TW200836749A - Compositions including triciribine and bortezomib and derivatives thereof and methods of use thereof - Google Patents

Abstract

This application relates to combination therapies including triciribine and related compounds and bortezomib and derivatives thereof analogs and compositions with reduced toxicity for the treatment and prevention of tumors, cancer, and other disorders associated with abnormal cell proliferation.

Description

200836749 IX. INSTRUCTIONS: INSTRUCTIONS STATEMENT This application claims the benefit of U.S. Provisional Patent Application Serial No. 60/879,344, filed on Jan. 9, 2008, the disclosure of which is hereby incorporated by reference. data. v

FIELD OF THE INVENTION The present application relates to combination therapies comprising tricineribine and related compounds with bortezomib and its derivative analogs and for treatment and pretreatment 10 A composition that reduces toxicity against tumors, cancer, and other conditions associated with abnormal cell proliferation. BACKGROUND OF THE INVENTION Cancer is the abnormal growth of cells. Despite the limitations of space, the nutrients shared by other cells or the body's signal to stop the regeneration of the cells ^ cells still: rapid regeneration. The shape of cancer cells is usually different from healthy cells, and their function is not normal and can spread into many parts of the body. The abnormal growth of tissue, called a tumor, is a population of cells that can grow uncontrollably and divide. The tumor can be benign (non-cancerous) or malignant (cancerous). Benign tumors tend to grow slowly and do not disperse. Malignant tumors can grow, invade and destroy nearby normal groups and can spread throughout the body. The classification of it's cancer is based on the fluid or organization or formation of its origin. ❹布—Cutter. General categories: cancer, sarcoma, lymphoma, white hill disease, and (four) tumors, 5 200836749 5 10 & some kinds of table tf sales organization and money classification system. Carcinomas are cancers found on the body of the body 22, that is, on the surface of the covering palace, gland or body structure or as they are: For example, the cancer of the stomach is called a cancer. Many cancers affect organs or glands involved in secretion, such as the mammary glands that produce breast milk. Carcinoma is the main cause of fineness of all cancer cases to 9 ()%. Sarcomas are self-connecting tissues, such as cartilage, fat, muscle, tendon, and bone, and malignant tumors that grow. The most common sarcoma, a tumor on the bone, usually exists as a young person. Examples of sarcomas include osteosarcoma (bone) and chondrosarcoma (cartilage). Lymphoma refers to a cancer that originates from the knot or gland of a lymphocyte (which functions to produce a white gray ball and a corpus callosum) or n (such as the brain and breast). Lymphomas fall into two categories: 萑. Hodgkin’s lymphoma and non-Hodgkin's lymphoma. Leukemia, also known as blood cancer, is a bone marrow cancer that prevents the bone marrow from producing normal red blood cells and white blood cells and platelets. White blood cells are needed to fight infection. Red blood cells are needed to prevent anemia. Platelets prevent the body from easily licking and bleeding. Examples of leukemia include acute myeloid leukemia, chronic myeloid leukemia, acute lymphocytic leukemia, and chronic lymphocytic leukemia. The terms "myeloid," and "lymphocyte," refer to the type of cell involved. Finally, the myeloma line grows in the plasma cells of the bone marrow. In some cases, the myeloma cells can accumulate in a bone and form a single tumor, i.e., a cell tumor. In other cases, however, the myeloma cells can accumulate in many bones, thereby forming a bone tumor, also known as a multi-myxoma. Tumor induction and evolution are usually the result of changes in the accumulation of the tumor cell genome. Such changes include inactivation of a cell growth inhibitory gene or a tumor suppressor gene, and activation of a cell growth promoting gene or oncogene. 20 200836749 Many activated cell oncogenes have been identified in animal models to date, however, only a few of these genes have been shown to be associated with human cancer (Weinberg), 19 series 9 Oncogenes and the Molecular Origins of Cancer Cold Spring Harbor, NY? Stanbridge and Nowell 1990 Cell 63 5 867-874, Godwin et al., 1992 Oncogenes and antioncogenes in gynecological malignancies. In WJ Hoskins, CA Perez and RC Young (eds)? Gynecological oncology '.principles and pp 87-116, Lippincott , Philadelphia). The activation of oncogenes in human cancer can result from factors such as increased gene replication or structural changes. These factors can cause many cellular actions, for example, which can lead to overexpression of the gene product. Several oncogenes involved in human cancer can be activated via gene overexpression. It is clear that the continuous genetic abnormality of cancer cells leads to the regulation of normal cell proliferation, differentiation, and programmed cell death to a conductive 15 pathway defect (Hanahan, D. and RA Weinberg, Cell, 2000. 100(1) :ρ· 57-700). This in turn leads to a fundamental defect in cell physiology, which can lead to malignancy. These defects include: a) self-sufficiency of growth signals (ie, growth factor receptor tyrosine acid, such as EGFR overexpression and downstream signal transduction pathways such as Ras/Raf/Mek/Erk% and 20 Ras/PI3K/ Abnormal activation of Akt), b) resistance to growth signals (ie, low expression of TGFB and its receptors), c) escape from apoptosis (ie loss of pro-apoptotic p53; pro-survival Bcl-2) Overexpression; survival pathways, such as high activation of the mediator's survival pathway by P13K/Akt, d) sustained angiogenesis (ie, high secretion of VEGF) and f) tissue invasion and metastasis (ie, cell 7 200836749) And original metastatic integrin) (along 1^11,0.&11 (1 in human Weinberg, Cell, 2000·100(1): ρ· 57-700) 〇 receptor tyrosine kinase, such as EGFR , ErbB2, VERFR, and insulin-like growth factor I receptor (IGF-1R), closely involved in many human cancers, 5 including the formation of colorectal cancer, pancreatic cancer, breast cancer, and ovarian cancer (Khaleghpour et al., 2004, Carcinogenesis) , 25:241-8·, Sekharam et al., 2003, Cancer Res, 63:7708-16). Ligand Such as EGF, VEGF and IGF-1, binding to their receptors can promote the stimulation of endogenous tyrosine kinase activity, specific tyrosine in the 10 cell body pulp domain of these receptors Chemotactic and recruitment of signaling proteins that can trigger a variety of complex signal transduction pathways (Olayioye et al, 2000, Embo J 19: 3159-3167, Porter et al, 1998, Oncogene 17: 1343-52). Tumor viability and tumorigenic pathways, such as these Ras/Raf/Mek/Erk%, JAK/STAT3, and PI3K/Akt pathways, although all three pathways involve colon, pancreas, breast, and ovary Tumor formation, but these pathways by Akt have been shown to be important in malignant degeneration, including many steps in cell proliferation, anti-apoptosis/survival, invasion and metastasis, and angiogenesis (Datta et al., 1999). , Genes Dev, 13:2905-2927). 20 Akt is a serine/threonine protein kinase (also known as PKB) with three family members: AkU, Akt2 and Akt3. Cells stimulated by growth or survival factors Lipase kinase, phosphoinositide-3 -OH-kinase (PI3K), which phosphorylates phosphoinositide-4,5-bisphosphate (PIP2) to form PIP3, which complements the cell membrane with Akt, which can be borrowed from Thr3 0 8 and S er47 3 (Akt 1), Thr3 0 8 25 and Ser474 (Akt2) and Thr308 and Ser472 (Akt3) phosphorylation reaction 200836749 to activate Akt (Datta, SR· et al. Genes Dev, 1999, 13(22): ρ· 2905 -27). Therefore, PI3K can activate Akt by phosphorylating PIP2 and converting it to PIP3. The phosphatase, guanidine, dephosphorylates ΡΙΡ3 to form ΡΙΡ2, thus preventing Akt activation. 5 Most human cancers contain highly activated Akt (Datta et al., 1999,

Genes Dev 13: 2905-2927; Bellacosa et al, 1995 · Int J Cancer, 64: 280-2855; Sun et al, 2001, Am J Pathol, 159: 431-437). In more detail, in 57%, 32%, 27%, and 36% of human colorectal cancer, pancreatic cancer, breast cancer, and ovarian cancer, Akt is overexpressive and/or highly active at 10 degrees (Roy et al. 2002, Carcinogenesis, 23: 201-205; Altomare et al, 2003, J Cell Biochem, 88: 470·476; Sim et al, 2001, Cancer Res 61: 5985-5991; Stal et al, 2003, Breast Cancer 5: R37-R44; Cheng et al., 1992, Proc Natl Acad Sci USA, 89: 9267-9271; Yuan et al., 2000, Oncogene, 15 19:2324-2330). The high activation of Art results from the amplification and/or overexpression of Akt itself and genetic alterations upstream of Akt, including the overexpression of receptor tyrosine kinases and/or their ligands (Khaleghpour et al, 2004, Carcinogenesis, 25: 241-248; Sekharam et al, 2003, Cancer Res, 63: 7708-7716; Cohen et al, 1998, Biochem Soc 20 Symp, 63: 199-210; Muller et al, 1998, Biochem Soc Symp, 63: 149-157; Miller et al, 1995, J Virol, 69: 4390-4398; Slamon et al, 1987, Science, 235: 177-182; Andmlis et al, 1998, J Clin Oncol, 16:1340 -1349) and deletion of the phosphatase PTEN. It has been demonstrated that ectopic presentation of Akt induces malignant deformation and promotes cell 25 survival (Sun et al., 2001, Am J Pathol, 159: 431-437; Cheng et al. 9 200836749, 1997, Oncogene, 14: 2793-2801 And the division of the Akt pathway inhibits cell growth and induces cell death (Jetzt et al., 2003, Cancer Res, 63: 6697-6706) and preclinically demonstrates Akt's conceptual validation of tumor formation. Current treatments for cancer and related diseases are limited in effectiveness and have many 5 serious unintended side effects. Although many anticancer drugs have proven clinically effective, severe systemic toxicity usually halts the clinical development of successful chemotherapeutic agents. In addition, overexpression of receptor tyrosine kinases (such as EGFR) and their ligands (such as IGF-1), excessive expression and/or loss of Akt in PTEN (which all lead to high activation of Akt) and cancer The patient's poor prognosis, resistance to chemotherapy, and shortened survival were associated. Current research strategies emphasize the search for effective treatments with low risk. Therefore, the triclinide compound or a derivative thereof and stilzomicin or a salt or derivative thereof can be used as a potential combination therapy for treating tumors, cancer, and abnormal cell proliferation. 15 EMBODIMENT: 5 § 1 SUMMARY OF THE INVENTION The present invention provides a treatment of a tumor or cancer in a patient and which can limit systemic toxicity, including triclinbine, tricineridine phosphate and related compounds with bortezomib and its derivatives. Novel treatment options. The present invention is found in the following substrates: 20 The synergistic effect of the cytotoxic effect of a tumor or cancer overexpressing Akt kinase on TCN and related compounds with the combination of bortezomib salt or a derivative thereof and a derivative thereof The effect is particularly sensitive. Unlike previous prior art and experience, the inventors have identified a successful method for the treatment of tumors and cancer with tromethine and bortezomib and its derivatives, by one or the following group 10 200836749 combination: (1) Administering such drugs to patients who show increased sensitivity to tromethine and bortezomib and its salts or derivatives; (9) using such doses that minimize the toxicity of such drugs but still exhibit efficacy; Or (10) using the dosing regimen that minimizes the toxicity of such drugs. In the present invention - the illustrated embodiment, the present invention encompasses a composition comprising: (1) a compound of formula I-IV:

11 200836749 a ruler 2, R3' and I' are each independently hydrogen, optionally substituted phosphorus g or two salts (including mono-, di- or tri-disc acid salts or diazepam An acid group (which includes a low carbonic acid group); a burnt group (which includes a low-calcining group); a brewing amine '% _' which includes a silk-burning base; a sulfonate, which includes a continuation-helmet soil, wherein the benzene The group may be optionally substituted by one or more substituents as described, for example, in the definition of the aryl group as taught herein; optionally substituted aryl sulfhydryl; lipid, its (iv) lipid; amino acid, · Carbohydrates, months, or cholesterol; or in vivo, R2, R3, and R5,

Other pharmaceutically acceptable eight R and R independently of the compound of the mono- or di-phosphate are hydrogen, optionally substituted phosphate; the sulfhydryl group includes low carbonic acid Base); _, a base (which includes a low-cracking base); an aromatic & water-oxygenated olefinic smoke such as polytrimethylene, a selectively substituted aragonite yellow base, which comprises a phospholipid; an amine a base acid; a carbohydrate; a peptide; or a cholesterol 15 sterol, or other pharmaceutically acceptable release group. In one embodiment, the compound is administered as a 5'-phosphate ether lipid or a 5,-ether lipid;

Ri and R2 are each independently a linear, optionally substituted linear, branched or cyclic alkyl group (which includes a hindered alkyl group), an alkenyl or alkynyl group, a c〇-alkyl group, a c〇_alkenyl group. , CO-alkynyl, CO-aryl or heteroaryl, co-alkyloxyalkyl, c〇_ 20 aryloxyalkyl, CO-substituted aryl, sulfonyl, alkanesulfonyl, arylsulfonate Base, aromatic burning base; (ϋ) Formula V compound.,: 12 200836749

Wherein R 1 ' R 2 , R 3 , R 4 , and 5 are each independently oxime, selectively halogenated, substituted straight bond, branched or cyclic alkyl (which includes lower alkyl), alkoxy, 5 diluted Or block, aryl, CO. alkyl, CO-alkenyl, CO-alkynyl, CO-aryl or heteroaryl, CO-alkoxyalkyl, CO-aryloxyalkyl, CO-substituted Aryl, sulfonyl, alkanesulfonyl, arylsulfonyl, aralkylsulfonyl; and (iii) a pharmaceutically acceptable carrier. In another illustrative embodiment of the invention, the invention encompasses a composition comprising TCN, TCN-P, TCN-PM or the like, a combination thereof and a salt of pentapizozime or a salt or derivative thereof. In further clarified embodiments, the invention encompasses a method of treating a tumor or cancer in a mammal comprising administering to the mammal an effective amount of a composition comprising 15 compounds: (1) a compound of formula I-IV. 200836749

5 wherein R 2 ', r 3 ' and r 5 ' are each independently hydrogen, optionally substituted phosphate or phosphonate (which includes mono-, di- or triphosphate or stabilized phosphate prodrugs); (which includes a lower fluorenyl group); an alkyl group (which includes a lower alkyl group); a decylamine, a phthalic acid, which is a sulphur-based or aryl-alkyl group, which includes a fluorenyl group and a benzyl group. Wherein the phenyl group is optionally substituted with one or more substituents as described, for example, in the definition of the aryl group as taught in 10; optionally substituted azulsulfonyl; a lipid comprising a phospholipid; Amino acid; carbohydrate hydration 14 200836749 Physic, skin, or cholesterol, or other pharmaceutics in which anaerobic 2, r3, and R5, which are independently hydrazine or mono-, di- or tri- sulphate, can be obtained in vivo. Acceptable leaving group; wherein R and Ry are independently hydrogen, optionally substituted phosphate 丨醯5 group (which includes lower sulfhydryl groups); decylamine, alkyl group (which includes lower alkyl aromatic poly Oxidized olefins, such as polyethylene glycol, optionally substituted aryl sulphate; lipids, including phospholipids; amino acids; carbon a compound; a form; or a cholesterol; or other pharmaceutically acceptable exfoliating group. In one embodiment, the compound is administered as a 5,- squaric acid ether lipid or a 5,_yed lipid; 1 〇Rl&R2 are independent W, optionally substituted linear, branched or cyclic alkyl (which includes lower alkyl), alkenyl or alkynyl, c〇-alkyl, c〇-alkenyl, CQ alkynyl, CO. aryl or heteroaryl, Cq_calciferous alkyl, c〇_f oxyalkyl, CO-substituted aryl, syl-, sulfonyl, arylsulfonyl, aryl sulphonic acid ; 15 and (11) formula: ^ compound:

—N

Vr4 \

Ο HR HRs ς Μ,

:shed

Ri, I, r3, r4, and R5 are each independently oxime, selectively halogenated, substituted 15 200836749 linear, branched or cyclic alkyl (which includes a low carbon alkyl), alkoxy, alkenyl or Fast, aryl, CO-, ketone, CO-alkenyl, CO-alkynyl, CO-aryl or heteroaryl, CO-alkoxyalkyl, CO-aryloxyalkyl, CO-substituted aromatic Base, continued base, academic achievements SS base, arylsulfonyl, aromatic sulfonyl. In another illustrative embodiment, the invention encompasses a method of treating a tumor or cancer in a mammal comprising administering to the mammal an effective amount comprising TCN, TCN-P, TCN-PM or a combination thereof and A composition of bortezomib or a salt or derivative thereof. The methods available for treating tumors or cancer are particularly affected by the toxic effects of TCN, TCN-P, 10 TCN-PM and/or related compounds. In another embodiment, a method for treating a tumor in a mammal, particularly a human, comprising (1) obtaining a biological sample from the tumor; (ii) determining whether the tumor overexpresses Akt kinase, and (iii) A combination of triclinbine, trichostatin phosphate or a related compound and bortezomib or a salt or derivative thereof 15 as described herein is used to treat the tumor which overexpresses Akt kinase. In another embodiment, the degree of AkT kinase expression can be determined, for example, by assaying the phosphorylated Akt kinase contained in the tumor or cancer using an antibody that detects the phosphorylated form. In another embodiment, the extent of Ak t expression can be determined by examining the tumor or cancer cells obtained from the patient and comparing to the extent of the control tissue. In a particular embodiment, the Akt has at least 2, 2.5, 3 or 5 folds of over-sounding performance in the cancer sample compared to the control tissue. In a particular embodiment, the overexpressing Akt kinase can be a highly active and phosphorylated Akt kinase. In another aspect of the invention, there is provided a dosing regimen that limits the toxic side effects of TCN and related compounds. In another embodiment, the 16200836749 regimen may minimize or remove toxic side effects including, but not limited to, hepatotoxicity, thrombocytopenia, hyperglycemia, diarrhea, Gastritis and / or fever. In another embodiment, administration of TCN, TCN-P, TCN-PM, or related compounds provides at least a portion, such as at least 15, 20, or 30% or complete, in at least 15, 20, or 25% of 5 patients. In vivo reaction. In another embodiment, there is provided an effective amount of TCN, TCN-P, TCN-PM or related compound and bortezomib and a derivative thereof for administration to a patient diagnosed with a tumor according to a dosing schedule A method of treating the subject, the dosing schedule comprising administering the drugs about once a week for about 3 weeks, and then not administering the drugs for a week. In another embodiment, a dosing regimen of 10 mg/m2 or less of TCN, TCN-P, TCN-PM or related compounds with tezomib and its derivatives is administered to the patient once a week. A method of treating a tumor or cancer in a patient. In another embodiment, the tricitidine compound and bortezomib and its derivatives can be administered in a single large dose in a short period of time, for example, about 5, 10 or 15 minutes. In a further embodiment, a dosing schedule wherein the troxaribine compound and bortezomib and its derivatives are administered by continuous infusion, which takes at least 24, 48, 72, 96 or 120 hours, is provided. In a particular embodiment, the continuous administration can be repeated at least once a week, once every two weeks, and/or once a month. In other embodiments, the trastolimide compound and bortezomib and its derivatives can be administered at least once every three weeks. In other embodiments, the compounds can be administered at least once a week for at least 2, 3, 4 or 5 days. In other embodiments, the patient may be administered a sirolimus and samimi as disclosed herein, and the traitor is effective in causing the tumor to subside. Administration of the tromethamine compound to borteide (tetra) and its derivatives can result in at least a portion, such as at least 15, 20 or 30% or complete in vivo reaction, in at least 15 to 30% of such patients. In a particular embodiment, the 5,10, 15 patient can be administered at least 2, 5, 1 G, ^, as, or as, for example, mg of the trichostatin compound and samizomib and its derivatives. Administration of the triclinide compound _zomib and its derivatives can be carried out according to any of the treatment regimens disclosed herein. In a particular embodiment, the dosing regimen can include administration of less than 20 mg/m2 of Qushebin and Yuzomi and its derivatives. In the examples, less than 1 mg/m 2 of the trichostatin compound and _zomi and its derivatives may be administered once a week. In other embodiments, the patient may be administered a dose of less than 2 mg/m2, 5 mg/m2, a sulki/m2, and/or 15 dg/m of the triclinide compound and bortezomib and Its derivatives. In another embodiment, a dose of less than 1 mg/m2 may be administered to the patient by continuous rounds, which may take at least 5 days. In a particular embodiment, the tromethamine compound and bortezomib and its derivatives as disclosed herein are useful for the treatment of pancreatic cancer, prostate cancer, colorectal cancer, and/or ovarian cancer. In another embodiment, 'the troxamycin compound of the invention and rottenzomib and its derivatives and/or treatment regimen can be used to prevent and/or treat cancer, meat 20 tumors, lymphomas, leukemias, and/or Myeloma. In other embodiments of the invention, the triclinide compound and bortezomib and its derivatives can be used to treat solid tumors. In still other embodiments, the tromethamine compound and bortezomib and its derivatives and compositions disclosed herein can be used to treat tumors or cancers such as, but not limited to, cancers of the following organs or tissues: breast, prostate , 18 200836749 11"The intestines (which are secretly secreted to the colon), the urinary bladder, the bladder, the nails:: gold: lymphoma, melanoma, kidney, adrenal gland, pancreas, pharynx, bismuth, and/or In particular embodiments, the tromethamine complex and bortezomib and its derivatives are useful for the treatment of pancreatic cancer, breast cancer, and/or osteogenesis cancer. Further embodiments of the invention中中,文=之曲西立滨化合物和辨佐米 and its derivatives can be used

