Field of the Invention The present invention relates to combinations and methods for treatment or prevention of neoplasia disorders in a mammal using two or more components with at least one component being an antiangiogenesis agent.

Background of the Invention A neoplasm, or tumor, is an abnormal, unregulated, and disorganized proliferation of cell growth. A neoplasm is malignant, or cancerous, if it has properties of destructive growth, invasiveness and metastasis. Invasiveness refers to the local spread of a neoplasm by infiltration or destruction of surrounding tissue, typically breaking through the basal laminas that define the boundaries of the tissues, thereby often entering the body's circulatory system. Metastasis typically refers to the dissemination of tumor cells by lymphotics or blood vessels. Metastasis also refers to the migration of tumor cells by direct extension through serous cavities, or subarachnoid or other spaces.

Through the process of metastasis, tumor cell migration to other areas of the body establishes neoplasms in areas away from the site of initial appearance.

Cancer is now the second leading cause of death in the United States and over 8,000,000 persons in the United States have been diagnosed with cancer. In 1995, cancer accounted for 23.3% of all deaths in the United States. (See U. S. Dept. of Health and Human Services,

National Center for Health Statistics, Health United States 1996-97 and Injury Chartbook 117 (1997)).

Cancer is not fully understood on the molecular level. It is known that exposure of a cell to a carcinogen such as certain viruses, certain chemicals, or radiation, leads to DNA alteration that inactivates a "suppressive"gene or activates an"oncogene".

Suppressive genes are growth regulatory genes, which upon mutation, can no longer control cell growth.

Oncogenes are initially normal genes (called prooncogenes) that by mutation or altered context of expression become transforming genes. The products of transforming genes cause inappropriate cell growth. More than twenty different normal cellular genes can become oncogenes by genetic alteration. Transformed cells differ from normal cells in many ways, including cell morphology, cell-to-cell interactions, membrane content, cytoskeletal structure, protein secretion, gene expression and mortality (transformed cells can grow indefinitely).

Cancer is now primarily treated with one or a combination of three types of therapies: surgery, radiation, and chemotherapy. Surgery involves the bulk removal of diseased tissue. While surgery is sometimes effective in removing tumors located at certain sites, for example, in the breast, colon, and skin, it cannot be used in the treatment of tumors located in other areas, such as the backbone, nor in the treatment of disseminated neoplastic conditions such as leukemia.

Chemotherapy involves the disruption of cell replication or cell metabolism. It is used most often in the treatment of breast, lung, and testicular cancer.

The adverse effects of systemic chemotherapy used in the treatment of neoplastic disease is most feared by patients undergoing treatment for cancer. Of these adverse effects nausea and vomiting are the most common and severe side effects. Other adverse side effects include cytopenia, infection, cachexia, mucositis in patients receiving high doses of chemotherapy with bone marrow rescue or radiation therapy; alopecia (hair loss ); cutaneous complications (see M. D. Abeloff, et al: Alopecia and Cutaneous Complications. P. 755-56. In Abeloff, M. D., Armitage, J. O., Lichter, A. S., and Niederhuber, J. E. (eds) Clinical Oncology. Churchill Livingston, New York, 1992, for cutaneous reactions to chemotherapy agents), such as pruritis, urticaria, and angioedema; neurological complications; pulmonary and cardiac complications in patients receiving radiation or chemotherapy; and reproductive and endocrine complications.

Chemotherapy-induced side effects significantly impact the quality of life of the patient and may dramatically influence patient compliance with treatment.

Additionally, adverse side effects associated with chemotherapeutic agents are generally the major dose- limiting toxicity (DLT) in the administration of these drugs. For example, mucositis, is one of the major dose limiting toxicity for several anticancer agents, including the antimetabolite cytotoxic agents 5-FU, methotrexate, and antitumor antibiotics, such as doxorubicin. Many of these chemotherapy-induced side effects if severe, may lead to hospitalization, or

require treatment with analgesics for the treatment of pain.

The adverse side effects induced by chemotherapeutic agents and radiation therapy have become of major importance to the clinical management of cancer patients.

U. S. Patent No. 5,854,205 describes an isolated endostatin protein that is an inhibitor of endothelial cell proliferation and angiogenesis. U. S. Patent No.

5,843,925 describes a method for inhibiting angiogenesis and endothelial cell proliferation using a <BR> <BR> <BR> <BR> 7-[substituted amino]-9-[(substituted glycyl0amido]-6- demethyl-6-deoxytetracycline. U. S. Patent No. 5,863,538 describes methods and compositions for targeting tumor vasculature of solid tumors using immunological and growth factor-based reagents in combination with chemotherapy and radiation. U. S. Patent No. 5,837,682 describes the use of fragments of an endothelial cell proliferation inhibitor, angiostatin. U. S. Patent No.

5,861,372 describes the use of an aggregate endothelial inhibitor, angiostatin, and it use in inhibiting angiogenesis. U. S. Patent No. 5,885,795 describes methods and compositions for treating diseases mediated by undesired and uncontrolled angiogenesis by administering purified angiostatin or angiostatin derivatives. PCT/GB97/00650 describes the use of cinnoline derivatives for use in the production of an antiangiogenic and/or vascular permeability reducing effect. PCT/US97/09610 describes administration of an anti-endogin monoclonal antibody, or fragments thereof, which is conjugated to at least one angiogenesis

inhibitor or antitumor agent for use in treating tumor and angiogenesis-associated diseases. PCT/IL96/00012 describes a fragment of the Thrombin B-chain for the treatment of cancer. PCT/US95/16855 describes compositions and methods of killing selected tumor cells using recombinant viral vectors.

Ravaud, A. et al. describes the efficacy and tolerance of interleukin-2 (IL-2), interferon alpha-2a, and fluorouracil in patients with metastatic renal cell carcinoma.. J. Clin. Oncol. 16, No. 8,2728-32,1998.

Stadler, W. M. et al. describes the response rate and toxicity of oral 13-cis-retinoic acid added to an outpatient regimen of subcutaneous interleukin-2 and interferon alpha in patients with metastatic renal cell carcinoma. J. Clin. Oncol. 16, No. 5,1820-25,1998.

Rosenbeg, S. A. et al. describes treatment of patients with metastatic melanoma using chemotherapy with cisplatin, dacarbazine, and tamoxifen alone or in combination with interleukin-2 and interferon alpha-2b.

J. Clin. Oncol. 17, No. 3,968-75,1999. Tourani, J-M. et al describes treatment of renal cell carcinoma using interleukin-2, and interferon alpha-2a administered in combination with fluorouracil. J. Clin. Oncol. 16, No. 7, 2505-13,1998. Majewski, S. describes the anticancer action of retinoids, vitamin D3 and cytokines (interferons and interleukin-12) as related to the antiangiogenic and antiproliferative effects.

J. Invest. Dermatol. 108, No. 4,571,1997. Ryan, C. W. describes treatment of patients with metastatic renal cell cancer w*ith GM-CSF, Interleukin-2, and interferon- alpha plus oral cis-retinoic acid in patients with

metastatic renal cell cancer. J. Invest. Med. 46, No. 7, 274A, 1998. Tai-Ping, D. describes potential anti- angiogenic therapies. Trends Pharmacol. Sci. 16, No. 2, 57-66,1995. Brembeck, F. H. describes the use of 13- cis retinoic acid and interferon alpha to treat UICC stage III/IV pancreatic cancer. Gastroenterology 114, No. 4, Pt. 2, A569,1998. Brembeck, F. H. describes the use of 13-cis retinoic acid and interferon alpha in patients with advanced pancreatic carcinoma. Cancer 83, No. 11,2317-23,1998. Mackean, M. J. describes the use of roquinimex (Linomide) and alpha interferon in patients with advanced malignant melanoma or renal carcinoma. Br. J. Cancer 78, No. 12,1620-23,1998 Jayson, G. C. describes the use of interleukin 2 and interleukin-interferon alpha in advanced renal cancer.

Br. J. Cancer 78, No. 3,366-69,1998. Abraham, J. M. describes the use of Interleukin-2, interferon alpha and 5-fluorouracil in patients with metastatic renal carcinoma. Br. J. Cancer 78, Suppl. 2,8,1998. Soori, G. S. describes the use of chemo-biotherapy with chlorambucil and alpha interferon in patients with non- hodgkins lymphoma. Blood 92, No. 10, Pt. 2 Suppl. 1, 240b, 1998. Enschede, S. H. describes the use of interferon alpha added to an anthracycline-based regimen in treating low grade and intermediate grade non- hodgkin's lymphoma. Blood 92, No. 10, Pt. 1 Suppl. 1, 412a, 1998. Schachter, J. describes the use of a sequential multi-drug chemotherapy and biotherapy with interferon alpha, a four drug chemotherapy regimen and GM-CSF. Cancer Biother. Radiopharm. 13, No. 3,155-64, 1998.

Mross, K. describes the use of retinoic acid, interferon alpha and tamoxifen in metastatic breast cancer patients. J. Cancer Res. Clin. Oncology. 124 Suppl. 1 R123,1998. Muller, H. describes the use of suramin and tamoxifen in the treatment of advanced and metastatic pancreatic carcinoma. Eur. J. Cancer 33, Suppl. 8, S50,1997. Rodriguez, M. R. describes the use of taxol and cisplatin, and taxotere and vinorelbine in the treatment of metastatic breast cancer. Eur. J. Cancer 34, Suppl. 4, S17-S18,1998. Formenti, C. describes concurrent paclitaxel and radiation therapy in locally advanced breast cancer patients. Eur. J. Cancer 34, Suppl. 5, S39,1998. Durando, A. describes combination chemotherapy with paclitaxel (T) and epirubicin (E) for metastatic breast cancer. Eur. J. Cancer 34, Suppl. 5, S41,1998. Osaki, A. describes the use of a combination therapy with mitomycin-C, etoposide, doxifluridine and medroxyprogesterone acetate as second-line therapy for advanced breast cancer. Eur. J. Cancer 34, Suppl. 5, S59, 1998. Lode, H. et al. describes Synergy between an antiangiogenic integrin alpha v antagonist and an antibody-cytokine fusion protein eradicates spontaneous tumor metastasis. Proc. Nat. Acad. Sci. USA., 96 (4), 1591-1596,1999. Giannis, A. et al describes Integrin antagonists and other low molecular weight compounds as inhibitors of angiogenesis: new drugs in cancer therapy.

Angew. Chem. Int. Ed. Engl. 36 (6), 588-590,1997.

Takada, Y. et al describes the structures and functions of integrins. Jikken Igaku 14 (17), 2317-2322,1996.

Varner, J. et al. Tumor angiogenesis and the role of

vascular cell integrin alphavbeta3. Impt. Adv. Onc., 69- 87 Ref: 259.1996.

The use of TNP-470 and minocycline in combination with cyclophasphamide, CDDP, or thiotepa have been observed to substantially increase the tumor growth delay in one pre-clinical solid tumor model. (Teicher, B. A. et al., Breast Cancer Research and Treatment, 36 : 227-236,1995). Additionally, improved results were observed when the antiangiogenesis agents were used in combination with cyclophosphamide and fractionated radiation therapy. (Teicher, B. A. et al., European Journal of Cancer 32A (14): 2461-2466,1996).

Neri et al. examined the use of AG-3340 in combination with carboplatin and taxol for the treatment of cancer. (Neri et al., Proc Am Assoc Can Res, Vol 39, 89 meeting, 302 1998). U. S. Patent No. 5,837,696 describes the use of tetracycline compounds to inhibit cancer growth. WO 97/48,685 describes various substituted compounds that inhibit metalloproteases. EP 48/9,577 describes peptidyl derivatives used to prevent tumor cell metastasis and invasion. WO 98/25,949 describes the use of N5-substituted 5-amino-1,3,4- thiadiazole-2-thiols to inhibit metallopreteinase enzymes. WO 99/21,583 describes a method of inhibiting metastases in patients having cancer in which wildtype p53 is predominantly expressed using a combination of radiation therapy and a selective matrix metalloproteinase-2 inhibitor. WO 98/33,768 describes arylsulfonylamino hydroxamic acid derivatives in the treatment of cancer. WO 98/30,566 describes cyclic sulfone derivatives useful in the treatment of cancer.

WO 98/34,981 describes arylsulfonyl hydroxamic acid derivatives useful in the treatment of cancer. WO 98/33,788 discloses the use of carboxylic or hyroxamic acid derivatives for treatment of tumors. WO 97/41,844 describes a method of using combinations of angiostatic compounds for the prevention and/or treatment of neovascularization in human patients. EP 48/9,579 describes peptidyl derivatives with selective gelatinase action that may be of use in the treatment of cancer and to control tumor metastases.

WO 98/11,908 describes the use of carboxylic or hyroxamic acid derivatives and a cyclosporin in combination therapy for treating mammals suffering from arthritic disease.

WO 98/03,516 describes phasphinate based compounds useful in the treatment of cancer.

WO 95/23,811 describes novel carbocyclic compounds which inhibit platelet aggregation.

WO 93/24,475 describes sulphamide derivatives may be useful in the treatment of cancer to control the development of metastases.

WO 98/16,227 describes a method of using [Pyrozol- 1-yl] benzenesulfonamides in the treatment of and prevention of neoplasia.

WO 98/22,101 describes a method of using [Pyrozol- l-yl] benzenesulfonamides as anti-angiogenic agents.

Description of the Invention A method for treating or preventing a neoplasia disorder in a mammal, including a human, in need of such treatment or prevention is

provided. The method comprises treating the mammal with a therapeutically effective amount of a combination comprising two or more components, the first component is an integrin antagonist, the second component is a MMP inhibitor, and the additional component or components is optionally selected from (a) an antiangiogenesis agent; (b) an antineoplastic agent; (c) an adjunctive agent; (d) an immunotherapeutic agent; (e) a device; (f) a vaccine; (g) an analgesic agent; and (h) a radiotherapeutic agent; provided that the additional component (s) is other than the integrin antagonist selected as the first component and the matrix metalloproteinase inhibitor selected as the second component.

In one embodiment the combination comprises a MMP inhibitor, an integrin antagonist and an antineoplastic agent.

Besides being useful for human treatment, the present invention is also useful for veterinary treatment of companion animals, exotic animals and farm animals, including mammals, rodents, and the like. More preferred animals include horses, dogs, and cats.

The methods and combinations of the present invention may be used for the treatment or prevention of neoplasia disorders including, but not limited to acral lentiginous melanoma, actinic keratoses, adenocarcinoma, adenoid cycstic carcinoma, adenomas, adenosarcoma, adenosquamous carcinoma, astrocytic tumors, bartholin gland carcinoma, basal cell carcinoma, bronchial gland

carcinomas, capillary, carcinoids, carcinoma, carcinosarcoma, cavernous, cholangiocarcinoma, chondosarcoma, choriod plexus papilloma/carcinoma, clear cell carcinoma, cystadenoma, endodermal sinus tumor, endometrial hyperplasia, endometrial stromal sarcoma, endometrioid adenocarcinoma, ependymal, epitheloid, Ewing's sarcoma, fibrolamellar, focal nodular hyperplasia, gastrinoma, germ cell tumors, glioblastoma, glucagonoma, hemangiblastomas, hemangioendothelioma, hemangiomas, hepatic adenoma, hepatic adenomatosis, hepatocellular carcinoma, insulinoma, intaepithelial neoplasia, interepithelial squamous cell neoplasia, invasive squamous cell carcinoma, large cell carcinoma, leiomyosarcoma, lentigo maligna melanomas, malignant melanoma, malignant mesothelial tumors, medulloblastoma, medulloepithelioma, melanoma, meningeal, mesothelial, metastatic carcinoma, mucoepidermoid carcinoma, neuroblastoma, neuroepithelial adenocarcinoma nodular melanoma, oat cell carcinoma, oligodendroglial, osteosarcoma, pancreatic polypeptide, papillary serous adenocarcinoma, pineal cell, pituitary tumors, plasmacytoma, pseudosarcoma, pulmonary blastoma, renal cell carcinoma, retinoblastoma, rhabdomyosarcoma, sarcoma, serous carcinoma, small cell carcinoma, soft tissue carcinomas, somatostatin-secreting tumor, squamous carcinoma, squamous cell carcinoma, submesothelial, superficial spreading melanoma, undifferentiatied carcinoma, uveal melanoma, verrucous carcinoma, vipoma, well differentiated carcinoma, and Wilm's tumor.

The methods and combinations of the present invention provide one or more benefits. Combinations of MMP inhibitors and integrin antagonists with the compounds, compositions, agents and therapies of the present invention are useful in treating and preventing neoplasia disorders. Preferably, the MMP inhibitors and integrin antagonists and the compounds, compositions, agents and therapies of the present invention are administered in combination at a low dose, that is, at a dose lower than has been conventionally used in clinical situations.

A benefit of lowering the dose of the compounds, compositions, agents and therapies of the present invention administered to a mammal includes a decrease in the incidence of adverse effects associated with higher dosages. For example, by the lowering the dosage of a chemotherapeutic agent such as methotrexate, a reduction in the frequency and the severity of nausea and vomiting will result when compared to that observed at higher dosages. Similar benefits are contemplated for the compounds, compositions, agents and therapies in combination with the antiangiogenesis agents of the present invention.

By lowering the incidence of adverse effects, an improvement in the quality of life of a patient undergoing treatment for cancer is contemplated.

Further benefits of lowering the incidence of adverse effects include an improvement in patient compliance, a reduction in the number of hospitalizations needed for the treatment of adverse effects, and a reduction in the

administration of analgesic agents needed to treat pain associated with the adverse effects.

Alternatively, the methods and combination of the present invention can also maximize the therapeutic effect at higher doses.

When administered as a combination, the therapeutic agents can be formulated as separate compositions which are given at the same time or different times, or the therapeutic agents can be given as a single composition.

When used as a therapeutic the compounds described herein are preferably administered with a physiologically acceptable carrier. A physiologically acceptable carrier is a formulation to which the compound can be added to dissolve it or otherwise facilitate its administration. Examples of physiologically acceptable carriers include, but are not limited to, water, saline, physiologically buffered saline. Additional examples are provided below.

The term"pharmaceutically acceptable"is used adjectivally herein to mean that the modified noun is appropriate for use in a pharmaceutical product.

Pharmaceutically acceptable cations include metallic ions and organic ions. More preferred metallic ions include, but are not limited to appropriate alkali metal salts, alkaline earth metal salts and other physiological acceptable metal ions. Exemplary ions include aluminum, calcium, lithium, magnesium, potassium, sodium and zinc in their usual valences.

Preferred organic ions include protonated tertiary amines and quaternary ammonium cations, including in part, trimethylamine, diethylamine, N, N'-

dibenzylethylenediamine, chloroprocaine, choline, diethanolamine, ethylenediamine, meglumine (N- methylglucamine) and procaine. Exemplary pharmaceutically acceptable acids include without limitation hydrochloric acid, hydrobromic acid, phosphoric acid, sulfuric acid, methanesulfonic acid, acetic acid, formic acid, tartaric acid, maleic acid, malic acid, citric acid, isocitric acid, succinic acid, lactic acid, gluconic acid, glucuronic acid, pyruvic acid oxalacetic acid, fumaric acid, propionic acid, aspartic acid, glutamic acid, benzoic acid, and the like.

A compound of the present invention can be formulated as a pharmaceutical composition. Such a composition can then be administered orally, parenterally, by inhalation spray, rectally, or topically in dosage unit formulations containing conventional nontoxic pharmaceutically acceptable carriers, adjuvants, and vehicles as desired. Topical administration can also involve the use of transdermal administration such as transdermal patches or iontophoresis devices. The term parenteral as used herein includes subcutaneous injections, intravenous, intramuscular, intrasternal injection, or infusion techniques. Formulation of drugs is discussed in, for example, Hoover, John E., Remington's Pharmaceutical Sciences, Mack Publishing Co., Easton, Pennsylvania; 1975. Another example of includes Liberman, H. A. and Lachman, L., Eds., Pharmaceutical Dose Forms, Marcel Decker, New York, N. Y., 1980.

Injectable preparations, for example, sterile injectable aqueous or oleaginous suspensions can be formulated according to the known art using suitable dispersing or wetting agents and suspending agents. The sterile injectable preparation can also be a sterile injectable solution or suspension in a nontoxic parenterally acceptable dilutent or solvent, for example, as a solution in 1,3-butanediol. Among the acceptable vehicles and solvents that can be employed are water, Ringer's solution, and isotonic sodium chloride solution. In addition, sterile, fixed oils are conventionally employed as a solvent or suspending medium. For this purpose any bland fixed oil can be employed including synthetic mono-or diglycerides. In addition, fatty acids such as oleic acid find use in the preparation of injectables. Dimethyl acetamide, surfactants including ionic and non-ionic detergents, polyethylene glycols can be used. Mixtures of solvents and wetting agents such as those discussed above are also useful.

Suppositories for rectal administration of the drug can be prepared by mixing the drug with a suitable nonirritating excipient such as cocoa butter, synthetic mono-di-or triglycerides, fatty acids and polyethylene glycols that are sold at ordinary temperatures but liquid at the rectal temperature and will therefore melt in the rectum and release the drug.

Solid dosage forms for oral administration can include capsules, tablets, pills, powders, and granules.

In such solid dosage forms, the compounds of this invention are ordinarily combined with one or more

adjuvants appropriate to the indicated route of administration. If administered per os, a contemplated aromatic sulfone hydroximate inhibitor compound can be admixed with lactose, sucrose, starch powder, cellulose esters of alkanoic acids, cellulose alkyl esters, talc, stearic acid, magnesium stearate, magnesium oxide, sodium and calcium salts of phosphoric and sulfuric acids, gelatin, acacia gum, sodium alginate, polyvinylpyrrolidone, and/or polyvinyl alcohol, and then tableted or encapsulated for convenient administration.

Such capsules or tablets can contain a controlled- release formulation as can be provided in a dispersion of active compound in hydroxypropylmethyl cellulose. In the case of capsules, tablets, and pills, the dosage forms can also comprise buffering agents such as sodium citrate, magnesium or calcium carbonate or bicarbonate.

Tablets and pills can additionally be prepared with enteric coatings.

For therapeutic purposes, formulations for parenteral administration can be in the form of aqueous or non-aqueous isotonic sterile injection solutions or suspensions. These solutions and suspensions can be prepared from sterile powders or granules having one or more of the carriers or diluents mentioned for use in the formulations for oral administration. A contemplated aromatic sulfone hydroximate inhibitor compound can be dissolved in water, polyethylene glycol, propylene glycol, ethanol, corn oil, cottonseed oil, peanut oil, sesame oil, benzyl alcohol, sodium chloride, and/or various buffers. Other adjuvants and modes of

administration are well and widely known in the pharmaceutical art.

Liquid dosage forms for oral administration can include pharmaceutically acceptable emulsions, solutions, suspensions, syrups, and elixirs containing inert diluents commonly used in the art, such as water.

Such compositions can also comprise adjuvants, such as wetting agents, emulsifying and suspending agents, and sweetening, flavoring, and perfuming agents.

The amount of active ingredient that can be combined with the carrier materials to produce a single dosage form varies depending upon the mammalian host treated and the particular mode of administration.

The present invention further includes kits comprising a MMP inhibitor, and integrin antagonist and optionally an antineoplastic agent.

The term"treatment"refers to any process, action, application, therapy, or the like, wherein a mammal, including a human being, is subject to medical aid with the object of improving the mammal's condition, directly or indirectly.

The term"inhibition,"in the context of neoplasia, tumor growth or tumor cell growth, may be assessed by delayed appearance of primary or secondary tumors, slowed development of primary or secondary tumors, decreased occurrence of primary or secondary tumors, slowed or decreased severity of secondary effects of disease, arrested tumor growth and regression of tumors, among others. In the extreme, complete inhibition, is referred to herein as prevention or chemoprevention.

The term"prevention"includes either preventing

the onset of clinically evident neoplasia altogether or preventing the onset of a preclinically evident stage of neoplasia in individuals at risk. Also intended to be encompassed by this definition is the prevention of initiation for malignant cells or to arrest or reverse the progression of premalignant cells to malignant cells. This includes prophylactic treatment of those at risk of developing the neoplasia.

The term"angiogenesis"refers to the process by which tumor cells trigger abnormal blood vessel growth to create their own blood supply, and is a major target of cancer research. Angiogenesis is believed to be the mechanism via which tumors get needed nutrients to grow and metastasize to other locations in the body.

Antiangiogenic agents interfere with these processes and destroy or control tumors.

Angiogenesis is an attractive therapeutic target because it is a multi-step process that occurs in a specific sequence, thus providing several possible targets for drug action. Examples of agents that interfere with several of these steps include thrombospondin-1, angiostatin, endostatin, interferon alpha and compounds such as matrix metalloproteinase (MMP) inhibitors that block the actions of enzymes that clear and create paths for newly forming blood vessels to follow; compounds, such as avp3 inhibitors, that interfere with molecules that blood vessel cells use to bridge between a parent blood vessel and a tumor; agents, such as specific COX-2 inhibitors, that prevent the growth of cells that form new blood vessels; and

protein-based compounds that simultaneously interfere with several of these targets.

Antiangiogenic therapy may offer several advantages over conventional chemotherapy for the treatment of cancer.

Antiangiogenic agents have low toxicity in preclinical trials and development of drug resistance has not been observed (Folkman, J., Seminars in Medicine of the Beth Israel Hospital, Boston 333 (26): 1757-1763,1995). As angiogenesis is a complex process, made up of many steps including invasion, proliferation and migration of endothelial cells, it can be anticipated that combination therapies will be most effective. Kumar and Armstrong describe anti-angiogenesis therapy used as an adjunct to chemotherapy, radiation therapy, or surgery.

(Kumar, CC, and Armstrong, L., Tumor-induced angiogenesis: a novel target for drug therapy?, Emerging Drugs (1997), 2,175-190).

The phrase"therapeutically-effective"is intended to qualify the amount of each agent that will achieve the goal of improvement in neoplastic disease severity and the frequency of neoplastic disease over treatment of each agent by itself, while avoiding adverse side effects typically associated with alternative therapies.

A"therapeutic effect"or"therapeutic effective amount"is intended to qualify the amount of an anticancer agent required to relieve to some extent one or more of the symptoms of a neoplasia disorder, including, but is not limited to: 1) reduction in the number of cancer cells; 2) reduction in tumor size; 3) inhibition (i. e., slowing to some extent, preferably

stopping) of cancer cell infiltration into peripheral organs; 3) inhibition (i. e., slowing to some extent, preferably stopping) of tumor metastasis; 4) inhibition, to some extent, of tumor growth; 5) relieving or reducing to some extent one or more of the symptoms associated with the disorder; and/or 6) relieving or reducing the side effects associated with the administration of anticancer agents.

The phrase"combination therapy" (or"co-therapy") embraces the administration of a metalloproteinase inhibitor, an integrin antagonist and optionally an antineoplastic agent as part of a specific treatment regimen intended to provide a beneficial effect from the co-action of these therapeutic agents. The beneficial effect of the combination includes, but is not limited to, pharmacokinetic or pharmacodynamic co-action resulting from the combination of therapeutic agents.

Administration of these therapeutic agents in combination typically is carried out over a defined time period (usually minutes, hours, days or weeks depending upon the combination selected)."Combination therapy" generally is not intended to encompass the administration of two or more of these therapeutic agents as part of separate monotherapy regimens that incidentally and arbitrarily result in the combinations of the present invention."Combination therapy"is intended to embrace administration of these therapeutic agents in a sequential manner, that is, wherein each therapeutic agent is administered at a different time, as well as administration of these therapeutic agents, or at least two of the therapeutic agents, in a

substantially simultaneous manner. Substantially simultaneous administration can be accomplished, for example, by administering to the subject a single capsule having a fixed ratio of each therapeutic agent or in multiple, single capsules for each of the therapeutic agents. Sequential or substantially simultaneous administration of each therapeutic agent can be effected by any appropriate route including, but not limited to, oral routes, intravenous routes, intramuscular routes, and direct absorption through mucous membrane tissues. The therapeutic agents can be administered by the same route or by different routes.

For example, a first therapeutic agent of the combination selected may be administered by intravenous injection while the other two therapeutic agents of the combination may be administered orally. Alternatively, for example, all three therapeutic agents may be administered orally or all three therapeutic agents may be administered by intravenous injection. The sequence in which the therapeutic agents are administered is not narrowly critical."Combination therapy"also can embrace the administration of the therapeutic agents as described above in further combination with other biologically active ingredients (such as, but not limited to, a second and different antineoplastic agent) and non-drug therapies (such as, but not limited to, surgery or radiation treatment). Where the combination therapy further comprises radiation treatment, the radiation treatment may be conducted at any suitable time so long as a beneficial effect from the co-action of the combination of the therapeutic agents and

radiation treatment is achieved. For example, in appropriate cases, the beneficial effect is still achieved when the radiation treatment is temporally removed from the administration of the therapeutic agents, perhaps by days or even weeks.

The phrases"low dose"or"low dose amount", in characterizing a therapeutically effective amount of the antiangiogenesis agent and the antineoplastic agent or therapy in the combination therapy, defines a quantity of such agent, or a range of quantity of such agent, that is capable of improving the neoplastic disease severity while reducing or avoiding one or more antineoplastic-agent-induced side effects, such as myelosupression, cardiac toxicity, alopecia, nausea or vomiting.

The phrase"adjunctive therapy"encompasses treatment of a subject with agents that reduce or avoid side effects associated with the combination therapy of the present invention, including, but not limited to, those agents, for example, that reduce the toxic effect of anticancer drugs, e. g., bone resorption inhibitors, cardioprotective agents; prevent or reduce the incidence of nausea and vomiting associated with chemotherapy, radiotherapy or operation; or reduce the incidence of infection associated with the administration of myelosuppressive anticancer drugs.

The phrase an"immunotherapeutic agent"refers to agents used to transfer the immunity of an immune donor, e. g., another person or an animal, to a host by inoculation. The term embraces the use of serum or gamma gobulin containing performed antibodies produced

by another individual or an animal; nonspecific systemic stimulation; adjuvants; active specific immunotherapy; and adoptive immunotherapy. Adoptive immunotherapy refers to the treatment of a disease by therapy or agents that include host inoculation of sensitized lymphocytes, transfer factor, immune RNA, or antibodies in serum or gamma globulin.

The phrase a"device"refers to any appliance, usually mechanical or electrical, designed to perform a particular function.

The phrase a"vaccine"includes agents that induce the patient's immune system to mount an immune response against the tumor by attacking cells that express tumor associated antigens (TAAs).

The phrase"multi-functional proteins"encompass a variety of pro-angiogenic factors that include basic and acid fibroblast growth factors (bFGF and aFGF) and vascular permeability factor/vascular endothelial growth factor (VPF/VEGF) (Bikfalvi, A. et al., Endocrine Reviews 18: 26-45,1997). Several endogenous antiangiogenic factors have also been characterized as multi-functional proteins and include angiostatin <BR> <BR> <BR> <BR> <BR> (O'Reilly et al., Cell (Cambridge, Mass) 79 (2): 315-328, 1994), endostatin (O'Reilly et al, Cell (Cambridge, Mass) 88 (2): 277-285,1997), interferon. alpha.

(Ezekowitz et al, N. Engl. J. Med., May 28,326 (22) 1456-1463,1992), thrombospondin (Good et al, Proc Nat2 Acad Sci USA 87 (17): 6624-6628,1990; Tolsma et al, J Cell Biol 122 (2): 497-511,1993), and platelet factor 4 (PF4) (Maione et al, Science 247: (4938): 77-79,1990).

The phrase an"analgesic agent"refers to an agent that relieves pain without producing anesthesia or loss of consciousness generally by altering the perception of nociceptive stimuli.

The phrase a"radiotherapeutic agent"refers to the use of electromagnetic or particulate radiation in the treatment of neoplasia.

The term"pBATT"embraces"or"Protein-Based Anti- Tumor Therapies,"refers to protein-based therapeutics for solid tumors. The PBATTs are including proteins that have demonstrated efficacy against tumors in animal models or in humans. The protein is then modified to increase its efficacy and toxicity profile by enhancing its bioavailability and targeting.

