WO2006089150A2 - Agents anti-angiogeniques comportant de l'aldesleukine - Google Patents

Agents anti-angiogeniques comportant de l'aldesleukine Download PDF

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WO2006089150A2
WO2006089150A2 PCT/US2006/005720 US2006005720W WO2006089150A2 WO 2006089150 A2 WO2006089150 A2 WO 2006089150A2 US 2006005720 W US2006005720 W US 2006005720W WO 2006089150 A2 WO2006089150 A2 WO 2006089150A2
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aldesleukin
ril
amine
cells
des
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PCT/US2006/005720
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WO2006089150A3 (fr
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Sharon Lea Aukerman
Kimberly Denis-Mize
Laurence Elias
Bahija Jallal
Daniel Menezes
Gary W. Witherell
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Novartis Vaccines And Diagnostics Inc.
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Priority to JP2007556337A priority Critical patent/JP2008530239A/ja
Priority to AU2006214138A priority patent/AU2006214138A1/en
Priority to CA002598448A priority patent/CA2598448A1/fr
Priority to MX2007010037A priority patent/MX2007010037A/es
Priority to EP06735400A priority patent/EP1853302A2/fr
Priority to BRPI0608880-5A priority patent/BRPI0608880A2/pt
Publication of WO2006089150A2 publication Critical patent/WO2006089150A2/fr
Publication of WO2006089150A3 publication Critical patent/WO2006089150A3/fr

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • A61K38/19—Cytokines; Lymphokines; Interferons
    • A61K38/20—Interleukins [IL]
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/50—Pyridazines; Hydrogenated pyridazines
    • A61K31/502—Pyridazines; Hydrogenated pyridazines ortho- or peri-condensed with carbocyclic ring systems, e.g. cinnoline, phthalazine
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/40—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with one nitrogen as the only ring hetero atom, e.g. sulpiride, succinimide, tolmetin, buflomedil
    • A61K31/403—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with one nitrogen as the only ring hetero atom, e.g. sulpiride, succinimide, tolmetin, buflomedil condensed with carbocyclic rings, e.g. carbazole
    • A61K31/404—Indoles, e.g. pindolol
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/44—Non condensed pyridines; Hydrogenated derivatives thereof
    • A61K31/4412—Non condensed pyridines; Hydrogenated derivatives thereof having oxo groups directly attached to the heterocyclic ring
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/505—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim
    • A61K31/517—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim ortho- or peri-condensed with carbocyclic ring systems, e.g. quinazoline, perimidine
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • A61K38/19—Cytokines; Lymphokines; Interferons
    • A61K38/20—Interleukins [IL]
    • A61K38/2013—IL-2
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P37/00—Drugs for immunological or allergic disorders
    • A61P37/02—Immunomodulators
    • A61P37/04—Immunostimulants
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P9/00—Drugs for disorders of the cardiovascular system
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P9/00—Drugs for disorders of the cardiovascular system
    • A61P9/02—Non-specific cardiovascular stimulants, e.g. drugs for syncope, antihypotensives
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P9/00—Drugs for disorders of the cardiovascular system
    • A61P9/12—Antihypertensives

Definitions

  • the present invention relates to methods of therapy for diseases associated with abnormal cellular proliferation.
  • recombinant IL-2 is combined with antangiogenic agents for use in the treatment of cancer.
  • Capillaries reach into almost all tissues of the human body and supply tissues with oxygen and nutrients as well as removing waste products. Under typical conditions, the endothelial cells lining the capillaries do not divide, and capillaries, therefore, do not normally increase in number or size in a human adult. Under certain conditions, however, such as when a tissue is damaged, or during certain parts of the menstrual cycle, the capillaries begin to proliferate rapidly. This process of forming new capillaries from pre-existing blood vessels is known as angiogenesis or neovascularization. See Folkman, J. Scientific American 275, 150-154 (1996). Angiogenesis during wound healing is an example of pathophysiological neovascularization during adult life.
  • the additional capillaries provide a supply of oxygen and nutrients, promote granulation tissue, and aid in waste removal. After termination of the healing process, the capillaries normally regress. Lymboussaki, A. "Vascular Endothelial Growth Factors and their Receptors in Embryos, Adults, and in Tumors" Academic Dissertation, University of Helsinki, Molecular/Cancer Biology Laboratory and Department of Pathology, Haartman Institute, (1999).
  • Angiogenesis also plays an important role in the growth of cancer cells. It is known that once a nest of cancer cells reaches a certain size, roughly 1 to 2 mm in diameter, the cancer cells must develop a blood supply in order for the tumor to grow larger as ⁇ iltusion will not be sufficient to supply the cancer cells with enough oxygen and nutrients. Thus, inhibition of angiogenesis is expected to halt the growth of cancer cells.
  • VEGF vascular endothelial growth factor
  • EGF epidermal growth factor
  • FGF fibroblast growth factor
  • PDGF platelet derived growth factor
  • anti-tumor immunity involves activation of multiple cell types in the immune system, the most efficient being cytolytic T lymphocytes.
  • cytolytic T lymphocytes To induce specific, anti-tumor T lymphocyte mediated immune response, recognition of not only the tumor antigen of interest, but also costimulatory interactions between specific ligands present on either the tumor cell or the antigen presenting cell, and the target T lymphocytes are required.
  • This second, co-stimulatory signal may also be provided by soluble factors, such as cytokines or other peptide molecules which bind to specific, cell surface receptors and initiate various signal transduction pathways, resulting in augmentation of effector function.
  • Interleukin-2 is a T lymphocyte-derived cytokine which binds to specific receptors present on T cells and natural killer (NK) cells, and will activate them for tumor cytolysis, cytokine secretion, and other effector functions.
  • Both CD4 and CD8 T lymphocytes express the receptor for IL-2, and develop increased cytolytic effector and cytokine synthetic function after exposure to the biologic.
  • the functional effects of IL-2 are mediated via engagement with the IL-2 receptor (IL- 2R), which is structurally composed of three subunits: ⁇ , ⁇ , and a common ⁇ chain that is shared with other cytokine receptors (Caligiuri et al., J. Exp. Med.
  • T cells express the high affinity IL-2R ⁇ , whereas NK cells, monocytes, and macrophages express predominantly the intermediate-affinity form, IL-2R ⁇ (Caligiuri et al., J. Exp. Med. 1990 171(5): 1509-1526; Voss et al., J. Exp. Med. 1992 176(2):531-541; Theze et al., Immunol. Today 1996 17(10):481-486; Nagler et al, J. Exp. Med. 1990 171(5): 1527-1533).
  • IL-2 activates the JAKs (JAKl, JAK3), which phosphorylate key tyrosines on IL-2R ⁇ , which serve as docking sites for downstream signaling molecules, including She and Stat5, which then catalyze the activation of two distinct pathways: Shc/Ras/Raf/MAPK, and the JAK/STAT5, which translocates to the nucleus and directly regulates a family of gene-regulating transcription factors- STATs (O'Shea, J.J., Immunity 1997 7(1):1-11).
  • Grb-2/Sos complex and activates the Ras/Raf/MAPK pathway, and Grb-2/Gab-2, which activates the phosphatidylinositol 3-kinase (PI3K) pathway. Together these signaling proteins regulate gene transcription factors, ultimately controlling cell growth, division, differentiation and immune activity.
  • PI3K phosphatidylinositol 3-kinase
  • IL- 2 also induces anti-apoptotic effects by regulating pro-mitogenic genes leading to increased bcl-2, bcl-xl, c-myb, which affects cell cycle control through activation of cdk 2,4,6 and inhibition of p27kipl or may be negatively regulated by SOCS (suppressors of cytokine signaling) (Smith, K.A., Science 1988 240(4856): 1169-1176; Theze et al., Immunol. Today 1996 17(10):481-486).
  • SOCS suppressors of cytokine signaling
  • recombinant human IL-2 when administered to tumor-bearing animals for periods of 10 to 14 days can result in regression of tumor burdens, long-term survival, and increased resistance to tumor regrowth.
  • Analysis of splenic lymphocytes obtained from these animals has shown that the anti-tumor effects are due at least in part to augmentation of cytolytic T cell function. This effect includes activation of both direct cytolysis of tumor cells by the CTL, as well an increased synthesis of other, T lymphocyte derived cytokines, which may have further direct or indirect anti-tumor effects as well.
  • RRCC renal cell carcinoma
  • RTK small molecule receptor tyrosine kinase
  • BAY 43-9006 is a potent inhibitor of Raf- 1 , a member of the Raf/MEK/ERK signaling pathway (Richly et al., Int. J. Clin. Pharmacol. Ther.
  • SUl 1248 is a highly potent, selective RTK inhibitor of VEGFR-I- 3, PDGFR ⁇ , cKIT, FLT3 and PDGFR ⁇ (Abrams et al., MoI Cancer Ther. 2003 2(5):471-478; Mendel et al., Clin. Cancer Res. 2003 9(l):327-337; O'Farrell et al., Clin. Cancer Res. 2003 9(15):5465-5476).
  • SU11248 has shown antitumor activity in a number of advanced solid tumors (RCC, neuroendocrine, stromal and adenocarninomas) (Faivre et al., J. Clin. Oncol.
  • Clinical data of SUl 1248 also indicate that the drug is generally tolerated with manageable toxicities, which include fatigue, lymphopenia, neutropenia, hyperlipasemia (Faivre et al., J. Clin. Oncol. 2006 24(l):23-35; Motzer et al., J. Clin. Oncol. 2006 24(1): 16-24).
  • VHL von Hippel-Lindau tumor suppressor gene
  • Vascular endothelial growth factor represents a good target for treatment of clear-cell kidney cancer because mutations in the von Hippel-Lindau tumor- suppressor gene, result in overproduction of this growth factor by the tumors.
  • a randomized, double-blind, placebo-controlled study of the humanized monoclonal antibody (bevacizumab) in patients with metastatic clear-cell renal cancer was conducted by Yang et al.
  • Recombinant human interleukin-2 (aldesleukin) was approved by the FDA for the treatment of metastatic renal cancer based on the results of several multicenter trials in 255 patients who received an intermittent high dose bolus regimen. In these trials, objective responses were seen in 15% of patients, and median survival was 16.3 months (Fisher and Rosenberg 2000 Cancer J Sci Am 6Sl :S55-57). Owing to the significant toxicity associated with this regimen, a series of phase I and phase II trials employing rIL-2 at different doses and using different routes of administration were undertaken. A recent review of the efficacy of rIL-2 as a single agent indicated an overall response of 15% in more than 1700 patients with metastatic RCC who were treated in this series of studies.
  • rIL-2 may also prevent tumor proliferation by causing secondary cytokine release (IFN ⁇ ) from activated NK cells.
  • IFN ⁇ secondary cytokine release
  • Saraya KLA Balkwill FR. Temporal sequence and cellular origin of interleukin-2 stimulated cytokine gene expression. British Journal of Cancer. 1993;67(3):514-21. The response rates and outcomes of clear cell renal cancer patients can be improved. Clinical trials employing novel drug combinations are needed.
  • the combination of rIL2 and antiangiogenic agents, such as bevacizumab may have additive effects that could translate to added clinical benefit for patients with metastatic renal cell carcinoma, as well as other cancers. Another advantage to the complimentary approach to cancer regression is that it provides a platform less easily bypassed by resistance mutations.
  • a single point mutation in the disease state may render it unaffected by a drug resulting in even harsher strains of the disease in future generations.
  • Novel methods and mechanisms for treating patients having disorders associated with abnormal proliferation that are resistant to, or inadequately treated by conventional approaches, utilizing agents targeting immune response mechanisms in the body and disease-state substrates, are needed.
  • the present invention provides such therapeutic agents, and further provides other related advantages.
  • the present invention is based in part on unique combination therapies using small molecule receptor tyrosine kinase inhibitors and rIL-2 with non-overlapping toxicities.
  • melanomas and RCC are generally responsive to immunotherapy such as rIL-2
  • rational combinations of rIL-2 with potential receptor tyrosine kinase inhibitors, based on pharmacology of the individual agents, have been evaluated herein.
  • the invention provides clinically applicable drug schedules to circumvent potential drug interactions based on the toxicological profile of these agents.
  • the potential of adverse pharmacokintic/pharmacodynamic interations and adverse pharmacological interactions are also elucidated.
  • the effects of rIL-2 and small molecule receptor tyrosine kinase inhibitors, such as BAY 43-9006, on T cells and the impact on IL-2 -mediated signaling pathways (MAPK and JAK/STAT5) is described.
  • the present invention expands the indication of the anti-tumor efficacy of IL-2 compounds, such as recombinant IL-2 (rIL-2, also known as aldesleukin) against cancer cell lines that respond poorly to conventional therapies, resulting in increased long-term survival and immunity to tumor rechallenge.
  • IL-2 compounds such as recombinant IL-2 (rIL-2, also known as aldesleukin) against cancer cell lines that respond poorly to conventional therapies, resulting in increased long-term survival and immunity to tumor rechallenge.
  • rIL-2 also known as aldesleukin
  • the immunostimulotory effects of rIL-2 aim to alleviate existing side effects caused by administration of antiangiogenic compositions for co-administration therewith.
  • Methods of treating a subject suffering from abnormal cellular proliferation, particularly renal cell carcinoma using a combination of an IL-2 compound, such as rIL-2 and at least one antiangiogenic agent is provided.
  • IL-2 compound such as rIL-2
  • small molecules of the present invention are listed in Tables 1-5. Particularly preferred molecules are N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-aminoethyl-N-(2-aminoethyl)-2-a small molecule.
  • Aldesleukin and antiangiogenic agents may be administered together or separately as individual pharmaceutical compositions. If administered separately, Aldesleukin can be administered prior to, concurrent with, or subsequent to the antiangiogenic agent.
  • aldesleukin with antiangiogenic agent such as 6,7-bis(2-methoxyethoxy)-N-(3- ethynylphenyl)quinazolm-4-amine, 6-(3-mo ⁇ holinopropoxy)-N-(3-chloro-4- fluorophenyl)-7-methoxyquinazolin-4-amine, N-(2-(dimethylamino)ethyl)-5-((5- fluoro-2-oxoindolin-3-ylidene)methyl)-2,4-diethyl-lH-pyrrole-3-carboxamide, N-(4- chlorophenyl)-4-((pyridin-4-yl)methyl)phthalazin-l -amine, and l-(4-(2- (methylcarbamoyl) ⁇ yridin-4-yloxy
  • the antiangiogenic agents preferably one of 6,7-bis(2- methoxyethoxy)-N-(3-ethynylphenyl)quinazolin-4-amine, 6-(3-morpholinopropoxy)- N-(3-chloro-4-fluorophenyl)-7-methoxyquinazolin-4-amine, N-(2-
  • antiangiogenic proteins such as monoclonal antibodies capable of inhibiting VEGF.
  • antiangiogenic proteins of the present invention are bevacizumab or VEGF-Trap.
  • the protein is an EGF inhibitor, such as cetuximab.
  • administering potentiates the effectiveness of the antiangiogenic agent, resulting in a positive/synergistic therapeutic response that is improved with respect to that observed with the inhibitor alone.
  • a therapeutic package suitable for commercial sale for treating a patient suffering from cancer comprising a container, a therapeutically effective amount of aldesleukin, and a therapeutically effective amount of an antiangiogenic agent and/or small molecule, preferably listed in Tables 1-5, such as 6,7-bis(2-methoxyethoxy)-N-(3-ethynylphenyl)quinazolin-4-amine, 6-(3- morpholinopropoxy)-N-(3-chloro-4-fluorophenyl)-7-methoxyquinazolin-4-amine, N- (2-fdimethvlamino ') ethvlV5-ff5-fli ] nro-2-oxoindolin-3-ylidene)methyl)-2,4-diethyl- lH-pyrrole-3-carboxamide, N-(4-chlorophenyl)-4-((pyridin-4-yl)methyl)phthal
  • kits comprises a combination of medicaments for the treatment of a patient suffering from cancer, comprising: (a) aldesleukin, and (b) an antiangiogenic agent selected from 6,7-bis(2- methoxyethoxy)-N-(3-ethynylphenyl)quinazolin-4-amine, 6-(3-morpholinopropoxy)- N-(3-chloro-4-fluorophenyl)-7-methoxyquinazolin-4-amine, N-(2- (dimethylamino)ethyl)-5-((5-fluoro-2-oxoindolin-3-ylidene)methyl)-2,4-diethyl-lH- pyrrole-3-carboxamide, N-(4-chlorophenyl)-4-((pyridin-4-yl)methyl)phthalazin-l- amine, or l-(4-(2-(methylcarbamoyl)pyr
  • Figure 1 shows single agent activity of rIL-2, BAY 43-9006 or SUl 1248 in IL-2-responsive, T-cell competent murine tumor models.
  • B16-F10 melanoma (2 x 10 6 ), CT26 colon (2 x 10 6 ) or RENCA renal carcinoma (Ix 10 6 ) cells were implanted s.c. into the right flank of female C57BL6 or BALB/c mice. Treatments were initiated when tumors were established to a mean size of 50 - 225 mm 3 , as outlined in methods. Mice were randomized into treatment cohorts (10 mice/group). rIL-2 was administered daily subcutaneously (0.2-3 mg/kg/d).
  • FIG. 1 illustrates the mean tumor growth inhibition (calculated as [l-(mean tumor volume of treated group/mean tumor volume of vehicle group) x 100]) of single agents in the B 16-F 10, CT26 and RENCA tumor model between days 10-14. The data is compiled from multiple independent studies with each study of 10 mice/group. * denotes statistical significance vs. Vehicle treatment (p ⁇ 0.05, ANOVA).
  • Figure 2 shows efficacy of concomitant rIL-2 and BAY 43-9006 therapy in the B16-F10 murine melanoma model.
