| Antagonists of neuropilin receptor function and use thereof -> Monitor Keywords |
|
|
 
Antagonists of neuropilin receptor function and use thereofRelated Patent Categories: Chemistry: Natural Resins Or Derivatives; Peptides Or Proteins; Lignins Or Reaction Products Thereof, Proteins, I.e., More Than 100 Amino Acid Residues, Blood Proteins Or Globulins, E.g., Proteoglycans, Platelet Factor 4, Thyroglobulin, Thyroxine, Etc., Globulins, Immunoglobulin, Antibody, Or Fragment Thereof, Other Than Immunoglobulin Antibody, Or Fragment Thereof That Is Conjugated Or AbsorbedAntagonists of neuropilin receptor function and use thereof description/claimsThe Patent Description & Claims data below is from USPTO Patent Application 20080076906, Antagonists of neuropilin receptor function and use thereof. Brief Patent Description - Full Patent Description - Patent Application Claims
FIELD OF THE INVENTION [0002] The present invention relates to antagonists of neuropilin receptor function and use thereof in the treatment of cancer, particularly metastatic cancer, and angiogenic diseases. BACKGROUND OF THE INVENTION [0003] Cancer, its development and treatment is a major health concern. The standard treatments available are few and directed to specific types of cancer, and provide no absolute guarantee of success. Most treatments rely on an approach that involves killing off rapidly growing cells in the hope that rapidly growing cancerous cells will succumb, either to the treatment, or at least be sufficiently reduced in numbers to allow the body's system to eliminate the remainder. However most, of these treatments are non-specific to cancer cells and adversely effect non-malignant cells. Many cancers although having some phenotype relationship are quite diverse. Yet, what treatment works most effectively for one cancer may not be the best means for treating another cancer. Consequently, an appreciation of the severity of the condition must be made before beginning many therapies. In order to most effective, these treatments require not only an early detection of the malignancy, but an appreciation of the severity of the malignancy. Currently, it can be difficult to distinguish cells at a molecular level as it relates to effect on treatment. Thus, methods of being able to screen malignant cells and better understand their disease state are desirable. [0004] While different forms of cancer have different properties, one factor which many cancers share is that they can metastasize. Until such time as metastasis occurs, a tumor, although it may be malignant, is confined to one area of the body. This may cause discomfort and/or pain, or even lead to more serious problems including death, but if it can be located, it may be surgically removed and, if done with adequate care, be treatable. However, once metastasis sets in, cancerous cells have invaded the body and while surgical resection may remove the parent tumor, this does not address other tumors. Only chemotherapy, or some particular form of targeting therapy, then stands any chance of success. [0005] The process of tumor metastasis is a multistage event involving local invasion and destruction of intercellular matrix, intravasation into blood vessels, lymphatics or other channels of transport, survival in the circulation, extravasation out of the vessels in the secondary site and growth in the new location (Fidler, et al., Adv. Cancer Res. 28, 149-250 (1978), Liotta, et al., Cancer Treatment Res. 40, 223-238 (1988), Nicolson, Biochim. Biophy. Acta 948, 175-224 (1988) and Zetter, N. Eng. J. Med. 322, 605-612 (1990)). Success in establishing metastatic deposits requires tumor cells to be able to accomplish these steps sequentially. Common to many steps of the metastatic process is a requirement for motility. The enhanced movement of malignant tumor cells is a major contributor to the progression of the disease toward metastasis. Increased cell motility has been associated with enhanced metastatic potential in animal as well as human tumors (Hosaka, et al., Gann 69, 273-276 (1978) and Haemmerlin, et al., Int. J. Cancer 27, 603-610 (1981)). [0006] Identifying