs in adult resß1ratory distress syndrome. [0037] Substituted aminopyrimidines, as a class, have been disclosed as inhibiting binding of VLA-4 to VCAM-1 and, accordingly, exhibit anti- inflammatory properties.29 While these compounds possess antagonist properties to such binding, enhanced bioavailabiiity of these compounds would augment their efficacy. SUMMARY OF THE INVENTION [0038] This invention is directed to the discovery that certain N-[2-N",N"- diethylamino-5-aminosuifonylphenylpyrimidin-4-yl]-p-carbomyloxy- phenylalanine compounds possess unexpectedly superior bioavailability, as measured by their AUC, as compared to other substituted aminopyrimidine compounds previously disclosed. [0039] In one of its composition aspects, this invention is directed to a compound of formula (I): wherein each X is independently fluoro, chloro or bromo; p is 0 or an integer from 1-3; R1 is selected from the group consisting of methyl and ethyl; R2 is selected from the group consisting of lower alkyl, lower alkenyl, and lower alkylenecycloalkyl; and pharmaceuticaHy acceptable salts thereof. [0040] In a preferred embodiment, this invention provides compounds of Formula II: wherein each X is independently selected from the group consisting of fluoro and chloro, m is an integer equal to 1 or 2; R2 is selected from the group consisting of lower alkyl, lower alkenyl, and lower alkylenecycloalkyl; and pharmaceutically acceptable salts thereof. [0041] In a particularly preferred embodiment, this invention provides compounds of Formula III: wherein each X is independently fluoro or chloro; n is zero or one; R2 is -CH2-R" where R1 is selected from the group consisting of hydrogen, methyl or -CH=CH2; and pharmaceutically acceptable salts thereof. [0042] In another particularly preferred embodiment, this invention provides compounds of Formula (IV): wherein each X is independently fluoro, chloro or bromo; p is 0 or an integer from 1-3; R1 is selected from the group consisting of methyl and ethyl; R2 is lower alkynyl; and pharmaceutically acceptable salts thereof. [0043] In yet another particularly preferred embodiment, this invention provides compounds of Formula V: wherein each X is independently selected from the group consisting of fluoro and chloro, m is an integer equal to 1 or 2; R2 is lower alkynyl; and pharmaceutically acceptable salts thereof. [0044] In still another particularly preferred embodiment, this invention provides compounds of Formula VI: wherein each X is independently fluoro or chloro; n is zero or one; R2 is lower alkynyl; and pharmaceutically acceptable salts thereof. R2 is preferably propargyl in any of one of Formula IV, V or VI.. [0045] N-[2-N\N"-diethylamino-5-aminosulfonylphenylpyrimidin-4-yl]-p- carbomyloxyphenylalanine compounds within the scope of this invention include those set forth in Table.I as follows.. [0046] Specific compounds within the scope of this invention include the following. As used below, these compounds are named based on proß1onic acid derivatives but, alternatively, these compounds could have been named based on N-[2-N\N"-diethylarnino-5-aminosulfoaylphenylpyrimidin-4-yl]-p- carbomyIoxy-phenylalanine derivatives. [0047] 2-{2-diethylamino-5-[(4-chlorobenzenesulfonyl)methylamino]- pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)proß1onicacid; [0048] 2-{2-diethylamino-5-[(4-fluor"obenzenesulfonyl)methylamino]- pyrirnidin-4-yIamino}-3-(4-dimethylcarbamoyloxyphenyl)proß1onicacid; [0049] 2-{2-diethylamino-5-[(3,4-difluorobenzenesulfonyl)methylamino]- pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)proß1onicacid; [0050] 2-{2-diethylamino-5-[(3,4-dichlorobenzenesulfonyl)methylamino]- pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)proß1onic acid; [0051] 2-{2-diethylamino-5-[(benzenesulfonyl)methylamino]- pyrinudin-4-ylamino}-3-(4-dimethylcarbanioyloxyphenyl)propionic acid; [0052] 2-{2-diediylamino-5-[(2-fluorobeiizenesulfonyl)methylamino]- pyrimidin-4-ylamino}-3-(4"dimethylcarbamoyloxyphenyl)propionicacid; [0053] 2-{2-diethylamino-5-[(3-fluorobenzenesulfonyl)methylamino]- pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)propionicacid; [0054] 2-{2-diethylamino-5-[(4-fluorobenzenesulfonyl)isopropylamino]- pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)propionicacid; [0055] 2-{2-diethylammo-5-[(4-fluorobenzenesulfonyl)ethylamino]- pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)propionicacid; [0056] 2-{2-diethylamino-5-[(3,4-difluorobenzenesulfoayl)isopropylamino]- pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)propionicacid; [0057] 2-{2-diethylamino-5-[(4-chlorobenzenesulfonyI)isopropylamino]- pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyplienyl)propionicacid; [0058] 2-{2-diethylamino-5-[(3,4-difluorobenzenesulfonyl)ethylamino]- pyrimidin^ylamino}-3-(4-dimethylcarbamoyloxyphenyl)propionicacid; [0059] 2-{2-diethylamino-5-[(4-chlorobenzenesulfonyl)ethylamino]- pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)propionicacid; [0060] 2-{2-diethylamino-5-[(4-fluorobenzenesulfonyl)cylclopropylmethyl- amino]pyrimidin-4-ylamino}-3-(4-dimethylcarbatnoyloxyphenyl)propionic acid; [0061] 2-{2-diethylamino-5-[(3,5-difiuorobenzenes\iifoDyl)methylamino]- pyrimidin^-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)propionicacid; [0062] 2-{2-diethylamino-5-[(3,5-difluorobenzenesulfonyl)ethylamino]- pyrimidin-4-yIamino}-3-(4-dimethylcarbamoyloxypb.enyl)propionicacid; [0063] 2-{2-diethylaiiiino-5-[(2,4-difluorobenzeaesulfonyI)methylamino]- pyrimidia-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)propionicacid; [0064] 2-{2-diethylamino-5-[(2,4-difluorobenzenesulfonyl)ethylamino]- pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)proß1onicacid; [0065] 2-{2-diethylamino-5-[(3,5-dichlorobenzenesulfonyl)methylamino]- pyrimidm-4-ylamino}-3-(4-dimethylcarbamoyloxyj)henyl)proß1onicacid; [0066] 2-{2-diethylamino-5-[(3,5-dichlorobenzenesulfonyl)ethylamino]- pyrimidin-4-ylamino} -3-(4-dimethylcarbamoyloxyphenyl)proß1onic acid; [0067] 2-{2-diethylamino-5-[(4-fluorobenzenesulfonyl)-n-propylamino]- pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxypheiiyl)proß1onicacid; [0068] 2-{2-diethylamino-5-[(4-fluorobenzenesulfonyl)allylamino]- pyrunidin-4-ylamino}-3-(4-dimethylcarbamoyloxypheiiyl)proß1oiiicacid; [0069] 2-{2-diethylamino-5-[(4-fluorobenzenesulfonyl)isobotylamino]- pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)proß1onicacid; [0070] 2-{2-diethylamino-5-[(4-fluorobenz;enesulfonyl)-n-butylamino]- pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)proß1onicacid; [0071] 2-{2-diethylamino-5-[(2,6-difluorobenzenesulfonyl)methylamino]- pyrimidin-4-ylamino} -3-(4-dimethylcarbamoyloxyphenyl)proß1onic acid; [0072] 2-{2-diethylamino-5-[(2,3-difluorobenzenesulfonyl)ethylamino]- pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)proß1onicacid; [0073] 2-{2-Diethylamino-5-[(4-fluorobenzenesulfonyl)propargylamino] pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)proß1onicacid; [0074] 2-{2-Diethylamino-5-[(2,4-difluorobenzenesulfonyl)propargylamino] pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)proß1onicacid; [0075] 2-{2-Diethylamino-5-[(4-fluorobenzenesulfonyl)-(2-trisfiuoroethyl)- amino]pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)proß1onic acid; and pharmaceutically acceptable salts thereof. [0076] In another aspect, this invention provides pharmaceutical compositions comprising a pharmaceutically acceptable carrier and a therapeutically effective amount of the compounds defined herein. [0077] In one of its method aspects, this invention is directed to a method for treating a disease mediated at least in part by a4 integrin, preferably VLA-4, in a patient, which method comprises administering a pharmaceutical composition comprising a pharmaceutically acceptable carrier and a therapeutically effective amount of a compound of this invention. [0078] The compounds and pharmaceutical compositions of this invention are useful for treating disease conditions mediated at least in part by a4 integrins, where the a4 integrin is preferably VLA-4 or leucocyte adhesion. Such disease conditions include, by way of example, asthma, Alzheimer"s disease, atherosclerosis, AIDS dementia, diabetes (including acute juvenile onset diabetes), inflammatory bowel disease (including ulcerative colitis and Crohn"s disease), multiple sclerosis, rheumatoid arthritis, tissue transplantation, tumor metastasis, meningitis, encephalitis, stroke, and other cerebral traumas, nephritis, retinitis, atoß1c dermatitis, psoriasis, myocardial ischemia and acute leukocyte-mediated lung injury such as that which occurs in adult resß1ratory distress syndrome. [0079] Other disease conditions include, but are not limited to, inflammatory conditions such as erythema nodosum, allergic conjunctivitis, optic neuritis, uveitis, allergic rhinitis Anky losing spondylitis, psoriatic arthritis, vasculitis, Reiter"s syndrome, systemic lupus erythematosus, progressive systemic sclerosis, polymyositis, dermatomyositis, Wegner"s granulomatosis, aortitis, sarcoidosis, lymphocytopenia, temporal arteritis, pericarditis, myocarditis, congestive heart failure, polyarteritis nodosa, hypersensitivity syndromes, allergy, hypereosinophilic syndromes, Churg-Strauss syndrome, chronic obstructive pulmonary disease, hypersensitivity pneumonitis, chronic active hepatitis, interstitial cystitis, autoimmune endocrine failure, primary biliary cirrhosis, autoimmune aplastic anemia, chronic persistent hepatitis and thyroiditis. [0080] In a preferred embodiment, the disease condition mediated by a4 integrin is an inflammatory disease. DETAILED DESCRIPTION OF THE INVENTION [0081] As above, this invention relates to compounds which inhibit leukocyte adhesion and, in particular, leukocyte adhesion mediated at least in part by a4 integrins, preferably VLA-4. However, prior to describingjhis invention in further detail, the following terms will first be defined. Definitions} [0082] Unless otherwise stated, the following terms used in the specification and claims have the meanings given below: [0083] As used herein, "lower alkyl" refers to monovalent alkyl groups having from 1 to 5 carbon atoms including straight and branched chain alkyl groups. This term is exemplified by groups such as methyl, ethyl, iso- propyl, n-propyl, n-butyl, iso-butyl, sec-butyl, r-butyl, n-pentyl and the like. "Lower alkyl" may be optionally substituted with a halogen, such as chloro, fluoro, bromo and the like. [0084] The term "lower alkylene" refers to divalent alkylene groups of from 1 to 4 carbon atoms including straight and branched chain alkylene groups. This term is exemplied by groups such as methylene, ethylene, n-propylene, iso-propyiene (-CH2CH(CH3)- and -CH(CH3)CHr) and the like. [0085] The term "lower alkynyl" refers to an alkynyl group preferably having from 2 to 6 carbon atoms and having at least 1 site and preferably only 1 site of alkynyl unsaturation (i.e., -OC). This term is exemplified by groups such as acetyl (-OCH), propargyl (-CH2-OCH), 3-butynyl (- CH2CH2OCH3) and the like. [0086] The term "lower cycloalkyl" refers to cyclic alkyl groups of from 3 to 6 carbon atoms having a single cyclic ring including, by way of example, cyclopropyl, cyclobutyl, cyclopentyl and cyclohexyl. [0087] The term "lower alkylenecycloalkyl" refers to the group consisting of a lower alkylene-lower cycloalkyl, as defined herein. Such groups are exemplified by methylenecyclopropyl (-CH2-cyclopropyl), ethylenecyclopropyl and the like. [0088] "Pharmaceutically acceptable carrier" means a carrier that is useful in preparing a pharmaceutical composition that is generally safe, non-toxic and neither biologically nor otherwise undesirable, and includes a carrier that is acceptable for veterinary use as well as human pharmaceutical use. "A pharmaceutically acceptable carrier" as used in the specification and claims includes both one and more than one such carrier. [0089] "Treating" or "treatment" of a disease includes: (1) preventing the disease, i.e., causing the clinical symptoms of the disease not to develop in a mammal that may be exposed to or predisposed to the disease but does not yet experience or display symptoms of the disease, (2) inhibiting the disease, i.e., arresting or reducing the development of the disease or its clinical symptoms, or (3) relieving the disease, i.e., causing regression of the disease or its clinical symptoms. [0090] A "therapeutically effective amount" means the amount of a compound that, when administered to a mammal for treating a disease, is sufficient to effect such treatment for the disease. The "therapeutically effective amount" will vary depending on the compound, the disease and its severity and the age, weight, etc., of the mammal to be treated. [0091] "Pharmaceutically acceptable salt" refers to pharmaceutically acceptable salts of a compound of Formula I which salts are derived from a variety of organic and inorganic counter ions well known in the art and include, by way of example only, sodium, potassium, calcium, magnesium, ammonium, tetraalkylammonium, and the like; and when the molecule contains a basic functionality, salts of organic or inorganic acids, such as hydrochloride, hydrobromide, tartrate, mesylate, acetate, maleate, oxalate and the like. [0092] Integrins are a large family of homologous transmembrane linker proteins that are the principal receptors on animal cells for binding most extracellular matrix proteins, such as collagen, fibronectin, and laminin. The integrins are heterodimers comprised of an a chain and a ß chain. To date, twenty different integrin heterodimers, made from 9 different a subunits and 14 different ß subunits, have been identified. The term " a 4 integrins" refers to the class of heterodimer, enzyme-linked cell-surface receptors that contain the a 4 subunit paired with any of the ß subunits. VLA-4 is an example of an a 4 integrin, and is a heterodimer of the a 4 and ßt subunits, and is also referred to as a4 ß, integrin. Compound Preparation [0093] The compounds of this invention can be prepared from readily available starting materials using the methods and procedures set forth in the examples below. These methods and procedures outline specific reaction protocols for preparing N-[2-N",N"-diethyIainino-5-aminosulfonylphenyl- yrimidin-4-yl]-p-carbomyloxy-phenylalanine compounds. Compounds within the scope not exemplified in these examples and methods are readily prepared by appropriate substitution of starting materials which are either commercially available or well known in the art. [0094] Other procedures and reaction conditions for preparing the compounds of this invention are described in the examples set forth below. Additionally, other procedures for preparing compounds useful in certain aspects of this invention are disclosed in U.S. Patent 6,492,372; the disclosure of which is incorporated herein by reference in its entirety. Pharmaceutical Formulations [0095] When employed as Pharmaceuticals, the compounds of this invention are usually administered in the form of pharmaceutical compositions. These compositions can be administered by a variety of routes including oral, rectal, transdermal, subcutaneous, intravenous, intramuscular, and intranasal. These compositions are effective by both injectable and oral delivery. Such compositions are prepared in a manner well known in the pharmaceutical art and comprise at least one active compound. [0096] This invention also includes pharmaceutical compositions which contain, as the active ingredient, one or more of the compounds of formula I- VII above associated with pharmaceutically acceptable carriers. In making the compositions of this invention, the active ingredient is usually mixed with an exciß1ent, diluted by an exciß1ent or enclosed within such a carrier which can be in the form of a capsule, sachet, paper or other container. The excipient employed is tyß1cally an exciß1ent suitable for administration to human subjects or other mammals. When the exciß1ent serves as a diluent, it can be a solid, semi-solid, or liquid material, which acts as a vehicle, carrier or medium for the active ingredient. Thus, the compositions can be in the form of tablets, ß1lls, powders, lozenges, sachets, cachets, elixirs, suspensions, emulsions, solutions, syrups, aerosols (as a solid or in a liquid medium), ointments containing, for example, up to 10% by weight of the active compound, soft and hard gelatin capsules, suppositories, sterile injectable solutions, and sterile packaged powders. [0097] In preparing a formulation, it may be necessary to mill the active compound to provide the appropriate particle size prior to combining with the other ingredients. If the active compound is substantially insoluble, it ordinarily is milled to a particle size of less than 200 mesh. If the active compound is substantially water soluble, the particle size is normally adjusted by milling to provide a substantially uniform distribution in the formulation, e.g., about 40 mesh. [0098] Some examples of suitable exciß1ents include lactose, dextrose, sucrose, sorbitol, mannitol, starches, gum acacia, calcium phosphate, alginates, tragacanth, gelatin, calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, water, syrup, and methyl cellulose. The formulations can additionally include: lubricating agents such as talc, magnesium stearate, and mineral oil; wetting agents; emulsifying and suspending agents; preserving agents such as methyl- and propylhydroxy- benzoates; sweetening agents; and flavoring agents. The compositions of the invention can be formulated so as to provide quick, sustained or delayed release of the active ingredient after administration to the patient by employing procedures known in the art. [0099] The compositions are preferably formulated in a unit dosage form, each dosage containing from about 5 to about 100 mg, more usually about 10 to about 30 mg, of the active ingredient. The term "unit dosage forms" refers to physically discrete units suitable as unitary dosages for human subjects and other mammals, each unit containing a predetermined quantity of active material calculated to produce the desired therapeutic effect, in association with a suitable pharmaceutical exciß1ent. [00100] The active compound is effective over a wide dosage range and is generally administered in a pharmaceutically effective amount. It, will be understood, however, that the amount of the compound actually administered will be determined by a physician, in the light of the relevant circumstances, including the condition to be treated, the chosen route of administration, the actual compound administered, the age, weight, and response of the individual patient, the severity of the patient"s symptoms, and the like. [00101] For preparing solid compositions such as tablets, the principal active ingredient is mixed with a pharmaceutical exciß1ent to form a solid preformulation composition containing a homogeneous mixture of a compound of the present invention. When referring to these preformulation compositions as homogeneous, it is meant that the active ingredient is dispersed evenly throughout the composition so that the composition may be readily subdivided into equally effective unit dosage forms such as tablets, ß1lls and capsules. This solid preformulation is then subdivided into unit dosage forms of the type described above containing from, for example, 0.1 to about 500 mg of the active ingredient of the present invention. [00102] The tablets or ß1lls of the present invention may be coated or otherwise compounded to provide a dosage form affording the advantage of prolonged action. For example, the tablet or ß1ll can comprise an inner dosage and an outer dosage component, the latter being in the form of an envelope over the former. The two components can be separated by an enteric layer which serves to resist disintegration in the stomach and permit the inner component to pass intact into the duodenum or to be delayed in release. A variety of materials can be used for such enteric layers or coatings, such materials including a number of polymeric acids and mixtures of polymeric acids with such materials as shellac, cetyl alcohol, and cellulose acetate. [00103] The liquid forms in which the novel compositions of the present invention may be incorporated for administration orally or by injection include aqueous solutions suitably flavored syrups, aqueous or oil suspensions, and flavored emulsions with edible oils such as cottonseed oil, sesame oil, coconut oil, or peanut oil, as well as elixirs and similar pharmaceutical vehicles. [00104] Compositions for inhalation or insufflation include solutions and suspensions in pharmaceutically acceptable, aqueous or organic solvents, or mixtures thereof, and powders. The liquid or solid compositions may contain suitable pharmaceutically acceptable exciß1ents as described supra. Preferably the compositions are administered by the oral or nasal resß1ratory route for local or systemic effect. Compositions in preferably pharmaceutically acceptable solvents may be nebulized by use of inert gases. Nebulized solutions may be breathed directly from the nebulizing device or the nebulizing device may be attached to a face masks tent, or intermittent positive pressure breathing machine. Solution, suspension, or powder compositions may be administered, preferably orally or nasally, from devices which deliver the formulation in an appropriate manner. [00105] The following formulation examples illustrate the pharmaceutical compositions of the present invention. Formulation Example 1 [00106] Hard gelatin capsules containing the following ingredients are prepared: Quantity Ingredient (mg/capsule) Active Ingredient 30.0 Starch 305.0 Magnesium stearate 5.0 The above ingredients are mixed and filled into hard gelatin capsules in 340 mg quantities. Formulation Example 2 [00107] A tablet formula is prepared using the ingredients below: Quantity Ingredient (mg/tablet) Active Ingredient 25.0 Cellulose, microcrystalline 200.0 Colloidal silicon dioxide 10.0 Stearic acid 5.0 [00108] The components are blended and compressed to form tablets, each weighing 240 mg. Formulation Example 3 [00109] A dry powder inhaler formulation is prepared containing the following components: Ingredient Weight % Active Ingredient 5 Lactose 95 [00110] The active mixture is mixed with the lactose and the mixture is added to a dry powder inhaling appliance. Formulation Example 4 [00111] Tablets, each containing 30 mg of active ingredient, are prepared as follows: Quantity Ingredient (mg/tahlet) Active Ingredient 30.0 mg Starch 45.0 mg Microcrystalline cellulose 35.0 mg Polyvinylpyrrolidone (as 10% solution in water) 4.0 mg Sodium carboxymethyl starch 4.5 mg Magnesium stearate 0.5 mg laic l.Q mg Total 120 mg [00112] The active ingredient, starch and cellulose are passed through a No. 20 mesh U.S. sieve and mixed thoroughly. The solution of polyvinyl- pyrrolidone is mixed with the resultant powders, which are then passed through a 16 mesh U.S. sieve. The granules so produced are dried at 50° to 60°C and passed through a 16 mesh U.S. sieve. The sodium carboxymethyl starch, magnesium stearate, and talc, previously passed through a No. 30 mesh U.S. sieve, are then added to the granules which, after mixing, are compressed on a tablet machine to yield tablets each weighing 120 mg. Formulation Example 5 [00113] Capsules, each containing 40 mg of medicament are made as follows: Quantity Ingredient (mg/capsule) Active Ingredient 40.0 mg Starch 109.0 mg Magnesium stearate 1.0 mg Total 150.0 mg [00114] The active ingredient, starch, and magnesium stearate are blended, passed through a No. 20 mesh U.S. sieve, and filled into hard gelatin capsules in 150 mg quantities. Formulation Example 6 [00115] Suppositories, each containing 25 mg of active ingredient are made as follows: Ingredient Amount Active Ingredient 25 mg Saturated fatty acid glycerides to 2,000 mg [00116] The active ingredient is passed through a No. 60 mesh U.S. sieve and suspended in the saturated fatty acid glycerides previously melted using the minimum heat necessary. The mixture is then poured into a suppository mold of nominal 2.0 g capacity and allowed to cool. Formulation Example 7 [00117] Suspensions, each containing 50 mg of medicament per 5.0 mL dose are made as follows: Ingredient Amount Active Ingredient 50.0 mg Xanthan gum 4.0 mg Sodium carboxymethyl cellulose (11%) Macrocrystalline cellulose (89%) 50.0 mg Sucrose 1.75 g Sodium benzoate 10.0 mg Flavor and Color q.v. Purified water 5.0 mL [00118] The medicament, sucrose and xanthan gum are blended, passed through a No. 10 mesh U.S. sieve, and then mixed with a previously made solution of the microcrystalline cellulose and sodium carboxymethyl cellulose in water. The sodium benzoate, flavor, and color are diluted with some of the water and added with stirring. Sufficient water is then added to produce the required volume. Formulation Example 8 Quantity Ingredient (mg/capsule) Active Ingredient 15.0 mg Starch 407.0 mg Magnesium stearate 3-0 nig Total 425.0 mg [00119] The active ingredient, starch, and magnesium stearate are blended, passed through a No. 20 mesh U.S. sieve, and filled into hard gelatin capsules in 425 mg quantities. Formulation Example 9 [00120] An intravenous formulation may be prepared as follows: Tngredient Quantity Active Ingredient 250.0 mg Isotonic Saline 1000 mL Formulation Example 10 * [00121] A toß1cal formulation may be prepared as follows: Tngredient Quantity Active Ingredient 1-10 g Emulsifying Wax 30 g Liquid Paraffin 20 g White Soft Paraffin 2 to 100 g [00122] The white soft paraffin is heated until molten. The liquid paraffin and emulsifying wax are incorporated and stirred until dissolved. The active ingredient is added and stirring is continued until dispersed. The mixture is then cooled until solid. [00123] Another preferred formulation employed in the methods of the present invention employs transdermal delivery devices ("patches"). Such transdermal patches may be used to provide continuous or discontinuous infusion of the compounds of the present invention in controlled amounts. The construction and use of transdermal patches for the delivery of pharmaceutical agents is well known in the art. See, e.g., U.S. Patent 5,023,252, issued June 11, 1991, herein incorporated by reference. Such patches may be constructed for continuous, pulsatile, or on demand delivery of pharmaceutical agents. [00124] Direct or indirect placement techniques may be used when it is desirable or necessary to introduce the pharmaceutical composition to the brain. Direct techniques usually involve placement of a drug delivery catheter into the host"s ventricular system to bypass the blood-brain barrier. One such implantable delivery system used for the transport of biological factors to specific anatomical regions of the body is described in U.S. Patent 5,011,472 which is herein incorporated by reference. [00125] Indirect techniques, which are generally preferred, usually involve formulating the compositions to provide for drug latentiation by the conversion of hydrophilic drugs into liß1d-soluble drugs. Latentiation is generally achieved through blocking of the hydroxy, carbonyl, sulfate, and primary amine groups present on the drug to render the drug more lipid soluble and amenable to transportation across the blood-brain barrier. Alternatively, the delivery of hydrophilic drugs may be enhanced by intra-arterial infusion of hypertonic solutions which can transiently open the blood-brain barrier. Utility [00126] The compounds of this invention inhibit, in vivo, adhesion of leukocytes to endothelial cells mediated at least in part by a4 integrins, where the a4 integrin is preferably VLA-4, by competitive binding to a4 integrins, preferably VLA-4. Accordingly, the compounds of this invention can be used in the treatment of mammalian diseases mediated by a4 integrins, where the a4 integrin is preferably VLA-4, or leucocyte adhesion. Such diseases include inflammatory diseases in mammalian patients such as asthma, Alzheimer"s disease, atherosclerosis, AIDS dementia, diabetes (including acute juvenile onset diabetes), inflammatory bowel disease (including ulcerative colitis and Crohn"s disease), multiple sclerosis, rheumatoid arthritis, tissue transplantation, tumor metastasis, meningitis, encephalitis, stroke, and other cerebral traumas, nephritis, retinitis, atopic dermatitis, psoriasis, myocardial ischemia and acute leukocyte-mediated lung injury such as that which occurs in adult resß1ratory distress syndrome. [00127] The amount administered to the mammalian patient will vary depending upon what is being administered, the purpose of the administration, such as prophylaxis or therapy, the state of the patient, the manner of administration, and the like. In therapeutic applications, compositions are administered to a patient already suffering from a disease in an amount sufficient to cure or at least partially arrest the symptoms of the disease and its complications. An amount adequate to accomplish this is defined as "therapeutically effective dose." Amounts effective for this use will depend on the disease condition being treated as well as by the judgment of the attending clinician depending upon factors such as the severity of the inflammation, the age, weight and general condition of the patient, and the like. [00128] The compositions administered to a patient are in the form of pharmaceutical compositions described above. These compositions may be sterilized by conventional sterilization techniques, or may be sterile filtered. The resulting aqueous solutions may be packaged for use as is, or lyophilized, the lyophilized preparation being combined with a sterile aqueous carrier prior to administration. The pH of the compound preparations tyß1cally will be between 3 and 11. more preferably from 5 to 9 and most preferably from 7 to 8. It will be understood that use of certain of the foregoing exciß1ents, carriers, or stabilizers will result in the formation of pharmaceutical salts. [00129] The therapeutic dosage of the compounds of the present invention will vary according to, for example, the particular use for which the treatment is made, the manner of administration of the compound, the health and condition of the patient, and the judgment of the prescribing physician. For example, for intravenous administration, the dose will tyß1cally be in the range of about 20 µg to about 500 µg per kilogram body weight, preferably about 100 µg to about 300 µg per kilogram body weight. Suitable dosage ranges for intranasal administration are generally about 0.1 pg to 1 mg per kilogram body weight. Effective doses can be extrapolated from dose-response curves derived from in vitro or animal model test systems. [00130] The following synthetic and biological examples are offered to illustrate this invention and are not to be construed in any way as limiting the scope of this invention. Unless otherwise stated, all temperatures are in degrees Celsius. EXAMPLES [00131] In the examples below, the following abbreviations have the following meanings. If an abbreviation is not defined, it has its generally accepted meaning. AUC = Area under the curve BSA = bovine serum albumin DMAP = 4-N,N-limethylarriinopyridine DMSO = dimethylsulfoxide EDC = l-(3-dimethylaminopropyl)-3- ethylcarbodiimide hydrochloride EDTA = Ethylenediamine tetraacetic acid EtOAc = ethyl acetate EtOH = ethanol bd = broad doublet bs = broad singlet d = doublet m = multiplet t = triplet s = singlet eq or eq. = equivalent Example 1 Preparation of 2-{2-diethylamino-5-[(4-fluorohenzenesulfony1)Tnethylamino] pyriniidin-4-v1amino}-3-(;4-dimethylcarhamoy1oxypheny1)propionic acid [00132] General. Flash chromatography was performed using a Biotage Flash 75L, using 800 g KP-Sil silica cartridges (32-63/xM, 60 angstrom, 500- 550 m2/g). R1S are reported for analytical thin layer chromatography, using EM Science Silica Gel F(254) 250 µM thick plates for normal phase, and Watman MKC18F 200 µM thick plates for reverse phase. [00133] Step 1: Preparation of 2,4-Dichloro-5-nitropyrimidine. 