  • B16-F10 cells (2 x 10 6 cells) were implanted subcutaneously in the right flank of female C57BL6 mice. Mice were randomized into groups when tumors were ⁇ 50 mm 3 , and treatments initiated on day 0 with either vehicle, days 0-6 ( ⁇ ), rIL-2 (3.3 mg/kg, s.c, days 0-6) ( ⁇ ) or BAY 43-9006 (30 mg/kg, p.o. days 0-6) (x) or combined rIL-2 (3.3 mg/kg, s.c, days 0-6) + BAY 43- 9006 (30 mg/kg, p.o. days 0-6) ( ⁇ ).
  • Figures 3 A and 3B show efficacy of sequential rIL-2 and BAY 43-9006 therapy in the B 16-F 10 murine melanoma model.
  • B 16-F 10 cells (2 x 10 6 cells) were implanted subcutaneously in the right flank of female C57BL6 mice. Mice were randomized into groups when tumors were -50 mm3, and treatments initiated on day 0.
  • Figure 3A shows results from sequential regimen with rIL-2 administered first, then BAY 43-9006. Panel illustrates: vehicle ( ⁇ ) days 0-6, 7-13; rIL-2 (3.3 mg/kg, s.c, days 0-6) ( ⁇ ) or BAY 43-9006 (30 mg/kg, p.o.
  • FIG. 3B shows results from sequential regimen with BAY 43-9006 administered first, followed by rIL-2.
  • Panel illustrates: vehicle ( ⁇ ) days 0-6, 7-13; BAY 43-9006 (30 mg/kg, p.o. days 0-6) (x); rIL-2 (3.3 mg/kg, s.c, days 7-13) (D) or combined BAY 43-9006 (30 mg/kg, p.o. days 0-6) + rIL-2 (3.3 mg/kg, s.c, days 7-13) (A).
  • FIG 4 shows efficacy of concomitant rIL-2 and SUl 1248 therapy in the B16-F10 murine melanoma model.
  • B16-F10 cells (2 x 10 6 cells) were implanted subcutaneously in the right flank of female C57BL6 mice. Mice were randomized into groups when tumors were ⁇ 50 mm 3 , and treatments initiated on day 0 with either vehicle, days 0-6 (0), rIL-2 (3.3 mg/kg, s.c, days 0-6) (D) or SUl 1248 (40 mg/kg, p.o. days 0-6) (O) or combined rIL-2 (3.3 mg/kg, s.c, days 0-6) + SUl 1248 (40 mg/kg, p.o. days 0-6) (A).
  • Figures 5 A and 5B show efficacy of sequential rIL-2 and SUl 1248 therapy in the Bl 6-F 10 murine melanoma model.
  • Bl 6-F 10 cells (2 x 10 6 cells) were implanted subcutaneously in the right flank of female C57BL6 mice. Mice were randomized into groups when tumors were ⁇ 50 mm J , and treatments initiated on day 0.
  • Figure 5A shows results from sequential regimen with rIL-2 administered first, then SUl 1248. Panel illustrates: vehicle (0) days 0-6, 7-13; rIL-2 (3.3 mg/kg, s.c, days 0-6) ( ⁇ ) or SUl 1248 (40 mg/kg, p.o.
  • FIG. 5B shows results from sequential regimen with SUl 1248 administered first, followed by rIL-2.
  • Panel illustrates: vehicle (0) days 0-6, 7-13; SUl 1248 (40 mg/kg, p.o. days 0-6) (O); rIL-2 (3.3 mg/kg, s.c, days 7-13) ( ⁇ ) or combined SUl 1248 (40 mg/kg, p.o. days 0-6) + rIL-2 (3.3 mg/kg, s.c, days 7-13) ( ⁇ ).
  • Figure 6 shows efficacy of sequential rIL-2 and BAY 43-9006 therapy in the CT26 murine colon adenocarcinoma model.
  • CT26 cells (2 x 10 6 cells) were implanted subcutaneously in the right flank of female BALB/c mice. Mice were randomized into groups when tumors were -225 mm 3 , and treatments initiated on day 0.
  • Panel illustrates the sequential regimen with rIL-2 administered first, then BAY 43- 9006: vehicle ( ⁇ ) days 0-6, 7-13; rIL-2 (1 mg/kg, s.c, days 0-6) (D) or BAY 43- 9006 (40 mg/kg, p.o. days 7-13) ( ⁇ ) or combined rIL-2 (1 mg/kg, s.c, days 0-6) + BAY 43-9006 (40 mg/kg, p.o. days 7-13) (•).
  • Figure 7 shows efficacy of concomitant treatment of rIL-2 and SUl 1248 in the CT26 murine colon adenocarcinoma model.
  • CT26 cells (2 x 10 6 cells) were implanted subcutaneously in the right flank of female BALB/c mice. Mice were randomized into groups when tumors were -225 mm 3 , and treatments initiated on day 0.
  • Panel illustrates: vehicle ( ⁇ ) days 0-6; rIL-2 (1 mg/kg, s.c, days 0-6) (D) or SUl 1248 (40 mg/kg, p.o. days 0-6) (O) or combined rIL-2 (1 mg/kg, s.c, days 0-6) + SUl 1248 (40 mg/kg, p.o. days 0-6) (•).
  • Figures 8 A and 8B shows efficacy of sequential rIL-2 and SUl 1248 therapy in the CT26 murine colon adenocarcinoma model.
  • CT26 cells (2 x 10 6 cells) were implanted subcutaneously in the right flank of female BALB/c mice. Mice were randomized into groups when tumors were -225 mm 3 , and treatments initiated on day 0.
  • Figure 8A shows results from the sequential regimen with rIL-2 administered first, then SUl 1248: vehicle ( ⁇ ) days 0-6, 7-13; rIL-2 (1 mg/kg, s.c, days 0-6) (D) or SUl 1248 (40 mg/kg, p.o.
  • FIG. 8B shows results irom sequential regimen with SUl 1248 administered first, followed by rIL-2.
  • Panel illustrates: vehicle ( ⁇ ) days 0-6, 7-13; SUl 1248 (40 mg/kg, p.o. days 0-6) (O); rIL-2 (3.3 mg/kg, s.c, days 7-13) (D) or combined SUl 1248(40 mg/kg, p.o. days 0-6) + rIL-2 (3.3 mg/kg, s.c, days 7-13) (A).
  • Figure 9 shows efficacy of concomitant rIL-2 and BAY 43-9006 therapy in the murine RCC RENCA tumor model.
  • RENCA cells (1 x 10 6 cells) were implanted subcutaneously in the right flank of female BALB/c mice. Mice were randomized into groups when tumors were ⁇ 50 mm 3 , and treatments initiated on day 0 with either vehicle, days 0-8 ( ⁇ ), rIL-2 (1 mg/kg, s.c, days 0-4, 7-11) ( ⁇ ) or BAY 43-9006 (30 mg/kg, p.o. days 0-8) (x) or combined rIL-2 (1 mg/kg, s.c, days 0-4, 7-11) + BAY 43-9006 (30 mg/kg, p.o. days 0-8 ( ⁇ ).
  • Figure 10 shows efficacy of concomitant treatment of rIL-2 and SUl 1248 in the therapy in the murine RCC RENCA tumor model.
  • RENCA cells (1 x 10 6 cells) were implanted subcutaneously in the right flank of female BALB/c mice. Mice were randomized into groups when tumors were ⁇ 50 mm 3 , and treatments initiated on day 0.
  • Panel illustrates: vehicle ( ⁇ ) days 0-6; rIL-2 (1 mg/kg, s.c, days 0-6) ( ⁇ ) or SUl 1248 (40 mg/kg, p.o. days 0-6) (x) or combined rIL-2 (1 mg/kg, s.c, days 0-6) + SUl 1248 (40 mg/kg, p.o. days 0-6) (A).
  • cancers include, for example, CML, AML, breast, gastric, endometrial, salivary gland, adrenal, non-small cell lung, pancreatic, renal, rectal, skin, melanoma, multiple myeloma, brain/CNS, cervix, nasopharynx, malignant mesothelioma, hypopharynx, gastroinstestinal carcinoid, peritoneum, omentum, mesentery, gallbladder, testis, esophageal, lung, thyroid, ovarian, peritoneal, prostate, head and neck, bladder, colon, colorectal, lymphomas, and glioblastomas.
  • the methods described herein are useful in the treatment of any such cancer.
  • Therapy with a combination of aldesleukin and at least one antiangiogenic agent in the manner set forth herein causes a physiological response that is beneficial with respect to treatment of cancers whose unabated proliferating cells are highly dependent on vascularization, such as VEGF.
  • One embodiment of the invention provides a method of treating a cancer patient suffering from hypotension from the administration of aldesleukin, comprising: co-administering to the patient a therapeutically effective amount of an antiangiogenic agent to attenuate hypotension. Another more particular embodiment thereof comprises amelioration of cancer in the patient.
  • Another embodiment of the invention provides a method of treating a cancer patient suffering from hypertension from the administration of an antiangiogenic agent, comprising: co-administering to the patient a therapeutically effective amount of aldesleukin to attenuate hypertension.
  • Another more particular embodiment thereof comprises amelioration of cancer in the patient.
  • the cancer is susceptible to inhibition of angiogenesis and/or immune stimulation.
  • said antiangiogenic agent is selected from 6,7-bis(2-methoxyethoxy)-N-(3- ethynylphenyl)quinazolin-4-amine, 6-(3 -morpholinopropoxy)-N-(3 -chloro-4- fluorophenyl)-7-methoxyquinazolin-4-amine, N-(2-(dimethylamino)ethyl)-5-((5- fluoro-2-oxoindolin-3-ylidene)methyl)-2,4-diethyl-lH-pyrrole-3-carboxamide, N-(4- chlorophenyl)-4-((pyridin-4-yl)methyl)phthalazin- 1 -amine, or 1 -(4-(2- (methylcarbamoyl)pyridin-4-yloxy)phenyl)-3-(4-chloro-3- (trifluoromethyl)phenyl
  • Another embodiment of the invention provides a method of treating a patient suffering from cancer comprising administering to said patient aldesleukin and a compound selected from 6,7-bis(2-methoxyethoxy)-N-(3-ethynylphenyl)quinazolin-4- amine, 6-(3-morpholinopropoxy)-N-(3-chloro-4-fluorophenyl)-7-methoxyquinazolin- 4-amine, N-(2-(dimethylamino)ethyl)-5-((5-fluoro-2-oxoindolin-3-ylidene)methyl)- 2,4-diethyl- 1 H-pyrrole-3 -carboxamide, N-(4-chlorophenyl)-4-((pyridin-4- yl)methyl)phthalazin- 1 -amine, or 1 -(4-(2-(methylcarbamoyl)pyridin-4-yloxy)phenyl)- 3-(4-ch
  • Another embodiment of the invention provides a method of increasing efficacy of aldesleukin in a cancer patient comprising first administering an antiangiogenic agent in a dose capable of inducing hypoxia in the patient then administering the aldesleukin.
  • Another embodiment provides a method of increasing efficacy of an antiangiogenic agent in a cancer patient comprising reducing nitric oxide synthase by administration of aldesleukin to the patient.
  • Another embodiment provides a method of increasing efficacy of an antiangiogenic agent in a cancer patient comprising administration aldesleukin to the patient wherein, nitric oxide synthase is thereby reduced by administration of aldesleukin to the patient.
  • said antangiogenic agent is selected from 6,7-bis(2-methoxyethoxy)-N-(3-ethynylphenyl)quinazolin-4-amine, 6-(3- morpholinopropoxy)-N-(3-chloro-4-fluorophenyl)-7-methoxyquinazolin-4-amine, N- (2-(dimethylamino)ethyl)-5-((5-fluoro-2-oxoindolin-3-ylidene)methyl)-2,4-diethyl- 1 H-pyrrole-3 -carboxamide, N-(4-chlorophenyl)-4-((pyridin-4-yl)methyl)phthalazin- 1 - amine, or 1 -(4-(2-(methylcarbamoyl)pyridin-4-yloxy)phenyl)-3 -(4-chloro-3 - (trifluoromethyl)phenyl)urea
  • Another embodiment of the invention provides a method for treating a patient suffering from cancer, comprising the steps of first administering to said patient a therapeutically effective amount of aldesleukin followed by administration of an antiangiogenic agent (such as) selected from 6,7-bis(2-methoxyethoxy)-N-(3- ethynylphenyl)quinazolin-4-amine, 6-(3-morpholinopropoxy)-N-(3-chloro-4- fluorophenyl)-7-methoxyquinazolm-4-amine, N-(2-(dimethylamino)ethyl)-5-((5- fluoro-2-oxoindolin-3-ylidene)methyl)-2,4-diethyl-lH-pyrrole-3-carboxamide, N-(4- chlorophenyl)-4-((pyridin-4-yl)methyl)phthalazin-l -amine, or l-(4-(2- (methylcarbamoyl)
  • Another embodiment of the invention provides a method for treating a patient suffering from cancer, comprising the steps of first administering to said patient a therapeutically effective amount of an antiangiogenic agent, such as, 6,7-bis(2- methoxyethoxy)-N-(3-ethynylphenyl)quinazolin-4-amine, 6-(3-morpholinopropoxy)- N-(3-chloro-4-fluorophenyl)-7-methoxyquinazolin-4-amine, N-(2- (dimethylamino)ethyl)-5-((5-fluoro-2-oxoindolin-3-ylidene)methyl)-2,4-diethyl-lH- pyrrole-3 -carboxamide, N-(4-chloro ⁇ henyl)-4-((pyridin-4-yl)methyl)phthalazin- 1 - amine, or l-(4-(2-(methylcarbamoyl)pyridin-4-yl
  • Another embodiment of the invention provides a method for treating a patient suffering from cancer, comprising separately administering to said patient a therapeutically effective amount of an antiangiogenic agent and aldesleukin according to a dosing schedule, wherein the aldesleukin is administered from 1 to 3 times daily in a dose between about 9 and about 130 MIU/day for a period of at least 3 consecutive days, optionally followed by a rest period of at least 3 consecutive days.
  • said antiangiogenic agent is administered from 1 to 6 times every 2-3 weeks.
  • said antiangiogenic agent is a VEGF inhibitor.
  • the antiangiogenic agent is selected from 6,7-bis(2-methoxyethoxy)-N-(3-ethynylphenyl)quinazolin-4-amine, 6-(3- morpholinopropoxy)-N-(3-chloro-4-fluorophenyl)-7-methoxyquinazolin-4-amine, N- (2-(dimethylamino)ethyl)-5-((5-fluoro-2-oxoindolin-3-ylidene)methyl)-2,4-diethyl- 1 H-pyrrole-3 -carboxamide, N-(4-chlorophenyl)-4-((pyridin-4-yl)methyl)phthalazin- 1 - amine, or 1 -(4-(2-(methylcarbamoyl)pyridin-4-yloxy)phenyl)-3-(4-chloro-3- (trifluoromethyl)phenyl)urea
  • the aldesleukin is administered subcutaneously and said rest period is absent.
  • the aldesleukin is administered 3 times daily in a dose of about 30-100 MIU/day.
  • the aldesleukin is administered for a period of 5 consecutive days followed by a 9-day rest period.
  • said dosing schedule is repeated for at least two courses.
  • each course consists of 2-5 day treatments followed by a rest period of 9-15 days.
  • the aldesleukin is administered 3 times for the first day and once daily each proceeding day.
  • said dosing schedule is repeated for at least two courses, or 3 courses, or 4 courses, or 5 courses, or 6 courses, or 7 courses, or 8 courses, or 9 courses, or 10 courses.
  • the cancer is colon cancer, renal cell carcinoma or malignant melanoma.
  • At least one compound selected from acetaminophen, meperidine, indomethacin, ranitidine, nizatidine, diastop, loperamide, diphenhydramine, or furosemide is also administered to said patient.
  • said antangiogenic agent is 6,7-bis(2-methoxyethoxy)-N-(3-ethynylphenyl)quinazolin-4- amine, 6-(3 -morpholinopropoxy)-N-(3 -chloro-4-fluorophenyl)-7-methoxyquinazolin- 4-amine, N-(2-(dimethylammo)ethyl)-5-((5-fluoro-2-oxoindolin-3-ylidene)methyl)- 2,4-diethyl- 1 H-pyrrole-3 -carboxamide, N-(4-chlorophenyl)-4-((pyridin-4- yl)methyl)phthalazin-l -amine, or l-(4-(2-(methylcarbamoyl)pyridin-4-yloxy)phenyl)- 3 -(4-chloro-3 -(trifluoromethyl
  • onfffirin p from cancer comprising administering aldesleukin and: a tautomer thereof, a pharmaceutically acceptable salt thereof, or a pharmaceutically acceptable salt of the tautomer.
  • Ri is alkyl, -aryl(Rj) p , or heterocyclyl;
  • R 2 is H or alkyl; or,
  • Ri and R 2 are bound together to form R] -2 ;
  • R 3 is H, -CN, -OH, halogen, alkyl, aryl, alkoxy, -NR a R b , -C(O)R c , -S(O) n Ra, or heterocyclyl;
  • R 4 Is H, -CN, -OH, halogen, alkyl, aryl, -0-(CH 2 ) q -R g , -O-(CH 2 ) q -O-Re, -NR 3 R b , -S(O) n R d , or -heterocyclyl-R f ;
  • R 5 is H, -CN, -OH, halogen, alkyl, aryl, -0-(CH 2 ) q -R g , -O-(CH 2 ) q -O-Re, -NR 3 R b ,
  • R 6 is H, -CN, -OH, halogen, alkyl, aryl, alkoxy, -NR a R b , -C(O)R c , -S(O) n R d , or heterocyclyl
  • R 7 is H, -OH, halogen, alkyl, aryl, alkoxy, -NR 3 R b , -S(O) n R d , or heterocyclyl
  • each R a and R b is independently H, alkyl, -C(O)R 0 , aryl, heterocyclyl, or alkoxy; or,
  • R 3 and R b are bound together to form Ri -2 ; each R 0 is independently H, alkyl, alkoxy, -C(O)alkyl, -C(O)aryl, -CHO, aryl, or heterocyclyl; each Rj is independently H, alkyl, alkenyl, aryl, or -NR 3 R b ; each R 3 is independently H or alkyl;
  • Rf is H, halogen, -OH, -CN, -(CH 2 ) q NR a R h , alkoxy, -C(O)R 0 , -(CH 2 ) q CH 3 .