"Angiostatin"is a 38 kD protein comprising the first three or four kringle domains of plasminogen and was first described in 1994 (O'Reilly, M. S. et al., Cell (Cambridge, Mass.) 79 (2): 315-328,1994). Mice bearing primary (Lewis lung carcinoma-low metastatic) tumors did not respond to angiogenic stimuli such as bFGF in a corneal micropocket assay and the growth of metastatic tumors in these mice was suppressed until the primary tumor was excised. The factor responsible for the inhibition of angiogenesis and tumor growth was designated mouse angiostatin. Angiostatin was also shown to inhibit the growth of endothelial cells in vitro.

Human angiostatin can be prepared by digestion of plasminogen by porcine elastase (O'Reilly, et al., Cell 79 (2): 315-328,1994) or with human metalloelastase (Dong et al., Cell 88,801-810,1997). The angiostatin

produced via porcine elastase digestion inhibited the growth of metastases and primary tumors in mice.

O'Reilly et al (Cell 79 (2): 315-328,1994) demonstrated that human angiostatin inhibited metastasis of Lewis lung carcinoma in SCID mice. The same group (O'Reilly, M. S. et al., Nat. Med. (N. Y.) 2 (6): 689-692,1996) subsequently showed that human angiostatin inhibited the growth of the human tumors PC3 prostate carcinoma, clone A colon carcinoma, and MDA-MB breast carcinoma in SCID mice. Human angiostatin also inhibited the growth of the mouse tumors Lewis lung carcinoma, T241 fibrosarcoma and M5076 reticulum cell carcinoma in C57B1 mice.

Because these enzymatically-prepared angiostatins are not well characterized biochemically, the precise composition of the molecules is not known.

Angiostatins of known composition can be prepared by means of recombinant DNA technology and expression in heterologous cell systems. Recombinant human angiostatin comprising Kringle domains one through four (K1-4) has been produced in the yeast Pichia pastoris (Sim et al., Cancer Res 57: 1329-1334,1997). The recombinant human protein inhibited growth of endothelial cells in vitro and inhibited metastasis of Lewis lung carcinoma in C57B1 mice. Recombinant murine angiostatin (K1-4) has been produced in insect cells (Wu et al., Biochem Biophys Res Comm 236: 651-654,1997).

The recombinant mouse protein inhibited endothelial cell growth in vitro and growth of primary Lewis lung carcinoma in vivo. These experiments demonstrated that the first four kringle domains are sufficient for

angiostatin activity but did not determine which kringle domains are necessary.

Cao et al. (J. Biol. Chem. 271: 29461-29467,1996), produced fragments of human plasminogen by proteolysis and by expression of recombinant proteins in E. coli.

These authors showed that kringle one and to a lesser extent kringle four of plasminogen were responsible for the inhibition of endothelial cell growth in vitro.

Specifically, kringles 1-4 and 1-3 inhibited at similar concentrations, while K1 alone inhibited endothelial cell growth at four-fold higher concentrations.

Kringles two and three inhibited to a lesser extent.

More recently Cao et al. (J Biol Chem 272: 22924-22928, 1997), showed that recombinant mouse or human kringle five inhibited endothelial cell growth at lower concentrations than angiostatin (K1-4). These experiments demonstrated in vitro angiostatin-like activity but did not address in vivo action against tumors and their metastases.

World patent applications WO 95/29242 A1, WO 96/41194 A1, and WO 96/35774 A2 describe the expression, purification, and characterization of angiostatin. WO 95/29242 A1 951102 discloses purification of a protein from blood and urine by HPLC that inhibits proliferation of endothelial cells. The protein has a molecular weight between 38 kilodaltons and 45 kilodaltons and an amino acid sequence substantially similar to that of a murine plasminogen fragment beginning at amino acid number 79 of a murine plasminogen molecule. WO 96/41194 A1 961219, discloses compounds and methods for the diagnosis and monitoring of angiogenesis-dependent

diseases. WO 96/35774 A2 961114 discloses the structure of protein fragments, generally corresponding to kringle structures occurring within angiostatin. It also discloses aggregate forms of angiostatin, which have endothelial cell inhibiting activity, and provides a means for inhibiting angiogenesis of tumors and for treating angiogenic-mediated diseases.

"Endostatin"is a 20-kDa (184 amino acid) carboxy fragment of collagen XVIII, is an angiogenesis inhibitor produced by a hemangioendothelioma (O'Reilly, M. S. et al., Cell (Cambridge, Mass.) 88 (2): 277-285,1997); and WO 97/15666). Endostatin specifically inhibits endothelial proliferation and inhibits angiogenesis and tumor growth. Primary tumors treated with non-refolded suspensions of E. coli-derived endostatin regressed to dormant microscopic lesions. Toxicity was not observed and immunohistochemical studies revealed a blockage of angiogenesis accompanied by high proliferation balanced by apoptosis in tumor cells.

"Interferon. alpha." (IFN. alpha.) is a family of highly homologous, species-specific proteins that possess complex antiviral, antineoplastic and immunomodulating activities (Extensively reviewed in the monograph"Antineoplastic agents, interferon alfa", American Society of Hospital Pharmacists, Inc., 1996).

Interferon. alpha. also has anti-proliferative, and antiangiogenic properties, and has specific effects on cellular differentiation (Sreevalsan, in"Biologic Therapy of Cancer", pp. 347-364, (eds. V. T. DeVita Jr., S. Hellman, and S. A. Rosenberg), J. B. Lippincott Co, Philadelphia, PA, 1995).

Interferon. alpha. is effective against a variety of cancers including hairy cell leukemia, chronic myelogenous leukemia, malignant melanoma, and Kaposi's sarcoma. The precise mechanism by which IFN. alpha. exerts its anti-tumor activity is not entirely clear, and may differ based on the tumor type or stage of disease. The anti-proliferative properties of IFN. alpha., which may result from the modulation of the expression of oncogenes and/or proto-oncogenes, have been demonstrated on both tumor cell lines and human tumors growing in nude mice (Gutterman, J. U., Proc.

Natl. Acad. Sci., USA 91: 1198-1205,1994).

Interferon is also considered an anti-angiogenic factor, as demonstrated through the successful treatment <BR> <BR> <BR> <BR> <BR> of hemangiomas in infants (Ezekowitz et al, N. Engl. J.<BR> <BR> <BR> <BR> <BR> <BR> <BR> <P>Med., May 28,326 (22) 1456-1463,1992) and the effectiveness of IFN. alpha. against Kaposi's sarcoma (Krown, Semin Oncol 14 (2 Suppl 3): 27-33,1987). The mechanism underlying these anti-angiogenic effects is not clear, and may be the result of IFN. alpha. action on the tumor (decreasing the secretion of pro-angiogenic factors) or on the neo-vasculature. IFN receptors have been identified on a variety of cell types (Navarro et al., Modern Pathology 9 (2): 150-156,1996).

United States Patent 4,530,901, by Weissmann, describes the cloning and expression of IFN-. alpha.-type molecules in transformed host strains. United States Patent 4,503,035, Pestka, describes an improved processes for purifying 10 species of human leukocyte interferon using preparative high performance liquid chromatography. United States Patent 5,231,176,

Goeddel, describes the cloning of a novel distinct family of human leukocyte interferons containing in their mature form greater than 166 and no more than 172 amino acids.

United States Patent 5,541,293, by Stabinsky, describes the synthesis, cloning, and expression of consensus human interferons. These are non-naturally occurring analogues of human (leukocyte) interferon- . alpha. assembled from synthetic oligonucleotides. The sequence of the consensus interferon was determined by comparing the sequences of 13 members of the IFN-. alpha. family of interferons and selecting the preferred amino acid at each position. These variants differ from naturally occurring forms in terms of the identity and/or location of one or more amino acids, and one or more biological and pharmacological properties (e. g., antibody reactivity, potency, or duration effect) but retain other such properties.

"Thrombospondin-1" (TSP-1) is a trimer containing three copies of a 180 kDa polypeptide. TSP-1 is produced by many cell types including platelets, fibroblasts, and endothelial cells (see Frazier, Curr Opin Cell Biol 3 (5): 792-799,1991) and the cDNA encoding the subunit has been cloned (Hennessy, et al., 1989, J Cell Biol 108 (2): 729-736; Lawler and Hynes, J Cell Biol 103 (5): 1635-1648,1986). Native TSP-1 has been shown to block endothelial cell migration in vitro and neovascularization in vivo (Good et al, Proc Nat1 Acad Sci USA 87 (17): 6624-6628,1990). Expression of TSP-1 in tumor cells also suppresses tumorigenesis and tumor-induced angiogenesis (Sheibani and Frazier, Proc

Natl Acad Sci USA 92 (15) 6788-6792,1995; Weinstat- Saslow et al., Cancer Res 54 (24): 6504-6511,1994). The antiangiogenic activity of TSP-1 has been shown to reside in two distinct domains of this protein (Tolsma <BR> <BR> <BR> <BR> <BR> et al, J Cell Biol 122 (2): 497-511,1993). One of these domains consists of residues 303 to 309 of native TSP-1 and the other consists of residues 481 to 499 of TSP-1.

Another important domain consists of the sequence CSVTCG which appears to mediate the binding of TSP-1 to some tumor cell types (Tuszynski and Nicosia, Bioessays 18 (1): 71-76,1996). These results suggest that CSVTCG, or related sequences, can be used to target other moieties to tumor cells. Taken together, the available data indicate that TSP-1 plays a role in the growth and vascularization of tumors. Subfragments of TSP-1, then, may be useful as antiangiogenic components of chimeras and/or in targeting other proteins to specific tumor cells. Subfragments may be generated by standard procedures (such as proteolytic fragmentation, or by DNA amplification, cloning, expression, and purification of specific TSP-1 domains or subdomains) and tested for antiangiogenic or anti-tumor activities by methods known <BR> <BR> <BR> <BR> <BR> in the art (Tolsma et al, J Cell Biol 122 (2): 497-511,<BR> <BR> <BR> <BR> <BR> <BR> <BR> <BR> 1993; Tuszynski and Nicosia, Bioessays 18 (1): 71-76, 1996).

The phrase"integrin antagonist"includes agents that impair endothelial cell adhesion via the various integrins. Integrin antagonists induce improperly proliferating endothelial cells to die, by interfering with molecules that blood vessel cells use to bridge between a parent blood vessel and a tumor.

Adhesion forces are critical for many normal physiological functions. Disruptions in these forces, through alterations in cell adhesion factors, are implicated in a variety of disorders, including cancer, stroke, osteoporosis, restenosis, and rheumatoid arthritis (A. F. Horwitz, Scientific American, 276: (5): 68-75,1997).

Integrins are a large family of cell surface glycoproteins which mediate cell adhesion and play central roles in many adhesion phenomena. Integrins are heterodimers composed of noncovalently linked alpha and beta polypeptide subunits. Currently eleven different alpha subunits have been identified and six different beta subunits have been identified. The various alpha subunits can combine with various beta subunits to form distinct integrins.

One integrin known as avb3 (or the vitronectin receptor) is normally associated with endothelial cells and smooth muscle cells. AVb3 integrins can promote the formation of blood vessels (angiogenesis) in tumors.

These vessels nourish the tumors and provide access routes into the bloodstream for metastatic cells.

The avb3 integrin is also known to play a role in various other disease states or conditions including tumor metastasis, solid tumor growth (neoplasia), osteoporosis, Paget's disease, humoral hypercalcemia of malignancy, angiogenesis, including tumor angiogenesis, retinopathy, arthritis, including rheumatoid arthritis, periodontal disease, psoriasis, and smooth muscle cell migration (e. g. restenosis).

Tumor cell invasion occurs by a three step process: 1) tumor cell attachment to extracellular matrix; 2) proteolytic dissolution of the matrix; and 3) movement of the cells through the dissolved barrier. This process can occur repeatedly and can result in metastases at sites distant from the original tumor.

The avb3 integrin and a variety of other av- containing integrins bind to a number of Arg-Gly-Asp (RGD) containing matrix macromolecules. Compounds containing the RGD sequence mimic extracellular matrix ligands and bind to cell surface receptors. Fibronectin and vitronectin are among the major binding partners of avb3 integrin. Other proteins and peptides also bind the avb3 ligand. These include the disintegrins (M.

Pfaff et al., Cell Adhes. Commun. 2 (6): 491-501,1994), peptides derived from phage display libraries (Healy, J. M. et al., Protein Pept. Lett. 3 (1): 23-30,1996; Hart, S. L. et al., J. Biol. Chem. 269 (17): 12468-12474, 1994) and small cyclic RGD peptides (M. Pfaff et al., J.

Biol. Chem., 269 (32): 20233-20238,1994). The monoclonal antibody LM609 is also an avb3 integrin

antagonist (D. A. Cheresh et al., J. Biol. Chem., 262 (36): 17703-17711,1987).

AVb3 inhibitors are being developed as potential anti-cancer agents. Compounds that impair endothelial cell adhesion via the avb3 integrin induce improperly proliferating endothelial cells to die.

The avb3 integrin has been shown to play a role in melanoma cell invasion (Seftor et al., Proc. Natl. Acad.

Sci. USA, 89: 1557-1561,1992). The avb3 integrin expressed on human melanoma cells has also been shown to promote a survival signal, protecting the cells from apoptosis (Montgomery et al., Proc. Natl. Acad. Sci.

USA, 91: 8856-8860,1994).

Mediation of the tumor cell metastatic pathway by interference with the avb3 integrin cell adhesion receptor to impede tumor metastasis would be beneficial.

Antagonists of avb3 have been shown to provide a therapeutic approach for the treatment of neoplasia (inhibition of solid tumor growth) because systemic administration of avb3 antagonists causes dramatic regression of various histologically distinct human tumors (Brooks et al., Cell, 79: 1157-1164,1994).

The adhesion receptor identified as integrin avb3 is a marker of angiogenic blood vessels in chick and man. This receptor plays a critical role in angiogenesis or neovascularization. Angiogenesis is characterized by the invasion, migration and proliferation of smooth muscle and endothelial cells by

new blood vessels. Antagonists of avb3 inhibit this process by selectively promoting apoptosis of cells in the neovasculature. The growth of new blood vessels, also contributes to pathological conditions such as diabetic retinopathy (Adonis et al., Amer. J. Ophthal., 118: 445-450,1994) and rheumatoid arthritis (Peacock et al., J. Exp. Med., 175:, 1135-1138,1992). Therefore, avb3 antagonists can be useful therapeutic targets for treating such conditions associated with neovascularization (Brooks et al., Science, 264: 569- 571,1994).

The avb3 cell surface receptor is also the major integrin on osteoclasts responsible for the attachment to the matrix of bone. Osteoclasts cause bone resorption and when such bone resorbing activity exceeds bone forming activity, osteoporosis (a loss of bone) results, which leads to an increased number of bone fractures, incapacitation and increased mortality.

Antagonists of avb3 have been shown to be potent inhibitors of osteoclastic activity both in vitro (Sato et al., J. Cell. Biol., 111: 1713-1723,1990) and in vivo (Fisher et al., Endocrinology, 132: 1411-1413, 1993). Antagonism of avb3 leads to decreased bone resorption and therefore assists in restoring a normal balance of bone forming and resorbing activity. Thus it would be beneficial to provide antagonists of osteoclast avb3which are effective inhibitors of bone resorption

and therefore are useful in the treatment or prevention of osteoporosis.

PCT Int. Appl. WO 97/08145 by Sikorski et al., discloses meta-guanidine, urea, thiourea or azacyclic amino benzoic acid derivatives as highly specific ab integrin antagonists.

PCT Int. Appl. WO 96/00574 A1 960111 by Cousins, R. D. et. al., describe preparation of 3-oxo-2,3,4,5- tetrahydro-lH-1,4-benzodiazepine and-2-benzazepine derivatives and analogs as vitronectin receptor antagonists.

PCT Int. Appl. WO 97/23480 A1 970703 by Jadhav, P. K. et. al. describe annelated pyrazoles as novel integrin receptor antagonists. Novel heterocycles including 3- [1- [3- (imidazolin-2-ylamino) propyl] indazol- 5-ylcarbonylamino]-2- (benzyl oxycarbonylamino) propionic acid, which are useful as antagonists of the avb3 integrin and related cell surface adhesive protein receptors.

PCT Int. Appl. WO 97/26250 A1 970724 by Hartman, G. D. et al., describe the preparation of arginine dipeptide mimics as integrin receptor antagonists.

Selected compounds were shown to bind to human integrin avb3 with EIB <1000 nM and claimed as compounds, useful for inhibiting the binding of fibrinogen to blood platelets and for inhibiting the aggregation of blood platelets.

PCT Int. Appl. WO 97/23451 by Diefenbach, B. et. al. describe a series of tyrosine-derivatives used as alpha v-integrin inhibitors for treating tumors,

osteoporosis, osteolytic disorder and for suppressing angiogenesis.

PCT Int. Appl. WO 96/16983 A1 960606. by Vuori, K. and Ruoslahti, E. describe cooperative combinations of avb3 integrin ligand and second ligand contained within a matrix, and use in wound healing and tissue regeneration. The compounds contain a ligand for the avb3 integrin and a ligand for the insulin receptor, the PDGF receptor, the IL-4 receptor, or the IGF receptor, combined in a biodegradable polymeric (e. g. hyaluronic acid) matrix.

PCT Int. Appl. WO 97/10507 A1 970320 by Ruoslahti, E; and Pasqualini, R. describe peptides that home to a selected organ or tissue in vivo, and methods of identifying them. A brain-homing peptide, nine amino acid residues long, for example, directs red blood cells to the brain. Also described is use of in vivo panning to identify peptides homing to a breast tumor or a melanoma.

PCT Int. Appl. WO 96/01653 A1 960125 by Thorpe, Philip E.; Edgington, Thomas S. describes bifunctional ligands for specific tumor inhibition by blood coagulation in tumor vasculature. The disclosed bispecific binding ligands bind through a first binding region to a disease-related target cell, e. g. a tumor cell or tumor vasculature; the second region has coagulation-promoting activity or is a binding region for a coagulation factor. The disclosed bispecific binding ligand may be a bispecific (monoclonal) antibody, or the two ligands may be connected by a

(selectively cleavable) covalent bond, a chemical linking agent, an avidin-biotin linkage, and the like.

The target of the first binding region can be a cytokine-inducible component, and the cytokine can be released in response to a leukocyte-activating antibody; this may be a bispecific antibody which crosslinks activated leukocytes with tumor cells.

Nonlimiting examples of integrin antagonists that may be used in the present invention are identified in Table 1, below.

Table No. 1. Examples of Integrin antagonists Compound'Trade/Mode of Reference Dosage l ResearchAction Name 2 (S)- L-748415 Vitronectin Benzenesulfonam antagonist ido)-3- [4- [2- (3,4,5,6- tetrahydropyrim idin-2- ylamino) ethoxy ]benzamido] prop ionic acid Merk KGaA Compoun d 125 Ethyl beta- [ [2- Vitronectin WO 97/08145 [ [ [3- antagonist [ (3,4,5,6,- tetrahydro-2H- azepin-7- yl) amino] phenyl ] carbonyl] am ino] acetyl]- amino]pyridine- 3-propanoic acid O- [9, 10- Vitronectin WO 97/34865 dimethoxy-antagonist Compound Trade/ Mode of Reference Dosage Research Action Name 1,2,3,4,5,6- hexahydro-4- [(1,4,5,6- tetrahydro-2- pyrimidinyl) hydrazono]-8- benz(e)azulenyl ]-N- [(phenylmethoxy )carbonyl]-DL- homoserine2,3- dih ydroxypropyl ester (2S)-Vitronectin EP 796855 Benzoylcarbonyl antagonist amino-3- [2- ( (4S)- (3- (4,5- dihydro-lH- imidazol-2- ylamino)-pro pyl)-2,5-dioxo- imidazolidin-l- yl)- acetylamino]- propionate S-836 Vitronectin antagonist; Angiogenesi s inhibitor; solid tumors (S)-2- [7- [N- SB-223245 Vitronectin (Benzimidazol-antagonist; 2-ylmethyl)-N-Angiogenesi methylcarbamoyl s inhibitor ]-4-methyl-3- oxo-2,3,4,5- tetrahydro-lH- 1,4- benzodiazepin- 2-yl]acetic Compound Trade/Mbde of Reference Dosage Research Action Name acid SD-983 Vitronectin antagonist; Angiogenesi s inhibitor Isoxaoline Vitronectin WO 96/37492 0.001-10 derivatives receptor mg/kg/ antagonist day; 0.01- 0.5 (pref. 0.01-0.1) mg/kg/day intra- nasally (2S)-Vitronectin EP 796855 Bensoylcarbonyl antagonist amino-3- [2- ( (4S)- (3- (4,5- dihydro-lH- imidazol-2- ylamino)- propyl)-2,5- dioxo- imidazolindin- l-yl)- acetylamino]- propionate Benzazulene Vitronectin WO 97/34865 deriviatives; antagonist 0- [9, 10- dimethoxy- 1,2,3,4,5,6- hexahydro-4- [ (1,4,5,6- tetrahydro-2- pyrimidinyl) hydrazono]-8- benz (e) azzuleny 1]-N- [ (phenylmethoxy )carbonyl]-DL- homoserine 2,3- dih ydroxypropyl Compound Trade/Mode of Reference Dosage Research Action Name ester Imunoglobulin abcix-GPIIb IIIa Recomended G, (human-mouse imab; receptor dosage: monoclonal c7E3 ReoPro antagonist; Intra- clone p7E3VHhC Vitronectin venous gamma 4 Fab antagonist bolus of fragment anti- 0.25 humanmg/kg, glycoprotein followed IIb/IIIa by 10 receptor), jjg/min for disulfide with 12 hrs. human-mouse monoclonal c7E3 clone p7E3VkhCk light chain- Arg-Gly-Asp-D-cRGDfV Apoptosis phe-Val penta- agonist; peptide Vitronectin antagonist vitro-Vitronectin Orally nectin antagonist active antag- onist Further examples of integrin antagonists can be found in the following documents: WO 98/07432 WO 98/16227 WO 97/36862 WO 97/36861 WO 9736859WO97/36858USWO 5639765 WO 5639765WO98/22500WOUS 98/20897 WO 98/18764 WO 98/14192 WO 98/08840 WO 98/04913 WO 97/48395 WO 9744333 WO 98/00395 WO 97/41102 WO 97/34865 WO 97/39028 WO 97/37655 WO 97/33887 EP 97/26250WO97/24124WOWO 97/24122 WO97/24119WO97/23480WOWO97/24336 97/23451 EP 765660 WO 97/14716 EP 77/1818 WO 97/01540 WO 96/37492 EP 741133 US 5565449 wo 96/26190 EP 727425 US 5627197 DE 4439846 EP 711770 EP 710657 WO 96/06087 WO 96/00730 WO 96/00574 WO 95/23811 US 5464855 WO 95/28426 JP 07242645 JP 07206860 EP 645376 WO 95/07712 WO 95/00544 AU 9464771 EP 614664 WO 94/21607 WO 94/15936 JP 06128289 WO 9411739 WO 93/08174 EP 537654 | EP 5229366WO92/07870WOUS 92/00995 EP 381033 WO 98/08518 US 5721210 EP 820991 EP 820988 WO 97/48444 WO 97/41844 WO 97/45447 WO 97/45137 US 5686570 US 5686568 US 5686571 US 5686569 US 5686567 US 5686566 WO 97/41149 DE 19613933 WO 97/35615 WO 97/25031 US 5639726 WO 97/18838 WO 97/11718 US 5612311 EP 77/0622 WO 97/08203 WO 97/06791 WO 97/03094 WO 96/40781 WO 5536814US5510332WOUS 96/07734 WO 96/00581WO95/34641WOWO 95/30438 DE 668278EP656348DEEP 4336758 EP 623615 DE 4310643 AU 9459185 WO 94/01152 CA 2120303 EP 632053 EP 618225 WO 94/18981 WO 06116289WO94/05310EPJP 58/9181 EP 5491129WO93/25218WOUS 93/20229 US 5225531 EP 570352 EP 570352 WO 92/09200 WO 445796WO91/07977EPEP 410767 US 384362US5663297EPEP 372486 US 9003983WO89/05155DEWO 19548798 DE 19653645DE9653646DEDE 19653647 DE 19654483 DE 4439846 EP 683173 EP 537654 EP 645376 EP 0710657 EP 727425 EP 741133 EP 771565 EP 0846702 EP 853084 JP 07285992 JP 09169742JP9235239JPJP 09316000 JP 08183752JP183788US5574026JP WO 9525543WO95/28426WOWO 95/32710 WP 96/26190W96/32945WOWO 97/12625 WO 97/16197WO97/21726WO97/22596WO WO 97/24336WO98/25892WOWO 98/25601 WO 97/33576WO98/00144WO98/00395WO WO 98/08518WO98/08840WOWO 98/10795 WO 98/11223WO98/12226WOWO 98/13071 WO 98/13354WO98/14192WOWO 98/15278 WO 98/18460WO98/18461WOWO 98/18764 WO 98/23608WO98/23613WO

The following individual references each hereby incorporated by reference herein, describe various integrin antagonists suitable for use in the invention described herein, and processes for their manufacture: WO 98/07432 WO 98/16227 WO 97/36862 WO 97/36861 WO 97/36860 WO 97/36859 WO 97/36858 US 5639765 WO 5639765WO98/22500WOUS 98/20897 WO 98/14192WO98/08840WOWO 98/04913 WO 97/48395 WO 97/44333 WO 98/00395 WO 97/41102 WO 97/39028WO97/37655WOWO 97/33887 EP 97/26250WO97/24124WOWO 97/24122 WO 97/24336 WO 97/24119 WO 97/23480 WO 97/23451 EP 76/5660 WO 97/14716 EP 771818 WO 97/01540 WO 96/37492 EP 74/1133 US 5565449 WO 96/26190 EP 5627197DE4439846EPUS 711770 EP 96/06087WO96/00730WOWO 96/00574 WO 95/23811 US 5464855 WO 95/28426 JP 07242645 JP 64/5376WO95/07712WOEP 95/00544 AU 61/4664WO94/21607WOEP 94/15936 JP 94/11739WO93/08174EPWO 537654 EP 52/29366WO92/07870WOUS 92/00995 EP 98/08518USWO EP 820991 EP 97/48444WO97/41844WOWO 97/45447 WO 5686570US5686568USUS 5686571 US 5686567US5686566WOUS 97/41149 DE 97/35615WO97/25031USWO 5639726 WO 97/11718US5612311EPWO 770622 WO 97/06791WO97/03094WOWO 96/40781 WO 5536814US5510332WOUS 96/07734 WO96/00581WO95/34641WOWO96/05304 95/30438 DE 66/8278EP656348DEEP 4336758 EP 43/10643AU94/59185NODE 94/01152 CA 63/2053EP618225WOEP 94/18981 WO 06/116289WO94/05310EPJP 58/9181 EP5491129WO93/25218WO93/20229US U.S. 570352EP57/0352WOEP 92/09200 WO 445796WO91/07977EPEP 410767 US 384362USEP EP 37/2486 US 90/03983WO89/05155DEWO 19548798 DE 19653645DE19653646DEDE 19653647 DE 4439846EP683173EPDE 537654 EP 0710657EP727425EPEP 741133 EP 0846702EP853084JPEP 07285992 JP 09169742JP09235239JPJP 09316000 JP 08183752JP08183788USJP 5574026 WO 95/25543WO95/28426WOWO 95/32710 WP 96/26190WO96/32945WOWO 97/12625 WO97/16197WO97/21726WOWO97/15666 97/22596 WO 97/24336WO98/25892WOWO 98/25601 WO 97/33576WO98/00144WOWO 98/00395 WO 98/08518WO98/08840WOWO 98/10795 WO 98/11223WO98/12226WOWO 98/13071 WO 98/13354WO98/14192WOWO 98/15278 WO 98/18460WO98/18461WO98/18764WO WO 98/23608WOWO 98/23613

The following individual references each hereby incorporated by reference herein, describe additional integrin antagonists suitable for use in the invention described herein, and processes for their manufacture: WO 99/45927WO99/44994USWO 5955572 US 99/38849WO99/37683WOWO 99/37621 WO 928793US5925655USEP 5919792 WO 99/31099US5912234WOWO 99/31061 WO99/30713WO99/30709WOWO99/31061 99/26945 WO 99/15507WO99/15506WOWO 99/15178 WO 99/11626WO99/06049WOWO 99/05107 US 5843906WO98/54217USUS 5840961 WO 5773646US5773644WOUS 98/33919 WO 98/30542EP854145EPWO 854140 EP 5773412US5766591USUS 5760028 US 98/15278US5741796WOWO 98/10795 WO 97/08145

The Vitaxin used in the therapeutic combinations of the present invention can be prepared in the manner set forth in WO 98/33,919.

Some Preferred integrin antagonists that may be used in the present invention are listed in the following references hereby each individually incorporated by reference, herein: U. S. Patent No. 5,773,644; U. S. Patent No. 5,773,646; Patent Application Serial No. U. S. 092/89,140; U. S.

Patent No. 5,852,210; U. S. Patent No. 5,843,906; U. S.

Patent Application Serial No. 091/41,547; U. S. Patent No. 5,952,381; U. S. Patent Application No. 092/88,742; Patent Application Serial No. U. S. 600/03,277; Patent Application Serial No. U. S. 087/13,555; Patent Application Serial No. U. S. 092/15,229; Patent Application Serial No. U. S. 090/34,758; Patent Application Serial No. U. S. 092/61,822; WO 98/33919.