5-Nitrouracil, was treated with phosphorous oxychloride and N,N- dimethylaniline, according to the procedure of Whittaker (J. Chem. Soc. 1951, 1565), to give the title compound, which is also available from City Chemical (West Haven, CT). [00134] Step 2: Preparation of 2-(2-diethylamino-5-nitropyrimidin-4- ylamino)-3-(4-hydroxyphenyl)proß1onic acid, r-butyl ester. To a solution of 2-amino-3-(4-hydroxyphenyl)proß1onic acid, (30.6 g, 0.129 mol) inTHF (250 mL) at -10 °C was added 2,4-DichIoro-5-nitropyrimidine (25g, 0.129 mol), keeß1ng the temperature below 5 °C during the addition. Once the addition was complete, N,N-diisopropylethylamine (33.7 mL, 0.194 mol) was added dropwise. After stirring for 1 h at -10 °C, diethylamine (66.73 mL, 0.645 mol) was added slowly, and then the reaction mixture was warmed to room temperature overnight. The reaction mixture was diluted with diethyl ether (500 mL), and the organic layer was washed with 0.2 N citric acid (3 x 150 mL), water (1 x 150 mL), and 10% K2CO3 (3 x 150 mL). The organic phase was dried (Na2SO4), filtered, and concentrated in vacuo to yield a yellow residue. The residue was purified by flash chromatography (20% EtOAc/hexanes on silica gel) to yield 37.39 g (67%) the title compound as a yellow foam. Rf=0.21 (25% EtOAc/hexanes on silica gel). [00135] Step 3: Preparation of 2-(2-diethyIammo-5-nitropyrimidin-4- ylamino)-3-(4-dimethylcarbamoyloxyphenyl)propionic acid /-butyl ester. To a solution of 2-(2-diethylamino-5-nitropyrimidin-4-ylamino)-3-(4- hydroxy-phenyl)propionic acid /-butyl ester (31.80 g, 0.074 mol) in CH2C12 (600 mL) was added DMAP (9.00 g, 0.074 mol). Afl:er 5 minutes triethylamine (10.23 mL, 0.074 mol) was added dropwise. N,N- dimethylcarbamyl chloride (13.83 mL, 0.110 mol) was added dropwise, and the reaction was heated to reflux overnight. The reaction mixture was concentrated in vacuo and taken up in EtOAc (1 L). The organic phase was washed with 0.5 M citric acid (3 x 250 mL), sat. NaHCO3 (3 x 250 mL), brine (1 x 250 mL), dried (MgSO4), filtered, and concentrated in vacuo to yield 37.0 g (99%) the title compound as a white solid. [00136] Step 4: Preparation of 2-(2-diethylamiuo-5-aminopyrimidin-4- yIamino)-3-(4-dimethylcarbamoyloxyphenyl)propionic acid /-butyl ester. A mixture of 2-(2-diemylarnino-5-nitropyrimidin-4-ylamino)-3-(4- dimethylcarbamoyl-oxyphenyl)propionic acid /-butyl ester(37.0 g, 0.073 mol) and 10% Pd/C (3.8 g, 10 wt% Pd) in EtOH (250 mL) was shaken under 60 psi hydrogen until TLC (50% EtOAc/hexanes on silica gel) showed 100% conversion to product (48 hours). The reaction mixture was then filtered through a Celite plug and concentrated in vacuo to yield 32.0 g (92%) the title compound as a violet foam. [00137] Step 5: Preparation of 2-{2-diethylamino-5-[(4- fluorobenzenesulfonyl) amino]-pyrimidin-4-ylamino}-3-(4- dimethylcarbamoyloxyphenyl) propionic acid /-butyl ester. A pyridine (120 mL) solution of 2-(2-diemylanamo-5-aminopyrimidin-4-ylarnino)-3-(4- dimethylcarbamoyloxy-phenyOpropionic acid /-butyl ester(32.0 g, 0.067 mol) was cooled to -20 °C with a dry ice/CH3CN bath. The mixture stirred for 30 minutes, and then p-fluorobenzenesulfonyl chloride (13.18 g, 0.067 mol) was added slowly. The reaction was stirred at -20 °C for 4.5 his, and then 3- dimethylaminopropyl amine (8.52 mL, 0.067 mol) was added, and then the mixture was allowed to warm to room temperature overnight. The reaction was concentrated in vacua. The residue was taken up in EtOAc (1 L), and the organic phase was washed with 0.5 M citric acid (3 x 900 mL), water (1 x 900 mL), sat. NaHCO3 (3 x 900 mL), brine (1 x 900 mL), dried (MgSO4), filtered, and concentrated in vacuo to yield a brown residue. The residue was purified by flash chromatography (50% EtOAc/hexanes on silica gel) to yield 33.04g (77%) the title compound as a yellow foam. Rf= 0.54 (3:2 EtOAc/hexanes on silica gel). [00138] Step 6: Preparation of 2-{2-diethylammo-5-[(4- fluorobenzenesulfonyl) methylamino]-pyrimidin-4-ylamino}-3-(4- dimethylcarbamoyloxyphenyl) propionic acid /-butyl ester. To a solution of 2-{2^iemylan^o-5-[(4-fluorobenzenesuIfonyl)arnino]-pyrirnidin-4- ylamino}-3-(4-dimethyl-carbamoyloxyphenyl)propionic acid /-butyl ester (33.04 g, 0.052 mol) in acetone (510 mL) was added K2CO3, (8.69 g, 0.063 mol), and the mixture was stirred for 10 min at room temperature. Dimethyl sulfate (5.95 mL, 0.063 mol) was then added slowly, and the reaction mixture was stirred overnight at room temperature. The reaction mixture was concentrated in vacuo, and the residue was taken up in EtOAc (600 mL). The organic phase was washed with water (2 x 400 mL), brine (2 x 400 mL), dried MgSO4, filtered, and concentrated in vacuo. The residue was purified by flash chromatography (2:1 hexanes/EtOAc on silica gel) to yield 28.69 g (85%) the title compound as a white solid. [00139] Step 7: Preparation of 2-{2-diethylamino-5-[(4- fluorobenzenesulfonyl) methyIamino]pyrimidin-4-ylamino}-3-(4- dimethylcarbamoyloxyphenyl) propionic acid hydrochloride. A formic acid (500 mL) solution of 2-{2-diethylamino-5-[(4-fluorobenzenesulfonyl) methylamino]-pyrimidin-4-ylamino}-3-(4-dimethylcarbainoyloxyphenyl) propionic acid /-butyl ester (28.69 g, 0.044 mol) was heated to 70 °C for 2 h, and then concentrated in vacuo. The residue was dissolved again in formic acid (500 mL), and then heated again at 70 °C for 2 h, and then concentrated again in vacuo. The residue was dissolved again in formic acid (500 mL), and then heated again at 70 °C for 1 h. The solution was reduced in volume by 90%, and then treated with 1.0 M HC1 (44 mL, 0.044 mol) and distilled water (490 mL). The resulting homogeneous solution was concentrated in vacuo, and then distilled water (100 mL) was added, and the homogenous solution was lyophilized over 14 days to yield 26.76 g (96%) the title compound, as a white solid. "H NMR (CD3OD) d 7.96-7.92 (m, 2H), 7.45-7.25 (m, 4H), 7.06-6.95 (m, 3H), 5.00-4.93 (m, 1H), 3.55-3.40 (m, 5H), 3.34-3.20 (m,, 2H), 3.15-3.05 (m, 5H), 3.07-3.00 (m, 3H), 1.22 (bs, 6H) 13C NMR (CD3OD) d 17L6, 168.3, 154.5, 144.4, 137.9, 135.1, 135.0, 134.1, 125.5, 120.6, 120.3, 39.6, 39.2, 39.1, 15.2 MS m/z 589 (MH+) Example 2 Preparation of 2-{2-diethylamino-5-[(4-chlorobenzenesulfonyl)methylamino] pyrimidin-4-ylamino}-3-(4-dimethylcarhamoyloxyphenyl)propionic acid [00140] Steps 1, 2, 3, 4, 6 and 7 were performed as for Example 1. Step 5 was performed using 4-chlorobenzenesulfonyl chloride in place of 4- fluorobenzenesulfonyl chloride. "H NMR (CD3OD) d 7.88-7.85 (m, 2H), 7.72-7.69 (m, 2H), 7.39-7.25 (m, 2H), 7.14-6.92 (m, 3H), 5.00-4.85 (m, 1H), 3.60-3.50 (m, 1H), 3.37-3.28 (m, 6H), 3.15-3.07 (m, 6H), 3.01 (bs, 3H), 1.22 (bs, 6H) 13C NMR (CD3OD)d 208.6, 145.3, 134.9, 128.8, 124.9, 124.5, 124.4, 116.3,50.2,30.4,30.0,6.0 MS m/z 605(MH+) Example 3 Preparation of 2-{2-diethyIamino-5-[(3,4-difluorohenzenesulfony1) methy1amino]pyrimidin-4-ylamino}-3-(4-dimethylcarhamoyloxyphenyl) propionic acid. [00141] Steps 1, 2, 3, 4, 6 and 7 were performed as for Example 1. Step 5 was performed using 3,4-difluorobenzenesulfonyl chloride in place of 4- fluorobenzenesulfonyl chloride. 1H NMR (CD3OD) d 7.84-7.77 (m, 1H), 7.67 (bs, 1H), 7.58-7.53 (m, 1H), 7.37-7.34 (m, 1H), 7.22-7.18 (m, 1H), 7.08-7.02 (m, 3H), 4.83-4.76 (m, 1H), 3.55-3.54 (m, 4H), 3.35-3.33 (m, 1H), 3.23-3.12 (m, 6H), 3.03-2.99 (m, 3H), 1.19(bs,6H) 13C NMR (CD3OD) d 178.3, 177.8, 163.2, 162.6, 159.3, 159.1, 155.9, 155.7, 154.3, 153.0, 152.5, 152.4, 138.4, 138.1, 134.0, 129.5, 125.3, 122.4, 122.2, 121.7, 121.4, 115.3, 59.3, 46.0, 42.4., 41.9, 40.4, 39.9, 39.2, 39.1, 15.76 MS m/z 607.2 (MH+) Example 4 Preparation of 2-{2-diethylamino-5-[(3r4-dichlorohenzenesulfonyn methy1amino]pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl) propionic acid [00142] Steps 1, 2, 3, 4, 6 and 7 were performed as for Example 1. Step 5 was performed using 3,4-dichlorobenzenesulfonyl chloride in place of 4- fluorobenzenesulfonyl chloride. *H NMR (CD3OD) d 8.00-7.98 (m, 1H), 7.83-7.74 (m, 2H), 7.37-7.34 (m, 1H), 7.21-7.20 (m, 1H), 7.10-7.02 (m, 3H), 4.85-4.83 (m, 1H), 3.55-3.53 (m, 2H), 3.35-3.33 (m, 1H), 3.21-3.12 (m, 6H), 3.04-2.99 (m, 6H), 1.19 (bs, 6H) l3C NMR (CD3OD) d 176.4, 166.2, 161.7, 161.2, 158.0, 157.8, 152.8, 151.5, 150.5, 140.2, 139.8, 139.5, 136.8, 135.8, 133.9, 132.6, 132.0, 129.8, 123.8, 113.7, 113.4, 57.8, 44.6, 40.8, 40.4, 38.7, 38.3, 37.7, 37.5, 14.1 MS m/z 639.1 (MH+) Example 5 Preparation of 2-{2-diethylamino-5-[(benzenesulfonyl)rnethylainino] pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxypheny1)propionic acid [00143] Steps 1, 2, 3, 4, 6 and 7 were performed as for Example 1. Step 5 was performed using benzenesulfonyl chloride in place of 4- fluorobenzenesulfonyl chloride. 1H NMR (CD3OD) d 8.14 (bs, 1H), (7.85-7.84 (m, 1H), 7.8-7.78 (m, 1H), 7.69-7.66 (mf 2H), 7.40-7.37 (m, 1H), 7.21-7.195 (m, 1H), 7.04-7.03 (m, 2H), 7.95-7.90 (m, 1H), 5.52 (bs, 1H), 3.54-3.53 (m, 2H), 3.36-3.33 (m, 6H), 3.13-3.12 (m, 3H), 3.01-3.00 (m, 3H), 1.20-1.17 (m, 6H) 13C NMR (CD3OD) d 165.9, 152.8, 136.7, 135.8, 132.6, 131.6, 130.2, 123.8, 44.7, 37.5, 14.0 MS m/z 571.2 (MH+) Example 6 Preparation of 2-{2-diethylamino-5-[(2-fluorobenzenesulfonyl)methylaniino] pyrimidin-4-y1amino}-3-(4-dimethylcarbamoyloxyphenyl)propionic acid [00144] Steps 1, 2, 3, 4, 6 and 7 were performed as for Example 1. Step 5 was performed using 2-fluorobenzenesulfonyl chloride in place of 4- fluorobenzenesulfonyl chloride. 1H NMR (CD3OD) d 8.31 (bs, 1H), 7.94-7.85 (m, 2H), 7.57-7.44 (m, 3H), 7.34-7.30 (m, 1H), 7.15-7.12 (m, 2H), 5.00-4.85 (1H), 3.63-3.62 (m, 4H), 3.50-3.42 (m, 1H), 3.34-3.30 (m, 4H), 3.29-3.22 (m, 4H), 3.11-3.10 (m, 2H), 1.28 (bs, 6H) 13C NMR (CD3OD) d 176.5, 166.4, 163.1, 160.4, 159.7, 157.7, 152.8, 151.5, 150.7, 138.5, 138.3, 136.7, 133.7, 132.5, 132.2, 127.1, 123.7, 119.9, 119.6, 113.4, 57.8, 44.6, 40.6, 39.0, 38.4, 37.7, 37.5, 14.1 Example 7 Preparation of 2-{2-diethylamino-5-[(3-fluorobenzenesulfonyl) methylamino]pyrimidin-4-y1amino}-3-(4-dimethyIcairhamoyIoxypheny1\|) propionic acid [00145] Steps 1, 2, 3, 4, 6 and 7 were performed as for Example 1. Step 5 was performed using 3-fluorobenzenesulfonyl chloride in place of 4- fluorobenzenesulfonyl chloride. 1H NMR (CD3OD) d 8.15-8.12 (bs, 1H), 7.72-7.68 (m, 1H), 7.63-7.60 (m, 1H), 7.53-7.52 (m, 1H), 7.38-7.35 (m, 1H), 7.21-7.20 (m, 1H), 7.10-6.99 (m, 3H), 4.87-4.86 (m, 1H), 3.54-3.53 (m, 4H), 3.35-3.34 (m, 3H), 3.15- 3.12 (m, 4H), 3.05-3.00 (m, 4H), 1.20 (bs, 6H) l3C NMR (CD3OD)d 166.1, 153.1, 136.9, 134.1, 132.8, 126.5, 124.1, 123.2, 122.9, 117.7, 117.4, 103.4, 45.0, 38.0, 14.3 MS m/z 589.2 (MH+) Example 8 Preparation of 2-{2-diethylamino-5-[(4-fluorobenzenesulfony\|) isopropylamino]pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl) propionic acid [00146] Steps 1, 2 and 3 were performed as for Example 1. Thereafter, Steps 4 and 6 were accomplished in one pot, according to the following procedure. Thereafter, Steps 5 and 7 were performed as for Example 1. 1H NMR (CD3OD) d 8.20-8.16 (m, 1H), 7.95-7.84 (m, 2H), 7.36-7.25 (m, 3H), 7.24-7.15 (m, 3H), 7.07-6.98 (m, 3H), 5.07-5.05 (m, 1H), 4.90-4.86 (m, 1H), 4.65-4.62 (m, 1H), 4.49-4.41 (m, 1H), 3.63-3.56 (m, 3H), 3.38- 3.31 (m, 2H), 3.27-3.11 (m, 2H), 3.00-2.99 (m, 3H), 1.27-1.21 (m, 6H), 1.05-0.99 (m, 6H) 13C NMR (CD3OD) d 175.8, 175.5, 169.6, 166.3, 165.9, 163.5, 163.4, 157.7, 153.0, 152.9, 152.3, 138.1, 136.4, 136.1, 133.1, 133.0, 133.0, 132.9, 132.7, 132.3, 123.8, 118.8, 118.7, 118.5, 118.4, 107.5, 57.6, 57.2, 54.7, 44.7, 38.7, 38.1, 37.6, 37.5, 23.0, 22.9, 22.2, 22.0, 14.1, 14.0 [00147] Alternative one-pot procedure for the preparation of 2-(2- diethylamino-5-isopropylaminopyrimidin-4-yl)-3-(4- dimethylcarbamoyloxypkenyl) propionic acid /-butyl ester. A mixture of 2-(2-diethylamino-5-nitropyrimidin-4-ylamino)-3-(4- dimethylcarbamoyloxyphenyl)propionic acid r-butyl ester (5.0 g, 0.010 mol), glacial acetic acid (10 drops), acetone (2.19 mL, 0.030 mol), and platinum oxide (0.250 g, 5 wt%) in EtOH (15 mL) was hydrogenated at 45 psi hydrogen until TLC (50% EtOAc/hexanes) showed 100% conversion to product (20 hours). The reaction mixture was then filtered through a Celite plug and concentrated in vacuo to yield a brown residue. The residue was purified by flash chromatography (4:1 EtOAc/hexanes) to yield 3.54 g (70%) 9 as a purple foam. Example 9 Preparation of 2-{2-diethylamino-5-[(4-fluorobenzenesulfonyl)ethylamino] pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)propionic acid [00148] Steps 1, 2, 3, 4, 5 and 7 were performed as for Example 1. Step 6 was performed using ethyl iodide in place of dimethyl sulfate. 1H NMR (CDC13) d 0.89 (t, J = 7.2, 1.8H), 1.06 (t,J = 7.1, 1.2H), 1.10- 1.30 (m, 6H), 2.97 (s, 3H), 3.05 (s, 3H), 3.10-3.90 (m, 8H), 4.82 (q, J = 5.4, 0.6H), 4.91 (q, J = 6.1, 0.4H), 6.80-7.45 (m, 8H), 7.77 (m, 2H), 12.44 (bs, 1H) MS m/z 603.3 (MH+) Example 10 Preparation of 2-{2-diethyIamino-5-[(3,4-difluorobenzenesuIfonyl) isopropylamino]pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl) propiortic acid [00149] Steps 1, 2, 3, 4, 6 and 7 were performed as for Example 8. Step 5 was performed using 3,4-difluorobenzenesulfonyl chloride in place of 4- fluorobenzenesulfonyl chloride. 1H NMR (CD3OD) d 8.20-8.19 (m, 1H), 7.84-7.78 (m, 1H), 1.10-1.64 (m, 1H), 7.54-7.48 (m, 1H), 7.39-7.31 (m, 1H), 7.20-7.17 (m, 1H), 7.05-6.96 (m, 2H), 4.91-4.89 (m, 1H), 4.70-4.68 (m, 1H), 4.48-4.41 (m, 2H), 3.60- 3.58 (m, 3H), 3.34-3.33 (m, 1H), 3.27-3.20 (m, 1H), 3.09-3.08 (m, 2H), 2.98-2.97 (m, 2H), 1.28-1.19 (m, 6H), 1.06-0.98 (m, 6H), 0.83-0.81 (m, 1H) 13C NMR (CD3OD) d 177.6, 177.2, 167.9, 164.9, 164.8, 159.2, 159.1, 155.7, 154.5, 154.4, 152.4, 152.3, 140.4, 140.3, 137.8, 134.3, 133.9, 129.3, 129.2, 125.4, 122.6, 122.5, 122.4, 122.2, 121.5, 121.2, 109.1, 59.5, 59.1, 56.7, 56.6, 46.4, 46.3, 39.6, 39.3, 39.2, 24.7, 24.5, 23.9, 23.6, 15.7, 15.6 Example 11 Preparation of 2-{2-diethylamino-5-[(4-chlorobenzenesulfonyl) isopropylamino]pyrimidin-4-ylaminQ}-3-(4-dimethylcarbamoyloxyphenyl) propionic acjd [00150] Steps 1, 2, 3, 4, 6 and 7 were performed as for Example 8. Step 5 was performed using 4-chlorobenzenesulfonyI chloride in place of 4- fluorobenzenesulfonyl chloride. 1H NMR (CD3OD) d 8.18-8.17 (m, 1H), 7.85-7.78 (m, 1H), 7.62-7.58 (m, 1H), 7.38-7.35 (m, 1H), 7.34-7.24 (m, 1H), 7.17-7.16 (m, 1H), 7.10-7.05 (m, 2H), 7.04-6.98 (m, 2H), 4.98-4.87 (m, 1H), 4.73-4.68 (m, 1H), 4.55- 4.38 (m, 2H), 3.70-3.52 (m, 3H), 3.40-3.30 (m, 1H), 3.28-3.18 (m, 1H), 3.17-3.08 (m, 2H), 3.05-2.98 (m, 2H), 1.25-1.20 (m, 6H), 1.04-0.96 (m, 6H), 0.80-0.77 (m, 1H) 13C NMR (CD3OD) d 175.7, 175.5, 166.2, 165.8, 169.6, 163.5, 163.4, 157.6, 152.9, 152.8, 138.0, 136.3, 136.1, 133.1, 133.0, 132.9, 132.7, 132.2, 123.8, 118.8, 118.6, 118.5, 118.5, 118.3, 107.5, 57.6, 57.2, 54.7, 44.6, 38.6, 38.1, 37.6, 37.5, 22.9, 22.8, 22.2, 21.9, 14.1, 13.9 Example 12 Preparation of 2-{2-diethylamino-5-[(3,4-difluorohenzenesuifonyn ethylamino]pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyIoxyphenyl) propionic acid [00151] Steps 1, 2, 3, 4, 6 and 7 were performed as for Example 9. Step 5 was performed using 3,4-difiuorobenzensulfonyl chloride in place of 4- fluorobenzenesulfonyl chloride. 1H NMR (CD3OD) d 8.15-8.14 (m, 1H), 7.80-7.75 (m, 1H), 7.73-7.62 (m, 1H), 7.60-7.49 (m, 1H), 7.30-7.18 (m, 1H), 7.16-7.00 (m, 2H), 5.58-5.50 (m, 1H), 4.90-4.83 (m, 1H), 5.78-5.70 (m, 1H), 3.85-3.75 (m, 1H), 3.65- 3.54 (m, 3H), 3.40-3.23 (m, 5H), 3.18-3.10 (m, 3H), 3.05-2.98 (m, 3H), 1.25-1.15 (m, 3H), 1.18-1.05 (t, 1.5H), 1.02-1.00 (t, 1.5H) 13C NMR (CD3OD) d 165.8, 152.7, 145.7, 136.4, 136.3, 132.5, 132.2, 127.5, 123.6, 120.7, 120.4, 81.4, 57.0, 44.3, 38.5, 38.1, 37.4, 14.9, 14.6, 14.1, 14.0 MS m/z 621.5 (MH+) Example 13 Preparation of 2-{2-diethylamino-5-[(4-ch1orobenzenesulfony1)ethylamino] pyrimidin-4-ylaTnino}-3-r4-dimethylcarhamoyloxyphenynpropionicacid [00152] Steps 1, 2, 3, 4, 6 and 7 were performed as for Example 9. Step 5 was performed using 4-chlorobenzenensulfonyl chloride in place of 4- fluorobenzenesulfonyl chloride. 1H NMR (CD3OD) d 8.15-8.14 (m, 1H), 7.84-7.79 (m, 1H), 7.67-7.61 (m, 1H), 7.37-7.33 (m, 1H), 7.22-7.18 (m, 1H), 7.14-7.13 (m, 1H), 7.06-7.00 (m, 3H), 4.80-4.75 (m, 1H), 4.18-4.10 (m, 1H), 3.65-3.30 (m, 3H), 3.28- 3.20 (m, 3H), 3.18-3.08 (m, 2H), 3.03-2.98 (m, 2H), 2.05-2.04 (m, 1H), 1.30-1.16 (m, 9H), 1.10-1.08 (t, 1.5H), 0.99-0.95 (t, 1.5H) 13C NMR (CD3OD) d 176.2, 176.1, 166.7, 162.7, 162.3, 157.6, 152.9, 142.0, 138.8, 136.5, 132.8, 132.5, 132.0, 131.8, 123.8, 111.7, 111.4,57.9, 57.8, 44.9, 38.9, 38.3, 37.8, 37.7, 15.1, 14.9, 14.3, 14.2 MS m/z 619 A (MH+) Example 14 Preparation of 2-{2-djethylamino-5-[(4-fluorobenzenesulfonyl) cylclopropy1methyIamino]pyrimidin-4-ylamino}-3-(4-dimethylcarhamoyl- oxyphenyl)propionic acid [00153] Steps 1, 2, 3, 4, 5 and 7 were performed as for Example 1. Step 6 was performed using bromomethylcyclopropane and cesium carbonate in place of dimethyl sulfate and potassium carbonate. lH NMR (CDC13) d -0.2-0.2 (m, 2.4H), 0.2-0.45 (m, 1.6H), 0.54 (m, 0.6H), 0.85 (m, 0.4H), 1.00-1.40 (m, 6H), 2.80-3.80 (m, 14H), 4.79 (q, J = 5.5, 0.6H), 4.91 (q, / = 6.3, 0.4H), 6.70-7.40 (m, 8H), 7.77 (m, 2H), 10.26 (bs, 1H) MS m/z 629.2 (MH+) Example IS Preparation of 2-{2-diethylamino-5-[(3r5-difluorobenzenesulfonyl) methylamino] pyrimidm-4-ylamino}-3-(4-dimethylcarbamoy1oxyphey1) propionic acid [00154] Steps 1, 2, 3, 4, 6 and 7 were performed as for Example 1. Step 5 was performed using 3,5-difluorobenzenesulfonyl chloride in place of 4- fluorobenzenesulfonyl chloride. 1H NMR (CD3OD) d 7.68-7.67 (m, 1H), 7.67-7.56 (m, 2H), 7.42-7.40 (m, 2H), 7.31-7.30 (m, 1H), 7.26-7.23 (m, 2H), 5.20-4.90 (m, 1H), 4.35-4.33 (m, 1H), 3.78-3.74 (m, 4H), 3.57-3.54 (2H), 3.38-3.33 (m, 2H), 3.26-3.21 (m, 2H), 2.41-2.39 (m, 2H), 2.26-2.25 (m, 2H), 1.50-1.38 (m, 6H) I3C NMR (CD3OD) d 162.5, 162.3, 159.2, 159.0, 148.0, 146.1, 132.2, 127.8, 127.7, 127.6, 118.9, 109.1, 109.0, 108.7, 108.6, 106.2, 105.8,52.5, 39.6,34.1,32.9,9.5 Example 16 Preparation of 2-{2-diethylamino-5-[(3,5-difluorobenzenesiiIfonyl) ethylamino]pyrimidin-4-ylamino}-3-(4-dimethylrarbamoyloxyphenyI) propionic acid [00155] Steps 1,2, 3, 4, 5 and 7 were performed as for example 15. Step 6 was performed using ethyl iodide in place of dimethyl sulfate. 1H NMR (CD3OD) d 7.45-7.43 (m, 1H), 7.42-7.18 (m, 2H), 7.21-7.16 (m, 2H), 7.07-7.06 (m, 1H), 7.04-6.97 (m, 2H), 5.51 (bs, 1H), 4.86-4.82 (m, 1H), 4.72-4.66 (m, 1H), 3.84-3.77 (m, 1H), 3.59-3.50 (m, 3H), 3.34-3.31 (m, 2H), 3.12-3.10 (m, 3H), 2.99-2.96 (m, 3H), 1.22-1.14 (m, 9H), 1.10- 1.05 (t, 1.5H), 0.97-0.95 (t, 1.5H) 13C NMR (CD3OD) d 159.9, 150.9, 150.1, 134.0, 130.0, 129.7, 121.2, 107.9, 86.7, 42.0, 41.9, 36.3, 35.2, 35.1, 12.8, 12.5, 11.9, 11.8, Example 17 Preparation of 2-{2-diethylamino-5-[(2.4-difluoroben7enesnlfonyl) methylamino]pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxypheny]) propionic acid [00156] Steps 1, 2, 3, 4, 6 and 7 were performed as for Example 1. Step 5 was performed using 2,4-difluorobenzenesulfonyl chloride in place of 4- fluorobenzenesulfonyl chloride. 1H NMR (CD3OD) d 8.16-8.11 (m, 1H), 7.59-7.56 (m, 2H), 7.48-7.45 (m, 2H), 7.26-7.24 (m, 3H), 5.21-5.16 (m, 1H), 3.79-3.77 (m, 4H), 3.57-3.54 (m, 3H), 3.48-3.46 (m, 2H), 3.44-3.34 (m, 3H), 3.22-3.21 (m, 3H), 1.45- 1.44 (m,6H) 13C NMR (CDC1) d 180.2, 170.3, 166.6, 150.3, 129.0, 128.9, 128.7, 125.9, 125.4, 117.5, 117.4, 116.5, 114.8, 107.7, 107.4, 95.5, 90.8, 68.0, 65.1, 55.7, 50.8, 37.6, 36.4, 31.9, 31.7, 31.6, 13.2, 9.4, 8.3, 7.8 Example 18 Preparation of 2-{2-diethylamino-5-[(2r4-difhiorobenzenesulfonyI) ethylamino] pyrimidin-4-ylamino}-3-(4-dimethyIcarhamoyloxyphenyl) propionic acid [00157] Steps 1, 2, 3, 4, 5 and 7 were performed as for Example 17. Step 6 was performed using ethyl iodide in place of dimethyl sulfate. 1H NMR (CD3OD) d 8.15 (bs, 1H), 7.91-7.76 (m, 1H), 7.32-7.30 (m, 2H), 7.20-7.19 (m, 2H), 7.04-7.00 (m, 2H), 4.84-4.83 (m, 1H), 4.74-4.67 (m, 1H), 4.14-4.07 (m, 1H), 3.92-3.82 (m, 1H), 3.51-3.49 (m, 3H), 3.34-3.31 (m, 3H), 3.12-2.99 (m, 2H), 2.98-2.97 (m, 2H), 2.03-2.02 (m, 1H), 1.26- 1.17 (m, 6H), 1.10-1.06 (t, 1.5H), 1.03-0.98 (t, 1.5H) 13C NMR (CD3OD) d 173.6, 173.3, 171.4, 167.7, 164.3, 161.2, 159.9, 159.3, 157.1, 156.7, 155.2, 152.4, 151.0, 150.3, 134.0, 133.3, 133.1, 132.9, 130.0, 123.2, 122.9, 122.8, 121.3, 121.2, 112.0, 111.8, 111.6, 111.5, 107.7, 107.2, 106.0, 105.9, 105.6, 105.2, 60.0, 54.8, 42.0, 36.5, 35.9, 35.3, 35.1, 19.3, 13.0, 12.9, 12.7, 11.9, 11.8 Example 19 Preparation of 2-{2-diethyIamino-5-[(3 f 5-dichlorobenzenesulfonyl) methylamino]pyriini(iin-4-ylamino}-3-(4-dimethylcarhamoyloxyphenyn propionic acid [00158] Steps 1, 2, 3, 4, 6 and 7 were performed as for Example 1. Step 5 was performed using 3,5-dichlorobenzenesulfonyl chloride in place of 4- fluorobenzenesulfonyl chloride. 1H NMR (CD3OD) d 7.84-7.82 (m, 1H), 7.76-7.75 (m, 3H), 7.34-7.32 (m, 1H), 7.19-7.10 (m, 1H), 7.03-7.00 (m, 2H), 5.50 (bs, 1H), 4.83-4.82 (m, 1H), 4.74-7.73 (m, 1H), 3.55-3.38 (m, 4H), 3.34-3.32 (m, 2H), 3.15-3.11 (m, 4H), 3.02-2.99 (m, 3H), 1.18-1.15 (m, 6H) 13C NMR (CD3OD)d 157.1, 155.2, 150.1, 149.7, 140.1, 135.9, 134.3, 132.9, 130.0, 129.9, 126.0, 121.2, 110.7, 55.2, 54.8, 42.0, 38.5, 38.1, 36.5,35.9,35.2,35.1, 11.9 MS m/z 639.1 (MH+) Example 20 Preparation of 2-{2-diethy1amino-5-(Y3r5-riir,h1orohenzenesulfonyl) ethyIamino]pyrimidin-4-ylamino}-3-(4-dirnethylcarbamQyloxyphenyl) propionic acid [00159] Steps 1, 2, 3, 4, 5 and 7 were performed as for Example 19. Step 6 was performed using ethyl iodide in place of dimethyl sulfate. 1H NMR (CD3OD) d 8.15 (bs, 1H), 7.84-7.84-7.79 (m, 1H), 1.16-1.1 A (m, 2H), 7.33-7.30 (m, 1H), 7.22-7.11 (m, 2H), 7.04-6.98 (m, 1H), 5.51 (bs, 1H), 4.86-4.82 (m, 1H), 4.72-4.67 (m, 1H), 3.77-3.75 (m, 1H), 3.60-3.50 (m, 3H), 3.34-3.29 (m, 2H), 3.27-3.22 (m, 2H), 3.12-3.11 (m, 2H), 2.99- 2.98 (m, 2H), 1.23-1.14 (m, 6H), 1.10-1.05 (t, 1.5H), 0.99-0.94 (t, 1.5H) l3C NMR (CD3OD)d 173.6, 173.4, 163.7, 159.9, 159.3, 157.3, 156.8, 155.2, 155.1, 152.1, 150.8, 150.2, 141.4, 141.2, 135.9, 134.0, 132.7, 130.0, 129.7, 125.8, 125.7, 121.3, 121.2, 107.9, 107.4, 54.8, 54.7, 42.0, 36.4, 35.8, 35.3, 35.1, 12.8, 12.5, 11.9, 11.8 MS m/z 653.2 (MH+) Example 21 Preparation of 2-{2-diethylarnino-5-[ propylamino] pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl) propionic acid [00160] Steps 1, 2, 3, 4, 5 and 7 were performed as for Example 1. Step 6 was performed using 1-propyl iodide in place of dimethyl sulfate. 1H NMR (CDC13) d 0.75 (m, 3H), 1.00-1.50 (m, 8H), 3.00 (s, 3H), 3.08 (s, 3H), 3.20-3.70 (m, 8H), 4.79 (q, / = 6.3, 0.6H), 4.91 (q, / = 6.6, 0.4H), 5.73 (bs, 0.6H), 5.92 (bs, 0.4H), 6.90-7.45 (m, 7H), 7.76 (m, 2H) MS m/z 617.2 (MH+) Example 22 Preparation of 2-{2-diefhylamino-5-[(4-fluorobenzenesnlfnnyna1ly1amino] pyrimidin-4-ylamino}-3-(4-dimethy1carbamoyloxyphenynpropionic acid [00161] Steps 1, 2, 3, 4, 5 and 7 were performed as for Example 1. Step 6 was performed using allyl bromide in place of dimethyl sulfate. 1H NMR (CDC13) d 1.20 (m, 6H), 2.98 (s, 3H), 3.06 (s, 3H), 3.10-4.30 (m, 8H), 4.75- 4.95 (m, 1H), 5.07 (m, 2H), 5.48 (m, 0.6H), 5.67 (m, 0.4H), 6.90-7.45 (m, 8H), 7.76 (m, 2H), 11.07 (bs, 1H) MS m/z 615.2 (MH+) Example 23 Preparation of 2-{2-diethylamino-5-[(4-fluorobenzenesulfonyl)isobotylamino] pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)propionic acid [00162] Steps 1, 2, 3, 4, 5 and 7 were performed as for Example 1. Step 6 was performed using isobutyl iodide in place of dimethyl sulfate. MS m/z 631.2 (MH+) Example 24 Preparation of 2-{2-diefhy1amino-5-[(4-fluorobenzenesuilfonyl)-n-burylamino] pyrimidin-4-ylamino}-3-(4-dimethylcarhamoyloxyphenyl)propionic acid [00163] Steps 1, 2, 3, 4, 5 and 7 were performed as for Example 1. Step 6 was performed using 1-butyl iodide in place of dimethyl sulfate. 1H NMR (CDC13) d o.82 (q, / = 7.1, 3H), 1.05-1.40 (m, 10H), 3.01 (s, 3H), 3.10 (s, 3H), 3.15-3.80 (m, 8H), 4.75 (q, J = 6.3, 0.6H), 4.91 (q, / = 5.9, 0.4H), 5.79 (d, / = 5.4, 0.6H), 5.91 (d, J = 6.6, 0.4H), 7.00-7.40 (m, 7H), 7.77 (m, 2H) Example 25 Preparation of 2-{2-djethylamino-5-[(2 r5-difluorobenzenesulfony 1) methylamino] pyrimidin-4-ylamino}-3-(4-dimethylcarhamoyIoxyphenyI) propionic acid [00164] Steps 1, 2, 3, 4, 6 and 7 were performed as for Example 1. Step 5 was performed using 2,6-difluorobenzenesulfonyl chloride in place of 4- fluorobenzenesulfonyl chloride. 1H NMR (CD3OD) d 8.38-8.37 (m, 1H), 7.99-7.95 (m, 1H), 7.55-7.54 (m, 2H), 1.50-1 A2 (m, 2H), 7.27-7.22 (m, 2H), 5.08-5.06 (m, 1H), 3.76-3.74 (m, 4H), 3.59-3.54 (m, 3H), 3.49-3.42 (m, 4H), 3.36-3.34 (m, 2H), 3.23- 3.21 (m,2H), 1.40 (bs, 6H) l3C NMR (CD3OD)d 161.4, 159.2, 155.8, 153.1, 148.1, 147.1, 133.6, 132.0, 127.8, 119.0, 111.1, 110.8, 110.7, 108.5, 105.8, 94.8, 86.4, 66.7, 54.0, 52.8, 39.7, 35.8, 34.2, 33.7, 32,9, 32.8, 9.4 Example 26 Preparation of 2-{2-diethylamino-5-[(2r3-difhiorobenzenesulfonyl) ethyiamino] pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl) propionic. acid. [00165] Steps 1, 2, 3, 4, 6 and 7 were performed as for Example 1. Step 5 was performed using 2,3-difluorobenzenesulfonyl chloride in place of 4- fluorobenzenesulfonyl chloride. 2,3-Difluorobenzenesulfonyl chloride was prepared by the following procedure. 1H NMR (CD3OD) d 8.32 (bs, 1H), 7.90-7.80 (m, 2H), 7.59-7.48 (m, 3H), 7.27-7.23 (m, 2H), 5.09-5.08 (m, 1H), 3.77-3.70 (m, 4H), 3.60-3.51 (m, 3H), 3.50-3.42 (m, 2H), 3.39-3.31n (m, 3H), 3.32-3.18 (m, 2H), 1.43-1.41 (m, 6H) 13C NMR (CD3OD) d 170.4, 160.8, 158.1, 156.1, 153.0, 151.6, 150.5, 148.9, 148.2, 147.3, 147.2, 143.9, 143.5, 142.6, 141.1, 140.9, 131.8, 127.7, 125.1, 123.8, 120.8, 120.6, 119.2, 40.5, 35.7, 33.4, 32.9, 32.7, 9.0 [00166] Preparation of 2,3-Difluorobenzenesulfony] Chloride. The following procedure was executed using two flasks. In the first flask, 2,3- difluoroaniline (2.0 g, 0.015 mol) was dissolved in concentrated HC1 (15.9 mL), and the resulting solution was cooled to -5 °C, using an ice/NaCl bath. A solution of sodium nitrite (1.18 g, 0.017 mol) in distilled water (13.6 mL) was added in portions with stirring, while maintaining the temperature below 0 °C, and the mixture was stirred for 10 min. In the second flask, thionyl chloride (5.08 mL, 0.069 mol) was added dropwise to distilled water (30.6 mL), which had been pre-cooled to -5 °C, using an ice/NaCl bath. The resulting solution was allowed to warm to room temperature, and then Cu(I)Cl (0.08 g, 0.77 mmol) was added, and then the reaction mixture was re-cooled to -5 °C. With continued cooling and stirring, the contents of the first flask were added in 2 mL portions to the contents of the second flask, and the mixture was stirred for 30 min, during which time a precipitate formed. The precipitate was isolated by filtration, rinsed with cold water, and stored under vacuum to give 3.25 g (98%) 10 as a white solid. Example 27 Preparation of 2-{2-Diethylamino-5-[(4-fluorohenzenesulfonyI) propargylamino]pyrimjdin-4-ylamino}-3-(4-dimethylcarhamoyloxyphenyl) propionic acid [00167] Steps 1, 2, 3, 4, 5 and 7 were performed as for Example 1. Step 6 was performed using propargyl bromide in place of dimethyl sulfate. lH NMR (CDCl3) d 1.15 (m, 6H), 2.27 (d, J = 2.1, 1H), 2.97 (s, 3H), 3.06 (s, 3H), 3.10-3.70 (m, 6H), 3.75 (dd, / =17.7, 2.0, 0.6H), 3.95 (dd, / = 18.1, 2.0, 0.4H), 4.51 (dd, / =19.5, 2.2, 0.6H), 4.54 (dd, / = 18.1, 2.2, 0.4H), 4.79 (q, / = 5.9, 0.6H), 4.88 (q, / = 6.6, 0.4H), 6.42 (bd, 0.4H), 6.65 (bs, 0.6H), 6.85-7.30 (m, 6H), 7.52 (s, 0.6H), 7.56 (s, 0.4H), 7.85 (m, 2H), 8.20 (bs, 1H) MS m/z 613.2 (MH+) Example 28 Preparation of 2-{2-Diethylamino-5-[(2t4- difluorobenzenesulfonyt)propargylamino]pyrimidin-4-ylarnino}-3-(4- dimethylcarbamoyloxyphenyPpropionic acid [00168] Steps 1, 2, 3, 4, 5 and 7 were performed as for Example 17. Step 6 was performed using propargyl bromide in place of dimethyl sulfate. 