  • each Rg is independently H, halogen, -C(O)R 0 , aryl, heterocyclyl, or -NR a R b ;
  • R h is H, or -(CH 2 ) q S(O) n Rd; each Rj is independently H, halo, alkyl, alkenyl, alkynyl, or -0(CH 2 ) q -R g ; each n is independently 0, 1, or 2; each p is independently 0, 1, 2, or 3; each q is independently 0, 1, or 2; Ri -2 has the general structure as shown:
  • said compound is:
  • said compound is:
  • R- 2U R 22 , R 23 , and R 24 are each independently H, -CN, -OH, halogen, alkyl, aryl, alkoxy, -NR a R b , -C(O)R 2 , -S(O) n R d , or heterocyclyl;
  • R 25 is H, alkyl, or -C(O)R 2 ;
  • R 2 6 and R 27 are each independently H, -OH, halogen, alkyl, -NR a R b , -C(O)R 2 , or
  • R 28 is H, -CH 3 , or halogen; each R a and R b is independently H, alkyl, -C(O)R 2 , aryl, heterocyclyl, or alkoxy; each R 2 is independently H, alkyl, alkoxy, -NH 2 , -NH(alkyl), -N(alkyl) 2 , aryl, or heterocyclyl; each Ra is independently H, alkyl, alkenyl, aryl, or -NR a Rt,; each n is independently O, 1 , or 2; and each q is independently O, 1 , or 2; or a tautomer thereof, a pharmaceutically acceptable salt thereof, or a pharmaceutically acceptable salt of the tautomer.
  • said compound is:
  • said compound is:
  • t au t omer thereof a pharmaceutically acceptable salt thereof, or a pharmaceutically acceptable salt of the tautomer.
  • Another embodiment of the invention provides a method of decreasing the toxicity associated with the administration of aldesleukin to a cancer patient comprising administering a quinolinone, preferably an antiangiogenic agent, preferably selected from 6,7-bis(2-methoxyethoxy)-N-(3-ethynylphenyl)quinazolin-4- amine, 6-(3-morpholinopropoxy)-N-(3-chloro-4-fluorophenyl)-7-methoxyquinazolin- 4-amine, N-(2-(dimethylamino)ethyl)-5-((5-fluoro-2-oxoindolin-3-ylidene)methyl)- 2,4-diethyl- 1 H-pyrrole-3 -carboxamide, N-(4-chlorophenyl)-4-((pyridin-4- yl)methyl)phthalazin- 1 -amine, or 1 -(4-(2-(methylcarbamoyl)
  • Another embodiment of the invention provides a method of decreasing the toxicity associated with the administration of an antiangiogenic agent, preferably selected from 6,7-bis(2-methoxyethoxy)-N-(3-ethynylphenyl)quinazolin-4-amine, 6- (3-mo ⁇ holinopropoxy)-N-(3-chloro-4-fluorophenyl)-7-methoxyquinazolin-4-amine, N-(2-(dimethylamino)ethyl)-5-((5-fluoro-2-oxoindolin-3-ylidene)methyl)-2,4-diethyl- lH-pyrrole-3-carboxamide, N-(4-chlorophenyl)-4-((pyridin-4-yl)methyl)phthalazin-l- amine, or 1 -(4-(2-(methylcarbamoyl)pyridin-4-yloxy)phenyl)-3-(4-chloro-3- (
  • Another embodiment of the invention provides a method of decreasing IL-2 resistance in a patient suffering from cancer comprising administering an antiangiogenic agent, preferably selected from 6,7-bis(2-methoxyethoxy)-N-(3- ethynylphenyl)quinazolin-4-amine, 6-(3 -morpholinopropoxy)-N-(3 -chloro-4- fluorophenyl)-7-methoxyquinazolin-4-amine, N-(2-(dimethylamino)ethyl)-5-((5- fluoro-2-oxoindolin-3-ylidene)methyl)-2,4-diethyl-lH-pyrrole-3-carboxamide, N-(4- chlorophenyl)-4-((pyridin-4-yl)methyl)phthalazin-l -amine, or l-(4-(2- (methylcarbamoyl)pyridin-4-yloxy)phenyl)-3-(4-
  • Another embodiment of the invention provides a method for treating a patient suffering from cancer, comprising the steps of first administering to said patient a therapeutically effective amount of aldesleukin followed by administration of an antiangiogenic agent, preferably selected from 6,7-bis(2-methoxyethoxy)-N-(3- ethynylphenyl)quinazolin-4-amine, 6-(3 -morpholinopropoxy) -N-(3 -chloro-4- fluoiOphenyl)-7-methoxyquinazolin-4-amine, N-(2-(dimethylamino)ethyl)-5-((5- fiuoro-2-oxoindolin-3 -ylidene)methyl)-2,4-diethyl- 1 H-pyrrole-3 -carboxamide, N-(4-
  • Another embodiment of the invention provides a method for treating a patient suffering from cancer, comprising separately administering to said patient a therapeutically effective amount of an antiangiogenic agent, preferably selected from 6,7-bis(2-methoxyethoxy)-N-(3-ethynylphenyl)quinazolin-4-amine, 6-(3- mo ⁇ holino ⁇ ropoxy)-N-(3-chloro-4-fluorophenyl)-7-methoxyquinazolin-4-amine, N- (2-(dimethylamino)ethyl)-5-((5-fluoro-2-oxoindolin-3-ylidene)methyl)-2,4-diethyl- lH- ⁇ yrrole-3-carboxamide, N-(4-chlorophenyl)-4-((pyridm-4-yl)methyl)phthalazin-l- amine, or 1 -(4-(2-(methylcarbamoyl)pyridin-4-y
  • the dose is between about 30 and about 100 MIU/day. More specifically the dose is between about 30 and about 60 MIU/day. More specifically the dose is between about 30 and about 40 MIU/day. More specifically the dose is between about 17 and about 30 MIU/day. More specifically the dose is between about 9 and about 30 MIU/day.
  • said antiangiogenic agent preferably selected from 6,7-bis(2-methoxyethoxy)-N-(3-ethynylphenyl)quinazolin-4-amine, 6- (3-morpholinopropoxy)-N-(3-chloro-4-fluorophenyl)-7-methoxyquinazolin-4-amine, N-(2-(dimethylamino)ethyl)-5-((5-fluoro-2-oxoindolin-3-ylidene)methyl)-2,4-diethyl- lH- ⁇ yrrole-3-carboxamide, N-(4-chlorophenyl)-4-((pyridin-4-yl)methyl)phthalazin-l- amine, or l-(4-(2-(methylcarbamoyl)pyridin-4-yloxy)phenyl)-3-(4-chloro-3-
  • said antiangiogenic agent preferably selected from 6,7- bis(2-methoxyethoxy)-N-(3 -ethynylphenyl)quinazolin-4-amine, 6-(3 - morpholinopropoxy)-N-(3-chloro-4-fluorophenyl)-7-methoxyquinazolin-4-amine, N- (2-(dimethylamino)ethyl)-5-((5-fluoro-2-oxoindolin-3-ylidene)methyl)-2,4-diethyl- lH-pyrrole-3-carboxamide, N-(4-chlorophenyl)-4-((pyridin-4-yl)methyl)phthalazin-l- amine, or 1 -(4-(2-(methylcarbamoyl)pyridin-4-yloxy)phenyl)
  • the antiangiogenic agent preferably selected from 6,7- bis(2-methoxyethoxy)-N-(3-ethynyl ⁇ henyl)quinazolin-4-amine, 6-(3- morpholinopropoxy)-N-(3-chloro-4-fluoro ⁇ henyl)-7-methoxyquinazolin-4-amine, N- (2-(dimethylamino)ethyl)-5-((5-fluoro-2-oxoindolin-3-ylidene)methyl)-2,4-diethyl- 1 H-pyrrole-3 -carboxamide, N-(4-chlorophenyl)-4-((pyridin-4-yl)methyl)phthalazin- 1 - amine, or l-(4-(2-(methylcarbamoyl)pyridin-4-yloxy)phenyl)-3-(4-chloro-3- (trifluoromethyl)phenyl)ure
  • said bevacizumab is administered in a dose between 1 and 12 mg/kg once every 12-16 days.
  • the antiangiogenic agent is administered within 72 hours of the administration of rIL-2; wherein "within” is meant to indicate before or after, as in: the antiangiogenic agent is administered 72 hours before or 72 hours after the administration of rIL-2.
  • the antiangiogenic agent is administered within 14 days of the administration of rIL-2.
  • the antiangiogenic agent is administered within 7 days of the administration of rIL-2.
  • the antiangiogenic agent is administered within 6 days of the administration of rIL-2.
  • the antiangiogenic agent is administered within 5 days of the administration of rIL-2.
  • the antiangiogenic agent is administered within 4 days of the administration of rIL-2. In another embodiment, the antiangiogenic agent is administered within 48 hours of the administration of rIL-2. In another embodiment, the antiangiogenic agent is administered within 36 hours of the administration of rIL-2. In another embodiment, the antiangiogenic agent is administered within 24 hours of the administration of rlL- 2. In another embodiment, the antiangiogenic agent is administered within 12 hours of the administration of rIL-2. In another embodiment, the antiangiogenic agent is administered within 6 hours of the administration of rIL-2. In another embodiment, the antiangiogenic agent is administered within 1 hour of the administration of rIL-2.
  • the antiangiogenic agent is administered at the same time as the administration of rIL-2. In another embodiment, the antiangiogenic agent is administered in combination with rIL-2. In a more particular embodiment thereof the antiangiogenic agent is selected from 6,7-bis(2-methoxyethoxy)-N-(3 ⁇ ethynylphenyl)quinazolin-4-amine, 6-(3-morpholinopropoxy)-N-(3-chloro-4- fiuorophenyl)-7-methoxyquinazolin-4-amine, N-(2-(dimethylamino)ethyl)-5-((5- fluoro-2-oxoindolin-3-ylidene)methyl)-2,4-diethyl-lH-pyrrole-3-carboxamide, N-(4-
  • Another embodiment of the invention provides a method of treating a patient suffering from cancer comprising co-administering aldesleukin and an antiangiogenic agent, preferably selected from 6,7-bis(2-methoxyethoxy)-N-(3- ethynylphenyl)quinazolin-4-amine, 6-(3-morpholinopropoxy)-N-(3-chloro-4- fluorophenyl)-7-methoxyquinazolin-4-amine, N-(2-(dimethylamino)ethyl)-5 ⁇ ((5- fluoro-2-oxoindolin-3-ylidene)methyl)-2,4-diethyl-lH-pyrrole-3-carboxamide, N-(4- chloro ⁇ henyl) ⁇ 4-((pvridin-4-yl)methyl)phthalazin-l -amine, or l-(4-(2- (methylcarbainoyl)pyridin-4-yloxy)phenyl
  • Another embodiment of the invention provides a therapeutic package suitable for commercial sale for treating a patient suffering from cancer, comprising a container, a therapeutically effective amount of aldesleukin, and a therapeutically effective amount of an antiangiogenic agent, preferably selected from 6,7-bis(2- methoxyethoxy)-N-(3-ethynylphenyl)quinazolin-4-amine, 6-(3-mo ⁇ holinopropoxy)- N-(3-chloro-4-fluorophenyl)-7-methoxyquinazolin-4-amine, N-(2- (dimethylamino)ethyl)-5-((5-fluoro-2-oxoindolin-3-ylidene)methyl)-2,4-diethyl-lH- pyrrole-3 -carboxamide, N-(4-chlorophenyl)-4-((pyridin-4-yl)methyl) ⁇ hthalazin- 1 - amine, or 1 -(4
  • a therapeutic package thereof wherein said patient is suffering from renal cell carcinoma.
  • a therapeutic package thereof further comprising written matter instructing that the patient receive treatment with aldesleukin prior to treatment with the antiangiogenic agent, preferably selected from 6,7-bis(2-methoxyethoxy)-N-(3-ethynylphenyl)quinazolin-4-amine, 6-(3- morpholinopropoxy)-N-(3-chloro-4-fluorophenyl)-7-methoxyquinazolin-4-amine, N- (2-(dimethylamino)ethyl)-5-((5-fluoro-2-oxoindolin-3-ylidene)methyl)-2,4-diethyl- lH-pyrrole-3-carboxamide, N-(4-chlorophenyl)-4-((pyridin-4-yl)methyl)phthalazin-l- amine, or l-(4-(2-(methylcarbamoyl)pyr
  • Another embodiment provides a pharmaceutical composition
  • a pharmaceutical composition comprising a an antiangiogenic agent, preferably selected from 6,7-bis(2-methoxyethoxy)-N-(3- ethynylphenyl)quinazolin-4-amine, 6-(3-morpholinopropoxy)-N-(3-chloro-4-
  • Clear cell renal cell carcinoma is associated with increased vascularization, particularly VEGF expression.
  • VEGF is a protein that plays a crucial role in tumor angiogenesis (the formation of new blood vessels to the tumor) and maintaining established tumor blood vessels. It binds to specific receptors on blood vessels to stimulate extensions to existing blood vessels. Some but not all cases of RCCa have increased serum VEGF levels.
  • VEGF increases vascular permeability and treatment with anti- VEGF has been noted to lead to increased blood pressure of some patients. It is therefore likely that some of the major toxicities of anti-VEGF (e.g. hypertension) and rIL-2 (e.g. hypotension) would be mutually counteracted and lead to an overall improved therapeutic index if the combinations of the present invention were administered to patients suffering from cancer, such as RCCa.
  • anti-VEGF e.g. hypertension
  • rIL-2 e.g. hypotension
  • aldesleukin with bevacizumab or cetuximab yield a higher proportion and/or better durability of responses compared to the agents administered alone.
  • the particular aldesleukin dosing regimens disclosed herein provide for intermittent stimulation of natural killer (NK) cell activity and decreased risk of rIL-2- related side effects that can be associated with long term exposure to rIL-2 dosing.
  • NK natural killer
  • hypotension typically associated with rIL-2 dosing is offset by the administration of the antiangiogenic compositions of the present invention, since they are generally associated with hypertension.
  • hypoxia typically associated with VEGF inhibitors will increase sensitivity to rIL-2 treatment efficacy, while simultaneously providing palliative benefits as well.
  • Another embodiment of the invention provides the use of aldesleukin in the manufacture of a medicament for treating cancer, wherein said medicament is for
  • an antiangiogenic agent preferably selected from 6,7-bis(2-methoxyethoxy)-N-(3-ethynylphenyl)quinazolin-4-amine, 6- (3-morpholinopropoxy)-N-(3-chloro-4-fluorophenyl)-7-methoxyquinazolin-4-amine, N-(2-(dime1hylamino)ethyl)-5-((5-fluoro-2-oxoindolin-3-ylidene)methyl)-2,4-diethyl- 1 H-pyrrole-3 -carboxamide, N-(4-chloropb.enyl)-4-((pyridin-4-yl)methyl)phtb.alazin- 1 - amine, or l-(4-(2-(methylcarbamoyl)pyridin-4-yloxy)phenyl)-3-(4-chloro-3- (
  • said antiangiogenic agent preferably selected from 6,7-bis(2-methoxyethoxy)-N-(3- ethynylphenyl)quinazolin-4-amine, 6-(3-morpholinopropoxy)-N-(3-chloro-4- fluorophenyl)-7-methoxyquinazolin-4-amine, N-(2-(dimethylamino)ethyl)-5-((5- fluoro-2-oxomdolin-3-ylidene)methyl)-2,4-diethyl-lH-pyrrole-3-carboxamide, N-(4- chlorophenyl)-4-((pyridin-4-yl)methyl)phthalazin- 1 -amine, or 1 ⁇ (4-(2- (methylcarbamoyl)pyridin-4-yloxy)phenyl)-3-(4-chloro ⁇ 3- (trifluoromethyl)phenyl)urea is a
  • an antiangiogenic agent preferably selected from 6,7-bis(2-methoxyethoxy)-N-(3- ethynylphenyl)quinazolin-4-amine, 6-(3-morpholino ⁇ ropoxy)-N-(3-chloro-4- fluorophenyl)-7-methoxyquinazolin-4-amine, N-(2-(dimethylamino)ethyl)-5-((5- fluoro-2-oxoindolin-3-ylidene)methyl)-2,4-diethyl-lH-pyrrole-3-carboxamide, N-(4- chlorophenyl)-4-((pyridin-4-yl)methyl)phthalazin- 1 -amine, or 1 ⁇ (4-(2- (methylcarbamoyl)pyridin-4-yloxy)phenyl)-3-(4-chloro-3-
  • Preferred molecules associated with angiogenesis, modulated (such as inhibition) by the compositions of the present invention include: Vascular Endothelial Growth Factor (VEGF), Fibroblast Growth Factor (FGF), Interleukin-8 (IL-8), Angiogenin, Angiotropin, Epidermal Growth Factor (EGF), Platelet Derived Endothelial Cell Growth Factor, Transforming Growth Factor a (TGF-a),
  • VEGF Vascular Endothelial Growth Factor
  • FGF Fibroblast Growth Factor
  • IL-8 Interleukin-8
  • Angiogenin Angiotropin
  • EGF Epidermal Growth Factor
  • TGF-a Transforming Growth Factor a
  • TGF-b Transforming Growth Factor b
  • Nitric Oxide a compound that modulates TGF-b
  • modulation such as potentiation
  • Thrombospondin, Angiostatin, and Endostatin is contemplated within the present invention.