10 15

Become a related disease. In the embodiment, the method for treating white disease by infusion of trifluralin compound _zomizol and its derivatives via hydrazine, and at least Γ, 4 „8 7-2 or 9 hours is provided. Other embodiments may repeat a continuous infusion, such as at least once every 2, 3, or 4 weeks. Γ Specific implementation of the financial, health (4) method of reversing, cancer, U-cell hyperplasia related disorders, the method includes An effective amount of a selective amount of a tromethamine compound and bortezomib and a derivative thereof, which are selectively pharmaceutically acceptable carriers. In the present aspect, the far Quercetin compound and the stone pentozomib and The derivative money composition can be formed and can form part of the same composition, or be provided in a different composition suitable for administration at the same time. In other embodiments, the literary disclosure reveals The trichostatin compound No. 20 zomi and its derivatives can be used to treat tumors or carcinomas that are resistant to - or a variety of known anticancer drugs, including the embodiments disclosed herein or the cancer and compounds. - In the examples, the Quxi Libinization disclosed in the text The dosage of the compound and quetstazole and its derivatives can effectively treat patients with drug-resistant tumors or cancers. 'The drug-resistant tumor or cancer is, for example, a multi-drug resistant tumor or 19 200836749 cancer, which includes, but does not Limited to: tumors or cancers that are only resistant to yew, anti-rapamycin, tamoxifen, anti-cisplatin, and/or anti-gefitinib (iressa). In the present invention, there is provided a method comprising the addition of a tresibine compound and a sirolimus and a derivative thereof, or a tumor thereof, which is effective for treating a host, which is disclosed in the article which is effective for the main treatment. a pharmaceutical composition of the triclinic compound and bortezomib and its derivatives of other diseases associated with cancer, and abnormal cell proliferation. In another embodiment, a method for treating a tumor or cancer is provided. The method comprises administering to an individual in need of treatment an effective amount of a compound, or a salt, isomer, prodrug or ester thereof, as disclosed herein, wherein the cancer is, for example, a carcinoma, sarcoma, lymphoma, leukemia or myeloma. The compound or a salt thereof The isomer, prodrug or ester may optionally be provided as a pharmaceutically acceptable composition comprising a suitable carrier, such as water, which composition is adapted to be administered to a subject in need thereof, 15 The compound can be administered selectively or in combination with at least the external therapeutic agent to treat the tumor or cancer. The scope of the invention also includes a compound or a salt thereof which is selectively disclosed in the context of a pharmaceutically acceptable carrier, a prodrug or a sirolimus compound and a bortezomib thereof and a salt thereof, a prodrug or a drug thereof, which are disclosed in the context of treating a tumor or a drug, and optionally in a pharmaceutically acceptable carrier Use of Cui to prepare a drug for treating cancer or tumor. Brief Description of the Drawing Figure 1 illustrates API-2 (Cucidine) as an Akt inhibitor derived from NCI diversity, a Diversity Set. Applicable to the identification of the object. The first eight knives 20 200836749 The clarification of the chemical structure of API-2 (Quxi Libin). Figure (1) shows that Αρμ2 inhibits the phosphorylation of ΑΚΤ2 in ΝΙΗ3Τ3 cells transformed by ΑΚΤ2. Treatment of wild-type ΑΚΤ2-transformed νιη3Τ cells with ΑΡΙ-2 (1 μΜ), time-consuming, and anti-phosphorylation of Akt_T3〇8 and ·S473 antibody 5 (top group and intermediate group) for immunoblotting analysis. The bottom group gives the performance of the total AKT2. In the ic diagram, it is shown that api-2 can inhibit three variants of Akt. HEK293 cells were transfected with HA-Akt1, -AKT2 and -AKT3 and treated with apl2 (1 μΜ) or wortmannin in (15-) before EGF stimulation, so that the cells were solubilized and resistant. ΗΑAnti-1 steroid immunoprecipitation. The immunoprecipitates were subjected to an in vitro kinase assay (top) and immunoblot analysis was performed using an anti-phospho-Akt-T308 (bottom) antibody. The middle group shows the expressiveness of the metastatically infected Aktl, AKT2 and AKT3. Dijon) The clarification of API-2 does not inhibit Akt in vitro. A live 15 in vitro kinase assay of the recombinantly active AKT2 recombinant peptone in a kinase buffer containing ΙμΜ AH-2 (lane 3). Figure 2 illustrates that API-2 does not inhibit PI3K, PDK, and closely related members of the AGC kinase family. Figure 2A illustrates an in vitro PI3K kinase assay. The serum of ΗΕΚ293 cells was deficient and treated with Αρι 2 (丨 or wortmannin (15 μΜ) for 30 minutes before EGF stimulation. The immunoprecipitates were subjected to in vitro kinase assay using ρι_4_ρ as the 20 matrix. Figure 2B is dark The effect of API-2 on the activation of PDK1 in vitro (the top group), the solid circle indicates the inhibition by API-2. The open circle indicates the positive control stamOsporine, which It is an effective PDK1 inhibitor (IC50=5nM), and the inhibition is carried out. The bottom group is 妒21 200836749

Myc_PDK4 was transferred to immunoblotting analysis of HEK293 cells infected with wortmannin or API-2 prior to EGF stimulation. The immunoblot method is detected by the designated antibody. Figure 2C is an illustration of the immunoblotting of PKCa phosphorylation using anti-phosphorylated PKC α T638 (top) and assembly 5 PKCa (bottom) antibodies after treatment with API-2 or a non-selective PKC inhibitor Ro31-8220. Point analysis. Figure 2D shows an in vitro SGK kinase assay. HEK293 cells were metastasized via HA-SGK transfer and treated with API-2 or wortmannin prior to EGF stimulation. The in vitro kinase assay was performed using the MBP as the substrate (top) with the HA_SGK immunoprecipitate. The bottom panel represents the table 10 of the HA-SGK that has been transferred. Figure 2E illustrates the results of the PKA kinase assay. The immunopurified PKA was incubated in ADB buffer (Upstate Biotechnology Inc) containing the indicated inhibitor (API-2 or AKAI) and matrix, Kemptide. The kinase activity was quantified. The 2F figure represents the Western dot method. Treatment of OVCAR3 cells with API-2 took time to specify. Immunoblots of cell lysates were performed with the indicated anti-phospho antibodies (Groups 15 to 4) and anti-actin antibodies (bottom). Figure 3 is a diagram showing that API-2 inhibits Akt activity and cell growth and can induce apoptosis of human cancer cells with high Akt. Figure 3A shows Western blotting. After treatment with API-2, the phosphorylation of Akt was detected by anti-phospho-Akt-T308 antibody in designated human cancer cell lines. These dot methods were then probed with anti-total Akt antibody 20 (the bottom panel). Figure 3B shows a cell proliferation assay. The cell lines shown in the figure were treated with different doses of API-2, which took 24 hours and 48 hours, and then analyzed by the CellTiter 96 Cell Proliferation Assay Kit (Promega). Figure 3C provides an analysis of apoptosis. Cells were treated with API-2 and stained with annexin V and PI, and then analyzed by FACScan. 22 200836749 Figure 4 shows that API-2 inhibits downstream targets of Akt and displays anti-tumor organisms in cancer cell lines with high Akt in mouse xenografts. Figure 4A shows that API-2 inhibits Akt phosphorylation of potatorin, Bad, AFX and GSK-3/5. After treatment with API-2, OVAR3 5 cells were lysed and the ink spots were immunized with the indicated antibodies. Figure 4B shows that API-2 inhibits tumor growth. Tumor cells were injected subcutaneously into nude mice with low Akt cell content on the left and high Akt cell content on the right. When the tumors reached an average size of about 100 to 150 mm 3 , the animals were treated with vehicle or 1 mg/kg/day of API-2. Each measured value represents the average of 10 tumors. Figure 4C is a graphical representation of mice treated with 10 API.2 or vehicle (control) with OVCAR3 (right) and OVCAR5 (left) xenografts. Fig. 4D shows the tumor size (bottom) and weight (top) at the end of the experiment. In Figure 4E, anti-phospho-Akt_S473 (top) and anti-AKT2 (lower) antibodies were used for tumor lysis in treated (T3 and T4) and untreated (T1 and T2) OVCAR-3-derived tumors. The product was analyzed for 15 points. Figure 5 shows that API-2 (Cucidine) inhibits in vitro kinase activity. In vitro kinase assays were performed in recombinant kinases containing PDK1 and Akt in a kinase buffer containing phospholipid creatinine-3,4,5-P3 (PIP3), API-2 and tissue protein as matrix. After 30 minutes of incubation, the aliquots were separated by SDS-PAGE and exposed to 20 in the film. Figures 6a to c provide mRNA and amino acid sequences of human Aktl, as well as restriction enzyme positions. Figures 7a to d provide mRNA and amino acid sequences of human Akt2, as well as restriction enzyme positions. 23 200836749 Figures 8a to C provide mRNA and amino acid sequences of human Akt3, as well as restriction enzyme positions. Figure 9 is a graph showing the effect of bortezomib synergistically enhanced API-2 (TCN) on these viable myeloma cells. H929 cells (derived multiple bone 5 myeloma) were treated with 0, 2.5, 5, 10, 20 or 40 nM bortezomib only in the presence of ADIμΜ ADI-2 or only 0, 2.5, 5, 10, 20 or 4 (^1\/[eight 01-2 treatment. Calibrate and map cell viability (Fig. 9A). U266 cells (derived multiple myeloma) are only 0 or in the presence of ΙΟηΜ API-2 , 2.5, 5, 10, 20 or 40 μΜ ADI-2 treatment or treatment with only 〇, 2.5, 5, 10, 20 or 40 nM boron for 10 zomi. Calibrate and plot cell viability (Fig. 9B). It indicates the synergistic effect of bortezomib and API-2 (TCN) on the enveloped cell lymphoma cells. The Jeko-Ι cells (enveloped cell lymphoma) are treated only in the presence of 5 μΜ API-2. , 2.5, 5, 1〇, 20, 40 or 801^ bortezomib treatment or treatment with 0, 1.25, 2.5, 5, 10, 20 or 4(^]^1 八 1 1 2 15 . Calibrate and plot cell viability [Embodiment 3 Detailed Description of the Preferred Embodiments In contrast to the prior art and experience, the inventors have determined to successfully use a combination of tricilide compound and bortezomib and its derivatives to treat swelling. And 2 〇 cancer methods by one or a combination of the following: (1) Patients who exhibit enhanced sensitivity only to the tromethamine compound and/or bortezomib and its derivatives according to the following diagnostic test Administration of triclinib and bortezomib and its derivatives; (ii) use of such doses which minimize the toxicity of the triclinide compound and/or bortezomib and its derivatives but still have efficacy Or (m) 24 200836749 The dosing regimen which minimizes the toxicity of the triclinide compound and/or bortezomib and its derivatives. Definitions As used herein, the term "compound of the invention" A compound of the formula ι-ν - 5 and combinations thereof. As used herein, the terms "cancer" and "cancerous" refer to or describe a cell growth that is typically uncontrolled (ie, a proliferative disorder). Physiological conditions of mammals of a characteristic. Examples of such proliferative disorders include cancers, such as carcinomas, lymphomas, blastomas, sarcomas, and leukemias; and other cancers as described herein. Specific examples include breast cancer, Adenocarcinoma, colon cancer, squamous cell carcinoma, small cell lung cancer, non-small cell lung cancer, gastrointestinal cancer, pancreatic cancer, cervical cancer, ovarian cancer, liver cancer (eg liver cancer), bladder cancer, colorectal cancer, uterus Endometrial cancer, kidney cancer, and thyroid cancer. Other non-limiting examples of cancer are basal cell carcinoma; biliary tract cancer; bone 15 cancer; brain and CNS cancer; choriocarcinoma; connective tissue cancer; esophageal cancer; Cancer; ^ ^ cancer of the head and neck; gastric cancer; intraepithelial neoplasia; laryngeal cancer; lymphoma, including Hodgkin's and non-Hodgkin's lymphoma; melanoma; myeloma; Tumor; oral cancer (eg lip, tongue, mouth, and pharynx); pancreatic cancer; retinoblastoma; rhabdomyosarcoma; rectal cancer; respiratory cancer; 20 sarcoma; skin cancer; gastric cancer, testicular cancer; Cancer of the urinary system; and other cancers and sarcomas. As used herein, the term "tumor" refers to all neoplastic cell growth and proliferation (whether malignant or benign), and all precancerous and cancerous cells and tissues. For example, a particular cancer may be characterized by a solid agglomeration. If present, 25 200836749 The solid tumor mass can be a primary tumor mass. Primary tumor mass refers to the growth of cancer cells within the tissue resulting from the transformation of normal cells of the tissue. In the case of large recordings, the primary tumor mass can be determined by the presence of a cyst (which can be found by visual or palpation methods) or by the shape irregularity, structure or weight of the tissue. However, some primary tumors are not tactile and can only be detected by medical imaging techniques, such as x-rays (e. g., mammography), or by needle aspiration. These techniques are used more often in early detection. Molecular and phenotypic analysis of cancer cells within the tissue usually confirms whether the cancer is an endogenous cancer of the tissue or whether the lesion is caused by a metastasis from another site. Unless otherwise specified, as used herein, the term "alkyl," includes, for example, a saturated straight chain, a branched chain or a ring system, a first, second or second hydrocarbon, and more specifically, Including methyl, trifluoromethyl, ethyl, propyl, isopropyl, cyclopropyl, butyl, isobutyl, tert-butyl, pentyl, cyclopentyl, 15 isopentyl, neopentyl Base, hexyl, isohexyl, cyclohexyl, cyclohexylmethyl, 3-methylpentyl, 2,2-dimethylbutyl, and 2,3-dimethylbutyl. Substituted by, for example, a substituent such as halogen (F, cl, Br or 1) (for example Cf3, 2_Br_ethyl, ch2F, CH2Q, CH2CF3 or CF2CF3), hydroxyl (for example ch2OH), amine group (for example CH2NH2, 20) CH2NHCH3 or CH2N(CH3)2, an amine group, an arylamine group, an alkoxy group, an aryloxy group 'nitro group, an azide group (for example, CH2N3), a cyano group (for example, ch2cn), a sulfonic acid, a sulfate group, a phosphonic acid , phosphate or phosphonate, and if necessary, as known to those skilled in the art, for example, as Greene's in Protective Groups in Organic Svnthegig| John Wiley and Sons 2nd Edition (1991) 26 200836749 shows that the special group may be unprotected or protected, and the document is incorporated herein by reference. Unless otherwise specified, the term "lower alkyl," means (^ To a saturated straight chain, a branched chain or, if appropriate, a ring alkyl group (for example a cyclopropyl 5 group) which includes both substituted and unsubstituted forms. The term "alkylamino" or "arylamino", Included are amine groups each having one or two alkyl or aryl substituents. 10 15 20 octacalcin and synthetic alpha, cold, 7 or (tetra) acid and include, but money: amine found in egg * f (10) Acid, ie, glycine, alanine, aminic acid, leucine, isoleucine, methionine, phenylpropyl «, color _, bribe, thief, threonine, cysteamine Acid, yoghurt, linoleic acid, lysine, acetaminophen, lysine, lysine, arginine, and histidine. In the preferred embodiment, the amine group is present. Or, the axis The base acid may be a derivative of the following: acrylamido, aryl, leucine, iso(tetra), amidoxime, phenylpropanyl, tryptamine, methylthio(tetra)ylamine , ceramide, thiol sulfhydryl, cysteamine sulfhydryl, sulfazone, glutamine amidoxime, glutamine amide, aspartame, lysine diamine, base Amidoxime, amide, μ(tetra)yl, benzyl, /3.isoamine oxime, oxime oxime, phenyldiamine aryl, /3-tryptamine thiol+methylthio-yl, ^ Glycine base, ^ silk ship soil base, ^ sulphide base ^ money _ base, W amine thiol, ^ aspartame 0 aspartame, valerin, arginine, Or /3-histidine group. When the term "amino acid" is used, its 27 200836749 is considered to be a specific and independent disclosure of each of the natural or synthetic amino acid esters in the D and L configuration, including but not limited to :α, /3, τ or 6 glycine, alanine, valine, leucine, isoleucine, methionine, phenylalanine, tryptophan, lysine, serine , sulphate, cysteamine 5, tyrosine, aspartic acid, glutamic acid, aspartate, glutamate, lysine, arginine and histidine. Further, as used herein, the term "protected" includes a group that is intended to be added to an oxygen, nitrogen, sulfur or phosphorus atom to prevent its further reaction or for other uses. A variety of oxygen and nitrogen protecting groups are familiar to the organic Synthetic Skills 10 (see Greene and Wuts, Protective Groups in Orgam V g^nthesis, 3rd edition, John Wiley & Sons, Inc., New York, NY. 1991) 〇 Unless otherwise specified, as used herein The term "aryl" includes phenyl, biphenyl or naphthyl, and is preferably phenyl. The aryl group may be optionally substituted with a molecular group of 15: for example, i-, amino, acryl, arylamine, alkoxy, oxy, nitro, cyano, sulfonic acid, sulfate 'phosphonic acid, can acid or phosphonate, and if necessary, as known to those skilled in the art, for example as Greene et al. in Protective Groups in Orphic Svnth^ic Tnhn

As taught in Wiley and Sons, 3rd edition (1999), these molecular groups may be unprotected or protected. The term "alkaryl," or "alkylaryl, includes alkyl having an aryl substituent. The term "aralkyl" or "arylalkyl" includes aryl having an alkyl substituent. & The term "dentate" as used herein, includes chloro, bromo, iodo, and fluoro. 28 200836749 δ海 the term "mercapto" includes a carboxylic acid ester, wherein the non-base group of the ester group is selected from a linear, branched or cyclic alkyl or lower alkyl group; an alkoxyalkyl group, including Methyl methyl; aralkyl, including a benzyl group; an aryloxyalkyl group, such as a phenoxy fluorenyl group; an aryl group, which optionally includes a halogen, (^ to a decyl group or a 5 Cl to C4 gas group) a substituted phenyl group; a sulfonate such as an alkyl or aralkylsulfonic acid group which includes a methylsulfonyl group, a mono-, di- or triphosphate, a trityl group or a monomethoxytriphenylmethyl group, substituted a benzyl group, a trialkylcarbenyl group (e.g., dimethyl-tert-butylformamyl) or a diphenylmethyl decyl group. The aryl group in the esters preferably contains a phenyl group. "Mercapto" refers to a fluorenyl group wherein the non-carbonyl 10 molecule group is a lower alkyl group. As used herein, "substantially free" or "substantially lacking" in terms of mirror image purity, is meant to include at least 85 or 9 〇 ❻ /., preferably 95 to 98 5% by weight and more preferably 99 to 1 重量% by weight of the composition of the mirror image isomer. In a preferred embodiment In the methods and compounds of the invention, the 15 compounds are substantially free of other mirror image isomers. Similarly, the term "isolated" means at least 85 or 9 % by weight, preferably 95. Up to 98% by weight and more preferably 99 to 100% by weight of the compound and the compound composition containing the chemical species or the mirror image isomer thereof. The term "independent" is used herein to mean the variable. The system is applied separately and varies independently from application to application. Therefore, in a compound (such as R"XYR"), wherein R""is independently carbon or nitrogen,", both R" may be carbon, two R" It may be nitrogen, or one R" may be carbon and the other R, which is nitrogen. The term "pharmaceutically acceptable salt or prodrug" as used throughout this specification is described once it is administered to a patient. This compound 29 200836749 5 10 15 Meituan (1) Acceptance form (editory, meaning, s| or related μ) Accepts the spin-in acceptable inorganic: organic base and acid salt. Suitable salts include derivatives Salt from the following: It is known to us in medicine Metals of many other acids, such as unloading, 'surstrial metals, such as lack of town. Pharmaceutically acceptable prodrugs, mean compounds that can be hydrolyzed or oxidized, for example, to form a compound of the invention. Typical examples of prodrugs include compounds having a biologically labile protecting group on a functional molecular group of the active compound. Prodrugs include oxidative, reduction, amination, deamination, enzymatic, de-based , hydrolysis, dehydrolysis, alkylation, dealkylation, deuteration, deuteration, phosphorylation, dephosphorylation to produce a compound of the active compound. Unless otherwise specified, the term "pharmaceutically acceptable" is used herein. Accepted esters' include those that are suitable for contact with the host within the scope of normal medical judgment and that are not toxic, irritating, allergic, etc., which are commensurate with reasonable benefit/risk ratios and are effective for their use. An ester of one or more compounds of interest. The term "patient, as used herein, refers to an animal, preferably a mammal, preferably a human. Mammals may include non-human mammals including, but not limited to, pigs, goats, sheep, cattle ( Bovine animals, deer, lynx, 20 horses, monkeys; and other non-human primates, such as dogs, cats, rats, mice, rabbits, or any other animal known or disclosed herein. The invention provides the use of a combination of TCN, TCN-P, TCN-oxime and related compounds with bortezomib and its derivatives for the treatment of a proliferative disorder. 30 200836749 Unless otherwise specified, as used herein The term "tricilidine compound" and "tricilidine and related compounds" refer to a compound having the following structure:

5 wherein R 2 , R 3 , and R 5 are each independently hydrogen, optionally substituted phosphate or phosphonate (which includes mono-, di- or triphosphate or stabilized phosphate prodrugs); (including low carbon fluorenyl); alkyl (which includes low sulphur); decylamine, sulphuric acid, which includes alkyl or aryl, and fluorenyl, including thiol and benzyl Wherein the phenyl group is optionally substituted with, for example, a substituent as described in the definition of the aryl group of 10 herein; a optionally substituted arylsulfonyl group; a lipid comprising a phospholipid; an amino acid Carbohydrate; peptide; 31 200836749 or cholesterol" or other pharmaceutically acceptable cleavage groups in which I, %, and R5, independently η or a single = one or two salt salts; And R is independently hydrogen, a selectively substituted phosphate; a ruthenium consists of low-stone anthracene &base; a guanamine, a burnt base (which includes a low-carbon base); and an aromatic take 5 such as polyethylene Alcohol, optionally substituted, sinter-filled February mussels, including dish fats; amino acids; carbohydrates; peptides; or cholesterol; It pharmaceutically acceptable leaving group. In one embodiment, the compound is administered as a 5'-phosphate ether lipid or a 5,-ether lipid.