factors that are associated with onset of tumor metastasis is extremely important. In addition, to using such factors for diagnosis and prognosis, those factors are an important site for identifying new compounds that can be used for treatment and as a target for treatment identifying new modes of treatment such as inhibition of metastasis is highly desirable. [0007] Tumor angiogenesis is essential for both primary tumor expansion and metastatic tumor spread, and angiogenesis itself requires ECM degradation (Blood et al., Biochim. Biophys. Acta 1032:89-118 (1990)). Thus, malignancy is a systemic disease in which interactions between the neoplastic cells and their environment play a crucial role during evolution of the pathological process (Fidler, I. J., Cancer Metastasis Rev. 5:29-49 (1986)). [0008] There is mounting evidence that VEGF may be a major regulator of angiogenesis (reviewed in Ferrara, et al., Endocr. Rev., 13, 18-32 (1992); Klagsbrun, et al., Curr. Biol., 3, 699-702 (1993); Ferrara, et al., Biochem. Biophjs. Res. Commun., 161, 851-858 (1989)). VEGF was initially purified from the conditioned media of folliculostellate cells (Ferrara, et al., Biochem. Biophjs. Res. Commun., 161, 851-858 (1989)) and from a variety of tumor cell lines (Myoken, et al., Proc. Natl. Acad. Sci. USA, 88:5819-5823 (1991); Plouet, et al., EMBO. J., 8:3801-3806 (1991)). VEGF was found to be identical to vascular permeability factor, a regulator of blood vessel permeability that was purified from the conditioned medium of U937 cells at the same time (Keck, et al., Science, 246:1309-1312 (1989)). VEGF is a specific mitogen for endothelial cells (EC) in vitro and a potent angiogenic factor in vivo. The expression of VEGF is up-regulated in tissue undergoing vascularization during embryogenesis and the female reproductive cycle (Brier, et al., Development, 114:521-532 (1992); Shweiki, et al., J. Clin. Invest., 91:2235-2243 (1993)). High levels of VEGF are expressed in various types of tumors, but not in normal tissue, in response to tumor-induced hypoxia (Shweiki, et al., Nature 359:843-846 (1992); Dvorak et al., J. Exp. Med., 174:1275-1278 (1991); Plate, et al., Cancer Res., 53:5822-5827; Ikea, et al., J. Biol. Chem., 270:19761-19766 (1986)). Treatment of tumors with monoclonal antibodies directed against VEGF resulted in a dramatic reduction in tumor mass due to the suppression of tumor angiogeneis (Kim, et al., Nature, 382:841-844 (1993)). VEGF appears to play a principle role in many pathological states and processes related to neovascularization. Regulation of VEGF expression in affected tissues could therefore be key in treatment or prevention of VEGF induced neovascularization/angiogenesis. [0009] VEGF exists in a number of different isoforms that are produced by alternative splicing from a single gene containing eight exons (Ferrara, et al., Endocr. Rev., 13:18-32 (1992); Tischer, et al., J. Biol. Chem., 806:11947-11954 (1991); Ferrara, et al., Trends Cardio Med., 3:244-250 (1993); Polterak, et al., J. Biol. Chem., 272:7151-7158 (1997)). Human VEGF isoforms consists of monomers of 121, 145, 165, 189, and 206 amino acids, each capable of making an active homodimer (Polterak et al., J. Biol. Chem., 272:7151-7158 (1997); Houck, et al., Mol. Endocrinol., 8:1806-1814 (1991)). The VEGF.sub.121 and VEGF.sub.165 isoforms are the most abundant. VEGF.sub.121 is the only VEGF isoforms that does not bind to heparin and is totally secreted into the culture medium. VEGF.sub.165 is functionally different than VEGF.sub.121 in that it binds to heparin and cell surface heparin sulfate proteoglycans (HSPGs) and is only partially released into the culture medium (Houck, et al., J. Biol. Chem., 247:28031-28037 (1992); Park, et al., Mol. Biol. Chem., 4:1317-1326 (1993)). The remaining isoforms are entirely associated with cell surface and extracellular matrix HSPGs (Houck, et al., J. Biol. Chem., 247:28031-28037 (1992); Park, et al., Mol. Biol. Chem., 4:1317-1326 (1993)). [0010] VEGF receptor tyrosine kinases, KDR/Flk-1 and/or Flt-1, are mostly expressed by EC (Terman, et al., Biochem. Biophys. Res. Commun., 