  • Ri is alkyl, -aryl(RO P , or heterocyclyl
  • R 2 is H or alkyl
  • Ri and R 2 are bound together to form Ri -2 ;
  • R 3 is H, -CN, -OH, halogen, alkyl, aryl, alkoxy, -NR a R b , -C(O)Rc, -S(O) n Rd, or heterocyclyl;
  • R 4 Is H, -CN, -OH, halogen, alkyl, aryl, -0-(CH 2 ) q -R g , -0-(CH 2 ) C1 -O-R 6 , -NR a R b ,
  • R 5 is H, -CN, -OH, halogen, alkyl, aryl, -0-(CH 2 ) q -R g> -0-(CH 2 ) q -0-Re, -NR a R b , -S(O) n Rd, or heterocyclyl;
  • R 6 is H, -CN, -OH, halogen, alkyl, aryl, alkoxy, -NR 3 Rb, -C(O)R 0 , -S(O) n Rd, or heterocyclyl;
  • R 7 is H, -OH, halogen, alkyl, aryl, alkoxy, -NR 3 R b , -S(O) n R d , or heterocyclyl; each R a and R b is independently H, alkyl, -C(O)R 0 , aryl, heterocyclyl, or alkoxy; or, R a and R b are bound together to form Ri -2 ; each R 0 is independently H, alkyl, alkoxy, -C(O)alkyl, -C(O)aryl, -CHO, aryl, or heterocyclyl; each Rd is independently H, alkyl, alkenyl, aryl, or -NR a R b ; each R e is independently H or alkyl; each R f is H, halogen, -OH, -CN, -(CH 2 ) q NR a R h , alkoxy, -C(O)Rc
  • Rh is H, or -(CH 2 ) q S(O) n Rd; each Ri is independently H, halo, alkyl, alkenyl, alkynyl, or -0(CH 2 ) q -R g ; each n is independently O, 1, or 2; each p is independently O, 1 , 2, or 3;
  • Ri -2 has the general structure as shown:
  • R 2 is H.
  • Ri is -aryl(Ri) p .
  • Ri is -aryl(Rj) P; aryl within Rj is phenyl, p within Ri is 2 and both Ri groups within Ri are halo.
  • Ri is -aryl(Ri) p
  • aryl within Ri is phenyl
  • p within Ri is 1 and R
  • groups within Ri is alkynyl, preferably ethynyl.
  • Ri is -aryl(Rj) P) aryl within Ri is phenyl, p within Ri is 2, one R; group within Ri is halo and the other R; group within Ri is - O(CH 2 ) q -R g .
  • q within Ri is 1 and R g within R] is halophenyl.
  • Ri is -aryl(Ri) p
  • p within Ri is 1 and K ⁇ within Ri is alkynyl.
  • Ri and R 2 are bound together to form Ri -2 :
  • R 8 is H
  • X is N
  • R 9 is -C(O)NHR b
  • R b within R 9 is -phenyl-O-CH 2 (CH 3 ) 2
  • R 3 and R 6 are H.
  • R 4 is -O-(CH 2 ) q -R g .
  • q within R 4 is 1 and R g is H.
  • R 4 is -O-(CH 2 ) q -R g
  • R g within R 4 is heterocyclyl.
  • R 4 and R 5 are each -O- (CH 2 ) q -O-R e .
  • q within both R 4 and R 5 is 2 and R e within both R 4 and R 5 is methyl.
  • R 4 is -heterocyclyl-R f and R 5 is H.
  • said heterocyclyl within R 4 is furanyl.
  • R f within R 4 is -(CH 2 ) q NHR h .
  • R h is -(CH 2 ) q NHR h .
  • R 5 is -O-(CH 2 ) q -R g .
  • R g within R 5 is heterocyclyl.
  • R 7 is H.
  • R 3 , R 6 , and R 7 are all H. INDOLINONE
  • Ri i is alkyl, aryl or heterocyclyl
  • Ri 2 , Ri 3 Ri 4 , and Ri 5 are each independently H, -CN, -OH, halogen, alkyl, aryl, alkoxy, -NR a R b , -C(O)R 2 , -S(O) n Rd, or heterocyclyl; each R 3 and R b is independently H, alkyl, -C(O)R 2 , aryl, heterocyclyl, or alkoxy; each R z is independently H, alkyl, alkoxy, -NH 2 , -NH(alkyl), -N(alkyl) 2 , aryl, or heterocyclyl; each Rd is independently H, alkyl, alkenyl, aryl, or -NR 3 R b ", each n is independently 0, 1, or 2; and each q is independently 0, 1, or 2.
  • Rn is heterocyclyl.
  • Rn is Ri i a :
  • Ri 6 , Ri 7 and Rj 8 are each independently H, -CN, -OH, halogen, alkyl, aryl, alkoxy, - NR 3 Rb, -C(O)R 2 , -S(O) n R d , or heterocyclyl.
  • R 7 is -C(O)-(CH 2 ) p -N(H)( 2-r) (alkyl) r , wherein p is O, 1, 2, 3, 4, or 5 and r is 0, 1 , or 2.
  • Ri i is Ri ia
  • RB is F.
  • Ri i is R l la
  • Ri 7 is -(CH 2 ) t COOH, wherein t is 1, 2, 3, or 4.
  • Rn is R l la
  • R 6 and R 8 are both methyl.
  • Rn is Rn 3
  • Ri 7 is H.
  • Ri i is aryl. More particular still, Ri i is substituted or unsubstituted phenyl. In another embodiment R is iodide.
  • dotted line represents an optional placement of an additional bond
  • R 2 I, R 22 , R 23 , and R 24 are each independently H, -CN, -OH, halogen, alkyl, aryl, alkoxy, -NR 3 Rb, -C(O)R 2 , -S(O) n Rd, or heterocyclyl;
  • R 25 is H, alkyl, or -C(O)R 2 ;
  • R 2 6 and R 27 are each independently H, -OH, halogen, alkyl, -NR 3 R b , -C(O)R 2 , or
  • R 28 is H, -CH 3 , or halogen; each R 3 and Rb is independently H, alkyl, -C(O)R 2 , aryl, heterocyclyl, or alkoxy;
  • references to a certain element such as hydrogen or H is meant to include all isotopes of that element.
  • an R group is defined to include hydrogen or H, it also includes deuterium and tritium.
  • antiangiogenic agent any small molecule, more specifically, any compound having a molecular weight less than 1,100 g/mol that has been shown or will be shown to suppress angiogensis in a system.
  • VEGF Vascular Endothelial Growth Factor
  • FGF Fibroblast Growth Factor
  • IL-8 Interleukin-8
  • Angiogenin Angiotropin
  • EGF Epidermal Growth Factor
  • EGF Platelet Derived Endothelial Cell Growth Factor
  • TGF-a Transforming Growth Factor b
  • Nitric Oxide Nitric Oxide
  • compositions of the present invention include: 6,7-bis(2-methoxyethoxy)-N-(3 -ethynylphenyl)quinazolin-4-amine, 6-(3 - mo ⁇ holinopropoxy)-N-(3-chloro-4-fluorophenyl)-7-methoxyquinazolin-4-amine, N- (2-(dimethylamino)ethyl)-5-((5-fluoro-2-oxoindolin-3-ylidene)methyl)-2,4-diethyl- 1 H- ⁇ yrrole-3-carboxamide, N-(4-chlorophenyl)-4-((pyridin-4-yl)methyl)phthalazin- 1 - amine, and l-(4-(2-(methylcarbamoyl)pyridin-4-yloxy)phenyl)-3-(4-chloro-3- (trifluoromethyl)phenyl)urea.
  • an effective amount of a compound to treat renal cell carcinoma may be an amount necessary to cause regression of tumor growth in renal cells.
  • the effective amount may vary, depending, for example, upon the condition treated, weight of the subject and severity of the disease. One of skill in the art can readily determine the effective amount empirically without undue experimentation.
  • an effective amount for treatment refers to an amount sufficient to palliate, ameliorate, stabilize, reverse, slow or delay progression of a condition such as a disease state.
  • a “subject” or “patient” is meant to describe a human or vertebrate animal including a dog, cat, pocket pet, marmoset, horse, cow, pig, sheep, goat, elephant, giraffe, chicken, lion, monkey, owl, rat, squirrel, slender loris, and mouse.
  • a “pocket pet” refers to a group of vertebrate animals capable of fitting into a commodious coat pocket such as, for example, hamsters, chinchillas, ferrets, rats, guinea pigs, gerbils, rabbits and sugar gliders.
  • pharmaceutically acceptable ester refers to esters, which hydrolyze in vivo and include those that break down readily in the human body to leave the parent compound or a salt thereof. Suitable ester groups include, for example, those derived from pharmaceutically acceptable aliphatic carboxylic acids, particularly alkanoic, alkenoic, cycloalkanoic and alkanedioic acids, in which each
  • esters include, but are not limited to, formates, acetates, propionates, butyrates, acrylates and ethylsuccinates.
  • the compounds of the present invention can be used in the form of salts as in "pharmaceutically acceptable salts" derived from inorganic or organic acids.
  • These salts include but are not limited to the following: acetate, adipate, alginate, citrate, aspartate, benzoate, benzenesulfonate, bisulfate, butyrate, camphorate, camphorsulfonate, digluconate, cyclopentanepropionate, dodecylsulfate, ethanesulfonate, glucoheptanoate, glycerophosphate, hemisulfate, heptanoate, hexanoate, fumarate, hydrochloride, hydrobromide, hydroiodide,
  • 2-hydroxyethanesulfonate lactate, maleate, methanesulfonate, nicotinate, 2-napth- alenesulfonate, oxalate, pamoate, pectinate, sulfate, 3-phenylpropionate, picrate, pivalate, propionate, succinate, tartrate, thiocyanate, p-toluenesulfonate and undecanoate.
  • the basic nitrogen-containing groups can be quaternized with such agents as loweralkyl halides, such as methyl, ethyl, propyl, and butyl chloride, bromides, and iodides; dialkyl sulfates like dimethyl, diethyl, dibutyl, and diamyl sulfates, long chain halides such as decyl, lauryl, myristyl and stearyl chlorides, bromides and iodides, aralkyl halides like benzyl and phenethyl bromides, and others. " Water or oil-soluble or dispersible products are thereby obtained.
  • loweralkyl halides such as methyl, ethyl, propyl, and butyl chloride, bromides, and iodides
  • dialkyl sulfates like dimethyl, diethyl, dibutyl, and diamyl sulfates
  • long chain halides
  • prodrugs refers to those prodrugs of the compounds of the present invention which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of humans and lower animals with undue toxicity, irritation, allergic response, and the like, commensurate with a reasonable benefit/risk ratio, and effective for their intended use, as well as the zwitterionic forms, where possible, of the compounds of the invention.
  • prodrug refers to compounds that are rapidly transformed in vivo to yield the parent compound of the above formula, for example by hydrolysis in blood. A thorough discussion is provided in T. Higuchi and V. Stella, Pro-drugs as Novel Delivery Systems, Vol. 14 of the A.C.S. Symposium Series, and in Edward B. Roche, ed., Bioreversible Carriers in Drug Design, American Pharmaceutical
  • halo refers to F, Cl, Br, or I atoms.
  • alkyl refers to substituted and unsubstituted alky groups such as methyl, ethyl, propyl, butyl, pentyl, hexyl, heptyl, octyl, nonyl, decyl, undecyl, dodecyl and the like.
  • the phrase also includes branched chain isomers of straight chain alkyl groups, including but not limited to, the following which are provided by way of example: -CH(CH 3 ) 2 , -CH(CH 3 )(CH 2 CH 3 ), -CH(CH 2 CH 3 ) 2 , -C(CH 3 ) 3 , -C(CH 2 CH 3 ) 3) -CH 2 CH(CH 3 ) 2 , -CH 2 CH(CH 3 )(CH 2 CH 3 ), -CH 2 CH(CH 2 CH 3 ) 2 , -CH 2 C(CH 3 ) 3 , -CH 2 C(CH 2 CH 3 ) 3 , -CH(CH 3 )CH(CH 3 )(CH 2 CH 3 ), -CH 2 CH 2 CH(CH 3 ) 2 , -CH 2 CH 2 CH(CH 3 )(CH 2 CH 3 ), -CH 2 CH 2 CH(CH 3 )S, -CH 2 CH 2 C(CH 3 ) 3 ,
  • the phrase also includes cyclic alkyl groups such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and cyclooctyl and such rings substituted with straight and branched chain alkyl groups as defined above.
  • cyclic alkyl groups such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and cyclooctyl and such rings substituted with straight and branched chain alkyl groups as defined above.
  • polycyclic alkyl groups such as, but not limited to, adamantyl norbornyl, and bicyclo[2.2.2]octyl and such rings substituted with straight and branched chain alkyl groups as defined above.
  • alkyl also includes groups in which one or more bonds to a carbon(s) or hydrogen(s) are replaced by a bond to non-hydrogen and non-carbon atoms such as, but not limited to, a halogen atom in halides such as F, Cl, Br, and I; and oxygen atom in groups such as hydroxyl groups, alkoxy groups, aryloxy groups, and ester groups; a sulfur atom in groups such as thiol groups, alkyl and aryl sulfide groups, sulfone groups, sulfonyl groups, and sulfoxide groups; a nitrogen atom in groups such as amines, amides, alkylamines, dialkylamines, arylamines, alkylarylamines, diarylamines, N-oxides, imides, and enamines; a silicon atom in groups such as in trialkylsilyl groups, dialkylarylsilyl groups, alkyldiarylsilyl groups
  • aryl refers to substituted and unsubstituted aryl groups that do not contain heteroatoms.
  • the phrase includes, but is not limited to, groups such as phenyl, biphenyl, anthracenyl, naphthenyl by way of example.
  • Aryl groups also include those in which one of the aromatic carbons is bonded to a non-carbon or non- hydrogen atoms described above (in the alkyl definition) and also includes aryl groups in which one or more aromatic carbons of the aryl group is bonded to a substituted
  • aryl includes, but is not limited to tolyl, and hydroxyphenyl among others.
  • Alkenyl groups are those limited to having 2 to 15 carbon atoms and as many as 4 additional heteroatoms as described above. More preferred alkenyl groups have from 2 to 5 carbon atoms and as many as 2 heteroatoms.
  • alkoxy refers to substituted or unsubstituted alkoxy groups of the formula -O-alkyl, wherein the point of attachment is the oxy group and the alkyl group is as defined above.
  • Alkoxy groups are those limited to having 1 to 20 carbon atoms and as many as 5 additional heteroatoms, including the oxygen atom. More preferred alkoxy groups have from 1 to 5 carbon atoms and as many as 2 heteroatoms, including the oxygen atom.
  • alkynyl groups in which a non-carbon or non-hydrogen atom is bonded to a carbon triple bonded to another carbon and those in which a non-carbon or non-hydrogen atom is bonded to a carbon not involved in a triple bond to another carbon.
  • Alkynyl groups are those limited to having 2 to 15 carbon atoms and as many as 4 additional heteroatoms as described
  • More preferred alkynyl groups have from 2 to 5 carbon atoms and as many as 2 heteroatoms.
  • heterocyclyl refers to both aromatic and nonaromatic ring compounds including monocyclic, bicyclic, and polycyclic ring compounds such as, but not limited to, quinuclidyl, containing 3 or more ring members of which one or more is a heteroatom such as, but not limited to, N, O, and S.
  • heterocyclyl groups include, but are not limited to: unsaturated 3 to 8 membered rings containing 1 to 4 nitrogen atoms such as, but not limited to pyrrolyl, pyrrolinyl, imidazolyl, pyrazolyl, pyridyl, dihydropyridyl, pyrimidyl, pyrazinyl, pyridazinyl, triazolyl (e.g. 4H-l,2,4-triazolyl, lH-l,2,3-triazolyl, 2H-l,2,3-triazolyl etc.), tetrazolyl, (e.g.
  • saturated 3 to 8 membered rings containing 1 to 4 nitrogen atoms such as, but not limited to, pyrrolidinyl, imidazolidinyl, piperidinyl, piperazinyl; condensed unsaturated heterocyclic groups containing 1 to 4 nitrogen atoms such as, but not limited to, indolyl, isoindolyl, indolinyl, indolizinyl, benzimidazolyl, quinolyl, isoquinolyl, indazolyl, benzotriazolyl; unsaturated 3 to 8 membered rings containing 1 to 2 oxygen atoms such as, but not limited to furanyl; unsaturated 3 to 8 membered rings containing 1 to 2 oxygen atoms and 1 to 3 nitrogen atoms such as, but not limited to, oxazolyl, isoxazolyl, oxadiazolyl (e.g.
  • unsaturated 3 to 8 membered rings containing 1 to 3 sulfur atoms and 1 to 3 nitrogen atoms such as, but not limited to, thiazolyl, isothiazolyl, thiadiazolyl (e.g.
  • Ri i is alkyl, aryl or heterocyclyl
  • Ri 2 , Ri 3 Ri 4 , and R 15 are each independently H, -CN, -OH, halogen, alkyl, aryl, alkoxy, -NR a R b , -C(O)R 2 , -S(O) n Rd, or heterocyclyl; each R a and Rb is independently H, alkyl, -C(O)R 2 , aryl, heterocyclyl, or alkoxy; each R z is independently H, alkyl, alkoxy, -NH 2 , -NH(alkyl), -N(alkyl) 2 , aryl, or heterocyclyl; each Rd is independently H, alkyl, alkenyl, aryl, or -NR a R b ; each n is independently 0, 1, or 2; and each q is independently 0, 1, or 2; or a tautomer thereof, a pharmaceutically acceptable salt thereof, or a pharmaceutically acceptable salt of the tautomer.
  • Ri 1 is Rj i a :
  • Ri 6, Ri 7 and Ri 8 are each independently H, -CN, -OH, halogen, alkyl, aryl, alkoxy, - NR 3 Rb, -C(O)R 2 , -S(O) n Rd, or heterocyclyl.