Ri and R2 are each independently a linear, optionally substituted straight chain, a branched chain or a 0^ group (which includes a lower alkyl group), an alkenyl group or an alkynyl group, a c〇_alkyl group, a c〇-ene group. Base, CO-alkynyl, CO-aryl or heteroaryl, c〇-alkoxyalkyl, CO-aryloxyalkyl, CO-substituted aryl, sulfonyl, alkanesulfonyl, arylsulfonate A arylsulfonyl group; in one embodiment, R2, and r3 are hydrogen. In another embodiment, r2, and I' are hydrogen. In yet another embodiment, r2, r3, and r5 are hydrogen. In still other embodiments, R2, R3, r5, and the heart are converted to hydrogen. In another embodiment, the triclinide compound has the structure: 32 200836749

Wherein R3 is H, optionally substituted straight chain, branched or cyclic alkyl (which includes lower alkyl), alkenyl or alkynyl, NH2, NHR4, N(R4)2, aryl, alkane An oxyalkyl group, an aryloxyalkyl group or a substituted aryl group; and 5 each R4 is independently fluorene; a fluorenyl group comprising a lower fluorenyl group; an alkyl group comprising a lower alkyl group such as, but not limited to, a methyl group , ethyl, propyl and cyclopropyl; alkenyl; alkynyl; cycloalkyl; alkoxy; alkoxyalkyl; hydroxyalkyl or aryl. In a sub-embodiment, R3 is a linear C1 to C11 alkyl, isopropyl, tert-butyl or phenyl group. In one embodiment, the trichostatin compounds provided herein have the following structure: 33 200836749

In another embodiment, the trichostatin compounds provided herein have the following structure:

In another embodiment, the trichostatin compounds provided herein have the following structure: 34 200836749

Wherein R6 is fluorene, alkyl (which includes lower alkyl), alkenyl, alkynyl, alkoxyalkyl, hydroxyalkyl, aralkyl, cycloalkyl, NH2, NHR4, NR4R4, CF3, CH2OH, CH2F , CH2C, CH2CF3, C(Y3)3, C(Y3)2C(Y3)3, 5 C(=0)0H, C(=0)0R4, c(=0)-alkyl, C(=0) -aryl, C(=0)-alkoxyalkyl, C(=0)NH2, C(=0)NHR4, C(=0)N(R4)2, wherein each Y3 is independently H or halogen; Each R4 is independently hydrazine; a fluorenyl group, which includes a lower fluorenyl group; an alkyl group including a lower alkyl group such as, but not limited to, a methyl group, an ethyl group, a propyl group, and a cyclopropyl group; a cycloalkyl group; an alkoxy group; an alkoxyalkyl group; a hydroxyalkyl group or an aryl group. In a sub-embodiment, R6 is ethyl, CH2CH2OH or CH2-phenyl. In another embodiment, the trichostatin compounds provided herein have the following structure: 35 200836749

Wherein R7 is hydrazine, halogen, alkyl (which includes lower alkyl), alkenyl, alkynyl, alkoxy, alkoxyalkyl, hydroxyalkyl, cycloalkyl, nitro, cyano, OH, OR4 , NH2, NHR4, NR4R4, SH, SR4, CF3, CH2OH, 5 CH2F, CH2Cn, CH2CF3, C(Y3)3, C(Y3)2C(Y3)3, C(=0)0H, C(=0) 0R4, C(=0)_alkyl, C(=0)_aryl, C(=0)·alkoxyalkyl, C(=0)NH2, C(=0)NHR4, C(=0) N(R4)2 or N3, wherein each Y3 is independently H or halogen; and each R4 is independently oxime; fluorenyl, which includes lower fluorenyl; alkyl, including 10 lower alkyl such as, but not limited to: Methyl, ethyl, propyl and cyclopropyl; alkenyl; alkynyl; cycloalkyl; alkoxy; alkoxyalkyl; hydroxyalkyl. In a sub-embodiment, r7 is methyl, ethyl, phenyl, chloro or νη2. In another embodiment, the trichostatin compounds provided herein have the following structure: 36 200836749

CH

In another embodiment, the trichostatin compounds provided herein have the following structure:

As used herein and unless otherwise specified, the term "bortezomib and its derivatives" refers to a compound of formula V: 37 200836749

among them

Ri, R!, R3, R4 and R5 are each independently oxime, optionally halogenated, substituted linear, branched or cyclic alkyl (which includes lower alkyl), alkoxy, alkenyl or alkyne Base, aryl, CO-alkyl, CO-alkenyl, CO-alkynyl, c〇-aryl or heteroaryl, CO-alkoxyalkyl, CO-aryloxyalkyl, CO-substituted aryl , sulfonyl, alkanesulfonyl, arylsulfonyl, aralkyl sulfonyl. In another embodiment, the bortezomib and its derivatives have the structure of the following formula VI:

10

Ri, Ri, R3, R4 and R5 are each independently oxime, optionally halogenated, substituted linear, branched or cyclic alkyl (which includes lower alkyl), alkoxy, alkenyl or alkynyl , aryl, CO-alkyl, CO-alkenyl, CO-alkynyl, CO- 38 200836749 aryl or heteroaryl, CO-alkoxyalkyl, CO-aryloxyalkyl, CO-substituted aryl , continued sulfhydryl, sulphur-burning base, aromatic Si-based, aromatic sulfonyl sulfhydryl. In another embodiment, the bortezomib and its derivatives have the structure of the following formula VI:

It will be appreciated that the compounds disclosed herein may contain a palm center. Such palm center may have a (R) or (S) configuration or may be a mixture thereof. Thus, the compounds provided herein may be mirror image-isomerically pure or may be a mixture of stereoisomerism or diastereoisomers. It is to be understood that the disclosure of the compounds herein encompasses racemic, optically active, polymorphic or stereoisomeric forms or mixtures thereof, preferably having the useful properties described herein, and methods for preparing optically active forms and Methods for determining activity using standard assays described herein or using other similar assays that are well known in the art are well known in the art. Examples of methods which can be used to obtain the optical isomers of these compounds include the following: i) Physical separation of crystals: A technique for manually separating macroscopic crystals of individual mirror image isomers. If the crystals of individual mirror image isomers exist, that is, the substance is a crystal group and the crystals are visually different, ii) simultaneous crystallization: only when the solution of the racemate is a solid crystal group, 20 Techniques for crystallizing individual mirror image isomers from the solution; 39 200836749 m) Enzymatic resolution method • Partial or complete separation of the racemate by using enzymes for different rates of reaction of the mirror image isomers technology. Enzymatic asymmetric synthesis method. At least one step of the synthesis method is an synthesis technique using an enzymatic reaction to obtain a mirror-isomerized pure or enriched 5-component precursor of a desired mirror image isomer; The chemical non-synthesis method produces asymmetry (that is, the palmity) of the oil τ, and synthesizes the desired mirror image isomer from the non-pseudo-precursor precursor, which can use the palm touch The medium or the palmarity aid; (7) the material separation method, the racemic compound is reacted with the mirror image isolating agent (the pair of palm gracils) to convert the individual mirror image isomers into non-correspondence The technology of isomers. However, the more prominent structures of the present invention are different by chromatography or crystallization to separate the diastereomers formed, and the pair of palmitic adjuvants are later removed to obtain the desired mirror structure. · 15 V11) second and second order asymmetry transformation: a diastereomer that balances the self-raceomer to the diastereomers from the desired smectomer The preferential crystallization of the non-sub-image in the solution, which is predominant or from the desired mirror image, will disturb the equilibrium so that, in principle, all of the material is converted from crystalline to non-mirrible. Isomer. And then the technique of releasing the desired mirror image from the diastereomer; (4) kinetic resolution: the technique refers to the use of the mirror isomers and the palmar, non-external or Partial or complete resolution or further resolution of part of the racemate by the reaction of the catalyst; 40 Part of the compound that was resolved in 200836749; ix) Mirror specific synthesis from a non-racemic precursor: The synthesis technique of the non-pivoted starting material to obtain the desired mirror image isomer, and its stereochemical integrity is not impaired or minimally damaged during the synthesis process; - 5 x) palm liquid chromatography: A technique for separating such mirror image isomers of a racemate in a liquid mobile phase by the interaction of a mirror image isomer with a stationary phase. The stationary phase may be made of a palm material or the mobile phase may contain another pair of palms that can cause different interactions; xi) for the palm gas layer method, volatilizing the racemate and borrowing the mirror isomer a technique for separating the mirror image isomers in a gaseous 10 mobile phase with a different interaction with a column containing a fixed non-racemic palmitic absorber phase; xii) using a palmitic solvent extraction method, A technique in which the mirror image isomer is preferentially dissolved in a specific palm solvent to separate the mirror image isomers; xiii) a technique in which the racemate is attached to the membrane barrier via a method of carrying it on a palm film. The barrier typically separates the two miscible fluids (where one contains the racemate) and the driving force, such as concentration or pressure differential, can result in preferential transport via the membrane barrier. In some embodiments, the system provides trizidine, triclinide squanate (TCN-P), triclinbine 5,-phosphate (TCN_P), triclinib 5, _phosphoric acid 20 Salt monohydrate (TCN_PM) or Quercetin DMF addition ^ (TCM-DMF). The TCN can be synthesized by any of the techniques known to those skilled in the art, for example, as described in Tetrahedron Letters, 49: 4757-4760 (197i), by any technique known to those skilled in the art, such as, for example, a U.S. patent. TcN_p was prepared by the method described in No. 4,123,524. 兮41 200836749 The synthesis method of TCN-DMF is described, for example, in INSERM, vol. 81: 37-82 (1978), which can be disclosed, for example, in the following materials. Other compounds related to TCN as described herein are synthesized by methods: Gudmundsson et al., 2001, Nucleosides Nucleotides Nucleic 5 Acids 20: 1823-1830; Porcari et al., 2000, J Med Chem 43: 2457-2463; Porcari et al. Human, 1999, Nucleosides Nucleotides, 18: 2475-2497; Porcari et al., 2000, J Med Chem, 43: 2438-2448; Porcari et al., 2003, Nucleosides Nucleotides Nucleic Acids, 22: 2171-2193; Porcari et al. 2004, 10 Nucleosides Nucleotides Nucleic, Acids, 23:31-39; Schweinsberg et al., 1981, Biochem Pharmacol 30: 2521-2526; Smith et al., 2004, Bioorg Med Chem Lett 14: 3517-3520; Townsend et al. 1986, Nucleic Acids Symp Ser 1986: 41-44; and/or Wotring et al., 1986, Cancer Treat Rep 70: 491-7. 15 pharmaceutically acceptable salts and prodrugs are sufficient for the formation of a stable or acidic compound. In the case of a toxic acid or a salt, the compound is preferably administered as a pharmaceutically acceptable salt. The pharmaceutically acceptable salts include those derived from pharmaceutically acceptable inorganic or organic bases and acids. Suitable salts include those derived from The pharmacy is well known to us as the metal of many acids, such as the clock and sodium; and soil testing metals, such as dance and town. In more detail, examples of pharmaceutically acceptable salts are organic using acid. An acid addition salt which forms a physiologically acceptable anion such as a sulphate, mesylate, acetate, citrate, malonate, tartrate, succinate, benzoate, ascorbate, alpha - ketoglutarate, 42 200836749 and alpha-glycerol deacidification. Suitable inorganic salts can also be formed, including sulfates, sulfites, bicarbonates, and carbonates. A standard procedure well known in the art can be used to obtain a forkable salt on a vehicle, for example, by reacting a sufficient compound, such as an amine, with five suitable acids to obtain a physiologically acceptable Anion. It is also possible to prepare a metal for carboxylic acid (for example, sodium, potassium or lithium) or an alkaline earth metal (for example, calcium). Any of the nucleotic acids described herein may be added with a nucleotate prodrug to increase the activity, bioavailability, stability or altered properties of the sorbic acid. Many nucleotide prodrugs are known for their coordination systems. In general, the alkylation, deuteration or other lipophilic modification of the 10 mono-, di- or triphosphate of the nucleotide increases the stability of the acid; acid stability. Examples of substituents which may replace one or more hydrogens on the phosphate molecular group are alkyl groups, aryl groups, steroids, carbohydrates (including sugars), dimercaptoglycerols and alcohols. A number of such substituents are described in R. Jones and ν Bischofberger, Antiviral 15 Research, 27 (1995) 1-17. Either of these can be used in combination with the disclosed nucleoside acid to obtain the desired effect. In one embodiment, the triclinide or related compound is provided as a 5,-hydroxy lipophilic prodrug. Non-limiting examples of U.S. patents which disclose covalently incorporation into the nucleotide, preferably at the 5,-0H position of the nucleotide or lipophilic formulation, include U.S. Patent No. 5,149,794 (issued to Yatvin et al. on September 22, 1992); 5,194,654 (issued to Hostetler et al., March 16, 1993); 5,223,263 (issued to Hostetler, etc., June 29, 1993) Person); No. 5,256,641 (issued to Yatvin et al. on October 26, 1993); No. 5,411,947 (issued to Hostetler et al. on May 2, 1995); 43 200836749 No. 5,463,092 (October 31, 1995) To Hostetler et al.); 5,543,389 (Yatvin et al., August 6, 1996); 5,543,390 (Attavin et al., August 6, 1996); 5,543,391 (August 6, 1996) To Yatvin et al.; and 5,554,728 (issued to 5 Basava et al. on September 1, 1996), all of which are incorporated herein by reference. The disclosure of a lipophilic substituent or a lipophilic preparation which can be linked to the tromethine or related compound of the present invention includes WO 89/02733, WO 90/00555, WO 91/16920, WO 91/18914, WO 93/00910, WO 94/26273, WO/15132, EPO 350 287, 10 EP 93917054.4, and WO 91/19721. Further non-limiting examples of derivatives of tricineribine or related compounds are derivatives containing substituents as described in the following publications. These derivatized triclinide or related compounds can be used in the conditions described herein or as an antiviral agent, including anti-HIV or anti-HBV agents. The publications are as follows: 15 Ho, 1973, Cancer Res. 33, 2816-2820; Holy, 1993 Isopolar phosphorous-modified nucleotide analogues, in: De Clercq (ed.)? Advances in Antiviral Drug Design, Vol. I5 JAI Press , pp. 179-231; Hong et al., 1976, Blochem Biophys Rs Commun 88: 1223-1229; Hong et al 5 1980? J Med Chem 20 28: 171-177; Hostetler et al.? 1990, Biol Chem 266: 11714 -11717, Hostetler et al., 1994, Antiviral Res 24:59-67; Hostetler et al., 1994, Antimicrobial Agents Chemother 38: 2792-2797; Hunston et al., 1984, J Med Chem 27:440-444 ; Ji et al., 1990, J Med Chem 33: 2264-2270; Jones et al. 5 1984, J Chem Soc Perkin 44 200836749

Trans 1:1471-1474; Juodka et al., 1974, Coll Czech Chem Comm 39:363-968; Kataoka et al., 1989, Nucleic Acids Res Sym Ser 21:1-2; Kataoka et al.? 1991, Heterocycles 32: 1351-1356; Kinchington et al., 1992, Antiviral Chem Chemother 3: 107-112;

5 Kodama et al., 1989, Jpn J Cancer Res 80: 679-685; Korty et al, 1979, Naunyn-Schmiedeberg 5s Arch Pharmacol 310: 103-111; Kumar et al., 1990, J Med Chem 33: 2368- 2375; LeBec et al., 1991, Tetrahedron Lett 32: 6553-6556; Lichtenstein et al., 1960, Biol Chem 235: 457-465; Lucthy et al., 1981, Mitt Geg, 10 Lebensmittelunters Hyg 72: 131-133 (Chem. Abstr. 95, 127093);

McGuigan et al., 1989, Nucleic Acids Res 17: 6065-6075; McGuigan et al, 1990, Antiviral Chem Chemother 1:107-113; McGuigan et al 5 1990, Antiviral Chem Chemother 1:355-360; McGuigan et al. 5 1990, Antiviral Chem Chemother 1:25-33; 15 McGuigan et al. 5 1991? Antiviral Res 15:255-263; McGuigan et al., 1992, Antiviral Res 17:311-321; McGuigan et al., 1993, Antiviral Chem Chemother 4: 97-101; McGuigan et al. 9 1993, J Med Chem 36: 1048-1052. The hydrogen phosphate alkylate derivative of the anti-HIV agent AZT is less toxic than the 20 parent nucleotide analog. Antiviral Chem Chemother 5: 271-277; Meyer et al, 1973, Tetrahearon Lett 269-272; Nagyvary et al, 1973, Biochern Biophys Res Commun 55: 1072-1077; Namane et al.? 1992, J Med Chem 35 :3939-3044; Nargeot et al., 1983, Proc· Natl. Acad. Sci. USA 80:2395-2399; 45 200836749