187:1579-1586 (1992); Shibuya, et al., Oncogene, 5:519-524 (1990); De Vries, et al., Science, 265:989-991 (1992); Gitay-Goran, et al., J. Biol. Chem., 287:6003-6096 (1992); Jakeman, et al., J. Clin. Invest., 89:244-253 (1992)). It appears that VEGF activities such as mitogenicity, chemotaxis, and induction of morphological changes are mediated by KDR/Flk-1 but not Flt-1, even though both receptors undergo phosphorylation upon binding of VEGF (Millauer, et al., Cell, 72:835-846 (1993); Waltenberger, et al., J. Biol. Chem., 269:26988-26995 (1994); Seetharam, et al., Oncogene, 10:135-147 (1995); Yoshida, et al., Growth Factors, 7:131-138 (1996)). Recently, Soker et al., identified a new VEGF receptor which is expressed on EC and various tumor-derived cell lines such as breast cancer-derived MDA-MB-231 (231) cells (Soker, et al., J. Biol. Chem., 271:5761-5767 (1996)). This receptor requires the VEGF isoform to contain the portion encoded by exon 7. For example, although both VEGF.sub.121 and VEGF.sub.165R bind to KDR/Flk-1 and Flt-1, only VEGF.sub.165 binds to the new receptor. Thus, this is an isoform-specific receptor and has been named the VEGF.sub.165 receptor (VEGF.sub.165R). It will also bind the 189 and 206 isoforms. VEGF.sub.165R has a molecular mass of approximately 130 kDa, and it binds VEGF.sub.165 with a Kd of about 2.times.10.sup.-10M, compared with approximately 5.times.10.sup.-12M for KDR/Flk-1. In structure-function analysis, it was shown directly that VEGF.sub.165 binds to VEGF.sub.165R via its exon 7-encoded domain which is absent in VEGF.sub.121 (Soker, et al., J. Biol. Chem., 271:5761-5767 (1996)). However, the function of the receptor was unclear. [0011] Identifying the alterations in gene expression which are associated with malignant tumors, including those involved in tumor progression and angiogenesis, is clearly a prerequisite not only for a full understanding of cancer, but also to develop new rational therapies against cancer. [0012] A further problem arises, in that the genes characteristic of cancerous cells are very often host genes being abnormally expressed. It is quite often the case that a particular protein marker for a given cancer while expressed in high levels in connection with that cancer is also expressed elsewhere throughout the body, albeit at reduced levels. [0013] The current treatment of angiogenic diseases is inadequate. Agents which prevent continued angiogenesis, e.g, drugs (TNP-470), monoclonal antibodies, antisense nucleic acids and proteins (angiostatin and endostatin) are currently being tested. See, Battegay, J. Mol. Med., 73, 333-346 (1995); Hanahan et al., Cell, 86, 353-364 (1996); Folkman, N. Engl. J. Med., 333, 1757-1763 (1995). Although preliminary results with the antiangiogenic proteins are promising, there is still a need for identifying genes encoding ligands and receptors involved in angiogenesis for the development of new antiangiogenic therapies. SUMMARY OF THE INVENTION [0014] We have isolated a cDNA encoding the VEGF.sub.165R gene (SEQ ID NO: 1) and have deduced the amino acid sequence of the receptor (SEQ ID NO:2) We have discovered that this novel VEGF receptor is structurally unrelated to Flt-1 or KDR/Flk-1 and is expressed not only by endothelial cells but by non-endothelial cells, including surprisingly tumor cells. [0015] In ascertaining the function of the VEGF.sub.165R we have further discovered that this receptor has been identified as a cell surface mediator of neuronal cell guidance and called neuropilin-1. Kolodkin et al., Cell 90:753-762 (1997). We refer to the receptor as VEGF.sub.165R/NP-1 or NP-1. [0016] In addition to the expression cloning of VEGF.sub.165R/NP-1 cDNA we isolated another human cDNA clone whose predicted amino acid sequence was 47% homologous to that of VEGF.sub.165R/NP-1 and over 90% homologous to rat neuropilin-2 (NP-2) which was recently cloned (Kolodkin, et al., Cell 90, 753-762 (1997)). [0017] Our results indicate that VEGF.sub.165R/NP-1 and NP-2 are expressed by both endothelial and tumor cells. (FIG. 19) We have shown that endothelial cells expressing both KDR and