  • said compound is:
  • oxygen atoms such as benzodioxolyl (e.g. 1,3-benzodioxoyl, etc.); unsaturated 3 to 8 membered rings containing an oxygen atom and 1 to 2 sulfur atoms such as, but not limited to, dihydrooxathiinyl; saturated 3 to 8 membered rings containing 1 to 2 oxygen atoms and 1 to 2 sulfur atoms such as 1,4-oxathiane; unsaturated condensed rings containing 1 to 2 sulfur atoms such as benzothienyl, benzodithiinyl; and unsaturated condensed heterocyclic rings containing an oxygen atom and 1 to 2 oxygen atoms such as benzoxathiinyl.
  • benzodioxolyl e.g. 1,3-benzodioxoyl, etc.
  • unsaturated 3 to 8 membered rings containing an oxygen atom and 1 to 2 sulfur atoms such as, but not limited to, dihydroo
  • Heterocyclyl group also include those described above in which one or more S atoms in the ring is double-bonded to one or two oxygen atoms (sulfoxides and sulfones).
  • heterocyclyl groups include tetrahydrothiophene, tetrahydrothiophene oxide, and tetrahydrothiophene 1,1- dioxide.
  • Preferred heterocyclyl groups contain 5 or 6 ring members.
  • More preferred heterocyclyl groups include morpholine, piperazine, piperidine, pyrrolidine, imidazole, pyrazole, 1,2,3-triazole, 1,2,4-triazole, tetrazole, thiomorpholine, thiomorpholine in which the S atom of the thiomorpholine is bonded to one or more O atoms, pyrrole, homopiperazine, oxazolidin-2-one, pyrrolidin-2-one, oxazole, quinuclidine, thiazole, isoxazole, furan, and tetrahydrofuran.
  • Heterocyclyl also refers to those groups as defined above in which one of the ring members is bonded to a non-hydrogen atom such as described above with respect to substituted alkyl groups and substituted aryl groups. Examples, include, but are not limited to, 2- methylbenzimidazolyl, 5-methylbenzimidazolyl, 5-chlorobenzthiazolyl, 1 -methyl piperazinyl, and 2-chloropyridyl among others. Heterocyclyl groups are those limited to having 2 to 15 carbon atoms and as many as 6 additional heteroatoms as described above. More preferred heterocyclyl groups have from 3 to 5 carbon atoms and as many as 2 heteroatoms.
  • substituted as applied to an undefined, yet well known in the art group, such as phenyl, will have the same meaning with respect to the optional appendages as described in the definition of alkyl.
  • the invention also includes isotopically-labeled compounds, that are structurally identical to those disclosed above, but for the fact that one or more atoms are replaced by an atom having an atomic mass or mass number different from the atomic mass or mass number usually found in nature.
  • isotopes that can be incorporated into compounds of the invention include isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorous, sulfur, fluorine and chlorine, such as 2 H, 3 H, 13 C, 14 C, 15 N, 18 0, 17 0, 31 P, 32 P, 35 S, 18 F and 36 Cl, respectively.
  • Compounds of the present invention, prodrugs thereof, and pharmaceutically acceptable salts of said compounds and of said prodrugs that contain the aforementioned isotopes and/or other isotopes of other atoms are within the scope of this invention.
  • Certain isotopically labeled compounds of the present invention, for example those into which radioactive isotopes such as H and C are incorporated, are useful in drug and/or substrate tissue distribution assays. Tritiated, i.e., 3 H, and carbon-14, i.e., 14 C, isotopes are particularly preferred for their ease of preparation and detectability.
  • isotopically labeled compounds of this invention and prodrugs thereof can generally be prepared by carrying out known or referenced procedures and by substituting a readily available isotopically labeled reagent for a non-isotopically labeled reagent.
  • IL-2 or "interleukin-2” indicates a lymphocyte that is produced by normal peripheral blood lymphocytes and is present in the body at low concentrations.
  • IL-2 was first described by Morgan et al. (1976) Science 193:1007- 1008 and originally called T-cell growth factor because of it's ability to induce proliferation of stimulated T lymphocytes. It is a protein with a reported molecular weight in the range of 13,000 to 17,000 (Gillis and Watson (1980) J. Exp. Med. 159: 1709) and has an isoelectric point in the range of 6-8.5.
  • IL-2 is intended to encompass any source of IL-2, including mammalian sources such as, e.g., mouse, rat, rabbit, primate, pig, pocket pet, and human, and may be native or obtained by recombinant techniques, such as recombinant IL-2 polypeptides produced by microbial hosts.
  • the IL-2 may be the native polypeptide sequence, or can be a variant of the native IL-2 polypeptide as described herein below, so long as the variant IL-2 polypeptide retains the IL-2 biological activity of interest as defined herein.
  • the IL-2 polypeptide or variant thereof is derived from a human source, and includes human IL-2 that is , recombinantly produced, such as recombinant human IL-2 polypeptides produced by microbial hosts, and variants thereof that retain the IL-2 biological activity of interest.
  • Any pharmaceutical composition comprising IL-2 as a therapeutically active component can be used to practice the present invention.
  • compositions of the present invention may be administered in conjunction with other anticancer agents.
  • compositions will either be formulated together as a combination therapeutic or administered separately.
  • Anticancer agents for use with the invention include, but are not limited to, one or more of the following set forth below: A. Kinase Inhibitors
  • EGFR Epidermal Growth Factor Receptor
  • small molecule quinazolines for example gefitinib (US 5457105, US 5616582, and US 5770599), ZD-6474 (WO 01/32651), erlotinib (Tarceva®, US 5,747,498 and WO 96/30347), and lapatinib (US 6,727,256 and WO 02/02552); Vascular Endothelial Growth Factor Receptor (VEGFR) kinase inhibitors, including SU-11248 (WO 01/60814), SU 5416 (US 5,883,113 and WO 99/61422), SU 6668 (US 5,883,113 and WO 99/61422), CHIR-258 (US 6,605,617 and US 6,774,237), vatalanib or PTK-787 (US 6,258,812), VEGF-787 (US 6,258,812), VEGF-787 (US 6,258,812), VEGF-787 (US 6,
  • Estrogen-targeting agents for use in anticancer therapy in conjunction with the compositions of the present invention include Selective Estrogen Receptor Modulators (SERMs) including tamoxifen, toremifene, raloxifene; aromatase inhibitors including Arimidex® or anastrozole; Estrogen Receptor Downregulators (ERDs) including Faslodex® or fulvestrant.
  • SERMs Selective Estrogen Receptor Modulators
  • ESDs Estrogen Receptor Downregulators
  • Androgen-targeting agents for use in anticancer therapy in conjunction with the compositions of the present invention include flutamide, bicalutamide, finasteride, aminoglutethamide, ketoconazole, and corticosteroids.
  • inhibitors for use as anticancer agents in conjunction with the compositions of the present invention include protein farnesyl transferase inhibitors " including tipifarnib or R-115777 (US 2003134846 and WO 97/21701), BMS-214662, AZD-3409, and FTI-277; topoisomerase inhibitors including merbarone and diflomotecan (BN-80915); mitotic kinesin spindle protein (KSP) inhibitors including SB-743921 and MKI-833; protease modulators such as bortezomib or Velcade® (US 5,780,454), XL-784; and cyclooxygenase 2 (COX-2) inhibitors including nonsteroidal antiinflammatory drugs I (NSAIDs).
  • protein farnesyl transferase inhibitors including tipifarnib or R-115777 (US 2003134846 and WO 97/21701), BMS-214662, AZD-3409, and FTI-277
  • Alkylating agents for use in conjunction with the compositions of the present invention for anticancer therapeutics include VNP-40101M or cloretizine, oxaliplatin (US 4,169,846, WO 03/24978 and WO 03/04505), glufosfamide, mafosfamide, etopophos (US 5,041,424), prednimustine; treosulfan; busulfan; irofluven (acylfulvene); penclomedine; pyrazoloacridine (PD-115934); O6-benzylguanine; decitabine (5-aza-2-deoxycytidine); brostallicin; mitomycin C (MitoExtra); TLK-286 (Telcyta®); temozolomide; trabectedin (US 5,478,932); AP-5280 (Platinate formulation of Cisplatin); porfiromycin; and clearazide (meclorethamine).
  • Chelating agents for use in conjunction with the compositions of the present invention for anticancer therapeutics include tetrathiomolybdate (WO 01/60814); RP- 697; Chimeric T84.66 (cT84.66); gadofosveset (Vasovist®); deferoxamine; and bleomycin optionally in combination with electorporation (EPT).
  • tetrathiomolybdate WO 01/60814
  • RP- 697 Chimeric T84.66 (cT84.66); gadofosveset (Vasovist®); deferoxamine; and bleomycin optionally in combination with electorporation (EPT).
  • EPT electorporation
  • Biological response modifiers for use in conjunction with the compositions of the present invention for anticancer therapeutics include staurosprine and macrocyclic analogs thereof, including UCN-01, CEP-701 and midostaurin (see WO 02/30941, WO 97/07081, WO 89/07105, US 5,621,100, WO 93/07153, WO 01/04125, WO 02/30941, WO 93/08809, WO 94/06799, WO 00/27422, WO 96/13506 and WO 88/07045); squalamine (WO 01/79255); DA-9601 (WO 98/04541 and US 6,025,387); alemtuzumab; interferons (e.g.
  • interleukins specifically IL-2 or aldesleukin as well as IL-I, IL-3, IL-4, IL-5, IL-6, IL-7, IL-8, IL-9, IL- 10, IL- 11 , IL- 12, and active biological variants thereof having amino acid sequences greater than 70% of the native human sequence; altretamine ( ⁇ exalen®); SU 101 or leflunomide (WO 04/06834 and US 6,331,555); imidazoquinolines such as resiquimod and imiquimod (US 4,689,338, 5,389,640, 5,268,376, 4,929,624, 5,266,575, 5,352,784, 5,494,916, 5,482,936, 5,346,905, 5,395,937, 5,238,944, and 5,525,612); and SMIPs, including benzazoles, anthraquinones, thiosemicarbazones, and tryptanthrins (WO 04
  • Anticancer vaccines for use in conjunction with the compositions of the present invention include Avicine® (Tetrahedron Letters 26, 1974 2269-70); oregovomab (OvaRex®); Theratope® (STn-KL ⁇ ); Melanoma Vaccines; GI-4000 series (GI-4014, GI-4015, and GI-4016), which are directed to five mutations in the Ras protein; GlioVax-1; MelaVax; Advexin® or INGN-201 (WO 95/12660); Sig/E7/LAMP-1, encoding ⁇ PV-16 E7; MAGE-3 Vaccine or M3TK (WO 94/05304); HER-2VAX; ACTIVE, which stimulates T-cells specific for tumors; GM-CSF cancer vaccine; and Listeria monocytogenes-based vaccines.
  • J. Antisense Therapy :
  • Anticancer agents for use in conjunction with the compositions of the present invention also include antisense compositions, such as AEG-35156 (GEM-640); AP- 12009 and AP-11014 (TGF-beta2-specific antisense oligonucleotides); AVI-4126; AVI-4557; AVI-4472; oblimersen (Genasense®); JFS2; aprinocarsen (WO 97/29780); GTI-2040 (R2 ribonucleotide reductase mRNA antisense oligo) (WO 98/05769); GTI-2501 (WO 98/05769); liposome-encapsulated c-Raf antisense oligodeoxynucleotides (LErafAON) (WO 98/43095); and Sirna-027 (RNAi-based therapeutic targeting VEGFR-I mRNA).
  • AEG-35156 GEM-640
  • IL-2 compound is "aldesleukin” or "Proleukin®", manufactured by Chiron Corporation of Emeryville, California.
  • the IL-2 in this formulation is a recombinantly produced, unglycosylated human IL-2 mutein which differs from the native human IL-2 amino acid sequence in having the initial alanine residue eliminated and the cysteine residue at position 125 replaced by a serine residue (referred to as des-alanyl-1, serine-125 human interleukin-2).
  • This IL-2 mutein can be expressed in E. coli, and subsequently purified by diafiltration and cation exchange chromatography as described in U.S. Patent No. 4,931,543.
  • Aldesleukin has been compared to native (Jurkat) IL-2 in vitro. No significant differences have been seen and in vivo induction of cytolytic cells in mice and serum half-life following IV administration is equivalent for aldesleukin and native (Jurkat) IL-2; although there are beneficial attributes of the form of IL-2 encompassed by aldesleukin and therefore reference to "aldesleukin" encompasses only that composition and not all possible forms of the IL-2 protein.
  • the pharmaceutical compositions useful in the methods of the invention may comprise biologically active variants of IL-2.
  • Such variants should retain the desired biological activity of the native polypeptide such that the pharmaceutical composition comprising the variant polypeptide has the same therapeutic effect as the pharmaceutical composition comprising the native polypeptide when administered to a subject. That is, the variant polypeptide will serve as a therapeutically active component in the pharmaceutical composition in a manner similar to that observed for the native polypeptide.
  • Methods are available in the art for determining whether a variant polypeptide retains the desired biological activity, and hence serves as a therapeutically active component in the pharmaceutical composition.
  • Biological activity can be measured using assays specifically designed for measuring activity of the native polypeptide or protein, including assays described in the present inventions.
  • antibodies raised against a biologically active native polypeptide can be tested for their ability to bind to the variant polypeptide, where effective binding is indicative of a polypeptide having a conformation similar to that of the native polypeptide.
  • the IL-2 biological activity of interest is the ability of IL-2 to activate and/or expand natural killer (NK) cells to mediate lymphokine activated killer (LAK) activity and antibody-dependent cellular cytotoxicity (ADCC).
  • NK natural killer
  • LAK lymphokine activated killer
  • ADCC antibody-dependent cellular cytotoxicity
  • an IL-2 variant for example, a mutein of human IL-2
  • Assays to determine IL-2 activation or expansion of NK cells and mediation of LAC or ADCC activity are well known in the art.
  • Suitable biologically active variants of native or naturally occurring IL-2 can be fragments, analogues, and derivatives of that polypeptide.
  • fragment is intended a polypeptide consisting of only a part of the intact polypeptide sequence and structure, and can be a C-terminal deletion or N-terminal deletion of the native polypeptide.
  • analogue is intended an analogue of either the native polypeptide or of a fragment of the native polypeptide, where the analogue comprises a native polypeptide sequence and structure having one or more amino acid substitutions, insertions, or deletions.
  • “Muteins”, such as those described herein, and peptides having one or more peptoids (peptide mimics) are also encompassed by the term analogue (see International Publication No. WO 91/04282).
  • “derivative” is intended any suitable modification of the native polypeptide of interest, of a fragment of the native polypeptide, or of their respective analogues, such as glycosylation, phosphorylation, polymer conjugation (such as with polyethylene glycol), or other addition of foreign moieties, so long as the desired biological activity of the native polypeptide is retained.
  • Methods for making polypeptide fragments, analogues, and derivatives are generally available in the art.
  • amino acid sequence variants of the polypeptide can be prepared by mutations in the cloned DNA sequence encoding the native polypeptide of interest.
  • Methods for mutagenesis and nucleotide sequence alterations are well known in the art. See, for example, Walker and Gaastra, eds. (1983) Techniques in Molecular Biology (MacMillan Publishing Company, New York); Kunkel (1985) Proc. Natl. Acad. Sci. USA 82:488-492; Kunkel et al. (1987) Methods Enzymol. 154:367-382; Sambrook et al. ( 1989) Molecular Cloning: A Laboratory Manual (Cold Spring
  • variants of the IL-2 polypeptide of interest modifications are made such that variants continue to possess the desired activity. Any mutations made in the DNA encoding the variant polypeptide must not place the sequence out of reading frame and preferably will not create complementary regions that could produce a secondary mRNA structure.
  • Biologically active variants of IL-2 will generally have at least 70%, preferably at least 80%, more preferably about 90% to 95% or more, and most preferably about 98% or more amino acid sequence identity to the amino acid sequence of the reference polypeptide molecule, which serves as the basis for comparison.
  • a biologically active variant thereof will have at least 70%, preferably at least 80%, more preferably about 90% to 95% or more, and most preferably about 98% or more sequence identity to the amino acid sequence for human IL-2.
  • a biologically active variant of a native polypeptide of interest may differ from the native polypeptide by as few as 1-15 amino acids, as few as 1-10, such as 6-10, as few as 5, as few as 4, 3, 2, or even 1 amino acid residue.
  • sequence identity is intended the same amino acid residues are found within the variant polypeptide and the polypeptide molecule that serves as a reference when a specified, contiguous segment of the amino acid sequence of the variants is aligned and compared to the amino acid sequence of the reference molecule.
  • the percentage sequence identity between two amino acid sequences is calculated by determining the number of positions at which the identical amino acid residue occurs in both sequences to yield the number of matched positions, dividing the number of matched positions by the total number of positions in the segment undergoing comparison to the reference molecule, and multiplying the result by 100 to yield the percentage of sequence identity.
  • the determination of percent identity between any two sequences can be accomplished using a mathematical algorithm.
  • naturally or non-naturally occurring variants of IL-2 have amino acid sequences that are at least 70%, preferably 80%, more preferably, 85%, 90%, 91%, 92 %, 93%, 94% or 95% identical to the amino acid sequence to the reference molecule, for example, the native human IL-2, or to a shorter portion of the reference IL-2 molecule. More preferably, the molecules are 96%, 97%, 98% or 99% identical.
  • Percent sequence identity is determined using the Smith- Waterman homology search algorithm using an affined gap search with a gap open penalty of 12 and a gap extension penalty of 2, BLOSUM matrix of 62.
  • a variant may, for example, differ by as few as 1 to 10 amino acid residues, such as 6-10, as few as 5, as few as 4, 3, 2, or even 1 amino aid residue.
  • the contiguous segment of the variant amino acid sequence may have additional amino acid residues or deleted amino acid residues with respect to the reference amino acid sequence.