Nelson et al., 1987, J Am Chem Soc 109: 4058-4064; Nerbonne, et al., 1984, Nature 301: 74-76; Neumann et al, 1989, J Am Chem Soc 111: 4270_4277; Ohno et al ·, 1991, Oncology 48:451-455. Palomino et al. 5 1989, J Med Chem 5 32:622-625; Perkins et al.? 1993, Antiviral Res 20 (Supp. I): 84; Piantadosi et al· , 1991, J Med Chem 34: 1408-1414; Pompon et al" 1994, Antiviral Chem Chemother 5: 91-98; Postemark, 1974, Anu Rev Pharmacol 14: 23-33; Prisbe et al" 1986, J Med Chem 29 :671-675; Pucch et al., 1993, Antiviral 10 Res 22: 155-174; Pugaeva et al., 1969, Gig Trf Prof Zabol 13: 47-48 (Chem. Abstr. 72, 212); Robins, 1984 , Robins, 1984, Pharm Res 11-18; Rosowsky et al., 1982, J Med Chem 25: 171-178; Ross, 1961, Biochem Pharm 8: 235-240; Ryu et al., 1982, J Med Chem 25 :1322-1329; Safthill et al., 1986, 15 Chem Biol Interact 57:347-355; Saneyoshi et al., 1980, Chem Pharm Bull 28:2915-2923; Sastry et al., 1992, Mol Pharmacol 41:441 -445; Shaw et al.? 1994, 9th Annual AA PS Meeting, San Diego, CA (Abstract); Shuto et al., 1987, Tetrahedron Lett 28: 199-202; Shuto et al. 5 1988, Chem 2 〇 Phann Bull 36: 209-217. A preferred phosphate prodrug group is S-mercapto-2-thioethyl, which is also known as "SATE". Additional Examples of Prodrugs Can Be Used The following patents and patent applications are described in the following patents and patent applications: U.S. Patent Nos. 5,614,548, 5,512,671, 5,770,584, 5,962,437, 5,223,263, 5,817,638 46 200836749, 6,252,060 , Nos. 6,448,392, 5,411,947, 5,744,592, 5,484,809, 5,827,831, 5,696,277, 6,022,029, 5,780,617, 5,194,654, 5,463,092, 5,744,461, 4,444,766, 4,562,179 5, 5,599,205, 4,493,832, 4,221,732, 5,116,992, 6,429,227, 5,149,794, 5,703,063, 5,888,990, 4,810,697 No. 5,512,671, 6,030,960, 2004/0259845, 6,670,341, 2004/0161398, 2002/082242, 5,512,671, 10 2002/0082242, and or PCT publication WO 90/11079, WO 96/39197, and/or WO 93/08807. In vivo efficacy/dosing regimen In another aspect of the invention, there is provided a dosing regimen that limits the toxic side effects of TCN and related compounds. In one embodiment, such administrations 15 may minimize toxic side effects including, but not limited to, hepatotoxicity, thrombocytopenia, hyperglycemia, vomiting, hypocalcemia, Anemia, hypoalbuminemia, inhibition of gastric myeloid activity, hypertriglyceridemia, high amylase whiteness, diarrhea, osteitis and/or fever. In another embodiment, the administration of TCN, TCN-P or related compounds with borte 20 zomid and its derivatives can provide at least partial or complete in vivo response in at least 15 to 20% of patients, in a particular implementation In one embodiment, a portion of the response can be at least 15, 20, 25, 30, 35, 40, 50, 55, 60, 65, 70, 75, 80 or 85% regression of the tumor. In other embodiments, in at least 15, 20, 25, 30, 35, 40, 50, 55, 60, 65, 7〇, 75, 80, 85 47 200836749 or 90% of patients treated with the treatment The reaction was found. In other embodiments, such response rates can be obtained by any of the treatment regimens disclosed herein. In other embodiments, a method of treating a condition diagnosed as having cancer is administered by administering to the patient an effective amount of 5 TCN, TCN-P or related compound and zosami according to a dosing schedule. a derivative thereof, which comprises administering the triclinide compound and/or rototizomib or a salt or a derivative thereof once a week for 3 weeks, and then not administering the drug within a week period (also That is, via the 28-day cycle). In other embodiments, such a 28 day period can be repeated for at least 2, 3, 4, or 5 times or until the tumor is clearly found to resolve 10 . In a further embodiment, a 42-day cycle is provided, the compound disclosed in the Chinese may be administered once a week for 4 weeks, and then the tromethine compound and/or frotezomib is not administered during 2 weeks. Its derivatives. In other embodiments, such a 42-day cycle can be repeated for at least 2, 3, 4 or 5 or until until the tumor is clearly found to subside. In a particular embodiment, less than 50, less than 25, or less than 10 mg/m2 of TCN, TCN-P, TCN-PM, or related compounds can be administered according to a 42 day period. In other specific embodiments, 2, 3, 4, 5, 6, 7, 8, 9, 1 or 11 mg/m2 of TCN, TCN-P, TCN-PM or may be administered according to a 42 day period. Related compounds. In another specific embodiment, bortezomib or a derivative thereof is administered at a dose of from about 1 mg/m 2 to about 50 mg/m 2 . In a particular embodiment, shedzomib or a salt thereof can be administered at 0.1, 0.5, 1, 5, 1 〇, 15, 20, 25, 30, 35 or 40 mm/m2 depending on the 42 day period. In another embodiment, there is provided a method of treating cancer in a patient, which is administered to the patient at a dose of 10 mg/m2 or less of TCN, TCN-P, TCN-PM or related compound once a week. And a dosing regimen of less than about 30 mg of stilzomib and its derivatives. In a particular embodiment, 0.5, 1, 1.5, 2, 2.5, 3, 3.5, 4, 4.5, 5, 5.5, 6, 6.5, 7, 5, 7.5, 8, 8.5, 9, 9.5 can be administered once a week. Or 10 mg/m 2 of TCN, TCN-P, TCN-PM or related compounds as disclosed herein. In another specific embodiment, 0.1, 0.5, 1, 5, 10, 15, 20, 25, 30, 35 or 40 mg/m2 of bortezomib or a derivative thereof can be administered once a week. In an embodiment of the invention, the compounds disclosed herein may be administered simultaneously in a single large dose over a short period of time, for example about 5, 10, 10 15, 20, 30 or 60 minutes. In a further embodiment, a dosage regimen wherein the compounds are administered by continuous infusion over a period of at least 24, 48, 72, 96 or 120 hours is provided. In a particular embodiment, the administration of the tricineridine compound and/or bortezide 15 m and its derivatives can be repeated by repeated or bulk injections at a specific frequency as follows: at least once a week, at least once every two weeks. Once, every 3 weeks at least once, at least once every month, at least once every 5 weeks, at least once every 6 weeks, at least once every 8 weeks, at least once every 10 weeks, and/or at least once every 12 weeks. The type and frequency of administration can be combined to create a dosing cycle using any of the methods disclosed herein. The spirulina compound and/or bortezomib and its derivatives may be repeatedly administered by a specific administration cycle, for example, by a large injection every two weeks, which takes 3 weeks. The dosing cycle can be administered for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 18 or 24 months. Alternatively, the patient can be administered at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 15 or 20 dosing cycles. The triclinide compound and/or rototizomib and its derivatives may be administered according to any of the combinations of 49 200836749 disclosed herein. For example, the tricineridine compound may be administered once a week, every 3 weeks. Or stilbene and its derivatives for a total of 3 cycles. In further embodiments, the compounds may be administered separately daily, at least 2, 3, 4, 5, 6, 7, 8, 9, or 10 days. After such administration, the western compound and/or ketotiz and its derivatives were not administered in the same _. TCN, TCN-P, TCN-PM and related compounds and derivatives thereof are disclosed in the literature, and the dosage thereof can effectively cause tumor regression. 10 The administration of the TCN, TCN-P, TCN-PM or related compounds with bortezomib and its derivatives may be at least partially, such as at least 15, 20 or 30% or completely, in at least 15 to 20% of such patients. In vivo reaction. In a particular embodiment, the patient can be administered at least 2, 5, 10, 15, 2, 3, or 5 Torr. In a particular embodiment, 15 may be administered to the patient at least about 〇·5,; 1, 15, 2, 2 5, 3, 3 5, 4, 4·5, 5, 5.5, 6, 6.5, 7, 7.5 , 8, 8, 5, 9, 9, 5, 10, 12, 15, , 7, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, TCN, TCN-P, TCN-PM or related 20 compounds disclosed in the text 90, 95, 100, 150, 165, 175, 200, 250, 300 or 35 mg/m2. In a particular embodiment, the patient can be administered 5, 10, 15, 20, 25, 30, 35 or 40 mg/m2 of bortezomib. Administration of the compound can be carried out according to the treatment regimen disclosed herein. In a particular embodiment, the dosing regimen comprises administering less than about 20 mg/m2 of TCN and related compounds to less than about 50 200836749 30 mg of bortezomib simultaneously, continuously or over a period of time. In one embodiment, tcn or related compounds of less than 20 mg/m2 and stonepentezomib of less than about 1 mg/m2 may be administered simultaneously once a week. In another embodiment, TCN or related compounds of less than 2 mg/m 2 may be administered once a week and less than about 3 mg of bromide 5 zomi may be administered in the next week. In further embodiments, the patient may be administered 2 mg/m 2, 5 mg/m 2, 103⁄4 g/m 2, and/or 15 mg/m 2 of TCN or related compound and less than about 30 ' 25, 20 , 15, 10, 5,; 1, 〇·5 or 〇·1 mg/m 2 of bortezomib or a salt or derivative thereof. In another embodiment, the patient 10 can be administered less than 10 g/m2 of tripellilide compound and less than about 30 mg/m2 of bortezomib via continuous infusion for a period of at least 5 days. The invention provides any combination of the types of administration, frequency, number of cycles and dosages disclosed herein. Screening of the I Population In another aspect of the invention, methods are provided for identifying cancers or tumors that are susceptible to the toxic effects of Tricinem 15 (TCN) and related compounds. In an embodiment, there is provided a method of treating a cancerous 3 Ge tumor in a mammal by (1) obtaining a biological sample from the tumor; (9) determining whether the cancer or tumor overexpresses Akt kinase or a high activation and acidification Ahsaki; and (d) treatment of the cancer or tumor using tricineribine or a related compound as described herein. In an embodiment, the biological sample can be a biopsy. In the embodiment of the basin, the biological sample can be a fluid, a cell, and/or aspirate obtained from the tumor or cancer. The biological sample can be obtained according to any technique known to those skilled in the art. In one embodiment, the biopsy can be taken to obtain a biological sample. 51 200836749 5 10 20 A procedure in which a living slice removes tissue or cells from the body for testing. Some live cuts can be performed in the physician's house, while other live slices need to be done in the west. In addition, some biopsies require an anesthetic to paralyze this area a, while other living sections do not require any sedation. Endoscopic biopsies can be performed in a particular embodiment. Such biopsies are passed through a natural body hole (ie, a rectum) or a small incision (ie, an arthroscope) through a fiber optic endoscope (long, thin tube suitable for viewing at the end of the eye with the closest focus). get on. The endoscope is used to view an abnormal or suspicious area of the organ to obtain a small amount of tissue suitable for the study. Endoscopy procedures are used for organs or body areas that are to be developed and/or treated. The physician can insert the endoscope into the uterine intestine (endoscopic gastrointestinal examination), bladder (cytoscopic examination), abdominal cavity (abdominal Wei examination), _ cavity _ examination), scarf f« position (longitudinal g examination ) or ^ and the stagnation system (laryngeoscopy and bronchoscopy). In another example, bone marrow biopsy can be performed. This living body can be itched from the sternum: _m) or the sacral bone (in any head region of the pelvis on the lower back region). Insert a long hard needle into the person; Γ and provide a local anesthetic to this occasion, which is uncomfortable/internal, and the remaining cytoplasm is studied; sliced (from the _ petal (1) trousers riding the core of the living body of the core":: In the case of resection or incision of the mammal ==::: When the skin part is wider or deeper, usually this step is checked and sutured _=* knife) to remove the full thickness of the skin for tumor progression When the pot is called cut ~ * sutured with a line of wounds. When removing the entire swollen it is privately a resectable biopsy technique. If only the part of the tumor 52 200836749 is removed, it is called incision In vivo biopsy techniques. For example, when melanoma (a type of skin cancer) is suspected, resected biopsy is usually a method that is often preferred. In still other embodiments, fine needle aspiration (FNA) biopsy can be used* 5 This biopsy involves the use of fine needles to remove small pieces from the tumor. Sometimes a local anesthetic is used to itch the area, but the test rarely causes discomfort and leaves no sputum. FNA does not apply, such as suspicious Diagnosis of pigmented nevus, instrument | bit is available, for example A biopsy is close to the large lymph nodes of the melanoma to see if the melanoma has metastasized (diffusion). A computerized tomography scan (CT or CAT scan) can be used to introduce the needle into a tumor in an internal organ such as the lung or liver. In other embodiments, drilling, shaving, and/or biopsy of the skin may be performed. Drilling biopsy includes removal using a biopsy instrument using a short cylinder of removable tissue or a "core" core. Deep skin sample. After administration of local anesthetic 15 drench, the instrument is rotated on the surface of the skin until it passes through all layers (including the dermis, epidermis, and the most superficial part of the subcutaneous tissue (fat)). A shaved biopsy includes removing the layer by shaving off the top layer of the skin. A local anesthetic is also used for shaving biopsy. Skin biopsy involves removing the skin sample for examination under a microscope to determine, for example, 20 Whether melanoma is present. The biopsy is performed under local anesthesia. In a particular embodiment, a method is provided to determine whether the tumor overexpresses Akt The overexpression of Akt kinase may indicate the state of this acidification. The hyperphosphorylation of Akt can be detected according to the methods described herein. In one embodiment, tumor biopsies and control tissues can be compared. The control tissue may be a normal tissue obtained from a mammal obtaining the biopsied or a normal tissue obtained from a healthy mammal. Akt excitatory hyper-expressive or highly carbonated may be assayed to understand the Akt of the tumor in vivo. Whether the content and/or Akt kinase filling is higher than the control tissue, such as Akt 5 kinase, which is at least about 1.5, 2, 2, 25, 2.5, 2.75, 3, 3.25 higher than the Akt kinase content in the control tissue. 3.5, 3.75, 4, 4.25, 4.5, 4.75, 5, 5.5, 6, 7, 8, 9, or 10 times. In one embodiment, the invention provides a method of detecting abnormal Akt kinase expression in a patient or a biological sample obtained from the patient, which is immunized with 10 specific antibodies against Akt kinase or an antigenic portion thereof. The interacting molecule contacts cells, cell extracts, serum or other samples or biological samples obtained from such patients and screens for immunocompetent molecule-A kt kinase complex formation, wherein the relative cellular cells The high presence of the complex indicates abnormal cells that can express or overexpress Akt. In one embodiment, the amount of high Akt kinase present in cells or cell extracts can be screened. In another embodiment, the degree of expression of the gene encoded by Akt kinase can be screened to detect abnormal expression of Akt in the cell at the gene level, wherein the transcriptional expression product (ie, mRNA) is compared with normal cells. High levels indicate abnormal cells. In a particular embodiment, real-time PCR and its 20 pCR program can be used to determine transcriptional activity. In one embodiment, the mRNA can be obtained from a patient's cell or a biological sample obtained from the patient and can selectively produce cDNA. The mRNA or cDNA can then be contacted with a gene probe capable of hybridizing and/or amplifying all or part of the nucleotide sequence encoding the Akt kinase or its complementary nucleotide sequence, and then detecting the mRNA or cDNA content, 54 200836749 The presence of high levels of mRNA or cDNA compared to the normal control group can be assessed. Still another embodiment of the present invention encompasses the use of antibodies (single or multiple strains) of Akt kinase in a quantitative or semi-quantitative diagnostic kit to determine the relative amount of Akt kinase in a patient's suspected cancer cell, the kit All reagents required to perform the assay can be included. In one embodiment, a kit for using the reagents and materials required for performing an ELISA assay is provided. The reagent may include, for example, a buffer for washing, an antibody dilution buffer, a blocking buffer, a cell staining solution, a developing solution, a stop solution, an acid-protein specific antibody, an anti-Pan protein 10-specific antibody, and two Secondary antibodies, and distilled water. The kit may also include instructions for use and may be selectively automated or semi-automated or in a form that is compatible with automated machinery or software. In one embodiment, a phospho-serine-473Akt antibody capable of detecting an activated form of AKT (phosphorylated Akt of serine 474) can be used as an antibody in a diagnostic kit. See, for example, Yuan et al. 15 (2000) uFrequent Activation of AKT2 and induction of apoptosis by inhibition of phosphinositide-3-OH kinase/Akt pathway in human ovarian cancer,’’ Oncogene 19:2324-2330 o

Akt kinase

Akt, also known as PKB3' represents a subfamily of serine/threonine kinases. 20 Three members have been identified in this subfamily, AKT1, AKT2, and AKT3. In the PI3K-dependent manner, Akt is activated by extracellular stimuli (Datta, S. R., et al. Genes Dev 13: 2905-2927, 1999). Complete activation of Akt requires Thr. phosphorylation in the activation loop and selenate reaction of Ser473 in the C-terminal activation domain. Akt is a PTEN tumor suppressor without 55 200836749

Adjusted to advantage. Mutations within PTEN have been identified in various tumors that can lead to activation of the Akt pathway (Datta, S. R., et al. Genes Dev. / 3: 2905-2927, 1999). In addition, amplification, overexpression and/or activation of Akt has been detected in many human malignancies (Datta, SR, et al. Genes Dev. 5 /3: 2905-2927, 1999, Cheng, J. Q., And Nicosia, SV AKT signal transduction pathway in oncogenesis. In Schwab D, editor. Encyclopedic Reference of Cancer. Berlin Heidelberg and New York: Springer; 2001. pp35_7). Akt, particularly ectopic expression of the apoplastic Akt, induces cell survival and malignant transformation, whereas inhibition of Akt 10 activity stimulates apoptosis in a range of mammalian cells (Datta, SR, et al. Genes Dev · /3:2905-2927, 1999, Cheng, JQ? and Nicosia, SV AKT signal transduction pathway in oncogenesis. In Schwab D? editor. Encyclopedic Reference of Cancer. Berlin Heidelberg and New York: Springer; 2001. 15 PP35-7 , Sun, M., et al. Am J. Path, 159: 431-437, 2001,

Cheng, J.Q., et al. Oncogene, 14: 2793-2801, 1997). Moreover, activation of Akt has been shown to be associated with tumor invasiveness and chemical resistance (West, K.A., et al. Drug Resist· Updat., 5: 234-248, 2002). Activation of this Akt pathway plays a key role in malignant transformation and chemical resistance by inducing cell remnant, growth, migration, and angiogenesis. The present invention provides methods for determining the overexpression and/or high activation of Akt kinase and the amount of Akt kinase. The Akt kinase can be any known Akt kinase or a related kinase thereof including, but not limited to, Aktl, Akt2, Akt3. The mRNA and amino acid sequences of human Aktl, Akt2, 56 200836749 and Akt3 are elucidated in Figures 6a-c, > 7a-d, and 8a-c, respectively. Cockpit assay immunological assay 5 In one embodiment, a method for detecting Akt-excited cells in a mammalian animal or in a biological sample obtained from the mammal is provided, It is obtained by contacting cells, cell extracts or serum or other samples or biological samples obtained from the mammal with immunologically interacting molecules specific for Akt kinase or its antigenic portion, and screening for immunological interactions. The extent to which the 10 molecule-Akt kinase complex is formed is then determined whether the complex is present in large amounts relative to normal cells. The immunologically interacting molecule may be a molecule having specificity and binding affinity for Akt kinase or an antigenic portion thereof or a homolog thereof or a derivative thereof. In one embodiment, the immunointeractive molecule can be an immunoglobulin 15 protein molecule. In other embodiments, the immunoglobulin molecules can be antibody fragments, single chain antibodies, and/or deimmunized molecules, including humanized antibodies and T cell associated antigen binding molecules (TABMs). In a specific embodiment, the antibody can be a monoclonal antibody. In another specific embodiment, the antibody can be a plurality of antibodies. The immunologically interacting molecule is specific for Akt kinase 20 or, more specifically, an epitope or epitope on Akt kinase. An epitope or epitope on an Akt kinase includes a portion of the molecule that is guided by an immune response. The antigenic epitope or epitope antigen may be a B cell epitope antigen or, if appropriate, a T cell epitope antigen. In the embodiment, the present antibody is a lin-serine 473 Akt antibody. 57 200836749 兔明之-Example provides a method for diagnosing mammalian growth or cancer-like growth, in which there is abnormal gamma in the genus, and using A kt spike binding effective amount of the A kt kinase or The antibody specific to the body or the surface antigen is contacted with the biological sample obtained from the donor cell extract or from the patient, and then the amount of the kinase-antibody complex is determined or determined. Among them is the determination of the presence of this southern content complex with normal cells =.乂 10 Any of a number of methods known to those skilled in the art can be utilized. For example, in the detection of human Akt kinase, antibodies usually do not = two from: human: animals, such as primates, livestock animals (such as Mian Ping cattle, pigs, Shanping, horses), laboratory test animals (such as mouse guinea pigs) , rabbits, and/or accompanying animals (eg, dogs, antibodies can also be: recombinantly produced in prokaryotic or eukaryotic host cells, and can be used in vitro on cells or tissue biopsies. Mainly, if the antibody is appropriately deimmunized or used for human use, the antibody can be labeled, for example, by a nuclear tag, and the patient is vaccinated by a thief and the nucleus is accumulating. The antibody can be a 20 f ticket. Thus, the 'inventive invention' provides such deimmunized forms suitable for imaging cancer in humans or non-type I patients, in the case of antibody-producing 歳 kinases, The anti-system shot method is attached to the autonomic material (which includes human tissue) or from the cell Μ5. The enzyme can be isolated from the organism by any suitable method. For example, the separation method can be Use the following Ak Any one or more of the properties of t kinase: surface charge properties, size, density, and its affinity for another entity (eg, another protein shell or chemical compound to which it is associated or associated). Thus, for example, Akt kinase can be isolated from the biological fluid by any one or more of the following methods: ultracentrifugation, ion exchange chromatography (eg, anion exchange chromatography, cation exchange chromatography), electrophoresis 5 (eg, poly Acrylamide gel electrophoresis, isoelectric focusing method), size separation method (such as gel filtration method, ultrafiltration method) and affinity media separation method (such as immunoaffinity separation method, including, but not limited to, magnetic Bead separation method (such as Dynabead (trademark) separation method, immunochromatography, immunoprecipitation method). The method for isolating Akt kinase from the biological fluid can maintain the type 10 epitope antigen present on the kinase and, therefore, is best Avoid using techniques that can cause denaturation of the enzyme. In another embodiment, any one or more of affinity separation, gel filtration, and/or ultrafiltration can be used. Fluid separation kinases. Standard procedures known in the art can be used, for example, by Kohler and Milstein (Kohler and Milstein, Nature 256: 495-499, 1975; 15 Kohler and Milstein, Eur J. Immunol. 6(7) :511-519, 1976), Coligan et al. ("Current Protocols in Immunology, John Wiley & Sons, Inc" 1991-1997) or Toyama et al. (Monoclonal Antibody, Experiment Manual", published by Kodansha Scientific, 1987) The procedure described is followed by an immunological reaction and subsequent preparation of a monoclonal antibody 20 per plant. Basically, animals are immunized by Akt kinase-containing biological fluids or parts thereof or recombinant Akt kinases by standard methods to produce cells capable of producing antibodies, particularly somatic cells (eg, B lymphocytes) capable of producing antibodies. . These cells can then be removed from the immunized animal to immunize the cells. In certain embodiments, fragments of Akt kinase can be used to generate anti-59 200836749 bodies. This fragment can be associated with a carrier. The vector may be any material that is typically high molecular weight, either non- or low immunogenic (e. g., hapten), which is naturally or artificially linked to enhance its immunogenicity. The immortalization of cells capable of producing 5 antibodies can be carried out using methods well known in the art. For example, the cell immortalization reaction can be carried out by the transformation method using Epstein-Barr virus (EB V) (Kozbor et al., Methods in Enzymology 121: 140, 1986). In another embodiment, a cell fusion method (described in Coligan et al., 1991-1997, which is widely used in the manufacture of monoclonal antibodies) is used to immunize the antibody-producing cells. In the present method, an antibody-producing somatic cell having a potential to produce an antibody, particularly a B cell, is fused by a myeloma cell line. These somatic cells may be derived from the animal that has been exposed to the antigen, preferably the lymph nodes, spleen and peripheral blood of carious animals such as mice and rats. In a specific embodiment, mouse spleen cells can be used. In other embodiments, rat, free, 15 sheep or goat cells can also be used. A fusion procedure suitable for the production of hybridomas has been established (Kohler and Milstein, 1976, supra; Shulman et al, Nature 276: 269-270, 1978; Volk et al, J. Virol. 42(1): 220-227, 1982 A lymphoma tumor is developed for a specific myeloma cell line. Many myeloma cell lines can also be used to make fusion cell hybrids, including, for example, P3 multiplied by 20 63-Ag8, P3 multiplied by 63-AG8.653, P3/NSl-Ag4-l (CN-l),