1H NMR (CDC13) d 1.16 (q, / = 7.5, 6H), 2.27 (m, 1H), 2.99 (s, 3H), 3.09 (s, 3H), 3.10-3.70 (m, 6H), 4.04 (dd, / =17.7, 2.4, 0.6H), 4.24 (dd, J = 17.9, 2.2, 0.4H), 4.47 (m, 1H), 4.81 (q, / = 5.9, 0.6H), 4.89 (q, / = 6.3, 0.4H), 6.27 (d, / = 7.5, 0.4H), 6.41 (d, J = 5.7, 0.6H), 6.90-7.10 (m, 4H), 7.16 (d, J = 8.3, 1H), 7.28 (d, / = 8.3, 1H), 7.55 (bs, 1H), 7.66 (s, 0.6H), 7.67 (s, 0.4H), 7.81 (m, 1H) Example 29 Preparation of 2-{2-diethylamino-5-[(4-fluorobenzenesulfonyl)-(2,2r2-trifluoroethyl)amino]p yrimidin-4-ylamino}-3-(4-dimethylcarbamoyl-oxyphenyl)propionic acid [00169] Steps 1, 2, 3, 4, 5 and 7 were performed as for Example 1. Step 6 was performed using 2,2,2-trifluoroethyl triflate and cesium carbonate in place of dimethyl sulfate and potassium carbonate. lR NMR (CDC13) 6 1.14 (m, 6H), 2.98 (s, 3H), 3.06 (s, 3H), 3.10-4.20 (m, 8H), 4.80 (q, J = 5.9, 0.6H), 4.87 (q, J = 6.2, 0.4H), 6.09 (d, J = 5.9, 0.4H), 6.18 (bd, 0.6H), 6.80-7.50 (m, 7H), 7.55 (bs, 1H), 7.77 (m, 2H); MS m/z 657.2 (MH+) Example A a4ß1 Iategrin Adhesion Assay: Jurkat™ Cell Adhesion to Human Plasma Fibronectin Procedure: [00170] 96 well plates (Costar 3590 EIA plates) were coated with human fibronectin (Gibco/BRL, cat #33016-023) at a concentration of 10 /Ag/mL overnight at 4°C. The plates were then blocked with a solution of bovine serum albumin (BSA; 0.3%) in saline. Jurkat™ cells (maintained in log phase growth) were labeled with Calcein AM according to the manufacturer"s instructions, and suspended at a concentration of 2 x 106 cells/mL in Hepes/Saline/BSA. The cells were then exposed to test and control compounds for 30 minutes at room temperature before transfer to individual wells of the fibronectin coated plate. Adhesion was allowed to occur for 35 minutes at 37°C. The wells were then washed by gentle aspiration and pipetting with fresh saline. Fluorescence associated with the remaining adherent cells was quantified using a fluorescence plate reader at EX 485/EM 530. [00171] Cell cultures were prepared by first splitting the stationary phase Jurkat™ cells at 1:10 on day one, and 1:2 on day two to perform assay on day 3. The cells split 1:10 on day one were split 1:4 on day 3 for a day 4 assay. [00172] The assay plates were prepared by first making a working solution of Gibco/BRL Human Fibronectin (cat # 33016-023) In PBS+ + , at 10 µg/mL. A Costar 3590 EIA plate was then coated with 50 µL/well for 2 hours at room temperature (thought it can also be left overnight at 4 °C). Finally the plate was asperated and blocked with Hepes/Saline Buffer, 100 µL/well, for 1 hour at RT followed by washing three times with 150 µL of PBS+ +. [00173] Compound dilutions were accomplished by preparing 1:3 serial dilutions of compounds as follows. For each plate (4 compounds/plate) 600 µL were added to 4 Bio-Rad Titertubes in a Titertube rack. Enough compound was added to each appropriate tube to give a 2X concentration using methods well known in the art. Using Falcon Fiexiplates, 100 µL of Hepes/Saline buffer or human serum were added to rows B through G. A multi-channel pipetter set to 180 µL was used to with four tips spaced evenly the pipetter. Each set of four tubes was mixed 5 times and 180 µL of 2X compound was transferred to the first column of each compound dilution in Row B, leaving Row A empty. 180 µL were added to the other wells in Row A. Serial dilutions were performed down the plate by transferring 50 µL to the next dilution and mixing 5 times, changing tips each time after mixing. Dilutions were stopped at Row F. Row G had no compound present. [00174] A 20 µg/mL solution in Hepes/Saline buffer or human serum, of 21/6 antibody was the positive control and was set aside in a reagent trough to add to cell suspension plate. [00175] The cell staining was accomplished by first harvesting the log-phase Jurkat™ cells by centrifugation in 50 mL tubes (1100 rpm for 5 minutes). The cells were resuspended in 50 mL PBS + +, spun, and resuspend in 20 mL PBS+ +. The ceils were stained by adding 20 µL of Calcein AM for 30 minutes R.T. The volume was brought to 50 mL with Hepes/Saline buffer and the cells were counted, spun, and resuspend to 2 x 106 cells/mL in Hepes/Saline buffer or human serum. [00176] The compounds were incubated using the following procedure. In a new flexiplate, 65 µL of stained cells were added to Rows B through H. Then 65 pL of 2X compounds were added to the appropriate rows following the plate setup and mixed 3X. 65µL of 2X-21/6 antibody were added to Row H and mixed 3X. Finally the plate was incubated at room temperature for 30 minutes. [00177] Fibronectin adhesion was measured using a fluorescent plate reader at EX 485/EM 530 after the following work up procedure. After incubation, the cells were mixed 3X and 100 µL were transferred to the Fibronectin coated plates and incubated at 37 °C for about 35 minutes. Each plate was washed, row by row, by gently pipetting 100 µL of R.T. PBS+ + down the sides of the wells and turning the plate 90 degrees to aspirate. This procedure was repeated for a total of 3 washes. Each well was filled with 100 µL after washing by pipetting down the side of the well. [00178] An IC50 value was calculated for each compound, both in the presence of the human serum and in the absence of human serum. IC50 is concentration at which the growth or activity is inhibited by 50%. The data is presented in the following tables. Cell Adhesion to Human Plasma Fibronectin (Without the human serum) Example B In vitro Saturation Assay For Determining Binding of Candidate Compounds to a4ß1p, [00181] The following describes an in vitro assay to determine the plasma levels needed for a compound to be active in the Experimental Autoimmune Encephalomyelitis ("EAE") model, described in the next example, or in other in vivo models. [00182] Log-growth Jurkat cells are washed and resuspended in normal animal plasma containing 20 µg/mL of the 15/7 antibody (Yednock, et al.,J. Biol. Chem., (1995) 270(48):28740). [00183] The Jurkat cells are diluted two-fold into either normal plasma samples containing known candidate compound amounts in various concentrations ranging from 66 µM to 0.01 µM, using a standard 12 point serial dilution for a standard curve, or into plasma samples obtained from the peripheral blood of candidate compound-treated animals. [00184] Cells are then incubated for 30 minutes at room temperature, washed twice with phosphate-buffered saline ("PBS") containing 2% fetal bovine serum and lmM each of calcium chloride and magnesium chloride (assay medium) to remove unbound 15/7 antibody. [00185] The ceils are then exposed to phycoerythrin-conjugated goat F(ab")2 anti-mouse IgG Fc (Immunotech, Westbrook, ME), which has been adsorbed for any non-specific cross-reactivity by co-incubation with 5% serum from the animal species being studied, at 1:200 and incubated in the dark at 4°C for 30 minutes. [00186] Cells are washed twice with assay medium and resuspended in the same. They are then analyzed with a standard fluorescence activated cell sorter ("FACS") analysis as described in Yednock et al. J. Biol. Chem., 1995, 270:28740. [00187] The data is then graßhed as fluorescence versus dose, e.g., in a normal dose-response fashion. The dose levels that result in the upper plateau of the curve represent the levels needed to obtain efficacy in an in vivo model. [00188] This assay may also be used to determine the plasma levels needed to saturate the binding sites of other integrins, such as the a9ß1 pi integrin, which is the integrin most closely related a4ß1 (Palmer et al, 1993, J. Cell Bio., 123:1289). Such binding is predictive of in vivo utility for inflammatory conditions mediated by a9ß1, integrin, including by way of example, airway hyper-responsiveness and occlusion that occurs with chronic asthma, smooth muscle cell proliferation in atherosclerosis, vascular occlusion following angioplasty, fibrosis and glomerular scarring as a result of renal disease, aortic stenosis, hypertrophy of synovial membranes in rheumatoid arthritis, and inflammation and scarring that occur with the progression of ulcerative colitis and Crohn"s disease. [00189] Accordingly, the above-described assay may be performed with a human colon carcinoma cell line, SW 480 (ATTC #CCL228) transfected with cDNA encoding a9 integrin (Yokosaki et al., 1994, J. Biol. Chem., 269:26691), in place of the Jurkat cells, to measure the binding of the a9ß1 integrin. As a control, SW 480 cells which express other a and ß1 subunits may be used. [00190] Accordingly, another aspect of this invention is directed to a method for treating a disease in a mammalian patient, which disease is mediated by a9ß1, and which method comprises administering to said patient a therapeutically effective amount of a compound of this invention. Such compounds are preferably administered in a pharmaceutical composition described herein above. Effective daily dosing will depend upon the age, weight, condition of the patient which factors can be readily ascertained by the attending clinician. However, in a preferred embodiment, the compounds are administered from about 20 to 500 µg/kg per day. Example C Cassette Dosing and Serum Analysis for determination of Bioavailability [00191] The oral bioavailability was screened by dosing rats with a cassette, i.e. mixture of 6 compounds per dosing solution. The cassette included 5 test articles and a standard compound, for a total dose of 10 mg/kg. Each compound/test article was converted to the sodium salt with equimolar 1 N NaOH and dissolved in water at 2 mg/mL. The cassette was prepared by mixing equal volumes of each of the six solutions. The cassette dosing solution was mixed well and then the pH was adjusted to 7.5-9. The dosing solution was prepared the day before the study and stirred overnight at room temperature. [00192] Male Sprague Dawley (SD) rats from Charles River Laboratories, 6- 8 weeks old were used in this screen. Rats were quarantined for at least one day and had continuous access to food and water. On the night before the administration of the cassette, the rats were fasted for aßproximately 16 h. [00193] Four SD rats were assigned in each cassette. A single dose of the dosing solution was administered orally to each rat. The dosing volume (5 mL/kg) and time were recorded and rats were fed 2 h after dosing. [00194] Blood samples were collected via cardiac puncture at the following time points: 4 h, 8 h and 12 h. Immediately prior to blood collection, rats were anesthetized with CO2 gas within 10-20 seconds. After the 12-hour samples were collected, the rats were euthanized via CO2 asphyxiation followed by cervical dislocation. [00195] Blood samples were kept in heparinized microtainer tubes under sub-ambient temperature (4 °C) before they were processed. Blood samples were centrifuged (10000 rpm for 5 minutes) and plasma samples were removed and stored in a -20 °C freezer until analyzed for drug levels. Drug levels in the plasma were analyzed using the following protocol for direct plasma precipitation. [00196] The in vivo plasma samples were prepared in a 1.5 mL 96-well plate, by adding, in order, 100 µL of the test plasma, 150 µL of methanol, followed by vortexing for 10-20 seconds. 150 µL of 0.05 ng//µL of an Internal Standard in acetonitrile were added and vortexed for 30 seconds. [00197] The standard curve samples were prepared in a 1.5 mL 96-well plate, by adding, in order, 100 µL of control mouse plasma, followed by 150 µL of methanol and vortexing for 10-20 seconds. 150 µL of 0.05 ng/µL of an Internal Standard in acetonitrile were added and vortexed for 30 seconds. The samples were spiked with 0-200 ng (10 concentrations) of the compound of interest in 50% methanol to obtain a standard curve range of 0.5 ng/mL - 2,000 ng/mL. Again, the sample was vortexed for 30 seconds. [00198] The samples were then spun for 20-30 minutes at 3000 rpm in an Eppendorf microfuge before 80-90% of supernatant was transferred into a clean 96-well plate. The organic solvent was then evaßorated until the samples were dry (under N2 at 40°C/ 30-60 min (ZymarkTurbovaß)). [00199] The residue was then dissolved in 200 - 600 L mobile phase (50% CH3OH/0.1% TFA). LC/MS/MS was then run using a PE-Sciex aßi-3000 triple quadurpole mass spectrometer (SN0749707), Perkin-Elmer, Series200auto-sampler, and shimadzu 10A pump. Acquisition was done with • PE-Sciex Analyst (vl.l) and data analysis and quantification were accomplished using PE-Sciex Analyst (vl.l). A 5-50 µL sample volume was injected onto a reverse phase ThermoHypersil DASH-18 column (Keystone 2.0 x 20 mm, 5 µm, PN: 8823025-701) using a mobile phase of 25% CH3OH, 0.1% TFA-100% CH3OH, 0.1% TFA. The run time was about 8 minutes at a flow rate of about 300 µL/minutes. [00200] The Area Under the Curve (AUC) was calculated using the linear traßezoidal rule from t=0 to the last sampling time t, (see Handbook of Basic Pharmacokinetics, Wolfgang A. Ritschel and Gregory L. Kearns, 5th ed, 1999). AUC0?tx = S((Cn + Cn+1)/2)) • (tn+1 - tr) [µg/mL)h] [00201] In the case of the cassette dosing paradigm, samples at 4, 8 and 12 h post extravascular dosing, the AUC was calculated from t = 0 to t = 12 h. The AUC°?12h values were calculated for each individual animal and the average AUC°?12h are reported in the table below. Example D Asthma Models [00203] Inflammatory conditions mediated by a4ß1 integrin include, for example, eosinophil influx, airway hyper-responsiveness and occlusion that occurs with chronic asthma. The following describes animal models of asthma that were used to study the in vivo effects of the compounds of this invention for use in treating asthma. Rat Asthma Model [00204] Following the procedures described by Chaßman et al, Am J. Resp. Crit. Care Med,. 153 4, A219 (1996) and Chaßman et al, Am. J. Resp. Crit Care Med 155:4, A881 (1997), both of which are incorporated by reference in their entirety. Ovalbumin (OA; 10 µg/mL) were mixed with aluminum hydroxide (10 mg/mL) and injected (i.p.) in Brown Norway rats on day 0. Injections of OA, together with adjuvant, were repeated on days 7 and 14. On day 21, sensitized animals were restrained in plastic tubes and exposed (60 minutes) to an aerosol of OA (10 mg/kg) in a nose-only exposure system. Animals will be sacrificed 72 hours later with pentobarbital (250 mg/kg, i.p.). The lungs were lavaged via a tracheal cannula using 3 aliquots (4 mL) of Hank"s solution (HBSS x 10, 100 mL; EDTA 100 mM, 100 mL; HEPES 1 M, 25 mL; made up to 1 L with H20); recovered cells were pooled and the total volume of recovered fluid adjusted to 12 mL by addition of Hank"s solution. Total cells were counted (Sysmex microcell counter F-500, TOA Medical Electronics Otd., Jaßan) and smears were made by diluting recovered fluid (to aßproximately 106 cells/mL) and pipetting an aliquot (100 µL) into a centrifuge (Cytospin, Shandon, U.K.). Smears were air dried, fixed using a solution of fast green in methanol (2 mg/mL) for 5 seconds and stained with eosin G (5 seconds) and thiazine (5 seconds) (Diff-Quick, Browne Ltd. U.K.) in order to differentiate eosinophils, neutrophils, macrophages and lymphocytes. A total of 500 cells per smear were counted by light microscopy under oil immersion (x 100). Compounds of this invention were formulated into a 0.5% carboxymethylcellulose and 2% Tween80 suspension and administered orally to rats which had been sensitized to the allergen, ovalbumin. Compounds which inhibited allergen- induced leucocyte accumulation in the airways of actively sensitized Brown Norway rats were considered to be active in this model. Mouse Asthma Model [00205] Compounds were also evaluated in a mouse model of acute pulmonary inflammation following the procedures described by, Kung et ai., Am J. Respir. Cell Mol. Biol. 13:360-365, (1995) and Schneider etal., (1999). Am J. Respir. Cell Mol. Biol. 20:448-457, (1999), which are each incorporated by reference in their entirety. Female Black/6 mice (8-12 weeks of age) were sensitized on day 1 by an intraßeritoneal injection (i.p.) of 0.2 mL ova/alum mixture containing 20 µg of ova (Grade 4, Sigma) and 2 mg inject Alum (pierce). A booster injection was administered on day 14. Mice are challenged on days 28 and 29 with aerosolized 1 % ova (in 0.9% saline) for 20 minutes. Mice are euthanized and bronchaveolar lavage samples (3 mL) are collected on day 30, 48 hours post first challenge. Eosinophils were quantified by a FACs/FITC staining method. Compounds of this invention were formulated into a 0.5% carboxymethylcellulose and 2% Tween80 suspension and administered orally to mice which had been sensitized to the allergen, ovalbumin. Compounds which inhibited allergen-induced leucocyte accumulation in the airways of actively sensitized C57BL/6 mice were considered to be active in this model. Sheep Asthma Model [00206] This model employs the procedures described by Abraham et al., J.Clin, Invest, 93:776-787 (1994) and Abraham et al., Am J. Respir Crit Care Med 156:696-703 (1997), both of which are incorporated by reference in their entirety. Compounds of this invention have been evaluated by intravenous (saline aqueous solution), oral (2% Tween80, 0.5% carboxymethylcellulose), and aerosol administration to sheep which are hypersensitive to Ascaris suum antigen. Compounds which decrease the early antigen-induced bronchial response and/or block the late-phase airway response, e.g. have a protective effect against antigen-induced late responses and airway hyper-responsiveness ("AHR"), are considered to be active in this model. [00207] Allergic sheep which are shown to develop both early and late bronchial responses to inhaled Ascaris suum antigen were used to study the airway effects of the candidate compounds. Following topical anesthesia of the nasal passages with 2% lidocaine, a balloon catheter was advanced through one nostril into the lower esophagus. The animals were then incubated with a cuffed endotracheal tube through the other nostril with a flexible fiberoptic bronchoscope as a guide. [00208] Pleural pressure was estimated according to Abraham (1994). Aerosols (see formulation below) were generated using a disposable medical nebulizer that provided an aerosol with a mass median aerodynamic diameter of 3.2 µm as determined with an Andersen cascade impactor. The nebulizer was connected to a dosimeter system consisting of a solenoid valve and a source of compressed air (20 psi). The output of the nebulizer was directed into a plastic T-piece, one end of which was connected to the inspiratory port of a piston respirator. The solenoid valve was activated for 1 second at the beginning of the inspiratory cycle of the respirator. Aerosols were delivered at VT of 500 mL and a rate of 20 breaths/minute. A 0.5 % sodium bicarbonate solution only was used as a control. [00209] To assess bronchial responsiveness, cumulative concentration- response curves to carbachol was generated according to Abraham (1994). Bronchial biopsies were taken prior to and following the initiation of treatment and 24 hours after antigen challenge. Bronchial biopsies were preformed according to Abraham (1994). [00210] An in vitro adhesion study of alveolar macrophages were also performed according to Abraham (1994), and a percentage of adherent cells calculated. Aerosol Formulation [00211] A solution of the candidate compound in 0.5% sodium bicarbonate/saline (w/v) at a concentration of 30.0 mg/mL is prepared using the following procedure: Procedure: 1. Add 0.5g sodium bicarbonate into a 100 mL volumetric flask. 2. Add approximately 90.0 mL saline and sonicate until dissolved. 3. Q.S. to 100.0 mL with saline and mix thoroughly. [00213] B. Preparation of 30.0 mg/mL Candidate Compound: 10.0 mL Procedure: 1. Add 0.300 g of the candidate compound into a 10.0 mL volumetric flask. 2. Add approximately 9.7 mL of 0.5% sodium bicarbonate / saline stock solution. 3. Sonicate until the candidate compound is completely dissolved. 4. Q.S. to 10.0 mL with 0.5% sodium bicarbonate / saline stock solution and mix thoroughly. Example E 10-Day Toxicity Study on C57B6 Mice [00214] A 10-day study was conducted to evaluate the toxicity of compounds of the present invention to female C57B6 mice. The compound was administered by gavage at five dose levels, 0 (vehicle control), 10, 30, 100, 300 and 1000 mg/kg (mpk), with five mice in each dose level. The dose volume for all levels was 10 mL/kg. Dose solutions or suspensions were prepared in 2% Tween 80 in 0.5% carboxymethyl cellulose (CMC) and new dose solutions or suspensions were prepared every two - three days. In-life observations included body weights (study day 1, 2, 3, 5, 7, 8 and 11), daily cageside clinical observations (1-2/day) and periodic (study day -1, 2 and 9) functional observation battery. [00215] At termination, blood samples were collected by cardiac puncture for clinical pathology (hematology and clinical chemistry) and drug levels. The EDTA blood samples were analyzed for total white blood cell count, red blood cell count, hemoglobin, hematocrit, erythrocyte indices (MCV, MCH, MCHC), platelets and a WBC five part differential (neutrophil, lymphocytes, monocytes, eosinophils and basophils). Heparinized plasma samples were analyzed for alanine transaminase, aspartate transaminase, alkaline phosphatase, total bilirubin, albumin, protein, calcium, glucose, urea nitrogen, creatinine, cholesterol and triglycerides. [00216] After blood collection, the carcass was necropsied and organs (liver, spleen, kidneys, heart and thymus) were weighed. Tissue samples; brain, salivary glands, thymus, heart, lung, liver, kidney, adrenal spleen, stomach, duodenum, ileum, colon and uterus/ovary, were collected and formalin fixed. Tissues from the vehicle control and 300 and 1000 mpk group animals were processed to H & E stained glass slides and evaluated for histopathological lesions. [00217] Body weight changes, absolute and relative organ weights and clinical pathology results were analyzed for statistical significant differences compared to the vehicle controls by Dunnet"s multiple comparison test using Prism software. The functional observation battery results were analyzed for differences using the Dunnet"s, Fisher"s exact tests and dose trend effects by the Cochran-Mantel-Haenszel correlation test using SAS software. [00218] Using a conventional oral formulation, compounds of this invention would be active in this model. Example F Adjuvant-Induced Arthritis in Rats [00219] Adjuvant induced arthritis ("ALA") is an animal model useful in the study of rheumatoid arthritis (RA), which is induced by injecting M. tuberculosis in the base of the tail of Lewis rats. Between 10 and 15 days following injection, animals develop a severe, progressive arthritis. [00220] Generally, compounds are tested for their ability to alter hind paw swelling and bone damage resulting from adjuvant-induced edema in rats. To quantitate the inhibition of hind paw swelling resulting from AIA, two phases of inflammation have been defined: (1) the primary and secondary injected hind paw, and (2) the secondary uninjected hind paw, which generally begins developing about eleven days from the induction of inflammation in the injected paw. Reduction of the latter type of inflammation is an indication of immunosuppressive activity. Cf. Chang, Arth. Rheum., 20, 1135-1141 (1977). [00221] Using an animal model of RA, such as AIA, enables one to study the cellular events involved in the early stages of the disease. CD44 expression on macrophages and lymphocytes is up-regulated during the early development of adjuvant arthritis, whereas LFA-1 expression is up-regulated later in the development of the disease. Understanding the interactions between adhesion molecules and endothelium at the earliest stages of adjuvant arthritis could lead to significant advances in the methods used in the treatment of RA. WE CLAIM: 1. A compound of formula (I): wherein each X is independently fluoro, chloro or bromo; p is 0 or an integer from 1-3; R1 is selected from the group consisting of methyl and ethyl; R2 is selected from the group consisting of monovalent (C1 to C5) lower alkyl, monovalent (C2 to C3) lower alkenyl, and lower alkylene cycloalkyl, where lower alkylene refers to a divalent (C1 to C4) lower alkylene group which is bound to cycloalkyl which refers to a monovalent (C3 to C6) cycloalkyl group and pharmaceutical ly acceptable salts thereof. 2. A compound of Formula II: wherein each X is independently selected from the group consisting of fluoro and chloro, m is an integer equal to 1 or 2; R2 is selected from the group consisting of monovalent (C1 to C5) lower alkyl, monovalent (C2 to C3) lower alkenyl, and lower alkylene cycloalkyl, where lower alkylene refers to a divalent (C1 to C4) lower alkylene group which is bound to cycloalkyl which refers to a monovalent (C3 to C6) cycloalkyl group and pharmaceutically acceptable salts thereof. wherein each X is independently fluoro or chloro; n is zero or one; R2 is -CH2-R" where R" is selected from the group consisting of hydrogen, methyl or -CH=CH2; and pharmaceutically acceptable salts thereof. 4. A compound as claimed in any one of claims 1, 2 or 3, wherein R2 is CH3. 5. A compound as claimed in claim 3, wherein X is F or Cl, and n is 0. 6. A compound selected from the group consisting of: 2-{2-diethylamino-5-[(4-chlorobenzenesulfonyl)tnethylatnino]-pyritnidin-4-ylamino}-3- (4-ditnethylcarbamoyloxyphenyl)propionic acid; 2-{2-dietliylamino-5-[(4-fluorobenzenesulfonyl)methylainino]-pyrimidin-4-ylainino}-3- (4-dimethylcarbatnoyloxyphenyl)propionic acid; 2-{2-diethylamino-5-[(3,4-difluorobenzenesulfonyl)methylamino]-pyrimidin-4-ylamino } -3-(4-dimethylcarbamoyloxyphenyl)propionic acid; 2-{2-diethylamino-5-[(3,4-dichlorobenzenesulfonyl)methylamino]-pyrimidin-4-ylamiao }-3-(4-dimethylcarbamoyloxyphenyl)propionic acid; 2-{2-diethylamino-5-[(benzenesulfonyl)methylamino]-pyrimidin-4-ylamino}-3-(4-dime thylcarbamoyloxyphenyl)propionic acid; 2-{2-diethylamino-5-[(2-fluorobenzeaesulfonyl)methylamino]-pyritnidin-4-ylamino}-3- (4-dimethylcarbamoyloxyphenyl)propionic acid; 2-{2-diethylamino-5-[(3-fluorobenzenesulfonyl)methylamino]-pyrimidin-4-ylamino}-3- (4-dimethylcarbamoyloxyphenyl)propionic acid; 2-{2-diethylamino-5-[(4-fluorobenzenesulfonyl)isopropylamino]-pyrimidin-4-ylamino}- 3-(4-dimethylcarbamoyloxyphenyl)propionic acid; 2-{2-diethylamino-5-[(4-fluorobenzenesulfonyl)ethylamino]-pyrimidin-4-ylamino}-3-( 4-dimethylcarbamoyloxyphenyl)propionic acid; 2-{2-diethylamino-5-[(3,4-difluorobenzenesuIfonyl)isopropylatnino]-pyrimidin-4-ylatni no}-3-(4-dimethylcarbamoyloxyphenyl)propionic acid; 2-{2-diethylamino-5-[(4-chlorobenzenesulfonyl)isopropylamino]-pyrimidin-4-ylamino}- 3-(4-ditnethylcarbamoyloxyphenyl)propionic acid; 2-{2-diethyIamino-5-[(3,4-difluorobenzenesulfonyl)ethylamino]-pyrimidin-4-yIamino}- 3-(4-dimethylcarbamoyloxyphenyI)propionic acid; 2-{2-diethylamino-5-[(4-chlorobenzenesulfonyl)ethylamino]-pyrimidin-4-ylamino}-3-( 4-dimethylcarbamoyloxyphenyl)propionic acid; 2-{2-diethylamino-5-[(4-fluorobenzenesulfonyl)cylclopropylmethyl-amino]pyrimidin-4- ylamino}-3-(4-dimethylcarbamoyloxyphenyl)propionic acid; 2-{2-diethylamino-5-[(3,5-difluorobenzenesulfonyl)methylamino]-pyrimidin-4-ylamino } -3-(4-dimethylcarbamoyloxyphenyl)propionic acid 2-{2-diethyIamino-5-[(3,5-difluorobenzenesuIfonyl)ethylamino]-pyrimidin-4-ylamino}- 3-(4-dimethylcarbamoyloxyphenyl)propionic acid; 2-{2-diethylamino-5-[(2,4-difluorobenzenesulfonyl)methylamino]-pyrimidin-4-ylamino } -3-(4-dimethylcarbamoyloxyphenyl)propionic acid; 2-{2-diethylamino-5-[(2,4-difluorobenzenesulfonyl)ethylamino]-pyrimidin-4-ylamino}- 3-(4-dimethylcarbarnoyloxyphenyl)propionic acid; 2-{2-diethylamino-5-[(3,5-dichlorobenzenesulfonyl)tnethylamino]-pyrimidin-4-ylamino }-3-(4-dimethylcarbamoyloxyphenyl)propionic acid; 2-{2-diethylamino-5-[(3,5-dichlorobenzenesulfonyl)ethylamino]-pyrimidin-4-ylamino}- 3-(4-dimethylcarbamoyloxyphenyl)propionic acid; 2-{2-diethylamino-5-[(4-fluorobenzenesulfonyl)-n-propylamino]-pyrimidin-4-ylamino}- 3-(4-dimethylcarbamoyloxyphenyl)propionic acid; 2-{2-diethylamino-5-[(4-fluorobenzenesulfonyl)allylami]no]-pyrimidin-4-ylamino}-3-(4- dimethylcarbamoyloxyphenyl)propionic acid; 2-{2-diethylamino-5-[(4-fluorobenzenesulfonyl)isobotylamino]-pyrimidin-4-ylamino}- 3-(4-dimethylcarbamoyloxyphenyl)propionic acid 2-{2-diethylamino-5-[(4-fluorobenzenesulfonyl)-n-butylamino]-pyrimidin-4-ylamino}- 3-(4-dimethylcarbamoyloxyphenyl)propionic acid; 2-{2-diethylamino-5-[(2,5-difluorobenzenesulfonyl)methylamino]-pyrimidin-4-ylamino }-3-(4-dimethylcarbamoyloxyphenyl)propionic acid; 2-{2-diethyIamino-5-[(2,3-difluorobenzenesulfonyl)ethylamino]-pyrimidin-4-ylamino}- 3-(4-dimethylcarbamoyloxyphenyl)propionic acid; 2-{2-Diethylamino-5-[(4-fluorobenzenesulfonyl)-(2-trisfluoroethyl)- amino]pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)propionic acid; and pharmaceutically acceptable salts thereof. 7. A pharmaceutical composition comprising a pharmaceutically acceptable carrier and a therapeutical ly effective amount of a compound as claimed in any one of claims 1-3, 5, or 6. 8. A pharmaceutical composition as claimed in claim 7 for treating a disease mediated by cu integrin in a patient. 9. A pharmaceutical composition as claimed in claim 8, whereinthe disease mediated by 04 integrin is an inflammatory disease/ 10. A compound of formula (IV): wherein each X is independently fluoro, chloro or bromo; p is 0 or an integer from 1-3; Rl is selected from the group consisting of methyl and ethyl; R2 is lower alkynyl; and pharmaceutical ly acceptable salts thereof. 11. A compound of Formula V: wherein each X is independently selected from the group consisting of fluoro and chloro, m is an integer equal to 1 or 2; R2 is lower alkynyl; and pharmaceutically acceptable salts thereof. 12. A compound of Formula VI: wherein each X is independently fluoro or chloro; n is zero or one; R2 is lower alkynyl; and pharmaceutically acceptable salts thereof. 13. A compound as claimed in any one of claims 10, 11 or 12, wherein R2 is propargyl. 14. A compound as claimed in claim 12, wherein X is F or Cl, and n is 0. 15. A compound selected from the group consisting of: 2-{2-Diethylamino-5-[(4-fluorobenzenesulfonyl)propargylamino] pyrimidin-4- ylamino}-3-(4-dimethylcarbamoyloxyphenyl)propionic acid, 2-{2-Diethylamino-5-[(2,4-difluorobenzenesulfonyl)propargylamino] pyrimidin-4- ylamino}-3-(4-dimethylcarbamoyloxyphenyl)propionic acid; and pharmaceutically acceptable salts thereof. 16. A pharmaceutical composition comprising a pharmaceutically acceptable carrier and a therapeutically effective amount of a compound as claimed in any one of claims 10-12, 14 or 15. 17. A pharmaceutical composition as claimed in claim 16 for treating a disease mediated by cu integrin in a patient. 18. A pharmaceutical composition as claimed in claim 17, wherein the disease mediated by cu integrin is an inflammatory disease. 19. A pharmaceutical composition as claimed in claim 17, wherein the disease is rheumatoid arthritis. Disclosed are compounds which bind 4 integrins, where the 4integrin is preferably VLA-4. Certain of these com- pounds also inhibit leukocyte adhesion and. in particular, leukocyte adhesion mediated by 4integrins, where the 4 integrin is preferably VLA-4. Such compounds are useful in the treatment of inflammatory diseases in a mammalian patient, e.g., human, such as asthma, Alzheimer"s disease, atherosclerosis, AIDS dementia, diabetes, inflammatory bowel disease, rheumatoid arthritis, tissue transplan- tation, tumor metastasis and myocardial ischemia. The compounds can also be administered for the treatment of inflammatory brain diseases such as multiple sclerosis.