  • the contiguous segment used for comparison to the reference amino acid sequence will include at least twenty (20) contiguous amino acid residues, and may be 30, 40, 50, or more amino acid residues. Corrections for sequence identity associated with conservative residue substitutions or gaps can be made (see Smith-Waterman homology search algorithm).
  • polypeptide having IL-2 activity depends on a number of factors. As ionizable amino and carboxyl groups are present in the molecule, a particular polypeptide may be obtained as an acidic or basic salt, or in neutral form. All such preparations that retain their biological activity when placed in suitable environmental conditions are included in the definition of polypeptides having IL-2 activity as used herein.
  • the primary amino acid sequence of the polypeptide may be augmented by derivatization using sugar moieties (glycosylation) or by other supplementary molecules such as lipids, phosphate, acetyl groups and the like. It may also be augmented by conjugation with saccharides. Certain aspects of such augmentation are accomplished through post-translational processing systems of the producing host; other such modifications may be introduced in vitro. In any event, such modifications are included in the definition of an IL-2 polypeptide used herein so long as the IL-2 activity of the polypeptide is not destroyed. It is expected that such modifications may quantitatively or qualitatively affect the activity, either by enhancing or diminishing the activity of the polypeptide, in the various assays. Further, individual amino acid residues in the chain may be modified by oxidation, reduction, or other derivatization, and the polypeptide may be cleaved to obtain fragments that retain activity.
  • IL-2 variants are suitable for use as a therapeutically active component of a pharmaceutical composition used in the methods of the present invention.
  • the IL-2 or variants thereof for use in the methods of the present invention may be from any source, but preferably is recombinant IL-2.
  • recombinant IL-2 is intended interleukin-2 that has comparable biological activity to native-sequence IL-2 and that has been prepared by recombinant DNA techniques as described, for example, by Taniguchi et al.
  • IL-2 nucleic Acids Research 11 :4307-4323 or mutationally altered IL-2 as described by Wang et al. (1984) Science 224:1431-1433.
  • the gene coding for IL-2 is cloned and then expressed in transformed organisms, preferably a microorganism, and most preferably E. coli, as described herein.
  • the host organism expresses the foreign gene to produce IL-2 under expression conditions.
  • Synthetic recombinant IL-2 can also be made in eukaryotes, such as yeast or human cells. Processes for growing, harvesting, disrupting, or extracting the IL-2 from cells are substantially described in, for example, U.S. Patent Nos.
  • EP 136,489 (which discloses one or more of the following alterations in the amino acid sequence of naturally occurring IL-2: Asn26 to Gln26; Trpl21 to Phel21; Cys58 to Ser58 or Ala58, CyslO5 to SerlO5 or AlalO5; Cysl25 to Serl25 or Alal25; deletion of all residues following Arg 120; and the Met-1 forms thereof); and the recombinant IL-2 muteins described in European Patent Application No. 83306221.9, filed October 13, 1983 (published May 30, 1984 under Publication No. EP 109,748), which is the equivalent to Belgian Patent No. 893,016, and commonly owned U.S. Patent No.
  • 4,752,585 (which discloses the following variant IL-2 proteins: alalO4 serl25 IL- 2, alalO4 IL-2, alalO4 alal25 IL-2, vallO4 ser!25 IL-2, vallO4 IL-2, vallO4 alal25 IL-2, des-alal alalO4 serl25 IL-2, des-alal alalO4 IL-2, 10 des-alal alalO4 alal25 IL-2, des-alal vallO4 serl25 IL-2, des-alal vallO4 IL-2, des-alal vallO4 alal25 IL-2, des-alal des-pro2 alalO4 serl25 IL-2, des-alal des-pro2 alalO4 serl25 IL-2, des-alal des-pro2 alalO4 serl25 IL-2, des-alal des-pro2 alalO4 IL- 2, des-alal des-pro2 alalO4 aial25 IL
  • Patent No. 4,931 ,543 (which discloses the IL-2 mutein des-alanyl- 1 , serine 30 125 human IL-2 used in the examples herein, as well as the other IL-2 muteins). Also see European Patent Publication No. EP 200,280 (published December 10, 1986), which discloses recombinant IL-2 muteins wherein the methionine at position 104 has been replaced by a conservative amino acid.
  • Examples include the following muteins: ser4 des-ser5 alalO4 IL-2; des- alal des-pro2 des-thr3 des-ser4 des-ser5 alalO4 alal25 IL-2; des-alal des-pro2 des-thr3 des-ser4 des-ser5 glulO4 serl25 IL-2; des-alal des-pro2 des-thr3 des ser4 des-ser5 glulO4 IL-2; des-alal des-pro2 des-thr3 des-ser4 des-ser5 glul04 alal25 IL-2; des-alal des-pro2 des-thr3 des-ser4 des-ser5 des-ser6 alalO4 alal25 IL-2; des-alal des-pro2 des-thr3 des-ser4 des-ser5 des-ser6 alalO4 alal25 IL-2; des-alal des-pro2 des-thr3 des-ser4 des
  • IL-2 muteins include the those disclosed in WO 99/60128 (substitutions of the aspartate at position 20 with histidine or isoleucine, the asparagine at position 88 with arginine, glycine, or isoleucine, or the glutamine at positionl26 with leucine or gulatamic acid), which reportedly have selective activity for high affinity IL-2 receptors expressed by cells expressing T cell receptors in preference to NK cells and reduced IL-2 toxicity; the muteins disclosed in U.S Patent No. 5,229, 109
  • WO 00/04048 (corresponding to the first 30 amino acids of IL-2, which contains the entire a-helix A of IL-2 and interacts with the b chain of the IL-2 receptor), which reportedly stimulates NK cells and induction of LAK cells; and a mutant form of the IL-2 pl-30 peptide also disclosed in WO 00/04048 (substitution of aspartic acid at position 20 with lysine), which reportedly is unable to induce vascular bleeds but remains capable of generating LAK cells.
  • IL-2 can be modified with polyethylene glycol to provide enhanced solubility and an altered pharmokinetic profile (see U.S. Patent No. 4,766,106).
  • IL-2 as used herein is also intended to include IL-2 fusions or conjugates comprising IL-2 fused to a second protein or covalently conjugated to polyproline or a water soluble polymer to reduce dosing frequencies or to improve IL- 2 tolerability.
  • the IL-2 (or a variant thereof as defined herein) can be fused to human albumin or an albumin fragment using methods known in the art (see WO 01/79258).
  • the IL-2 can be covalently conjugated to polyproline or polyethylene glycol homopolymers and polyoxyethylated polyols, wherein the homopolymer is unsubstituted or substituted at one end with an alkyl group and the poplyol is unsubstituted, using methods known in the art (see, for example, U.S.
  • compositions comprising IL-2 as the therapeutically active component can be used in the methods of the invention.
  • Such pharmaceutical compositions are known in the art and include, but are not limited to, those disclosed in U.S. Patent Nos. 4,745,180; 4,766,106; 4,816,440; 4,894,226; 4,931 ,544; and 5,078,997.
  • liquid, lyophilized, or spray- dried compositions comprising IL-2 or variants thereof that are known in the art may be prepared as an aqueous or nonaqueous solution or suspension for subsequent administration to a subject in accordance with the methods of the invention.
  • compositions will comprise IL-2 or variants thereof as a therapeutically or prophylactically active component.
  • therapeutically or prophylactically active component is intended the IL-2 or variants thereof is specifically incorporated into the composition to bring about a desired therapeutic or prophylactic response with regard to treatment, prevention, or diagnosis of a disease or condition within a subject when the pharmaceutical composition is administered to that subject.
  • the pharmaceutical compositions comprise appropriate stabilizing agents, bulking agents, or both to minimize problems associated with loss of protein stability and biological activity during preparation and storage.
  • the IL-2 containing pharmaceutical compositions useful in the methods of the invention are compositions comprising stabilized monomelic IL-2 or variants thereof, compositions comprising multimeric IL-2 or variants thereof, and compositions comprising stabilized lyophilized or spray-dried IL-2 or variants thereof.
  • compositions comprising stabilized monomeric IL-2 or variants thereof are disclosed in International Publication No. WO 01/24814, entitled “Stabilized Liquid Polypeptide-Containing Pharmaceutical Compositions.”
  • IL-2 is intended the protein molecules are present substantially in their monomer form, not in an aggregated form, in the pharmaceutical compositions described herein. Hence covalent or hydrophobic oligomers or aggregates of IL-2 are not present.
  • the IL-2 in these liquid compositions is formulated with an amount of an amino acid base sufficient to decrease aggregate formation of IL-2 during storage.
  • the amino acid base is an amino acid or a combination of amino acids, where any given amino acid is present either in its free base form or in its salt form.
  • Preferred amino acids are selected from the group consisting of arginine, lysine, aspartic acid, and glutamic acid.
  • compositions further comprise a buffering agent to maintain phi of the liquid compositions within an acceptable range for stability of IL-2, where the buffering agent is an acid substantially free of its salt form, an acid in its salt form, or a mixture of an acid and its salt fond.
  • the acid is selected from the group consisting of succinic acid, citric acid, phosphoric acid, and glutamic acid.
  • Such compositions are referred to herein as stabilized monomeric IL-2 pharmaceutical compositions.
  • the amino acid base in these compositions serves to stabilize the IL-2 against aggregate formation during storage of the liquid pharmaceutical composition, while use of an acid substantially free of its salt form, an acid in its salt form, or a mixture of an acid and its salt form as the buffering agent results in a liquid composition having an osmolality that is nearly isotonic.
  • the liquid pharmaceutical composition may additionally incorporate other stabilizing agents, more particularly methionine, a nonionic surfactant such as polysorbate 80, and EDTA, to further increase stability of the polypeptide.
  • Such liquid pharmaceutical compositions are said to be stabilized, as addition of amino acid base in combination with an acid substantially free of its salt fond, an acid in its salt fonn, or a mixture of an acid and its salt form, results in the compositions having increased storage stability relative to liquid pharmaceutical compositions formulated in the absence of the combination of these two components.
  • These liquid pharmaceutical compositions comprising stabilized monomelic IL-2 may either be used in an aqueous liquid form, or stored for later use in a frozen state, or in a dried form for later reconstitution into a liquid fond or other form suitable for administration to a subject in accordance with the methods of present invention.
  • liquid pharmaceutical composition or fonnulation is dried either by freeze drying (i.e., lyophilization; see, for example, Williams and Polli (1984) J. Parenteral Sci. Technol. 38:48 59), spray drying (see Masters (1991) in Spray- Drying Handbook (Sth ed; Longman Scientific and Technical, Essez, U.K.), pp. 491-676; Broadhead et al. (1992) Drug Devel. Ind. Pharm. 18:1169-1206; and Mumenthaler et al. (1994) Pharm. Res. 11 :12- 20, or air drying (Carpenter and Crowe (1988) Cryobiology 25:459-470; and Roser (1991) Biopharm. 4:47-53).
  • freeze drying i.e., lyophilization
  • spray drying see Masters (1991) in Spray- Drying Handbook (Sth ed; Longman Scientific and Technical, Essez, U.K.), pp. 491-676; Broadhead et al. (19
  • IL-2 formulations that comprise IL-2 in its nonaggregated monomeric state include those described in Whittington and Faulds (1993) Drugs 46(3):446- 514.
  • These formulations include the recombinant IL-2 product in which the recombinant IL- 2 mutein Teceleukin (unglycosylated human IL-2 with a methionine residue added at the amino-terminal) is formulated with 0.25% human serum albumin in a lyophilized powder that is reconstituted in isotonic saline, and the recombinant IL-2 mutein Bioleukin (human IL-2 with a methionine residue added at the amino-terminal, and a substitution of the cysteine residue at position 125 of the human IL-2 sequence with alanine) fonnulated such that 0.1 to 1.0 mg/ml IL-2 mutein is combined with acid, wherein the fonnulation has a pH of 3.0 to 4.0, 10
  • multimeric IL-2 examples of pharmaceutical compositions comprising multimeric IL-2 are disclosed in commonly owned U.S. Patent No.4,604,377.
  • multimeric is intended the protein molecules are present in the pharmaceutical composition in a microaggregated form having an average molecular association of 10-50 molecules. These multimers are present as loosely bound, physically associated IL-2 molecules. A lyophilized form of these compositions is available commercially under the tradename Proleukin® IL-2 (Chiron Corporation, Emeryville, California).
  • the lyophilized formulations disclosed in this reference comprise selectively oxidized, microbially produced recombinant IL-2 in which the recombinant IL-2 is admixed with a water soluble carrier such as mannitol that provides bulk, and a sufficient amount of sodium dodecyl sulfate to ensure the solubility of the recombinant IL-2 in water.
  • a water soluble carrier such as mannitol that provides bulk
  • compositions are suitable for reconstitution in aqueous injections for parenteral administration and are stable and well tolerated in human patients.
  • the IL-2 When reconstituted, the IL-2 retains its multimeric state.
  • Such lyophilized or liquid compositions comprising multimeric IL-2 are encompassed by the methods of the present invention.
  • Such compositions are referred to herein as multimeric IL-2 pharmaceutical compositions.
  • the methods of the present invention may also use stabilized lyophilized or spray-dried pharmaceutical compositions comprising IL-2, which may be reconstituted into a liquid or other suitable form for administration in accordance with methods of the invention.
  • Such pharmaceutical compositions are disclosed in International Publication No. W O 01/49274 entitled "Methods for Pulmonary Del vefy of InterleuklTz-2.”
  • These compositions may further comprise at least one bulking agent, at least one agent in an amount sufficient to stabilize the protein during the drying process, or both.
  • IL-2 protein or variants thereof retains its monomelic or multimeric form as well as its other key properties of quality, purity, and potency following lyophilization or spray-drying to obtain the solid or dry powder form of the composition.
  • preferred carrier materials for use as a bulking agent include glycine, mannitol, alanine, valine, or any combination thereof, most preferably glycine.
  • the bulking agent is present in the formulation in the range of 0% to about 10% (w/v), depending upon the agent used.
  • Preferred carrier materials for use as a stabilizing agent include any sugar or sugar alcohol or any amino acid.
  • Preferred sugars include sucrose, trehalose, raffinose, stachyose, sorbitol, glucose, lactose, dextrose or any combination thereof, preferably sucrose.
  • the stabilizing agent is a sugar, it is present in the range of about 0% to about 9.0% (w/v), preferably about 0.5% to about 5.0%, more preferably about 1.0% to about 3.0%, most preferably about 1.0%.
  • the stabilizing agent is an amino acid, it is present in the range of about 0% to about 1.0% (w/v), preferably about 0.3% to about 0.7%, most preferably about 0.5%.
  • These stabilized lyophilized or spray- dried compositions may optionally comprise methionine, ethylenediaminetetracetic acid (EDTA) or one of its salts such as disodium EDTA or other chelating agent, which protect the IL-2 or variants thereof against methionine oxidation. Use of these agents in this manner is described in U.S. Application Serial No. 09/677,643, herein incorporated by reference.
  • the stabilized lyophilized or spray- dried compositions may be formulated using a buffering agent, which maintains the pH of the pharmaceutical composition within an acceptable range, preferably between about pH 4.0 to about pH 8.5, when in a liquid phase, such as during the formulation process or following reconstitution of the dried form of the composition. Buffers are chosen such that they are compatible with the drying process and do not affect the quality, purity, potency, and stability of the protein during processing and upon storage.
  • IL-2 pharmaceutical compositions represent suitable compositions for use in the methods of the invention.
  • any pharmaceutical composition comprising an IL-2 compound as a therapeutically active component is encompassed by the methods of the invention.
  • kits or package containing at least one combination composition of the invention, accompanied by instructions for use.
  • the kit comprises each of the drugs, along with instructions for use.
  • the drug components may be packaged in any manner suitable for administration, so long as the packaging, when considered along with the instructions for administration, clearly indicates the manner in which each of the drug components is to be administered.
  • each of the drug components of the combination may be combined into a single administrable dosage form such as a single composition.
  • kits comprising rIL-2 and an antiangiogenic agent
  • the kit may be organized by any appropriate time period, such as by day.
  • a representative kit may comprise unit dosages of each of rIL-2 and the antiangiogenic agent. If each of the drugs is to be administered twice daily, then the kit may contain, corresponding to Day 1, two rows of unit dosage forms of each of rIL-2 and the antiangiogenic agent, with instructions for the timing of administration.
  • the kit may contain, corresponding to Day 1, two rows of unit dosage forms of each of rIL-2 and the antiangiogenic agent, with instructions for the timing of administration.
  • one or more of the drugs differs in the timing or quantity of drug to be administered in comparison to the other drug members of the combination, then such would be reflected in the packaging and instructions.
  • exemplary Day 1 packaging might correspond to unit dosage forms of rIL-2 as "Day 1, Dose 1", along with dosage forms for the antiangiogenic agent corresponding to "Day 1, Dose 2".
  • the packaging may be in any form commonly employed for the packaging of pharmaceuticals, and may utilize any of a number of features such as different colors, wrapping, tamper-resistant packaging, blister paks, dessicants, and the like.
  • compositions for Co-administration with rIL-2 A Compositions for Co-administration with rIL-2 A. COMPOUNDS
  • Scheme 1 describes a modular method of synthesizing a myriad of substituted quinazoline compounds.
  • Reference to AG indicates an activating group such as, for example a halide, triflate, or ketone, wherein as many as four AG groups may be present. Since the 4-chloro position, shown in the third step, is more active than positions 5-8 on the benzyl ring, displacement with NHRR' may proceed without much concomitant displacement of AG in any of positions 5-8. Subsequent displacement of the AG group for R" in the final step may proceed in the presence of a weak to moderate base.
  • the AG group(s) may be modified to yield the desired product, for example a NO 2 group may be reduced with Fe and AcOH in EtOH and H 2 O to yield an amino substituent, which may be further substituted, for example by reductive amination with paraformaldehyde.