Sp2/0-Agl4 and S194/5.XXO.BU.1. By Kohler and

These Sp3/0-Agl4 myeloma cell lines can be formed by multiplying P3 by 63-Ag8 and NS-1 cell lines as described by Milstein (1976, supra), Shulman et al. (1978, supra). The S194/5.XXO.BU.1 cell line is reported by Trowbridge 60 200836749 (J. Exp. Med. 148(1): 313-323, 1978). Methods for producing hybrids of spleen or lymph node cells and myeloma cells capable of producing antibodies typically include a ratio of 10:1 in the presence of one agent or group of agents (chemical, viral or electrical) that promotes cell membrane fusion (Although the ratio can vary from about 2: j 5 to about 1:1) mixed somatic and myeloma cells. Fusion methods are described (Kohler and Milstein, 1975, supra; Kohler and Milstein, 1976, supra; Gefter et al, Somatic Cell Genet. 3 231-236, 1977; Volk et al., 1982, supra). The fusion enhancers used by their researchers were Sendai virus and polyethylene glycol (PEG). In a particular embodiment, a method of selecting a fused cell hybrid is provided from the remaining unfused cells, particularly unfused myeloma cells. In general, the selection of the fused cell hybrid can be carried out by culturing the cells in a medium that maintains the growth of the hybridoma but prevents the growth of the unfused myeloma cells, which can generally continue to divide indefinitely. . Such somatic cells used for fusion 15 in in vitro cultures do not maintain long-term viability. It takes several weeks to selectively culture the fused cell hybrids. At the beginning of the period, it is necessary to identify such hybrids which produce the desired antibodies, whereby the hybrids can be subsequently colonized and propagated. In general, about 10% of the obtained hybrid can produce the desired antibody, but it is not uncommon from about 1 to about 30%. Hybrids capable of producing antibodies can be detected by any of several 20 standard assay methods, such as, for example, by Kennet et al. (Monoclonal Antibodies and Hybridomas: A New Dimension in Biological Analyses, pp 376-384, Plenum Enzyme-linked immunoassay and radioimmunoassay techniques described in Press, New Yark, 1980) and by FACS analysis (0, Reilly et al. 61 200836749, Biotechniques 25: 824-830, 1998). Once the desired fusion cell hybridization site has been selected and cloned in individual cell lines capable of producing antibodies, each cell line can be propagated by either of two standard methods. A suspension of such hybridoma cells can be injected into a tissue-receptive animal. The injected animal can then form a tumor that secretes a specific monoclonal antibody produced by the hybrid cell hybrid. Body fluids of the animal, such as serum or ascites fluid, can be released to provide a high concentration of monoclonal antibodies. Alternatively, each of the cell lines can be propagated in vitro in a laboratory culture vessel. A high concentration of a single specific 10 strain of early antibody can be collected by decantation, filtration or telecentric method and then purified. These cell lines can then be tested by any suitable immunoassay to appreciate the specificity of the relevant Akt kinase. For example, cell strains can be divided into several wells and cultured in a number of wells, and then analyzed by enzyme-linked immunosorbent assay (ELISA), indirect fluorescent antibody technique, and the like, and clarified from each well. The cell strain (group) producing a monoclonal antibody capable of recognizing the target LIM kinase but not recognizing the non-target epitope antigen can be identified and then directly cultured or injected into a histocompatible animal to form a tumor and be produced and collected. Purify the necessary antibodies. Accordingly, the present invention provides a method for detecting Akt kinase or a 20-segment, variant or derivative thereof in a sample, which comprises contacting the sample with an antibody or a fragment or derivative thereof and a normal control group (high content thereof) The Akt kinase is assayed for comparison to detect the content of the complex comprising the antibody and Akt kinase or a fragment, variant or derivative thereof, and any method capable of determining the formation of the complex can be used. For example, an antibody of the invention having a molecule associated with a reporter group can be used in an immunoassay. Such immunoassays include, but are limited to, radioimmunoassay (RIA), enzyme-linked immunosorbent assay (EUSa), immunohistochemistry (I), and Western blotting methods are well known to those skilled in the art. The immunization inspection method may also include a competitive verification method. The invention covers both qualitative and quantitative 5 immunoassays. Suitable immunoassay techniques are described, for example, in U.S. Patent Nos. 4,016,043, 4,424,297, and 4,018,653. These assays include both non-competitive and single-site and dual-site assays as well as traditional competitive assays. These assays also include the direct binding of the labeled antigen-binding molecule 10 to the antigen of interest. The present invention further provides a method for quantifying the degree of expression or activation of Akt protein in a cell or tissue sample of an animal, such as a human cancer patient or an individual suspected of having cancer. In one embodiment, the invention provides a method for quantitatively quantifying the degree of expression or activation of Akt proteins using an imaging system. The imaging system can be used to receive, enhance, and process images of cells or tissue samples stained with Akt protein negative specificity stains to determine the level of AKT protein expressed in cells or tissue samples obtained from such animals. Or degree of activation. In an embodiment of such methods of the invention, a calibration curve for the expression of AKT1 and AKT2 20 proteins of at least two cell lines exhibiting different levels of AKT protein can be produced. This calibration curve can then be used to quantitatively determine the AKT protein content expressed in a cell or tissue sample. A similar calibration curve suitable for activating AKT protein can be made using reagents specific to these activation properties. It can also be used to determine changes in AKT content and activation status before and after treatment of clinical cancer. In a particular embodiment of the methods of the invention, the AKT protein expression in a cell or tissue sample can be quantified using an enzyme-linked immunosorbent assay (ELISA) to determine the amount of prion protein in the sample. . Such methods are described, for example, in U.S. Patent Publication No. 2002/0015974. In other embodiments, an enzyme immunoassay can be used to detect the Akt5 kinase. In such assays, the enzyme is typically bound to a second antibody by glutaraldehyde or periodate. It is generally preferred to use a matrix which can be used in conjunction with such specific enzymes once hydrolyzed by the corresponding enzyme produces a detectable color change. It is also possible to use a matrix which produces fluorescence which produces a fluorescent product rather than a chromogen matrix. The enzyme-labeled antibody can be added to the first antibody-antigen complex to make it adhere, and then the excess reagent is washed away. A solution containing the appropriate substrate can then be added to the antibody-antigen-antibody complex. The matrix can be reacted with an enzyme linked to the second antibody to produce a qualitatively visible signal, which can generally be further spectrally measured to express the antigen content present in the sample. Alternatively, luminescent compounds, such as luciferin, rhodamine, and lanthanide (铕 15 (EU)), can be chemically coupled to the antibody without altering their binding ability. When activated by illumination of a particular wavelength of light, the fluorescent dye-labeled antibody absorbs light energy, induces an excitable state within the molecule, and then illuminates in a characteristic color that can be visually detected by an optical microscope. The fluorescently labeled antibody is bound to the first antibody-antigen complex. After the unbound reagent 20 is washed away, the remaining tertiary composite is then exposed to light of a suitable wavelength. The observed fluorescence indicates the presence of the relevant antigen. The Immunofluorescence Assay (IFMA) has been well established in this art and is particularly suitable for use in this method. However, other reporter group molecules such as radioisotopes, chemiluminescent or bioluminescent molecules can also be used. 64 200836749 In certain embodiments, antibodies to Akt kinase can also be used to perform ELISA vector assays for Akt kinase in serum or other circulating fluids. This is carried out by immobilizing the anti-Akt kinase antibody to a solid support and contacting it with a biological extract such as serum, blood, lymph or other body fluids, cell extracts or fine cells. A labeled anti-Akt kinase antibody can then be used to detect the immobilized Akt kinase. This assay can vary in many ways and all variations are within the scope of the present invention and are known to those skilled in the art. The method can be used, for example, in a serum-based assay to rapidly detect and quantify Akt kinase levels. In one embodiment, an Akt Elisa assay kit can be used in the present invention. Example 10 A Cellular Activation of Signaling ELISA for Akt S473 from SuperArray Bi〇science can be used in the present invention. In one embodiment, the antibody can be an anti-pan antibody that recognizes AktS473. The EHsa assay kit contains anti-Akt antibodies and additional reagents including, but not limited to, wash buffers, antibody dilution buffers, blocking buffers, cell staining solutions, imaging solutions, stop solutions, secondary antibodies, and Distilled water. Nucleotide Detection In another embodiment, a method for detecting Akt kinase by detecting the degree of expression of a polynucleic acid encoding an Akt kinase in a cell is provided. Any suitable technique known to those skilled in the art can be used to determine the performance of the 2 〇 multicore. In one embodiment, a 5 hex polynucleic acid of Akt can be used as a probe in the northern ink spot of the RNA extract from the cell. In other embodiments, in the nucleic acid amplification reaction (such as RTPCR), the nucleic acid extract of the animal can be used in combination with the message of the polynucleic acid encoded by the beer, and the anti-message sequence or its flanking sequence. Nuclear 65 200836749 Glycosyl acid primer. A variety of automated solid phase detection techniques can be used by those skilled in the art, such as those described by Fodor et al. (Science 251: 767-777, 1991) and Kazal et al. (Nature Medicine 2: 753-759, 1996). In other embodiments, methods are provided for detecting RNA transcripts encoding 5 Akt kinase. Cellular assays for Akt kinase RNA can be suspected of isolating RNA, for example, from human cancer tissue. RNA can be isolated by methods known in the art, for example using TRIZOL reagent (GIBCO-BRL/Life Technologies, Gaithersburg, Md). Oligo-dT or a random sequence oligonucleotide and a sequence-specific oligonucleotide can be used as primers in the reverse transcriptase reaction to prepare the first cDNAs from the isolated RNA. The first strand of cDNA formed in the PCR reaction can then be amplified by sequence specific oligonucleotide oxalate to produce an amplified product. Polymerase chain reaction or "PCR" refers to a procedure or technique in which a large number of preselected nucleic acids, RNA and/or DNA fragments are amplified as described, for example, in U.S. Patent No. 5,683,195. In general, the use is derived from the relevant area.

A sequence message at or near the end of the sequence to design a nucleotide primer. The sequence of these primers is identical or similar to the reverse strand of the template to be amplified. PCR can be used to amplify specific RNA sequences and cDNAs transcribed from total cellular RNA. In general, see Mullis et al. (Quant. Biol. 51:263, 1987; Erlich, eds., 20 PCR Technology, Stockton Press, NY, 1989). Thus, amplification of a specific nucleic acid sequence by PCR relies on an oligonucleotide or "introduction" having a reserving nucleotide sequence that is traced back to the arrangement of the relevant gene or protein sequence, For example, sequence comparison of mammalian Akt kinase genes. For example, a primer that is expected to bind to the anti-message strand and prepare another primer for the 66 200836749 message strand that is expected to bind to a cDNA molecule encoding the Akt kinase can be prepared. To detect the amplified product, the reaction mixture is typically subjected to agarose gel electrophoresis or other suitable hybrid technique and the relative presence of the kinase-specific amplified DNA of the Akt is detected. Akt kinase-amplified DNA can be detected, for example, by using a specific oligonucleotide probe for Southern hybridization or by comparing its 5 electrophoretic mobility to DNA standard mobility of known molecular weight. The isolation, purification and characterization of the amplified Akt kinase DNA is carried out by gel excision or elution of fragments (see, for example, Lawn et al, Nucleic Acids Res. 2:6103, 1981; Goeddel et al. Nucleic Acids Res 8:4057-1980), the amplified product is selected in a selection site of a suitable vector, such as 10 pCRII vector (Invitrogen), sequence analysis of the selected insert is performed and the DNA sequence is compared Known sequences with LIΜ kinase. The relative amount of LIM kinase mRNA and cDNA can then be determined. In one embodiment, time-consuming PCR can be used to determine the transfer of Akt nucleotides. The determination of transcriptional activity also includes the determination of potential transcriptional activity based on the availability of mRNA transcripts. Real-time PCR and other PCR programs use many chemical methods that can be used to detect PCR products, including DNA-binding fluorophores (5' endonuclease), adjacent lining and hairpin-type nucleotide probes, and spontaneous fluorescence A combination of amplicons. These chemical methods and real-time PCR are discussed in the following literature: for example, Mackay et al, Nucleic Acids Res 30(6): 1292-1305, 2002; Walker, J. Biochem. Mol. Toxicology 15(3): 121-127, 2001; Lewis et al, J. Pathol. 195: 66-71, 20 (H. In another embodiment, the step of identifying the abnormal expression of Akt is to have 67 200836749 σ complementary court from the 6a_e The nucleus (4) probe of the sequence of the nuclear (4) sequence of the 7a-d or 8a_c diagram or the sequence of the fragment of the ♦ fragment is contacted with the nuclear self-purification sequence isolated from the biological sample, and then hybridized for the sequence. The probe is used to detect the sequence and compare the result with the normal sample. The probe can be used to detect the hybridization of the probe for the biological sample by using any detectable agent. The /1⁄4 needle can be ^ 'For example, radioisotopes, biotin, fluorescent dyes, buns loosening agents, enzymes, haptens or protein labels for available antibodies. Can be borrowed from any slavery', including spectrometry, photochemistry, biochemistry, immunization Chemical method, radioisotope method or chemical method, and evaluation can be detected 10 The following techniques can also be used to detect the probe: such as oligomer restriction technique, dot ink assay, reverse dot ink assay, multi-enzyme assay, and 5, nuclear enzyme assay. Any of a wide variety of widely used DNA array technologies, including macroarrays, microarrays, and DNA microchip technology to detect the probe. The nutrient-nuclear acid probe typically comprises a pair of selected from 6a-c, 15 7a-d, and 8a-c At least 14, 15 ' 16, 18, 20, 25 or 28 nucleotide oxalates are hybridized to the nucleotides of the map or to fragments thereof. It is generally preferred not to use probes greater than about 25 or 28 nucleotides in length. The oligonucleotide probe is used to identify Akt nucleotide sequences.Kinok assay 20 Any suitable kinase assay known in the art can be used to determine the activity of such Akt kinases. For example, Hogg et al. Nc〇gene 1994 9:98-96), Mills et al. (J. Biol. Chem. 1992 267:16000-006) and

The method described in Tomizawa et al. 2001 (FEBS Lett. 2001 492:221-7), Schmandt et al. (J. Immunol. 1994, 152: 96-105), but not limited to 200836749 herein. In addition, the serine, threonine and tyrosine kinase assays are described in Ausubel et al. (Short Protocols in Molecular Biology, 1999 unit 17-6).

Akt kinase assays typically employ an Akt polypeptide, a labeled donor substrate, and a receptor matrix that is specific or non-specific for Akt. In such assays, Akt can transfer the labeled molecular group from the donor matrix to the acceptor matrix and the kinase activity is determined by the amount of labeled molecular groups transferred from the donor matrix to the acceptor matrix. A variety of expression systems can be used to make an Akt polypeptide that can be purified from a cell, can be in the form of a split or undivided recombinant 10 fusion protein and/or can have a non-Akt polypeptide sequence, for example, in its N- or The end has a His tag or /3-galactosidase. If a cancerous cell line is used as a source of Akt to be assayed, Akt activity can be assayed in these cancerous cell lines. A donor matrix suitable for Akt assays includes any molecule that is susceptible to dephosphorylation by Akt, such as 7-labeled ATP and ATP analogs, 15 wherein the label is 33p, 32p, 35s or any other radioisotope or suitable Fluorescent mark. Acceptor matrices suitable for Akt assays include any polypeptide or other molecule that is susceptible to a guanidination reaction by Akt. The receptor matrix can be derived from a fragment of an in vivo marker of Akt. The acceptor matrix fragment may be 8 to 5 amino acids in length, typically 10 to 30 amino acids, and preferably has about 1 〇, 12, 15, 20 18, 20 and 25 amino acids. A range of different peptides or other molecules can be used to determine additional receptor matrices based on the assay. Once the reaction is carried out, the target of the receptor matrix suitable for TTK can be purified from the other components of the reaction. The purification reaction is usually carried out by molecular interaction, wherein the receptor matrix is biotinylated and can be used to identify the specific antibody of the receptor matrix by streptavidin 69 200836749 or specificity. Purification by interaction. The reaction can be carried out under various conditions, such as on a solid carrier, in a gel, in a solution, or in a living cell. The choice of detection method depends on the type of label used for the donor molecule and can include, for example, autoradiography, scintigraphy, scanning, or fluorescence radiography to perform an assay that incorporates radiation or fluorescence. Methods of Treatment The compounds and pharmaceutical compositions provided herein can be used to treat a condition, including tumors, cancer, and other conditions associated with abnormal cell proliferation. In one embodiment, the compounds of the invention are useful for treating cancer, sarcoma, lymphoma, white disease, and/or myeloma. In other embodiments of the invention, the compounds disclosed herein can be used to treat solid tumors. The compounds of the invention are useful in the treatment of cancer, such as, but not limited to, cancer of the following organs or tissues: breast, prostate, lung, bronchi, colon, urinary system, bladder, non-Hodgkin's lymphoma, melanoma , kidney tumor, kidney system, pancreas, pharynx, thyroid, stomach, brain, multiple myeloma, esophagus, liver, intrahepatic biliary tract, neck, larynx, acute myeloid leukemia, chronic lymphocytic leukemia, soft tissue (such as heart ), Hodgkin's lymphoma, testicular, small intestine, chronic myelogenous leukemia, acute lymphocytic leukemia, anal, anal canal, 20 anal and rectal system, thyroid, female genital, gallbladder, pleura, eye, nose, nasal cavity, Middle ear, nasopharyngeal, ureter, peritoneum, omentum, mesentery, and gastrointestinal tract, high degree glioma, glioblastoma, colon, rectum, pancreas, stomach cancer, hepatocellular carcinoma; head and neck cancer, carcinoma Renal cell carcinoma; adenocarcinoma; sarcoma; hemangioendothelioma; lymphoma; leukemia, sickle 70 200836749 edema. In further embodiments, the compounds of the invention may be used to treat dermatological conditions including, but not limited to, malignant diseases: angiosarcoma, blood stasis, basal cell carcinoma, squamous cell carcinoma, malignant melanoma, and cardo Western sarcoma; and non-malignant diseases or conditions, such as psoriasis, 5 lymphatic hyperplasia, childhood hemangioma, Stitch _ Weberga exhibition (4) syndrome, vulgaris, fibrone on neuroma, nodular sclerosis, abdominal granuloma, Recessive dystrophic macrobubble epidermal loosening 'venous ulcers, acne, acne scars, eczema, infectious soft palate, sebum leakage keratosis, and actinic keratosis. Compositions comprising such compounds of the invention may be used to treat these cancers and other cancers at any stage from the discovery of the cancer to the advanced stage. In addition, compositions comprising the compounds of the invention may be used to treat primary cancer and its metastasis. In other embodiments of the invention, the compounds described herein are useful for treating cancers which include, but are not limited to, the cancers disclosed in Table 1 below. □ Table 1: Cancer 1 - □ Adult acute lymphoblastic leukemia Children acute lymphoblastic leukemia Adult acute sputum leukemia Children Acute myeloid leukemia Adrenal cortical cancer Children Adrenal cortical cancer AIDS Related cancer AIDS Associated lymphoma Anal cancer Child cerebellum Astrocytoma cerebral astrocytoma □ basal cell carcinoma □ hair cell leukemia head and neck cancer adult liver cell (liver) cancer (primary) child liver cell (liver) cancer (primary) adult ho Kidney Lymphoma Childhood Hodgkin's Lymphoma Pregnancy Hodgkin's Lymphoma Hypopharyngeal Children Hypothalamus and Optic Nerve Glioma 眼 Intraocular Melanoma Teng Island Cell Carcinoma (Endocrine Pancreas) Posi Sarcoma (Kaposi's Sarcoma;) Kidney Cell Carcinoma---~~-- 71 200836749 Extrahepatic biliary tract cancer Bladder cancer Child bladder cancer Bone cancer Osteosarcoma/malignant fibrous histiocytoma Child brain stem glioma Adult brain tumor Child brain tumor, brain stem glioma cerebellar brain tumor child astrocytoma cerebral brain tumor child astrocytoma / malignant Brain tumors in children with glioma, brain tumors in children with ependymoma, brain tumors on the cerebral blastoma of children, primordial neuroectodermal tumors in children, brain tumors, optic nerve bundles and hypothalamic gliomas in children with brain tumors Breast cancer Child breast cancer Male breast cancer Child bronchial adenoma / Carcinoid Burkitfs Lymphoma □ Child carcinoid gastrointestinal carcinoid tumor Unknown primary CNS cancer Child Cerebellar astrocytoma Children Cerebral astrocytoma / malignant glioma children glioma children kidney cancer laryngeal cancer children laryngeal cancer adult acute lymphoblastic leukemia acute lymphoblastic leukemia children primary lymphoma adult acute myeloid leukemia children acute bone Leukemia chronic lymphocytic leukemia chronic visceral leukemia B cell leukemia lip and oral cancer adult liver cancer (primary) childhood liver cancer (primary) non-small cell lung cancer small cell lung cancer AIDS associated lymphoma Burkitt's lymphoma Skin T-cell lymphoma, see verrucous granuloma and Nishizawa (SSz Ary) syndrome adult Hodgkin's lymphoma child Hodgkin's lymphoma pregnancy Hodgkin's lymphoma adult non-Hodgkin's lymphoma child non-Hodgkin's lymphoma pregnancy non-Hodgkin's Lymphoma primary central nervous system lymphoma 大 macroglobulinemia, bone of Waldenstrdm's malignant fibrous histiocytoma / osteosarcoma children neural tube blastoma melanoma intraocular (eye) melanin Mole cancer cell carcinoma Malignant mesothelioma 72 200836749 Cervical cancer Child mesothelioma Child cancer Recessive primary metastatic squamous Chronic lymphocytic leukemia Carcinoma Chronic spinal cord hyperplasia Children multiple endocrine neoplasia Formation of syndromes, colorectal cancer, multiple myeloma/plasma neoplasms, colorectal cancer, squamous cell granuloma, T-cell lymphoma, see myelodysplastic syndrome, verrucous granuloma and zeazel syndrome, myeloproliferative/myeloproliferative □ Endometrial cancer disease children ependymoma chronic myeloid leukemia esophageal cancer adult acute osteomyelemia Esophageal cancer in children with acute myeloid leukemia Ewing's family tumor multiple myeloma children gonadal ectodermal tumors chronic myeloproliferative disorders gonadal blastoma tumors □ nasal and paranasal sinus cancer extrahepatic biliary tract cancer nasopharyngeal carcinoma Eye cancer, intraocular melanoma, nasopharyngeal carcinoma, retinoblastoma, neuroblastoma, gallbladder carcinoma, adult, non-Hodgkin's lymphoma, gastric cancer, non-Hodgkin's lymphoma, childhood, gastric cancer, non-Hodge Jin's gastrointestinal carcinoma, lymphoma, ectodermal cell tumor, non-small cell lung cancer, children's gonadal ectodermal cells, childhood oral cancer, oral cancer, lip and oropharyngeal cancer, blastocytic tumor, osteosarcoma, bone, malignant fibrous tissue, surname Nursing trophoblastic tumor cell tumor Adult glioma Child ovarian cancer Child brain stem glioma Ovarian epithelial cancer Child cerebral glioma Ovarian blast cell tumor Astrocytoma Ovarian low malignant potential tumor Child visual nerve bundle and hypothalamus □ Pancreatic cancer glioma children visceral cancer □ skin cancer (melanin Islet cell pancreatic cancer Mock cell skin cancer tumor paranasal sinus and nasal cancer small cell lung cancer parathyroid carcinoma small intestine cancer penis cancer adult soft tissue sarcoma pheochromocytoma 73 200836749 pineal gland blastoma and supratentorial ϋ 4tU< Trfr .rfe

Squamous cell carcinoma, see itchy skin (non-melanoma) A stable primary metastatic squamous cell carcinoma, gastric cancer, children with gastric cancer, supratentorial primitive neuroectodermal tumor, □ T-cell lymphoma, see verrucous granuloma and Xizelai syndrome group 癌 癌 癌 儿童 thymoma thymoma and thymoma thyroid cancer children 曱 腺 腺 盂 盂 盂 盂 盂 细胞 细胞 细胞 细胞 细胞 细胞 细胞 细胞 细胞 细胞 细胞 细胞 细胞 □ □ □ □ 成人 成人 成人 成人 成人 成人 成人 成人 成人Type of primary site cancer, ureter and renal pelvis, unusual cancer, transitional cell carcinoma, urethral cancer, endometrial uterine cancer, uterine sarcoma, vaginal cancer, children's optic nerve bundle, and hypothalamic glioma, female genital cancer, huadente Wilm's Tumor primordial neuroectodermal tumor pituitary tumor water cell tumor / multiple myeloma pleural and lung blastoma breast cancer pregnancy ten years of pregnancy Hodgkin Lymphoma, pregnancy, non-Hodgkin's lymphoma, primary central nervous system, lymphoma, prostate cancer, renal cell (kidney) cancer, childhood kidney cell (kidney) cancer Lunar sputum and ureteral transitional cell carcinoma retinoblastoma children rhabdomyosarcoma salivary gland cancer children salivary gland carcinosarcoma, eus family tumor sarcoma, Kaposi's sarcoma adult soft tissue sarcoma uterine sarcoma West Zeray syndrome skin cancer (non-melanoma) Child Skin Cancer In a further embodiment of the invention, the compounds disclosed herein are useful for treating angiogenesis-associated diseases. Angiogenic small molecules include thalidomide, which has a function of inhibiting NFkB in the genus 74 200836749; 2-methoxyestradiol, which affects microtubule activation and hypoxia-inducible factor (HIF la Activation; cyclooxygenase 2 (COX2) inhibitors; and low doses of conventional chemotherapeutic agents, including cyclophosphamide, onioncycline analogs, and vinca alkaloids (vincristine 5 ( Vincristine), vinblastine (D'Amato, RJ et al. (1994) Proc. Natl. Acad. Sci. USA 91, 3964-3968, D, Amato, RJ et al. (1994) Proc· Natl· Acad Sci. USA91, 4082-4085). In addition, specific tyrosine kinase inhibitors can indirectly reduce angiogenesis by reducing VEGF and other pro-angiogenic factors produced by tumors and stromal cells. Herceptin, imatinib, (Glivec), and lressa (Bergers, G. et al. (2003) J(10)r(10)/ 〇/(10) 111, 1287-1295, Ciardiello, F. Et al. (2001) Clinical Cancer Research 7? 1459-1465, Plum, SM et al. (2003) C/Wca/ 9, 4619-4626) o 15 Nearly 'angiogenesis inhibitors have evolved into animal models. Angiogenesis inhibitors represent promising treatments for various cancers. Recently, Avastin, which is an anti-vascular endothelial growth factor (VEGF) , has been shown to be a single agent in advanced renal cell carcinoma to prolong life and to use chemotherapy in advanced colon cancer to prolong life (Yang, JC et al. 20 (2003) New England Journal of Medicine 349, 427-434,