More preferred integrin antagonists that may be used in the present invention include, but are not limited to (3R)-N- [ [5- [ (1,4,5,6-tetrahydro-5-hydroxy-2- pyrimidinyl)amino]- 3-pyridinyl]carbonyl] glycyl-3- (3-bromo-5- chloro-2-hydroxyphenyl)-b-alanine;

(3R)-N-[(1,6-dihydro-6-oxo-5-[(1,4,5,6- tetrahydro-5-hydroxy-2-pyrimidinyl) amino]-3- <BR> <BR> pyridinyl] carbonyl] glycyl-3- (3-bromo-5-chloro- 2-hydroxyphenyl)-b-alanine;

(3R)-N- [3-amino-5- [ (1,4,5,6-tetrahydro-5- hydroxy-2-pyrimidinyl) amino] benzoyl} glycyl-3- (3-bromo-5-chloro-2-hydroxyphenyl)-b-alanine;

(3R)-N- [3- [ (hydroxyamino) carbonyl]-5-<BR> <BR> [ (1,4,5,6-tetrahydro-5-hydroxy)-2- pyrimidinyl) amino] benzoyl] glycyl-3- (3-bromo-5- chloro-2-hydroxyphenyl)-b-alanine; (3R)-N- [3- [ (4-, 5-dihydro-lH-imidazol-2- yl) amino] benzoyl] glycyl-3- (3-bromo-5-chloro-2- hydroxyphenyl)-b-alanine; (3R)-N- [3- [(aminoiminomethyl)amino]benzoyl]glycyl-3-(3- bromo-5-chloro-2-hydroxyphenyl)-b-alanine;

(3R)-N- [3-hydroxy-5- [ (1,4,5,6-tetrahydro-5- hydroxy-2-pyrimidinyl) amino] benzoyl] glycyl-3- (3-bromo-5-chloro-2-hydroxyphenyl)-b-alanine; (3R)-N- [3-hydroxy-5- [ (1,4,5,6-tetrahydro-5- hydroxy-2-pyrimidinyl) amino] benzoyl] glycyl-3- (3,5-dichloro-2-hydroxyphenyl)-b-alanine; (3R)-N- [3-hydroxy-5- [ (1,4,5,6-tetrahydro-5- hydroxy-2-pyrimidinyl) amino] benzoyl] glycyl-3- (5-bromo-3-chloro-2-hydroxyphenyl)-b-alanine;

(3R)-N- [3-hydroxy-5- [ (1,4,5,6-tetrahydro-5- hydroxy-2-pyrimidinyl) amino] benzoyl] glycyl-3- <BR> <BR> (3-bromo-5-chloro-2-hydroxyphenyl)-b-alanine; b-[3-[[[3-[[4,5-dihydro-1H-imidazol-2- yl)amino] phenyl] sulfonyl] amino] phenyl]-3,5- difluorobenzenepropanoic acid; 3, 5-difluoro-b-[3-[[[3-[(1, 4,5,6-tetrahydro-2- pyrimidinyl) amino] benzoyl] amino] methyl] phenyl] benzenepropanoic acid;

(2E)-3-[3-ethyl-4-[[3-[(1, 4,5,6-tetrahydro-2- pyrimidinyl) amino] benzoyl] amino] phenyl]-2- propenoic acid; (2E)-3- [3- [2- [3- [ (4, 5-dihydro-lH-imidazol-2- yl) amino] phenyl]-2-oxoethoxy] phenyl]-2- propenoic acid;

(10S)-10,11-dihydro-3-[3-(2- pyridinylamino) propoxy]-5H- dibenzo [a, d] cycloheptene-10-acetic acid ; (2S)-7-[[(lH-benzimidazol-2- ylmethyl) methylamino] carbonyl]-2,3,4,5- tetrahydro-4-methyl-3-oxo-1H-1,4- benzodiazepine-2-acetic acid; (2S)-2,3,4,5-tetrahydro-4-methyl-7- [ [ [ (5- methyl-lH-imidazo [4,5-b] pyridin-2- yl] methyl] amino] carbonyl]-3-oxo-1H-1, 4- benzodiazepine-2-acetic acid;

(bR)-b-[[[(3R)-2-oxo-3-[2-(1, 5,6,7-tetrahydro- 1,8-naphthyridin-2-yl) ethyl]-1- pyrrolidinyl] acetyl] amino]-lH-indole-3- pentanoic acid;

I24) Vitaxin antibody (Ixsys); I25) Merck KGaA EMD-121974, cyclo [RGDf-N (Me) V-];

Still more preferred integrin antagonists include but are not limited to (10S)-10,11-dihydro-3- [3- (2- pyridinylamino) propoxy]-5H- dibenzo [a, d] cycloheptene-10-acetic acid;

(2S)-7-[[(lH-benzimidazol-2- ylmethyl) methylamino] carbonyl]-2,3,4,5- tetrahydro-4-methyl-3-oxo-lH-1,4- benzodiazepine-2-acetic acid; (2S)-2,3,4,5-tetrahydro-4-methyl-7- [ [ [ (5- methyl-lH-imidazo[4,5-b] pyridin-2- <BR> <BR> yl] methyl] amino] carbonyl]-3-oxo-lH-1,4- benzodiazepine-2-acetic acid; (bR)-b-[[[(3R)-2-oxo-3-[2-(1, 5,6,7-tetrahydro- 1,8-naphthyridin-2-yl)ethyl]-1- pyrrolidinyl] acetyl] amino]-lH-indole-3- pentanoic acid;

I24) Vitaxin antibody (Ixsys); I25) Merck KGaA EMD-121974, cyclo [RGDf-N (Me) V-];

The phrase"matrix metalloproteinase inhibitor"or "MMP inhibitor"includes agents that specifically inhibit a class of enzymes, the zinc metalloproteinases (metalloproteases). The zinc metalloproteinases are involved in the degradation of connective tissue or connective tissue components. These enzymes are released from resident tissue cells and/or invading inflammatory or tumor cells. Blocking the action of zinc metalloproteinases interferes with the creation of paths for newly forming blood vessels to follow.

Examples of MMP inhibitors are described in Golub, LM, Inhibition of Matrix Metalloproteinases: Therapeutic Applications (Annals of the New York Academy of Science, Vol 878). Robert A. Greenwald and Stanley Zucker (Eds.), June 1999), and is hereby incorporated by reference.

Connective tissue, extracellular matrix constituents and basement membranes are required

components of all mammals. These components are the biological materials that provide rigidity, differentiation, attachments and, in some cases, elasticity to biological systems including human beings and other mammals. Connective tissues components include, for example, collagen, elastin, proteoglycans, fibronectin and laminin. These biochemicals makeup, or are components of structures, such as skin, bone, teeth, tendon, cartilage, basement membrane, blood vessels, cornea and vitreous humor.

Under normal conditions, connective tissue turnover and/or repair processes are controlled and in equilibrium. The loss of this balance for whatever reason leads to a number of disease states. Inhibition of the enzymes responsible loss of equilibrium provides a control mechanism for this tissue decomposition and, therefore, a treatment for these diseases.

Degradation of connective tissue or connective tissue components is carried out by the action of proteinase enzymes released from resident tissue cells and/or invading inflammatory or tumor cells. A major class of enzymes involved in this function are the zinc metalloproteinases (metalloproteases).

The metalloprotease enzymes are divided into classes with some members having several different names in common use. Examples are: collagenase I (MMP-1, fibroblast collagenase; EC 3.4.24.3); collagenase II (MMP-8, neutrophil collagenase; EC 3.4.24.34), collagenase III (MMP-13), stromelysin 1 (MMP-3; EC 3.4.24.17), stromelysin 2 (MMP-10; EC 3.4.24.22), proteoglycanase, matrilysin (MMP-7), gelatinase A

(MMP-2,72kDa gelatinase, basement membrane collagenase; EC 3.4.24.24), gelatinase B (MMP-9,92kDa gelatinase; EC 3.4.24.35), stromelysin 3 (MMP-11), metalloelastase (MMP-12, HME, human macrophage elastase) and membrane MMP (MMP-14). MMP is an abbreviation or acronym representing the term Matrix Metalloprotease with the attached numerals providing differentiation between specific members of the MMP group.

The uncontrolled breakdown of connective tissue by metalloproteases is a feature of many pathological conditions. Examples include rheumatoid arthritis, osteoarthritis, septic arthritis; corneal, epidermal or gastric ulceration; tumor metastasis, invasion or angiogenesis; periodontal disease; proteinuria; Alzheimer's Disease; coronary thrombosis and bone disease. Defective injury repair processes also occur.

This can produce improper wound healing leading to weak repairs, adhesions and scarring. These latter defects can lead to disfigurement and/or permanent disabilities as with post-surgical adhesions.

Matrix metalloproteases are also involved in the biosynthesis of tumor necrosis factor (TNF) and inhibition of the production or action of TNF and related compounds is an important clinical disease treatment mechanism. TNF-a, for example, is a cytokine that at present is thought to be produced initially as a 28 kD cell-associated molecule. It is released as an active, 17 kD form that can mediate a large integer of deleterious effects in vitro and in vivo. For example, TNF can cause and/or contribute to the effects of inflammation, rheumatoid arthritis, autoimmune disease,

multiple sclerosis, graft rejection, fibrotic disease, cancer, infectious diseases, malaria, mycobacterial infection, meningitis, fever, psoriasis, cardiovascular/pulmonary effects such as post-ischemic reperfusion injury, congestive heart failure, hemorrhage, coagulation, hyperoxic alveolar injury, radiation damage and acute phase responses like those seen with infections and sepsis and during shock such as septic shock and hemodynamic shock. Chronic release of active TNF can cause cachexia and anorexia. TNF can be lethal. <BR> <BR> <BR> <BR> <BR> <BR> <P> TNF-a convertase is a metalloproteinase involved in<BR> <BR> <BR> <BR> <BR> <BR> <BR> <BR> the formation of active TNF-a. Inhibition of TNF-a<BR> <BR> <BR> <BR> <BR> <BR> <BR> <BR> <BR> convertase inhibits production of active TNF-a.

Compounds that inhibit both MMPs activity have been disclosed in, for example PCT Publication WO 94/24140.

Other compounds that inhibit both MMPs activity have also been disclosed in WO 94/02466. Still other compounds that inhibit both MMPs activity have been disclosed in WO 97/20824.

There remains a need for effective MMP and TNF-a convertase inhibiting agents. Compounds that inhibit MMPs such as collagenase, stromelysin and gelatinase have been shown to inhibit the release of TNF (Gearing et al. Nature 376,555-557 (1994)). McGeehan et al., Nature 376,558-561 (1994) also reports such findings.

MMPs are involved in other biochemical processes in mammals as well. Included is the control of ovulation, post-partum uterine involution, possibly implantation, cleavage of APP (ß-Amyloid Precursor Protein) to the amyloid plaque and inactivation of al-protease inhibitor

(a-PI). Inhibition of these metalloproteases permits the control of fertility and the treatment or prevention of Alzheimers Disease. In addition, increasing and maintaining the levels of an endogenous or administered serine protease inhibitor drug or biochemical such as a 1-PI supports the treatment and prevention of diseases such as emphysema, pulmonary diseases, inflammatory diseases and diseases of aging such as loss of skin or organ stretch and resiliency.

Inhibition of selected MMPs can also be desirable in other instances. Treatment of cancer and/or inhibition of metastasis and/or inhibition of angiogenesis are examples of approaches to the treatment of diseases wherein the selective inhibition of stromelysin (MMP-3), gelatinase (MMP-2), or collagenase III (MMP-13) are the relatively most important enzyme or enzymes to inhibit especially when compared with collagenase I (MMP-1). A drug that does not inhibit collagenase I can have a superior therapeutic profile.

Inhibitors of metalloproteases are known. Examples include natural biochemicals such as tissue inhibitor of metalloproteinase (TIMP), a2-macroglobulin and their analogs or derivatives. These are high molecular weight protein molecules that form inactive complexes with metalloproteases. An integer of smaller peptide-like compounds that inhibit metalloproteases have been described. Mercaptoamide peptidyl derivatives have shown ACE inhibition in vitro and in vivo. Angiotensin converting enzyme (ACE) aids in the production of angiotensin II, a potent pressor substance in mammals

and inhibition of this enzyme leads to the lowering of blood pressure.

Thiol group-containing amide or peptidyl amide- based metalloprotease (MMP) inhibitors are known as is shown in, for example, WO 95/12389. Thiol group- containing amide or peptidyl amide-based metalloprotease (MMP) inhibitors are also shown in WO 96/11209. Still furhter Thiol group-containing amide or peptidyl amide- based metalloprotease (MMP) inhibitors are shown in U. S.

Patent No. 4,595,700. Hydroxamate group-containing MMP inhibitors are disclosed in a number of published patent applications that disclose carbon back-boned compounds, such as in WO 95/29892. Other published patents include WO 97/24117. Additionally, EP 0 780 386 further discloses hydroxamate group-containing MMP inhibitors.

WO 90/05719 disclose hydroxamates that have a peptidyl back-bones or peptidomimetic back-bones. WO 93/20047 also discloses hydroxamates that have a peptidyl back- bones or peptidomimetic back-bones. Additionally, WO 95/09841 discloses disclose hydroxamates that have peptidyl back-bones or peptidomimetic back-bones. And WO 96/06074 further discloses hydroxamates that have peptidyl back-bones or peptidomimetic back-bones.

Schwartz et al., Progr. Med. Chem., 29: 271-334 (1992) also discloses disclose hydroxamates that have peptidyl back-bones or peptidomimetic back-bones. Furthermore, Rasmussen et al., Pharmacol. Ther., 75 (1): 69-75 (1997) discloses hydroxamates that have peptidyl back-bones or peptidomimetic back-bones. Also, Denis et al., Invest. <BR> <BR> <BR> <P>New Drugs, 15 (3) : 175-185 (1997) discloses hydroxamates

that have a peptidyl back-bones or peptidomimetic back- bones as well.

One possible problem associated with known MMP inhibitors is that such compounds often exhibit the same or similar inhibitory effects against each of the MMP enzymes. For example, the peptidomimetic hydroxamate known as batimastat is reported to exhibit ICso values of about 1 to about 20 nanomolar (nM) against each of MMP-1, MMP-2, MMP-3, MMP-7, and MMP-9. Marimastat, another peptidomimetic hydroxamate was reported to be another broad-spectrum MMP inhibitor with an enzyme inhibitory spectrum very similar to batimastat, except that marimastat exhibited an ICso value against MMP-3 of 230 nM. Rasmussen et al., Pharmacol. Ther., 75 (1) : 69- 75 (1997).

Meta analysis of data from Phase I/II studies using marimastat in patients with advanced, rapidly progressive, treatment-refractory solid tumor cancers (colorectal, pancreatic, ovarian, prostate), indicated a dose-related reduction in the rise of cancer-specific antigens used as surrogate markers for biological activity. The most common drug-related toxicity of marimastat in those clinical trials was musculoskeletal pain and stiffness, often commencing in the small joints in the hands, spreading to the arms and shoulder. A short dosing holiday of 1-3 weeks followed by dosage reduction permits treatment to continue. Rasmussen et al., Pharmacol. Ther., 75 (1): 69-75 (1997). It is thought that the lack of specificity of inhibitory effect among the MMPs may be the cause of that effect.

In view of the importance of hydroxamate MMP inhibitor compounds in the treatment of several diseases and the lack of enzyme specificity exhibited by two of the more potent drugs now in clinical trials, it would be beneficial to use hydroxamates of greater enzyme specificity. This would be particularly the case if the hydroxamate inhibitors exhibited limited inhibition of MMP-1 that is relatively ubiquitous and as yet not associated with any pathological condition, while exhibiting quite high inhibitory activity against one or more of MMP-2, MMP-9 or MMP-13 that are associated with several pathological conditions.

Non-limiting examples of matrix metalloproteinase inhibitors that may be used in the present invention are identified in Table No. 2, below.

Table No. 2. Matrix metalloproteinase inhibitors. Compound Trade Name Reference Dosage Biphenyl WO 97/18188 hydroxamate AG-3067 Winter Conf. (Agouron Med. Bio- Pharm. organic Inc.) Chem. 1997 January, 26- 31 AG-3340 WO 97/20824 50 mg/kg (Agouron treatment Pharm. of Lewis Inc.) lung Compound Trade Name Reference Dosage carcinomas in test animals AG-2024 (Agouron Pharm. Inc.) AG-3365 (Agouron Pharm. Inc.) 3 (S)-N-hydroxy- WO 97/20824. In female 4- (4- [4- FEBS (1992) Lewis rats, (imidazol-1-296 (3): 263 arthritis yl) phenoxy] benze model: dose nesulfonyl)-2,2- of 25 dimethyl-mg/kg/day tetrahydro-2H-gave 97.5% 1,4-thiazine-3- weight loss carboxamide, and inhibition derivatives thereof Heteroaryl WO 98/17643 succinamides derivatives AG-3296 (Agouron Pharm. Inc.) AG- Compound Trade Name Reference Dosage 3287 (Agour on Pharm. Inc.) AG-3293 (Agouron Pharm. Inc.) AG-3294 (Agouron Pharm. Inc.) AG-3067 Winter Conf (Agouron Med Bio- Pharm. organic Chem Inc.) 1997 January 26-31 08184432R,4S)-4-EP hydroxy-2- isobutyl-5- mercapto-N- [ (1S)-2,2- dimethyl-1- methylcarbamoylp ropyl] pentanamide N-alkyl, N-WO 98/16520 phenylsulfonyl- N'-hydroxamic acid derivatives of heteroaryl Compound Trade Name Reference Dosage carboxylic acids Novel N-alkyl, WO 98/16514 N- phenylsulfonyl- N'-hydroxamic acid derivatives of heteroaryl carboxylic acids Novel N-alkyl, WO 98/16506 N- phenylsulfonyl- N'-hydroxamic acid derivatives of cycloalkane carboxylic acids Novel N-alkyl, WO 98/16503 N- phenylsulfonyl- N'-hydroxamic acidderivatives of anthranilic acid sulfonamido-EP 03/98753 hydroxamic acid derivatives TIMP-3: WO 95/09918 polynucleotides encoding endogenous (human) peptides Compound Trade Name Reference Dosage (3alpha, WO 93/23075 5beta, 6alpha, 7al phabeta)-4',4'- (hexahydro-2,2- dimethyl-1,3- benzodioxole-5, 6-diyl) bis (2,6- piperazinedione) and derivatives thereof BE-16627B WO 91/08222. Int. J. Cancer 1994 58 5 730- 735 (2S)-4- (4- (4- WO 96/15096 chlorophenyl) phe nyl)-4-oxo-2- (2- phthalimidoethyl ) butanoic acid Bay-12-WO 96/15096 10 to 400 9566 mg/day 4-oxo-2- (2- WO 97/43238 phthalimidoethyl ) alkanoic acid derivatives Novel 4- (4- WO 97/43237 Alkynylphenyl) 4-oxobutanoic Compound Trade Name Reference Dosage acid derivatives Substituted 4-WO 96/15096 biarylbutyric or 5- biarylpentanoic acids and derivatives Substituted 4-WO 98/22436 biphenyl-4- hydroxybutyric acid derivatives 2R, S)-HONH-CO- J Med Chem CH (i-Bu)-CO-Ala- 1998 41 3 Gly-NH2,339-345 batimastat; BB-WO 90/05719 15 to 135 94; Hydroxamic mg/m2 acid based administer- collagenase ed intra- inhibitors pleurally Hydroxamic acid WO 90/05719 based collagenase inhibitors marimastat BB-WO 94/02447 5 to 800 mg 2516; Hydroxamic daily acid derivatives alpha-cycloalkyl Bio-organic analogs of Med Chem marimastat Lett 1998 8 11 1359- Compound Trade Name Reference Dosage 1364 GI-245402 (BB-2983) Hydroxamic acid WO 94/21625 derivatives Succinyl WO 95/32944 hydroxamic acid, N-formyl-N- hydroxyamino carboxylicacid and succinic acid amide derivatives hydroxamic acid, WO 97/19053 N-formyl-N- hydroxyamino and carboxylic acid derivatives, pseudopeptide WO 97/19050 hydroxamic and carboxylic acid derivatives from the corresponding lactone and alpha-amino acid Succinic acid WO 97/03966. amide GB 95/00111. derivatives GB 95/00121. Hydroxamic acid WO 97/02239 Compound Trade Name Reference Dosage derivatives Succinamidyl WO 96/33165 (alpha substituted) hydroxamic acid derivatives 96/25156(2S,3R)-3-[2,2-WO dimethyl-lS- (thiazol-2- ylcarbamoyl) pro- pylcarbamoyl]-5- methyl-2- (prop- 2-enyl) hexano- hydroxanic acid and derivatives thereof Hydroxamic or WO 96/16931 carboxylic acid derivatives hydroxamic and WO 96/06074 carboxylic acids 2- [ (IS)-I- ( (lR)- WO 98/23588 2-[[1,1- biphenyl]-4- ylmethylthio]-l- [(1S)-2,2- dimethyl-1- (methylcarbamoyl ) propylcarbamoyl ] ethylcarbamoyl) Compound Trade Name Reference Dosage -4- (1,3-dioxo- 1,3- dihydroisoindol- 2-yl) butylthio]- acetate, and derivatives thereof Hydroxamic acid WO 95/09841 derivatives as inhibitors of cytokine production Hydroxamic acid WO 94/24140 derivatives Aromatic or WO 95/19956 heteroaryl substituted hydroxamic or carboxylic acid derivatives Hydroxamic acid WO 95/19957 Doses are derivatives preferably 1 to 100 mg/kg. Hydroxamic acid WO 95/19961 Doses are and carboxylic preferably acid derivatives 1 to 100 mg/kg. Butanediamide, BB-1433 At 50 mg/kg N1-bid. p. o. Compound Trade Name Reference Dosage [l(cyclohexyl- inhibited methyl)-2 bone (methylamino)-2-mineral oxoethyl]-N4,3- density dihydroxy-2- (2- loss methylpropyl)-, [2R [N1 (S*), 2R*, 3 S*]]- tetracycline EP 733369 D-penicill- analogs and D-amine penicillamine reduced allergic encephaliti s symptom scores in a dose dependent manner at 27,125 and 375 mug with complete inhibition CDP-845 Biochem Pharmacol 1990 39 12 2041-2049 succinamide WO 95/04033 oral derivatives bioavail- ability by Compound Trade Name Reference Dosage murine pleural cavity assay in the presence of gelatinase: Between 73% and 100% inhibition was displayed at 10 mg/kg for six of the compounds. The seventh displayed 100% inhibition at 80 mg/kg. Peptidyl WO 94/25435. derivatives WO 94/25434 Mercaptoalkyl-WO 97/19075 peptidyl compounds having an imidazole substituent mercaptoalkyl-WO 97/38007. Compound Trade Name Reference Dosage peptide WO 95/12389. derivatives WO 96/11209. Mercaptoalkyl-WO 97/37974 amide derivatives arylsulfonyl-WO 97/37973. hydrazine WO 95/12389 derivatives N-acetylthio-WO 96/35714 lacetyl-N- (3- phthalimidopropy l)-L-leucyl-L- phenylalanine N- methylamide 2-acetylsulfany-WO 96/35712 dosages of 1-5-phthalimido-about 0.5 pentanoyl-L-mg to 3.5 g leucineN- (2- per day for phenylethyl)-the amide treatment of inflam- mation 5-phthalimido-WO 96/35711 pentanoyl-L- leucyl-L- phenylalanineN- methylamide peptidyl WO 98/06696 derivatives 4- [4- WO 98/05635 Compound Trade Name Reference Dosage (methoxycarbonyl methoxy)-3,5- dimethylphenyl]- 2-methyl-1 (2H)- phthalazinone, and hydroxamic and carboxylic acid derivatives thio-substituted WO 97/12902 peptides Mercaptoamides WO 97/12861 Peptidyl WO 96/35687 derivatives having SH or acylo groups which are amides, primary amides or thioamides D-5410 (Chiro- science Group plc) WO 95/13289 CH-104, (Chiro- science Group plc) D-2163 (Chiro Compound Trade Name Reference Dosage Science Ltd.) D-1927 (Chiro Science Ltd.) Dermastat (Colla- Genex Phar- maceu- tical Inc.) Metastat (Colla- Genex) Osteostat (Colla- Genex Phar- maceu- tical Inc.) doxy-Gingival cycline; crevicular Roche; fluid Periostat collagenase is reported to be inhibited Compound Trade Name Reference Dosage at concentra- tions of 5- 10 microg /ml or 15- 30 microM 2S, 5R, 6S-3-WO 97/18207 aza-4-oxo-10- oxa-5-isobutyl- 2- (N- methylcarbox- amido)- [10] paracyclopha ne-6-N- hydroxycarboxami de hydroxamic acid WO 96/33176 and amino- carboxylate compounds N-hydroxamic WO 96/33166 derivatives of succinamide Macrocyclic J Med Chem amino 1998 41 11 carboxylates 1749-1751 SE-205 (Du Bio-organic Pont Merck Med Chem Pharm Co.) Lett 1998 8 7 837-842. Compound Trade Name Reference Dosage J Med Chem 1998 41 11 1745-1748 macrocyclic matrix metalloprotease- 8 inhibitors Hydroxamic acid WO 95/22966 and carboxylic acid derivatives succinamid US 5256657 derivatives mercaptosulfide WO 95/09833 derivatives sulfoximine and WO 95/09620 sulfodiimine derivatised peptides water soluble WO 96/33968 MMP inhibitors hydantoin EP 06/40594 derivatives Piperazine WO 98/27069 derivatives GI-155704A J Med Chem 1994 37 5 674. Bioorganic Med Chem Lett 1996 6 Compound Trade Name Reference Dosage 16 1905- 1910 Cyclic imide EP 05/20573 derivatives. 3- (mercapto- WO 97/48685 methyl) hexa- hydro-2,5- pyrazinedione derivatives beta-WO 96/40738 mercaptoketone and beta- mercaptoalcohol derivatives ilomastat US 5114953. eye drops MPI; GM-Cancer Res containing 6001; 1994 54 17 ilomastat Galardin 4715-4718 (800 microg/ml) Cyclic and WO 97/18194 heterocyclic N- substituted alpha- iminohydroxamic and carboxylic acids Aminomethyl-EP 703239 phosphonic and aminomethyl- phosphinic acids Compound Trade Name Reference Dosage derivatives 3-Mercapto-WO 98/12211 acetylamino-1,5- substituted-2- oxo-azepan derivatives 2-substituted WO 94/04531 indane-2- mercaptoacetyl- amide tricyclic derivatives Ro-2756 (Roche Holding AG) Ro-26-4325 (Roche Holding AG) Ro-26-5726 (Roche Holding AG) Ro-26-6307 (Roche Holding AG) Ro-31-9790 J Am Soc mono- (Roche Nephrol 1995 arthritis Holding 6 3 904. in rat: 100 Compound Trade Name Reference Dosage AG) Inflamm Res mg/kg/day 1995 44 8 345-349 substituted and WO 92/09556 unsubstituted hydroxamates (specifically N- [D, L-2-isobutyl- 3- (N'-hydroxy- carbonyl-amido)- propanoyl] trypto phanmethylamide) GM6001, N- (2 (R)- WO 95/24921 2- (hydroxyaminocar bonylmethyl)-4- methylpentanoyl) -L-tryptophan methylamide. Oligonucleotice (c-jun) Sulfated WO 98/11141 polysaccharides KB-R7785; Life Sci KB-R8301; 1997 61 8 KB-R8845 795-803 Fas ligand WO 97/09066 solubilization inhibitor gelastatin AB, Compound Trade Name Reference Dosage KRIBB KT5-12 Faseb J 1998 (Kotobuki 12 5 A773 Seiyaku Co (4482) Ltd.) 2-(N2-[(2R)-2-GB(N2-[(2R)-2-GB 23/18789 (2-hydroxyamino- 2-oxoethyl)-5- (4- methoxyphenoxy) p entanoyl]-L- phenylalanylamin o) ethanesulfonam ide, and carboxylic acid derivatives thereof Chromone EP 758649 2- derivatives Pyrolylthio -chromone in a murine melanoma model produced 37% inhibition at 100 mg/kg Esculetin EP 719770 derivatives, Compound Trade Name Reference Dosage substituted and WO 92/09563 unsubstituted hyroxyureas and reverse hydroxamates Synthetic MMP WO 94/22309 inhibitors (ex. N- (D, L-2- isobutyl-3- (N'- hydroxycarbonyla mido) propanoyl) t ryptophan methylamide) Reverse WO 95/19965 in female hydroxamates and mice hydroxyureas infected w/murine melanoma- init 80 mu g followed by 150 mg/kg/day N-US 5629343 (mercaptoacyl)- aryl derivatives of leucine and phenylalanine N-carboxyalkyl WO 95/29689 derivatives Substituted GB 22/82598 Inflammatio Compound Trade Name Reference Dosage cyclic n is stated derivatives to be effectively treated by oral administrat ion of 0.01 to 50 mg/kg Substituted n-GB 22/72441 carboxyalkyldi- peptides (2S, 4R)-2-WO 97/11936 methyl-4- (phenylamino- carbonylmethyl- aminocarbonyl)- 6- (4-propyl- phenyl) hexanoic acid, and carboxylic acid derivatives Substituted US 5403952 cyclic derivatives Thiol WO 98/03166 sulfonamide metalloprotease inhibitors Thiol sulfone WO 98/03164 metalloprotein- Compound Trade Name Reference Dosage ase inhibitors formulations WO 97/47296 containing vanadium compounds and N- acetylcysteine NSC- 683551; COL-3 (National Cancer Institute) BB-3644 (Neures Ltd.) Arylsulfonamido-CGS-Int Congr 600 mg tid substituted 27023A; Inflamm Res (Ph I- hydroxamic acids CGS-25966 Assoc 1994 colorectal 7th Abs 73. and EP-00606046 melanoma patients); 100 mg/kg in food in osteoarthri tis model rabbits alpha-WO 97/22587 Substituted arylsulfonamido hydroxamic acid Compound Trade Name Reference Dosage derivatives Arylsulfonamido-US 5455258 active at substituted 30 mg/kg in hydroxamic acids in vivo assay Arylsulfonamido-WO 96/00214 substituted hydroxamic acids 2S, 3S)-N-WO 98/14424 hydroxy-5- methyl-2- [2- (2- methoxyethoxy) et hoxymethyl]-3- (N- [ (1S)-1- (N- methylcarbamoyl) -2- phenylethyl] carb amoyl) hexanamide and Hydroxamic acid deriva- tives arylsulfonamido-WO 96/40101 in tumor substituted model mice: hydroxamic acids administere d for 7 to 17 days at a dosage of 30 mg/kg twice daily Aryl (sulfide, WO 97/49679 Compound Trade Name Reference Dosage sulfoxide and sulfone) derivatives Phenylsulfon-WO 97/45402 amide derivatives Arylsulfonamido-EP 757037 aminoacid derivative A1PDX (Oregon Health Sciences University) futoenone Bio-organic analogs Med Chem Lett 1995 5 15 1637- 1642 debromohymeni-WO 96/40147 preferred aldisine and 1-30 mg/day related compounds amide WO 96/40745 derivatives of 5-amino-1,3,4- thiadiazolones 3S- (4- (N- WO 94/21612 hydroxylamino)- 2R- isobutylsuccinyl )amino-l- Compound Trade Name Reference Dosage methoxymethyl- 3,4- dihydrocarbostyr il and deriviatives therof Carbostyryl JP 8325232 derivatives OPB-3206 (Otsuka Pharmaceutical Co, Ltd.) Arylsulfonyl WO 96/33172 hydroxamic acid derivatives Cyclic sulfone EP 818442 derivatives arylsulfonamido WO 96/27583 N-hydroxamic acid derivatives of butyric acid Arylsulfonyl-WO 98/07697 amino hydroxamic acid derivatives phosphinate-WO 98/03516 based derivatives cyclopentyl-WO 92/14706 substituted glutaramide derivatives Compound Trade Name Reference Dosage N-hydroxamic WO 97/49674 acidsuccinamide derivatives Thiadiazole WO 97/48688 amide MMP inhibitors. (S)-1-[2-WO 97/40031 [ [ [ (4,5-Dihydro- 5-thioxo-1,3,4- thiadiazol-2- yl)amino3- carbonyl] amino]- l-oxo-3- (pentafluoro- phenyl)propyl]- 4- (2-pyridinyl)- piperazine hydroxamic acid WO 97/32846 derivatives of pyrrolidone-3- acetamide. alpha-WO 98/17645 arylsulfonamido- N-hydroxamic acid derivatives beta-WO 98/13340 Sulfonylhydrox- amic acids Hydroxamic acid US 5712300 derivatives Compound Trade Name Reference Dosage PNU-99533 (Pharmacia & UpJohn Inc.) PNU-143677 (Pharmacia & UpJohn Inc.) POL-641 (Poli- farma) Peptidomimetic WO 96/20,18. inhibitors WO 96/29313. WO 98/08814. WO 98/08815. WO 98/08850. WO 98/08822. WO 98/08823. WO 98/08825. WO 98/08827. 2R)-N- ()-caprol-WO 96/29313 rheumatoid hydroxycarboxami actam-arthritis: demethyldecanoic (3S)-amine female acid amide of subject- 1N-50 mg po (carbomethoxy-for 2 yrs; methyl) male subject- 70 mg po daily for 5 Compound Trade Name Reference Dosage yrs; corneal ulcer: male subject 0 10 mg in saline soln for 2 months, 2 times/day 3-(N-[(N-WO 96/20918 Hydroxyaminocarb onyl)methyl]-N- isobutylaminocar bonyl)-2- (R)- isobutylpro- panoyl-L- phenylalanine amide N-hydroxy-WO 98/08853 phosphinic acid amides N'-arylsulfonyl WO 98/08850 derivatives of spirocyclic-N- hydroxycarbox- amides N'-arylsulfonyl WO 98/08827 derivatives of thiazepinone and Compound Trade Name Reference Dosage azepinone-N- hydroxycarbox- amides Substituted WO 98/08825 piperazine derivatives N'-arylsulfonyl WO 98/08823 derivatives of pyrimidine, thiazepine and diazepine-N- hydroxycarbox- amides Substituted WO 98/08815 pyrrolidine derivatives Substituted WO 98/08814 heterocycles Substituted 1,3- WO 09/08822 diheterocyclic derivatives substituted 5-WO 98/25949 amino-1,2,4- thiadiazole-2- thiones Hydroxamic acid WO 97/24117 derivatives which inhibit TNF production. 6-methoxy-WO 97/37658 Compound Trade Name Reference Dosage 1,2,3,4- tetrahydro- norharman-1- carboxylic acid RS-130830 Arthritis Rheum 1997 40 9 SUPPL. S128 Aralkyl MMP WO 96/16027 inhibitors (ex. N- (2R- carboxymethyl-5- (biphen-4- yl) pentanoyl)-L- t-butylglycine- N'- (pyridin-4- yl) carboxamide) Ro-32-3555 (Roche Holding AG) Ro-32-1278 (Roche Holding AG) Ro-32-1541 (Roche Holding AG) Ro-31-3790 Arthritic Compound Trade Name Reference Dosage (Roche model rats: Holding Protection AG) of cartilage degradation following oral administrat ion; ED50 = 10 mg/kg po (3R, 11S)-N- WO 95/04735 hydroxy-5- methyl-3- (10- oxo-1,9- diazatricyclo- (11.6.1.014,19) e icosa- 13 (20), 14 (19), 15 ,17-tetraen- 11- ylcarbamoyl) hexa namide and derivatives thereof Bridged indoles WO 96/23791 (Roche Holding AG) substituted EP 780386 phenylsulfonyl acetamide, propionamide and Compound Trade Name Reference Dosage carboxamide compounds 5- (4'-biphenyl)- WO 97/23465 5- [N- (4- nitrophenyl) piperazinyl] barbituric acid Malonic acid EP 716086 based matrix metalloproteinas e inhibitors phenyl WO 95/12603 carboxamide derivatives Malonic acid EP 716086 based mmp inhibitors (specifically 2- (4-acetylamino- benzoyl)-4- methylpentanoic acid) Hydroxyl amine Ro-31-EP 236872 derivatives 4724; Ro- 31-7467;

The following individual patent references listed in Table No. 3 below, hereby individually incorporated by reference, describe various MMP inhibitors suitable

for use in the present invention described herein, and processes for their manufacture.