  • the AG group(s) in the starting material may be a substituent desired in the final product (as in R"), such as, for example an alkoxy group(s) or a substituted alkyl group(s).
  • a number functionalized 2-nitrobenzamide starting materials are available, which are easily converted to the 2-aminobenzamide starting material in the presence of a reducing agent, such as H 2 /Pd/C in EtOH.
  • Scheme 2a is performed as a one-pot procedure, with reagents in step a being NH 4 OH, CuCl in H 2 O.
  • reagents in step a being NH 4 OH, CuCl in H 2 O.
  • R 2 -R 5 are as defined herein.
  • reaction may be heated to enhance the yield, depending on reactivity of the particular starting materials.
  • R 9 as shown in scheme 2, is H, -OH, -CN, alkyl, aryl, heterocyclyl, alkoxy, or -NR 3 R b as defined herein. It is contemplated that the above structure may replace Formula II to allow substitution at R ⁇ >, whereby all other substituents are as defined herein.
  • the filter residue is dissolved in about 200 ml hot methanol, the solution is treated with 0.75 g activated carbon and filtered Via a Hyfio Super CeI, and the pH of the filtrate is adjusted to about 2.5 with 7 ml 3N methanolic HCl.
  • the filtrate is evaporated to about half the original volume and ether added until slight turbidity occurs; cooling then leads to the precipitation of crystals.
  • the crystallizate is filtered off, washed with a mixture of methanol/ether (1:2) as well as ether, dried for 8 h at 110° C. under HV, and equilibrated for 72 h at 20° C. and in room atmosphere.
  • Messenger RNA from human Jurkat cell line is used to create double stranded cDNA, which is hybridized into pBR322 plasmids.
  • a clone containing the IL-2 gene is identified using a 32 P-labeled oligonucleotide probe corresponding to a short length IL-2 base sequence.
  • the gene is inserted into a region of the pBR322 plasmid that has a convenient restriction site.
  • the appropriate promoter and ribosome-binding site is inserted in front of the IL-2 gene, and the resulting expression clone encodes a modified recombinant rIL-2.
  • Proleukin ® is as a lyophilized cake in 5 cc vials containing 1.3 mg of protein. Vials of Proleukin ® for injection are reconstituted with 1.2 niL of Sterile Water for Injection, USP. The diluent is directed against the side of the vial to avoid excess foaming, swirling contents gently until completely dissolved, while avoiding shaking. When reconstituted each mL contains 1.1 mg (18 million IU) of Proleukin ® . Reconstituted Proleukin® is suitable for intravenous injection directly or may be diluted as necessary in volumes of 50 mL to 500 mL with 5% Dextrose Injection, USP with 0.1% Albumin Human, USP. When diluting, the Albumin Human, USP is added to the 5% Dextrose Injection, USP prior to the addition of the reconstituted Proleukin ® .
  • rIL-2 recombinant human interIeukin-2; MIU: million International Units; CR: complete response; PR: partial response; NS: not stated; i.v.: intravenous.
  • Antiangiogenic agent is given again on day 1 and then every 2 weeks continuously in an 8-weeks treatment cycle.
  • Treatment with rIL-2 is continued for 6 consecutive weeks (days 1 - 42, Monday- Friday of each week) followed by a 2-week rest period resulting in an 8- week treatment cycle.
  • Example 4 Combination Therapy with rIL-2 and Small Molecule Receptor Tyrosine Kinase
  • Inhibitors BAY 43-9006 and SU11248 A Material and Methods Drugs
  • Recombinant human Interleukin-2 (Proleukin ® , Aldesleukin/rIL-2); 18 MIU/ ml, Chiron Corporation, Emeryville, CA) was reconstituted with sterile water for injection and formulated in 5% dextrose prior to administration.
  • Vincristine (vincristine sulfate) was from Mayne Pharma Ltd (Mulgrave, Australia).
  • CHIR-258 is 4-amino-5-fluoro-3-[5-(4-methylpiperazin- 1 -yl)- 1 H-benzimidazol-2-yl]quinolin- 2(lH)-one (Chiron).
  • BAY 43-9006 (Sorafanib/Nexavar®) (Riedl et al., Proc. Am. Assoc. Cancer Res. 2001 42(Abs 4956); Lowinger et al., Curr. Pharm. Des. 2002 8(25):2269-2278; WO 9932455) and SUl 1248 (Sunitinib/Sutent®) (Sun et al., J. Med. Chem. 2003 46(7):1116-1119; WO 0160814) were synthesized and purified in- house according to published procedures and patents.
  • BAY 43- 9006 or SUl 1248 (20 mM) were prepared in DMSO, and aliquots will be stored at - 20 0 C prior to use.
  • all drugs were diluted in optimal culture medium.
  • BAY 43-9006 was formulated in 100% PEG 400 vehicle, whereas, SUl 1248 dosing solutions were prepared in 5 mM citrate buffer. All other chemicals used were of research grade.
  • CTLL-2 IL-2 dependent T cell line
  • B16-F10 melanoma CT26 colon
  • renal carcinoma RENCA renal carcinoma RENCA
  • CTLL-2 were grown in RPMIl 640 supplemented with 10% FBS (fetal bovine serum, Gibco Life Technologies, Gaithersburg, MD), 2 mM L-glutamine, 1 mM sodium pyruvate, 25 mM HEPES, 0.5 nM rIL-2, 2 mM /3-mercaptoethanol.
  • RENCA cells were cultured in a media containing EMEM with 10% FBS, 2% IOOX vitamin, 1% 200 mM glutmine,l% 100 mM NaPy, 1% nonessential amino acids.
  • media contained EMEM, 10% FBS, 2% vitamins, 1% 200 mM glutamine, 1% 100 mM sodium pyruvate, 1% nonessential amino acids.
  • B16-F10 cells were grown in RPMI 1640 with 10% FBS, 1% nonessential amino acids, 1% 10OmM sodium pyruvate 2% vitamins; 2 mM l-glutamine; 2% sodium bicarbonate.
  • Yac-1 cells were cultured in RPMI + 10% FBS and subcultured 1-2 days prior to assay to ensure log- phase growth. Cells were maintained as suspension or adherent cultures in a humidified atmosphere at 37 0 C and 5% CO 2 . Cells were used in exponential growth phase (not exceeding 6-8 passages) with viability >98% (assessed using trypan blue staining) and determined free of mycoplasma.
  • mice Female BALB/c or C57BL6 mice (4-6 week-old, 18-22 g) were obtained from Charles River (Wilmington, MA) and acclimated for 1 week in pathogen-free enclosures prior to the start of the study. Animals received sterile rodent chow and water ad libitum and were housed in sterile filter-top cages with 12 hour light/dark cycles. All experiments were under the guidelines of the Association for Assessment and Accreditation of Laboratory Animal Care International.
  • B16-F10 (2 x 10 6 ), CT26 (2 x 10 6 ) or RENCA (1 x 10 6 ) cells were harvested, washed three times, and resuspended in PBS. Mice were shaved on the flank, and implanted (0.2 ml) subcutaneously (s.c.) into the right flank of mice.
  • B16-F10 tumor model C57BL6 mice were used, whereas CT26 and RENCA tumors were implanted in BALB/c mice. Treatments were initiated when tumors were established to a mean size of 50-250 mm 3 (day 0) as outlined in specific study designs. Mice were randomized into treatment cohorts (typically 10 mice/group).
  • rIL-2 was administered daily s.c. (0.2-3 mg/kg/day) on days 0-6 or 7-13 or days 0-4, 7-11.
  • BAY 43-9006 or SUl 1248 (1-100 mg/kg) were administered daily (for 5-12 days) as a solution via oral gavage, starting day 0 or day 7. All monotherapy and drug combinations at selected doses, outlined in individual studies were well tolerated.
  • Tumor volumes and body weights were assessed 2-3 times weekly. Caliper measurements of tumors were converted into mean tumor volume (mm 3 ) using the formula: 1 A (length (mm) x [width (mm)] 2 ). Tumor growth inhibition (TGI) was calculated as [l-(mean tumor volume of treated group/mean tumor volume of control group) x 100]. Responses were defined as either a complete response (CR, no measurable tumor), or partial response (PR 5 50-99% tumor volume reduction) compared to tumor volume for each animal at treatment initiation. Tumor growth delay analysis was calculated as: [(number of days for a treated group to reach a mean tumor volume of 1000 mm 3 ) - (number of days for the control group to reach a mean tumor volume of 1000 mm 3 )].
  • mice were treated with a single s.c. dose of rIL-2 (6 mg/kg, 0.2 ml) or BAY 43-9006 (20 mg/kg, p.o., 0.2 ml) and blood was collected at various times after drug administration. Plasma levels of BAY 43- 9006 and rIL-2 were determined using HPLC or an ELISA bioassay.
  • pERK detection was done with a mouse antibody to pERK (1 : 1000, Cell Signaling, Beverly, MA) and incubated at 4 0 C overnight.
  • Detection of pSTAT5 antibody (1 : 1000 Upstate) and pAKT (1 : 1000 Cell Signaling) was performed with the same amount of protein by probing with appropriate anti-phosphotyrosine antibodies for 2 hours at room temperature.
  • the membranes were then incubated for 1 hour at room temperature with 1 : 5000 horseradish peroxidase-conjugated anti-rabbit IgG (Jackson Immunoresearch, West Grove, PA).
  • blots were stripped and re-probed with anti- ERK (Cell Signaling), anti-STAT5 (BD Biosciences), and anti-AKT (Cell Signaling) antibodies to measure total ERK, STAT5 and AKT protein, respectively. Proteins were detected using enhanced chemiluminescence (ECL; Amersham Biosciences, Buckinghamshire, England) and visualized after exposure to Kodak film. Scanning densitometry was performed to quantify band intensities. The amount pERK, pSTAT5 or pAKT was normalized to total ERK, STAT5 or AKT protein levels, and compared with vehicle or untreated controls.
  • ECL enhanced chemiluminescence
  • Murine T cells were detected with a rabbit anti-CD3 antibody (1 :60 dilution, Dako Norden A/S, Glostrup Denmark), and monocytes/macrophages were detected using F4/80 (Serotec).
  • mouse tumors were stained for Ki-67 using a monoclonal rat anti-mouse antibody (1: 15 dilution, DAKO). Heat-induced epitope retrieval was performed using CC 1 (Ventana Medical Systems). Samples were then incubated with the appropriate secondary antibodies (goat anti-rabbit IgG biotinylated antibody, 1 : 100 dilution, Jackson ImmunoReseach Laboratories).
  • a horseradish peroxidase-labelled streptavidin biotin system with 3-3'-diaminobenzidine chromogen (Ventana Medical Systems) was used for localizing the antibodies. Sections were counterstained with nuclear fast red to enhance visualization of tissue morphology.
  • the Ventana Bluemap kit - an alternative NBT/BCIP staining method was used due to observed melanin deposits in H&E tissues, which aided in visualization of T cells in tumors.
  • CTLL-2 T cells were preincubated with or without BAY 43-9006 (3 ⁇ M) at 37 0 C for 2 hours. The cells were then washed and plated in 96-well microtiter plates, 5,000 cells/well in culture media with serial dilutions of rhIL-2 (1 pM to 100 nM). At the end of the incubation period (72 hours at 37 0 C), cell viability was determined by a tetrazolium dye assay using cell proliferation reagent WST-I (Roche Applied Science, Indianapolis, IN).
  • CTLL-2 5000 cells/well; RENCA: 1500/well; B16-F10: 1000/well; MV4;11 : 5000/well
  • CTLL-2 is an IL-2 dependent cell line, and hence for these cells, the cytoxicity assay was conducted in media containing 5 nM rIL-2.
  • % specific lysis 100 x ((mean experimental - mean spontaneous release)/(mean maximal release - mean spontaneous release)). The spontaneous release was determined from wells containing labeled target cells and no effector cells, and maximal release was determined from wells containing labeled target cells in 1% Triton X-100.
  • CTLL-2 Activates JAK/STAT and MAPK Signaling in CTLL-2 Cells in vitro
  • CTLL-2 cells IxIO 7 cells
  • rIL-2 Activates JAK/STAT and MAPK Signaling in CTLL-2 Cells in vitro
  • CTLL-2 cells IxIO 7 cells
  • key signaling pathways MAPK, STAT5, AKT were evaluated using western blot analyses.
  • IL-2R ⁇ KD 10 "11 M
  • IL-2 ⁇ KD 10 "9 M
  • cells were exposed to free anti-IL-2 ⁇ antibody (>1000-fold excess; 10 nM) incubated for 1 hour prior to addition of rIL-2.
  • serum starved CTLL-2 cells were activated with 10 nM rIL-2, and IL-2 signaling was examined at various times from 10 minutes up to 48 hours.
  • IL-2R downstream phosphorylation of ERK1/2, STAT5 and AKT was evaluated in rIL-2-treated cells using Western blot analysis.
  • the relative levels of pERK, pSTAT5 or pAKT were compared to the total protein levels for ERK, STAT5 or AKT, respectively.
  • pERKl/2 Activation of pERKl/2 was observed following exposure of CTLL-2 cells with rIL-2.
  • Phospho ERK was slightly activated at 1 pM at 2 hours however, maximal activation was seen at > 100 pM concentrations.
  • the JAK/STAT5 pathway was maximally activated at 1 pM concentration; and increasing concentrations of rlL- 2 did not change levels of pSTAT5 (up to 100 nM).
  • the basal pAKT pathway in CTLL-2 appeared to be activated in T cells under serum starved conditions. Furthermore pAKT levels remained largely unchanged at the rIL-2 concentrations tested (1 pM to 100 nM; using a pAKT antibody to phosphorylation site 483).
  • a blocking antibody to IL-2R ⁇ was used to confirm that the IL-2 signal pathways are mediated specifically by the binding to the IL-2R.
  • the pSTAT5 levels in CTLL-2 cells were analyzed by Western blot. CTLL cells were serum starved and treated with excess free anti-IL-2R ⁇ antibody (10 nM, > 1000-fold) for 1 hour prior to treatment with rIL-2 (0.1 pM to 10 nM). In the absence of the blocking IL-2R ⁇ antibody, pSTAT5 was activated at 1 pM following rIL-2 treatment, however, in the presence of IL-2R ⁇ inhibition, pSTAT5 signaling was abrogated (> 95% inhibition), confirming the requirement of the IL-2R ⁇ in STAT5 signaling.
  • rIL-2 Activation of pSTAT5 is Rapid and Sustained in CTLL-2 Cells To evaluate the time of onset and duration of IL-2R signaling responses in
  • CTLL-2 cells serum starved cells were treated in vitro with rIL-2 (10 nM) and effects on phosphorylation of ERKl /2, STAT5 and AKT was evaluated using Western blot analyses. Phosphorylation of STAT5 was activated in minutes ( ⁇ 10 minutes) after addition of rIL-2 to CTLL-2 cells and the duration of pSTAT5 response was maintained up to 48 hours. Activation of the MAPK pathway (pERK) pathway by rIL-2 appeared to be slightly delayed and was activated by 1 hour. The intensity of pERK was lower than the maximal response obtained by stimulation of serum starved CTLL-2 cells with PMA (50 ng/ml) + ionomycin (0.4 ⁇ g/ml) for 15 minutes.
  • PMA 50 ng/ml
  • ionomycin 0.4 ⁇ g/ml
  • pERK levels were sustained up to 24 hours and returned to background levels by 48 hours. No discernable effects were observed on the pAKT, confirming that signaling via PI-3K/AKT pathway was continually active in CTLL-2 cells.
  • BAY-43-9006 is a potent inhibitor of Raf-1, which also inhibits both wild-type and mutant BRAF.
  • BAY-43-9006 inhibits multiple kinases particularly VEGF2, 3; PDGFR ⁇ ; FLT3 and cKIT (Wilhelm, SM et al. Cancer Res 2004; and Chiron's kinase profiling data), and inhibits to some extent Lck and Fyn, two kinases that are involved in T cell functional responses.
  • Lck and Fyn two kinases that are involved in T cell functional responses.
  • BAY 43-9006 Since the concentration of BAY 43-9006 required to inhibit the MAPK pathway in different cell types is quite variable, the effects of various concentrations of BAY 43-9006 (ranging from 0 to 20 ⁇ M) were evaluated on serum starved CTLL- 2 cells for 2 hours with or without PMA + Ionomycin stimulation. At the end of the incubation period, treated CTLL-2 cells were then lysed and the protein lysates were subjected to Western blot analyses for determination of levels of pERK. BAY 43- 9006 substantially inhibited pERK at concentrations > 1 ⁇ M in murine CTLL-2 cells and the human Jurkat T cell line. The effects of BAY 43-9006 treatment on CTLL-2 cells did not alter pSTAT5 and pAKT levels in cells indicating that the JAK/STAT5 pathway and PI-3K/AKT in T cells were largely unaffected.
  • rIL-2 and BAY 43-9006 were studied.
  • the IL-2 mediated T cell signaling effects with concomitant or sequential regimens of the two drugs were investigated in CTLL-2 cells.
  • Serum starved CTLL-2 cells were treated with BAY 43-9006 (3 ⁇ M) for 2 hours and then treated with either vehicle, rIL-2 (10 nM) or PMA+Ionomycin.
  • rIL-2 (10 nM, 2 hours) treatment followed by BAY 43-9006 (3 ⁇ M, 2 hours) was also investigated.
  • the effects of BAY 43-9006 treatment on murine tumor cell lines- B16-F10 melanoma, CT26 colon and the RCC RENCA model was determined.
  • the murine cell lines were selected based on their responsiveness to in vivo rIL-2 therapy in immunocompetent models (T-/NK-/monocyte-/macrophage-competent mice), where the effects of combined rIL-2 and BAY 43-9006 therapy could be investigated.
  • Serum starved B16-F10 melanoma and RENCA cells were exposed to a range of concentrations of BAY 43-9006 from 0 to 20 ⁇ M.