Kabbinavat, F. et al. (2003) J0urnai of Clinical 〇nc〇1〇gy 21, 60_65). Cortical-related diseases include, but are not limited to, inflammation, autoimmune diseases, and infectious diseases; angiogenesis-dependent cancers include, for example, solid tumors, 75 200836749 hematogenous tumors (such as leukemia), and tumor metastasis; , for example, blood stasis, auditory neuroma, neurofibromatosis, trachoma, and purulent granuloma; rheumatoid arthritis; psoriasis; wet therapy; ocular angiogenic diseases such as diabetic retinopathy, retinopathy of prematurity, Macular degeneration, 5 corneal transplant rejection, neovascular glaucoma, posterior fibrous tissue hyperplasia, redness, Osler-Weber syndrome; myocardial angiogenesis, plaque neovascularization; microvascular dilatation; hemophilia Joints; angiofibroma, and wound granulation. In addition, the compositions of the present invention may be used to treat diseases such as, but not limited to, intestinal adhesions, atherosclerosis, Pang, and 10 hypertrophic scars (i.e., neoplasms). The compositions of the invention may also be used to treat diseases which have angiogenesis due to pathogenic consequences, such as R〇chde minalia quintosa, Helobacter pylori, tuberculosis, and leprosy. Drug-Resistant Tumor or Cancer Treatment #本舍明k provides a compound useful for the treatment of anti-sex cancer, including the cancer and triclinibine compound and/or bortezomib and its derivatives disclosed in the above. Multidrug resistance (MDR) occurs in human cancer and can be a major obstacle to the success of chemotherapy. Multidrug resistance is a phenomenon in which tumor cells exposed to cytotoxic agents form cross-resistance to a range of structurally and functionally unrelated compounds. In addition, MDR can occur endogenously in certain cancers that have not been exposed to chemotherapeutic agents before. Thus, in one embodiment, the invention provides a method for treating a patient having an anti-musk cancer, such as a multi-drug resistant cancer, by administering TCN, TCN-P, TCN-PM as disclosed herein Or related compounds with shedzomib and its derivatives. In a particular embodiment, TCN, TCN-P, TCN-PM and related compounds and bortezomib and its derivatives 76 200836749 can be used to treat only Taxol, rapamycin, Temo Cefend, splatin, and/or gefitinib (Aressa) resistant cancer. 5 10 15 20

In her case, as disclosed in the article, TCN, TCN-P, TCN-PM = Mesh-complex and (4) Zomi and its projections can be examined for cancer: the kidneys, the adrenal glands, the pancreas, the liver, And/or other cancers. The rice and its sputum, and the t-like towel of the present invention can be administered together with other cytotoxic agents. In the other and in the same ', the case of the tresibine compound and bortezomib and its derivatives: 2: 'When used to treat solid tumors, another example of the use of light compounds - examples In the text, the trebutine chemotherapeutic agent disclosed in the text. Zomi and its biological and composition can be combined with at least one additional stream.兮^— A backing may be provided with the compound disclosed herein or in the form of a portion of the same composition that may be formed at the same time or may be administered at appropriate or different times. In the case of the application, the <t-tomizine and its extension τ are not disclosed as such a sirolimus compound and boron, or may be used in combination with an anti-angiogenic agent to enhance their effectiveness. An anti-angiogenic agent and other cytotoxic drugs, such as Quxi Li... In the case of the sputum, when used to treat solid tumors, the medicinal substances and thiozomib and its derivatives can be selected Self-sufficient (only to be 'these agents are not limited to · IL-12, retinoic acid 77 200836749 (retinoids), interferon, angiostatin, endostatin, sali mad, gold Thrombospondin-1, thrombospondin-2, captopryl, anti-neoplastic agents (such as alpha interferon), COMP (cyclophosphamide, vinorelbine, amesu Meth5 (methotrexate) and prednisone (prednisone), scorpion etoposide, mBACOD (americ sulphate, breomycin, cranberry, cyclophosphamide, periwinkle Neobase and dexamethasone (dexamethasone), PRO-MACE/MOPP (prednisone, Amesu 嗓 ( (w/ ercofu rescue agent), cranberry, cyclophosphamide, scorpion B 10 Fork/mechlorethamine, new periwinkle, procarbazine, vinblastine, vinblastine, blood Angiostatin, TNP-470, pentosan, polysulfate, platelet factor 4, vasstatin, LM-609, SU-1 (H, CM-101, Techgalan, Shaly sinensis, SP-PG, and radiation. In other embodiments, the 15th compound and composition disclosed herein may be administered together with or in turn: for example, an anti-mitotic drug, such as a cytoskeleton The elemental calibration drug, which includes podophylotoxin or vinca flower bioassay (vinca new test, periwinkle test); antimetabolite drug (such as 5-fluorouridine, dextran, cytarabine (cytarabine), gemcitabine, guanidine-like 20 substances, such as pentostatin (Pentostatin, Amesu); alkylating agent or nitrogen fenfluent (such as nitrosourea, cyclophosphamide or AI Fukusamine (if〇sphamide); a drug that can be labeled with DNA, such as the anthracycline drug, adriamycin, cranberry, pan-eimycin (pharmorubici... or epirubicin; A drug that will be labeled with a topoisomerase, such as G. sinensis; hormone 78 2008 36749 and hormone agonists or antagonists, such as estrogen, antiestrogens (temoxifene and related compounds) and androgens, flutamide, leuprorelin, goserelin , cyprotrone or octreotide; a drug 5 that can modulate signaling in tumor cells, including antibody derivatives, such as oxetine; alkylating drugs, such as platinum drugs (stepping, carding) Carboplatin, oxaliplatin, paraplatin or sulfhydryl urea; drugs that potentially affect tumor metastasis, such as matrix metalloproteinase inhibitors; gene therapy agents and anti-message agents; antibodies Therapeutic agents; other biologically active compounds of marine origin, especially 10 drops of didemnin, such as aplidine; steroid analogs, especially dexamethasone; antiemetics, including 511孓3 inhibitors (such as gramisetron or ondasetron, with steroids and their derivatives, especially dexamethasone. In still other embodiments, the compounds and compositions can be used with or in turn with the chemotherapeutic agents disclosed in Table 2 below.

Table 2: Chemotherapeutic Agents -13 -cis-retinoic acid-2-amino-6-mercaptopurine -2-CdA

-2-Chlorodeoxygenated gland-5-gas urinary β-sigma sigma-5-FU -6-TG -6-sulfur sulphide-6-weiki 嗓呤-6_ΜΡ -Accutane - radiant菌素-D - os 素 环 Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne Ne - Novaldex - Novantrone Octreotide - Octreotide Acetate - Oncospar 79 200836749 - Adrucil - Agrylin - Ala-Cort • Aldesleukin Alemtuzumab - Alitretinoin - Alkaban - AQ - Alkeran All-trans retinoic acid-α-interferon-hexamethine Altretamine - Amethopterin - Amifostine - Aminoglutethimide - Anagrelide - Anandron • Ana Anastrozole "Avabinosylcytosine • Alla (Ara)-C - Aranesp • Aredia - Arimidex - Aromasin

- Ascites _ Asparagine - ATRA - Avastin

-BCG

_BCNU - Bevacizumad - Bexarotene

• Bicalutamide - BiCNU _ Blenoxane - Bleomycin - Bortezomib - Busulfan - Busulfex - Tumor (Ontak) - Onxal - Onxal - Oprevelkin - Orapred - Orasone - Oxaliplatin - Pacific Paclitaxel - Papin Phosphonic Acid Salt (Pamidronate) - Pan Lei, Panretin - Pediapred - PEG interferon - Pegaspargase - Pegfilgrastin - PEG insert ( Intron) • PEG-L-aspartame-phenylalanine mustard-cisplatin (Platinol)

- cisplatin-AQ - Adrenal cortex (Predoneolone) - Prednisone - Predrome - Procrust - PiOleukin - with carmustine implant Prolifeprospan 20 - σPurnethol - Raloxifene Rheumatorex - Rimixan - Rituximab 'Ruo Roveron-A (a-2a interferon) - Rub ex - Rubidomycin hydrochloride - Sandostatin

-Good to get LAR 80 200836749

-C225 - Calcium Leucovorin - Campas - Camptosar - Camptothecin-11 - Capecitabine - Carac - Carboplatin - Carmustine - Carmustine - Carmustine wafer - Casodex - CCNU - CDDP - CeeNU - Cerubidine - Cetuximab • Chlorambucil - Cisplatin • Citrovorum factor • Cladribine - Cortisone - Cosmegen - CPT-11 - Cytadren - Cytarabine - Cytarabine - Cytosar - U - Cytoxan - Daccarbazine Dactinomycin -α Dabepoetin alfa - Daunomycin - Daunorubicin Daunubicin hydrochloride •Danobicin liposome-Dangsuo DaimoXome) - Decadron - Sargramostim - Solu-Cortef - Solu-Medrol • STI-571 • Streptozocin ) - Tamoxifen • Targretin, Targretin - Taxol - Taxotere - Temodar • Temozolomide - Teniper Ring (Teniposide) -TESPA - Thalidomide - Thalomid - TheraCys - Thioguanine - Thioguanine Pills - Thiofosamide Thioplex - Thiotepa - TICE • Toposar • Topotecan - Toremifene - Trastuzumab - Tretinoin - Destroy Trexall - Trisenox -TSAP -VCR •Velban -Velcade -VePesid -Vesanoid •Viadur - Changchun flower test - Changchun flower test sulphate salt - Vincasar Pfs 81 200836749 -6 - Solid layer (Delta-Cortef) _ De Pesson (Deltlasone) - Denileukin difitox - Dipselet (DepoCyt) - Dexamethasone - Dexamethasone acetate - Dexamethasone sodium - Dexasone Dexrazoxane - DHAD -DIC - Diode Ex) - Docetaxel - Doxil - Doxorubicin - Xiaohong per liposome - Droxia - DTIC - DTIC - Hemisphere - Duralone ) - Efudex - Eligard - Ellence • Eloxatin - Elspar - Emcyt - Epirubicin ) -ck Epoetin alfa - Erbitux - Erwinia L - aspartate - Estramustine - Ethyol - Etopo pine (Etopophos) - scorpion 臼 式 - 臼 臼 臼 - - 优 优 优 E E E E E E E E E E E E E E E E E E E E E E E E E E E E E E E E E E E E E E E E E E E E E E E E -VP-16 - Vumon - Xeloda - Zanosar - Zevalin - Zinecard - Zoladex Sitting Zoledronic acid ) - Zometa - Gliadel wafer - Glivec - CM-CSF - Goserelin - Granulocyte - Colony Stimulating Factor - Granulocyte Macrophage Colony Stimulation Factor-fluoromethyl ketone (Halotestin) - He Cancer - Hexadrol • Hexalen - hexamine melamine - HMM - Hycamtin - Hydrea - Hydrocort acetate · Hydrocorticone (corocortisone) Corticosterone Sodium phosphate corticosterone succinate 'hydrocorticosterone phosphate-Ibritumomab - Ibritumomab Tiuxetan • Idamycin ^ Idarubicin ) (Ifex) 82 200836749

- Exemestane - Fareston - Faslodex - Femara - Filgrastin - Fluxuridine - Fludara ) -Fluaroabine -Fluoroplex - Fluorouracil - Fluorescent Cancer (Fluorescent) - Fluoxymesterone - Fluoric Acid - Folinic Acid ) _FUDR - Fulvestrant - G-CSF - Gefitinib - Jesitabine - Gemtuzumab ozogamicin - Gemzar - Gleevec - Lupron ) - Lupron Depot - Matlane • Maxidex "Mechlorethamine - Mecamamine hydrochloride - Medralone - Medrol -Megace - Megestrol • Megestrol acetate - Melphalan - Mesna & Mesnex - Amesu-a-IFN-Isdfamide-IL-2I-IL-11 Imatinib-methionate-imidazolecarboxamide β-interferon-a 2b interferon (peg conjugate) -介百素-2 -介百素-11 -Introm A (a -2b interferon) - ercofu - Leukeran - Leukine - Liu Pei Lin - Changchun New Test ( Leurocristine) - Leustatin - Liposomal Ara-C - Liquid Pred Lomustine - L-DAM Sarcolysin " Meco Meticorten - mitomycin - schizophrenia - C - mitoxantrone M - prednisone (Prednisol) - MTC - MIX - nitrogen mustard (Mustargen) • Mustard (Mustine) - Trichoderma Mutamycin - Myleran - Irinotecan - Isotretinoin - Kidrolase - Lanacote - L-aspartate Aminase 83 200836749 - Amesu Cannes ^ CR " - methyl adrenocorticosterone - Mylocel - Letozole In certain embodiments, interferon (IFN) can be used with the present invention These compounds are used together. Suitable interferons include: a: _2a interferon, a-2b interferon, pegylated alpha interferon (which includes 0: _23 interferon & a-2b interference 5), interferon, 7 interference , τ interferon, ω interferon, INFERGEN (alphacon-l interferon) manufactured by InterMune, OMNIFERON (natural interferon) manufactured by Viragen, and Russian cloth made by Human Genome Science ALBUFERON, REBIF (yS-la interferon) manufactured by Ares-SeiOno, ω interferon 10 manufactured by BioMedicine, oral interferon alpha manufactured by Amarillo Biosciences, and interferon manufactured by InterMune, τ interferon, and / or τ -1 /3 interferon. In one embodiment, TCN, TCN-P, TCN-PM or related compounds and one or more compounds as disclosed herein may be used together with additional chemotherapeutic agents, such as the chemotherapeutic agents described herein or in Table 2, or in turn. To treat anti-15 drug cancers, such as multi-drug resistant cancer. Drug-resistant cancers can include colon cancer, bone cancer, kidney cancer, adrenal cancer, pancreatic cancer, liver cancer, and/or any other cancer known in the art or described herein. In one embodiment, the additional chemotherapeutic agent can be a P-glycoprotein inhibitor. In a specific non-limiting embodiment, the P-glycoprotein inhibitor can be selected from the group consisting of verapamil, cyclosporine (such as cyclosporin A), and temoximine. Calmodulin antagonist, dexverapamil, dexniguldipine, bespoda 84 200836749 (^^18卩〇(1&〇(?80 833), More than (1^1^〇(131*)(^/又-710), tariquidar (XR9576), zosuquidar (LY335979), laniquidar (Rl〇1933) And/or ONT-093. Pharmaceutical Composition 5 These compositions comprising the tricilide compound and bortezomib and its derivatives may be optionally administered with a pharmaceutical carrier or excipient. The pharmaceutical carrier for administration of a compound includes any such carrier known to those skilled in the art to be suitable for a particular mode of administration. The tresibine compound can be formulated together with bortezomib and its derivatives. The only active ingredient in the substance may be combined with bortezomib and its derivatives. Including the tromethamine compound and boron The composition of zomi and its derivatives may be suitable for oral, rectal administration, nasal administration, topical administration (including buccal and sublingual administration), vaginal administration or parenteral (including subcutaneous, intramuscular, intravenous, intradermal) Intraocular, intratracheal, intracisternal, intraperitoneal, and epidural. 15 These compositions are preferably administered intravenously. These compositions may preferably be presented in unit dosage form and may be prepared by conventional pharmaceutical techniques. These techniques include associating - or a plurality of compositions of the present invention with a plurality of pharmaceutical carriers or excipients. The isocyanine compound can be combined with stilzomib and its street organisms, and the like. The composition is prepared into a pharmaceutical preparation suitable for oral administration, such as a solution, a liquid, a lozenge, a dispersible lozenge, a pill, a capsule, a powder, a sustained release: a prescription or an ugly agent, or a domain which is difficult to be administered to the intestine a solution or solution, and a transdermal patch and a dry powder inhaler. In one embodiment, the above-described triclinide compound is adjusted to form a pharmaceutical composition using techniques and procedures well known in the art (see, For example Ansel Introduction to Pharmaceuti Cal Dosage Forms, 4th edition, 1985, 126). In such compositions, one or more of the effective concentrations of the compound or a pharmaceutically acceptable derivative thereof can be combined with one or more suitable pharmaceutical carriers. The compounds of the invention may be derivatized to the corresponding salts, esters, enol ethers or esters, condensates, ketals, orthoesters, semi-wakes, hemi-ketals, acids, solvates, hydrates or former Drugs. Once administered, the concentration of such compounds within the composition is effective to deliver an amount that can treat, prevent or ameliorate one or more of the symptoms of the target disease or condition. In one embodiment, the compositions 10 are formulated to be suitable for single dose administration. To modulate the composition, the weight-dispersed compound is dissolved, suspended, dispersed or mixed in a particular carrier at a concentration effective to alleviate, prevent, or ameliorate one or more symptoms. The composition suitable for oral administration can be provided in the form of a separate unit such as, but not limited to, a key, a caplet, a pill or a dragee, a capsule or a sachet, each containing a predetermined amount of the composition of the genus Or granules; solutions or suspensions in aqueous or non-aqueous liquids; or oil-in-water liquid emulsions or water-in-oil emulsions, or doughs, and the like. For example, by using a carrier, such as water, salt liquid, aqueous dextrose, glycerin, ethylene glycol, ethanol, etc., the pharmaceutical adjuvant to be dissolved, dispersed or mixed with triclinide and 20 is required to form a solution. Or a suspension to form a liquid pharmaceutically acceptable composition. If necessary, the pharmaceutical composition to be administered may also contain small amounts of non-toxic auxiliary substances such as wetting agents, emulsifiers, solubilizers, pH buffers, preservatives, flavoring agents, and the like, such as acetic acid, lemon, etc. Sodium, cyclodextrin derivatives, sorbitol needle monolaurate, 86 200836749 sodium alkanoate, triethanolamine oleate, and other such agents. Methods of preparing such dosage forms are known or known to those skilled in the art; for example, see Remington's Pharmaceutical Sciences, Mack Publishing Company, Easton, Pa., 15th Ed., 1975 〇5 Compositions of the Invention Suitable for Topical Administration in Oral Administration Included, for example, as a flavouring-based ingredient, typically a tablet of sucrose and acacia gum or tragacanth; having the tromethamine compound of the present invention and bortezomib and its derivatives in an inert ingredient such as gelatin And a fixative in glycerin, or sucrose and acacia; and a sputum 10 having the compositions of the invention administered in a suitable liquid carrier. Such tablets, pills, capsules, buccal tablets and the like may contain one or more of the following ingredients or compounds of similar nature: binding agents, lubricants, diluents, slip agents, decomposers, colorants, sweeteners, flavorings Agents, lubricants, emetic coatings, and film coatings. Examples of the binder include microcrystalline cellulose,

15 tragacanth, glucose solution, gum arabic mucilage, gelatin solution, molasses, polyethyl hydrazine 比 洛 洛 洛 洛 洛 洛 洛 洛 洛 洛 洛 洛 洛 洛 洛 洛 洛 洛 洛 洛 洛 洛 洛 洛 洛 洛 洛 洛 洛Things. Lubricants include talc, starch, stearin 20 or calcium stearate, lycopodium and stearic acid. Diluents include, for example, lactose, Yan sugar, house powder, kaolin, salt, mannitol, and = dicalcium. Spray agents include, but are not limited to, colloidal cerium oxide. The decomposing agent includes crosslinked sodium carboxymethyl cellulose, sodium starch glycolate, also 4 ^ ^, per bath acid, corn starch, potato starch, bentonite, methyl fiber xin, rob, summer fat and carboxymethyl cellulose. . Coloring agents include, for example, any water that has been approved for binding, a mixture of sex and c dyes, and the like; and suspended in a character of the water. Sweeteners include sucrose, lactose, mannitol, and artificial sweeteners such as saccharin, and any amount of spray-dried flavor. Flavoring agents include natural flavorings extracted from plants such as fruits and blends of synthetic compounds which produce a pleasant sensation such as, but not limited to, peppermint and methyl laurate. Wetting agents include propylene glycol monostearate, sorbitan monooleate, diethylene glycol monolaurate, and polyoxyethylene lauryl ether. The vomiting coating includes fatty acids, fats, waxes, shellac, ammoniated shellac and cellulose acetate vinegar. Film coatings include hydroxyethyl cellulose, sodium carboxymethyl cellulose, polyethylene glycol 4, and cellulose acetate phthalate. The composition suitable for topical administration to the skin can be provided as an ointment, emulsion, gel, and paste having one or more compositions administered in a pharmaceutically acceptable carrier. Compositions suitable for rectal administration may be presented as a suitable base, including suppositories such as cocoa butter or salicylic acid. 5 When the carrier is a solid, the composition suitable for nasal administration comprises the administration of a nasal inhalation (i.e., by inhalation through the nasal passages by means of a powder containing the powder close to the nose) Powders having a particle size in the range of, for example, from 2 to 500 microns, one or more of which may be incorporated into or absorbed into the nasal passages in an aqueous or oily solution. Compositions suitable for vaginal administration may be presented as a vaginal pill, tampon, emulsion, gel, paste, foam or spray containing one or more of these compositions and a suitable carrier. Compositions suitable for parenteral administration include aqueous and non-aqueous sterile injectable solutions which may contain antioxidants, buffers, bacteriostatic agents, and isotonicity of the formulation with blood of a predetermined recipient; Aqueous and non-aqueous sterile suspensions which may include suspending and thickening agents. Such compositions may be provided in unit or multi-dose containers (eg, sealed ampoules and vials) and may be stored in a lyophilized dry state immediately after application of a liquid carrier, such as water for injection. under. The ready-to-use injections and suspensions can be prepared from the previously described sterile powders, granules and lozenges.