Table No. 3. MMP inhibitors EP 189784 US 4609667 WO 98/25949 WO 98/25580 JP 10130257 WO 98/17655 WO 98/17645 US 5760027 US 5756545 WO 98/22436 WO 98/16514 WO 98/16506 WO 98/13340 WO 98/16520 WO 98/16503 WO 98/12211 WO 98/11908 WO 98/15525 WO 98/14424 WO 98/09958 WO 23/18789WO98/09940WOGB 98/09934 JP 98/08853WO98/06711WO98/05635WO WO 98/07697WO98/03516WOWO 98/03166 WO 23/17182WO98/05353WOGB 98/04572 WO 98/02578WO97/48688WOWO 97/48685 WO 97/47599WO97/43247WOWO 97/43240 WO 818443EP818442WO97/45402EP WO 97/44315WO97/38705USWO 5679700 WO 97/43245 WO 97/43239 WO 97/43237 JP 09227539 WO 97/42168 US 5686419 WO 97/37974 WO 97/36580 WO 97/24117US5646316WOWO 97/23459 WO 780386DE19548624WOEP 97/19068 WO 97/19050WO97/18188WOWO 97/18194 WO 97/18183 WO 97/17088 DE 19542189 WO 97/15553 WO 97/12902 WO 97/12861 WO 97/11936 WO 97/11693 WO 97/09066 JP 09025293 EP 75/8649 WO 97/03966 WO 97/03783 EP 75/7984 WO 97/02239 WO 96/40745 WO 96/40738 WO 96/40737 JP 08/311096 WO 96/40204 WO 96/40147 WO 96/38434 WO 96/35714 WO 96/35712 WO 96/35711 WO 96/35687 EP 74,3,070 WO 96/33968 WO 96/33165 WO 96/33176 WO 96/33172 WO 96/33166 WO 23/00190WO96/29313EPGB 73/6302 WO 733369WO96/26223WOEP 96/27583 WO 22/98423WO96/23791WOGB 96/23505 GB 22/97324 DE 19501032 WO 96/20918 US 5532265 EP 96/17838WO96/16931WOWO 96/16648 WO 716086WO96/15096JPEP 08104628 WO 96/13523 JP 08081443 WO 96/11209 EP 703239 WO 96/06074 WO 95/35276 WO 96/00214 WO 95/33731 WO 95/32944WO95/29892WOWO 95/29689 CA 95/24921WO95/24199WOWO 95/23790 WO 22/87023WO95/19965WOGB 95/19961 WO 95/19957WOWO WO 95/13380 WO 95/09918WO95/09841WOWO 95/09833 WO 95/08327GB22/82598WOWO 95/07695 WO 95/04735WO95/04033WOWO 95/02603 WO 626378WO94/25435WOEP 94/25434 WO 94/24140WO94/24140EPWO 622079 WO 06256209WO94/21625FRJP 27/03053 EP 606046 WO 94/12169 WO 94/11395 GB 22/72441 WO 94/07481 WO 94/04190 WO 94/00119 GB 22/68934 WO 94/02446 EP 575844 WO 93/24475 WO 93/24449 US 5270326 US 5256657 WO 93/20047 WO 93/18794 WO 93/14199 WO 93/14096 WO 93/13741 WO 93/09090 EP 53/2465 EP 532156 WO 93/00427 WO 92/21360 WO 92/09563 WO 92/09556 EP 48/9579 EP 489577 US 5114953 EP 45/5818 US 5010062 AU 90/53158 WO 97/19075 US 7488460 US 7494796 US 7317407 EP 277428 EP 23/2027 WO 96/15096 WO 97/20824 US 5837696

The Marimastat used in the therapeutic combinations of the present invention can be prepared in the manner set forth in WO 94/02,447.

The Bay-12-9566 used in the therapeutic combinations of the present invention can be prepared in the manner set forth in WO 96/15,096.

The AG-3340 used in the therapeutic combinations of the present invention can be prepared in the manner set forth in WO 97/20,824.

The Metastat used in the therapeutic combinations of the present invention can be prepared in the manner set forth in U. S. Patent No. 5,837,696.

The D-2163 used in the therapeutic combinations of the present invention can be prepared in the manner set forth in WO 97/19,075.

More preferred zinc matrix metalloproteinase inhibitors include those described in the individual U. S. Patent applications, PCT publications and U. S.

Patents listed below in Table No. 4, and are hereby individually incorporated by reference.

Table No. 4. More preferred zinc matrix metalloproteinase inhibitors U. S. Patent Application Serial Number 97/12,873 U. S. Patent Application Serial Number 97/12,874 U. S. Patent Application Serial Number 98/04,299 U. S. Patent Application Serial Number 98/04,273 U. S. Patent Application Serial Number 98/04,297 U. S. Patent Application Serial Number 98/04,300 U. S. Patent Application Serial Number 60/119,181 WO 94/02447 WO 96/15096 WO 97/20824 WO 97/19075 US 5837696

Even more preferred zinc matrix metalloproteinase inhibitors that may be used in the present invention include:

N-hydroxy-1-(4-methylphenyl)-4-[[4-[4- (trifluoromethyl) phenoxy] phenyl] sulfonyl]-4- piperidinecarboxamide monohydrochloride;

1-cyclopropyl-N-hydroxy-4-[[4-[4- (trifluoromethoxy) phenoxy] phenyl] sulfonyl]-4- piperidinecarboxamide monohydrochloride; N-hydroxy-1-(phenylmethyl)-4-[[4-[4 (trifluoromethoxy) phenoxy]-l- piperidinyl] sulfonyl]-4-piperidinecarboxamide monohydrochloride;

N-hydroxy-1-(4-pyridinylmethyl)-4-[[4-[4- (trifluoromethyl) phenoxy] phenyl] sulfonyl]-4- piperidinecarboxamide dihydrochloride; N-hydroxy-2,3-dimethoxy-6- [ [4- [4- (trifluoromethyl) phenoxy]-l- piperidinyl] sulfonyl] benzamide;

N-hydroxy-1-(4-pyridinylmethyl)-4-[[4-[4- (trifluoromethyl) phenoxy] phenyl] sulfonyl]-4- piperidinecarboxamide dihydrochloride; N-hydroxy-1-(3-pyridinylmethyl)-4-[[4-[4- (trifluoromethyl) phenoxy] phenyl] sulfonyl]-4- piperidinecarboxamide dihydrochloride;

N-hydroxy-1-(2-pyridinylmethyl)-4-[[4-[4- (trifluoromethyl) phenoxy] phenyl] sulfonyl]-4- piperidinecarboxamide monohydrochloride; British Biotech BB-2516 (Marimastat), N4- [2,2- dimethyl-l-[(methylamino) carbonyl] propyl]- N1,2-dihydroxy-3 (2-methylpropyl)-, [2S- [N4 (R*), 2R*, 3S*]]-);

Bayer Ag Bay-12-9566,4- [ (4'-chloro [l, l'- iphenyl]-4-yl) oxy]-2- [(phenylthio) methyl] butanoic acid; Agouron Pharmaceuticals AG-3340, N-hydroxy-2,2 dimethyl-4- [ [4- (4-pyridinyloxy) phenyl]- sulfonyl]-3-thiomorpholinecarboxamide;

M12) CollaGenex Pharmaceuticals CMT-3 (Metastat), 6-demethyl-6-deoxy-4- dedimethylaminotetracycline; M13) Chiroscience D-2163,2- [1S- ( [ (2R, S)- acetylmercapto-5-phthalimido] pentanoyl- L- leucyl)amino- 3-methylbutyl] imidazole; N-hydroxy-4-[[4-(phenylthio)phenyl]sulfonyl]- 1- (2-propynyl)-4-piperidinecarboxamide monohydrochloride; N-hydroxy-1-(2-methoxyethyl)-4-[[4-[4 (trifluoromethoxy) phenoxy] phenyl] sulfonyl]-4- piperidinecarboxamide monohydrochloride;

N-hydroxy-1-(2-methoxyethyl)-4-[[4-[4- (trifluoromethyl) phenoxy] phenyl] sulfonyl]-4- piperidinearboxamide; 1-cyclopropyl-N-hydroxy-4-[[4-[4- (trifluoromethyl) phenoxy] phenyl] sulfonyl]-4-

4- [ [4- (cyclohexylthio) phenyl] sulfonyl]-N- hydroxy-l- (2-propynyl)-4-piperidinecarboxamide monohydrochloride; 4- [ [4- (4- chlorophenoxy) phenyl] sulfonyl] tetrahydro-N- hydroxy-2H-pyran-4-carboxamide; N-hydroxy-4- [ [4- (4- methoxyphenoxy) phenyl) sulfonyl]-1-(2- propynyl)-4-piperidinecarboxamide;

1-cyclopropyl-4-[[4-[(4- fluorophenyl) thio] phenyl] sulfonyl]-N-hydroxy- 4-piperidinecarboxamide; 1-cyclopropyl-N-hydroxy-4-[[4- (phenylthio) phenyl] sulfonyl]-4- piperidinecarboxamide; tetrahydro-N-hydroxy-4- [ [4- (4- pyridinylthio) phenyl] sulfonyl]-2H-pyran-4- carboxamide;

tetrahydro-N-hydroxy-4- [ [4- [4- (trifluoromethyl) phenoxy] phenyl] sulfonyl]-2H- pyran-4-carboxamide.

Still more preferred MMP inhibitors include: N-hydroxy-1-(4-methylphenyl)-4-[[4-[4- (trifluoromethyl) phenoxy] phenyl] sulfonyl]-4- piperidinecarboxamide monohydrochloride;

1-cyclopropyl-N-hydroxy-4-[[4-[4- (trifluoromethoxy) phenoxy] phenyl] sulfonyl]-4- piperidinecarboxamide monohydrochloride; N-hydroxy-1-(phenylmethyl)-4-[[4-[4- (trifluoromethoxy) phenoxy]-l- piperidinyl] sulfonyl]-4-piperidinecarboxamide monohydrochloride;

N-hydroxy-1-(4-pyridinylmethyl)-4-[[4-[4- (trifluoromethyl) phenoxy] phenyl] sulfonyl]-4- piperidinecarboxamide dihydrochloride; N-hydroxy-2,3-dimethoxy-6- [ [4- [4- (trifluoromethyl) phenoxy]-l- piperidinyl] sulfonyl] benzamide;

N-hydroxy-1-(4-pyridinylmethyl)-4-[[4-[4- (trifluoromethyl) phenoxy] phenyl] sulfonyl]-4- piperidinecarboxamide dihydrochloride; N-hydroxy-1-(3-pyridinylmethyl)-4-[[4-[4- (trifluoromethyl) phenoxy] phenyl] sulfonyl]-4- piperidinecarboxamide dihydrochloride;

N-hydroxy-1-(2-pyridinylmethyl)-4-[[4-[4- (trifluoromethyl) phenoxy] phenyl] sulfonyl]-4- piperidinecarboxamide monohydrochloride; British Biotech BB-2516 (Marimastat), N4- [2,2- dimethyl-l-[(methylamino) carbonyl] propyl]- N1,2-dihydroxy-3 (2-methylpropyl)-, [2S- [N4 (R*), 2R*, 3S*]]-);

Bayer Ag Bay-12-9566,4- [ (4'-chloro [l, l'- iphenyl]-4-yl)oxy]-2- [ (phenylthio) methyl] butanoic acid; Agouron Pharmaceuticals AG-3340, N-hydroxy- 2,2- dimethyl-4- [ [4- (4- pyridinyloxy) phenyl] sulfonyl]- 3- thiomorpholinecarboxamide; M12) CollaGenex Pharmaceuticals CMT-3 (Metastat), 6-demethyl-6-deoxy-4-dedimethylaminotetracycline; M13) Chiroscience D-2163,2- [1S- ( [ (2R, S)- acetylmercapto-5-phthalimido] pentanoyl-L- leucyl) amino- 3-methylbutyl] imidazole.

The phrase"antineoplastic agents"includes agents that exert antineoplastic effects, i. e., prevent the development, maturation, or spread of neoplastic cells, directly on the tumor cell, e. g., by cytostatic or cytocidal effects, and not indirectly through mechanisms such as biological response modification. There are large numbers of antineoplastic agents available in commercial use, in clinical evaluation and in pre- clinical development, which could be included in the present invention for treatment of neoplasia by combination drug chemotherapy. For convenience of discussion, antineoplastic agents are classified into the following classes, subtypes and species: ACE inhibitors, alkylating agents, angiogenesis inhibitors, angiostatin, anthracyclines/DNA intercalators, anti-cancer antibiotics or antibiotic-type agents, antimetabolites, antimetastatic compounds, asparaginases, bisphosphonates, cGMP phosphodiesterase inhibitors, calcium carbonate, cyclooxygenase-2 inhibitors DHA derivatives, DNA topoisomerase,

endostatin, epipodophylotoxins, genistein, hormonal anticancer agents, hydrophilic bile acids (URSO), immunomodulators or immunological agents, integrin antagonists interferon antagonists or agents, MMP inhibitors, miscellaneous antineoplastic agents, monoclonal antibodies, nitrosoureas, NSAIDs, ornithine decarboxylase inhibitors, pBATTs, radio/chemo sensitizers/protectors, retinoids selective inhibitors of proliferation and migration of endothelial cells, selenium, stromelysin inhibitors, taxanes, vaccines, and vinca alkaloids.

The major categories that some preferred antineoplastic agents fall into include antimetabolite agents, alkylating agents, antibiotic-type agents, hormonal anticancer agents, immunological agents, interferon-type agents, and a category of miscellaneous antineoplastic agents. Some antineoplastic agents operate through multiple or unknown mechanisms and can thus be classified into more than one category.

A first family of antineoplastic agents which may be used in combination with the present invention consists of antimetabolite-type antineoplastic agents.

Antimetabolites are typically reversible or irreversible enzyme inhibitors, or compounds that otherwise interfere with the replication, translation or transcription of nucleic acids. Suitable antimetabolite antineoplastic agents that may be used in the present invention include, but are not limited to acanthifolic acid, aminothiadiazole, anastrozole, bicalutamide, brequinar sodium, capecitabine, carmofur, Ciba-Geigy CGP-30694, cladribine, cyclopentyl cytosine, cytarabine phosphate stearate, cytarabine conjugates, cytarabine ocfosfate, Lilly DATHF, Merrel Dow DDFC, dezaguanine, dideoxycytidine, dideoxyguanosine, didox, <BR> <BR> <BR> <BR> Yoshitomi DMDC, doxifluridine, Wellcome EHNA, Merck & Co. EX-015, fazarabine, finasteride, floxuridine, fludarabine phosphate, N- (2'-furanidyl)-5-fluorouracil, Daiichi Seiyaku FO-152, fluorouracil (5-FU), 5-FU- fibrinogen, isopropyl pyrrolizine, Lilly LY-188011, Lilly LY-264618, methobenzaprim, methotrexate, Wellcome MZPES, nafarelin, norspermidine, nolvadex, NCI NSC- 127716, NCI NSC-264880, NCI NSC-39661, NCI NSC-612567, Warner-Lambert PALA, pentostatin, piritrexim, plicamycin, Asahi Chemical PL-AC, stearate; Takeda TAC- 788, thioguanine, tiazofurin, Erbamont TIF, trimetrexate, tyrosine kinase inhibitors, tyrosine protein kinase inhibitors, Taiho UFT, toremifene, and uricytin.

Preferred antimetabolite agents that may be used in the present invention include, but are not limited to, those identified in Table No. 5, below.

Table No. 5. Antimetabolite agents Compound Common Company Reference Dosage Name/ Trade Name 1,3- anastrozole Zeneca EP 296749 1-mg/day Benzenediaceto ARIMIDEX# nitrile, alpha, alpha, alpha', a tetramethyl-5- (1H-1,2,4- triazol-1-ylme thyl)- Propanamide, bicalutamid Zeneca EP 100172 50 mg once N- [4-cyano-3- e; CASODEX# daily (trifluorometh yl)phenyl]-3- [ (4- fluorophenyl) sulfonyl]-2- hydroxy-2- methyl-, (+/- ) _ capecitabin Roche US 5472949 e Adenosine, 2-cladribine; Johnson & EP 173059 0.09 chloro-2'- 2-CdA; Johnson mg/kg/day LEUSTAT;for7deoxy-;2- chloro-2'-LEUSTA-days. deoxy-(beta)-TIN#; D-adenosine) LEUSTA-TINS in-jection; l LEUSTATINSR) ;RWJ- 26251; l 2(1H)-2(1H)-cytarabine Yamasa EP 239015 - 300 Pyrimidinone, ocfosfate; Corp mg/day for 4-amino-l- [5- ara CMP 2 weeks 0-stearyl [hydroxy (octad ester; C- ecyloxy) phosph 18-PCA; inyl]-beta-D-cytarabine arabinofuranos phosphate yl]-, stearate; monosodiumStarasid; Compound Common Company Reference Dosage Name/ TradeName salt YNK-01; CYTOSAR-U# 4-Azaandrost-4-Azaandrost-finasteride Merck EP 155096 PROPECIA#1-ene-17-; Co carboxamide, N- (1, 1- dimethylethyl) -3-oxo-, (5alpha,17beta ) _ fluorouraci S 4336381 1 (5-FU) Fludarabine fludarabine Southern US 4357324 25 mg/m2/d phosphate. phosphate; Research IV over a 9H-Purin-6-2-F-araAMP; Institute period of amine, 2-Fludara; ; Berlex approx- fluoro-9- (5-0- Fludara iv; imately 30 phosphono-Fludara minutes beta-D-Oral; NSC-daily for arabinofuranos 312887; SH-c yl) 573 ; SH- secutive 586;SU- 'commencez every28 days. gemcitabi Eli Lily US 4526988 ne N- (4- ( ( (2, 4- methotrexat Hyal JS 2512572 tropho- diamino-6-e iv, Hyal; Pharma-blastic pteridinyl) met HA + ceutical; diseases: hyl) methylamin methotrexat American 15 to 30 o) benzoyl)-L- e, Hyal; Home mg/d glutamic acid methotrexat Products; orally or e iv, HIT Lederle intra- Technolog; muscularly in a five- day course (repeated 3 to 5 times as needed) Luteinizing nafarelin Roche EP 21234 hormone- releasing Comprnund common Ccmpany Reference Dosage Name/ Trade Name factor (pig), 6- [3- (2- naphthalenyl)- D-alanine]- pentostatin Warner-JS 3923785 ; CI-825; Lambert DCF; deoxycoform ycin; Nipent; NSC-218321; Oncopent; Ethanamine, 2-toremifene; Orion EP 95875 60 mg/d [4- (4-chloro- FARESTONO Pharma 1,2-diphenyl- 1- butenyl) phenox y]-N, N- dimethyl-, (Z)- A second family of antineoplastic agents which may be used in combination with the present invention consists of alkylating-type antineoplastic agents. The alkylating agents are believed to act by alkylating and cross-linking guanine and possibly other bases in DNA, arresting cell division. Typical alkylating agents include nitrogen mustards, ethyleneimine compounds, alkyl sulfates, cisplatin, and various nitrosoureas. A disadvantage with these compounds is that they not only attack malignant cells, but also other cells which are naturally dividing, such as those of bone marrow, skin, gastro-intestinal mucosa, and fetal tissue. Suitable alkylating-type antineoplastic agents that may be used in the present invention include, but are not limited to, Shionogi 254-S, aldo-phosphamide analogues, altretamine, anaxirone, Boehringer Mannheim BBR-2207,

bestrabucil, budotitane, Wakunaga CA-102, carboplatin, carmustine (BiCNU), Chinoin-139, Chinoin-153, chlorambucil, cisplatin, cyclophosphamide, American Cyanamid CL-286558, Sanofi CY-233, cyplatate, dacarbazine, Degussa D-19-384, Sumimoto DACHP (Myr) 2, diphenylspiromustine, diplatinum cytostatic, Erba distamycin derivatives, Chugai DWA-2114R, ITI E09, elmustine, Erbamont FCE-24517, estramustine phosphate sodium, etoposide phosphate, fotemustine, Unimed G-6-M, Chinoin GYKI-17230, hepsul-fam, ifosfamide, iproplatin, lomustine, mafosfamide, mitolactol, mycophenolate, Nippon Kayaku NK-121, NCI NSC-264395, NCI NSC-342215, oxaliplatin, Upjohn PCNU, prednimustine, Proter PTT-119, ranimustine, semustine, SmithKline SK&F-101772, thiotepa, Yakult Honsha SN-22, spiromus-tine, Tanabe Seiyaku TA-077, tauromustine, temozolomide, teroxirone, tetraplatin and trimelamol.

Preferred alkylating agents that may be used in the present invention include, but are not limited to, those identified in Table No. 6, below.

Table No. 6. Alkylating agents Compound Common Company Reference Dosage Name/Trade Name Platinum, carboplatin; Johnson US 4657927.360 mg/m ( diammine [1, l PARAPLATIN @ Matthey US 4140707. squared) -cyclobu-I. V. on tanedicarbox day 1 ylato (2-)]-, every 4 (SP-4-2)-weeks. Carmustine, BiCNUS Ben Venue JAMA 1985; Preferred: 1,3-bis (2-Labora-253 (11): 150 to 200 chloroethyl) tories, 1590-1592. mcgm every 6 sourea wks. USetoposideBristol- 4564675 Compound Cammon Company Reference Dosage Name/Trade Name phosphate Myers Squibb thiotepa Platinum, cisplatin; Bristol-US 4177263 diamminedi-PLATINOL-AQ Myers chloro-, Squibb (SP-4-2)- dacarbazine DTIC Dome Bayer 2 to 4.5mg/kg/d ay for 10 days; 250mg/ square meter body surface/ day I. V. for 5 days every 3 weeks ifosfamide IFEX Bristol-4-5 g/m Meyers (square) Squibb single bolus dose, or 1.2-2 g/m (square) I. V. over 5 days. cyclophosph US 4537883 amide cis-Platinol Bristol- 20 mg/M2 diaminedichl Cisplatin Myers IV daily oroplatinum Squibb for a 5 day cycle.

A third family of antineoplastic agents which may be used in combination with the present invention consists of antibiotic-type antineoplastic agents.

Suitable antibiotic-type antineoplastic agents that may be used in the present invention include, but are not limited to Taiho 4181-A, aclarubicin, actinomycin D,

actinoplanone, Erbamont ADR-456, aeroplysinin derivative, Ajinomoto AN-201-II, Ajinomoto AN-3, Nippon Soda anisomycins, anthracycline, azino-mycin-A, bisucaberin, Bristol-Myers BL-6859, Bristol-Myers BMY- 25067, Bristol-Myers BMY-25551, Bristol-Myers BMY-26605, Bristol-Myers BMY-27557, Bristol-Myers BMY-28438, bleomycin sulfate, bryostatin-1, Taiho C-1027, calichemycin, chromoximycin, dactinomycin, daunorubicin, Kyowa Hakko DC-102, Kyowa Hakko DC-79, Kyowa Hakko DC- 88A, Kyowa Hakko DC89-A1, Kyowa Hakko DC92-B, ditrisarubicin B, Shionogi DOB-41, doxorubicin, doxorubicin-fibrinogen, elsamicin-A, epirubicin, erbstatin, esorubicin, esperamicin-Al, esperamicin-Alb, Erbamont FCE-21954, Fujisawa FK-973, fostriecin, Fujisawa FR-900482, glidobactin, gregatin-A, grincamycin, herbimycin, idarubicin, illudins, kazusamycin, kesarirhodins, Kyowa Hakko KM-5539, Kirin Brewery KRN-8602, Kyowa Hakko KT-5432, Kyowa Hakko KT- 5594, Kyowa Hakko KT-6149, American Cyanamid LL-D49194, Meiji Seika ME 2303, menogaril, mitomycin, mitoxantrone, SmithKline M-TAG, neoenactin, Nippon Kayaku NK-313, Nippon Kayaku NKT-01, SRI International NSC-357704, oxalysine, oxaunomycin, peplomycin, pilatin, pirarubicin, porothramycin, pyrindamycin A, Tobishi RA- I, rapamycin, rhizoxin, rodorubicin, sibanomicin, siwenmycin, Sumitomo SM-5887, Snow Brand SN-706, Snow Brand SN-07, sorangicin-A, sparsomycin, SS Pharmaceutical SS-21020, SS Pharmaceutical SS-7313B, SS Pharmaceutical SS-9816B, steffimycin B, Taiho 4181-2, talisomycin, Takeda TAN-868A, terpentecin, thrazine, tricrozarin A, Upjohn U-73975, Kyowa Hakko UCN-10028A, Fujisawa WF-3405, Yoshitomi Y-25024 and zorubicin.

Preferred antibiotic anticancer agents that may be used in the present invention include, but are not limited to, those agents identified in Table No. 7, below.

Table No. 7. Antibiotic anticancer agents Compound Commun Name/Company Reference Dosage Trade Name 4-Hexenoic mycopheno-Roche WO 91/19498 1 to 3 acid, 6- (1, 3- late mofetil gm/d dihydro-4- hydroxy-6- methoxy-7- methyl-3-oxo-5- isobenzofuranyl )-4-methyl-, 2- (4- morpholinyl) eth yl ester, (E)- mitoxan-US 4310666 trone doxorubicin US 3590028 Mitomycin Mutamycin Bristol-After and/or Myers full mitomycin-C Squibb hemato- Oncology/logical Immun-recovery ology from any previous chemo- therapy: 2 20 mg/m intra- venously as a single dose via a function -ing intra- venous catheter

A fourth family of antineoplastic agents which may be used in combination with the present invention consists of synthetic nucleosides. Several synthetic nucleosides have been identified that exhibit anticancer activity. A well known nucleoside derivative with strong anticancer activity is 5-fluorouracil (5-FU). 5- Fluorouracil has been used clinically in the treatment of malignant tumors, including, for example, carcinomas, sarcomas, skin cancer, cancer of the digestive organs, and breast cancer. 5-Fluorouracil, however, causes serious adverse reactions such as nausea, alopecia, diarrhea, stomatitis, leukocytic thrombocytopenia, anorexia, pigmentation, and edema. Derivatives of 5- fluorouracil with anti-cancer activity have been described in U. S. Pat. No. 4,336,381. Further 5-FU derivatives have been described in the following patents listed in Table No. 8, hereby individually incorporated by reference herein.

Table No. 8.5-Fu derivatives JP 50-50383 JP 50-50384 JP 50-64281 JP JP 53-84981 U. S. Pat. No. 4,000,137 discloses that the peroxidate oxidation product of inosine, adenosine, or cytidine with methanol or ethanol has activity against lymphocytic leukemia. Cytosine arabinoside (also referred to as Cytarabin, araC, and Cytosar) is a nucleoside analog of deoxycytidine that was first synthesized in 1950 and introduced into clinical medicine in 1963. It is currently an important drug in the treatment of acute myeloid leukemia. It is also

active against acute lymphocytic leukemia, and to a lesser extent, is useful in chronic myelocytic leukemia and non-Hodgkin's lymphoma. The primary action of araC is inhibition of nuclear DNA synthesis. Handschumacher, R. and Cheng, Y.,"Purine and Pyrimidine Antimetabolites", Cancer Medicine, Chapter XV-1,3rd Edition, Edited by J. Holland, et al., Lea and Febigol, publishers.

5-Azacytidine is a cytidine analog that is primarily used in the treatment of acute myelocytic leukemia and myelodysplastic syndrome.

2-Fluoroadenosine-5'-phosphate (Fludara, also referred to as FaraA) is one of the most active agents in the treatment of chronic lymphocytic leukemia. The compound acts by inhibiting DNA synthesis. Treatment of cells with F-araA is associated with the accumulation of cells at the G1/S phase boundary and in S phase; thus, it is a cell cycle S phase-specific drug. InCorp of the active metabolite, F-araATP, retards DNA chain elongation. F-araA is also a potent inhibitor of ribonucleotide reductase, the key enzyme responsible for the formation of dATP. 2-Chlorodeoxyadenosine is useful in the treatment of low grade B-cell neoplasms such as chronic lymphocytic leukemia, non-Hodgkins'lymphoma, and hairy-cell leukemia. The spectrum of activity is similar to that of Fludara. The compound inhibits DNA synthesis in growing cells and inhibits DNA repair in resting cells.

A fifth family of antineoplastic agents which may be used in combination with the present invention consists of hormonal agents. Suitable hormonal-type antineoplastic agents that may be used in the present

invention include, but are not limited to Abarelix; Abbott A-84861; Abiraterone acetate; Aminoglutethimide; anastrozole; Asta Medica AN-207; Antide; Chugai AG-041R; Avorelin; aseranox; Sensus B2036-PEG; Bicalutamide; buserelin; BTG CB-7598; BTG CB-7630; Casodex; cetrolix; clastroban; clodronate disodium; Cosudex; Rotta Research CR-1505; cytadren; crinone; deslorelin; droloxifene; dutasteride; Elimina; Laval University EM-800; Laval University EM-652; epitiostanol; epristeride; Mediolanum EP-23904; EntreMed 2-ME; exemestane; fadrozole; finasteride; flutamide; formestane;. Pharmacia & Upjohn FCE-24304; ganirelix; goserelin; Shire gonadorelin agonist; Glaxo Wellcome GW-5638; Hoechst Marion Roussel Hoe-766; NCI hCG; idoxifene; isocordoin; Zeneca ICI- 182780; Zeneca ICI-118630; Tulane University J015X; Schering Ag J96; ketanserin; lanreotide; Milkhaus LDI-200; letrozol; leuprolide; leuprorelin; liarozole; lisuride hydrogen maleate; loxiglumide; mepitiostane; Leuprorelin; Ligand Pharmaceuticals LG-1127; LG-1447; LG-2293; LG-2527; LG- 2716; Bone Care International LR-103; Lilly LY-326315; Lilly LY-353381-HC1; Lilly LY-326391; Lilly LY-353381; Lilly LY-357489; miproxifene phosphate; Orion Pharma MPV-2213ad; Tulane University MZ-4-71; nafarelin; nilutamide; Snow Brand NKS01; octreotide; Azko Nobel ORG- 31710; Azko Nobel ORG-31806; orimeten; orimetene; orimetine; ormeloxifene; osaterone; Smithkline Beecham SKB-105657; Tokyo University OSW-1; Peptech PTL-03001; Pharmacia & Upjohn PNU-156765; quinagolide; ramorelix; Raloxifene; statin; sandostatin LAR; Shionogi S-10364; Novartis SMT- 487; somavert; somatostatin; tamoxifen; tamoxifen methiodide; teverelix; toremifene; triptorelin; TT-232; vapreotide; vorozole; Yamanouchi YM-116; Yamanouchi YM-

511; Yamanouchi YM-55208; Yamanouchi YM-53789; Schering AG ZK-1911703; Schering AG ZK-230211; and Zeneca ZD- 182780.

Preferred hormonal agents that may be used in the present invention include, but are not limited to, those identified in Table No. 9, below.