Full Text

HETEROARYL COMPOUNDS WHICH INHIBIT LEUKOCYTE
ADHESION MEDIATED BY a4 INTEGRINS
BACKGROUND OF THE INVENTION
Field of the Invention
[0001] This invention relates to compounds which inhibit leukocyte adhesion
and, in particular, leukocyte adhesion mediated by a4 integrins, where the
a4 integrin is preferably VLA-4.
References
[0002] The following publications, patents and patent applications are cited
in this application as superscript numbers:
[0003] 1 Hemler and Takada, European Patent Application Publication
No. 330,506, published August 30, 1989
[0004] 2 Elices, et al., Cell, 60.577-584 (1990)
[0005]3 Springer, Nature, 246:425-434 (1990)
[0006] 4 Osborn, Cell, 62:3-6 (1990)
[0007] 5 Vedder, et al., Surgery, 106:509 (1989)
[0008] 6 Pretolani, et al., J. Exp. Med., 180:795 (1994)
[0009] 7 Abraham, et al., J. Clin. Invest., 21:776 (1994)
[0010] 8 Mulligan, et al., J. Immunology, 150_:2407 (1993)
[0011] 9 Cybulsky, et al., Science, 25.1:788 (1991)
[0012] l0 Li, et al., Arterioscler. Thromb., 12:197 (1993)
[0013] 11 Sasseville, et al., Am. J. Path., 144:27 (1994)
[0014] l2 Yang, et al., Proc. Nat. Acad. Science (USA), 90: 10494
(1993)
[0015] 13 Burkly, et al., Diabetes, 42:529 (1994)
[0016] H Baron, et al., J. Clin. Invest., 92.: 1700 (1994)
[0017] 15 Hamann, et al., /. Immunology, 152:3238 (1994)
[0018] l6 Yednock, et al.. Nature, 256:63 (1992)
[0019] 17 Baron, et al., J. Exp. Med., 121:57 (1993)
[0020] 18 van Dinther-Janssen, et al., J. Immunology, 142:4207 (1991)
[0021] 19 van Dinther-Janssen, et al., Annals. RheumaticDis., 52:672
(1993)
[0022]20 Elices, et al., /. Clin. Invest., 22:405 (1994)
[0023] n Postigo, et al., /. Clin. Invest., 89_: 1445 (1991)
[0024] 22 Paul, et al., Transpl. Proceed., 25:813 (1993)
[0025] B Okarhara, et al., Can. Res., 54:3233 (1994)
[0026]24 Paavonen, et al., Int. J. Can., 58:298 (1994)
[0027] M Schadendorf, et al., /. Path., 120:429 (1993)
[0028] 2S Bao, et al., Diff., 52:239 (1993)
[0029]27 Lauri, et al., British J. Cancer, 68:862 (1993)
[0030] M Kawaguchi, et al., Japanese J. Cancer Res., 82:1304 (1992)
[0031]29 Konradi, et al., PCT/US00/01686, filed, January 21, 2000.
[0032] All of the above publications are herein incorporated by reference in
their entirety to the same extent as if each individual publication was
specifically and individually indicated to be incorporated by reference in its
entirety.
State of the Art
[0033] VLA-4 (also referred to as a4ß1 integrin and CD49d/CD29), first
identified by Hemler and Takada1, is a member of the ß1 integrin family of
cell surface receptors, each of which comprises two subunits, an a chain and
a ß chain. VLA-4 contains an a4 chain and a ß1 chain. There are at least
nine ß1 integrins, all sharing the same ß1 chain and each having a distinct a
chain. These nine receptors all bind a different complement of the various
cell matrix molecules, such as fibronectin, laminin, and collagen. VLA-4,
for example, binds to fibronectin. VLA-4 also binds non-matrix molecules
that are expressed by endothelial and other cells. These non-matrix
molecules include VCAM-1, which is expressed on cytokine-activated
human umbilical vein endothelial cells in culture. Distinct eß1topes of VLA-
4 are responsible for the fibronectin and VCAM-1 binding activities and
each activity has been shown to be inhibited independently.2
[0034] Intercellular adhesion mediated by VLA-4 and other cell surface
receptors is associated with a number of inflammatory responses. At the site
of an injury or other inflammatory stimulus, activated vascular endothelial
cells express molecules that are adhesive for leukocytes. The mechanics of
leukocyte adhesion to endothelial cells involves, in part, the recognition and
binding of cell surface receptors on leukocytes to the corresponding cell
surface molecules on endothelial cells. Once bound, the leukocytes migrate
across the blood vessel wall to enter the injured site and release chemical
mediators to combat infection. For reviews of adhesion receptors of the
immune system, see, for example, Springer3 and Osborn.4
[0035] Inflammatory brain disorders, such as experimental autoimmune
encephalomyelitis (EAE), multiple sclerosis (MS) and meningitis, are
examples of central nervous system disorders in which the
endothelium/leukocyte adhesion mechanism results in destruction to
otherwise healthy brain tissue. Large numbers of leukocytes migrate across
the blood brain barrier (BBB) in subjects with these inflammatory diseases.
The leukocytes release toxic mediators that cause extensive tissue damage
resulting in impaired nerve conduction and paralysis.
[0036] In other organ systems, tissue damage also occurs via an adhesion
mechanism resulting in migration or activation of leukocytes. For example,
it has been shown that the initial insult following myocardial ischemia to
heart tissue can be further complicated by leukocyte entry to the injured
tissue causing still further insult (Vedder et al.).5 Other inflammatory or
medical conditions mediated by an adhesion mechanism include, by way of
example, asthma,6-8 Alzheimer"s disease, atherosclerosis,9-10 AIDS
dementia,11 diabetes12-14 (including acute juvenile onset diabetes),
inflammatory bowel disease15 (including ulcerative colitis and Crohn"s
disease), multiple sclerosis,16-17 rheumatoid arthritis,18-21 tissue
transplantation,22 tumor metastasis,23-28 meningitis, encephalitis, stroke, and
other cerebral traumas, nephritis, retinitis, atoß1c dermatitis, psoriasis,
myocardial ischemia and acute leukocyte-mediated lung injury such as that
which occurs in adult resß1ratory distress syndrome.
[0037] Substituted aminopyrimidines, as a class, have been disclosed as
inhibiting binding of VLA-4 to VCAM-1 and, accordingly, exhibit anti-
inflammatory properties.29 While these compounds possess antagonist
properties to such binding, enhanced bioavailabiiity of these compounds
would augment their efficacy.
SUMMARY OF THE INVENTION
[0038] This invention is directed to the discovery that certain N-[2-N",N"-
diethylamino-5-aminosuifonylphenylpyrimidin-4-yl]-p-carbomyloxy-
phenylalanine compounds possess unexpectedly superior bioavailability, as
measured by their AUC, as compared to other substituted aminopyrimidine
compounds previously disclosed.
[0039] In one of its composition aspects, this invention is directed to a
compound of formula (I):