1〇 15

2"" A pharmaceutical organic or inorganic solid carrier vehicle medium suitable for enteral or parenteral administration is used to prepare the compositions. Gelatin, lactose, house powder, bis acid town, talc, vegetable and animal fats and oils, gums, polyalkylene alcohols, water or other known carriers are all suitable as carrier media. The use of the trichostatin compound and sirolimus and its derivatives is used in combination with a pharmaceutically acceptable carrier medium and/or excipient. As a medium, ~ alpha carrier, bath, diluent or other liquid vehicle, = adjuvant, surfactant, isotonic agent, _ or emulsifier, = or suspension binder, lubricant, adjuvant , vehicle: agent, absorbent, surfactant, coloring agent, flavoring agent or sweetening agent, etc., in addition, 'these include triclinbine compound and (4) can be used together with pharmaceutically acceptable Shape agents and as needed, complaints: the choice of the substrate 'such as biodegradable into = hold:. "Pharmaceutically acceptable agents", including non-toxic solids:: sessile fillers, diluents, sputums or liquid dreams... materials or any type of formulation adjuvant. ..., and should be aware of such The composition of the composition is determined per patient according to reasonable medical judgment. The therapeutically effective dose for any particular ==2,200836749 may depend on various factors including, for example, the condition to be treated and the severity of the condition; The activity of a particular composition; the specific composition, age, weight, general health, sex, and daily diet used by the patient; the time of administration; the mode of administration; the rate of discharge of the particular compound used; the duration of treatment a similar factor that is well known to us in the art of medicinal techniques, such as the triclinide compound and/or quetiazide and its derivatives used together with the particular composition used, for example, in the technical scope of the art. Preferably, the composition is administered at a dose lower than that required to achieve the desired therapeutic effect, and then the dosage is gradually increased until the desired therapeutic effect is achieved. The uniformity of administration and dosage is preferably in the form of a dosage unit. The composition of the weekly preparation of the triclinide compound and bortezomib and its derivatives, as used herein, is in the form of a dosage unit, which means that it is suitable for treatment. The actual individual unit of the host's touch. Each dose should contain a group of early parental 1 to unit dosages, a sub-dose, or a suitable fraction, in association with the specific pharmaceutical carrier medium itself, in an amount expected to produce the desired therapeutic effect. Ingredients are administered. For example, a compound disclosed in m to 50 milliliters per day can reduce the volume of solid tumors in mice. The dosage may be derived from host factors such as finance, age, metabolism, tissue distribution, rate of absorption, and (iv) rate. In - two days, humans can be given an appointment. Revised in the text of 5 to 7 grams: Reduced compounds. You can selectively vote for humans every day! To the object. In the specific implementation of the money, every day to the humans to = 20 200836749 • 5 compounds. The therapeutically effective dose can depend on a number of factors such as those described above. In addition, it is preferred within the scope of the art to first use a relatively low dose of the composition and then add the agent until the desired therapeutic effect is achieved. A composition comprising a triclinide compound and bortezomib and a derivative thereof may be used in combination with a sustained release matrix which may be made of a material which is usually a polymer and which may be hydrolyzed by enzymatic or acid-based Or degraded by dissolution. Once in the body, the matrix acts by enzymes and body fluids. For example, the sustained release matrix is selected from biocompatible materials such as liposomes, polylactide (polylactic acid), polyglycolide (polymer of glycolic acid), polylactide 10 co-glycolide (lactic acid and ethanol). Copolymer of acid), polyanhydride, poly(ortho)ester, polypeptide, hyaluronic acid, collagen, chondroitin sulfate, carboxylic acid, fatty acid, phospholipid, polysaccharide, nucleic acid, polyamino acid, amino acid (such as phenyl) Alanine, lysine, isoleucine), polynucleotides, polyvinyl propylene, polyvinylpyrrolidone, and polyoxoxime. A preferred biodegradable substrate is a matrix of any of polylactide, polyglycolide or 15 • polylactide co-glycolide (copolymer of lactic acid and glycolic acid). The citrifloxacin compound and bortezomib and its derivatives can also be administered in the form of a liposome. As is known in the art, the liposomes are typically derived from phospholipids or other lipids. The lipophilic system is formed by crystallization of a mono- or multi-layered hydrated liquid dispersed in an aqueous medium. Any non-toxic, physiologically acceptable and metabolizable lipid capable of forming a liposome can be used. In addition to one or more of the compositions of the present invention, the liposome may contain a stabilizer, a preservative, an excipient, and the like. Examples of lipids are natural and synthetic phospholipids and phospholipids (lecithin). Methods of forming liposomes are known in the art. 91 200836749 An aerosol suitable for the preparation of a compound with bortezomib and its derivatives. It is suitable for the administration of the respiratory tract or in the form of a cup of aerosol or a solution in the form of a nebulizer, in the form of a stipulation, and can be used in combination with inert particles. In this case, the particles of the formulation have a diameter of less than 5 Å and a diameter of less than _ meters. w ^财, whose particle 10 includes the group of the triconide compound and boron # 佐米 and its enchantment can be used in its composition and / or procedures - to treat the above disease 'positive, for example, to use surgery Elevating or chemotherapeutic and one or more of the compositions of the present invention to treat a tumor, and then the patient may be administered an inactive state of the present invention or a plurality of compositions called a long sputum transfer, and the shirt is inhibited, inhibited or reduced. Growth of primary tumors. Further Example 15 The pharmaceutical compositions comprising the tricilide compound and bortezomib and its derivatives can be prepared according to known methods for preparing pharmaceutically useful compositions. Formulations are described in many sources that are well known to us and readily available to those skilled in the art. Such as Remington, s Pharmaceutical

Sciences (Martin EW [1995] Easton Pennsylvania, Mack 2(R) Publishing, Company, 19th Edition) describes formulations that can be used with the present invention. Formulations suitable for administration include, for example, aqueous sterile injection solutions which may contain an antioxidant, a buffer, a bacteriostatic agent, and a solute which renders the formulation isotonic with the blood of the intended recipient; and may include suspending agents and thickening Aqueous and non-aqueous sterile suspensions of the agent. These formulations may be supplied in unit or multi-dose 92 200836749 ^ 5 containers (eg sealed ampoules and vials) and may be stored in a cold liquid-dried form, such as water for injection, just prior to use. Dry) conditions. Injections and suspensions which can be used at any time can be prepared from sterile powders, granules, lozenges and the like. It will be appreciated that in addition to the ingredients specifically disclosed above, such formulations of the present invention may include other agents conventionally known in the art that have contemplated the type of formulation. • 10 For example, in terms of inhibiting the growth of cancerous cells, the present invention preferably combines at least one additional treatment, including but not limited to: chemotherapy, radiation therapy, and selective inhibition of Ras oncogenic signaling. Any other treatment known to those skilled in the art of treatment or management of the law, such as the administration of the drug. 15 • API-2 (Quxilide) salt can be administered. An example of a pharmaceutically acceptable salt is the use of an acid addition salt which forms a physiologically acceptable anion, such as a tosylate, an oxime sulfonate, an acetate, a salt, a malonate. , tartrate, succinate, benzoate, acid salt, alpha-ketoglutarate, and alpha-glycerol phosphate. It is also possible to form 2 suitable inorganic salts including hydrochloride, sulfate, sulphate and carbonate. 20 A barium acceptable salt can be obtained using standard procedures well known in the art, for example, by the use of a compound which is sufficient to produce a physiologically acceptable anion. It can also be used to test metals (such as sodium, potassium _) or soil-checking metals (for example, and salts. The citrifloxacin compound can be adapted to the selected (four) method, that is, oral. Or parenteral = 93 200836749 Various forms of pharmaceutical composition, intramuscular, topical or subcutaneous, and administered to patients (such as human or veterinary patients). The compound and bortezomib and its derivatives may be administered systemically with a pharmaceutically acceptable vehicle (i.e., a carrier), such as an inert diluent 5 or a ready-to-use plant, such as orally. Encapsulated in a hard or soft hub gelatin capsule, can be squeezed into a tablet or can be used directly with the patient's diet. For therapeutic oral administration, the compounds can be used in combination or in the form of the following: Use: Ingestible tablets, buccal tablets, buccal tablets, capsules, tinctures, suspensions, syrups, flakes, etc.

The percentage of the agent may of course vary and preferably may range from about 2 to about 60% by weight of the particular unit dosage form. Ingredients, therapeutically useful compositions of the active compound can be obtained in an effective amount. The tablets, buccal tablets, pills, capsules and the like may also contain the following components: arabic gum, corn starch or gelatin;

Binding agents, such as scutellaria, such as saponin II; 6 alginic acid; lubricants, fructose, lactose or pi 94 200836749 5 gin 10 15 20 acid methyl esters and benzopyrene Ingredients and seasonings, such as cherries and citrus seasonings. Of course, it is used for several times. [3] Any material prepared for any of the early dosage forms is musically acceptable and substantially non-toxic. In addition, the present invention can be used in sustained release formulations and devices. ^ Intravenous or intraperitoneal administration of the citrifloxacin compound and the _ card and its v. organism by infusion or injection. It may be selectively mixed with non-toxic surfactants in water to form such activators or solutions thereof. Dispersions can also be prepared in glycerol, liquid polyethylene glycols, mixtures of trisole and tris, and in oils. Under the conditions of storage and use, these preparations contain preservatives that prevent the growth of microorganisms. Such pharmaceutical dosage forms suitable for injection or infusion may include an aqueous aqueous solution or dispersion containing an active ingredient which is suitable for the preparation of a sterile injectable or infusible solution or suspension and optionally coated in the liposome or Sterile powder. The final dosage form must be sterile, fluid and stable under the conditions of manufacture and storage. The liquid carrier or vehicle may be a solvent or liquid dispersion medium including, for example, water, ethanol, polyol (eg, glycerol, propylene glycol, liquid polyethylene glycol, etc.), vegetable oil, non-toxic glycerin, and A suitable mixture. The proper fluidity can be maintained by, for example, by forming a liposome, by maintaining the desired particle size (in the case of a dispersion) or by using a surfactant. Microbial action can be prevented by various antibacterial and antifungal agents, for example, benzoic acid vinegar, chlorobutanol, hop, sorbic acid, thimerosal, and the like. In many cases, it may be preferred to include isotonic agents' such as sugars, buffers or gasified sodium. The absorption of such injectable compositions can be extended by the use of agents which delay absorption, such as aluminum monostearate 95 200836749 and gelatin, in such compositions. The sterile injectable solution is prepared by incorporating the necessary amount of the compound of the present invention and various other ingredients as appropriate into a suitable solvent, and if necessary, followed by filtration sterilization. For a sterile powder for the preparation of a sterile injectable solution, the preferred method is a vacuum drying and freeze drying technique which produces a powder having the active ingredient and any additional desired ingredients present in the previously sterilized solution. . For topical administration, the flucillin compound and bortezomib and its derivatives can be administered in pure form (i.e., liquid). However, they are administered to the skin in an amount appropriate to the composition or formulation of Group 10 and the dermatologically acceptable carrier (which may be a solid or a liquid). Useful solid carriers include finely divided solids such as talc, clay, microcrystalline cellulose, ceria, alumina, and the like. Useful liquid carriers include water, alcohol or ethylene glycol or a water-alcohol/ethylene glycol blend wherein the compounds of the present invention are selectively soluble or dispersed in an effective amount by means of a non-toxic surfactant. Adjuvants, such as perfumes and additional antimicrobial agents, may be added to optimize the properties of the particular application. The formed liquid composition applied by the self-absorbent pad can be used to impregnate the bandage and other dressings or spray it onto the infected area using a pump or aerosol sprayer. 20 A thickener, such as a synthetic polymer, a fatty acid, a fatty acid salt and an ester, a fatty alcohol, a modified cellulose or a modified mineral, may also be used in conjunction with the liquid carrier to form a direct application to the skin of the user. Can be spread, paste, gel, ointment, soap, etc. Useful dermatological compositions which can be used to deliver the compounds of the invention to the skin are disclosed in the following information. 96 200836749

Jacquet et al. (U.S. Patent No. 4,608,392), Geda (U.S. Patent No. 4,992,478), Smith et al. (U.S. Patent No. 4,559,157) and

Woltzman (U.S. Patent No. 4,820,508). The useful doses of these pharmaceutical compositions of the present invention can be determined by comparing their activity and in vivo activity in the animal model. Extrapolation of effective doses in mice, and other animals to humans is known in the art, for example, see U.S. Patent No. 4,938,949. In a non-limiting embodiment, the concentration of the active agent in the liquid composition (such as an emulsion) may range from about 0.1 to 25% by weight or from about 55 to 10% by weight. In one embodiment, the concentration in the semi-solid or solid composition (such as a gel or powder) may be from about 0.1 to 5% by weight, preferably from about 0.5 to 25 % by weight. In one embodiment, for an adult, a single dose suitable for injection, infusion or ingestion will typically vary between 5 and 15 mg and can be administered to 3 times per day to produce about 0.1 to 50 mg/ The dose of kilograms. A suitable non-limiting oral dose of the present invention may be between 7 and 45 mg per day, as appropriate for determining the weight of the individual 15. Accordingly, the present invention includes a pharmaceutical composition of a tripamicin compound and bortezomib and derivatives thereof or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier. A pharmaceutical composition suitable for oral, topical or parenteral administration comprising a quantity of a trichostatin compound and a bortezomib and a derivative thereof or a pharmaceutically acceptable salt thereof, constitutes a preferred embodiment of the present invention . In the context of the present invention, the dosage administered to a patient, particularly a human, should be sufficient to affect the patient's therapeutic response within a reasonable time frame. Those skilled in the art will understand that the dosage may depend on a variety of factors, including the health of the animal, 97 ‘the weight of the 36749 HAI animal, and the severity of the cancer and the stage of cancer. / The appropriate dose is such that the concentration of the tresibine compound and bortezomib and the two organisms in the known tumor tissue can affect the desired reaction, and the sputum 1 is the maximum inhibition of cancer cell growth. A dose that does not act as a side effect of the control. API-2 (or a pharmaceutically acceptable salt thereof) can be administered at different intervals, jN*, as determined by the ordinarily skilled artisan. Milk, Z is beneficial to the methods used to inhibit the growth of cancer cells, but not limited to: primates, such as baboons, chimpanzees, accommodating, human faces, 10 15 squirrels, wolves Sons, domesticated animals (such as pets), such as dogs, cats, days such as cattle ^, Vietnamese pot-bellied pigs, rabbits, and farm animals, on the cattle, horses, donkeys, pigs, sheep, and goats; typical 4 Beishikou, lion, old zebra, wild bee/cooma, rhinoceros, giraffe, antelope, sloth, gazelle, book seedling, hyena, kangaroo, kangaroo, kangaroo, possum, raccoon, bear and Whales ^ i ▲ Seals, sea lions, elephant seals, otters, porpoises, dolphins, meanings, the term "patients, and "patients," are used interchangeably in the text and have breastfeeding human and non-human mammal species. Similarly, the in vitro method of the present invention can be carried out on such cells. Standard technical identification known to the alum professional requires a patient to be treated using the method of the present invention. The following examples are used for clarification and are not limiting. Example Example 1: In Vitro Screening

: All cell lines are commercially available from ATCC 20 200836749 or as previously described (Cheng, J. Q., et al. 〇nc〇gene, 14: 2793-2801, 1997, West, KA, et al. Drug Resist. Updat., 5: 234-248, 2002, Satyamoorthy, K., et al. Cancer Res. 61: 7318-7324, 2001). The NCI structural demultiplexing molecular library is a repository of 1,992 compounds selected from the NCI drug library of approximately 140,000 compounds. Further information on the selection, structure, and activity of compounds in these diverse molecular libraries can be found on the NCI Developmental Therapeutics Program website.

Screening for inhibition of Akt-transformed cell growth: Inoculation of AKT2-transformed NIH3T3 cells or LXSN vector-infected NIH3T3 control cells 10 (Cheng, JQ, et al. Oncogene, 14: 2793-2801, 1997) Into the 96 well tissue culture plate. After treatment with 5 μM of the NCI diversity library compound, cell growth can be detected using the CellTier 96 One Solution Cell Proliferation kit (Promega). Compounds that inhibit AKT2 transfected cells but not LXSN-transfected NIH3T3 cells are considered to be suitable for Akt inhibitors and subjected to further analysis. In vitro protein kinase, cell survival and cell death assay: In vitro kinase assays were performed as previously described (see, for example, Jiang, K., Coppola, et al. Mol. Cell Biol, 20: 139-148, 2000). Cell survival was assayed by MTS (Promega). Apoptosis is detected by Annexin V, and the 20-line method is as described above (Jiang, K., Coppola, et al. Mol_ Cell Biol, 20: 139-148, 2000). Recombinant Akt and PDK1 are purchased from Upstate Biotechnology Inc 〇 Conclusion