Table No. 9. Hormonal agents Compound ReferenceCompany Dosage Name/ Name 2-EntreMed; EntreMed methoxyestradiol 2-ME N- (S)- A-84861 Abbott tetrahydrofuroyl -Gly-D2Nal- D4ClPhe-D3Pal- Ser-NMeTyr- DLys (Nic)-Leu- Lys (Isp)-Pro- DAla-NH2 raloxi- fene [3R-1- (2, 2- AG-041R Chugai WO 94/19322 Dimethoxyethyl)- 3- ( (4- methylphenyl) ami nocarbonylmethyl )-3- (N'- (4-me thylphenyl) ureid o)-indoline-2- one] AN-207 Asta WO 97/19954 Medica Ethanamine, 2-toremif-Orion EP 95875 60 mg/d [4- (4-chloro- ene; Pharma 1,2-diphenyl-1-FARESTONX butenyl) phenoxy] -N, N-dimethyl-, (Z)- Ethanamine, 2-tamoxifen Zeneca US 4536516 For [4- (1, 2- NOLVADEX (patients diphenyl-1-R) with CompanyReferenceDosageCompoundCommon Name/ Trade Name butenyl) phenoxy] breast -N, N-dimethyl-, cancer, (Z)-the recommen ded daily dose is 20-40 mg. Dosages greater than 20 mg per day should be divided (morning and evening) D-Alaninamide N-Antide; Ares-WO 89/01944 25 or acetyl-3- (2- ORF-23541 Serono 50microg naphthalenyl)-D-/kg sc alanyl-4-chloro- D-phenylalanyl- 3- (3- pyridinyl)-D- alanyl-L-seryl- N6- (3- pyridinylcarbony l)-L-lysyl-N6- (3-pyridinylca rbonyl)-D-lysyl- L-leucyl-N6- (l- methylethyl)-L- lysyl-L-prolyl- B2036-Sensus PEG; Somaver; Trovert 4-Methyl-2- [4- EM-800; Laval [2- (1-EM-652 Universi Compound Common Company Reference Dosage Name/ Name piperidinyl) etho ty xy] phenyl]-7- (pivaloyloxy)-3- [4- (pivaloylox y)phenyl]-2H-l- benzopyran letrozol US 4749346 goserelin US 4100274 3- [4- [1, 2- GW-5638 Glaxo Diphenyl-1(Z)- Wellcome butenyl]phenyl]- 2 (E)-propenoic acid Estra-1,3,5 (10)- ICI-Zeneca EP 34/6014 250mg/mt h diol, 7- [9- Faslodex; [ (4,4,5,5,5- ZD-182780 pentafluoro- pentyl) sulfinyl]- nonyl]-, (7alpha,17beta)- J015X Tulane Universi ty LG-1127; Ligand LG-1447 Pharmace uticals LG-2293 Ligand Pharmace uticals LG-2527; Ligand LG-2716 Pharmace uticals buser-Peptech elin, Peptech; des- lorelin, Peptech; PTL- 03001; trip- Compound Common Company Reference Dosage Name/ Trade Name torelin, Peptech LR-103 Bone Care Internat ional [2- (4- LY-326315 Lilly WO 9609039 Hydroxyphenyl)- 6- hydroxynaphthale n-l-yl][4- [2- (1- piperdinyl) ethox yjpheny l] methane hydrochloride LY-Lilly 353381- HC1 LY-326391 Lilly LY-353381 Lilly LY-357489 Lilly MPV-Orion EP 476944 0.3-300 2213ad Pharma mg Isobutyryl-Tyr-MZ-4-71 Tulane D-Arg-Asp-Ala-Universi Ile- (4-Cl)-Phe- ty Thr-Asn-Ser-Tyr- Arg-Lys-Val-Leu- (2- aminobutyryl)- Gln-Leu-Ser-Ala- Arg-Lys-Leu-Leu- Gln-Asp-Ile-Nle- Ser 4- guanidinobu tylamide Androst-4-ene-NKS01; Snow EP 300062 3,6,17-trione,14alpha-Brand 14-hydroxy-OHAT; 140HAT 3beta, 16beta, 17a OSW-1 lpha- Compound Commun Company Reference Dosage Name/ Trade trihydroxycholes t-5-en-22-one- 16-0- (2-0-4- methoxybenzoyl- beta-D-xy lopyranosyl)- (I- 3) (2-0-acetyl- alpha-L- arabinopyranosid e) Spiro [estra-4,9- Org-Akzo EP 289073 diene-31710; Nobel 17,2' (3'H)- Org-31806 furan]-3-one, 11- [4- (dimethylamino) p henyl]-4',5'- dihydro-6- methyl-, (6beta, llbeta, 17 beta)- Pharmaci(22RS)-N-(1,1,1-PNU- trifluoro-2-156765; a & phenylprop-2-FCE-28260Upjohn yl)-3-oxo-4-aza- 5alpha-androst- l-ene-17beta- carboxamide 1-[(benzofuran-Menarini 2yl)-4- chlorophenylmeth yl] imidazole Tryptamine Rhone-WO 96/35686 derivatives Poulenc Rorer Permanently Pharmos WO 95/26720 ionic derivatives of steroid hormones and their antagonists Compound Camas Company Reference Dosage Name/ Trade Novel Meiji WO 97/30040 tetrahydronaph Seika thofuranone derivatives SMT-487; Novartis 90Y- octreo- tide D-Phe-Cys-Tyr-D-TT-232 Trp-Lys-Cys-Thr- NH2 2-(lH-imidazol-YM-116 Yamanou- 4-ylmethyl)-9H-chi carbazole monohydrochlorid e monohydrate 4- [N- (4- YM-511 Yamanou- bromobenzyl)-N-chi (4- cyanophenyl) amin o]-4H-1,2,4- triazole 2-(lH-imidazol-YM-55208; Yamanou- 4-ylmethyl)-9H-YM-53789 chi carbazole monohydrochlorid e monohydrate ZK-Schering 1911703 AG ZK-230211 Schering AG abarelix Praecis Pharmace uticals Androsta-5,16- abira-BTG dien-3-ol, 17-terone (3-pyridinyl)-,acetate; acetate (ester), CB-7598; (3beta)-CB-7630 2,6- aminoglut Novartis US 3944671 Piperidinedione,ethimide; 3- (4-Ciba- Compound Conte Co » Wy Reference Dosage Name/ Trade Name aminophenyl)-3-16038; ethyl-Cytadren; Elimina; Orimeten; Orimet- ene; Orimetine 1,3- anastro-Zeneca EP 296749 lmg/day Benzenediacetoni zole; trile, alpha, alph Arimidex; a, alpha', alpha'-ICI- tetramethyl-5-D1033; (lH-1,2,4- ZD-1033 triazol-1-ylme thyl)- 5-Oxo-L-prolyl-avorelin; Medi-EP 23904 L-histidyl-L-Meterelin olanum tryptophyl-L- seryl-L-tyrosyl- 2-methyl-D- tryptophyl-L- leucyl-L- arginyl-N-ethyl- L-prolinamide Propanamide, N-bicalutam Zeneca EP 100172 [4-cyano-3-ide; (trifluoromethyl Casodex; )phenyl]-3- [ (4-Cosudex; fluorophenyl) ICI- sulfonyl]-2-176334 hydroxy-2- methyl-, (+/-)- Luteinizing busere-Hoechst GB 15/23623 200-600 hormone-lin; Hoe-Marion microg/d releasing factor 766; Roussel ay (pig),6-[O-Profact; (1,1- Receptal; dimethylethyl)-S-746766; D-serine]-9- (N- Suprecor; ethyl-L-Suprecur; prolinamide)-10-Supre- deglycinamide-fact; Suprefakt Compound Cammaa Company Reference Dosage Name/ Trade Name D-Alaninamide, cetro-Asta EP 29/9402 N-acetyl-3- (2- relix; Medica naphthalenyl)-D-SB-075; alanyl-4-chloro-SB-75 D-phenylalanyl- 3- (3-pyridinyl)- D-alanyl-L- seryl-L-tyrosyl- N5- (aminocarbonyl)- D-ol-L-leucyl-L- arginyl-L- prolyl- Phosphonic acid, clodro-Schering (dichloromethyle nate AG ne) bis-, disodium, disodiumsalt-Leiras; Bonefos; Clasto- ban; KCO- 692 Luteinizing deslore-Roberts US 4034082 hormone-lin; releasing factor gonado- (pig), 6-D-relin tryptophan-9- (N- analogue, ethyl-L-Roberts; prolinamide)-10-LHRH deglycinamide-analogue, Roberts; Somagard Phenol, 3- [1- [4- droloxi-Klinge EP 54168 [2-fene; FK- (dimethylamino) e 435; K- thoxy] phenyl]-2-060; K- phenyl-1-21060E; butenyl]-, (E)-RP 60850 [CA S] 4-Azaandrost-l-dutaster-Glaxo ene-17-ide; GG-Wellcome carboxamide, N-745; GI- (2,5- 198745 bis(trifluoromet Compound Commun Company Reference Dosage Name/ Name hyl) phenyl)-3- oxo-, ( 5alpha,17beta)- Androstan-17-ol, epitio-Shionogi US 3230215 2,3-epithio-, stanol; (2alpha, 3alpha, 5 10275-S; alpha,17beta)- epithioan drostan- ol; S- 10275; Thiobres- tin; Thiodrol Androsta-3,5- epriste-Smith-EP 289327 0.4- diene-3-ride; Kline 160mg/da carboxylic acid, ONO-9302; Beecham y 17-(((1,1-SK&F- dimethylethyl) am 105657; ino)carbonyl)-SKB- (17beta)-105657 estrone 3-O-estrone sulfamate 3-O- sulfamate 19-Norpregna-ethinyl Schering DE 1949095 1,3,5 (10)-trien- estradiol AG 20-yne-3,17- sulfon- diol, 3- (2- ate; J96; propanesulfonate Turister- ), (17alpha)-on Androsta-1,4- exemes-Pharmaci DE 3622841 5mg/kg a & dione, 6-FCE-24304 Upjohn methylene- Benzonitrile, 4-fadrozo-Novartis EP 165904 1 mg po bid tetrahydroimidaz Afema; o [1, 5-a] pyridin- Arensin; 5-yl)-, CGS- monohydrochlorid 16949; e CGS- 16949A; CGS- 20287; Compound Common Company Reference Dosage Name/ Trade Name fadrozole monohydro chloride 4-Azaandrost-l-finaster-Merck & EP 155096 5mg/day ene-17-ide; Co carboxamide,N-Andozac; (1,1-ChibroPro dimethylethyl)-scar; 3-oxo-, Finastid; (5alpha,MK-0906; MK-906; Procure; Prodel; Propecia; Proscar; Proskar; Protide; YM-152 Propanamide, 2-flutamide Schering US 4329364 methyl-N- [4- ; Plough nitro-3-Drogenil; (trifluoromethylEuflex; ) phenyl]- Eulexin; Eulexine; Flucinom; Flutamida ; Fugerel; NK-601; Odyne; Prostogen at; Sch- 13521 Androst-4-ene-formest-Novartis EP 346953 250 or 3,17-dione, 4-ane; 4-600mg/da hydroxy-HAD; 4-y po OHA; CGP- 32349; CRC- 82/01; Depot; Lentaron RocheEP[N-Ac-D-Nal,D-ganirel- 312052 Compound Common Company Reference Dosage Name/ Name pCl-Phe, D-Pal, D- ix; Org- hArg (Et) 2, hArg (E 37462; t) 2, D-Ala] GnRH- RS-26306 gonadore-Shire lin agonist, Shire Luteinizing goserel-Zeneca US 4100274 hormone-in; ICI- releasing factor 118630; (pig),Zoladex; (1, 1- zoladex dimethylethyl)-LA D-serine]-10- deglycinamide-, 2- (aminocarbonyl) h ydrazide hCG; Milkhaus gonadotro phin; LDI-200 human NIH chorionic gonadotro phin; hCG Pyrrolidine, 1-idoxifene BTG EP 260066 [2- [4- [1- (4-; CB- iodophenyl)-2-7386; CB- phenyl-1-7432; SB- butenyl] phenoxy] 223030 et hyl]-, (E)- isocord-Indena oin 2,4 (1H, 3H)- ketanse-Johnson EP 13612 Quinazolinedione rin; & , 3- [2- [4- (4- Aseranox; Johnson fluorobenzoyl)-Ketensin; 1-KJK-945; piperidinyl] ethy ketanse- 1]-rine; Perketan; R-41468; Compound Common. Company Reference Dosage Name/ Trade Name Serefrex; Serepr- ess; Sufrexal; Taseron L-Threoninamide, lanreot-Beaufour EP 215171 3- (2- ide;-Ipsen naphthalenyl)-D-Angiopept alanyl-L-in; BIM- cysteinyl-L-23014; tyrosyl-D-Dermopept tryptophyl-L-in; lysyl-L-valyl-L-Ipstyl; cysteinyl-, Somatul- cyclic (2-7)-ine; disulfide Somatul- ine LP Benzonitrile, letroz-Novartis EP 236940 2.5mg/da 4- ole; CGS-y triazol-1-20267; ylmethylene)bis- Femara Luteinizing leuprol-Atrix hormone-ide, releasing factor Atrigel; (pig), 6-D-leuprol- leucine-9- (N- ide, ethyl-L-Atrix prolinamid e)- 10- deglycinamide- Luteinizing leupror-Abbott US 4005063 3.75micr hormone-elin; og sc q releasing factor Abbott-28 days (pig), 6-D-43818; leucine-9- (N- Carcinil; ethyl-L-Enantone; prolinamide)-10-Leuplin; deglycinamide-Lucrin; Lupron; Lupron Depot; leuprol- ide, Compound ReferenceDosageCompany Name/ Trade Name Abbott; leuprol- ide, Takeda; leupror- elin, Takeda; Procren Depot; Procrin; Prostap; Prostap SR; TAP- 144-SR Luteinizing leupror-Alza hormone-elin, releasing factor DUROS; (pig), 6-D-leuprolid leucine-9- (N- e,DUROS; ethyl-L-leupror- prolinamid e)-elin 10- deglycinamide- 1H-liaro-Johnson EP 260744 300mg Benzimidazole, zole; & bid 5- [ (3- Liazal; Johnson chlorophenyl)-Liazol; IH-imidazol-l-liaro- ylmethyl]-zole fumarate; R-75251; R-85246; Ro-85264 Urea,N'-lisuride VUFB [ (8alpha)-9, 10- hydrogen didehydro-6-maleate; methylergolin-8-Cuvalit; yl]-N, N-diethyl-Dopergin; , (Z)-2-Dopergine butenedioate; Eunal; (1: 1) Lysenyl; Lysenyl Forte; Compound Common Company Reference Dosage Name/ Trade Name Revanil Pentanoic acid, loxiglumi Rotta WO 87/03869 4- [ (3, 4- de; CR-Research dichlorobenzoyl) 1505 amino]-5- [ (3- methoxypropyl) pentylamino]-5- oxo-, (+/-)- Androstane, 2,3- mepitiost Shionogi US 3567713 epithio-17- [ (l- ane; S- methoxycyclopent 10364; yl)oxy]-, Thioderon (2alpha, 3alpha, 5 alpha,17beta)- Phenol, 4- [1- [4- miproxife Taiho WO 87/07609 20mg/day [2-ne (dimethylamino) e phosphate thoxy] phenyl]-2-; DP-TAT- [4- (1- 59; TAT- methylethyl) 59 phenyl]-1- butenyl]-, dihydrogen phosphate (ester), (E)- Luteinizing nafarelin Roche EP 21/234 hormone- ; NAG, releasing factor Syntex; (pig), 6- [3- (2- Nasanyl; naphthalenyl)-D-RS-94991; alanine]-RS-94991- 298; Synarel; Synarela; Synrelina 2,4- nilutam-Hoechst US 4472382 Imidazolidinedio ide; Marion ne, Roussel dimethyl-3- [4- Niland- nitro-3-ron; (trifluoromethyl Notost- RU-)phenyl]-ran; 23908 Compound Commua Company Reference Dosage Name/ Trade Name obesity Lilly WO 96/24670 gene; diabetes gene; leptin L-Cysteinamide, octreot-Novartis EP 29/579 D-phenylalanyl-ide; L-cysteinyl-L-Longast- phenylalanyl-D-atina; tryptophyl-L-octreot- lysyl-L-ide threonyl-N- [2- pamoate; hydroxy-1-Sandost- (hydroxymethyl) p atin; ropyl]-, cyclic Sandostat (2-7)-in LAR; disulfide, [R-Sandost- (R*,R*)]- atina; Sandost- atine; SMS-201- 995 Pyrrolidine, 1-ormelox-Central DE 2329201 [2- (p- (7- ifene; Drug methoxy-2,2- 6720-Research dimethyl-3-CDRI; Inst. phenyl-4-Centron; chromanyl) Choie-7; phenoxy) ethyl]-, centchrom trans-an; Saheli 2-Oxapregna-4,6- osaterone Teikoku EP 193871 Hormone dione,17-Hipros; (acetyloxy)-6-TZP-4238 chloro- Pregn-4-ene-progester Columbia 3,20-dione one; Laborato Crinone ries Sulfamide, N, N- quinagol-Novartis EP 77754 diethyl-N'-ide; CV- (1,2,3,4,4a, 5,10 205-502; ,10a-octahydro-Nor- Compound Common Company Reference Dosage Name/ Name 6-hydroxy-1-prolac; propylbenzo [g] qu SDZ-205- inolin-3-yl)-, 502 (3alpha, 4aalpha, 10abeta)- (+/-)- L-Proline, 1-ramore-Hoechst EP 451791 (N2- (N- (N- (N- (N- lix; Hoe-Marion (N- (N- (N-acetyl- 013; Hoe-Roussel 3- (2- 013C; naphthalenyl)-D-Hoe-2013 alanyl)-4-chl oro-D- phenylalanyl)-D- tryptophyl)-L- seryl)-L- tyrosyl)-O- (6- deoxy-alpha-L- mannopyra nosyl)-D-seryl)- L-leucyl)-L- arginyl)-, 2- (aminocarbonyl) h ydrazide- somatosta Tulane tin Universi analogues ty Ethanamine, 2-tamoxi-Zeneca US 4536516 [4- (1, 2- fen; diphenyl-1-Ceadan; butenyl) phenoxy] ICI- -N, N-dimethyl-, 46474; (Z)-Kessar; Nolgen; Nolvadex; Tafoxen; Tamofen; Tamoplex; Tamoxas- ta ; Tamoxen; Tomaxen tamoxifen Pharmos methiod- Compound Common Company Reference Dosage Name/ Trade Name ide Ethanamine, 2-tamoxifen Douglas [4- (1,2- diphenyl-1- butenyl)phenoxy] -N, N-dimethyl-, (z)- D-Alaninamide, tevere-Asta N-acetyl-3- (2- lix; Medica naphthalenyl)-D-Antarelix alanyl-4-chloro- D-pheny lalanyl- 3- (3-pyridinyl)- D-alanyl-L- seryl-L-tyrosyl- N6- (aminocarbonyl)- D-lysyl-L- leucyl-N6- (l- methylethyl)-L- lysyl-L-prolyl- Ethanamine, 2-toremif-Orion EP 95875 60mg po [4- (4-chloro- ene; Pharma 1,2-diphenyl-l- Estrimex; butenyl) phenoxy] Fareston; -N, N-dimethyl-, FC-1157; (Z)-FC-1157a; NK-622 Luteinizing tripto-Debio-US 4010125 hormone-relin; pharm releasing factor ARVEKAP; (pig),6-D-AY-25650; tryptophan-BIM- 21003; EN-52104; Decap- eptyl; WY-42422 L-vapreot-Debio-EP 203031 500micro Tryptophanamide, ide; BMY-pharm g sc tid D-phenylalanyl-41606; L-cysteinyl-L-Octasta- tyrosyl-D-tin; RC- Compound Common Company Reference Dosage Name/ Name tryptophyl-L-160 lysyl-L-valyl- L-cysteinyl-, cyclic (2-7)- disulfide- 1H-vorozole; Johnson EP 293978 2.5mg/da Benzotriazole, R-76713; & y 6- [ (4- R-83842; Johnson chlorophenyl)-Rivizor IH-1,2,4- triazol-1- ylmethyl]-1- methyl- A sixth family of antineoplastic agents which may be used in combination with the present invention consists of a miscellaneous family of antineoplastic agents including, but not limited to alpha-carotene, alpha-difluoromethyl-arginine, acitretin, Biotec AD-5, Kyorin AHC-52, alstonine, amonafide, amphethinile, amsacrine, Angiostat, ankinomycin, anti-neoplaston A10, antineoplaston A2, antineoplaston A3, antineoplaston A5, antineoplaston AS2-1, Henkel APD, aphidicolin glycinate, asparaginase, Avarol, baccharin, batracylin, benfluron, benzotript, Ipsen-Beaufour BIM-23015, bisantrene, Bristo-Myers BMY-40481, Vestar boron-10, bromofosfamide, Wellcome BW-502, Wellcome BW-773, calcium carbonate, Calcet, Calci-Chew, Calci-Mix, Roxane calcium carbonate tablets, caracemide, carmethizole hydrochloride, Ajinomoto CDAF, chlorsulfaquinoxalone, Chemes CHX-2053, Chemex CHX-100, Warner-Lambert CI-921, Warner-Lambert CI-937, Warner-Lambert CI-941, Warner-Lambert CI-958, clanfenur, claviridenone, ICN compound 1259, ICN compound 4711, Contracan, Cell Pathways CP-461, Yakult

Honsha CPT-11, crisnatol, curaderm, cytochalasin B, cytarabine, cytocytin, Merz D-609, DABIS maleate, dacarbazine, datelliptinium, DFMO, didemnin-B, dihaematoporphyrin ether, dihydrolenperone, dinaline, distamycin, Toyo Pharmar DM-341, Toyo Pharmar DM-75, Daiichi Seiyaku DN-9693, docetaxel, Encore Pharmaceuticals E7869, elliprabin, elliptinium acetate, Tsumura EPMTC, ergotamine, etoposide, etretinate, EulexinS, Cell Pathways Exisulind@ (sulindac sulphone or CP-246), fenretinide, Merck Research Labs Finasteride, Florical, Fujisawa FR-57704, gallium nitrate, gemcitabine, genkwadaphnin, Gerimed, Chugai GLA-43, Glaxo GR-63178, grifolan NMF-5N, hexadecylphosphocholine, Green Cross HO-221, homoharringtonine, hydroxyurea, BTG ICRF-187, ilmofosine, irinotecan, isoglutamine, isotretinoin, Otsuka JI-36, Ramot K-477, ketoconazole, Otsuak K- 76COONa, Kureha Chemical K-AM, MECT Corp KI-8110, American Cyanamid L-623, leucovorin, levamisole, leukoregulin, lonidamine, Lundbeck LU-23-112, Lilly LY- 186641, Materna, NCI (US) MAP, marycin, Merrel Dow MDL- 27048, Medco MEDR-340, megestrol, merbarone, merocyanine derivatives, methylanilinoacridine, Molecular Genetics MGI-136, minactivin, mitonafide, mitoquidone, Monocal, mopidamol, motretinide, Zenyaku Kogyo MST-16, Mylanta, N- (retinoyl) amino acids, Nilandron; Nisshin Flour Milling N-021, N-acylated-dehydroalanines, nafazatrom, Taisho NCU-190, Nephro-Calci tablets, nocodazole derivative, Normosang, NCI NSC-145813, NCI NSC-361456, NCI NSC-604782, NCI NSC-95580, octreotide, Ono ONO-112, oquizanocine, Akzo Org-10172, paclitaxel, pancratistatin, pazelliptine, Warner-Lambert PD-111707,

Warner-Lambert PD-115934, Warner-Lambert PD-131141, Pierre Fabre PE-1001, ICRT peptide D, piroxantrone, polyhaematoporphyrin, polypreic acid, Efamol porphyrin, probimane, procarbazine, proglumide, Invitron protease nexin I, Tobishi RA-700, razoxane, retinoids, Encore Pharmaceuticals R-flurbiprofen, Sandostatin; Sapporo Breweries RBS, restrictin-P, retelliptine, retinoic acid, Rhone-Poulenc RP-49532, Rhone-Poulenc RP-56976, Scherring-Plough SC-57050, Scherring-Plough SC-57068, selenium (selenite and selenomethionine), SmithKline SK&F-104864, Sumitomo SM-108, Kuraray SMANCS, SeaPharm SP-10094, spatol, spirocyclopropane derivatives, spirogermanium, Unimed, SS Pharmaceutical SS-554, strypoldinone, Stypoldione, Suntory SUN 0237, Suntory SUN 2071, Sugen SU-101, Sugen SU-5416, Sugen SU-6668, sulindac, sulindac sulfone; superoxide dismutase, Toyama T-506, Toyama T-680, taxol, Teijin TEI-0303, teniposide, thaliblastine, Eastman Kodak TJB-29, tocotrienol, Topostin, Teijin TT-82, Kyowa Hakko UCN-01, Kyowa Hakko UCN-1028, ukrain, Eastman Kodak USB-006, vinblastine sulfate, vincristine, vindesine, vinestramide, vinorelbine, vintriptol, vinzolidine, withanolides, Yamanouchi YM-534, Zileuton, ursodeoxycholic acid, and Zanosar.

Preferred miscellaneous agents that may be used in the present invention include, but are not limited to, those identified in Table No. 10, below.

Table No. 10. Miscellaneous agents Caund ccc, cmce Dosage Name/ Trade Name Flutamide ; 2-EULEX) Schering 750 mg/d in methyl- N-(4- 3 8-hr Compound Common Compaqy Reference Dosage l Name/ Trade Name nitro-3-doses. (trifluoro- methyl) phenyl) propanamide Ketocon-US 4144346 azole leucovo-US 4148999 rin irinote-US 4604463 can levamis-GB 11/20406 ole megestrol US 4696949 paclita-US 5641803 xel Nilutamide Nilandron Hoechst A total 5,5-dimethyl Marion daily dose 3- (4-nitro 3-Roussel of 300 mg (trifluorometh for 30 days yl) phenyl) followed 2,4- thereafter imidazolidined by three ione tablets (50 mg each) once a day for a total daily dosage of 150 mg. Vinorel-EP 0010458 bine vinblas- tine vincris- tine Octreotide Sandosta-Sandoz s. c. or acetate L-tin Pharma-i. v. cysteinamide, ceuticals administrat D-ion phenylalanyl-Acromegaly: L-cysteinyl-L-50-300 phenylalanyl-mcgm tid. D-tryptophyl-Carcinoid L-lysyl-L-tumors: 100 Compound Common Company Reference Dosage Name/ Trade Name threonyl--600 NSAIDs- (2- mcgm/d hydroxy-1- (mean = 300 (hydroxymethyl mcgm/d) ) propyl)-, Vipomas: cyclic-200-300 disulfide; (R-mcgm in (R*, R*) first two acetate salt weeks of therapy Streptozocin Zanosar Pharmacia i. v. 1000 Streptozocin & Upjohn mg/M2 of 2-deoxy-2-body ( ( (methylnitro surface per samino) carbony week for weeks.1)amino)-two alpha (and beta)-D- glucopyranose) topotecan US 5004758 Selenium EP 804927 L-ACES@ J. R. selenomethioni Carlson ne Laborat- ories calcium carbonate sulindac ExisulandS US 5858694 sulfone ursodeoxycho US 5843929 lic acid Cell Pathways CP-461 Some additional preferred antineoplastic agents include those described in the individual patents listed in Table No. 11 below, and are hereby individually incorporated by reference.

Table No. 11. Antineoplastic agents

EP0882734EP00253738GB02/135425Ep0296749 WO 09/832762 EP 0236940 US 5338732 US 4418068 US 4692434 US 5464826 US 5061793 EP 0702961 EP 0702961 EP 0702962 EP 0095875 EP 0010458 EP 0321122 US 5041424 JP 60019790 WO 09/512606 US4,808614 US 4526988 CA 2128644 US 5455270 WO99/25344 WO 96/27014 US 5695966 DE 19547958 WO95/16693 WO 82/03395 US 5789000 US 5902610 EP189990 US 4500711 FR 24/74032 US 5925699 WO99/25344 US 4537883 US 4808614 US 5464826 US5366734 US 4767628 US 4100274 US 4584305 US4336381 JP 5050383 JP 5050384 JP 5064281 JP51146482 JP 5384981 US 5472949 US 5455270 US4140704 US 4537883 US 4814470 US 3590028 US4564675 US 4526988 US 4100274 US 4604463 US4144346 US 4749713 US 4148999 GB 11/20406 US4696949 US 4310666 US 5641803 US 4418068 US5,004758 EP 0095875 EP 0010458 US 4935437 US4,278689 US 4820738 US 4413141 US 5843917 US5,858694 US 4330559 US 5851537 US 4499072 US 5,217886 WO 98/25603 WO 98/14188 Table No. 12 provides illustrative examples of median dosages for selected cancer agents that may be used in combination with an antiangiogenic agent. It should be noted that specific dose regimen for the chemotherapeutic agents below depends upon dosing considerations based upon a variety of factors including the type of neoplasia; the stage of the neoplasm; the age, weight, sex, and medical condition of the patient; the route of administration; the renal and hepatic function of the patient; and the particular combination employed.

Table No. 12. Median dosages for selected cancer agents.

NAME OF CHEMOTHERAPEUTIC AGENT MEDIAN DOSAGE Asparaginase 10,000 units Bleomycin Sulfate 15 units Carboplatin 50-450 mg.

Carmustine 100 mg.

Cisplatin 10-50 mg.

Cladribine 10 mg.

Cyclophosphamide 100 mg.-2 gm.

(lyophilized) Cyclophosphamide (non-100 mg.-2 gm. lyophilized) Cytarabine (lyophilized 100 mg.-2 gm. powder) Dacarbazine 100 mg.-200 mg.

Dactinomycin 0.5 mg.

Daunorubicin 20 mg.

Diethylstilbestrol 250 mg.

Doxorubicin 10-150 mg.

Etidronate 300 mg.

Etoposide 100 mg.

Floxuridine 500 mg.

Fludarabine Phosphate 50 mg.

Fluorouracil 500 mg.-5 gm.

Goserelin 3.6 mg.

Granisetron Hydrochloride 1 mg.

Idarubicin 5-10 mg.

Ifosfamide 1-3 gm.

Leucovorin Calcium 50-350 mg.

Leuprolide 3.75-7.5 rng.

Mechlorethamine 10 mg.

Medroxyprogesterone 1 gm.

Melphalan 50 gm.

Methotrexate 20 mg.-l gm.

Mitomycin 5-40 mg.

Mitoxantrone 20-30 mg.

Ondansetron Hydrochloride 40 mg.

Paclitaxel 30 mg.

Pamidronate Disodium 30-90 mg.

Pegaspargase 750 units Plicamycin 2,500 mcgm.

Streptozocin 1 gm.

Thiotepa 15 mg. <BR> <BR> <BR> <BR> <P> Teniposide 50 mg.<BR> <BR> <BR> <BR> <BR> <BR> <P> Vinblastine 10 mg.

Vincristine 1-5 mg.

Aldesleukin 22 million units Epoetin Alfa 2,000-10,000 units Filgrastim 300-480 mcgm.

Immune Globulin 500 mg.-lO gm.

Interferon Alpha-2a 3-36 million units Interferon Alpha-2b 3-50 million units Levamisole 50 mg.

Octreotide 1,000-5,000 mcgm.

Sararamostim 250-500 mcam.

The anastrozole used in the therapeutic combinations of the present invention can be prepared in the manner set forth in U. S. Patent No. 4,935,437.

The capecitabine used in the therapeutic combinations of the present invention can be prepared in the manner set forth in U. S. Patent No. 5,472,949.

The carboplatin used in the therapeutic combinations of the present invention can be prepared in the manner set forth in U. S. Patent No. 5,455,270.

The Cisplatin used in the therapeutic combinations of the present invention can be prepared in the manner set forth in U. S. Patent No. 4,140,704.

The cyclophoshpamide used in the therapeutic combinations of the present invention can be prepared in the manner set forth in U. S. Patent No. 4,537,883.

The eflornithine (DFMO) used in the therapeutic combinations of the present invention can be prepared in the manner set forth in U. S. Patent No. 4,413,141.

The docetaxel used in the therapeutic combinations of the present invention can be prepared in the manner set forth in U. S. Patent No. 4,814,470.