wherein each X is independently fluoro, chloro or bromo;
p is 0 or an integer from 1-3;
R1 is selected from the group consisting of methyl and ethyl;
R2 is selected from the group consisting of lower alkyl, lower
alkenyl, and lower alkylenecycloalkyl;
and pharmaceuticaHy acceptable salts thereof.
[0040] In a preferred embodiment, this invention provides compounds of
Formula II:
wherein each X is independently selected from the group consisting
of fluoro and chloro,
m is an integer equal to 1 or 2;
R2 is selected from the group consisting of lower alkyl, lower
alkenyl, and lower alkylenecycloalkyl;
and pharmaceutically acceptable salts thereof.
[0041] In a particularly preferred embodiment, this invention provides
compounds of Formula III:
wherein each X is independently fluoro or chloro;
n is zero or one;
R2 is -CH2-R" where R1 is selected from the group consisting of
hydrogen, methyl or -CH=CH2;
and pharmaceutically acceptable salts thereof.
[0042] In another particularly preferred embodiment, this invention provides
compounds of Formula (IV):
wherein each X is independently fluoro, chloro or bromo;
p is 0 or an integer from 1-3;
R1 is selected from the group consisting of methyl and ethyl;
R2 is lower alkynyl;
and pharmaceutically acceptable salts thereof.
[0043] In yet another particularly preferred embodiment, this invention
provides compounds of Formula V:
wherein each X is independently selected from the group consisting
of fluoro and chloro,
m is an integer equal to 1 or 2;
R2 is lower alkynyl;
and pharmaceutically acceptable salts thereof.
[0044] In still another particularly preferred embodiment, this invention
provides compounds of Formula VI:
wherein each X is independently fluoro or chloro;
n is zero or one;
R2 is lower alkynyl;
and pharmaceutically acceptable salts thereof.
R2 is preferably propargyl in any of one of Formula IV, V or VI..
[0045] N-[2-N\N"-diethylamino-5-aminosulfonylphenylpyrimidin-4-yl]-p-
carbomyloxyphenylalanine compounds within the scope of this invention
include those set forth in Table.I as follows..
[0046] Specific compounds within the scope of this invention include the
following. As used below, these compounds are named based on proß1onic
acid derivatives but, alternatively, these compounds could have been named
based on N-[2-N\N"-diethylarnino-5-aminosulfoaylphenylpyrimidin-4-yl]-p-
carbomyIoxy-phenylalanine derivatives.
[0047] 2-{2-diethylamino-5-[(4-chlorobenzenesulfonyl)methylamino]-
pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)proß1onicacid;
[0048] 2-{2-diethylamino-5-[(4-fluor"obenzenesulfonyl)methylamino]-
pyrirnidin-4-yIamino}-3-(4-dimethylcarbamoyloxyphenyl)proß1onicacid;
[0049] 2-{2-diethylamino-5-[(3,4-difluorobenzenesulfonyl)methylamino]-
pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)proß1onicacid;
[0050] 2-{2-diethylamino-5-[(3,4-dichlorobenzenesulfonyl)methylamino]-
pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)proß1onic acid;
[0051] 2-{2-diethylamino-5-[(benzenesulfonyl)methylamino]-
pyrinudin-4-ylamino}-3-(4-dimethylcarbanioyloxyphenyl)propionic acid;
[0052] 2-{2-diediylamino-5-[(2-fluorobeiizenesulfonyl)methylamino]-
pyrimidin-4-ylamino}-3-(4"dimethylcarbamoyloxyphenyl)propionicacid;
[0053] 2-{2-diethylamino-5-[(3-fluorobenzenesulfonyl)methylamino]-
pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)propionicacid;
[0054] 2-{2-diethylamino-5-[(4-fluorobenzenesulfonyl)isopropylamino]-
pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)propionicacid;
[0055] 2-{2-diethylammo-5-[(4-fluorobenzenesulfonyl)ethylamino]-
pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)propionicacid;
[0056] 2-{2-diethylamino-5-[(3,4-difluorobenzenesulfoayl)isopropylamino]-
pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)propionicacid;
[0057] 2-{2-diethylamino-5-[(4-chlorobenzenesulfonyI)isopropylamino]-
pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyplienyl)propionicacid;
[0058] 2-{2-diethylamino-5-[(3,4-difluorobenzenesulfonyl)ethylamino]-
pyrimidin^ylamino}-3-(4-dimethylcarbamoyloxyphenyl)propionicacid;
[0059] 2-{2-diethylamino-5-[(4-chlorobenzenesulfonyl)ethylamino]-
pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)propionicacid;
[0060] 2-{2-diethylamino-5-[(4-fluorobenzenesulfonyl)cylclopropylmethyl-
amino]pyrimidin-4-ylamino}-3-(4-dimethylcarbatnoyloxyphenyl)propionic
acid;
[0061] 2-{2-diethylamino-5-[(3,5-difiuorobenzenes\iifoDyl)methylamino]-
pyrimidin^-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)propionicacid;
[0062] 2-{2-diethylamino-5-[(3,5-difluorobenzenesulfonyl)ethylamino]-
pyrimidin-4-yIamino}-3-(4-dimethylcarbamoyloxypb.enyl)propionicacid;
[0063] 2-{2-diethylaiiiino-5-[(2,4-difluorobenzeaesulfonyI)methylamino]-
pyrimidia-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)propionicacid;
[0064] 2-{2-diethylamino-5-[(2,4-difluorobenzenesulfonyl)ethylamino]-
pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)proß1onicacid;
[0065] 2-{2-diethylamino-5-[(3,5-dichlorobenzenesulfonyl)methylamino]-
pyrimidm-4-ylamino}-3-(4-dimethylcarbamoyloxyj)henyl)proß1onicacid;
[0066] 2-{2-diethylamino-5-[(3,5-dichlorobenzenesulfonyl)ethylamino]-
pyrimidin-4-ylamino} -3-(4-dimethylcarbamoyloxyphenyl)proß1onic acid;
[0067] 2-{2-diethylamino-5-[(4-fluorobenzenesulfonyl)-n-propylamino]-
pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxypheiiyl)proß1onicacid;
[0068] 2-{2-diethylamino-5-[(4-fluorobenzenesulfonyl)allylamino]-
pyrunidin-4-ylamino}-3-(4-dimethylcarbamoyloxypheiiyl)proß1oiiicacid;
[0069] 2-{2-diethylamino-5-[(4-fluorobenzenesulfonyl)isobotylamino]-
pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)proß1onicacid;
[0070] 2-{2-diethylamino-5-[(4-fluorobenz;enesulfonyl)-n-butylamino]-
pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)proß1onicacid;
[0071] 2-{2-diethylamino-5-[(2,6-difluorobenzenesulfonyl)methylamino]-
pyrimidin-4-ylamino} -3-(4-dimethylcarbamoyloxyphenyl)proß1onic acid;
[0072] 2-{2-diethylamino-5-[(2,3-difluorobenzenesulfonyl)ethylamino]-
pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)proß1onicacid;
[0073] 2-{2-Diethylamino-5-[(4-fluorobenzenesulfonyl)propargylamino]
pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)proß1onicacid;
[0074] 2-{2-Diethylamino-5-[(2,4-difluorobenzenesulfonyl)propargylamino]
pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)proß1onicacid;
[0075] 2-{2-Diethylamino-5-[(4-fluorobenzenesulfonyl)-(2-trisfiuoroethyl)-
amino]pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)proß1onic
acid;
and pharmaceutically acceptable salts thereof.
[0076] In another aspect, this invention provides pharmaceutical
compositions comprising a pharmaceutically acceptable carrier and a
therapeutically effective amount of the compounds defined herein.
[0077] In one of its method aspects, this invention is directed to a method for
treating a disease mediated at least in part by a4 integrin, preferably VLA-4,
in a patient, which method comprises administering a pharmaceutical
composition comprising a pharmaceutically acceptable carrier and a
therapeutically effective amount of a compound of this invention.
[0078] The compounds and pharmaceutical compositions of this invention are
useful for treating disease conditions mediated at least in part by a4 integrins,
where the a4 integrin is preferably VLA-4 or leucocyte adhesion. Such
disease conditions include, by way of example, asthma, Alzheimer"s disease,
atherosclerosis, AIDS dementia, diabetes (including acute juvenile onset
diabetes), inflammatory bowel disease (including ulcerative colitis and
Crohn"s disease), multiple sclerosis, rheumatoid arthritis, tissue
transplantation, tumor metastasis, meningitis, encephalitis, stroke, and other
cerebral traumas, nephritis, retinitis, atoß1c dermatitis, psoriasis, myocardial
ischemia and acute leukocyte-mediated lung injury such as that which occurs
in adult resß1ratory distress syndrome.
[0079] Other disease conditions include, but are not limited to, inflammatory
conditions such as erythema nodosum, allergic conjunctivitis, optic neuritis,
uveitis, allergic rhinitis Anky losing spondylitis, psoriatic arthritis, vasculitis,
Reiter"s syndrome, systemic lupus erythematosus, progressive systemic
sclerosis, polymyositis, dermatomyositis, Wegner"s granulomatosis, aortitis,
sarcoidosis, lymphocytopenia, temporal arteritis, pericarditis, myocarditis,
congestive heart failure, polyarteritis nodosa, hypersensitivity syndromes,
allergy, hypereosinophilic syndromes, Churg-Strauss syndrome, chronic
obstructive pulmonary disease, hypersensitivity pneumonitis, chronic active
hepatitis, interstitial cystitis, autoimmune endocrine failure, primary biliary
cirrhosis, autoimmune aplastic anemia, chronic persistent hepatitis and
thyroiditis.
[0080] In a preferred embodiment, the disease condition mediated by a4
integrin is an inflammatory disease.
DETAILED DESCRIPTION OF THE INVENTION
[0081] As above, this invention relates to compounds which inhibit leukocyte
adhesion and, in particular, leukocyte adhesion mediated at least in part by a4
integrins, preferably VLA-4. However, prior to describingjhis invention in
further detail, the following terms will first be defined.
Definitions}
[0082] Unless otherwise stated, the following terms used in the specification
and claims have the meanings given below:
[0083] As used herein, "lower alkyl" refers to monovalent alkyl groups
having from 1 to 5 carbon atoms including straight and branched chain alkyl
groups. This term is exemplified by groups such as methyl, ethyl, iso-
propyl, n-propyl, n-butyl, iso-butyl, sec-butyl, r-butyl, n-pentyl and the like.
"Lower alkyl" may be optionally substituted with a halogen, such as chloro,
fluoro, bromo and the like.
[0084] The term "lower alkylene" refers to divalent alkylene groups of from
1 to 4 carbon atoms including straight and branched chain alkylene groups.
This term is exemplied by groups such as methylene, ethylene, n-propylene,
iso-propyiene (-CH2CH(CH3)- and -CH(CH3)CHr) and the like.
[0085] The term "lower alkynyl" refers to an alkynyl group preferably
having from 2 to 6 carbon atoms and having at least 1 site and preferably
only 1 site of alkynyl unsaturation (i.e., -OC). This term is exemplified by
groups such as acetyl (-OCH), propargyl (-CH2-OCH), 3-butynyl (-
CH2CH2OCH3) and the like.
[0086] The term "lower cycloalkyl" refers to cyclic alkyl groups of from 3 to
6 carbon atoms having a single cyclic ring including, by way of example,
cyclopropyl, cyclobutyl, cyclopentyl and cyclohexyl.
[0087] The term "lower alkylenecycloalkyl" refers to the group consisting of
a lower alkylene-lower cycloalkyl, as defined herein. Such groups are
exemplified by methylenecyclopropyl (-CH2-cyclopropyl),
ethylenecyclopropyl and the like.
[0088] "Pharmaceutically acceptable carrier" means a carrier that is useful in
preparing a pharmaceutical composition that is generally safe, non-toxic and
neither biologically nor otherwise undesirable, and includes a carrier that is
acceptable for veterinary use as well as human pharmaceutical use. "A
pharmaceutically acceptable carrier" as used in the specification and claims
includes both one and more than one such carrier.
[0089] "Treating" or "treatment" of a disease includes:
(1) preventing the disease, i.e., causing the clinical symptoms of the
disease not to develop in a mammal that may be exposed to or predisposed to
the disease but does not yet experience or display symptoms of the disease,
(2) inhibiting the disease, i.e., arresting or reducing the development
of the disease or its clinical symptoms, or
(3) relieving the disease, i.e., causing regression of the disease or its
clinical symptoms.
[0090] A "therapeutically effective amount" means the amount of a
compound that, when administered to a mammal for treating a disease, is
sufficient to effect such treatment for the disease. The "therapeutically
effective amount" will vary depending on the compound, the disease and its
severity and the age, weight, etc., of the mammal to be treated.
[0091] "Pharmaceutically acceptable salt" refers to pharmaceutically
acceptable salts of a compound of Formula I which salts are derived from a
variety of organic and inorganic counter ions well known in the art and
include, by way of example only, sodium, potassium, calcium, magnesium,
ammonium, tetraalkylammonium, and the like; and when the molecule
contains a basic functionality, salts of organic or inorganic acids, such as
hydrochloride, hydrobromide, tartrate, mesylate, acetate, maleate, oxalate
and the like.
[0092] Integrins are a large family of homologous transmembrane linker
proteins that are the principal receptors on animal cells for binding most
extracellular matrix proteins, such as collagen, fibronectin, and laminin. The
integrins are heterodimers comprised of an a chain and a ß chain. To date,
twenty different integrin heterodimers, made from 9 different a subunits and
14 different ß subunits, have been identified. The term " a 4 integrins" refers
to the class of heterodimer, enzyme-linked cell-surface receptors that contain
the a 4 subunit paired with any of the ß subunits. VLA-4 is an example of an
a 4 integrin, and is a heterodimer of the a 4 and ßt subunits, and is also
referred to as a4 ß, integrin.
Compound Preparation
[0093] The compounds of this invention can be prepared from readily
available starting materials using the methods and procedures set forth in the
examples below. These methods and procedures outline specific reaction
protocols for preparing N-[2-N",N"-diethyIainino-5-aminosulfonylphenyl-
yrimidin-4-yl]-p-carbomyloxy-phenylalanine compounds. Compounds within
the scope not exemplified in these examples and methods are readily prepared
by appropriate substitution of starting materials which are either
commercially available or well known in the art.
[0094] Other procedures and reaction conditions for preparing the
compounds of this invention are described in the examples set forth below.
Additionally, other procedures for preparing compounds useful in certain
aspects of this invention are disclosed in U.S. Patent 6,492,372; the
disclosure of which is incorporated herein by reference in its entirety.
Pharmaceutical Formulations
[0095] When employed as Pharmaceuticals, the compounds of this invention
are usually administered in the form of pharmaceutical compositions. These
compositions can be administered by a variety of routes including oral,
rectal, transdermal, subcutaneous, intravenous, intramuscular, and intranasal.
These compositions are effective by both injectable and oral delivery. Such
compositions are prepared in a manner well known in the pharmaceutical art
and comprise at least one active compound.
[0096] This invention also includes pharmaceutical compositions which
contain, as the active ingredient, one or more of the compounds of formula I-
VII above associated with pharmaceutically acceptable carriers. In making
the compositions of this invention, the active ingredient is usually mixed with
an exciß1ent, diluted by an exciß1ent or enclosed within such a carrier which
can be in the form of a capsule, sachet, paper or other container. The
excipient employed is tyß1cally an exciß1ent suitable for administration to
human subjects or other mammals. When the exciß1ent serves as a diluent, it
can be a solid, semi-solid, or liquid material, which acts as a vehicle, carrier
or medium for the active ingredient. Thus, the compositions can be in the
form of tablets, ß1lls, powders, lozenges, sachets, cachets, elixirs,
suspensions, emulsions, solutions, syrups, aerosols (as a solid or in a liquid
medium), ointments containing, for example, up to 10% by weight of the
active compound, soft and hard gelatin capsules, suppositories, sterile
injectable solutions, and sterile packaged powders.
[0097] In preparing a formulation, it may be necessary to mill the active
compound to provide the appropriate particle size prior to combining with the
other ingredients. If the active compound is substantially insoluble, it
ordinarily is milled to a particle size of less than 200 mesh. If the active
compound is substantially water soluble, the particle size is normally adjusted
by milling to provide a substantially uniform distribution in the formulation,
e.g., about 40 mesh.
[0098] Some examples of suitable exciß1ents include lactose, dextrose,
sucrose, sorbitol, mannitol, starches, gum acacia, calcium phosphate,
alginates, tragacanth, gelatin, calcium silicate, microcrystalline cellulose,
polyvinylpyrrolidone, cellulose, water, syrup, and methyl cellulose. The
formulations can additionally include: lubricating agents such as talc,
magnesium stearate, and mineral oil; wetting agents; emulsifying and
suspending agents; preserving agents such as methyl- and propylhydroxy-
benzoates; sweetening agents; and flavoring agents. The compositions of the
invention can be formulated so as to provide quick, sustained or delayed
release of the active ingredient after administration to the patient by
employing procedures known in the art.
[0099] The compositions are preferably formulated in a unit dosage form,
each dosage containing from about 5 to about 100 mg, more usually about 10
to about 30 mg, of the active ingredient. The term "unit dosage forms"
refers to physically discrete units suitable as unitary dosages for human
subjects and other mammals, each unit containing a predetermined quantity
of active material calculated to produce the desired therapeutic effect, in
association with a suitable pharmaceutical exciß1ent.
[00100] The active compound is effective over a wide dosage range
and is generally administered in a pharmaceutically effective amount. It, will
be understood, however, that the amount of the compound actually
administered will be determined by a physician, in the light of the relevant
circumstances, including the condition to be treated, the chosen route of
administration, the actual compound administered, the age, weight, and
response of the individual patient, the severity of the patient"s symptoms, and
the like.
[00101] For preparing solid compositions such as tablets, the principal
active ingredient is mixed with a pharmaceutical exciß1ent to form a solid
preformulation composition containing a homogeneous mixture of a
compound of the present invention. When referring to these preformulation
compositions as homogeneous, it is meant that the active ingredient is
dispersed evenly throughout the composition so that the composition may be
readily subdivided into equally effective unit dosage forms such as tablets,
ß1lls and capsules. This solid preformulation is then subdivided into unit
dosage forms of the type described above containing from, for example, 0.1
to about 500 mg of the active ingredient of the present invention.
[00102] The tablets or ß1lls of the present invention may be coated or
otherwise compounded to provide a dosage form affording the advantage of
prolonged action. For example, the tablet or ß1ll can comprise an inner
dosage and an outer dosage component, the latter being in the form of an
envelope over the former. The two components can be separated by an
enteric layer which serves to resist disintegration in the stomach and permit
the inner component to pass intact into the duodenum or to be delayed in
release. A variety of materials can be used for such enteric layers or
coatings, such materials including a number of polymeric acids and mixtures
of polymeric acids with such materials as shellac, cetyl alcohol, and cellulose
acetate.
[00103] The liquid forms in which the novel compositions of the
present invention may be incorporated for administration orally or by
injection include aqueous solutions suitably flavored syrups, aqueous or oil
suspensions, and flavored emulsions with edible oils such as cottonseed oil,
sesame oil, coconut oil, or peanut oil, as well as elixirs and similar
pharmaceutical vehicles.
[00104] Compositions for inhalation or insufflation include solutions
and suspensions in pharmaceutically acceptable, aqueous or organic solvents,
or mixtures thereof, and powders. The liquid or solid compositions may
contain suitable pharmaceutically acceptable exciß1ents as described supra.
Preferably the compositions are administered by the oral or nasal resß1ratory
route for local or systemic effect. Compositions in preferably
pharmaceutically acceptable solvents may be nebulized by use of inert gases.
Nebulized solutions may be breathed directly from the nebulizing device or
the nebulizing device may be attached to a face masks tent, or intermittent
positive pressure breathing machine. Solution, suspension, or powder
compositions may be administered, preferably orally or nasally, from devices
which deliver the formulation in an appropriate manner.
[00105] The following formulation examples illustrate the
pharmaceutical compositions of the present invention.
Formulation Example 1
[00106] Hard gelatin capsules containing the following ingredients are
prepared:
Quantity
Ingredient (mg/capsule)
Active Ingredient 30.0
Starch 305.0
Magnesium stearate 5.0
The above ingredients are mixed and filled into hard gelatin
capsules in 340 mg quantities.
Formulation Example 2
[00107] A tablet formula is prepared using the ingredients below:
Quantity
Ingredient (mg/tablet)
Active Ingredient 25.0
Cellulose, microcrystalline 200.0
Colloidal silicon dioxide 10.0
Stearic acid 5.0
[00108] The components are blended and compressed to form tablets, each
weighing 240 mg.
Formulation Example 3
[00109] A dry powder inhaler formulation is prepared containing the
following components:
Ingredient Weight %
Active Ingredient 5
Lactose 95
[00110] The active mixture is mixed with the lactose and the mixture is
added to a dry powder inhaling appliance.
Formulation Example 4
[00111] Tablets, each containing 30 mg of active ingredient, are prepared as
follows:
Quantity
Ingredient (mg/tahlet)
Active Ingredient 30.0 mg
Starch 45.0 mg
Microcrystalline cellulose 35.0 mg
Polyvinylpyrrolidone
(as 10% solution in water) 4.0 mg
Sodium carboxymethyl starch 4.5 mg
Magnesium stearate 0.5 mg
laic l.Q mg
Total 120 mg
[00112] The active ingredient, starch and cellulose are passed through a No.
20 mesh U.S. sieve and mixed thoroughly. The solution of polyvinyl-
pyrrolidone is mixed with the resultant powders, which are then passed
through a 16 mesh U.S. sieve. The granules so produced are dried at 50° to
60°C and passed through a 16 mesh U.S. sieve. The sodium carboxymethyl
starch, magnesium stearate, and talc, previously passed through a No. 30
mesh U.S. sieve, are then added to the granules which, after mixing, are
compressed on a tablet machine to yield tablets each weighing 120 mg.
Formulation Example 5
[00113] Capsules, each containing 40 mg of medicament are made as
follows:
Quantity
Ingredient (mg/capsule)
Active Ingredient 40.0 mg
Starch 109.0 mg
Magnesium stearate 1.0 mg
Total 150.0 mg
[00114] The active ingredient, starch, and magnesium stearate are blended,
passed through a No. 20 mesh U.S. sieve, and filled into hard gelatin
capsules in 150 mg quantities.
Formulation Example 6
[00115] Suppositories, each containing 25 mg of active ingredient are made
as follows:
Ingredient Amount
Active Ingredient 25 mg
Saturated fatty acid glycerides to 2,000 mg
[00116] The active ingredient is passed through a No. 60 mesh U.S. sieve
and suspended in the saturated fatty acid glycerides previously melted using
the minimum heat necessary. The mixture is then poured into a suppository
mold of nominal 2.0 g capacity and allowed to cool.
Formulation Example 7
[00117] Suspensions, each containing 50 mg of medicament per 5.0 mL dose
are made as follows:
Ingredient Amount
Active Ingredient 50.0 mg
Xanthan gum 4.0 mg
Sodium carboxymethyl cellulose (11%)
Macrocrystalline cellulose (89%) 50.0 mg
Sucrose 1.75 g
Sodium benzoate 10.0 mg
Flavor and Color q.v.
Purified water 5.0 mL
[00118] The medicament, sucrose and xanthan gum are blended, passed
through a No. 10 mesh U.S. sieve, and then mixed with a previously made
solution of the microcrystalline cellulose and sodium carboxymethyl cellulose
in water. The sodium benzoate, flavor, and color are diluted with some of
the water and added with stirring. Sufficient water is then added to produce
the required volume.
Formulation Example 8
Quantity
Ingredient (mg/capsule)
Active Ingredient 15.0 mg
Starch 407.0 mg
Magnesium stearate 3-0 nig
Total 425.0 mg
[00119] The active ingredient, starch, and magnesium stearate are blended,
passed through a No. 20 mesh U.S. sieve, and filled into hard gelatin
capsules in 425 mg quantities.
Formulation Example 9
[00120] An intravenous formulation may be prepared as follows:
Tngredient Quantity
Active Ingredient 250.0 mg
Isotonic Saline 1000 mL
Formulation Example 10
*
[00121] A toß1cal formulation may be prepared as follows:
Tngredient Quantity
Active Ingredient 1-10 g
Emulsifying Wax 30 g
Liquid Paraffin 20 g
White Soft Paraffin 2 to 100 g
[00122] The white soft paraffin is heated until molten. The liquid paraffin
and emulsifying wax are incorporated and stirred until dissolved. The active
ingredient is added and stirring is continued until dispersed. The mixture is
then cooled until solid.
[00123] Another preferred formulation employed in the methods of the
present invention employs transdermal delivery devices ("patches"). Such
transdermal patches may be used to provide continuous or discontinuous
infusion of the compounds of the present invention in controlled amounts.
The construction and use of transdermal patches for the delivery of
pharmaceutical agents is well known in the art. See, e.g., U.S. Patent
5,023,252, issued June 11, 1991, herein incorporated by reference. Such
patches may be constructed for continuous, pulsatile, or on demand delivery
of pharmaceutical agents.
[00124] Direct or indirect placement techniques may be used when it is
desirable or necessary to introduce the pharmaceutical composition to the
brain. Direct techniques usually involve placement of a drug delivery
catheter into the host"s ventricular system to bypass the blood-brain barrier.
One such implantable delivery system used for the transport of biological
factors to specific anatomical regions of the body is described in U.S. Patent
5,011,472 which is herein incorporated by reference.
[00125] Indirect techniques, which are generally preferred, usually involve
formulating the compositions to provide for drug latentiation by the
conversion of hydrophilic drugs into liß1d-soluble drugs. Latentiation is
generally achieved through blocking of the hydroxy, carbonyl, sulfate, and
primary amine groups present on the drug to render the drug more lipid
soluble and amenable to transportation across the blood-brain barrier.
Alternatively, the delivery of hydrophilic drugs may be enhanced by
intra-arterial infusion of hypertonic solutions which can transiently open the
blood-brain barrier.
Utility
[00126] The compounds of this invention inhibit, in vivo, adhesion of
leukocytes to endothelial cells mediated at least in part by a4 integrins, where
the a4 integrin is preferably VLA-4, by competitive binding to a4 integrins,
preferably VLA-4. Accordingly, the compounds of this invention can be
used in the treatment of mammalian diseases mediated by a4 integrins, where
the a4 integrin is preferably VLA-4, or leucocyte adhesion. Such diseases
include inflammatory diseases in mammalian patients such as asthma,
Alzheimer"s disease, atherosclerosis, AIDS dementia, diabetes (including
acute juvenile onset diabetes), inflammatory bowel disease (including
ulcerative colitis and Crohn"s disease), multiple sclerosis, rheumatoid
arthritis, tissue transplantation, tumor metastasis, meningitis, encephalitis,
stroke, and other cerebral traumas, nephritis, retinitis, atopic dermatitis,
psoriasis, myocardial ischemia and acute leukocyte-mediated lung injury such
as that which occurs in adult resß1ratory distress syndrome.
[00127] The amount administered to the mammalian patient will vary
depending upon what is being administered, the purpose of the
administration, such as prophylaxis or therapy, the state of the patient, the
manner of administration, and the like. In therapeutic applications,
compositions are administered to a patient already suffering from a disease in
an amount sufficient to cure or at least partially arrest the symptoms of the
disease and its complications. An amount adequate to accomplish this is
defined as "therapeutically effective dose." Amounts effective for this use
will depend on the disease condition being treated as well as by the judgment
of the attending clinician depending upon factors such as the severity of the
inflammation, the age, weight and general condition of the patient, and the
like.
[00128] The compositions administered to a patient are in the form of
pharmaceutical compositions described above. These compositions may be
sterilized by conventional sterilization techniques, or may be sterile filtered.
The resulting aqueous solutions may be packaged for use as is, or
lyophilized, the lyophilized preparation being combined with a sterile
aqueous carrier prior to administration. The pH of the compound
preparations tyß1cally will be between 3 and 11. more preferably from 5 to 9
and most preferably from 7 to 8. It will be understood that use of certain of
the foregoing exciß1ents, carriers, or stabilizers will result in the formation of
pharmaceutical salts.
[00129] The therapeutic dosage of the compounds of the present invention
will vary according to, for example, the particular use for which the
treatment is made, the manner of administration of the compound, the health
and condition of the patient, and the judgment of the prescribing physician.
For example, for intravenous administration, the dose will tyß1cally be in the
range of about 20 µg to about 500 µg per kilogram body weight, preferably
about 100 µg to about 300 µg per kilogram body weight. Suitable dosage
ranges for intranasal administration are generally about 0.1 pg to 1 mg per
kilogram body weight. Effective doses can be extrapolated from
dose-response curves derived from in vitro or animal model test systems.
[00130] The following synthetic and biological examples are offered to
illustrate this invention and are not to be construed in any way as limiting the
scope of this invention. Unless otherwise stated, all temperatures are in