Akt signaling pathway, identification of small molecule inhibitors of ΑΠ-2: Akt in humans 99 200836749 The frequent changes in carcinoids are detected and the destruction of the Akt pathway induces apoptosis and inhibits tumor growth (Jetzt, A, et al. Cancer Res., 63: 697-706, 2003). Therefore, Akt is considered an attractive molecular target for the development of novel cancer therapies. In order to identify small molecule inhibitors (groups) of Akt, it was evaluated that the chemical library of 1,992 compounds from the NCI (NCI Diversity Molecular Library) is suitable for inhibiting AKT2 transfected cell growth rather than emptying media. Agent LXSN transfers the agent of infected NIH3T3 cells. Repeated experiments demonstrated that 32 compounds only inhibited the growth of AKT-2 transformed cells. One of the most potent compounds, API-2 (NCI identification number: NSC 154020), inhibits cell growth at a concentration of 50 nM. Figure 1A shows API-2, also known as the chemical structure of Tricinem (Schweinsberg, P. D., et al. Biochem Pharmacol, 30: 2521-2526, 1981) 'API-2 selective preferential inhibition The fact that AKT-2 transfected cells, but not untransformed parental cells, helps us determine whether API-2 is an inhibitor of AKT2 kinase. Accordingly, AKT2 was immunoprecipitated using an anti-AKT2 antibody derived from AKT-2 transmorphic NIH3T3 cells treated with API-2. The AKT2 immunoprecipitate was immunized with an anti-phosphorylated Akt antibody. As shown in Figure 1B, API-2 significantly inhibits the phosphorylation of sulphate-309 and serine-474, which are required for complete activation of AKT-2 (0&like, 8.11., et al. Genes Dev. 13: 2905-2927, 1999). Since the three variants of Akt have a high homology and similar structure, the effect of API-2 on their kinase activity can be assessed. The HEK293 cell line was infected with HA-Akt1, HA-AKT2 and HA-AKT3, and the serum was starved overnight and treated with API-2 before stimulation with EGF (50 ng/ml), which took 60 minutes. Three replicate experiments showed that API-2 inhibited EGF-induced kinase activity and sarcoylation of Aktl, AKT2, and AKT3 100 200836749 (Fig. 1C). However, in the in vitro kinase reaction, the kinase activity of recombinant pro-ogenic AKT2 (Myr_AKT2) was not inhibited by API-2 (Fig. 1D), which indicates that API-2 does not directly inhibit Akt in vitro and It is indicated that API-2 is neither a competitor to ATP nor a matrix competitor that can bind to the active site of Akt 5. API-2 does not inhibit the known upstream activation sword of Akt: it has been clearly documented that Akt can be activated via extracellular stimuli and intracellular signaling molecules, such as active Ras and Src, via a P13K-dependent manner. Thus, the API-2 inhibition of Akt can result from the labeling of upstream molecules (groups) of Akt. Since 10 PI3K and PDK1 are direct upstream regulators of Akt (Datta, S. R., et al. Genes Dev. 13: 2905-2927, 1999), it is possible to check whether API-2 can inhibit PI3K and/or PDK1. The serum of HEK293 cells was starved and then treated with API-2 or PI3K inhibitor, wortmannin, for 30 minutes prior to EGF stimulation. PI3K was immunoprecipitated with an anti-ρΙΙΟα antibody. The immunoprecipitates were subjected to an in vitro Κ3Κ kinase assay using PI-4-P 15 as a substrate. As shown in Fig. 2, the EGF-induced ΡΙ3Κ activity was inhibited by wortmannin and was not inhibited by ΑΡΙ-2. In order to evaluate the effect of ΑΡΙ-2 on PDK1, a assay in which recombinant PDK1 promotes the phosphorylation of threonine-309 of ΑΚΤ2 peptide is used in the presence of phospholipid erythritol-containing liposomes. As shown in Figure 2, the assay was strongly inhibited by the control PDK1 inhibitor, staurosporine (IC50 = 5ηΜ). Conversely, at the highest concentration tested (5·1 μΜ), API-2 showed only 21% inhibition of the assay. To further assess the effect of API-2 on PDK1 activation, after treatment of HEK293 cells with API_2, PDK1 was examined for serine-241, which is a residue that is itself phosphorylated and is important for its activity in 101 200836749 Autochemical acidification content (Datta, SR, et al. Genes Dev. 13: 2905-2927, 1999). Three replicate experiments demonstrated that the phosphorylation level of PDK1 was not inhibited by API-2 (Fig. 2B). However, the PI3K inhibitor, wortmannin, inhibits EGF-stimulated PDK1 (Fig. 2B). 5 The selectivity of API-2 for Akt is more than that of PKC, PKA, SGK, STAT, JNK, p38, and ERK. The Akt belongs to the AGC (PKA/PKG/PKC) kinase family. Also included are PKA, PKC, serum- and adrenal glucocorticoid-inducible kinase (SGK), p90 ribosomal S6 kinase, p70S6K, mitogen, and stress-activated protein 10 kinase and PKC- Related kinases. In the AGC kinase family, the protein structures of PKA, PKC and SGK are relatively close to Akt kinase and not other members. Therefore, the effect of API-2 on the enzymatic activity of these three kinases was examined next. HEK393 cells were transfected with HA-labeled PKA, PKCa or SGK. In vitro kinase assay and immunoblot analysis demonstrated that the kinase 15 activity of PKA and PKC α was inhibited by ΡΚΑΙ and R〇31-8220, respectively, which are PKC inhibitors, whereas API-2 showed no activity against them. Impact (2C and 2E). Moreover, serum-induced SGK kinase activity was attenuated by wortmannin but not by API-2 (Fig. 2D). In addition, it has been determined whether API-2 has an effect on other carcinogenic survival pathways. Western blot analysis using commercially available anti-phosphorylated antibodies showed that the acidification levels of Stat3, JNK, p38, and Erkl/2 were not affected by API-2 treatment (Fig. 2F. These data indicate that ΑΠ-2 can be Specific inhibition of Akt signaling pathways. API-2 inhibits cell growth and induces Akt overexpression/activation of apoptosis in human cancer cell lines: API-2 selectively inhibits this Akt pathway 102 200836749 Apoptosis in such tumor cells that inhibits proliferation and/or preferentially induces abnormal expression/activation of Akt. Since Akt is active in human malignancies that are usually caused by overexpression of Akt or PTEN mutations, API-2 can be used to treat cells that exhibit orthostatic Akt produced by overexpression of AKT2 • 5 (OVCAR3, OVCAR8, PNNC1 and AKT2-transformed NIH3T3) or by mutations in the PTEN gene Cells producing orthostatic Akt (PC-3, LNCaP, MDA-MB-468), and cells that do not respond to growth stimulation of IGF-1 (OVCAR5, DU-145, T47D, C0L0357, and LXSN-NIH3T3) 10 and borrowed to live Melanoma cells activated by IGF-1 of Akt but not responsive to growth stimulation of IGF-1 (Satyamoorthy, K., et al. Cancer Res. 61: 7318-7324, 2001). Immunoblotting analysis showed phosphorylation of Akt The content is only inhibited by API-2 in cells that exhibit elevated Akt or respond to IGF-1 stimulation (Fig. 3A). Therefore, compared to 15 with low Akt content, Akt expression In activated cells, API-2 can greatly inhibit the growth of fine cells. As shown in Figure 3, in Akt-expressing/activated cell lines, LNCaP, PC-3, OVCAR3, OVCA8, PANC1, MDA- API-2 treatment in MB-468 and WM35 inhibited cell proliferation of approximately 50 to 60%, whereas 20 DU145, OVCAR5, C0L0357, which showed low Akt content or did not respond to growth stimulation of IG F-1, Only about 10 to 20% inhibition was observed in T47D and WM852 cells. Furthermore, API-2 induced apoptosis in cell lines of 8x, 6x, 6x, and 3x, respectively: OVCAR3, OVCAR8, PANC1, and AKT2 -NIH3T3. OCCAR5, C0L0357 and LXSN_NIH3 T3 cells treated with API-2 and vehicle (DMSO) did not find fine 103 200836749 apoptosis obvious difference. Therefore, API-2 inhibits cell growth and preferentially induces apoptosis of cells capable of expressing abnormal Akt. API-2 inhibits downstream targets of Akt: Akt has been shown to exert its cellular effects via phosphorylation of many proteins (Datta, S.R., et al. 5 Genes Dev. 13:2905-2927, 1999). More than 20 proteins have been identified as Akt matrices, including members of the Forkhead family of proteins (FKHR, AFX, and FKHRL1), potato globulin (tuberlin)/TSC2, p70S6K, GSK-3/3, p21WAF1/Cipl, p27kipl, MDM2. Bad, ASK1, and ΙΚΚα, etc. Next, check if API-2 can suppress the downstream of Akt. Since the anti-squaric acid 10, sBad, AFX, and -GSK-3a/yS anti-systems are commercially available, the effect of API-2 on their phosphorylation reaction induced by Akt can be measured. After treatment with ΑΡΙ·2 (1 μΜ), OVAR3 cells were lysed and the ink spots were immunized with respective anti-phosphorylated antibodies. Figure 4 shows that ΑΡΙ-2 can greatly inhibit the degree of acidification of the potato globulin, which can lead to the stability and upregulation of the potato protein 15 (Dan, HC, et al. J. Biol. Chem., 277:35364 -35370, 2002). The degree of phosphorylation of Bad, GSK-3 /3, and AFX was partially attenuated by API-2. These data indicate that API-2 induces cell death and controls cell growth by inhibiting phosphorylation of its downstream targets. Different degrees of ΑΠ-2 inhibition in the downstream of Akt may result from the fact that the phosphorylation sites of these 20 targets are also controlled by other kinases (groups), for example, in addition to Akt, ΡΑΚΙ can also be Bad-serine- 136 Phosphorylation (Schurmann, A., et al. Mol. Cell. Biol 20: 453-461, 2000). Example 2: Anti-tumor activity of nude mouse tumor xenograft models As previously reported, tumor cells were taken to enable It was suspended in PBS and injected subcutaneously into the right flank and left flank (2χΐ〇6 cells/flank) of an 8-week-old female nude mouse at 104200836749 (Sirn, J., Blaskovic, et al. Cancer Res·, 59). :4919-4926, 1999). When the tumor reaches about 100 to 150 mm 3 , the animals are randomly sampled and administered intraperitoneally daily with 2 ml of the trichostatin compound and/or 5 onioncycline. A vehicle for the analog. The control animals received DMSO (20%) vehicle, whereas the treated animals were injected with a solution of API-2 (1 mg/kg/day) in 20% DMSO. API_2 inhibited energy. The tumor peak of Akt's Zen rats has been shown to be AKT1 and AKT2 in human ovarian and pancreatic cancer. Often 10 degrees of expression/activation and/or amplification (Cheng, JQ, and Nicosia, SV· AKT signal transduction pathway in oncogenesis· In Schwab D? Editor, Encyclopedic Reference of Cancer. Berlin Heidelberg and New York: Springer 2001· pp35-7). Inhibition of Akt pathway by PI3K, HSP70, Src and Farnesyltransferase inhibits Akt pathway and induces apoptosis (s〇lit, DB·, et al. Cancer Res., 63: 2139-2144, 2003, Xu, W" et al., Cancer Res., 63: 7777-7784, 2003). Recent studies have shown that intratumoral injection of adrenal adenovirus with dominant negative Akt can also significantly inhibit tumor growth with high Akt xenografts (Jetzt, A., et al. Cancer Res., 20 63: 697-706). , 2003). Because API-2 inhibits Akt signaling and induces apoptosis and cell growth control only in cancer cells with high Akt content, tumor growth with South Akt-1 in tumors is less than tumor growth with low Akt content. More sensitive to API-2. Accordingly, subcutaneously injected super-expressing cells (OVCAR3, OVCAR8, and PANC-1) were subcutaneously implanted into the right 105 200836749 of the mouse, and these cells exhibiting low Akt content (OVCAR5 and C0L0357) were implanted. Left flank in the abdomen. When tumors averaged about 1 to 15 mm 3 , the animals were randomized and treated intraperitoneally with vehicle or API-2 (1 mg/kg/day). As illustrated in Figure 4B, at 49 days after tumor implantation, 5 vehicle-treated VCAR_5 and C0L0357 tumors can grow to approximately 800-1,000 mm3. The OVCAR3, OVCAR8, and PANClIt tumors treated with the vehicle control were grown to about 7.900 mm 3 49 days after tumor implantation. API-2 inhibited tumor growth of 90% of OVCAR3, 88% of OVCAR8, and 80% of PANC1, respectively. Conversely, API-2 had minimal effect on the growth of OVCAR5 and 10 C0L0357 cells in nude mice (Fig. 4B to 4D and did not show data). At a dose of 1 mg/kg/day, ΑΡΙ·2 had no effect on blood glucose content, body weight, activity and food intake in mice. In the treated tumor samples, Akt activity was inhibited by ΑΡΙ-2 and did not alter the total Akt content (Fig. 4E). Taken together, these results indicate that API-2 selectively inhibits the growth of tumors with 15 high Akt levels. Example 3: TCN directly inhibits wild-type Akt kinase activity API-2 (TCN) directly inhibits wild-type Akt kinase activity induced by in vitro PDK1 (Fig. 1). This result confirms that API-2 is a direct Akt inhibitor and confirms that the basic mechanism may be that API-2 binds to the PH domain of Akt and / 20 or sulphate _308. In vitro kinase assays were performed using PDK1 and Akt in recombinants containing a lipid-filled inositol • 3,4,5-P3 (PIP3), API-2, and a kinase buffer as a matrix tissue protein. After incubation for 30 minutes, the reactants were separated by SDS-PAGE and exposed to the film. Example 4: TCN is effective against cancer-resistant cells 106 200836749 Testing tCN in cisplatin, paclitaxel, and tamoxifen-resistant A27〇cp, C-13, OVCAR433, and MCF7/TAM cells (API- 2) The utility. API_2 overcomes the resistance of these cells to cisplatin, paclitaxel, and tamoxifen. 5 The present invention has been described with reference to its preferred embodiment. Variations and modifications of the present invention will become apparent to those skilled in the art from this disclosure. All such variations and modifications are intended to be included within the scope of the present invention. BRIEF DESCRIPTION OF THE DRAWINGS Fig. 1 is a diagram showing the identification of a suitable substance of Akt inhibitor obtained from NCI Diversification 10 (Diversity Set). The first figure illustrates the chemical structure of API-2 (Quxi Libin). Figure 1B illustrates the phosphorylation procedure of Αρμ2 for inhibition of ΑΚΤ2 in ΝΙΗ3Τ3 cells transformed with ΑΚΤ2. Treatment of wild-type ΑΚΤ2-transformed ΝΙΗ3Τ3 cells with ΑΡΙ-2 (1 μΜ) was time-consuming, and immunoblot analysis was performed using anti-acidified Akt-T308 and -S473 antibody 15 (top group and intermediate group). . The bottom group shows the performance of the total AKT2. In the ic diagram, it is shown that API-2 inhibits three variants of Akt. HEK293 cells were transfected with HA-Akt1, -AKT2 and -AKT3 and treated with ΑΡΙ-2 (1 μΜ) or wortmannin (15 pM) before EGF stimulation, so that the cells were solubilized and resistant to anti-ha 20 body immune smothering. The immunoprecipitates were subjected to an in vitro kinase assay (top) and immunoblot analysis was performed using an anti-shitanylated Akt-T308 (bottom) antibody. The middle group shows the expressiveness of the metastatically infected Aktl, AKT2 and AKT3. Figure 1D illustrates that API-2 does not inhibit Akt in vitro. The activity of the intrinsic activity of the recombinant protein in the kinase buffer containing ΙμΜ ΑΠ-2 (lane 3) was carried out 107 200836749 In vitro kinase assay. Figure 2 illustrates that API-2 does not inhibit PI3K, PDK1, and closely related members of the AGC kinase family. Figure 2A illustrates an in vitro PI3K kinase assay. Serum deficiency of HEK293 cells and treatment with ΑΠ-2 (1 μΜ) 5 or wortmannin (15 μΜ) before EGF stimulation took 30 minutes. These immunoprecipitates were subjected to an in vitro kinase assay using ΡΙ-4-Ρ as a substrate. The second plot illustrates the effect of ΑΡΙ-2 on PDK1 activation in vitro (the top group), and the solid circle indicates inhibition by ΑΡΙ_2. The open circle indicates inhibition by the positive control staurosporine, which is a potent PDK1 inhibitor 10 (IC50 = 5 nM). The bottom group is

Immunoblot analysis of Myk-PDK4 metastatic infection and HEK293 cells treated with wortmannin or API-2 prior to EGF stimulation. The immunoblot method is detected by the designated antibody. Figure 2C illustrates the immunoblotting of PKCa phosphorylation using anti-phosphorylated PKC α T638 (top) and assembly 15 PKCa (bottom) antibodies after treatment with API-2 or non-selective PKC inhibitor Ro31_8220 analysis. Figure 2D shows an in vitro SGK kinase assay. HEK293 cells were metastasized via HA-SGK transfer and treated with API-2 or wortmannin prior to EGF stimulation. The in vitro kinase assay was performed using HAP as a substrate (top) with HA-SGK immunoprecipitates. The bottom panel represents the table 20 of the HA-SGK that has been transferred. Figure 2E illustrates the results of the PKA kinase assay. Immunopurified PKA was grown in ADB buffer (Upstate Biotechnology Inc) containing the indicated inhibitor (API-2 or AKAI) and matrix 'Kemptide'. The kinase activity was quantified. The 2F figure represents the Western dot method. Treatment of 0VCAR3 cells with API-2 took time to specify. Immunoblots of cell lysates were performed with the indicated anti-phospho antibodies (Nos. 1 108 200836749 to 4) and anti-actin antibodies (bottom). Figure 3 is a diagram showing that API-2 inhibits Akt activity and cell growth and can induce apoptosis of human cancer cells with high Akt. Figure 3A shows Western blotting. After treatment with API-2, the phosphorylation of Akt was detected by anti-phospho-5 Akt-T308 antibody in designated human cancer cell lines. These dot methods were then probed with anti-total Akt antibodies (the bottom panel). Figure 3B shows a cell proliferation assay. The cell lines shown in the figure were treated with different doses of API-2, which took 24 hours and 48 hours, and then analyzed by the CellTiter 96 Cell Proliferation Assay Kit (Promega). Figure 3C provides an analysis of apoptosis. 10 cells were treated with API-2 and stained with annexin V and PI, and then analyzed by FACScan. Figure 4 shows that API-2 inhibits downstream targets of Akt and displays anti-tumor organisms in cancer cell lines with high Akt in mouse xenografts. Figure 4A shows that API-2 inhibits Akt phosphorylation of potatorin, Bad, AFX and GSK-3 stones. After treatment with API-2, OVAR3 15 cells were lysed and the ink spots were immunized with the indicated antibodies. Figure 4B shows that API-2 inhibits tumor growth. Tumor cells were injected subcutaneously into nude mice with low Akt cell content on the left and high Akt cell content on the right. When the tumors reached an average size of about 1 to 150 mm 3 , the animals were treated with vehicle or 1 mg/kg/day of API-2. Each measured value represents the average of 10 tumors. Figure 4C is a graphical representation of mice treated with 20 API.2 or vehicle (control) with OVCAR3 (right) and OVCAR5 (left) xenografts. Fig. 4D shows the tumor size (bottom) and weight (top) at the end of the experiment. In Figure 4E, anti-phospho-Akt-S473 (top) and anti-AKT2 (bottom) antibodies were used in treated (T3 and T4) and untreated (T1 and T2) OVCAR-3-derived tumors. Cancer lysate exemption 109 200836749 Epidemic point analysis. Figure 5 shows that ΑΠ-2 (Cucidine) inhibits in vitro kinase activity. In vitro kinase assays were performed in recombinant kinases containing PDK1 and Akt in a kinase buffer containing phosphatidylinositol-3,4,5-Ρ3(ΡΙΡ3), ΑΡΙ-2 and histoprotein as matrix. After 30 minutes of incubation, the aliquots were separated by SDS-PAGE and exposed to the film. Figures 6a to c provide mRNA and amino acid sequences of human Aktl, as well as restriction enzyme positions. Figures 7a to d provide the mRNA and amino acid sequences of human Akt2, as well as the position of the restriction enzyme. Figures 8a to c provide mRNA and amino acid sequences of human Akt3, as well as restriction enzyme positions. Figure 9 is a graph showing the effect of bortezomib synergistically enhanced API-2 (TCN) on these viable myeloma cells. H929 cells (derived multiple bone myeloma) were treated with 0, 2.5, 5, 10, 20 or 40 nM bortezomib only in the presence of ADIμΜ ADI-2 or only 〇, 2.5, 5, 1〇 , 2〇 or 4〇0]^八〇1-2 treatment. Calibrate and plot cell viability (Figure 9A). U266 cells (derived multiple myeloma) are treated with 〇, 2.5, 5, 10, 20 or 40 μΜ ADI-2 only in the presence of ΙΟηΜ API-2 or only 0, 2.5, 5, 10, 20 or 40 nM boron was treated with 20 zomi. Calibrate and plot cell viability (Fig. 9B). Figure 1 shows the synergistic effect of bortezomib and API-2 (TCN) on enveloped cell lymphoma cells. Making Jeko-Ι cells (derived enveloped cell lymphoma) treated with 〇, 2.5, 5, 10, 20, 40 or 80 Μ bortezomib in the presence of 5 μΜ API-2 or via 〇, ΐ·25, 2·5, 5, 1〇, 2(^40μΜΑΡΙ-2 200836749 treatment. Calibrate and illustrate cell viability. [Main component symbol description] (none)

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Claims (1)

200836749 X. Patent application scope: 1. A composition comprising: (i) a compound of formula I-IV: 5 Μ ΚΙ Wherein R 2 ', R 3 ' and R 5 ' are each independently hydrogen, optionally substituted phosphate or phosphonate (which includes mono-, di- or triphosphate or diazepam 10 prodrugs); a group (which includes a lower fluorenyl group); an alkyl group (which includes a lower carbon group); an acid amine, a hydrazine group, which includes a decyl group or an aryl group; and a thiol group, 112 200836749 And a f group, wherein the phenyl group is optionally substituted with one or more substituents as described, for example, in the definition of an aryl group as taught herein; a selectively substituted sulfhydryl group; a lipid, including a lin Fatty; amino acid; carbohydrate; hydrazine; or biliary; or other pharmaceutically acceptable compounds in which R2, R3' and R5' are independently H or mono-, di- or tri-steanate Accepting a leaving group; wherein Rx and Ry are independently hydrogen, a selectively substituted phosphate; a fluorenyl group (which includes a lower sulfhydryl group); a guanamine, an alkyl group (which includes a lower alkyl group); An olefin such as polyethylene glycol, a selectively substituted aromatic sulfonyl alcohol; a lipid comprising a phospholipid; an amino acid; carbon water Thereof; peptide, or cholesterol; or other pharmaceutically acceptable leaving group. In one embodiment, the compound is administered as a 5,-phosphate ether lipid or a 5,-ether lipid; Ri and R 2 are each independently a linear, optionally substituted linear, branched or cyclic alkyl group ( It includes lower alkyl), alkenyl or alkynyl, CO-alkyl, co-alkenyl, co-alkynyl, co-aryl or heteroaryl, c〇-alkoxyalkyl, CO-aryloxy Alkyl, co-substituted aryl, sulfonyl, alkanesulfonyl, arylsulfonyl, aralkyl sulfonyl; (ii) a compound of formula V: 113 200836749 wherein l, R 2 , R 3 , R 4 , and R 5 are each independently oxime, optionally halogenated, substituted straight chain, branched or cyclic alkyl (which includes lower alkyl), alkoxy, alkenyl Or alkynyl, aryl, CO-alkyl, CO-alkenyl, CO-5 alkynyl, CO-aryl or heteroaryl, CO-alkoxyalkyl, CO-aryloxyalkyl, CO-substituted An aryl group, a fluorenyl group, a decyl group, a aryl group, an aralkyl sulfonyl group; and (iii) a pharmaceutically acceptable carrier. 10 2. The composition of claim 1, wherein the compound of formula I is triclinide. 3. The composition of claim 1, wherein the compound of formula I is triclinide phosphate. 4. The composition of claim 1, wherein the compound of formula I is tromethamine phosphonium monohydrate. 5. The composition of claim 1 wherein the bortezomib and its derivatives have the formula: % β% ..., ? ? 9« Η Ο γ wherein at least one of rLr5 is -Η. 20 6. The composition of claim 1 wherein the bortezomib and its 114 200836749 derivative have the formula: 7. The composition of claim 1, wherein the compound of formula I is present in a dose of at least 10 mg/m2. 5. The composition of claim 1, wherein the compound of formula II is present at a dose of at least 10 mg/m2. 9. The composition of claim 1 is suitable for parenteral administration. 10. The composition of claim 9, wherein the parenteral administration is intravenous administration. 10 11. The composition of claim 1 of the patent application, which is suitable for oral administration. 12. The composition of claim 1 of the patent application, which is suitable for topical administration. 13. A method of treating a tumor or cancer in a mammal comprising administering to the mammal an effective amount of a composition comprising: (1) a compound of formula I-IV, · 115 €13⁄4.200836749 01-3⁄4 Wherein R 2 ', R 3 ' and R 5 ' are each independently hydrogen, optionally substituted phosphate or phosphonate (which includes mono-, di- or triphosphate or stabilized phosphate prodrugs); It includes a low carbon fluorenyl group; an alkyl group (which includes a low carbon alkyl group); a decylamine, a sulfonic acid group, which includes a alkyl group or an aryl group; and a sulfhydryl group, which includes a methylsulfonyl group and a benzyl group, Wherein the phenyl group is optionally substituted with one or more substituents as described, for example, in the definition of the aryl group as taught herein; optionally substituted azulsulfonyl; lipids, including phospholipids; 200836749 Amino acid; carbohydrate; peptide; or cholesterol; or other pharmaceutically acceptable cleavage group in which R2', R3' and R5' are independently a compound of hydrazine or mono-, di- or tri-phosphate. Wherein Rx and Ry are independently hydrogen, optionally substituted phosphate; fluorenyl (which includes lower sulfhydryl); decylamine, alkyl (which includes lower alkyl); aromatic polyalkylene oxide, such as Polyethylene glycol, optionally substituted arylsulfonyl; lipids, including phospholipids; amino acids; carbohydrates; peptides; or cholesterol; or other pharmaceutically acceptable leaving groups. In one embodiment, the compound is administered as a 5'-phosphate ether lipid or a 5'-ether lipid; the heart and the oxime are each independently a linear, optionally branched, linear, branched or cyclic alkyl group ( It includes lower alkyl, alkenyl or alkynyl, CO-alkyl, CO-alkenyl, CO alkynyl, CO-aryl or heteroaryl, CO-alkoxyalkyl, CO_aryloxyalkyl , a CO-substituted aryl group, a sulfonyl group, an alkanesulfonyl group, an arylsulfonyl group, an arylalkylsulfonyl group; and (ii) a compound of the formula V: R!, R2, R3, R4, and R5 are each independently oxime, selectively halogenated, 117 200836749 ««n Μ chain or ring (tetra) group (which includes low wei group), alkoxy group, alkenyl group or alkynyl group, Aryl, CO-alkyl, CO-alkenyl, CO-silk, CO-aryl or heteroaryl, c〇-oxyalkyl, c〇_aryloxyalkyl, CO-substituted aryl, Sulfonyl, alkanesulfonyl, arylsulfonyl, 5-party calcined base. 14. The method of claim 13, wherein the compound of formula I-IV and the compound of formula V are administered simultaneously. 15. The method of claim 13, wherein the compound of formula I-IV is administered first, followed by the compound of formula V. 10 16. The method of claim 13, wherein the compound of formula V is administered first, followed by administration of the MV compound. 17. The method of claim 13, wherein the compound of formula I-IV is administered once a week for 3 weeks, and then the compound is not administered over a period of one week. 15 18. The method of claim 13, wherein the compound of formula ν is administered once a week for 3 weeks, and then the compound is not administered for a period of one week. Ϊ́9. The method of claim 14, wherein the dosing schedule is repeated at least twice. 20 20. The method of claim </RTI> wherein the dosing schedule is repeated at least four times. 21. The method of claim 1, wherein the treated tumor is a court, pancreas, ovary, and colorectal tumor. 22. The method of claim 13, wherein the compound of the formula IV-IV is administered at least 10 mg/m 2 118 200836749. 23. The method of claim 13, wherein the compound of formula I-IV is administered at 10 mg/m2 or less. 24. The method of claim 13, wherein at least 10 mg 5 /m 2 of the compound of formula V is administered. 25. The method of claim 13, wherein the compound of the formula V is administered at 10 mg/m 2 or less. 26. The method of claim 13, wherein the compound of the formula V10 is administered in an amount of 10 mg/m2 or less in a large number of intravenous injections of 3 to 5 seconds twice a week for 2 weeks. 119