The doxorubicin used in the therapeutic combinations of the present invention can be prepared in the manner set forth in U. S. Patent No. 3,590,028.

The etoposide used in the therapeutic combinations of the present invention can be prepared in the manner set forth in U. S. Patent No. 4,564,675.

The fluorouricil used in the therapeutic combinations of the present invention can be prepared in the manner set forth in U. S. Patent No. 4,336,381.

The gemcitabine used in the therapeutic combinations of the present invention can be prepared in the manner set forth in U. S. Patent No. 4,526,988.

The goserelin used in the therapeutic combinations of the present invention can be prepared in the manner set forth in U. S. Patent No. 4,100,274.

The irinotecan used in the therapeutic combinations of the present invention can be prepared in the manner set forth in U. S. Patent No. 4,604,463.

The ketoconazole used in the therapeutic combinations of the present invention can be prepared in the manner set forth in U. S. Patent No. 4,144,346.

The letrozole used in the therapeutic combinations of the present invention can be prepared in the manner set forth in U. S. Patent No. 4,749,713.

The leucovorin used in the therapeutic combinations of the present invention can be prepared in the manner set forth in U. S. Patent No. 4,148,999.

The levamisole used in the therapeutic combinations of the present invention can be prepared in the manner set forth in GB 11/20,406.

The megestrol used in the therapeutic combinations of the present invention can be prepared in the manner set forth in U. S. Patent No. 4,696,949.

The mitoxantrone used in the therapeutic combinations of the present invention can be prepared in the manner set forth in U. S. Patent No. 4,310,666.

The paclitaxel used in the therapeutic combinations of the present invention can be prepared in the manner set forth in U. S. Patent No. 5,641,803.

The Retinoic acid used in the therapeutic combinations of the present invention can be prepared in the manner set forth in U. S. Patent No. 4,843,096.

The tamoxifen used in the therapeutic combinations of the present invention can be prepared in the manner set forth in U. S. Patent No. 4,418,068.

The topotecan used in the therapeutic combinations of the present invention can be prepared in the manner set forth in U. S. Patent No. 5,004,758.

The toremifene used in the therapeutic combinations of the present invention can be prepared in the manner set forth in EP 00/095,875.

The vinorelbine used in the therapeutic combinations of the present invention can be prepared in the manner set forth in EP 00/010,458.

The sulindac sulfone used in the therapeutic combinations of the present invention can be prepared in the manner set forth in U. S. Patent No. 5,858,694.

The selenium (selenomethionine) used in the therapeutic combinations of the present invention can be prepared in the manner set forth in EP 08/04,927.

The ursodeoxycholic acid used in the therapeutic combinations of the present invention can be prepared in the manner set forth in WO 97/34,608. Ursodeoxycholic acid can also be prepared according to the manner set forth in EP 05/99,282. Finally, ursodeoxycholic acid can be prepared according to the manner set forth in U. S.

Patent No. 5,843,929.

Still more preferred antineoplastic agents include: anastrozole, calcium carbonate, capecitabine, carboplatin, cisplatin, Cell Pathways CP-461, cyclophosphamide, docetaxel, doxorubicin, etoposide, ExisulindO, fluorouracil (5-FU), fluoxymestrine, gemcitabine, goserelin, irinotecan, ketoconazole, letrozol, leucovorin, levamisole, megestrol,

mitoxantrone, paclitaxel, raloxifene, retinoic acid, tamoxifen, thiotepa, topotecan, toremifene, vinorelbine, vinblastine, vincristine, selenium (selenomethionine), ursodeoxycholic acid, sulindac sulfone and eflornithine (DFMO).

The phrase"taxane"includes a family of diterpene alkaloids all of which contain a particular eight (8) member"taxane"ring structure. Taxanes such as paclitaxel prevent the normal post division breakdown of microtubules which form to pull and separate the newly duplicated chromosome pairs to opposite poles of the cell prior to cell division. In cancer cells which are rapidly dividing, taxane therapy causes the microtubules to accumulate which ultimately prevents further division of the cancer cell. Taxane therapy also affects other cell processes dependant on microtubules such as cell motility, cell shape and intracellular transport. The major adverse side-effects associated with taxane therapy can be classified into cardiac effects, neurotoxicity, haematological toxicity, and hypersensitivity reactions. (See Exp. Opin. Thera.

Patents (1998) 8 (5), hereby incorporated by reference).

Specific adverse side-effects include neutropenia, alopecia, bradycardia, cardiac conduction defects, acute hypersensitivity reactions, neuropathy, mucositis, dermatitis, extravascular fluid accumulation, arthralgias, and myalgias. Various treatment regimens have been developed in an effort to minimize the side effects of taxane therapy, but adverse side-effects remain the limiting factor in taxane therapy.

Taxane derivatives have been found to be useful in treating refractory ovarian carcinoma, urothelial

cancer, breast carcinoma, melanoma, non-small-cell lung carcinoma, gastric, and colon carcinomas, squamous carcinoma of the head and neck, lymphoblastic, myeloblastic leukemia, and carcinoma of the esophagus.

Paclitaxel is typically administered in a 15-420 2 mg/m dose over a 6 to 24 hour infusion. For renal cell carcinoma, squamous carcinoma of head and neck, carcinoma of esophagus, small and non-small cell lung cancer, and breast cancer, paclitaxel is typically 2 administered as a 250 mg/m 24 hour infusion every 3 weeks. For refractory ovarian cancer paclitaxel is 2 typically dose escalated starting at 110 mg/m.

2 Docetaxel is typically administered in a 60-100 mg/M i. v. over 1 hour, every three weeks. It should be noted, however, that specific dose regimen depends upon dosing considerations based upon a variety of factors including the type of neoplasia; the stage of the neoplasm; the age, weight, sex, and medical condition of the patient; the route of administration; the renal and hepatic function of the patient; and the particular agents and combination employed.

In one embodiment, paclitaxel is used in the present invention in combination with a matrix metalloproteinase inhibitor, an integrin antagonist and with cisplatin, cyclophosphamide, or doxorubicin for the treatment of breast cancer. In another embodiment paciltaxel is used in combination with a matrix metalloproteinase inhibitor, an integrin antagonist, cisplatin or carboplatin, and ifosfamide for the treatment of ovarian cancer.

In another embodiment docetaxal is used in the present invention in combination with a matrix metalloproteinase inhibitor, an integrin antagonist and in combination with cisplatin, cyclophosphamide, or doxorubicin for the treatment of ovary and breast cancer and for patients with locally advanced or metastatic breast cancer who have progressed during anthracycline based therapy.

The following references listed in Table No. 13 below, hereby individually incorporated by reference herein, describe various taxanes and taxane derivatives suitable for use in the present invention, and processes for their manufacture.

Table No. 13. Taxanes and taxane derivatives EP683232EP639577EP627418EP EP 797988EP727492EP767786EP EP 5886026US5880131US5879929US US5869680US5871979US5854278US5871979 US5840748US5827831US5824701US5840930 US 5821263US5811292US5808113US US 5807888US5780653US5773461US US 5767282US5763628US5760252US US 5756776US5750737US5744592US US 5728850US5728725US5723634US US 5717115US5716981US5714513US US 5705508US5703247US5703117US US5693666US5688977US5684175US5700669 US 5679807US5677462US5675025US US 5654448US5654447US5646176US US 5637484US5635531US5631278US US 5622986US5618952US5616740US US 5614645US5614549USUS 5608102 US 5599820 US 5594157 US 5587489 US 5580899 US 5567614US5565478USUS 5560872 US 5547981US5539103USUS 5532363 US 5530020 US 5508447 US 5489601 US 5484809 US 5475011 US 5473055 US 5470866 US 5466834 US 5449790 US 5442065 US 5440056 US 5430160 US 5412116 US 5412092 US 5411984 US 5407816 US 5407674 US 5405972 US 5399726 US 5395850 US 5384399 US 5380916 US 5380751 US 5367086 US 5356928 US 5356927 US 5352806 US 5350866 US 5344775 US 5338872 US 5336785 US 5319112 US 5296506 US 5294737 US 5294637 US 5284865 US 5284864 US 5283253 US 5279949 US 5274137 US 5274124 US 5272171 US 5254703 US 5254580 US 5250683 US 5243045 US 5229526 US 5227400 US 5200534 US 5194635 US 5175,315 US 5136060 US 5015744 WO 98/38862 WO 95/24402 WO 93/21173 EP 681574 EP 681575 EP 568203 EP 642503 EP 667772 EP 668762 EP 679082 EP 681573 EP 688212 EP 690712 EP 690853 EP 710223 EP 534708 EP 534709 EP 605638 EP 669918 EP 855909 EP 605638 EP 428376 EP 428376 EP 534707 EP 605637 EP 679156 EP 689436 EP 690867 EP 605637 EP 690867 EP 687260 EP 690711 EP 400971 EP 690711 EP 400971 EP 690711 EP 884314 EP 568203 EP 534706 EP 428376 EP 534707 EP 400971 EP 669918 EP 605637 US 5015744 US 5175315 US 5243045 US 5283253 US 5250683 US 5254703 US 5274124 US 5284864 US 5284865 US 5350866 US 5227400 US 5229526 US 4876399 US 5136060 US 5336785 US 5710287 US 5714513 US 5717115 US 5721268 US 5723634 US 5728725 US 5728850 US 5739362 US 5760219 US 5760252 US 5384399 US 5399726 US 5405972 US 5430160 US 5466834 US 5489601 US 5539103US5574156USUS 5587489 US 5637732US5654447USUS 4942184 US 5157149US5202488USUS 5750736 US 5549830US5281727USUS 5019504 US 4857653 US 4924011 US 5733388 US 5696153 WO 93/06093 WO 93/06094 WO 94/10996 WO 9/10997 WO 94/11362 WO 94/15599 WO 94/15929 WO 94/17050 WO 94/17051 WO 94/17052 WO 94/20088 WO 94/20485 WO 94/21250 WO 94/21251 WO 94/21252 WO 94/21623 WO 94/21651 WO 95/03265 WO 97/09979 WO 97/42181 WO 99/09021WO93/06079USWO 5202448 US 4857653US4924011WOUS 97/15571 WO 5489589EP781778WOUS 96/11683 EP 747385US5422364WOEP 95/11020 EP 747372 WO 96/36622 US 5599820 WO 97/10234 WO 96/21658 WO 97/23472 US 5550261 WO 95/20582 WO 97/28156 WO 96/14309 WO 97/32587 WO 96/28435 WO 95/25728WO94/29288WOWO 96/00724 WO 694539WO95/24402WOEP 93/10121 WO 97/19086 WO 97/20835 WO 96/14745 WO 96/36335 I U. S. Patent No. 5,019,504 describes the isolation of paclitaxel and related alkaloids from culture grown Taxus brevifolia cells.

U. S. Patent No. 5,675,025 describes methods for synthesis of Taxol@, Taxol@ analogues and intermediates from baccatin III.

U. S. Patent No. 5,688,977 describes the synthesis of Docetaxel from 10-deacetyl baccatin III.

U. S. Patent No. 5,202,488 describes the conversion of partially purified taxane mixture to baccatin III.

U. S. Patent No. 5,869,680 describes the process of preparing taxane derivatives.

U. S. Patent No. 5,856,532 describes the process of the production of Taxol@.

U. S. Patent No. 5,750,737 describes the method for paclitaxel synthesis.

U. S. Patent No. 6,688,977 describes methods for docetaxel synthesis.

U. S. Patent No. 5,677,462 describes the process of preparing taxane derivatives.

U. S. Patent No. 5,594,157 describes the process of making Taxol@ derivatives.

Some preferred taxanes and taxane derivatives are described in the patents in Table No. 14 below, and are hereby individually incorporated by reference herein.

Table No. 14. Some preferred taxanes and taxane derivatives US 5015744 US 5136060 US 5175315 US 5200534 US 5194635 US 5227400 US 4924012 US 5641803 US 5059699 US 5157049 US 4942184 US 4960790 US 5202488 US 5675025 US 5688977 US 5750736 US 5684175 US 5019504 US 4814470 WO 95/01969

The phrase"retinoid"includes compounds which are natural and synthetic analogues of retinol (Vitamin A).

The retinoids bind to one or more retinoic acid receptors to initiate diverse processes such as reproduction, development, bone formation, cellular proliferation and differentiation, apoptosis, hematopoiesis, immune function and vision. Retinoids are required to maintain normal differentiation and proliferation of almost all cells and have been shown to reverse/suppress carcinogenesis in a variety of in vitro and in vivo experimental models of cancer, see (Moon et al., Ch. 14 Retinoids and cancer. In The Retinoids, Vol.

2. Academic Press, Inc. 1984). Also see Roberts et al.

Cellular biology and biochemistry of the retinoids. In The Retinoids, Vol. 2. Academic Press, Inc. 1984, hereby incorporated by reference), which also shows that vesanoid (tretinoid trans retinoic acid) is indicated for induction of remission in patients with acute promyelocytic leukemia (APL).

A synthetic description of retinoid compounds, hereby incorporated by reference, is described in: Dawson MI and Hobbs PD. The synthetic chemistry of retinoids: in The retinoids, 2 edition. MB Sporn, AB Roberts, and DS Goodman (eds). New York: Raven Press, 1994, pp 5-178.

Lingen et al. describe the use of retinoic acid and interferon alpha against head and neck squamous cell carcinoma (Lingen, MW et al., Retinoic acid and interferon alpha act synergistically as antiangiogenic and antitumor agents against human head and neck

squamous cell carcinoma. Cancer Research 58 (23) 5551- 5558 (1998), hereby incorporated by reference).

Iurlaro et al. describe the use of beta interferon and 13-cis retinoic acid to inhibit angiogenesis.

(Iurlaro, M et al., Beta interferon inhibits HIV-1 Tat- induced angiogenesis: synergism with 13-cis retinoic acid. European Journal of Cancer 34 (4) 570-576 (1998), hereby incorporated by reference).

Majewski et al. describe Vitamin D3 and retinoids in the inhibition of tumor cell-induced angiogenesis.

(Majewski, S et al., Vitamin D3 is a potent inhibitor of tumor cell-induced angiogenesis. J. Invest. Dermatology.

Symposium Proceedings, 1 (1), 97-101 (1996), hereby incorporated by reference.

Majewski et al. describe the role of retinoids and other factors in tumor angiogenesis. Majewski, S et al., Role of cytokines, retinoids and other factors in tumor angiogenesis. Central-European journal of Immunology 21 (4) 281-289 (1996), hereby incorporated by reference).

Bollag describes retinoids and alpha-interferon in the prevention and treatment of neoplastic disease.

(Bollag W. Retinoids and alpha-interferon in the prevention and treatment of preneoplastic and neoplastic diseases. Chemotherapie Journal, (Suppl) 5 (10) 55-64 (1996), hereby incorporated by reference.

Bigg, HF et al. describe all-trans retinoic acid with basic fibroblast growth factor and epidermal growth factor to stimulate tissue inhibitor of metalloproteinases from fibroblasts. (Bigg, HF et al., All-trans-retoic acid interacts synergystically with basic fibroblast growth factor and epidermal growth factor to stimulate the production of tissue inhibitor

of metalloproteinases from fibroblasts. Arch. Biochem.

Biophys. 319 (1) 74-83 (1995), hereby incorporated by reference).

Nonlimiting examples of retinoids that may be used in the present invention are identified in Table No. 15 below.

Table No. 15. Retinoids Compound Commun Company Reference Dosage Name/Trade Name CD-271 Adapaline EP 199636 Tretinoin Vesanoid Roche 45 trans Holdings mg/M/day retinoic as two acid evenly divided doses until complete remission 2,4,6,8- etretinate Roche US. 25-1.5 Nonatetraen isoetret-Holdings 4215215 mg/kg/day oic acid, in; Ro-10- 9- (4- 9359; Ro- methoxy-13-7652; 2,3,6- Tegison; trimethylph Tigason enyl)-3,7- dimethyl-, ethyl ester, (all-E)- Retinoic isotret-Roche US 4843096.5 to 2 acid, 13-inoin Holdings mg/kg/day cis-Accutane; Isotrex; Ro-4-3780; Roaccutan; Roaccutane Roche Ro-Roche 40-0655 Holdings Roche Ro-Roche 25-6760 Holdings Roche Ro-Roche 25-9022 Holdings Roche Ro-Roche 25-9716 Holdings Benzoic TAC-101 Taiho acid, 4-Pharmace [ [3, 5- utical bis (trimeth ylsilyl) ben zoyl] amino] Retinamide, fenretinid 50-400 N- (4- e 4-HPR; mg/kg/day hydroxyphen HPR; McN- yl)-R-1967 (2E, 4E, 6E)- LGD-1550 Ligand 20 7- (3, 5-Di- ALRT-1550; Pharma-microg/m2 tert-ALRT-550; ceuticas/day to butylphenyl LG-1550; 400 )-3-Allergan microg/m2 methylocta-USA/day 2,4,6- administe trienoic red as a acid single daily oral dose Molecular US Design 4885311 MDI-101 Molecular US Design 4677120 MDI-403 Benzoic bexarotene WO acid, 4- (1- LG-1064; 94/15901 (5,6,7,8- LG-1069; tetrahydro-LGD-1069; 3,5,5,8,8- Targretin; pentamethyl Targretin -2-Oral; naphthaleny Targretin 1) eth Topical enyl)-Gel Benzoic bexarotene R P acid, 4- (1-, soft gel Scherer (5,6,7,8- bexarotene tetrahydro-, Ligand; 3,5,8,8- bexaroten pentamethyl -2- naphthaleny 1)ethen yl)- (2E,4E)WO methyl-5-96/05165 [3- tetramethyl -5, 6, 7, 8- tetrahydro- naphthalen- 2-yl)- thiopen-2- yl]-penta- 2,4-dienoic acid SR-11262 Hoffmann F-La Roche Ltd BMS-181162 Bristol EP 476682 Myers Squibb N- (4- IIT Cancer hydroxyphen Research Research yl) retinami Institute 39, 1339- de 1346 (1979) AGN-193174 Allergan WO USA 96/33716

The following individual patent references listed in Table No. 16 below, hereby individually incorporated by reference, describe various retinoid and retinoid derivatives suitable for use in the present invention described herein, and processes for their manufacture.

Table No. 16. Retinoids US 4215215 US 4885311 US 4677120 US 4105681 US 5260059 US 4503035 US 5827836 US 3878202 US 4843096 WO 96/05165 WO 97/34869 WO 97/49704 EP 19/9636 WO 96/33716 WO 97/24116 WO 97/09297 | WO 98/36742 WO 97/25969 WO 96/11686 WO 94/15901 WO97/24116 CH 61/6134 DE 2854354 EP 579915 US 5547947 EP 552624 EP 728742 EP 331983 EP 476682

Some preferred retinoids include Accutane; Adapalene; Allergan AGN-193174; Allergan AGN-193676; Allergan AGN-193836; Allergan AGN-193109; Aronex AR-623; BMS-181162; Galderma CD-437; Eisai ER-34617; Etrinate; Fenretinide; Ligand LGD-1550; lexacalcitol; Maxia Pharmaceuticals MX-781; mofarotene; Molecular Design

MDI-101; Molecular Design MDI-301; Molecular Design MDI- 403; Motretinide; Eisai 4- (2- [5- (4-methyl-7- <BR> <BR> <BR> <BR> ethylbenzofuran-2-yl) pyrrolyl]) benzoic acid; Johnson & Johnson N- [4- [2-thyl-l- (lH-imidazol-l-yl) butyl] phenyl]- 2-benzothiazolamine; Soriatane; Roche SR-11262; Tocoretinate; Advanced Polymer Systems trans-retinoic acid; UAB Research Foundation UAB-8; Tazorac; TopiCare; Taiho TAC-101; and Vesanoid. cGMP phosphodiesterase inhibitors, including Sulindac sulfone (ExisulandO) and CP-461 for example, are apoptosis inducers and do not inhibit the cyclooxygenase pathways. cGMP phosphodiesterase inhibitors increase apoptosis in tumor cells without arresting the normal cycle of cell division or altering the cell's expression of the p53 gene.

Ornithine decarboxylase is a key enzyme in the polyamine synthesis pathway that is elevated in most tumors and premalignant lesions. Induction of cell growth and proliferation is associated with dramatic increases in ornithine decarboxylase activity and subsequent polyamine synthesis. Further, blocking the formation of polyamines slows or arrests growth in transformed cells. Consequently, polyamines are thought to play a role in tumor growth. Difluoromethylornithine (DFMO) is a potent inhibitor of ornithine decarboxylase that has been shown to inhibit carcinogen-induced cancer development in a variety of rodent models (Meyskens et al. Development of Difluoromethylornithine (DFMO) as a chemoprevention agent. Clin. Cancer Res. 1999 May, 5 (%): 945-951, hereby incorporated by reference, herein).

DFMO is also known as 2-difluoromethyl-2,5-

diaminopentanoic acid, or 2-difluoromethyl-2,5- diaminovaleric acid, or a- (difluoromethyl) ornithine; DFMO is marketed under the tradename Elfornithine@.

Therefore, the use of DFMO in combination with COX-2 inhibitors is contemplated to treat or prevent cancer, including but not limited to colon cancer or colonic polyps.

Populations with high levels of dietary calcium have been reported to be protected from colon cancer. In vivo, calcium carbonate has been shown to inhibit colon cancer via a mechanism of action independent from COX-2 inhibition. Further, calcium carbonate is well tolerated. A combination therapy consisting of calcium carbonate and a selective COX-2 inhibitor is contemplated to treat or prevent cancer, including but not limited to colon cancer or colonic polyps.

Several studies have focused attention on bile acids as a potential mediator of the dietary influence on colorectal cancer risk. Bile acids are important detergents for fat solubilization and digestion in the proximal intestine. Specific transprot processes in the apical domain of the terminal ileal enterocyte and basolateral domain of the hepatocyte account for the efficient conservation in the enterohepatic circulation.

Only a small fraction of bile acids enter the colon; however, perturbations of the cycling rate of bile acids by diet (e. g. fat) or surgery may increase the fecal bile load and perhaps account for the associated increased risk of colon cancer. (Hill MJ, Bile flow and colon cancer. 238 Mutation Review, 313 (1990).

Ursodeoxycholate (URSO), the hydrophilic 7-beta epimer of chenodeoxycholate, is non cytotoxic in a variety of

cell model systems including colonic epithelia. URSO is also virtually free of side effects. URSO, at doses of 15mg/kg/day used primarily in biliary cirrhosis trials were extremely well tolerated and without toxicity.

(Pourpon et al., A multicenter, controlled trial of ursodiol for the treatment of primary biliary cirrhosis.

324 New Engl. J. Med. 1548 (1991)). While the precise mechanism of URSO action is unknown, beneficial effects of URSO therapy are related to the enrichment of the hepatic bile acid pool with this hydrophilic bile acid.

It has thus been hypothesized that bile acids more hydrophilic than URSO will have even greater beneficial effects than URSO. For example, tauroursodeoxycholate (TURSO) the taurine conjugate of URSO. Non-steroidal anti-inflammatory drugs (NSAIDs) can inhibit the neoplastic transformation of colorectal epithelium. The likely mechanism to explain this chemopreventive effect is inhibition of prostaglandin synthesis. NSAIDs inhibit cyclooxygenase, the enzyme that converts arachidonic acid to prostaglandins and thromboxanes. However, the potential chemopreventive benefits of NSAIDs such as sulindac or mesalamine are tempered by their well known toxicities and moderately high risk of intolerance.

Abdominal pain, dispepsia, nausea, diarrhea, constipation, rash, dizziness, or headaches have been reported in up to 9% of patients. The elderly appear to be particularly vulnerable as the incidence of NSAID- induced gastroduodenal ulcer disease, including gastrointestinal bleeding, is higher in those over the age of 60; this is also the age group most likely to develop colon cancer, and therefore most likely to benefit from chemoprevention. The gastrointestinal side

effects associated with NSAID use result from the inhibition of cyclooxygenase-1, an enzyme responsible for maintenance of the gastric mucosa. Therefore, the use of COX-2 inhibitors in combination with URSO is contemplated to treat or prevent cancer, including but not limited to colon cancer or colonic polyps; it is contemplated that this treatment will result in lower gastrointestinal side effects than the combination of standard NSAIDs and URSO.

An additional class of antineoplastic agents that may be used in the present invention include nonsteroidal antiinflammatory drugs (NSAIDs). NSAIDs have been found to prevent the production of prostaglandins by inhibiting enzymes in the human arachidonic acid/prostaglandin pathway, including the enzyme cyclooxygenase (COX). However, for the purposes of the present invention the definition of an NSAID does not include the"cyclooxygenase-2 inhibitors"described herein. Thus the phrase"nonsteroidal antiinflammatory drug"or"NSAID"includes agents that specifically inhibit cyclooxygenase-1, without significant inhibition of cyclooxygenase-2; or inhibit cyclooxygenase-1 and cyclooxygenase-2 at substantially the same potency; or inhibit neither cyclooxygenase-1 or cyclooxygenase-2.

The potency and selectivity for the enzyme cyclooxygenase-1 and cyclooxygenase-2 can be determined by assays well known in the art, see for example, Cromlish and Kennedy, Biochemical Pharmacology, Vol. 52, pp 1777-1785,1996.

Examples of NSAIDs that can be used in the combinations of the present invention include sulindac, indomethacin, naproxen, diclofenac, tolectin,

fenoprofen, phenylbutazone, piroxicam, ibuprofen, ketophen, mefenamic acid, tolmetin, flufenamic acid, nimesulide, niflumic acid, piroxicam, tenoxicam, phenylbutazone, fenclofenac, flurbiprofen, ketoprofen, fenoprofen, acetaminophen, salicylate and aspirin.

The term"clinical tumor"includes neoplasms that are identifiable through clinical screening or diagnostic procedures including, but not limited to, palpation, biopsy, cell proliferation index, endoscopy, mammagraphy, digital mammography, ultrasonography, computed tomagraphy (CT), magnetic resonance imaging (MRI), positron emmission tomaagraphy (PET), radiography, radionuclide evaluation, CT-or MRI-guided aspiration cytology, and imaging-guided needle biopsy, among others. Such diagnostic techniques are well known to those skilled in the art and are described in Cancer Medicine 4th Edition, Volume One. J. F. Holland, R. C.

Bast, D. L. Morton, E. Frei III, D. W. Kufe, and R. R.

Weichselbaum (Editors). Williams & Wilkins, Baltimore (1997).

The term"tumor marker"or"tumor biomarker" encompasses a wide variety of molecules with divergent characteristics that appear in body fluids or tissue in association with a clinical tumor and also includes tumor-associated chromosomal changes. Tumor markers fall primarily into three categories: molecular or cellular markers, chromosomal markers, and serological or serum markers. Molecular and chromosomal markers complement standard parameters used to describe a tumor (i. e. histopathology, grade, tumor size) and are used primarily in refining disease diagnosis and prognosis after clinical manifestation. Serum markers can often

be measured many months before clinical tumor detection and are thus useful as an early diagnostic test, in patient monitoring, and in therapy evaluation.

Molecular Tumor Markers Molecular markers of cancer are products of cancer cells or molecular changes that take place in cells because of activation of cell division or inhibition of apoptosis. Expression of these markers can predict a cell's malignant potential. Because cellular markers are not secreted, tumor tissue samples are generally required for their detection. Non-limiting examples of molecular tumor markers that can be used in the present invention are listed in Table No. 1, below.

Table No. 1. Non-limiting Examples of Molecular Tumor Markers Tumor Marker | Breast p53 Breast, ErbB-2/Her-2 Ovarian Breast S phase and ploidy Breast pS2 Breast MDR2 Breast urokinase plasminogen activator Breast, myc family Colon, Lung Chromosomal Tumor Markers Somatic mutations and chromosomal aberrations have been associated with a variety of tumors. Since the

identification of the Philadelphia Chromosome by Nowel and Hungerford, a wide effort to identify tumor-specific chromosomal alterations has ensued. Chromosomal cancer markers, like cellular markers, are can be used in the diagnosis and prognosis of cancer. In addition to the diagnostic and prognostic implications of chromosomal alterations, it is hypothesized that germ-line mutations can be used to predict the likelihood that a particular person will develop a given type of tumor. Non-limiting examples of chromosomal tumor markers that can be used in the present invention are listed in Table No. 2, below.

Table No. 2. Non-limiting Examples of Chromosomal Tumor Markers Tumor Marker Breast lp36 loss Breast 6q24-27 loss Breast llq22-23 loss Breast llql3 amplification Breast TP53 mutation Colon Gain of chromosome 13 Colon Deletion of short arm of chromosome 1 Lung Loss of 3p Lung Loss of 13q Lung Loss of 17p Lung Loss of 9p Seroloaical Tumor Markers Serum markers including soluble antigens, enzymes and hormones comprise a third category of tumor markers.

Monitoring serum tumor marker concentrations during therapy provides an early indication of tumor recurrence and of therapy efficacy. Serum markers are advantageous for patient surveillance compared to chromosomal and cellular markers because serum samples are more easily obtainable than tissue samples, and because serum assays can be performed serially and more rapidly. Serum tumor markers can be used to determine appropriate therapeutic doses within individual patients. For example, the efficacy of a combination regimen consisting of chemotherapeutic and antiangiogenic agents can be measured by monitoring the relevant serum cancer marker levels. Moreover, an efficacious therapy dose can be achieved by modulating the therapeutic dose so as to keep the particular serum tumor marker concentration stable or within the reference range, which may vary depending upon the indication. The amount of therapy can then be modulated specifically for each patient so as to minimize side effects while still maintaining stable, reference range tumor marker levels. Table No.

3 provides non-limiting examples of serological tumor markers that can be used in the present invention.

Table No. 3. Non-limiting Examples of Serum Tumor Markers Cancer Type Marker Germ Cell Tumors a-fetoprotein (AFP) Germ Cell Tumors human chorinonic gonadotrophin (hCG) Germ Cell Tumors placental alkaline phosphatase(PLAP) Germ Cell Tumors lactate dehydrogenase (LDH) ProstateProstateprostate specific antigen (PSA) Breastcarcinoembryonic antigen (CEA) Breast MUC-1 antigen (CA15-3) BreastBreasttissue polypeptide antigen (TPA) BreastBreasttissue polypeptide specific antigen (TPS) BreastCYFRA 21.1 Breast soluble erb-B-2 Ovarian CA125 Ovarian OVX1 OvarianOvariancancer antigen CA72-4 Ovarian TPA Ovarian TPS Gastrointestinal CD44v6 Gastrointestinal CEA Gastrointestinal cancer antigen CA19-9 Gastrointestinal NCC-ST-439 antigen (Dukes C) Gastrointestinal cancer antigen CA242 Gastrointestinal soluble erb-B-2 Gastrointestinal cancer antigen CA195 Gastrointestinal TPA Gastrointestinal YKL-40 Gastrointestinal TPS Esophageal CYFRA 21-1 Esophageal TPA Esophageal TPS Esophageal cancer antigen CA19-9 Gastric Cancer CEA Gastric Cancer cancer antigen CA19-9 Gastric Cancer cancer antigen CA72-4 Lung neruon specific enolase (NSE) Lung CEA \Lung CYFRA 21-1 T Lung cancer antigen CA 125 Lung TPA Lung squamous cell carcinoma antigen (SCC) Pancreaticca19-9 Pancreatic ca50 Pancreatic cancer call9 Pancreaticcancer cal25 Pancreatic cancer CEA Pancreatic cancer Renal Cancer CD44v6 Renal Cancer E-cadherin Renal Cancer PCNA (proliferating cell nuclear antigen)

Examples Germ Cell Cancers Non-limiting examples of tumor markers useful in the present invention for the detection of germ cell cancers include, but are not limited to, a-fetoprotein (AFP), human chorionic gonadotrophin (hCG) and its beta

subunit (hCGb), lactate dehydrogenase (LDH), and placental alkaline phosphatase (PLAP).

AFP has an upper reference limit of approximately -10 kU/L after the first year of life and may be elevated in germ cell tumors, hepatocellular carcinoma and also in gastric, colon, biliary, pancreatic and lung cancers. AFP serum half life is approximately five days after orchidectomy. According to EGTM recommendations, AFP serum levels less than 1,000 kU/L correlate with a good prognosis, AFP levels between 1,000 and 10,000 kU/L, inclusive, correlate with intermediate prognosis, and AFP levels greater than 10,000 U/L correlate with a poor prognosis.