degrees Celsius.
EXAMPLES
[00131] In the examples below, the following abbreviations have the
following meanings. If an abbreviation is not defined, it has its generally
accepted meaning.
AUC = Area under the curve
BSA = bovine serum albumin
DMAP = 4-N,N-limethylarriinopyridine
DMSO = dimethylsulfoxide
EDC = l-(3-dimethylaminopropyl)-3-
ethylcarbodiimide hydrochloride
EDTA = Ethylenediamine tetraacetic acid
EtOAc = ethyl acetate
EtOH = ethanol
bd = broad doublet
bs = broad singlet
d = doublet
m = multiplet
t = triplet
s = singlet
eq or eq. = equivalent
Example 1
Preparation of 2-{2-diethylamino-5-[(4-fluorohenzenesulfony1)Tnethylamino]
pyriniidin-4-v1amino}-3-(;4-dimethylcarhamoy1oxypheny1)propionic acid
[00132] General. Flash chromatography was performed using a Biotage
Flash 75L, using 800 g KP-Sil silica cartridges (32-63/xM, 60 angstrom, 500-
550 m2/g). R1S are reported for analytical thin layer chromatography, using
EM Science Silica Gel F(254) 250 µM thick plates for normal phase, and
Watman MKC18F 200 µM thick plates for reverse phase.
[00133] Step 1: Preparation of 2,4-Dichloro-5-nitropyrimidine.
5-Nitrouracil, was treated with phosphorous oxychloride and N,N-
dimethylaniline, according to the procedure of Whittaker (J. Chem. Soc.
1951, 1565), to give the title compound, which is also available from City
Chemical (West Haven, CT).
[00134] Step 2: Preparation of 2-(2-diethylamino-5-nitropyrimidin-4-
ylamino)-3-(4-hydroxyphenyl)proß1onic acid, r-butyl ester. To a solution of
2-amino-3-(4-hydroxyphenyl)proß1onic acid, (30.6 g, 0.129 mol) inTHF
(250 mL) at -10 °C was added 2,4-DichIoro-5-nitropyrimidine (25g, 0.129
mol), keeß1ng the temperature below 5 °C during the addition. Once the
addition was complete, N,N-diisopropylethylamine (33.7 mL, 0.194 mol)
was added dropwise. After stirring for 1 h at -10 °C, diethylamine (66.73
mL, 0.645 mol) was added slowly, and then the reaction mixture was
warmed to room temperature overnight. The reaction mixture was diluted
with diethyl ether (500 mL), and the organic layer was washed with 0.2 N
citric acid (3 x 150 mL), water (1 x 150 mL), and 10% K2CO3 (3 x 150 mL).
The organic phase was dried (Na2SO4), filtered, and concentrated in vacuo to
yield a yellow residue. The residue was purified by flash chromatography
(20% EtOAc/hexanes on silica gel) to yield 37.39 g (67%) the title
compound as a yellow foam. Rf=0.21 (25% EtOAc/hexanes on silica gel).
[00135] Step 3: Preparation of 2-(2-diethyIammo-5-nitropyrimidin-4-
ylamino)-3-(4-dimethylcarbamoyloxyphenyl)propionic acid /-butyl ester.
To a solution of 2-(2-diethylamino-5-nitropyrimidin-4-ylamino)-3-(4-
hydroxy-phenyl)propionic acid /-butyl ester (31.80 g, 0.074 mol) in CH2C12
(600 mL) was added DMAP (9.00 g, 0.074 mol). Afl:er 5 minutes
triethylamine (10.23 mL, 0.074 mol) was added dropwise. N,N-
dimethylcarbamyl chloride (13.83 mL, 0.110 mol) was added dropwise, and
the reaction was heated to reflux overnight. The reaction mixture was
concentrated in vacuo and taken up in EtOAc (1 L). The organic phase was
washed with 0.5 M citric acid (3 x 250 mL), sat. NaHCO3 (3 x 250 mL),
brine (1 x 250 mL), dried (MgSO4), filtered, and concentrated in vacuo to
yield 37.0 g (99%) the title compound as a white solid.
[00136] Step 4: Preparation of 2-(2-diethylamiuo-5-aminopyrimidin-4-
yIamino)-3-(4-dimethylcarbamoyloxyphenyl)propionic acid /-butyl ester.
A mixture of 2-(2-diemylarnino-5-nitropyrimidin-4-ylamino)-3-(4-
dimethylcarbamoyl-oxyphenyl)propionic acid /-butyl ester(37.0 g, 0.073 mol)
and 10% Pd/C (3.8 g, 10 wt% Pd) in EtOH (250 mL) was shaken under 60
psi hydrogen until TLC (50% EtOAc/hexanes on silica gel) showed 100%
conversion to product (48 hours). The reaction mixture was then filtered
through a Celite plug and concentrated in vacuo to yield 32.0 g (92%) the
title compound as a violet foam.
[00137] Step 5: Preparation of 2-{2-diethylamino-5-[(4-
fluorobenzenesulfonyl) amino]-pyrimidin-4-ylamino}-3-(4-
dimethylcarbamoyloxyphenyl) propionic acid /-butyl ester. A pyridine
(120 mL) solution of 2-(2-diemylanamo-5-aminopyrimidin-4-ylarnino)-3-(4-
dimethylcarbamoyloxy-phenyOpropionic acid /-butyl ester(32.0 g, 0.067 mol)
was cooled to -20 °C with a dry ice/CH3CN bath. The mixture stirred for 30
minutes, and then p-fluorobenzenesulfonyl chloride (13.18 g, 0.067 mol) was
added slowly. The reaction was stirred at -20 °C for 4.5 his, and then 3-
dimethylaminopropyl amine (8.52 mL, 0.067 mol) was added, and then the
mixture was allowed to warm to room temperature overnight. The reaction
was concentrated in vacua. The residue was taken up in EtOAc (1 L), and
the organic phase was washed with 0.5 M citric acid (3 x 900 mL), water (1
x 900 mL), sat. NaHCO3 (3 x 900 mL), brine (1 x 900 mL), dried (MgSO4),
filtered, and concentrated in vacuo to yield a brown residue. The residue
was purified by flash chromatography (50% EtOAc/hexanes on silica gel) to
yield 33.04g (77%) the title compound as a yellow foam. Rf= 0.54 (3:2
EtOAc/hexanes on silica gel).
[00138] Step 6: Preparation of 2-{2-diethylammo-5-[(4-
fluorobenzenesulfonyl) methylamino]-pyrimidin-4-ylamino}-3-(4-
dimethylcarbamoyloxyphenyl) propionic acid /-butyl ester. To a solution
of 2-{2^iemylan^o-5-[(4-fluorobenzenesuIfonyl)arnino]-pyrirnidin-4-
ylamino}-3-(4-dimethyl-carbamoyloxyphenyl)propionic acid /-butyl ester
(33.04 g, 0.052 mol) in acetone (510 mL) was added K2CO3, (8.69 g, 0.063
mol), and the mixture was stirred for 10 min at room temperature. Dimethyl
sulfate (5.95 mL, 0.063 mol) was then added slowly, and the reaction
mixture was stirred overnight at room temperature. The reaction mixture
was concentrated in vacuo, and the residue was taken up in EtOAc (600 mL).
The organic phase was washed with water (2 x 400 mL), brine (2 x 400 mL),
dried MgSO4, filtered, and concentrated in vacuo. The residue was purified
by flash chromatography (2:1 hexanes/EtOAc on silica gel) to yield 28.69 g
(85%) the title compound as a white solid.
[00139] Step 7: Preparation of 2-{2-diethylamino-5-[(4-
fluorobenzenesulfonyl) methyIamino]pyrimidin-4-ylamino}-3-(4-
dimethylcarbamoyloxyphenyl) propionic acid hydrochloride. A formic
acid (500 mL) solution of 2-{2-diethylamino-5-[(4-fluorobenzenesulfonyl)
methylamino]-pyrimidin-4-ylamino}-3-(4-dimethylcarbainoyloxyphenyl)
propionic acid /-butyl ester (28.69 g, 0.044 mol) was heated to 70 °C for 2 h,
and then concentrated in vacuo. The residue was dissolved again in formic
acid (500 mL), and then heated again at 70 °C for 2 h, and then concentrated
again in vacuo. The residue was dissolved again in formic acid (500 mL),
and then heated again at 70 °C for 1 h. The solution was reduced in volume
by 90%, and then treated with 1.0 M HC1 (44 mL, 0.044 mol) and distilled
water (490 mL). The resulting homogeneous solution was concentrated in
vacuo, and then distilled water (100 mL) was added, and the homogenous
solution was lyophilized over 14 days to yield 26.76 g (96%) the title
compound, as a white solid.
"H NMR (CD3OD) d 7.96-7.92 (m, 2H), 7.45-7.25 (m, 4H), 7.06-6.95 (m,
3H), 5.00-4.93 (m, 1H), 3.55-3.40 (m, 5H), 3.34-3.20 (m,, 2H), 3.15-3.05
(m, 5H), 3.07-3.00 (m, 3H), 1.22 (bs, 6H)
13C NMR (CD3OD) d 17L6, 168.3, 154.5, 144.4, 137.9, 135.1, 135.0,
134.1, 125.5, 120.6, 120.3, 39.6, 39.2, 39.1, 15.2
MS m/z 589 (MH+)
Example 2
Preparation of 2-{2-diethylamino-5-[(4-chlorobenzenesulfonyl)methylamino]
pyrimidin-4-ylamino}-3-(4-dimethylcarhamoyloxyphenyl)propionic acid
[00140] Steps 1, 2, 3, 4, 6 and 7 were performed as for Example 1. Step 5
was performed using 4-chlorobenzenesulfonyl chloride in place of 4-
fluorobenzenesulfonyl chloride.
"H NMR (CD3OD) d 7.88-7.85 (m, 2H), 7.72-7.69 (m, 2H), 7.39-7.25 (m,
2H), 7.14-6.92 (m, 3H), 5.00-4.85 (m, 1H), 3.60-3.50 (m, 1H), 3.37-3.28
(m, 6H), 3.15-3.07 (m, 6H), 3.01 (bs, 3H), 1.22 (bs, 6H)
13C NMR (CD3OD)d 208.6, 145.3, 134.9, 128.8, 124.9, 124.5, 124.4,
116.3,50.2,30.4,30.0,6.0
MS m/z 605(MH+)
Example 3
Preparation of 2-{2-diethyIamino-5-[(3,4-difluorohenzenesulfony1)
methy1amino]pyrimidin-4-ylamino}-3-(4-dimethylcarhamoyloxyphenyl)
propionic acid.
[00141] Steps 1, 2, 3, 4, 6 and 7 were performed as for Example 1. Step 5
was performed using 3,4-difluorobenzenesulfonyl chloride in place of 4-
fluorobenzenesulfonyl chloride.
1H NMR (CD3OD) d 7.84-7.77 (m, 1H), 7.67 (bs, 1H), 7.58-7.53 (m, 1H),
7.37-7.34 (m, 1H), 7.22-7.18 (m, 1H), 7.08-7.02 (m, 3H), 4.83-4.76 (m,
1H), 3.55-3.54 (m, 4H), 3.35-3.33 (m, 1H), 3.23-3.12 (m, 6H), 3.03-2.99
(m, 3H), 1.19(bs,6H)
13C NMR (CD3OD) d 178.3, 177.8, 163.2, 162.6, 159.3, 159.1, 155.9,
155.7, 154.3, 153.0, 152.5, 152.4, 138.4, 138.1, 134.0, 129.5, 125.3,
122.4, 122.2, 121.7, 121.4, 115.3, 59.3, 46.0, 42.4., 41.9, 40.4, 39.9, 39.2,
39.1, 15.76
MS m/z 607.2 (MH+)
Example 4
Preparation of 2-{2-diethylamino-5-[(3r4-dichlorohenzenesulfonyn
methy1amino]pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)
propionic acid
[00142] Steps 1, 2, 3, 4, 6 and 7 were performed as for Example 1. Step 5
was performed using 3,4-dichlorobenzenesulfonyl chloride in place of 4-
fluorobenzenesulfonyl chloride.
*H NMR (CD3OD) d 8.00-7.98 (m, 1H), 7.83-7.74 (m, 2H), 7.37-7.34 (m,
1H), 7.21-7.20 (m, 1H), 7.10-7.02 (m, 3H), 4.85-4.83 (m, 1H), 3.55-3.53
(m, 2H), 3.35-3.33 (m, 1H), 3.21-3.12 (m, 6H), 3.04-2.99 (m, 6H), 1.19
(bs, 6H)
l3C NMR (CD3OD) d 176.4, 166.2, 161.7, 161.2, 158.0, 157.8, 152.8,
151.5, 150.5, 140.2, 139.8, 139.5, 136.8, 135.8, 133.9, 132.6, 132.0,
129.8, 123.8, 113.7, 113.4, 57.8, 44.6, 40.8, 40.4, 38.7, 38.3, 37.7, 37.5,
14.1
MS m/z 639.1 (MH+)
Example 5
Preparation of 2-{2-diethylamino-5-[(benzenesulfonyl)rnethylainino]
pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxypheny1)propionic acid
[00143] Steps 1, 2, 3, 4, 6 and 7 were performed as for Example 1. Step 5
was performed using benzenesulfonyl chloride in place of 4-
fluorobenzenesulfonyl chloride.
1H NMR (CD3OD) d 8.14 (bs, 1H), (7.85-7.84 (m, 1H), 7.8-7.78 (m, 1H),
7.69-7.66 (mf 2H), 7.40-7.37 (m, 1H), 7.21-7.195 (m, 1H), 7.04-7.03 (m,
2H), 7.95-7.90 (m, 1H), 5.52 (bs, 1H), 3.54-3.53 (m, 2H), 3.36-3.33 (m,
6H), 3.13-3.12 (m, 3H), 3.01-3.00 (m, 3H), 1.20-1.17 (m, 6H)
13C NMR (CD3OD) d 165.9, 152.8, 136.7, 135.8, 132.6, 131.6, 130.2,
123.8, 44.7, 37.5, 14.0
MS m/z 571.2 (MH+)
Example 6
Preparation of 2-{2-diethylamino-5-[(2-fluorobenzenesulfonyl)methylaniino]
pyrimidin-4-y1amino}-3-(4-dimethylcarbamoyloxyphenyl)propionic acid
[00144] Steps 1, 2, 3, 4, 6 and 7 were performed as for Example 1. Step 5
was performed using 2-fluorobenzenesulfonyl chloride in place of 4-
fluorobenzenesulfonyl chloride.
1H NMR (CD3OD) d 8.31 (bs, 1H), 7.94-7.85 (m, 2H), 7.57-7.44 (m, 3H),
7.34-7.30 (m, 1H), 7.15-7.12 (m, 2H), 5.00-4.85 (1H), 3.63-3.62 (m, 4H),
3.50-3.42 (m, 1H), 3.34-3.30 (m, 4H), 3.29-3.22 (m, 4H), 3.11-3.10 (m,
2H), 1.28 (bs, 6H)
13C NMR (CD3OD) d 176.5, 166.4, 163.1, 160.4, 159.7, 157.7, 152.8,
151.5, 150.7, 138.5, 138.3, 136.7, 133.7, 132.5, 132.2, 127.1, 123.7,
119.9, 119.6, 113.4, 57.8, 44.6, 40.6, 39.0, 38.4, 37.7, 37.5, 14.1
Example 7
Preparation of 2-{2-diethylamino-5-[(3-fluorobenzenesulfonyl)
methylamino]pyrimidin-4-y1amino}-3-(4-dimethyIcairhamoyIoxypheny1|)
propionic acid
[00145] Steps 1, 2, 3, 4, 6 and 7 were performed as for Example 1. Step 5
was performed using 3-fluorobenzenesulfonyl chloride in place of 4-
fluorobenzenesulfonyl chloride.
1H NMR (CD3OD) d 8.15-8.12 (bs, 1H), 7.72-7.68 (m, 1H), 7.63-7.60 (m,
1H), 7.53-7.52 (m, 1H), 7.38-7.35 (m, 1H), 7.21-7.20 (m, 1H), 7.10-6.99
(m, 3H), 4.87-4.86 (m, 1H), 3.54-3.53 (m, 4H), 3.35-3.34 (m, 3H), 3.15-
3.12 (m, 4H), 3.05-3.00 (m, 4H), 1.20 (bs, 6H)
l3C NMR (CD3OD)d 166.1, 153.1, 136.9, 134.1, 132.8, 126.5, 124.1,
123.2, 122.9, 117.7, 117.4, 103.4, 45.0, 38.0, 14.3
MS m/z 589.2 (MH+)
Example 8
Preparation of 2-{2-diethylamino-5-[(4-fluorobenzenesulfony|)
isopropylamino]pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)
propionic acid
[00146] Steps 1, 2 and 3 were performed as for Example 1. Thereafter,
Steps 4 and 6 were accomplished in one pot, according to the following
procedure. Thereafter, Steps 5 and 7 were performed as for Example 1.
1H NMR (CD3OD) d 8.20-8.16 (m, 1H), 7.95-7.84 (m, 2H), 7.36-7.25 (m,
3H), 7.24-7.15 (m, 3H), 7.07-6.98 (m, 3H), 5.07-5.05 (m, 1H), 4.90-4.86
(m, 1H), 4.65-4.62 (m, 1H), 4.49-4.41 (m, 1H), 3.63-3.56 (m, 3H), 3.38-
3.31 (m, 2H), 3.27-3.11 (m, 2H), 3.00-2.99 (m, 3H), 1.27-1.21 (m, 6H),
1.05-0.99 (m, 6H)
13C NMR (CD3OD) d 175.8, 175.5, 169.6, 166.3, 165.9, 163.5, 163.4,
157.7, 153.0, 152.9, 152.3, 138.1, 136.4, 136.1, 133.1, 133.0, 133.0,
132.9, 132.7, 132.3, 123.8, 118.8, 118.7, 118.5, 118.4, 107.5, 57.6, 57.2,
54.7, 44.7, 38.7, 38.1, 37.6, 37.5, 23.0, 22.9, 22.2, 22.0, 14.1, 14.0
[00147] Alternative one-pot procedure for the preparation of 2-(2-
diethylamino-5-isopropylaminopyrimidin-4-yl)-3-(4-
dimethylcarbamoyloxypkenyl) propionic acid /-butyl ester. A mixture of
2-(2-diethylamino-5-nitropyrimidin-4-ylamino)-3-(4-
dimethylcarbamoyloxyphenyl)propionic acid r-butyl ester (5.0 g, 0.010 mol),
glacial acetic acid (10 drops), acetone (2.19 mL, 0.030 mol), and platinum
oxide (0.250 g, 5 wt%) in EtOH (15 mL) was hydrogenated at 45 psi
hydrogen until TLC (50% EtOAc/hexanes) showed 100% conversion to
product (20 hours). The reaction mixture was then filtered through a Celite
plug and concentrated in vacuo to yield a brown residue. The residue was
purified by flash chromatography (4:1 EtOAc/hexanes) to yield 3.54 g (70%)
9 as a purple foam.
Example 9
Preparation of 2-{2-diethylamino-5-[(4-fluorobenzenesulfonyl)ethylamino]
pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)propionic acid
[00148] Steps 1, 2, 3, 4, 5 and 7 were performed as for Example 1. Step 6
was performed using ethyl iodide in place of dimethyl sulfate.
1H NMR (CDC13) d 0.89 (t, J = 7.2, 1.8H), 1.06 (t,J = 7.1, 1.2H), 1.10-
1.30 (m, 6H), 2.97 (s, 3H), 3.05 (s, 3H), 3.10-3.90 (m, 8H), 4.82 (q, J =
5.4, 0.6H), 4.91 (q, J = 6.1, 0.4H), 6.80-7.45 (m, 8H), 7.77 (m, 2H),
12.44 (bs, 1H)
MS m/z 603.3 (MH+)
Example 10
Preparation of 2-{2-diethyIamino-5-[(3,4-difluorobenzenesuIfonyl)
isopropylamino]pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)
propiortic acid
[00149] Steps 1, 2, 3, 4, 6 and 7 were performed as for Example 8. Step 5
was performed using 3,4-difluorobenzenesulfonyl chloride in place of 4-
fluorobenzenesulfonyl chloride.
1H NMR (CD3OD) d 8.20-8.19 (m, 1H), 7.84-7.78 (m, 1H), 1.10-1.64 (m,
1H), 7.54-7.48 (m, 1H), 7.39-7.31 (m, 1H), 7.20-7.17 (m, 1H), 7.05-6.96
(m, 2H), 4.91-4.89 (m, 1H), 4.70-4.68 (m, 1H), 4.48-4.41 (m, 2H), 3.60-
3.58 (m, 3H), 3.34-3.33 (m, 1H), 3.27-3.20 (m, 1H), 3.09-3.08 (m, 2H),
2.98-2.97 (m, 2H), 1.28-1.19 (m, 6H), 1.06-0.98 (m, 6H), 0.83-0.81 (m,
1H)
13C NMR (CD3OD) d 177.6, 177.2, 167.9, 164.9, 164.8, 159.2, 159.1,
155.7, 154.5, 154.4, 152.4, 152.3, 140.4, 140.3, 137.8, 134.3, 133.9,
129.3, 129.2, 125.4, 122.6, 122.5, 122.4, 122.2, 121.5, 121.2, 109.1, 59.5,
59.1, 56.7, 56.6, 46.4, 46.3, 39.6, 39.3, 39.2, 24.7, 24.5, 23.9, 23.6, 15.7,
15.6
Example 11
Preparation of 2-{2-diethylamino-5-[(4-chlorobenzenesulfonyl)
isopropylamino]pyrimidin-4-ylaminQ}-3-(4-dimethylcarbamoyloxyphenyl)
propionic acjd
[00150] Steps 1, 2, 3, 4, 6 and 7 were performed as for Example 8. Step 5
was performed using 4-chlorobenzenesulfonyI chloride in place of 4-
fluorobenzenesulfonyl chloride.
1H NMR (CD3OD) d 8.18-8.17 (m, 1H), 7.85-7.78 (m, 1H), 7.62-7.58 (m,
1H), 7.38-7.35 (m, 1H), 7.34-7.24 (m, 1H), 7.17-7.16 (m, 1H), 7.10-7.05
(m, 2H), 7.04-6.98 (m, 2H), 4.98-4.87 (m, 1H), 4.73-4.68 (m, 1H), 4.55-
4.38 (m, 2H), 3.70-3.52 (m, 3H), 3.40-3.30 (m, 1H), 3.28-3.18 (m, 1H),
3.17-3.08 (m, 2H), 3.05-2.98 (m, 2H), 1.25-1.20 (m, 6H), 1.04-0.96 (m,
6H), 0.80-0.77 (m, 1H)
13C NMR (CD3OD) d 175.7, 175.5, 166.2, 165.8, 169.6, 163.5, 163.4,
157.6, 152.9, 152.8, 138.0, 136.3, 136.1, 133.1, 133.0, 132.9, 132.7,
132.2, 123.8, 118.8, 118.6, 118.5, 118.5, 118.3, 107.5, 57.6, 57.2, 54.7,
44.6, 38.6, 38.1, 37.6, 37.5, 22.9, 22.8, 22.2, 21.9, 14.1, 13.9
Example 12
Preparation of 2-{2-diethylamino-5-[(3,4-difluorohenzenesuifonyn
ethylamino]pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyIoxyphenyl)
propionic acid
[00151] Steps 1, 2, 3, 4, 6 and 7 were performed as for Example 9. Step 5
was performed using 3,4-difiuorobenzensulfonyl chloride in place of 4-
fluorobenzenesulfonyl chloride.
1H NMR (CD3OD) d 8.15-8.14 (m, 1H), 7.80-7.75 (m, 1H), 7.73-7.62 (m,
1H), 7.60-7.49 (m, 1H), 7.30-7.18 (m, 1H), 7.16-7.00 (m, 2H), 5.58-5.50
(m, 1H), 4.90-4.83 (m, 1H), 5.78-5.70 (m, 1H), 3.85-3.75 (m, 1H), 3.65-
3.54 (m, 3H), 3.40-3.23 (m, 5H), 3.18-3.10 (m, 3H), 3.05-2.98 (m, 3H),
1.25-1.15 (m, 3H), 1.18-1.05 (t, 1.5H), 1.02-1.00 (t, 1.5H)
13C NMR (CD3OD) d 165.8, 152.7, 145.7, 136.4, 136.3, 132.5, 132.2,
127.5, 123.6, 120.7, 120.4, 81.4, 57.0, 44.3, 38.5, 38.1, 37.4, 14.9, 14.6,
14.1, 14.0
MS m/z 621.5 (MH+)
Example 13
Preparation of 2-{2-diethylamino-5-[(4-ch1orobenzenesulfony1)ethylamino]
pyrimidin-4-ylaTnino}-3-r4-dimethylcarhamoyloxyphenynpropionicacid
[00152] Steps 1, 2, 3, 4, 6 and 7 were performed as for Example 9. Step 5
was performed using 4-chlorobenzenensulfonyl chloride in place of 4-
fluorobenzenesulfonyl chloride.
1H NMR (CD3OD) d 8.15-8.14 (m, 1H), 7.84-7.79 (m, 1H), 7.67-7.61 (m,
1H), 7.37-7.33 (m, 1H), 7.22-7.18 (m, 1H), 7.14-7.13 (m, 1H), 7.06-7.00
(m, 3H), 4.80-4.75 (m, 1H), 4.18-4.10 (m, 1H), 3.65-3.30 (m, 3H), 3.28-
3.20 (m, 3H), 3.18-3.08 (m, 2H), 3.03-2.98 (m, 2H), 2.05-2.04 (m, 1H),
1.30-1.16 (m, 9H), 1.10-1.08 (t, 1.5H), 0.99-0.95 (t, 1.5H)
13C NMR (CD3OD) d 176.2, 176.1, 166.7, 162.7, 162.3, 157.6, 152.9,
142.0, 138.8, 136.5, 132.8, 132.5, 132.0, 131.8, 123.8, 111.7, 111.4,57.9,
57.8, 44.9, 38.9, 38.3, 37.8, 37.7, 15.1, 14.9, 14.3, 14.2
MS m/z 619 A (MH+)
Example 14
Preparation of 2-{2-djethylamino-5-[(4-fluorobenzenesulfonyl)
cylclopropy1methyIamino]pyrimidin-4-ylamino}-3-(4-dimethylcarhamoyl-
oxyphenyl)propionic acid
[00153] Steps 1, 2, 3, 4, 5 and 7 were performed as for Example 1. Step 6
was performed using bromomethylcyclopropane and cesium carbonate in
place of dimethyl sulfate and potassium carbonate.
lH NMR (CDC13) d -0.2-0.2 (m, 2.4H), 0.2-0.45 (m, 1.6H), 0.54 (m,
0.6H), 0.85 (m, 0.4H), 1.00-1.40 (m, 6H), 2.80-3.80 (m, 14H), 4.79 (q, J
= 5.5, 0.6H), 4.91 (q, / = 6.3, 0.4H), 6.70-7.40 (m, 8H), 7.77 (m, 2H),
10.26 (bs, 1H)
MS m/z 629.2 (MH+)
Example IS
Preparation of 2-{2-diethylamino-5-[(3r5-difluorobenzenesulfonyl)
methylamino] pyrimidm-4-ylamino}-3-(4-dimethylcarbamoy1oxyphey1)
propionic acid
[00154] Steps 1, 2, 3, 4, 6 and 7 were performed as for Example 1. Step 5
was performed using 3,5-difluorobenzenesulfonyl chloride in place of 4-
fluorobenzenesulfonyl chloride.
1H NMR (CD3OD) d 7.68-7.67 (m, 1H), 7.67-7.56 (m, 2H), 7.42-7.40 (m,
2H), 7.31-7.30 (m, 1H), 7.26-7.23 (m, 2H), 5.20-4.90 (m, 1H), 4.35-4.33
(m, 1H), 3.78-3.74 (m, 4H), 3.57-3.54 (2H), 3.38-3.33 (m, 2H), 3.26-3.21
(m, 2H), 2.41-2.39 (m, 2H), 2.26-2.25 (m, 2H), 1.50-1.38 (m, 6H)
I3C NMR (CD3OD) d 162.5, 162.3, 159.2, 159.0, 148.0, 146.1, 132.2,
127.8, 127.7, 127.6, 118.9, 109.1, 109.0, 108.7, 108.6, 106.2, 105.8,52.5,
39.6,34.1,32.9,9.5
Example 16
Preparation of 2-{2-diethylamino-5-[(3,5-difluorobenzenesiiIfonyl)
ethylamino]pyrimidin-4-ylamino}-3-(4-dimethylrarbamoyloxyphenyI)
propionic acid
[00155] Steps 1,2, 3, 4, 5 and 7 were performed as for example 15. Step 6
was performed using ethyl iodide in place of dimethyl sulfate.
1H NMR (CD3OD) d 7.45-7.43 (m, 1H), 7.42-7.18 (m, 2H), 7.21-7.16 (m,
2H), 7.07-7.06 (m, 1H), 7.04-6.97 (m, 2H), 5.51 (bs, 1H), 4.86-4.82 (m,
1H), 4.72-4.66 (m, 1H), 3.84-3.77 (m, 1H), 3.59-3.50 (m, 3H), 3.34-3.31
(m, 2H), 3.12-3.10 (m, 3H), 2.99-2.96 (m, 3H), 1.22-1.14 (m, 9H), 1.10-
1.05 (t, 1.5H), 0.97-0.95 (t, 1.5H)
13C NMR (CD3OD) d 159.9, 150.9, 150.1, 134.0, 130.0, 129.7, 121.2,
107.9, 86.7, 42.0, 41.9, 36.3, 35.2, 35.1, 12.8, 12.5, 11.9, 11.8,
Example 17
Preparation of 2-{2-diethylamino-5-[(2.4-difluoroben7enesnlfonyl)
methylamino]pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxypheny])
propionic acid
[00156] Steps 1, 2, 3, 4, 6 and 7 were performed as for Example 1. Step 5
was performed using 2,4-difluorobenzenesulfonyl chloride in place of 4-
fluorobenzenesulfonyl chloride.
1H NMR (CD3OD) d 8.16-8.11 (m, 1H), 7.59-7.56 (m, 2H), 7.48-7.45 (m,
2H), 7.26-7.24 (m, 3H), 5.21-5.16 (m, 1H), 3.79-3.77 (m, 4H), 3.57-3.54
(m, 3H), 3.48-3.46 (m, 2H), 3.44-3.34 (m, 3H), 3.22-3.21 (m, 3H), 1.45-
1.44 (m,6H)
13C NMR (CDC1) d 180.2, 170.3, 166.6, 150.3, 129.0, 128.9, 128.7, 125.9,
125.4, 117.5, 117.4, 116.5, 114.8, 107.7, 107.4, 95.5, 90.8, 68.0, 65.1,
55.7, 50.8, 37.6, 36.4, 31.9, 31.7, 31.6, 13.2, 9.4, 8.3, 7.8
Example 18
Preparation of 2-{2-diethylamino-5-[(2r4-difhiorobenzenesulfonyI)
ethylamino] pyrimidin-4-ylamino}-3-(4-dimethyIcarhamoyloxyphenyl)
propionic acid
[00157] Steps 1, 2, 3, 4, 5 and 7 were performed as for Example 17. Step 6
was performed using ethyl iodide in place of dimethyl sulfate.
1H NMR (CD3OD) d 8.15 (bs, 1H), 7.91-7.76 (m, 1H), 7.32-7.30 (m, 2H),
7.20-7.19 (m, 2H), 7.04-7.00 (m, 2H), 4.84-4.83 (m, 1H), 4.74-4.67 (m,
1H), 4.14-4.07 (m, 1H), 3.92-3.82 (m, 1H), 3.51-3.49 (m, 3H), 3.34-3.31
(m, 3H), 3.12-2.99 (m, 2H), 2.98-2.97 (m, 2H), 2.03-2.02 (m, 1H), 1.26-
1.17 (m, 6H), 1.10-1.06 (t, 1.5H), 1.03-0.98 (t, 1.5H)
13C NMR (CD3OD) d 173.6, 173.3, 171.4, 167.7, 164.3, 161.2, 159.9,
159.3, 157.1, 156.7, 155.2, 152.4, 151.0, 150.3, 134.0, 133.3, 133.1,
132.9, 130.0, 123.2, 122.9, 122.8, 121.3, 121.2, 112.0, 111.8, 111.6,
111.5, 107.7, 107.2, 106.0, 105.9, 105.6, 105.2, 60.0, 54.8, 42.0, 36.5,
35.9, 35.3, 35.1, 19.3, 13.0, 12.9, 12.7, 11.9, 11.8
Example 19
Preparation of 2-{2-diethyIamino-5-[(3 f 5-dichlorobenzenesulfonyl)
methylamino]pyriini(iin-4-ylamino}-3-(4-dimethylcarhamoyloxyphenyn
propionic acid
[00158] Steps 1, 2, 3, 4, 6 and 7 were performed as for Example 1. Step 5
was performed using 3,5-dichlorobenzenesulfonyl chloride in place of 4-
fluorobenzenesulfonyl chloride.
1H NMR (CD3OD) d 7.84-7.82 (m, 1H), 7.76-7.75 (m, 3H), 7.34-7.32 (m,
1H), 7.19-7.10 (m, 1H), 7.03-7.00 (m, 2H), 5.50 (bs, 1H), 4.83-4.82 (m,
1H), 4.74-7.73 (m, 1H), 3.55-3.38 (m, 4H), 3.34-3.32 (m, 2H), 3.15-3.11
(m, 4H), 3.02-2.99 (m, 3H), 1.18-1.15 (m, 6H)
13C NMR (CD3OD)d 157.1, 155.2, 150.1, 149.7, 140.1, 135.9, 134.3,
132.9, 130.0, 129.9, 126.0, 121.2, 110.7, 55.2, 54.8, 42.0, 38.5, 38.1,
36.5,35.9,35.2,35.1, 11.9
MS m/z 639.1 (MH+)
Example 20
Preparation of 2-{2-diethy1amino-5-(Y3r5-riir,h1orohenzenesulfonyl)
ethyIamino]pyrimidin-4-ylamino}-3-(4-dirnethylcarbamQyloxyphenyl)
propionic acid
[00159] Steps 1, 2, 3, 4, 5 and 7 were performed as for Example 19. Step 6
was performed using ethyl iodide in place of dimethyl sulfate.
1H NMR (CD3OD) d 8.15 (bs, 1H), 7.84-7.84-7.79 (m, 1H), 1.16-1.1 A (m,
2H), 7.33-7.30 (m, 1H), 7.22-7.11 (m, 2H), 7.04-6.98 (m, 1H), 5.51 (bs,
1H), 4.86-4.82 (m, 1H), 4.72-4.67 (m, 1H), 3.77-3.75 (m, 1H), 3.60-3.50
(m, 3H), 3.34-3.29 (m, 2H), 3.27-3.22 (m, 2H), 3.12-3.11 (m, 2H), 2.99-
2.98 (m, 2H), 1.23-1.14 (m, 6H), 1.10-1.05 (t, 1.5H), 0.99-0.94 (t, 1.5H)
l3C NMR (CD3OD)d 173.6, 173.4, 163.7, 159.9, 159.3, 157.3, 156.8,
155.2, 155.1, 152.1, 150.8, 150.2, 141.4, 141.2, 135.9, 134.0, 132.7,
130.0, 129.7, 125.8, 125.7, 121.3, 121.2, 107.9, 107.4, 54.8, 54.7, 42.0,
36.4, 35.8, 35.3, 35.1, 12.8, 12.5, 11.9, 11.8
MS m/z 653.2 (MH+)
Example 21
Preparation of 2-{2-diethylarnino-5-[ propylamino] pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)
propionic acid
[00160] Steps 1, 2, 3, 4, 5 and 7 were performed as for Example 1. Step 6
was performed using 1-propyl iodide in place of dimethyl sulfate.
1H NMR (CDC13) d 0.75 (m, 3H), 1.00-1.50 (m, 8H), 3.00 (s, 3H), 3.08 (s,
3H), 3.20-3.70 (m, 8H), 4.79 (q, / = 6.3, 0.6H), 4.91 (q, / = 6.6, 0.4H),
5.73 (bs, 0.6H), 5.92 (bs, 0.4H), 6.90-7.45 (m, 7H), 7.76 (m, 2H)
MS m/z 617.2 (MH+)
Example 22
Preparation of 2-{2-diefhylamino-5-[(4-fluorobenzenesnlfnnyna1ly1amino]
pyrimidin-4-ylamino}-3-(4-dimethy1carbamoyloxyphenynpropionic acid
[00161] Steps 1, 2, 3, 4, 5 and 7 were performed as for Example 1. Step 6
was performed using allyl bromide in place of dimethyl sulfate.
1H NMR (CDC13) d 1.20 (m, 6H), 2.98 (s, 3H), 3.06 (s, 3H), 3.10-4.30
(m, 8H), 4.75- 4.95 (m, 1H), 5.07 (m, 2H), 5.48 (m, 0.6H), 5.67 (m,
0.4H), 6.90-7.45 (m, 8H), 7.76 (m, 2H), 11.07 (bs, 1H)
MS m/z 615.2 (MH+)
Example 23
Preparation of 2-{2-diethylamino-5-[(4-fluorobenzenesulfonyl)isobotylamino]
pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)propionic acid
[00162] Steps 1, 2, 3, 4, 5 and 7 were performed as for Example 1. Step 6
was performed using isobutyl iodide in place of dimethyl sulfate.
MS m/z 631.2 (MH+)
Example 24
Preparation of 2-{2-diefhy1amino-5-[(4-fluorobenzenesuilfonyl)-n-burylamino]
pyrimidin-4-ylamino}-3-(4-dimethylcarhamoyloxyphenyl)propionic acid
[00163] Steps 1, 2, 3, 4, 5 and 7 were performed as for Example 1. Step 6
was performed using 1-butyl iodide in place of dimethyl sulfate.
1H NMR (CDC13) d o.82 (q, / = 7.1, 3H), 1.05-1.40 (m, 10H), 3.01 (s,
3H), 3.10 (s, 3H), 3.15-3.80 (m, 8H), 4.75 (q, J = 6.3, 0.6H), 4.91 (q, /
= 5.9, 0.4H), 5.79 (d, / = 5.4, 0.6H), 5.91 (d, J = 6.6, 0.4H), 7.00-7.40
(m, 7H), 7.77 (m, 2H)
Example 25
Preparation of 2-{2-djethylamino-5-[(2 r5-difluorobenzenesulfony 1)
methylamino] pyrimidin-4-ylamino}-3-(4-dimethylcarhamoyIoxyphenyI)
propionic acid
[00164] Steps 1, 2, 3, 4, 6 and 7 were performed as for Example 1. Step 5
was performed using 2,6-difluorobenzenesulfonyl chloride in place of 4-
fluorobenzenesulfonyl chloride.
1H NMR (CD3OD) d 8.38-8.37 (m, 1H), 7.99-7.95 (m, 1H), 7.55-7.54 (m,
2H), 1.50-1 A2 (m, 2H), 7.27-7.22 (m, 2H), 5.08-5.06 (m, 1H), 3.76-3.74
(m, 4H), 3.59-3.54 (m, 3H), 3.49-3.42 (m, 4H), 3.36-3.34 (m, 2H), 3.23-
3.21 (m,2H), 1.40 (bs, 6H)
l3C NMR (CD3OD)d 161.4, 159.2, 155.8, 153.1, 148.1, 147.1, 133.6,
132.0, 127.8, 119.0, 111.1, 110.8, 110.7, 108.5, 105.8, 94.8, 86.4, 66.7,
54.0, 52.8, 39.7, 35.8, 34.2, 33.7, 32,9, 32.8, 9.4
Example 26
Preparation of 2-{2-diethylamino-5-[(2r3-difhiorobenzenesulfonyl)
ethyiamino] pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)
propionic. acid.
[00165] Steps 1, 2, 3, 4, 6 and 7 were performed as for Example 1. Step 5
was performed using 2,3-difluorobenzenesulfonyl chloride in place of 4-
fluorobenzenesulfonyl chloride. 2,3-Difluorobenzenesulfonyl chloride was
prepared by the following procedure.
1H NMR (CD3OD) d 8.32 (bs, 1H), 7.90-7.80 (m, 2H), 7.59-7.48 (m, 3H),
7.27-7.23 (m, 2H), 5.09-5.08 (m, 1H), 3.77-3.70 (m, 4H), 3.60-3.51 (m,
3H), 3.50-3.42 (m, 2H), 3.39-3.31n (m, 3H), 3.32-3.18 (m, 2H), 1.43-1.41
(m, 6H)
13C NMR (CD3OD) d 170.4, 160.8, 158.1, 156.1, 153.0, 151.6, 150.5,
148.9, 148.2, 147.3, 147.2, 143.9, 143.5, 142.6, 141.1, 140.9, 131.8,
127.7, 125.1, 123.8, 120.8, 120.6, 119.2, 40.5, 35.7, 33.4, 32.9, 32.7, 9.0
[00166] Preparation of 2,3-Difluorobenzenesulfony] Chloride. The
following procedure was executed using two flasks. In the first flask, 2,3-
difluoroaniline (2.0 g, 0.015 mol) was dissolved in concentrated HC1 (15.9
mL), and the resulting solution was cooled to -5 °C, using an ice/NaCl bath.
A solution of sodium nitrite (1.18 g, 0.017 mol) in distilled water (13.6 mL)
was added in portions with stirring, while maintaining the temperature below
0 °C, and the mixture was stirred for 10 min. In the second flask, thionyl
chloride (5.08 mL, 0.069 mol) was added dropwise to distilled water (30.6
mL), which had been pre-cooled to -5 °C, using an ice/NaCl bath. The
resulting solution was allowed to warm to room temperature, and then
Cu(I)Cl (0.08 g, 0.77 mmol) was added, and then the reaction mixture was
re-cooled to -5 °C. With continued cooling and stirring, the contents of the
first flask were added in 2 mL portions to the contents of the second flask,
and the mixture was stirred for 30 min, during which time a precipitate
formed. The precipitate was isolated by filtration, rinsed with cold water,
and stored under vacuum to give 3.25 g (98%) 10 as a white solid.
Example 27
Preparation of 2-{2-Diethylamino-5-[(4-fluorohenzenesulfonyI)
propargylamino]pyrimjdin-4-ylamino}-3-(4-dimethylcarhamoyloxyphenyl)
propionic acid
[00167] Steps 1, 2, 3, 4, 5 and 7 were performed as for Example 1. Step 6
was performed using propargyl bromide in place of dimethyl sulfate.
lH NMR (CDCl3) d 1.15 (m, 6H), 2.27 (d, J = 2.1, 1H), 2.97 (s, 3H), 3.06
(s, 3H), 3.10-3.70 (m, 6H), 3.75 (dd, / =17.7, 2.0, 0.6H), 3.95 (dd, / =
18.1, 2.0, 0.4H), 4.51 (dd, / =19.5, 2.2, 0.6H), 4.54 (dd, / = 18.1, 2.2,
0.4H), 4.79 (q, / = 5.9, 0.6H), 4.88 (q, / = 6.6, 0.4H), 6.42 (bd, 0.4H),
6.65 (bs, 0.6H), 6.85-7.30 (m, 6H), 7.52 (s, 0.6H), 7.56 (s, 0.4H), 7.85 (m,
2H), 8.20 (bs, 1H)
MS m/z 613.2 (MH+)
Example 28
Preparation of 2-{2-Diethylamino-5-[(2t4-
difluorobenzenesulfonyt)propargylamino]pyrimidin-4-ylarnino}-3-(4-
dimethylcarbamoyloxyphenyPpropionic acid
[00168] Steps 1, 2, 3, 4, 5 and 7 were performed as for Example 17. Step 6
was performed using propargyl bromide in place of dimethyl sulfate.
1H NMR (CDC13) d 1.16 (q, / = 7.5, 6H), 2.27 (m, 1H), 2.99 (s, 3H), 3.09
(s, 3H), 3.10-3.70 (m, 6H), 4.04 (dd, / =17.7, 2.4, 0.6H), 4.24 (dd, J =
17.9, 2.2, 0.4H), 4.47 (m, 1H), 4.81 (q, / = 5.9, 0.6H), 4.89 (q, / = 6.3,
0.4H), 6.27 (d, / = 7.5, 0.4H), 6.41 (d, J = 5.7, 0.6H), 6.90-7.10 (m,
4H), 7.16 (d, J = 8.3, 1H), 7.28 (d, / = 8.3, 1H), 7.55 (bs, 1H), 7.66 (s,
0.6H), 7.67 (s, 0.4H), 7.81 (m, 1H)
Example 29
Preparation of
2-{2-diethylamino-5-[(4-fluorobenzenesulfonyl)-(2,2r2-trifluoroethyl)amino]p
yrimidin-4-ylamino}-3-(4-dimethylcarbamoyl-oxyphenyl)propionic acid
[00169] Steps 1, 2, 3, 4, 5 and 7 were performed as for Example 1. Step 6
was performed using 2,2,2-trifluoroethyl triflate and cesium carbonate in
place of dimethyl sulfate and potassium carbonate.
lR NMR (CDC13) 6 1.14 (m, 6H), 2.98 (s, 3H), 3.06 (s, 3H), 3.10-4.20 (m,
8H), 4.80 (q, J = 5.9, 0.6H), 4.87 (q, J = 6.2, 0.4H), 6.09 (d, J = 5.9,
0.4H), 6.18 (bd, 0.6H), 6.80-7.50 (m, 7H), 7.55 (bs, 1H), 7.77 (m, 2H);
MS m/z 657.2 (MH+)
Example A
a4ß1 Iategrin Adhesion Assay:
Jurkat™ Cell Adhesion to Human Plasma Fibronectin
Procedure:
[00170] 96 well plates (Costar 3590 EIA plates) were coated with human
fibronectin (Gibco/BRL, cat #33016-023) at a concentration of 10 /Ag/mL
overnight at 4°C. The plates were then blocked with a solution of bovine
serum albumin (BSA; 0.3%) in saline. Jurkat™ cells (maintained in log phase
growth) were labeled with Calcein AM according to the manufacturer"s