  • Phospho-ERK levels in cell lysates following drug exposure were determined by Western blot analyses.
  • BAY 43-9006 inhibited pERK levels in both cell lines tested (B16-F10 and RENCA) at very high concentrations of > 5 ⁇ M, with almost complete abolition of pERK seen at 20 ⁇ M.
  • proliferation assays were conducted by pre- incubating CTLL-2 cells (5000 cells/well) at concentrations of BAY 43-9006 (3 ⁇ M, 2 hours) that inhibit pERK. After a 2 hour incubation of cells with BAY 43-9006 (3 ⁇ M), cells were plated in 96-well plates and exposed to various concentrations of rlL- 2 (0 - 100 nM) for 72 hours. Untreated cells were given sham treatments prior to incubation with rIL-2 (at the same concentrations).
  • MV4;11 human FLT3 ITD AML cell line
  • BAY 43-9006 (0 to 50 ⁇ M)
  • B16-F10 cells were treated with vincristine (0 to 1 ⁇ M) to confirm cytotoxicity of agents and validity of assays.
  • the inhibitory concentrations of the drugs were expressed as an EC 5O value, which was defined as the concentration needed for a 50% reduction in proliferative response measured as absorbance of drug- treated cells vs. untreated/vehicle controls.
  • CTLL-2 5000 cells/well; RENCA: 1500/well; B16-F10: 1000/well; CT26: 1000 cells/well; MV4;11 : 5000/well
  • WST-I tetrazolium dye
  • a EC50 concentration needed for a 50% reduction in proliferative response measured as absorbance of drug-treated cells vs. untreated/vehicle controls.
  • b MV4;11 is a human acute myelogenous leukemia (AML) cell line that expresses a FLT3 Internal Tandem Duplication (ITD).
  • d Cell viability was assessed using the Promega Cell-Titer GloTM assay that measured ATP content of cells
  • BAY 43-9006 or SUl 1248 were needed to inhibit proliferation of CTLL-2 cells as well as various cells lines (B 16- FlO, RENCA, CT26), compared to the antimitotic agent vincristine (defined by the relative EC50's; see Table 9).
  • the cytotoxicity of BAY 43-9006 effects on RENCA cells was also examined using the BrdU assay to evaluate the effect on DNA synthesis (vs. antiproliferative activity using the mitochondrial tetrazolium dye assay).
  • the EC 5 O for BAY 43-9006 ( ⁇ 5 ⁇ M) on RENCA cells obtained using the BrdU method was similar to that observed with the WST-I assay.
  • a single oral dose of 30 mg/kg BAY 43-9006 in mice achieved a C max of about 5500 to 8000 ng/ml (-10 ⁇ M) at a t max of 2 hours.
  • the plasma elimination rate of BAY 43-9006 was fairly slow and the consequent half-life was about 4 hours.
  • the PK profile of rIL-2 following subcutaneous administration of 6 mg/kg demonstrated a C max of about 550 - 850 ng/ml at a t max of about 30 minutes.
  • the rlL- 2 ti /2 in mice was approximately 1 hour, with exposures of >50 ng/ml achieved for 4 hours.
  • rIL-2 and BAY 43-9006 treatment decreases immune effector function ex vivo Since inhibition of one or multiple T lymphocytic kinases (Lck, Fyn, Syk, Btk, Src, Tck2, MAPK, JAKs) may abrogate T cell expansion and immune effector function, the effects of rIL-2 and BAY 43-9006 on T cell proliferative and functional responses in vivo was investigated.
  • T lymphocytic kinases Lck, Fyn, Syk, Btk, Src, Tck2, MAPK, JAKs
  • BALB/c mice bearing RENCA tumors were treated with rIL-2 (lmg/kg/day, s.c. days 6-10), BAY 43-9006 (30 mg/kg/day, p.o., days 6-10) or combinations of rIL-2 and BAY 43-9006 administered either concomitantly or sequentially (rIL-2, lmg/kg/day, s.c, days 1-5 + BAY 43-9006, 30 mg/kg/day, p.o., days 6-10; or BAY 43-9006, 30 mg/kg/day, p.o., days 1-5 + rIL-2, lmg/kg/day, s.c, days 6-10).
  • Isolated splenocytes from treated mice were then subjected to ex vivo killing assays against Yac-1 target cells at various effector: target (E:T) ratios, and percent specific lysis of 51 Cr-labeled Yac-1 cells were determined.
  • Mice treated with rIL-2 (1 mg/kg) significantly increased splenocyte-mediated killing of Yac-1 targets compared to vehicle treatment (23% with rIL-2 vs. 0% with vehicle treatment).
  • T cells 1575 cells/ ⁇ l with vehicle
  • T cells 1550 cells/ ⁇ l vs. 664 cells/ ⁇ l with vehicle
  • a significantly increased ratio of CD4:CD8 cells was observed with rIL-2 therapy compared to vehicle treatment (5:3; rIL-2: Vehicle, i.e., 1.7-fold), as indicative of rIL-2 mechanism of action in expanding T cell numbers.
  • the relative numbers of non-T and monocytic cells (CD45+CD3-) following rIL-2 treatment were similar to vehicle treatment (837 cells/ ⁇ l vs. 911 cells/ ⁇ l with vehicle).
  • single agent BAY 43-9006 or BAY 43-9006 combined with rlL-
  • lymphocytes and monocytes had little impact or increased the absolute numbers of lymphocytes and monocytes (range of 2417-3577 cells/ ⁇ l vs. 2387 cells/ ⁇ l with vehicle), and also increased non- T cells and monocyte populations (1241-1563 cells/ ⁇ l vs. 837 cells/ ⁇ l with vehicle).
  • the effect of BAY 43-9006 therapy on total numbers of T cells was similar to vehicle treatment (1827 cells/ ⁇ l vs. 1550 cells/ ⁇ l with vehicle). Slightly increased total T cell numbers (including both CD4+ and CD8+ populations) observed with the sequential regimen of rIL-2 and BAY 43-9006 when commenced with rIL-2 (2081 T cells/ ⁇ l vs. 1550 cells/ ⁇ l with vehicle).
  • BAY 43-9006 or SUl 1248 therapy was determined.
  • tumor infiltrating T cells were detected with a mouse anti-CD3 antibody.
  • Ki67 staining With rIL-2 treatment, increased numbers of T cells were seen infiltrating both RENCA tumors (and B16-F10 tumors) compared to vehicle treatment. Generally, fewer T cells were detected in the Bayer 43-9006-treated group in the RENCA model.
  • n 10 BALB/c mice/group in each study.
  • CT26 tumor-bearing mice were treated when tumors were established to a mean size of -225 mm 3 .
  • b T/Cobs erv e d (O) % T/C
  • Vehicle 712 1.00 N/A N/A N/A
  • n 10 BALB/c mice/group in each study.
  • RENCA model tumored mice were treated tumors were established to a mean size of -50-70 mm 3 .
  • Concomitant regimens are presented as RENCA tumor model exhibits severe cachexia (animal wasting) making sequential administration of agents not possible, and not evaluable.
  • b T/Cobserved (O) % T/C

Abstract

L'invention concerne une polythérapie faisant appel à des compositions IL-2 et à des agents anti-angiogéniques pour traiter le cancer. En outre, l'invention concerne des méthodes de réduction de toxicités et d'augmentation d'efficacité associées à l'administration de compositions de IL-2 ou de compositions anti-angiogéniques.
PCT/US2006/005720 2005-02-18 2006-02-17 Agents anti-angiogeniques comportant de l'aldesleukine WO2006089150A2 (fr)

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AU2006214138A AU2006214138A1 (en) 2005-02-18 2006-02-17 Antiangiogenic agents with aldesleukin
CA002598448A CA2598448A1 (fr) 2005-02-18 2006-02-17 Agents anti-angiogeniques comportant de l'aldesleukine
MX2007010037A MX2007010037A (es) 2005-02-18 2006-02-17 Agentes antiangiogenicos con aldesleucina.
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Cited By (21)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP2011528676A (ja) * 2008-07-24 2011-11-24 バイエル ファーマ アクチエンゲゼルシャフト 免疫調節物質としてのスルホン置換キナゾリン誘導体、これらの調製及び薬剤としての使用
WO2012079075A1 (fr) 2010-12-10 2012-06-14 Concert Pharmaceuticals, Inc. Dérivés de phtalimide deutérés
CN101497607B (zh) * 2008-01-29 2012-11-28 上海百灵医药科技有限公司 舒尼替尼的合成方法
WO2013130849A1 (fr) 2012-02-29 2013-09-06 Concert Pharmaceuticals, Inc. Dérivés de phthalimide dioxopipéridinyle substitués
WO2013159026A1 (fr) 2012-04-20 2013-10-24 Concert Pharmaceuticals, Inc. Rigosertib deutéré
US8575184B2 (en) 2009-09-03 2013-11-05 Bristol-Myers Squibb Company Quinazolines as potassium ion channel inhibitors
WO2014066243A1 (fr) 2012-10-22 2014-05-01 Concert Pharmaceuticals, Inc. Formes solides de {s-3-(4-amino-1-oxo-isoindolin-2yl)(pipéridine-3,4,4,5,5-d5)-2,6-dione}
WO2014110322A2 (fr) 2013-01-11 2014-07-17 Concert Pharmaceuticals, Inc. Dérivés substitués de dioxopipéridinyl phtalimide
WO2014170910A1 (fr) 2013-04-04 2014-10-23 Natco Pharma Limited Procédé de préparation du lapatinib
US9045453B2 (en) 2008-11-14 2015-06-02 Concert Pharmaceuticals, Inc. Substituted dioxopiperidinyl phthalimide derivatives
US9145411B2 (en) 2012-08-02 2015-09-29 Asana Biosciences, Llc Substituted amino-pyrimidine derivatives
US9353123B2 (en) 2013-02-20 2016-05-31 Kala Pharmaceuticals, Inc. Therapeutic compounds and uses thereof
US9353122B2 (en) 2013-02-15 2016-05-31 Kala Pharmaceuticals, Inc. Therapeutic compounds and uses thereof
US9688688B2 (en) 2013-02-20 2017-06-27 Kala Pharmaceuticals, Inc. Crystalline forms of 4-((4-((4-fluoro-2-methyl-1H-indol-5-yl)oxy)-6-methoxyquinazolin-7-yl)oxy)-1-(2-oxa-7-azaspiro[3.5]nonan-7-yl)butan-1-one and uses thereof
US9790232B2 (en) 2013-11-01 2017-10-17 Kala Pharmaceuticals, Inc. Crystalline forms of therapeutic compounds and uses thereof
US9890173B2 (en) 2013-11-01 2018-02-13 Kala Pharmaceuticals, Inc. Crystalline forms of therapeutic compounds and uses thereof
US10253036B2 (en) 2016-09-08 2019-04-09 Kala Pharmaceuticals, Inc. Crystalline forms of therapeutic compounds and uses thereof
US10336767B2 (en) 2016-09-08 2019-07-02 Kala Pharmaceuticals, Inc. Crystalline forms of therapeutic compounds and uses thereof
US10766907B2 (en) 2016-09-08 2020-09-08 Kala Pharmaceuticals, Inc. Crystalline forms of therapeutic compounds and uses thereof
US10844039B2 (en) 2018-11-13 2020-11-24 Biotheryx, Inc. Substituted isoindolinones
CN114920704A (zh) * 2019-07-26 2022-08-19 暨南大学 一种苯基哌嗪喹唑啉类化合物或其药学上可接受的盐、制法与用途

Families Citing this family (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP2373165B1 (fr) * 2008-11-14 2013-10-30 Concert Pharmaceuticals, Inc. Dérivés de dioxopipéridinylphtalimide substitués
KR101478925B1 (ko) * 2013-12-31 2015-01-08 아주대학교산학협력단 프로테아좀 저해제와 로페라마이드를 유효성분으로 함유하는 암질환 예방 또는 치료용 약학조성물
CN108069913B (zh) * 2016-11-18 2022-03-01 陕西师范大学 双(吗啉基烷氧基)喹唑啉衍生物及其在抗肿瘤方面的用途
CN111499583B (zh) * 2020-05-22 2022-02-15 沈阳工业大学 喹唑啉衍生物及其作为抗肿瘤药物的应用
CN112538071A (zh) * 2020-11-30 2021-03-23 新乡医学院第一附属医院 用作raf-1抑制剂的化合物、制备方法和用途

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
SONI SANDEEP ET AL: "Treatment of pediatric ocular melanoma with high-dose interleukin-2 and thalidomide." JOURNAL OF PEDIATRIC HEMATOLOGY/ONCOLOGY : OFFICIAL JOURNAL OF THE AMERICAN SOCIETY OF PEDIATRIC HEMATOLOGY/ONCOLOGY. 2002 AUG-SEP, vol. 24, no. 6, August 2002 (2002-08), pages 488-491, XP009067555 ISSN: 1077-4114 *
STERNBERG C N: "Metastatic renal cell cancer treatments." DRUGS OF TODAY, vol. 39, no. Suppl. C, November 2003 (2003-11), pages 39-59, XP009067549 ISSN: 0025-7656 *

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US9643950B2 (en) 2012-10-22 2017-05-09 Concert Pharmaceuticals, Inc. Solid forms of {s-3-(4-amino-1-oxo-isoindolin-2-yl)(piperidine-3,4,4,5,5-d5)-2,6-dione}
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US9353122B2 (en) 2013-02-15 2016-05-31 Kala Pharmaceuticals, Inc. Therapeutic compounds and uses thereof
US10398703B2 (en) 2013-02-15 2019-09-03 Kala Pharmaceuticals, Inc. Therapeutic compounds and uses thereof
US9688688B2 (en) 2013-02-20 2017-06-27 Kala Pharmaceuticals, Inc. Crystalline forms of 4-((4-((4-fluoro-2-methyl-1H-indol-5-yl)oxy)-6-methoxyquinazolin-7-yl)oxy)-1-(2-oxa-7-azaspiro[3.5]nonan-7-yl)butan-1-one and uses thereof
US9833453B2 (en) 2013-02-20 2017-12-05 Kala Pharmaceuticals, Inc. Therapeutic compounds and uses thereof
US9861634B2 (en) 2013-02-20 2018-01-09 Kala Pharmaceuticals, Inc. Therapeutic compounds and uses thereof
US9353123B2 (en) 2013-02-20 2016-05-31 Kala Pharmaceuticals, Inc. Therapeutic compounds and uses thereof
US11369611B2 (en) 2013-02-20 2022-06-28 Kala Pharmaceuticals, Inc. Therapeutic compounds and uses thereof
US10285991B2 (en) 2013-02-20 2019-05-14 Kala Pharmaceuticals, Inc. Therapeutic compounds and uses thereof
US10758539B2 (en) 2013-02-20 2020-09-01 Kala Pharmaceuticals, Inc. Therapeutic compounds and uses thereof
WO2014170910A1 (fr) 2013-04-04 2014-10-23 Natco Pharma Limited Procédé de préparation du lapatinib
US9890173B2 (en) 2013-11-01 2018-02-13 Kala Pharmaceuticals, Inc. Crystalline forms of therapeutic compounds and uses thereof
US10975090B2 (en) 2013-11-01 2021-04-13 Kala Pharmaceuticals, Inc. Crystalline forms of therapeutic compounds and uses thereof
US10618906B2 (en) 2013-11-01 2020-04-14 Kala Pharmaceuticals, Inc. Crystalline forms of therapeutic compounds and uses thereof
US11713323B2 (en) 2013-11-01 2023-08-01 Kala Pharmaceuticals, Inc. Crystalline forms of therapeutic compounds and uses thereof
US9790232B2 (en) 2013-11-01 2017-10-17 Kala Pharmaceuticals, Inc. Crystalline forms of therapeutic compounds and uses thereof
US10160765B2 (en) 2013-11-01 2018-12-25 Kala Pharmaceuticals, Inc. Crystalline forms of therapeutic compounds and uses thereof
US10766907B2 (en) 2016-09-08 2020-09-08 Kala Pharmaceuticals, Inc. Crystalline forms of therapeutic compounds and uses thereof
US10253036B2 (en) 2016-09-08 2019-04-09 Kala Pharmaceuticals, Inc. Crystalline forms of therapeutic compounds and uses thereof
US11021487B2 (en) 2016-09-08 2021-06-01 Kala Pharmaceuticals, Inc. Crystalline forms of therapeutic compounds and uses thereof
US11104685B2 (en) 2016-09-08 2021-08-31 Kala Pharmaceuticals, Inc. Crystalline forms of therapeutic compounds and uses thereof
US10626121B2 (en) 2016-09-08 2020-04-21 Kala Pharmaceuticals, Inc. Crystalline forms of therapeutic compounds and uses thereof
US10336767B2 (en) 2016-09-08 2019-07-02 Kala Pharmaceuticals, Inc. Crystalline forms of therapeutic compounds and uses thereof
US10844039B2 (en) 2018-11-13 2020-11-24 Biotheryx, Inc. Substituted isoindolinones
US11352338B2 (en) 2018-11-13 2022-06-07 Biotheryx, Inc. Substituted isoindolinones
CN114920704A (zh) * 2019-07-26 2022-08-19 暨南大学 一种苯基哌嗪喹唑啉类化合物或其药学上可接受的盐、制法与用途
CN114920704B (zh) * 2019-07-26 2023-11-03 暨南大学 一种苯基哌嗪喹唑啉类化合物或其药学上可接受的盐、制法与用途

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BRPI0608880A2 (pt) 2010-02-02
AU2006214138A1 (en) 2006-08-24
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JP2008530239A (ja) 2008-08-07
EP1853302A2 (fr) 2007-11-14
KR20070108909A (ko) 2007-11-13
WO2006089150A3 (fr) 2006-11-02
MX2007010037A (es) 2008-02-15
CN101146549A (zh) 2008-03-19

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