HCG is synthesized in the placenta and is also produced by malignant cells. Serum hCG concentrations may be increased in pancreatic adenocarcinomas, islet cell tumors, tumors of the small and large bowel, hepatoma, stomach, lung, ovaries, breast and kidney.

Because some tumors only hCGb, measurement of both hCG and hCGb is recommended. Normally, serum hCG in men and pre-menopausal women is as high as-5 U/L while post- menopausal women have levels up to-10 U/L. Serum half life of hCG ranges from 16-24 hours. According to the EGTM, hCG serum levels under 5000 U/L correlate with a good prognosis, levels between 5000 and 50000 U/L, inclusively correlate with an intermediate prognosis, and hCG serum levels greater than 50000 U/L correlate with a poor prognosis. Further, normal hCG half lives correlate with good prognosis while prolonged half lives correlate with poor prognosis.

LDH is an enzyme expressed in cardiac and skeletal muscle as well as in other organs. The LDH-1 isoenzyme

is most commonly found in testicular germ cell tumors but can also occur in a variety of benign conditions such as skeletal muscle disease and myocardial infarction. Total LDH is used to measure independent prognostic value in patients with advanced germ cell tumors. LDH levels less than 1.5 x the reference range are associated with a good prognosis, levels between 1.5 and 10 x the reference range, inclusive, are associated with an intermediate prognosis, and levels more than 10 x the reference range are associated with a poor prognosis.

PLAP is a enzyme of alkaline phosphatase normally expressed by placental syncytiotrophoblasts. Elevated serum concentrations of PLAP are found in seminomas, non-seminomatous tumors, and ovarian tumors, and may also provide a marker for testicular tumors. PLAP has a normal half life after surgical resection of between 0.6 and 2.8 days.

Prostate Cancer A nonlimiting example of a tumor marker useful in the present invention for the detection of prostate cancer is prostate specific antigen (PSA). PSA is a glycoprotein that is almost exclusively produced in the prostate. In human serum, uncomplexed f-PSA and a complex of f-PSA with al-anthichymotrypsin make up total PSA (t-PSA). T-PSA is useful in determining prognosis in patients that are not currently undergoing anti-androgen treatment. Rising t-PSA levels via serial measurement indicate the presence of residual disease.

Breast Cancer Non-limiting examples of serum tumor markers useful in the present invention for the detection of breast

cancer include, but is not limited to carcinoembryonic antigen (CEA) and MUC-1 (CA 15.3). Serum CEA and CA15.3 levels are elevated in patients with node involvement compared to patients without node involvement, and in patients with larger tumors compared to smaller tumors.

Normal range cutoff points (upper limit) are 5-10 mg/L for CEA and 35-60 u/ml for CA15.3. Additional specificity (99.3%) is gained by confirming serum levels with two serial increases of more than 15%.

Ovarian Cancer A non-limiting example of a tumor marker useful in the present invention for the detection of ovarian cancer is CA125. Normally, women have serum CA125 levels between 0-35 kU/L; 99% of post-menopausal women have levels below 20 kU/L. Serum concentration of CA125 after chemotherapy is a strong predictor of outcome as elevated CA125 levels are found in roughly 80% of all patients with epithelial ovarian cancer. Further, prolonged CA125 half-life or a less than 7-fold decrease during early treatment is also a predictor of poor disease prognosis.

Gastrointestinal Cancers A non-limiting example of a tumor marker useful in the present invention for the detection of colon cancer is carcinoembryonic antigen (CEA). CEA is a glycoprotein produced during embryonal and fetal development and has a high sensitivity for advanced carcinomas including those of the colon, breast, stomach and lung. High pre- or postoperative concentrations (>2.5 ng/ml) of CEA are associated with worse prognosis than are low concentrations. Further, some studies in the literature

report that slow rising CEA levels indicates local recurrence while rapidly increasing levels suggests hepatic metastasis.

Lung Cancer Examples of serum markers useful in the present invention to monitor lung cancer therapy include, but are not limited to, CEA, cytokeratin 19 fragments (CYFRA 21-1), and Neuron Specific Enolase (NSE).

NSE is a glycolytic isoenzyme of enolase produced in central and peripheral neurons and malignant tumors of neuroectodermal origin. At diagnosis, NSE concentrations greater than 25 ng/mL are suggestive of malignancy and lung cancer while concentrations greater than 100 ng/mL are suggestive of small cell lung cancer.

CYFRA 21-1 is a tumor marker test which uses two specific monoclonal antibodies against a cytokeratin 19 fragment. At diagnosis, CYFRA 21-1 concentrations greater than 10 ng/mL are suggestive of malignancy while concentrations greater than 30 ng/mL are suggestive of lung cancer.

Accordingly, dosing of the matrix metalloproteinase inhibitor, integrin antagonist and antineoplastic agent may be determined and adjusted based on measurement of tumor markers in body fluids or tissues, particularly based on tumor markers in serum. For example, a decrease in serum marker level relative to baseline serum marker prior to administration of the matrix metalloproteinase inhibitor, integrin antagonist and antineoplastic agent indicates a decrease in cancer- associated changes and provides a correlation with inhibition of the cancer. In one embodiment, therefore, the method of the present invention comprises

administering the matrix metalloproteinase inhibitor, integrin antagonist and antineoplastic agent at doses that in combination result in a decrease in one or more tumor markers, particularly a decrease in one or more serum tumor markers, in the mammal relative to baseline tumor marker levels.

Similarly, decreasing tumor marker concentrations or serum half lives after administration of the combination indicates a good prognosis, while tumor marker concentrations which decline slowly and do not reach the normal reference range predict residual tumor and poor prognosis. Further, during follow-up therapy, increases in tumor marker concentration predicts recurrent disease many months before clinical manifestation.

In addition to the above examples, Table No. 4, below, lists several references, hereby individually incorporated by reference herein, that describes tumor markers and their use in detecting and monitoring tumor growth and progression.

Table No. 4. Tumor marker references.

European Group on Tumor Markers Publications Committee. Consensus Recommendations. Anticancer Research 19: 2785-2820 (1999) Human Cytogenetic Cancer Markers. Sandra R. Wolman and Stewart Sell (eds.). Totowa, New Jersey: Humana Press.

1997 Cellular Markers of Cancer. Carleton Garrett and Stewart Sell (eds.). Totowa, New Jersey: Human Press.

1995 Also included in the combination of the invention are the isomeric forms, prodrugs and tautomers of the described compounds and the pharmaceutically-acceptable salts thereof. Illustrative pharmaceutically acceptable salts are prepared from formic, acetic, propionic, succinic, glycolic, gluconic, lactic, malic, tartaric, citric, ascorbic, glucuronic, maleic, fumaric, pyruvic, aspartic, glutamic, benzoic, anthranilic, mesylic, stearic, salicylic, p-hydroxybenzoic, phenylacetic, mandelic, embonic (pamoic), methanesulfonic, ethanesulfonic, benzenesulfonic, pantothenic, toluenesulfonic, 2-hydroxyethanesulfonic, sulfanilic, cyclohexylaminosulfonic, algenic, b-hydroxybutyric, galactaric and galacturonic acids.

Suitable pharmaceutically-acceptable base addition salts of compounds of the present invention include metallic ion salts and organic ion salts. More preferred metallic ion salts include, but are not limited to appropriate alkali metal (group Ia) salts, alkaline earth metal (group IIa) salts and other physiological

acceptable metal ions. Such salts can be made from the ions of aluminum, calcium, lithium, magnesium, potassium, sodium and zinc. Preferred organic salts can be made from tertiary amines and quaternary ammonium salts, including in part, trimethylamine, diethylamine, N, N'-dibenzylethylenediamine, chloroprocaine, choline, diethanolamine, ethylenediamine, meglumine (N- methylglucamine) and procaine. All of the above salts can be prepared by those skilled in the art by conventional means from the corresponding compound of the present invention.

Administration Reaimen Any effective treatment regimen can be utilized and readily determined and repeated as necessary to effect treatment. In clinical practice, the compositions containing a MMP inhibitor and an integrin antagonist alone or in combination with other therapeutic agents are administered in specific cycles until a response is obtained.

For patients who initially present without advanced or metastatic cancer, a MMP inhibitor and an integrin angagonist may be given in combination with another MMP inhibitor and/or an integrin angagonist, a COX-2 inhibitor or one or more anticancer agents as an immediate initial therapy prior to surgery, chemotherapy, or radiation therapy, and as a continuous post-treatment therapy in patients at risk for recurrence or metastasis (for example, in adenocarcinoma of the prostate, risk for metastasis is based upon high PSA, high Gleason's score, locally extensive disease,

and/or pathological evidence of tumor invasion in the surgical specimen). The goal in these patients is to inhibit the growth of potentially metastatic cells from the primary tumor during surgery or radiotherapy and inhibit the growth of tumor cells from undetectable residual primary tumor.

For patients who initially present with advanced or metastatic cancer, an integrin antagonist in combination with a MMP inhibitor and/or one or more anticancer agents of the present invention is used as a continuous supplement to, or possible replacement for hormonal ablation. The goal in these patients is to slow or prevent tumor cell growth from both the untreated primary tumor and from the existing metastatic lesions.

In addition, the invention may be particularly efficacious during post-surgical recovery, where the present compositions and methods may be particularly effective in lessening the chances of recurrence of a tumor engendered by shed cells that cannot be removed by surgical intervention.

Combinations with Other Treatments The combination of MMP inhibitors and integrin antagonists may be used in conjunction with other treatment modalities, including, but not limited to surgery and radiation, hormonal therapy, antiangiogenic therapy, chemotherapy, immunotherapy, and cryotherapy.

The present invention may be used in conjunction with any current or future therapy.

The following discussion highlights some agents in this respect, which are illustrative, not limitative. A wide variety of other effective agents also may be used.

Surgery and Radiation In general, surgery and radiation therapy are employed as potentially curative therapies for patients under 70 years of age who present with clinically localized disease and are expected to live at least 10 years.

For example, approximately 70% of newly diagnosed prostate cancer patients fall into this category.

Approximately 90% of these patients (65% of total patients) undergo surgery, while approximately 10% of these patients (7% of total patients) undergo radiation therapy. Histopathological examination of surgical specimens reveals that approximately 63% of patients undergoing surgery (40% of total patients) have locally extensive tumors or regional (lymph node) metastasis that was undetected at initial diagnosis. These patients are at a significantly greater risk of recurrence.

Approximately 40% of these patients will actually develop recurrence within five years after surgery.

Results after radiation are even less encouraging.

Approximately 80% of patients who have undergone radiation as their primary therapy have disease persistence or develop recurrence or metastasis within five years after treatment. Currently, most of these surgical and radiotherapy patients generally do not receive any immediate follow-up therapy. Rather, for example, they are monitored frequently for elevated

Prostate Specific Antigen ("PSA"), which is the primary indicator of recurrence or metastasis prostate cancer.

Thus, there is considerable opportunity to use the present invention in conjunction with surgical intervention.

Hormonal Therapy Hormonal ablation is the most effective palliative treatment for the 10% of patients presenting with metastatic prostate cancer at initial diagnosis.

Hormonal ablation by medication and/or orchiectomy is used to block hormones that support the further growth and metastasis of prostate cancer. With time, both the primary and metastatic tumors of virtually all of these patients become hormone-independent and resistant to therapy. Approximately 50% of patients presenting with metastatic disease die within three years after initial diagnosis, and 75% of such patients die within five years after diagnosis. Continuous supplementation with NAALADase inhibitor based drugs are used to prevent or reverse this potentially metastasis-permissive state.

Among hormones which may be used in combination with the present inventive compounds, diethylstilbestrol (DES), leuprolide, flutamide, cyproterone acetate, ketoconazole and amino glutethimide are preferred.

Immunotherapy The MMP inhibitors and integrin angagonists of the present invention may also be used in combination with monoclonal antibodies in treating cancer. For example monoclonal antibodies may be used in treating prostate

cancer. A specific example of such an antibody includes cell membrane-specific anti-prostate antibody.

Antiangiogenic Therapy The MMP inhibitors and integrin antagonists of the present invention may also be used in combianation with other MMP inhibitors and integrin antagonists or other antiangiogenic agents in treating cancer. Antiangiogenic agents include but are not limited to MMP inhibitors, integrin antagonists, COX-2 inhibitors, angiostatin, endostatin, thrombospondin-1, and interferon alpha.

Examples of preferred antiangiogenic agents include, but are not limited to vitaxin, marimastat, Bay-12-9566, AG- 3340, metastat, celecoxib, rofecoxib, JTE-522, EMD- 121974, and D-2163 (BMS-275291).

The present invention may also be used with immunotherapies based on polyclonal or monoclonal antibody-derived reagents, for instance. Monoclonal antibody-based reagents are most preferred in this regard. Such reagents are well known to persons of ordinary skill in the art. Radiolabelled monoclonal antibodies for cancer therapy, such as the recently approved use of monoclonal antibody conjugated with strontium-89, also are well known to persons of ordinary skill in the art.

Cryotherap Cryotherapy recently has been applied to the treatment of some cancers. Methods and compositions of the present invention also could be used in conjunction with an effective therapy of this type.

All of the various cell types of the body can be transformed into benign or malignant neoplasia or tumor cells and are contemplated as objects of the invention.

A"benign"tumor cell denotes the non-invasive and non- metastasized state of a neoplasm. In man the most frequent neoplasia site is lung, followed by colorectal, breast, prostate, bladder, pancreas, and then ovary.

Other prevalent types of cancer include leukemia, central nervous system cancers, including brain cancer, melanoma, lymphoma, erythroleukemia, uterine cancer, and head and neck cancer. Examples 1 through 9 are provided to illustrate contemplated therapeutic combinations, and are not intended to limit the scope of the invention.

Illustrations The following non-limiting illustrative examples describe various cancer diseases and therapeutic approaches that may be used in the present invention, and are for illustrative purposes only. Preferred integrin antagonists of the below non-limiting <BR> <BR> <BR> <BR> illustrations include Compound I16, Compound I17,<BR> <BR> <BR> <BR> <BR> <BR> Compound I18, Compound 119, Compound I24, Compound I25,<BR> <BR> <BR> <BR> <BR> <BR> Compound I27, Compound 134, Compound 135, and Compound I36. Preferred MMP inhibitors of the below non-limiting illustrations include Compound M1, Compound M2, Compound M3, Compound M4, Compound M5, and Compound M7.

Example1 Lune Cancer In many countries including Japan, Europe and America, the number of patients with lung cancer is fairly large and continues to increase year after year and is the most frequent cause of cancer death in both men and women. Although there are many potential causes for lung cancer, tobacco use, and particularly cigarette smoking, is the most important. Additionally, etiologic factors such as exposure to asbestos, especially in smokers, or radon are contributory factors. Also occupational hazards such as exposure to uranium have been identified as an important factor. Finally, genetic factors have also been identified as another factor that increase the risk of cancer.

Lung cancers can be histologically classified into non-small cell lung cancers (e. g. squamous cell carcinoma (epidermoid), adenocarcinoma, large cell carcinoma (large cell anaplastic), etc.) and small cell lung cancer (oat cell). Non-small cell lung cancer (NSCLC) has different biological properties and responses to chemotherapeutics from those of small cell lung cancer (SCLC). Thus, chemotherapeutic formulas and radiation therapy are different between these two types of lung cancer.

Non-Small Cell Lung Cancer Where the location of the non-small cell lung cancer tumor can be easily excised (stage I and II disease) surgery is the first line of therapy and offers

a relatively good chance for a cure. However, in more advanced disease (stage IIIa and greater), where the tumor has extended to tissue beyond the bronchopulmonary lymph nodes, surgery may not lead to complete excision of the tumor. In such cases, the patient's chance for a cure by surgery alone is greatly diminished. Where surgery will not provide complete removal of the NSCLC tumor, other types of therapies must be utilized.

Today radiation therapy is the standard treatment to control unresectable or inoperable NSCLC. Improved results have been seen when radiation therapy has been combined with chemotherapy, but gains have been modest and the search continues for improved methods of combining modalities.

Radiation therapy is based on the principle that high-dose radiation delivered to a target area will result in the death of reproductive cells in both tumor and normal tissues. The radiation dosage regimen is generally defined in terms of radiation absorbed dose (rad), time and fractionation, and must be carefully defined by the oncologist. The amount of radiation a patient receives will depend on various consideration but the two most important considerations are the location of the tumor in relation to other critical structures or organs of the body, and the extent to which the tumor has spread. A prefered course of treatment for a patient undergoing radiation therapy for NSCLC will be a treatment schedule over a 5 to 6 week period, with a total dose of 50 to 60 Gy administered to the patient in a single daily fraction of 1.8 to 2.0 Gy,

5 days a week. A Gy is an abbreviation for Gray and refers to 100 rad of dose.

However, as NSCLC is a systemic disease, and radiation therapy is a local modality, radiation therapy as a single line of therapy is unlikely to provide a cure for NSCLC, at least for those tumors that have metastasized distantly outside the zone of treatment.

Thus, the use of radiation therapy with other modality regimens have important beneficial effects for the treatment of NSCLC.

Generally, radiation therapy has been combined temporally with chemotherapy to improve the outcome of treatment. There are various terms to describe the temporal relationship of administering radiation therapy in combination with integin antagonists, MMP inhibitors and chemotherapy, and the following examples are the preferred treatment regimens and are provided for illustration only and are not intended to limit the use of other combinations."Sequential"therapy refers to the administration of chemotherapy and/or MMP inhibitors and/or integrin antagonists and/or radiation therapy separately in time in order to allow the separate administration of either chemotherapy and/or integrin antagonists and/or MMP inhibitors, and/or radiation therapy."Concomitant"therapy refers to the administration of chemotherapy and/or an integrin antagonists, and/or MMP inhibitors and/or radiation therapy on the same day. Finally,"alternating therapy" refers to the administration of radiation therapy on the days in which chemotherapy and/or an integrin antagonist

and/or a MMP inhibitor would not have been administered if it was given alone.

It is reported that advanced non-small cell lung cancers do not respond favorably to single-agent chemotherapy and useful therapies for advanced inoperable cancers have been limited. (Journal of Clinical Oncology, vol. 10, pp. 829-838 (1992)).

Japanese Patent Kokai 5-163293 refers to some specified antibiotics of 16-membered-ring macrolides as a drug delivery carrier capable of transporting anthoracycline-type anticancer drugs into the lungs for the treatment of lung cancers. However, the macrolide antibiotics specified herein are disclosed to be only a drug carrier, and there is no reference to the therapeutic use of macrolides against non-small cell lung cancers.

WO 93/18,652 refers to the effectiveness of the specified 16-membered-ring macrolides such as bafilomycin, etc. in treating non-small cell lung cancers, but they have not yet been clinically practicable.

Pharmacology, vol. 41, pp. 177-183 (1990) describes that a long-term use of erythromycin increases productions of interleukins 1,2 and 4, all of which contribute to host immune responses, but there is no reference to the effect of this drug on non-small cell lung cancers.

Teratogenesis, Carcinogenesis, and Mutagenesis, vol. 10, pp. 477-501 (1990) describes that some of antimicrobial drugs can be used as an anticancer agent,

but does not refer to their application to non-small cell lung cancers.

In addition, interleukins are known to have an antitumor effect, but have not been reported to be effective against non-small cell lung cancers.

Any 14-or 15-membered-ring macrolides have not been reported to be effective against non-small cell lung cancers.

However, several chemotherapeutic agents have been shown to be efficacious against NSCLC. Preferred chemotherapeutic agents that can be used in the present invention against NSCLC include etoposide, carboplatin, methotrexate, 5-Fluorouracil, epirubicin, doxorubicin, taxol, inhibitor of normal mitotic activity; and cyclophosphamide. Even more preferred chemotherapeutic agents active against NSCLC include cisplatin, ifosfamide, mitomycin C, epirubicin, vinblastine, and vindesine.

Other agents that are under investigation for use against NSCLC include: camptothecins, a topoisomerase 1 inhibitor; navelbine (vinorelbine), a microtubule assebly inhibitor; gemcitabine, a deoxycytidine analogue; fotemustine, a nitrosourea compound; and edatrexate, a antifol.

The overall and complete response rates for NSCLC has been shown to increase with use of combination chemotherapy as compared to single-agent treatment.

Haskel CM: Chest. 99: 1325,1991; Bakowski MT: Cancer Treat Rev 10: 159,1983; Joss RA: Cancer Treat Rev 11:205,1984.

A preferred therapy for the treatment of NSCLC is a combination of therapeutically effective amounts of one or more integrin antagonists and/or MMP inhibitors in combination with one or more of the following combinations of antineoplastic agents: 1) itosfamide, cisplatin, etoposide; 2) cyclophoshamide, doxorubicin, cisplatin; 3) isofamide, carboplatin, etoposide; 4) bleomycin, etoposide, cisplatin; 5) isofamide, mitomycin, cisplatin; 6) cisplatin, vinblastine; 7) cisplatin, vindesine; 8) mitomycin C, vinblastine, cisplatin; 9) mitomycin C, vindesine, cisplatin; 10) isofamide, etoposide; 11) etoposide, cisplatin; 12) isofamide, mitomycin C; 13) flurouracil, cisplatin, vinblastine; 14) carboplatin, etoposide; or radiation therapy.

Accordingly, apart from the conventional concept of anticancer therapy, there is a strong need for the development of therapies practicably effective for the treatment of non-small cell lung cancers.

Small Cell Lune Cancer Approximately 15 to 20 percent of all cases of lung cancer reported worldwide is small cell lung cancer (SCLC). Ihde DC: Cancer 54: 2722,1984. Currently, treatment of SCLC incorporates multi-modal therapy, including chemotherapy, radiation therapy and surgery.

Response rates of localized or disseminated SCLC remain high to systemic chemotherapy, however, persistence of the primary tumor and persistence of the tumor in the associated lymph nodes has led to the integration of several therapeutic modalities in the treatment of SCLC.

A preferred therapy for the treatment of lung cancer is a combination of therapeutically effective amounts of one or more integrin antagonists and/or MMP inhibitors in combination with one or more of the following antineoplastic agents: vincristine, cisplatin, carboplatin, cyclophosphamide, epirubicin (high dose), etoposide (VP-16) I. V., etoposide (VP-16) oral, isofamide, teniposide (VM-26), and doxorubicin. Other preferred single-agents chemotherapeutic agents that may be used in the present invention include BCNU (carmustine), vindesine, hexamethylmelamine (altretamine), methotrexate, nitrogen mustard, and CCNU (lomustine). Other chemotherapeutic agents under investigation that have shown activity againe SCLC include iroplatin, gemcitabine, lonidamine, and taxol.

Single-agent chemotherapeutic agents that have not shown activity against SCLC include mitoguazone, mitomycin C, aclarubicin, diaziquone, bisantrene, cytarabine, idarubicin, mitomxantrone, vinblastine, PCNU and esorubicin.

The poor results reported from single-agent chemotherapy has led to use of combination chemotherapy.

A preferred therapy for the treatment of NSCLC is a combination of therapeutically effective amounts of one or more integrin antagonists and/or MMP inhibitors in combination with one or more of the following combinations of antineoplastic agents: 1) etoposide (VP- <BR> <BR> <BR> <BR> <BR> 16), cisplatin; 2) cyclophosphamide, adrianmycin<BR> <BR> <BR> <BR> <BR> <BR> <BR> [ (doxorubicin), vincristine, etoposide (VP-16)]; 3) Cyclophosphamide, adrianmycin (doxorubicin), vincristine; 4) Etoposide (VP-16), ifosfamide, cisplatin; 5)

etoposide (VP-16), carboplatin; 6) cisplatin, vincristine (Oncovin), doxorubicin, etoposide.

Additionally, radiation therapy in conjunction with the preferred combinations of integrin antagonists and MMP inhibitors and/or systemic chemotherapy is contemplated to be effective at increasing the response rate for SCLC patients. The typical dosage regimen for radiation therapy ranges from 40 to 55 Gy, in 15 to 30 fractions, 3 to 7 times week. The tissue volume to be irradiated is determined by several factors and generally the hilum and subcarnial nodes, and bialteral mdiastinal nodes up to the thoraic inlet are treated, as well as the primary tumor up to 1.5 to 2.0 cm of the margins.

Example 2 Colorectal Cancer Survival from colorectal cancer depends on the stage and grade of the tumor, for example precursor adenomas to metastatic adenocarcinoma. Generally, colorectal cancer can be treated by surgically removing the tumor, but overall survival rates remain between 45 and 60 percent. Colonic excision morbidity rates are fairly low and is generally associated with the anastomosis and not the extent of the removal of the tumor and local tissue. In patients with a high risk of reoccurrence, however, chemotherapy has been incorporated into the treatment regimen in order to improve survival rates.

Tumor metastasis prior to surgery is generally believed to be the cause of surgical intervention failure and up to one year of chemotherapy is required to kill the non-excised tumor cells. As severe toxicity is associated with the chemotherapeutic agents, only patients at high risk of recurrence are placed on chemotherapy following surgery. Thus, the incorporation of an antiangiogenesis inhibitor into the management of colorectal cancer will play an important role in the treatment of colorectal cancer and lead to overall improved survival rates for patients diagnosed with colorectal cancer.

A preferred combination therapy for the treatment of colorectal cancer is surgery, followed by a regimen of one or more chemotherapeutic agents and/or integrin antagonists and/or MMP inhibitors cycled over a one year time period. A more preferred combination therapy for the treatment of colorectal cancer is a regimen of one or more integrin antagonists and/or MMP inhibitors, followed by surgical removal of the tumor from the colon or rectum and then followed be a regimen of one or more chemotherapeutic agents and one or more integrin antagonists and/or MMP inhibitors, cycled over a one year time period. An even more preferred therapy for the treatment of colon cancer is a combination of therapeutically effective amounts of one or more integrin antagonists and/or COX-2 inhibitors.

A more preferred therapy for the treatment of colon cancer is a combination of therapeutically effective amounts of one or more integrin antagonists and/or COX-2 inhibitors in combination with the following

antineoplastic agents: fluorouracil, and Levamisole.

Preferably, fluorouracil and Levamisole are used in combination.

Example 3 Breast Cancer Today, among women in the United States, breast cancer remains the most frequent diagnosed cancer. One in 8 women in the United States are at risk of developing breast cancer in their lifetime. Age, family history, diet, and genetic factors have been identified as risk factors for breast cancer. Breast cancer is the second leading cause of death among women.

Different chemotherapeutic agents are known in art for treating breast cancer. Cytoxic agents used for treating breast cancer include doxorubicin, cyclophosphamide, methotrexate, 5- fluorouracil, mitomycin C, mitoxantrone, taxol, and epirubicin. CANCER SURVEYS, Breast Cancer volume 18, Cold Spring Harbor Laboratory Press, 1993.

In the treatment of locally advanced noninflammatory breast cancer, integrin antagonists and/or MMP inhibitors can be used to treat the disease in combination with other integrin antagonists and/or MMP inhibitors, or in combination with surgery, radiation therapy, antiangiogenic therapy or with chemotherapeutic agents. Preferred combinations of chemotherapeutic agents, radiation therapy and surgery that can be used in combination with the present invention include, but are not limited to the following

combinations: 1) doxorubicin, vincristine, radical mastectomy; 2) doxorubicin, vincristine, radiation therapy; 3) cyclophosphamide, doxorubicin, 5- flourouracil, vincristine, prednisone, mastecomy; 4) cyclophosphamide, doxorubicin, 5-flourouracil, vincristine, prednisone, radiation therapy; 5) cyclophosphamide, doxorubicin, 5-flourouracil, premarin, tamoxifen, radiation therapy for pathologic complete response; 6) cyclophosphamide, doxorubicin, 5- flourouracil, premarin, tamoxifen, mastectomy, radiation therapy for pathologic partial response; 7) mastectomy, radiation therapy, levamisole; 8) mastectomy, radiation therapy; 9) mastectomy, vincristine, doxorubicin, cyclophosphamide, levamisole; 10) mastectomy, vincristine, doxorubicin, cyclophosphamide; 11) mastecomy, cyclophosphamide, doxorubicin, 5- fluorouracil, tamoxifen, halotestin, radiation therapy; 12) mastecomy, cyclophosphamide, doxorubicin, 5- fluorouracil, tamoxifen, halotestin.

In the treatment of locally advanced inflammatory breast cancer, integrin antagonists and/or MMP inhibitors can be used to treat the disease in combination with other integrin antagonists and/or MMP inhibitors, or in combination with surgery, radiation therapy, antiangiogenic therapy or with chemotherapeutic agents. Preferred combinations of chemotherapeutic agents, radiation therapy and surgery that can be used in combination with the present invention include, but or not limited to the following combinations: 1) cyclophosphamide, doxorubicin, 5-fluorouracil, radiation therapy; 2) cyclophosphamide, doxorubicin, 5-

fluorouracil, mastectomy, radiation therapy; 3) 5- flurouracil, doxorubicin, clyclophosphamide, vincristine, prednisone, mastectomy, radiation therapy; 4) 5-flurouracil, doxorubicin, clyclophosphamide, vincristine, mastectomy, radiation therapy; 5) cyclophosphamide, doxorubicin, 5-fluorouracil, vincristine, radiation therapy; 6) cyclophosphamide, doxorubicin, 5-fluorouracil, vincristine, mastectomy, radiation therapy; 7) doxorubicin, vincristine, methotrexate, radiation therapy, followed by vincristine, cyclophosphamide, 5-florouracil; 8) doxorubicin, vincristine, cyclophosphamide, methotrexate, 5-florouracil, radiation therapy, followed by vincristine, cyclophosphamide, 5-florouracil; 9) surgery, followed by cyclophosphamide, methotrexate, 5- fluorouracil, predinsone, tamoxifen, followed by radiation therapy, followed by cyclophosphamide, methotrexate, 5-fluorouracil, predinsone, tamoxifen, doxorubicin, vincristine, tamoxifen; 10) surgery, followed by cyclophosphamide, methotrexate, 5- fluorouracil, followed by radiation therapy, followed by cyclophosphamide, methotrexate, 5-fluorouracil, predinsone, tamoxifen, doxorubicin, vincristine, tamoxifen; 11) surgery, followed by cyclophosphamide, methotrexate, 5-fluorouracil, predinsone, tamoxifen, followed by radiation therapy, followed by cyclophosphamide, methotrexate, 5-fluorouracil, doxorubicin, vincristine, tamoxifen;; 12) surgery, followed by cyclophosphamide, methotrexate, 5- fluorouracil, followed by radiation therapy, followed by cyclophosphamide, methotrexate, 5-fluorouracil,

predinsone, tamoxifen, doxorubicin, vincristine; 13) surgery, followed by cyclophosphamide, methotrexate, 5- fluorouracil, predinsone, tamoxifen, followed by radiation therapy, followed by cyclophosphamide, methotrexate, 5-fluorouracil, predinsone, tamoxifen, doxorubicin, vincristine, tamoxifen; 14) surgery, followed by cyclophosphamide, methotrexate, 5- fluorouracil, followed by radiation therapy, followed by cyclophosphamide, methotrexate, 5-fluorouracil, predinsone, tamoxifen, doxorubicin, vincristine; 15) surgery, followed by cyclophosphamide, methotrexate, 5- fluorouracil, predinsone, tamoxifen, followed by radiation therapy, followed by cyclophosphamide, methotrexate, 5-fluorouracil, doxorubicin, vincristine; 16) 5-florouracil, doxorubicin, cyclophosphamide followed by mastectomy, followed by 5-florouracil, doxorubicin, cyclophosphamide, followed by radtiation therapy.

In the treatment of metastatic breast cancer, integrin antagonists and/or COX-2 inhibitors can be used to treat the disease in combination with other antiangiogenic agents, or in combination with surgery, radiation therapy or with chemotherapeutic agents.

Preferred combinations of chemotherapeutic agents that can be used in combination with the integrin antagonists and/or MMP inhibitors of the present invention include, but are not limited to the following combinations: 1) cyclosphosphamide, methotrexate, 5-fluorouracil; 2) cyclophosphamide, adriamycin, 5-fluorouracil; 3) cyclosphosphamide, methotrexate, 5-flurouracil, vincristine, prednisone; 4) adriamycin, vincristine; 5)

thiotepa, adriamycin, vinblastine; 6) mitomycin, vinblastine; 7) cisplatin, etoposide.