instructions, and suspended at a concentration of 2 x 106 cells/mL in
Hepes/Saline/BSA. The cells were then exposed to test and control
compounds for 30 minutes at room temperature before transfer to individual
wells of the fibronectin coated plate. Adhesion was allowed to occur for 35
minutes at 37°C. The wells were then washed by gentle aspiration and
pipetting with fresh saline. Fluorescence associated with the remaining
adherent cells was quantified using a fluorescence plate reader at EX 485/EM
530.
[00171] Cell cultures were prepared by first splitting the stationary phase
Jurkat™ cells at 1:10 on day one, and 1:2 on day two to perform assay on
day 3. The cells split 1:10 on day one were split 1:4 on day 3 for a day 4
assay.
[00172] The assay plates were prepared by first making a working solution
of Gibco/BRL Human Fibronectin (cat # 33016-023) In PBS+ + , at 10
µg/mL.
A Costar 3590 EIA plate was then coated with 50 µL/well for 2 hours at
room temperature (thought it can also be left overnight at 4 °C). Finally the
plate was asperated and blocked with Hepes/Saline Buffer, 100 µL/well, for 1
hour at RT followed by washing three times with 150 µL of PBS+ +.
[00173] Compound dilutions were accomplished by preparing 1:3 serial
dilutions of compounds as follows. For each plate (4 compounds/plate) 600
µL were added to 4 Bio-Rad Titertubes in a Titertube rack. Enough
compound was added to each appropriate tube to give a 2X concentration
using methods well known in the art. Using Falcon Fiexiplates, 100 µL of
Hepes/Saline buffer or human serum were added to rows B through G. A
multi-channel pipetter set to 180 µL was used to with four tips spaced evenly
the pipetter. Each set of four tubes was mixed 5 times and 180 µL of 2X
compound was transferred to the first column of each compound dilution in
Row B, leaving Row A empty. 180 µL were added to the other wells in Row
A. Serial dilutions were performed down the plate by transferring 50 µL to
the next dilution and mixing 5 times, changing tips each time after mixing.
Dilutions were stopped at Row F. Row G had no compound present.
[00174] A 20 µg/mL solution in Hepes/Saline buffer or human serum, of
21/6 antibody was the positive control and was set aside in a reagent trough
to add to cell suspension plate.
[00175] The cell staining was accomplished by first harvesting the log-phase
Jurkat™ cells by centrifugation in 50 mL tubes (1100 rpm for 5 minutes).
The cells were resuspended in 50 mL PBS + +, spun, and resuspend in 20
mL PBS+ +. The ceils were stained by adding 20 µL of Calcein AM for 30
minutes R.T. The volume was brought to 50 mL with Hepes/Saline buffer
and the cells were counted, spun, and resuspend to 2 x 106 cells/mL in
Hepes/Saline buffer or human serum.
[00176] The compounds were incubated using the following procedure. In
a new flexiplate, 65 µL of stained cells were added to Rows B through H.
Then 65 pL of 2X compounds were added to the appropriate rows following
the plate setup and mixed 3X. 65µL of 2X-21/6 antibody were added to
Row H and mixed 3X. Finally the plate was incubated at room temperature
for 30 minutes.
[00177] Fibronectin adhesion was measured using a fluorescent plate reader
at EX 485/EM 530 after the following work up procedure. After incubation,
the cells were mixed 3X and 100 µL were transferred to the Fibronectin
coated plates and incubated at 37 °C for about 35 minutes. Each plate was
washed, row by row, by gently pipetting 100 µL of R.T. PBS+ + down the
sides of the wells and turning the plate 90 degrees to aspirate. This procedure
was repeated for a total of 3 washes. Each well was filled with 100 µL after
washing by pipetting down the side of the well.
[00178] An IC50 value was calculated for each compound, both in the
presence of the human serum and in the absence of human serum. IC50 is
concentration at which the growth or activity is inhibited by 50%. The data is
presented in the following tables.
Cell Adhesion to Human Plasma Fibronectin
(Without the human serum)
Example B
In vitro Saturation Assay For Determining Binding of
Candidate Compounds to a4ß1p,
[00181] The following describes an in vitro assay to determine the plasma
levels needed for a compound to be active in the Experimental Autoimmune
Encephalomyelitis ("EAE") model, described in the next example, or in other
in vivo models.
[00182] Log-growth Jurkat cells are washed and resuspended in normal
animal plasma containing 20 µg/mL of the 15/7 antibody (Yednock, et al.,J.
Biol. Chem., (1995) 270(48):28740).
[00183] The Jurkat cells are diluted two-fold into either normal plasma
samples containing known candidate compound amounts in various
concentrations ranging from 66 µM to 0.01 µM, using a standard 12 point
serial dilution for a standard curve, or into plasma samples obtained from the
peripheral blood of candidate compound-treated animals.
[00184] Cells are then incubated for 30 minutes at room temperature,
washed twice with phosphate-buffered saline ("PBS") containing 2% fetal
bovine serum and lmM each of calcium chloride and magnesium chloride
(assay medium) to remove unbound 15/7 antibody.
[00185] The ceils are then exposed to phycoerythrin-conjugated goat F(ab")2
anti-mouse IgG Fc (Immunotech, Westbrook, ME), which has been adsorbed
for any non-specific cross-reactivity by co-incubation with 5% serum from
the animal species being studied, at 1:200 and incubated in the dark at 4°C
for 30 minutes.
[00186] Cells are washed twice with assay medium and resuspended in the
same. They are then analyzed with a standard fluorescence activated cell
sorter ("FACS") analysis as described in Yednock et al. J. Biol. Chem.,
1995, 270:28740.
[00187] The data is then graßhed as fluorescence versus dose, e.g., in a
normal dose-response fashion. The dose levels that result in the upper
plateau of the curve represent the levels needed to obtain efficacy in an in
vivo model.
[00188] This assay may also be used to determine the plasma levels needed
to saturate the binding sites of other integrins, such as the a9ß1 pi integrin,
which is the integrin most closely related a4ß1 (Palmer et al, 1993, J. Cell
Bio., 123:1289). Such binding is predictive of in vivo utility for
inflammatory conditions mediated by a9ß1, integrin, including by way of
example, airway hyper-responsiveness and occlusion that occurs with chronic
asthma, smooth muscle cell proliferation in atherosclerosis, vascular
occlusion following angioplasty, fibrosis and glomerular scarring as a result
of renal disease, aortic stenosis, hypertrophy of synovial membranes in
rheumatoid arthritis, and inflammation and scarring that occur with the
progression of ulcerative colitis and Crohn"s disease.
[00189] Accordingly, the above-described assay may be performed with a
human colon carcinoma cell line, SW 480 (ATTC #CCL228) transfected with
cDNA encoding a9 integrin (Yokosaki et al., 1994, J. Biol. Chem.,
269:26691), in place of the Jurkat cells, to measure the binding of the a9ß1
integrin. As a control, SW 480 cells which express other a and ß1 subunits
may be used.
[00190] Accordingly, another aspect of this invention is directed to a method
for treating a disease in a mammalian patient, which disease is mediated by
a9ß1, and which method comprises administering to said patient a
therapeutically effective amount of a compound of this invention. Such
compounds are preferably administered in a pharmaceutical composition
described herein above. Effective daily dosing will depend upon the age,
weight, condition of the patient which factors can be readily ascertained by
the attending clinician. However, in a preferred embodiment, the compounds
are administered from about 20 to 500 µg/kg per day.
Example C
Cassette Dosing and Serum Analysis
for determination of Bioavailability
[00191] The oral bioavailability was screened by dosing rats with a cassette,
i.e. mixture of 6 compounds per dosing solution. The cassette included 5 test
articles and a standard compound, for a total dose of 10 mg/kg. Each
compound/test article was converted to the sodium salt with equimolar 1 N
NaOH and dissolved in water at 2 mg/mL. The cassette was prepared by
mixing equal volumes of each of the six solutions. The cassette dosing
solution was mixed well and then the pH was adjusted to 7.5-9. The dosing
solution was prepared the day before the study and stirred overnight at room
temperature.
[00192] Male Sprague Dawley (SD) rats from Charles River Laboratories, 6-
8 weeks old were used in this screen. Rats were quarantined for at least one
day and had continuous access to food and water. On the night before the
administration of the cassette, the rats were fasted for aßproximately 16 h.
[00193] Four SD rats were assigned in each cassette. A single dose of the
dosing solution was administered orally to each rat. The dosing volume (5
mL/kg) and time were recorded and rats were fed 2 h after dosing.
[00194] Blood samples were collected via cardiac puncture at the following
time points: 4 h, 8 h and 12 h. Immediately prior to blood collection, rats
were anesthetized with CO2 gas within 10-20 seconds. After the 12-hour
samples were collected, the rats were euthanized via CO2 asphyxiation
followed by cervical dislocation.
[00195] Blood samples were kept in heparinized microtainer tubes under
sub-ambient temperature (4 °C) before they were processed. Blood samples
were centrifuged (10000 rpm for 5 minutes) and plasma samples were
removed and stored in a -20 °C freezer until analyzed for drug levels. Drug
levels in the plasma were analyzed using the following protocol for direct
plasma precipitation.
[00196] The in vivo plasma samples were prepared in a 1.5 mL 96-well
plate, by adding, in order, 100 µL of the test plasma, 150 µL of methanol,
followed by vortexing for 10-20 seconds. 150 µL of 0.05 ng//µL of an
Internal Standard in acetonitrile were added and vortexed for 30 seconds.
[00197] The standard curve samples were prepared in a 1.5 mL 96-well
plate, by adding, in order, 100 µL of control mouse plasma, followed by 150
µL of methanol and vortexing for 10-20 seconds. 150 µL of 0.05 ng/µL of
an Internal Standard in acetonitrile were added and vortexed for 30 seconds.
The samples were spiked with 0-200 ng (10 concentrations) of the compound
of interest in 50% methanol to obtain a standard curve range of 0.5 ng/mL -
2,000 ng/mL. Again, the sample was vortexed for 30 seconds.
[00198] The samples were then spun for 20-30 minutes at 3000 rpm in an
Eppendorf microfuge before 80-90% of supernatant was transferred into a
clean 96-well plate. The organic solvent was then evaßorated until the
samples were dry (under N2 at 40°C/ 30-60 min (ZymarkTurbovaß)).
[00199] The residue was then dissolved in 200 - 600 L mobile phase (50%
CH3OH/0.1% TFA). LC/MS/MS was then run using a PE-Sciex aßi-3000
triple quadurpole mass spectrometer (SN0749707), Perkin-Elmer,
Series200auto-sampler, and shimadzu 10A pump. Acquisition was done with •
PE-Sciex Analyst (vl.l) and data analysis and quantification were
accomplished using PE-Sciex Analyst (vl.l). A 5-50 µL sample volume
was injected onto a reverse phase ThermoHypersil DASH-18 column
(Keystone 2.0 x 20 mm, 5 µm, PN: 8823025-701) using a mobile phase of
25% CH3OH, 0.1% TFA-100% CH3OH, 0.1% TFA. The run time was
about 8 minutes at a flow rate of about 300 µL/minutes.
[00200] The Area Under the Curve (AUC) was calculated using the linear
traßezoidal rule from t=0 to the last sampling time t, (see Handbook of Basic
Pharmacokinetics, Wolfgang A. Ritschel and Gregory L. Kearns, 5th ed,
1999).
AUC0?tx = S((Cn + Cn+1)/2)) • (tn+1 - tr) [µg/mL)h]
[00201] In the case of the cassette dosing paradigm, samples at 4, 8 and 12 h
post extravascular dosing, the AUC was calculated from t = 0 to t = 12 h.
The AUC°?12h values were calculated for each individual animal and the
average AUC°?12h are reported in the table below.
Example D
Asthma Models
[00203] Inflammatory conditions mediated by a4ß1 integrin include, for
example, eosinophil influx, airway hyper-responsiveness and occlusion that
occurs with chronic asthma. The following describes animal models of
asthma that were used to study the in vivo effects of the compounds of this
invention for use in treating asthma.
Rat Asthma Model
[00204] Following the procedures described by Chaßman et al, Am J. Resp.
Crit. Care Med,. 153 4, A219 (1996) and Chaßman et al, Am. J. Resp. Crit
Care Med 155:4, A881 (1997), both of which are incorporated by reference
in their entirety. Ovalbumin (OA; 10 µg/mL) were mixed with aluminum
hydroxide (10 mg/mL) and injected (i.p.) in Brown Norway rats on day 0.
Injections of OA, together with adjuvant, were repeated on days 7 and 14.
On day 21, sensitized animals were restrained in plastic tubes and exposed
(60 minutes) to an aerosol of OA (10 mg/kg) in a nose-only exposure system.
Animals will be sacrificed 72 hours later with pentobarbital (250 mg/kg,
i.p.). The lungs were lavaged via a tracheal cannula using 3 aliquots (4 mL)
of Hank"s solution (HBSS x 10, 100 mL; EDTA 100 mM, 100 mL; HEPES
1 M, 25 mL; made up to 1 L with H20); recovered cells were pooled and the
total volume of recovered fluid adjusted to 12 mL by addition of Hank"s
solution. Total cells were counted (Sysmex microcell counter F-500, TOA
Medical Electronics Otd., Jaßan) and smears were made by diluting
recovered fluid (to aßproximately 106 cells/mL) and pipetting an aliquot (100
µL) into a centrifuge (Cytospin, Shandon, U.K.). Smears were air dried,
fixed using a solution of fast green in methanol (2 mg/mL) for 5 seconds and
stained with eosin G (5 seconds) and thiazine (5 seconds) (Diff-Quick,
Browne Ltd. U.K.) in order to differentiate eosinophils, neutrophils,
macrophages and lymphocytes. A total of 500 cells per smear were counted
by light microscopy under oil immersion (x 100). Compounds of this
invention were formulated into a 0.5% carboxymethylcellulose and 2%
Tween80 suspension and administered orally to rats which had been
sensitized to the allergen, ovalbumin. Compounds which inhibited allergen-
induced leucocyte accumulation in the airways of actively sensitized Brown
Norway rats were considered to be active in this model.
Mouse Asthma Model
[00205] Compounds were also evaluated in a mouse model of acute
pulmonary inflammation following the procedures described by, Kung et ai.,
Am J. Respir. Cell Mol. Biol. 13:360-365, (1995) and Schneider etal.,
(1999). Am J. Respir. Cell Mol. Biol. 20:448-457, (1999), which are each
incorporated by reference in their entirety. Female Black/6 mice (8-12 weeks
of age) were sensitized on day 1 by an intraßeritoneal injection (i.p.) of 0.2
mL ova/alum mixture containing 20 µg of ova (Grade 4, Sigma) and 2 mg
inject Alum (pierce). A booster injection was administered on day 14. Mice
are challenged on days 28 and 29 with aerosolized 1 % ova (in 0.9% saline)
for 20 minutes. Mice are euthanized and bronchaveolar lavage samples (3
mL) are collected on day 30, 48 hours post first challenge. Eosinophils were
quantified by a FACs/FITC staining method. Compounds of this invention
were formulated into a 0.5% carboxymethylcellulose and 2% Tween80
suspension and administered orally to mice which had been sensitized to the
allergen, ovalbumin. Compounds which inhibited allergen-induced leucocyte
accumulation in the airways of actively sensitized C57BL/6 mice were
considered to be active in this model.
Sheep Asthma Model
[00206] This model employs the procedures described by Abraham et al.,
J.Clin, Invest, 93:776-787 (1994) and Abraham et al., Am J. Respir Crit
Care Med 156:696-703 (1997), both of which are incorporated by reference
in their entirety. Compounds of this invention have been evaluated by
intravenous (saline aqueous solution), oral (2% Tween80, 0.5%
carboxymethylcellulose), and aerosol administration to sheep which are
hypersensitive to Ascaris suum antigen. Compounds which decrease the early
antigen-induced bronchial response and/or block the late-phase airway
response, e.g. have a protective effect against antigen-induced late responses
and airway hyper-responsiveness ("AHR"), are considered to be active in this
model.
[00207] Allergic sheep which are shown to develop both early and late
bronchial responses to inhaled Ascaris suum antigen were used to study the
airway effects of the candidate compounds. Following topical anesthesia of
the nasal passages with 2% lidocaine, a balloon catheter was advanced
through one nostril into the lower esophagus. The animals were then
incubated with a cuffed endotracheal tube through the other nostril with a
flexible fiberoptic bronchoscope as a guide.
[00208] Pleural pressure was estimated according to Abraham (1994).
Aerosols (see formulation below) were generated using a disposable medical
nebulizer that provided an aerosol with a mass median aerodynamic diameter
of 3.2 µm as determined with an Andersen cascade impactor. The nebulizer
was connected to a dosimeter system consisting of a solenoid valve and a
source of compressed air (20 psi). The output of the nebulizer was directed
into a plastic T-piece, one end of which was connected to the inspiratory port
of a piston respirator. The solenoid valve was activated for 1 second at the
beginning of the inspiratory cycle of the respirator. Aerosols were delivered
at VT of 500 mL and a rate of 20 breaths/minute. A 0.5 % sodium
bicarbonate solution only was used as a control.
[00209] To assess bronchial responsiveness, cumulative concentration-
response curves to carbachol was generated according to Abraham (1994).
Bronchial biopsies were taken prior to and following the initiation of
treatment and 24 hours after antigen challenge. Bronchial biopsies were
preformed according to Abraham (1994).
[00210] An in vitro adhesion study of alveolar macrophages were also
performed according to Abraham (1994), and a percentage of adherent cells
calculated.
Aerosol Formulation
[00211] A solution of the candidate compound in 0.5% sodium
bicarbonate/saline (w/v) at a concentration of 30.0 mg/mL is prepared using
the following procedure:
Procedure:
1. Add 0.5g sodium bicarbonate into a 100 mL volumetric flask.
2. Add approximately 90.0 mL saline and sonicate until
dissolved.
3. Q.S. to 100.0 mL with saline and mix thoroughly.
[00213] B. Preparation of 30.0 mg/mL Candidate Compound: 10.0 mL
Procedure:
1. Add 0.300 g of the candidate compound into a 10.0 mL volumetric flask.
2. Add approximately 9.7 mL of 0.5% sodium bicarbonate / saline stock
solution.
3. Sonicate until the candidate compound is completely dissolved.
4. Q.S. to 10.0 mL with 0.5% sodium bicarbonate / saline stock solution
and mix thoroughly.
Example E
10-Day Toxicity Study on C57B6 Mice
[00214] A 10-day study was conducted to evaluate the toxicity of compounds
of the present invention to female C57B6 mice. The compound was
administered by gavage at five dose levels, 0 (vehicle control), 10, 30, 100,
300 and 1000 mg/kg (mpk), with five mice in each dose level. The dose
volume for all levels was 10 mL/kg. Dose solutions or suspensions were
prepared in 2% Tween 80 in 0.5% carboxymethyl cellulose (CMC) and new
dose solutions or suspensions were prepared every two - three days. In-life
observations included body weights (study day 1, 2, 3, 5, 7, 8 and 11), daily
cageside clinical observations (1-2/day) and periodic (study day -1, 2 and 9)
functional observation battery.
[00215] At termination, blood samples were collected by cardiac puncture
for clinical pathology (hematology and clinical chemistry) and drug levels.
The EDTA blood samples were analyzed for total white blood cell count, red
blood cell count, hemoglobin, hematocrit, erythrocyte indices (MCV, MCH,
MCHC), platelets and a WBC five part differential (neutrophil, lymphocytes,
monocytes, eosinophils and basophils). Heparinized plasma samples were
analyzed for alanine transaminase, aspartate transaminase, alkaline
phosphatase, total bilirubin, albumin, protein, calcium, glucose, urea
nitrogen, creatinine, cholesterol and triglycerides.
[00216] After blood collection, the carcass was necropsied and organs (liver,
spleen, kidneys, heart and thymus) were weighed. Tissue samples; brain,
salivary glands, thymus, heart, lung, liver, kidney, adrenal spleen, stomach,
duodenum, ileum, colon and uterus/ovary, were collected and formalin fixed.
Tissues from the vehicle control and 300 and 1000 mpk group animals were
processed to H & E stained glass slides and evaluated for histopathological
lesions.
[00217] Body weight changes, absolute and relative organ weights and
clinical pathology results were analyzed for statistical significant differences
compared to the vehicle controls by Dunnet"s multiple comparison test using
Prism software. The functional observation battery results were analyzed for
differences using the Dunnet"s, Fisher"s exact tests and dose trend effects by
the Cochran-Mantel-Haenszel correlation test using SAS software.
[00218] Using a conventional oral formulation, compounds of this invention
would be active in this model.
Example F
Adjuvant-Induced Arthritis in Rats
[00219] Adjuvant induced arthritis ("ALA") is an animal model useful in the
study of rheumatoid arthritis (RA), which is induced by injecting M.
tuberculosis in the base of the tail of Lewis rats. Between 10 and 15 days
following injection, animals develop a severe, progressive arthritis.
[00220] Generally, compounds are tested for their ability to alter hind paw
swelling and bone damage resulting from adjuvant-induced edema in rats. To
quantitate the inhibition of hind paw swelling resulting from AIA, two phases
of inflammation have been defined: (1) the primary and secondary injected
hind paw, and (2) the secondary uninjected hind paw, which generally begins
developing about eleven days from the induction of inflammation in the
injected paw. Reduction of the latter type of inflammation is an indication of
immunosuppressive activity. Cf. Chang, Arth. Rheum., 20, 1135-1141
(1977).
[00221] Using an animal model of RA, such as AIA, enables one to study
the cellular events involved in the early stages of the disease. CD44
expression on macrophages and lymphocytes is up-regulated during the early
development of adjuvant arthritis, whereas LFA-1 expression is up-regulated
later in the development of the disease. Understanding the interactions
between adhesion molecules and endothelium at the earliest stages of adjuvant
arthritis could lead to significant advances in the methods used in the
treatment of RA.
WE CLAIM:
1. A compound of formula (I):
wherein each X is independently fluoro, chloro or bromo;
p is 0 or an integer from 1-3;
R1 is selected from the group consisting of methyl and ethyl;
R2 is selected from the group consisting of monovalent (C1 to C5) lower alkyl,
monovalent (C2 to C3) lower alkenyl, and lower alkylene cycloalkyl, where lower alkylene refers
to a divalent (C1 to C4) lower alkylene group which is bound to cycloalkyl which refers to a
monovalent (C3 to C6) cycloalkyl group
and pharmaceutical ly acceptable salts thereof.
2. A compound of Formula II:
wherein each X is independently selected from the group consisting of fluoro and
chloro,
m is an integer equal to 1 or 2;
R2 is selected from the group consisting of monovalent (C1 to C5) lower alkyl,
monovalent (C2 to C3) lower alkenyl, and lower alkylene cycloalkyl, where lower alkylene refers
to a divalent (C1 to C4) lower alkylene group which is bound to cycloalkyl which refers to a
monovalent (C3 to C6) cycloalkyl group
and pharmaceutically acceptable salts thereof.
wherein each X is independently fluoro or chloro;
n is zero or one;
R2 is -CH2-R" where R" is selected from the group consisting of hydrogen, methyl or
-CH=CH2;
and pharmaceutically acceptable salts thereof.
4. A compound as claimed in any one of claims 1, 2 or 3, wherein R2 is CH3.
5. A compound as claimed in claim 3, wherein X is F or Cl, and n is 0.
6. A compound selected from the group consisting of:
2-{2-diethylamino-5-[(4-chlorobenzenesulfonyl)tnethylatnino]-pyritnidin-4-ylamino}-3-
(4-ditnethylcarbamoyloxyphenyl)propionic acid;
2-{2-dietliylamino-5-[(4-fluorobenzenesulfonyl)methylainino]-pyrimidin-4-ylainino}-3-
(4-dimethylcarbatnoyloxyphenyl)propionic acid;
2-{2-diethylamino-5-[(3,4-difluorobenzenesulfonyl)methylamino]-pyrimidin-4-ylamino
} -3-(4-dimethylcarbamoyloxyphenyl)propionic acid;
2-{2-diethylamino-5-[(3,4-dichlorobenzenesulfonyl)methylamino]-pyrimidin-4-ylamiao
}-3-(4-dimethylcarbamoyloxyphenyl)propionic acid;
2-{2-diethylamino-5-[(benzenesulfonyl)methylamino]-pyrimidin-4-ylamino}-3-(4-dime
thylcarbamoyloxyphenyl)propionic acid;
2-{2-diethylamino-5-[(2-fluorobenzeaesulfonyl)methylamino]-pyritnidin-4-ylamino}-3-
(4-dimethylcarbamoyloxyphenyl)propionic acid;
2-{2-diethylamino-5-[(3-fluorobenzenesulfonyl)methylamino]-pyrimidin-4-ylamino}-3-
(4-dimethylcarbamoyloxyphenyl)propionic acid;
2-{2-diethylamino-5-[(4-fluorobenzenesulfonyl)isopropylamino]-pyrimidin-4-ylamino}-
3-(4-dimethylcarbamoyloxyphenyl)propionic acid;
2-{2-diethylamino-5-[(4-fluorobenzenesulfonyl)ethylamino]-pyrimidin-4-ylamino}-3-(
4-dimethylcarbamoyloxyphenyl)propionic acid;
2-{2-diethylamino-5-[(3,4-difluorobenzenesuIfonyl)isopropylatnino]-pyrimidin-4-ylatni
no}-3-(4-dimethylcarbamoyloxyphenyl)propionic acid;
2-{2-diethylamino-5-[(4-chlorobenzenesulfonyl)isopropylamino]-pyrimidin-4-ylamino}-
3-(4-ditnethylcarbamoyloxyphenyl)propionic acid;
2-{2-diethyIamino-5-[(3,4-difluorobenzenesulfonyl)ethylamino]-pyrimidin-4-yIamino}-
3-(4-dimethylcarbamoyloxyphenyI)propionic acid;
2-{2-diethylamino-5-[(4-chlorobenzenesulfonyl)ethylamino]-pyrimidin-4-ylamino}-3-(
4-dimethylcarbamoyloxyphenyl)propionic acid;
2-{2-diethylamino-5-[(4-fluorobenzenesulfonyl)cylclopropylmethyl-amino]pyrimidin-4-
ylamino}-3-(4-dimethylcarbamoyloxyphenyl)propionic acid;
2-{2-diethylamino-5-[(3,5-difluorobenzenesulfonyl)methylamino]-pyrimidin-4-ylamino
} -3-(4-dimethylcarbamoyloxyphenyl)propionic acid
2-{2-diethyIamino-5-[(3,5-difluorobenzenesuIfonyl)ethylamino]-pyrimidin-4-ylamino}-
3-(4-dimethylcarbamoyloxyphenyl)propionic acid;
2-{2-diethylamino-5-[(2,4-difluorobenzenesulfonyl)methylamino]-pyrimidin-4-ylamino
} -3-(4-dimethylcarbamoyloxyphenyl)propionic acid;
2-{2-diethylamino-5-[(2,4-difluorobenzenesulfonyl)ethylamino]-pyrimidin-4-ylamino}-
3-(4-dimethylcarbarnoyloxyphenyl)propionic acid;
2-{2-diethylamino-5-[(3,5-dichlorobenzenesulfonyl)tnethylamino]-pyrimidin-4-ylamino
}-3-(4-dimethylcarbamoyloxyphenyl)propionic acid;
2-{2-diethylamino-5-[(3,5-dichlorobenzenesulfonyl)ethylamino]-pyrimidin-4-ylamino}-
3-(4-dimethylcarbamoyloxyphenyl)propionic acid;
2-{2-diethylamino-5-[(4-fluorobenzenesulfonyl)-n-propylamino]-pyrimidin-4-ylamino}-
3-(4-dimethylcarbamoyloxyphenyl)propionic acid;
2-{2-diethylamino-5-[(4-fluorobenzenesulfonyl)allylami]no]-pyrimidin-4-ylamino}-3-(4-
dimethylcarbamoyloxyphenyl)propionic acid;
2-{2-diethylamino-5-[(4-fluorobenzenesulfonyl)isobotylamino]-pyrimidin-4-ylamino}-
3-(4-dimethylcarbamoyloxyphenyl)propionic acid
2-{2-diethylamino-5-[(4-fluorobenzenesulfonyl)-n-butylamino]-pyrimidin-4-ylamino}-
3-(4-dimethylcarbamoyloxyphenyl)propionic acid;
2-{2-diethylamino-5-[(2,5-difluorobenzenesulfonyl)methylamino]-pyrimidin-4-ylamino
}-3-(4-dimethylcarbamoyloxyphenyl)propionic acid;
2-{2-diethyIamino-5-[(2,3-difluorobenzenesulfonyl)ethylamino]-pyrimidin-4-ylamino}-
3-(4-dimethylcarbamoyloxyphenyl)propionic acid;
2-{2-Diethylamino-5-[(4-fluorobenzenesulfonyl)-(2-trisfluoroethyl)-
amino]pyrimidin-4-ylamino}-3-(4-dimethylcarbamoyloxyphenyl)propionic acid; and
pharmaceutically acceptable salts thereof.
7. A pharmaceutical composition comprising a pharmaceutically acceptable carrier
and a therapeutical ly effective amount of a compound as claimed in any one of claims 1-3, 5, or
6.
8. A pharmaceutical composition as claimed in claim 7 for treating a disease
mediated by cu integrin in a patient.
9. A pharmaceutical composition as claimed in claim 8, whereinthe disease
mediated by 04 integrin is an inflammatory disease/
10. A compound of formula (IV):
wherein each X is independently fluoro, chloro or bromo;
p is 0 or an integer from 1-3;
Rl is selected from the group consisting of methyl and ethyl;
R2 is lower alkynyl;
and pharmaceutical ly acceptable salts thereof.
11. A compound of Formula V:
wherein each X is independently selected from the group consisting of fluoro and
chloro,
m is an integer equal to 1 or 2;
R2 is lower alkynyl;
and pharmaceutically acceptable salts thereof.
12. A compound of Formula VI:
wherein each X is independently fluoro or chloro;
n is zero or one;
R2 is lower alkynyl;
and pharmaceutically acceptable salts thereof.
13. A compound as claimed in any one of claims 10, 11 or 12, wherein R2 is
propargyl.
14. A compound as claimed in claim 12, wherein X is F or Cl, and n is 0.
15. A compound selected from the group consisting of:
2-{2-Diethylamino-5-[(4-fluorobenzenesulfonyl)propargylamino] pyrimidin-4-
ylamino}-3-(4-dimethylcarbamoyloxyphenyl)propionic acid,
2-{2-Diethylamino-5-[(2,4-difluorobenzenesulfonyl)propargylamino] pyrimidin-4-
ylamino}-3-(4-dimethylcarbamoyloxyphenyl)propionic acid; and
pharmaceutically acceptable salts thereof.
16. A pharmaceutical composition comprising a pharmaceutically acceptable carrier
and a therapeutically effective amount of a compound as claimed in any one of claims 10-12, 14
or 15.
17. A pharmaceutical composition as claimed in claim 16 for treating a disease
mediated by cu integrin in a patient.
18. A pharmaceutical composition as claimed in claim 17, wherein the disease
mediated by cu integrin is an inflammatory disease.
19. A pharmaceutical composition as claimed in claim 17, wherein the disease is
rheumatoid arthritis.
Disclosed are compounds which bind 4 integrins, where the 4integrin is preferably VLA-4. Certain of these com-
pounds also inhibit leukocyte adhesion and. in particular, leukocyte adhesion mediated by 4integrins, where the 4 integrin is preferably
VLA-4. Such compounds are useful in the treatment of inflammatory diseases in a mammalian patient, e.g., human, such as asthma,
Alzheimer"s disease, atherosclerosis, AIDS dementia, diabetes, inflammatory bowel disease, rheumatoid arthritis, tissue transplan-
tation, tumor metastasis and myocardial ischemia. The compounds can also be administered for the treatment of inflammatory brain
diseases such as multiple sclerosis.