Example 4 Prostate Cancer Prostate cancer is now the leading form of cancer among men and the second most frequent cause of death from cancer in men. It is estimated that more than 165,000 new cases of prostate cancer were diagnosed in 1993, and more than 35,000 men died from prostate cancer in that year. Additionally, the incidence of prostate cancer has increased by 50% since 1981, and mortality from this disease has continued to increase. Previously, most men died of other illnesses or diseases before dying from their prostate cancer. We now face increasing morbidity from prostate cancer as men live longer and the disease has the opportunity to progress.

Current therapies for prostate cancer focus exclusively upon reducing levels of dihydrotestosterone to decrease or prevent growth of prostate cancer. In addition to the use of digital rectal examination and transrectal ultrasonography, prostate-specific antigen (PSA) concentration is frequently used in the diagnosis of prostate cancer.

A preferred therapy for the treatment of prostate cancer is a combination of therapeutically effective amounts of one or more integrin antagonists and/or MMP inhibitors.

U. S. Pat. No. 4,472,382 discloses treatment of benign

prostatic hyperplasia (BPH) with an antiandrogen and certain peptides which act as LH-RH agonists.

U. S. Pat. No. 4,596,797 discloses aromatase inhibitors as a method of prophylaxis and/or treatment of prostatic hyperplasia.

U. S. Pat. No. 4,760,053 describes a treatment of certain cancers which combines an LHRH agonist with an antiandrogen and/or an antiestrogen and/or at least one inhibitor of sex steroid biosynthesis.

U. S. Pat. No. 4,775,660 discloses a method of treating breast cancer with a combination therapy which may include surgical or chemical prevention of ovarian secretions and administering an antiandrogen and an antiestrogen.

U. S. Pat. No. 4,659,695 discloses a method of treatment of prostate cancer in susceptible male animals including humans whose testicular hormonal secretions are blocked by surgical or chemical means, e. g. by use of an LHRH agonist, which comprises administering an antiandrogen, e. g. flutamide, in association with at least one inhibitor of sex steroid biosynthesis, e. g. aminoglutethimide and/or ketoconazole.

Prostate Specific Antigen One well known prostate cancer marker is Prostate Specific Antigen (PSA). PSA is a protein produced by prostate cells and is frequently present at elevated levels in the blood of men who have prostate cancer. PSA has been shown to correlate with tumor burden, serve as an indicator of metastatic involvement, and provide a parameter for following the response to surgery,

irradiation, and androgen replacement therapy in prostate cancer patients. It should be noted that Prostate Specific Antigen (PSA) is a completely different protein from Prostate Specific Membrane Antigen (PSMA). The two proteins have different structures and functions and should not be confused because of their similar nomenclature.

Prostate Specific Membrane Antigen (PSMA) In 1993, the molecular cloning of a prostate- specific membrane antigen (PSMA) was reported as a potential prostate carcinoma marker and hypothesized to serve as a target for imaging and cytotoxic treatment modalities for prostate cancer. Antibodies against PSMA have been described and examined clinically for diagnosis and treatment of prostate cancer. In particular, Indium-111 labelled PSMA antibodies have been described and examined for diagnosis of prostate cancer and itrium-labelled PSMA antibodies have been described and examined for the treatment of prostate cancer.

Example 5 Bladder Cancer The classification of bladder cancer is divided into three main classes: 1) superficial disease, 2) muscle-invasive disease, and 3) metastatic disease.

Currently, transurethral resection (TUR), or segmental resection, account for first line therapy of superficial bladder cancer, i. e., disease confined to

the mucosa or the lamina propria. However, intravesical therapies are necessary, for example, for the treatment of high-grade tumors, carcinoma in situ, incomplete resections, recurrences, and multifocal papillary.

Recurrence rates range from up to 30 to 80 percent, depending on stage of cancer.

Therapies that are currently used as intravesical therapies include chemotherapy, immuontherapy, bacille Calmette-Guerin (BCG) and photodynamic therapy. The main objective of intravesical therapy is twofold: to prevent recurrence in high-risk patients and to treat disease that cannot by resected. The use of intravesical therapies must be balanced with its potentially toxic side effects. Additionally, BCG requires an unimpaired immune system to induce an antitumor effect. Chemotherapeutic agents that are known to be inactive against superficial bladder cancer include Cisplatin, actinomycin D, 5-fluorouracil, bleomycin, and cyclophosphamide methotrxate.

In the treatment of superficial bladder cancer, integrin antagonists and/or MMP inhibitors can be used to treat the disease in combination with other integrin antagonists and/or MMP inhibitors, or in combination with surgery (TUR), chemotherapy and intravesical therapies.

A preferred therapy for the treatment of superficial bladder cancer is a combination of therapeutically effective amounts of one or more integrin antagonists and/or MMP inhibitors in combination with: thiotepa (30 to 60 mg/day), mitomycin

C (20 to 60 mg/day), and doxorubicin (20 to 80 mg/day).

A preferred intravesicle immunotherapeutic agent that may be used in the present invention is BCG. A preferred daily dose ranges from 60 to 120 mg, depending on the strain of the live attenuated tuberculosis organism used.

A preferred photodynamic therapuetic agent that may be used with the present invention is Photofrin I, a photosensitizing agent, administered intravenously. It is taken up by the low-density lipoprotein receptors of the tumor cells and is activated by exposure to visible light. Additionally, neomydium YAG laser activation generates large amounts of cytotoxic free radicals and singlet oxygen.

In the treatment of muscle-invasive bladder cancer, integrin antagonists and/or MMP inhibitors can be used to treat the disease in combination with other integrin antagonists and/or MMP inhibitors, or in combination with surgery (TUR), intravesical chemotherapy, antiangiogenic therapy, radiation therapy, and radical cystectomy with pelvic lymph node dissection.

A preferred radiation dose for the treatment of bladder cancer is between 5,000 to 7,000 cGY in fractions of 180 to 200 cGY to the tumor. Additionally, 3,500 to 4,700 cGY total dose is administered to the normal bladder and pelvic contents in a four-field technique. Radiation therapy should be considered only if the patient is not a surgical candidate, but may be considered as preoperative therapy.

A preferred combination of surgery and chemotherapeutic agents that can be used in combination with the integrin antagonists and/or MMP inhibitors of the present invention is cystectomy in conjunction with five cycles of cisplatin (70 to 100 mg/m (square)); doxorubicin (50 to 60 mg/m (square); and cyclophosphamide (500 to 600 mg/m (square).

A more preferred therapy for the treatment of superficial bladder cancer is a combination of therapeutically effective amounts of one or more integrin antagonists and/or MMP inhibitors.

An even more preferred combination for the treatment of superficial bladder cancer is a combination of therapeutically effective amounts of one or more integrin antagonists and/or MMP inhibitors in combination with one or more of the following combinations of antineoplastic agents: 1) cisplatin, doxorubicin, cyclophosphamide; and 2) cisplatin, 5- fluorouracil. An even more preferred combination of chemotherapeutic agents that can be used in combination with radiation therapy and integrin antagonists and/or MMP inhibitors is a combination of cisplatin, methotrexate, vinblastine.

Currently no curative therapy exists for metastatic bladder cancer. The present invention contemplates an effective treatment of bladder cancer leading to improved tumor inhibition or regression, as compared to current therapies.

In the treatment of metastatic bladder cancer, integrin antagonists and/or MMP inhibitors can be used to treat the disease in combination with other integrin

antagonists and/or MMP inhibitors, or in combination with surgery, radiation therapy, antiangiogenic therapy or with chemotherapeutic agents.

A preferred therapy for the treatment of metastatic bladder cancer is a combination of therapeutically effective amounts of one or more integrin antagonists and/or COX-2 inhibitors.

A more preferred combination for the treatment of metastatic bladder caner is a combination of therapeutically effective amounts of one or more integrin antagonists and/or MMP inhibitors in combination with one or more of the following combinations of antineoplastic agents: 1) cisplatin and methotrexate; 2) doxorubicin, vinblastine, cyclophoshamide, and 5-fluorouracil; 3) vinblastine, doxorubicin, cisplatin, methotrexate; 4) vinblastine, cisplatin, methotrexate; 5) cyclophosphamide, doxorubicin, cisplatin; 6) 5-fluorouracil, cisplatin.

Example 6 Pancreas Cancer Approximately 2% of new cancer cases diagnoses in the United States is pancreatic cancer. Pancreatic cancer is generally classified into two clinical types: 1) adenocarcinoma (metastatic and non-metastatic), and 2) cystic neoplasms (serous cystadenomas, mucinous cystic neoplasms, papilary cystic neoplasms, acinar cell systadenocarcinoma, cystic choriocarcinoma, cystic teratomas, angiomatous neoplasms).

Preferred combinations of therapy for the treatment of non-metastatic adenocarcinoma that may be used in the present invention include the use of integrin antagonists and/or MMP inhibitors along with preoperative bilary tract decompression (patients presenting with obstructive jaundice); surgical resection, including standard resection, extended or radial resection and distal pancreatectomy (tumors of body and tail); adjuvant radiation; and chemotherapy.

For the treatment of metastatic adenocarcinoma, a preferred combination therapy consists of integrin antagonists and/or MMP inhibitors of the present invention in combination with continuous treatment of 5- fluorouracil, followed by weekly cisplatin therapy.

A more preferred combination therapy for the treatment of cystic neoplasms is the use of integrin antagonists and/or MMP inhibitors along with resection.

Example 7 Ovarv Cancer Celomic epithelial carcinoma accounts for approximately 90% of ovarian cancer cases. A preferred therapy for the treatment of ovary cancer is a combination of therapeutically effective amounts of one or more integrin antagonists and/or MMP inhibitors.

Preferred single agents that can be used in combination with integrin antagonists and/or MMP inhibitors include, but are not limited to: alkylating agents, ifosfamide, cisplatin, carboplatin, taxol, doxorubicin, 5-fluorouracil, methotrexate, mitomycin,

hexamethylmelamine, progestins, antiestrogens, prednimustine, dihydroxybusulfan, galactitol, interferon alpha, and interferon gama.

Preferred combinations for the treatment of celomic epithelial carcinoma is a combination of therapeutically effective amounts of one or more integrin antagonists and/or MMP inhibitors in combination with one or more of the following combinations of antineoplastic agents: 1) cisplatin, doxorubicin, cyclophosphamide; 2) hexamthylmelamine, cyclosphamide, doxorubicin, cisplatin; 3) cyclophosphamide, hexamehtylmelamine, 5- flurouracil, cisplatin; 4) melphalan, hexamethylmelamine, cyclophosphamide; 5) melphalan, doxorubicin, cyclophosphamide; 6) cyclophosphamide, cisplatin, carboplatin; 7) cyclophosphamide, doxorubicin, hexamethylmelamine, cisplatin; 8) cyclophosphamide, doxorubicin, hexamethylmelamine, carboplatin; 9) cyclophosphamide, cisplatin; 10) hexamethylmelamine, doxorubicin, carboplatin; 11) cyclophosphamide, hexamethlmelamine, doxorubicin, cisplatin; 12) carboplatin, cyclophosphamide; 13) cisplatin, cyclophosphamide.

Germ cell ovarian cancer accounts for approximately 5% of ovarian cancer cases. Germ cell ovarian carcinomas are classified into two main groups: 1) dysgerminoma, and nondysgerminoma. Nondysgerminoma is further classified into teratoma, endodermal sinus tumor, embryonal carcinoma, chloricarcinoma, polyembryoma, and mixed cell tumors.

A preferred therapy for the treatment of germ cell carcinoma is a combination of therapeutically effective

amounts of one or more integrin antagonists and/or MMP inhibitors.

A more preferred therapy for the treatment of germ cell carcinoma is a combination of therapeutically effective amounts of one or more integrin antagonists and/or MMP inhibitors in combination with one or more of the following combinations of antineoplastic agents: 1) vincristine, actinomycin D, cyclophosphamide; 2) bleomycin, etoposide, cisplatin; 3) vinblastine, bleomycin, cisplatin.

Cancer of the fallopian tube is the least common type of ovarian cancer, accounting for approximately 400 new cancer cases per year in the United States.

Papillary serous adenocarcinoma accounts for approximately 90% of all malignancies of the ovarian tube.

A preferred therapy for the treatment of fallopian tube cancer is a combination of therapeutically effective amounts of one or more integrin antagonists and/or MMP inhibitors.

A more preferred therapy for the treatment of fallopian tube cancer is a combination of therapeutically effective amounts of one or more integrin antagonists and/or MMP inhibitors in combination with the following of antineoplastic agents: alkylating agents, ifosfamide, cisplatin, carboplatin, taxol, doxorubicin, 5-fluorouracil, methotrexate, mitomycin, hexamethylmelamine, progestins, antiestrogens, prednimustine, dihydroxybusulfan, galactitol, interferon alpha, and interferon gama.

An even more preferred therapy for the treatment of fallopian tube cancer is a combination of therapeutically effective amounts of one or more integrin antagonists and/or MMP inhibitors in combination with one or more of the following combinations of antineoplastic agents: 1) cisplatin, doxorubicin, cyclophosphamide; 2) hexamthylmelamine, cyclosphamide, doxorubicin, cisplatin; 3) cyclophosphamide, hexamehtylmelamine, 5-flurouracil, cisplatin; 4) melphalan, hexamethylmelamine, cyclophosphamide; 5) melphalan, doxorubicin, cyclophosphamide; 6) cyclophosphamide, cisplatin, carboplatin; 7) cyclophosphamide, doxorubicin, hexamethylmelamine, cisplatin; 8) cyclophosphamide, doxorubicin, hexamethylmelamine, carboplatin; 9) cyclophosphamide, cisplatin; 10) hexamethylmelamine, doxorubicin, carboplatin; 11) cyclophosphamide, hexamethlmelamine, doxorubicin, cisplatin; 12) carboplatin, cyclophosphamide; 13) cisplatin, cyclophosphamide.

Example 8 Central Nervous Svstem Cancers Central nervous system cancer accounts for approximately 2% of new cancer cases in the United States. Common intracranial neoplasms include glioma, meninigioma, neurinoma, and adenoma.

A preferred therapy for the treatment of central nervous system cancers is a combination of

therapeutically effective amounts of one or more integrin antagonists and/or MMP inhibitors.

A preferred therapy for the treatment of maligant glioma is a combination of therapeutically effective amounts of one or more integrin antagonists and/or MMP inhibitors in combination with one or more of the following combinations of therapies and antineoplastic agents:: 1) radiation therapy, BCNU (carmustine); 2) radiation therapy, methyl CCNU (lomustine); 3) radiation therapy, medol; 4) radiation therapy, procarbazine; 5) radiation therapy, BCNU, medrol; 6) hyperfraction radiation therapy, BCNU; 7) radiation therapy, misonidazole, BCNU; 8) radiation therapy, streptozotocin; 9) radiation therapy, BCNU, procarbazine; 10) radiation therapy, BCNU, hydroxyurea, procarbazine, VM-26; 11) radiation therapy, BNCU, 5- flourouacil; 12) radiation therapy, Methyl CCNU, dacarbazine; 13) radiation therapy, misonidazole, BCNU; 14) diaziquone; 15) radiation therapy, PCNU; 16) procarbazine (matulane), CCNU, vincristine. A preferred dose of radiation therapy is about 5,500 to about 6,000 cGY. Preferred radiosensitizers include misonidazole, intra-arterial Budr and intravenous iododeoxyuridine (IUdR). It is also contemplated that radiosurgery may be used in combinations with antiangiogenesis agents.

Biological Evaluation MMP Inhibitors 1. Pancreatic Cell (PC-3) Model: In this study, the test groups were a vehicle control, Compound M14, Compound M14 with cisplatin and cisplatin alone with n=10 for each group. The tumors were measured with a caliper and the volume calculated using the formula for the volume of an elipsoid. The cisplatin dose was 10 mpk administered by the intraperitonal route on day 8 post injection of tumor cells Compound M14,50 mpk, was first administered about 6: 00 pm the evening of the same day that the tumor cells were injected in the morning. The same dose of Compound M14 was administered bid for each following day. Tumor volume (mm3) was measured on day 25. The data below clearly show an improved response with the combination of the MMP inhibitor and cisplatin.

PC3 Model MMP Inhibitor Combination Study Results Agent Administered Tumor Volume at Day 25 PC3 Model (mm3) vehicle 860 cisplatin 630 Compound M14 480 Compound M14 110 with cisplatin

2. Breast Tumor Model: This study was carried out essentially as PC-3 model. MX-1 breast tumor pieces were implanted (with a trocar) into nude mice with n=10 per group. Dosing with Compound M14 (10 mpk or 50 mpk, PO bid) was initiated when the tumors reached a size of 60-120 mg. Dosing was continued for 26 days. Taxol was administered at a dose of 9 mpk for the first five days following the start of dosing by the interperitonal route. The tumors were measured using a caliper and the volume calculated using the formula for the volume of an elipsoid. The results tabulated below clearly show an improved response with combination therapy. An improved response is obtained with lower doses Compound M14.

MX-1 Model MMP Inhibitor Combination Study Results Agent Administered Tumor Volume at Day 25 (mm3) vehicle 1920 taxol 1280 Compound M14 960 @ 10 mpk Compound M14 1260 @ 50 mpk Compound M14 @ 50 mpk + 480 taxol @ 9 mpk Compound M14 @ 10 mpk + 240 taxol @ 9 mpk

3. MX-1 Adjuvant Model: Mice were implanted with MX-1 tumors and allowed to grow to 50-100 mm3. The animals were dosed with cyclophosphamide (100 or 80 mpk). This was considered Day 1. Two weeks later the animals were pair matched after tumor regression and dosing BID with the MMP inhibitor was begun until the end of the experiment.

Tumors were measured weekly. The endpoint for the study was a final tumor size of 1.5 g. Dose MMP Dose MDS sem (mpk) inhibitor (mpk) saline 23.9 1.3 cyclophosfamide 100 39.5 1.2 cyclophosfamide 80 37.2 1.5 cyclophosfamide 100 Compound 200 52.7 2.9 M14 cyclophosfamide 100 Compound 50 43.7 1.6 M14 cyclophosfamide 80 Compound 200 53.9 2.9 M14 cyclophosfamide 80 Compound 50 44.2 1.8 M14 MDS = mean days to tumor weight of 1.5 g 4. MX-1 breast tumor with taxol: Mice were implanted with MX-1 tumors and allowed to grow to 50-100 mg. The animals were pair matched and this was considered Day 1. Treatment with MMP inhibitor was begun BID on Day 1 until the end of the experiment.

Taxol was injected IP (15 or 9 mpk) QD for 5 days (days 1-5). Tumors were measured weekly until an endpoint of 1.5 g was reached.

Taxol MMP MMP MDS sem | Dose inhibitor inhibitor (mpk) Dose (mpk) vehicle 25.3 0.8

mmpi Compound 100 32.2 2.8 M14 mmpi Compound 20 34.7 3 M14 taxol + mmpi 18 56 11 taxol + mmpi 9 30.1 1.8 taxol + mmpi 18 Compound 100 61 l M14 taxol + mmpi 9 Compound 100 46.7 3.7 M14 taxol + mmpi 18 Compound 20 59.3 7 M14 taxol + mmpi 9 Compound 20 39.3 1.9 M14 MDS = 1.5 g 5. SK-mes tumor with Taxol Mice were implanted with SK-mes tumors and allowed to grow to 50-100 mg. The animals were pair matched and this was considered Day 1. Treatment with MMP inhibitor was begun BID on Day 1 until the end of the experiment. Taxol was injected IP (18 or 9 mpk) QD for 5 days (days 1-5). Tumors were measured weekly until an endpoint of 1.0 g was reached. Taxol MMP MMP MDS sem Dose inhibitor inhibitor (mpk) Dose (mpk) 21.2 2.1 mmpi Compound 100 24.7 1. 6 M14 mmpi Compound 20 18 1.1 M14 taxol 18 31.5 2.4 taxol 9 26.1 2. 3 taxol + mmpi 18 Compound 100 43 4 M14 taxol + mmpi 9 Compound 100 34.8 1.9 M14 taxol + mmpi 18 Compound 20 39.5 3.6 M14 taxol + mmpi 9 Compound 20 34.1 5.7 M14 MDS = 1.0 g 6. HT-29 tumor with Irinotecan Mice were implanted with HT-29 tumors and allowed to grow to 50-100 mg. The animals were pair matched and this was considered Day 1. Treatment with MMP inhibitor was begun BID on Day 1 until the end of the experiment. Irinotecan was injected IP (100 or 50 mpk) QD for 5 days (days 1-5). Tumors were measured weekly until an endpoint of 1.0 g was reached.

Irinotecan MMP MMP MDS SEM Dose inhibitor inhibitor (mpk) Dose (mpk) vehicle36.4 4.3 mmpimmpiCompound 100 37.9 5.0 M14 mmpi 20 36 4.2 Irinotecan 100 36.7 2.6 Irinotecan 50 38.1 3. 0 Irinotecan + 100 Compound 100 51.4 4. 4 mmpiM14 Irinotecan + 50Compound 100 44.4 4.0 mmpi M14 Irinotecan + 100 Compound 20 40.6 4.7 mmpiM14 Irinotecan + 2036.1Compound 3.0 mmpiM14

MDS = 1.0 g Integrin Antagonists 1.

Cancer cells were implanted subcutaneously in genetically engineered mice and grew large-volume tumors (>1,500 mm3). Subsequent administration of compound 17 reduced tumor growth by as much as 85 percent in a dose dependent manner. (Nickols A, et al. Inhibition of tumor growth and metastasis by an avp3 integrin antagonist. Presented at the 89 Annual Meeting of the American Association for Cancer Research, March, 1998.)

2.

In an additional experiment, tumor cells were implanted into mice; lung tumors of volumes greater than 2,000 mm3were developed. The mice were then separated into four groups, including a control group and three treatment groups: compound 17 alone; compound 17 with cisplatin (a cytotoxic drug); or cisplatin alone.

Compared to the control groups, the mice treated with combination compound 17/cisplatin therapy experienced more than an 80 percent reduction in tumor size. In comparison, the group receiving cisplatin alone experienced 50 percent reductions in tumor size and thecompound 17 group experienced 20-30 percent reductions. These studies indicate that compound 17 has prominent anti-tumor activity.

3. M21 human melanoma, rat Leydig testicular carcinoma, Lewis Lung and human xenograft models: To test the utility of an, b3 antagonists as single agents and in combination chemotherapy, the M21 human melanoma, rat Leydig testicular carcinoma, and the Lewis Lung carcinoma (LLC) model as well as other human tumor xenograft models were utilized. Tumor cells for implantation were taken from cells either grown in tissue culture (Leydig, M21) or serially passaged as tumors in mice and prepared as tumor brei (LLC). Mice were injected subcutaneously in the proximal dorsal midline with 5 x 106 tumor cells and administration of

test compound or vehicle was initiated the evening of the same day. Tumor volumes were measured at intervals over the course of the experiments. Tumors were measured with a vernier caliper and volumes were determined using the formula for the volume of a cylinder: tumor volume = width2 x length x 0.52. Blood was routinely drawn for plasma drug concentration 6 hours post-dosing on day 4 or 5 and again 12 hours post- dosing on the day of sacrifice. On the final day of the experiment, tumors were dissected free and weighed. The data are expressed, as the mean +/-SEM. Student's and Mann-Whitney tests were used to assess differences between means or medians using the InStat software package.

In the LLC model, compound 17 was administered continuously beginning on day 1 after implantation of the tumor cells, and the chemotherapeutic, cisplatin, was administered as a single intraperitoneal dose of 10 mg/kg on day 5. In this study, cisplatin alone significantly retarded the growth of the LLC tumor (p<0.05). Compound 17 (1 and 10 mg/kg, BID, PO) did not affect the growth of the primary tumor mass. However, the combination of compound 17 together with cisplatin resulted in an additive effect and a significant tumor growth delay (time to develop a tumor > 500 mm3 was: vehicle = 18.1 days; cisplatin = 22.4 days; cisplatin + compound 17 (10 mg/kg) = 27.3 days). The final tumor volume was also significantly reduced with the combination of cisplatin and compound 17 producing a reduction of final tumor volume of 68% in combination (p<0.05). Moreover, the combination of cisplatin and

compound 17 resulted in a 39% improvement in median survival time over vehicle controls and an enhancement over either agent alone (28 days for the vehicle group; 33 days for the cisplatin group; 33 days for the compound 17 at 10 mg/kg group; 38 days for the combination group). Similarly, compound 17 reduced tumor volume when given with cisplatin in a dose- sequencing protocol. The combination of a, b 3antagonist and chemotherapeutic agent was more efficacious than cisplatin alone, particularly when therapy with compound 17 (po, BID) was begun at the same time as cisplatin (once, IP on day 5) or 5 days later (p<0.05 or less for all).

In the M21 model, M21 human melanoma cells implanted subcutaneously into SCID mice developed tumors which grew to approximately 400 mm3 within 30 days.

Oral administration of compound compound 17 (BID) dose- dependently retarded the growth of these tumors when administered at the time of tumor implantation or beginning up to 21 days after implantation. Time to develop a tumor mass > 200mm3 was significantly lengthened in the group treated with the ab, antagonist (time to tumor volume > 200 mm3 was: vehicle = 15 days; compound I7,10 mg/kg = 27 days). These data clearly demonstrate the utility of compound compound 17 to inhibit the growth of pre-existing and established tumors. Moreover, compound compound 17 increased the antitumor efficacy of cisplatin when treatment with the avb3 antagonist was begun on day 1, prophylactically, or therapeutically, on day 14 or 17 (all combinations

significantly less than cisplatin alone, p<0.05).

Cisplatin was administered once by ip injection (10 mg/kg) on day 14. Final tumor weights were nearly identical in the combination treated groups, with clear enhancement of the effect of cisplatin treatment alone.

The results of this dose sequencing experiment establish the efficacy of compound 17 in combination therapy with cisplatin when administered before, concurrent with, or after cisplatin dosing.

The Rice 500 rat Leydig testicular tumor grows very quickly when implanted into the flank of SCID mice.

Compound 17 inhibited tumor growth dose-dependently when given in the drinking water at concentrations of 0.02 to 2 mg/ml. Tumor growth was reduced by about 50% at the 2 mg/ml dose in this aggressive model. Since the tumor does not express the avb3 integrin, the antitumor effects were likely to be produced by the inhibition of angiogenesis. Similar to the results seen in the M21 tumor model, compound 17 increased the effects of cisplatin in the Leydig tumor model. Indeed, the combination of cisplatin and compound 17 was almost 100% effective in preventing tumor growth over the 11 day course of the study. Dose-related inhibition of tumor growth by compound 17 (10 or 100 mg/kg, BID, PO) was also seen when the compound was given as monotherapy or in combination with cisplatin (10 mg/kg, ip once on day 5) (p<0.01 vs control). Therapeutic treatment with the ab3 antagonist was begun at the same time as cisplatin on day 5, with tumor volumes of about 200 mm3at the initiation of therapy. In a similar experiment, the

effects of compound I7, cisplatin and the combination were evaluated for potentiation of overall survival in the Leydig tumor mice. Survival was increased by either compound 17 or cisplatin alone when compared to vehicle treated controls (p<0.05). More importantly, the combination of the two agents almost doubled overall survival (from 17 to 29 days) (p<0.01 combination vs. cisplatin, p<0.001 combination vs. control). Thus, the ability of compound 17 to work alone or in combination therapy to prevent tumor growth clearly correlates with enhanced survival.

4. U251 Glioblastoma Model: compound 17 was evaluated in the human U251 glioblastoma model. The tumors were implanted onto the flanks of SCID mice and the mean tumor volume with time was calculated. In this model, at the dose tested (10 mg/kg, BID, PO), compound 17 produced little inhibition of tumor growth by itself when administered from day 14 through 44. The chemotherapeutic agent, BCNU (12 mg/kg) administered once a day on days 14,18 and 22, induced a regression of the tumors to the limit of detectability, but the tumors grew back. Combination treatment with BCNU and compound 17 regressed tumors to the limit of detectability throughout the period of treatment (compound 17 administered from day 14-44) and almost through the rest of the study. When the data are examined as time to tumor progression (days to 2 tumor doublings), there is clear enhancement by the drug combination over the antitumor effects of either agent alone (p<0.01). Moreover, the response rate (responders

to BCNU) is markedly enhanced and the duration of the response is increased 5-fold from 5 days to 25 days (p<0.01). These clinically relevant measurements of antitumor efficacy establish the antitumor efficacy of compound I7, especially when combined with standard of care chemotherapeutic agents.

5. A2780 Mouse Model: compound 17 prevents the growth of human ovarian carcinoma in SCID mice. The A2780 tumor line is another aggressive tumor model characterized by rapid growth. compound 17 treatment (10 mg/kg, BID, PO) was equally effective as cisplatin (10 mg/kg, ip once on day 20) in decreasing tumor growth. However, as seen in the other tumor models, compound 17 potentiated the effects of cisplatin, resulting in an 80% reduction vs control on day 30. Survival studies are now underway to characterize the survival benefit of combination therapy in this model.

6. Corneal Micropocket Assay: In this model, an intrastromal pocket is surgically created in the normally avascular cornea of female C57BL6 mice 1mm distance from the corneal-scleral junction. A slow release hydron polymer pellet containing an angiogenic growth factor (bFGF or VEGF) is inserted into the corneal pocket. The pocket is self sealing and antibiotic ointment is placed in the eye.

Five days later the eyes are examined under a slit lamp and the neovascular response is quantitated by measuring the average vessel length (VL) and the contiguous

circumferential zone (CH=clock hours where 1 CH = 30 degrees) and plugged into the formula of half an ellipse; Area (mm2) = 0.5 x 3.1416 x VL x CH x 0.4. compound 17 administered BID is a potent inhibitor of angiogenesis in the mouse corneal micropocket model. compound 17 dose-dependently inhibited the angiogenic response up to 42% with maximal inhibitory activity observed at doses of 10mg/kg, BID orally. Moreover, compound 17 inhibited angiogenesis induced by either bFGF or VEGF, the two predominant growth factors known to be produced by tumor cells in vivo. These data confirm the mechanism of action of compound 17 as direct inhibition of angiogenesis in vivo.

7. Metastasis Accurate quantitation of early-stage metastasis in animal models is typically hampered by the lack of sensitive and convenient assays to detect low numbers of tumor cells in a background of normal tissue.

Quantitation of late-stage metastasis by counting of visible foci or comparison of organ weights requires substantial tumor burden which can take 3-4 months to develop in conventional models of breast cancer, and generally cannot detect subtle differences. To develop a more quantitative metastasis model in which the effect of inhibitors on multiple stages of the metastatic process could be dissected, we have produced stable MDA- MB-435 breast carcinoma cell lines expressing a synthetic variant of green fluorescent protein (GFP).

The GFP-transfected cells are easily detected by flow cytometry, and fixation of the cells or the addition of

antibodies or exogenous substrates is not required. A highly aggressive clone was isolated from the lung of a SCID mouse implanted in the mammary fat pad with several GFP-expressing clones. This line, designated 435/GFP HAL-1, consistently generates substantial tumor burden in the lungs by 8-9 weeks compared with 12-16 weeks for the parent line. As few as 1 tumor cell in 200,000 host cells can be detected by flow cytometry, and fluorescent cells are detected in the lungs and blood as early as one week post-orthotopic implantation. compound 17 was administered at doses of 1,10, and 30 mg/kg, BID, orally following orthotopic surgical implantation of 435/GFP HAL-1 cells into the mammary fat pad of SCID mice. Eight weeks later, lungs were removed and weighed. Metastasis was quantitated using a semi- quantitative visible scoring method of gross metastases under a dissecting scope or, following dissection and disaggregation of lung tissue, by flow cytometry of GFP expressing cells. compound 17 administration dose- dependently reduced the spontaneous metastasis of 435 breast carcinoma cells to the lungs as determined either by direct visual counting or quantitation by flow cytometry. Doses of 10 and 30 mg/kg resulted in a 55% and 69% reduction in lung metastatic burden, respectively. However, compound 17 did not delay the growth of the primary tumor mass in this model.

Histological examination of lung sections from these studies revealed a dramatic reduction in the number of large macroscopic metastases and an increase in the presence of microscopic foci of metastases in the compound 17 treated animals.