Documents:

1533-kolnp-2004-granted-abstract.pdf

1533-kolnp-2004-granted-assignment.pdf

1533-kolnp-2004-granted-claims.pdf

1533-kolnp-2004-granted-correspondence.pdf

1533-kolnp-2004-granted-description (complete).pdf

1533-kolnp-2004-granted-form 1.pdf

1533-kolnp-2004-granted-form 18.pdf

1533-kolnp-2004-granted-form 3.pdf

1533-kolnp-2004-granted-form 5.pdf

1533-kolnp-2004-granted-gpa.pdf

1533-kolnp-2004-granted-letter patent.pdf

1533-kolnp-2004-granted-reply to examination report.pdf

1533-kolnp-2004-granted-specification.pdf

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Patent Number

218545

Indian Patent Application Number

01533/KOLNP/2004

PG Journal Number

14/2008

Publication Date

04-Apr-2008

Grant Date

02-Apr-2008

Date of Filing

12-Oct-2004

Name of Patentee

ELAN PHARMACEUTICALS, INC.

Applicant Address

800 GATEWAY BOULEVARD, SOUTH SAN FRANCISCO USA.

Inventors:

#	Inventor's Name	Inventor's Address
1	KONRADI ANDREI W	30,VICTORIA ROAD BURLINGAME USA.
2	SEMKO CHRISTOPHER M	4993 NORRIS ROAD FR34MONT USA.
3	XU YING-ZI	793 CEREZA DRIVE PALO ALTO USA
4	STUPI BRIAN P	421 TERRACE AVENUE P.O BOX 239 MOSS BEACH USA.
5	SMITH JENIFEER	APT.103 331 PHILIP DRIVE DAILY CITY USA.
6	THORSETT EUGENE D	326 PURISSIMA STREET HALF MOON BAY USA.
7	PLESS MICHAEL A	848 STELLE COURT SUNNYVALE USA.

PCT International Classification Number

A61K31/505

PCT International Application Number

PCT/US2003/017150

PCT International Filing date

2003-05-27

PCT Conventions:

#	PCT Application Number	Date of Convention	Priority Country
1	60/383,244	2002-05-24	U.S.A.