VEGF.sub.165R/NP-1 respond with increased chemotaxis towards VEGF.sub.165, not VEGF.sub.121, when compared to endothelial cells expressing KDR alone. While not wishing to be bound by theory, we believe that VEGF.sub.165R/NP-1 functions in endothelial cells to mediate cell motility as a co-receptor for KDR. [0018] We have also shown in the Boyden chamber motility assay that VEGF.sub.165 stimulates 231 breast carcinoma cell motility in a dose-response manner (FIG. 15A). VEGF.sub.121 had no effect motility of these cells (FIG. 15B). Since tumor cells such as, 231 cells, do not express the VEGF receptors, KDR or Flt-1, while not wishing to be bound by theory, we believe that tumor cells are directly responsive to VEGF.sub.165 via VEGF.sub.165R/NP-1. [0019] We have also analyzed two variants of Dunning rat prostate carcinoma cells, AT2.1 cells, which are of low motility and low metastatic potential, and AT3.1 cells, which are highly motile, and metastatic. Cross-linking and Northern blot analysis show that AT3.1 cells express abundant VEGF.sub.165R/NP-1, capable of binding VEGF.sub.165, while AT2.1 cells don't express VEGF.sub.165R/NP-1 (FIG. 18). Immunostaining of tumor sections confirmed the expression of VEGF.sub.165R/NP-1 in AT3.1, but not AT2.1 tumors (FIG. 17). Additionally, immunostaining showed that in subcutaneous AT3.1 and PC3 tumors, the tumor cells expressing VEGF.sub.165R/NP-1 were found preferentially at the invading front of the tumor/dermis boundary (FIG. 17). Furthermore, stable clones of AT2.1 cells overexpressing VEGF.sub.165R/NP-1 had enhanced motility in the Boyden chamber assay. These results indicate that neuropilin expression on tumor cells is associated with the motile, metastatic phenotype and angiogenesis, and thus is an important target for antiangiogenic and anticancer therapy. [0020] The present invention relates to antagonists of neuropilin (NP) receptor function that can be use to inhibit metastasis and angiogenesis. Antagonists of invention can block the receptor preventing ligand binding, disrupt receptor function, or inhibit receptor occurrence. Specific antagonists include, for example, compounds that bind to NP-1 or NP-2 and antibodies that specifically binds the receptor at a region that inhibits receptor function. For example, one can add an effective amount of a compound that binds to NP-1 to disrupt receptor function and thus inhibit metastasis. [0021] We have surprisingly discovered that members of the semaphorin/collapsins family are not only inhibitors of neuronal guidance but also inhibitors of endothelial and tumor cell motility in cells that express neuropilin. Accordingly, preferred antagonists include members of the semaphorin/collapsins family or fragments thereof that bind NP and have VEGF antagonist activity as determined, for example, by the human umbilical vein endothelial cell (HUVEC) proliferation assay using VEGF.sub.165 as set forth in Soker et al., J. Biol. Chem. 272, 31582-31588 (1997). Preferably, the semaphorin/collapsin has at least a 25% reduction in HUVEC proliferation, more preferably a 50% reduction, even more preferably a 75% reduction, most preferably a 95% reduction. [0022] VEGF antagonist activity of the semaphorin/collapsin may also be determined by inhibition of binding of labeled VEGF.sub.165 to VEGF.sub.165R as disclosed in Soker et al., J. Biol. Chem. 271, 5761-5767 (1996)) or to PAE/NP cells. Preferably, the portion inhibits binding by at least 25%, more preferably 50%, most preferably 75%. Continue reading about Antagonists of neuropilin receptor function and use thereof... Full patent description for Antagonists of neuropilin receptor function and use thereof Brief Patent Description - Full Patent Description - Patent Application Claims Click on the above for other options relating to this Antagonists of neuropilin receptor function and use thereof patent application. ###  How KEYWORD MONITOR works... a FREE service from FreshPatents1. Sign up (takes 30 seconds). 2. Fill in the keywords to be monitored. 3. Each week you receive an email with patent applications related to your keywords. Start now! - Receive info on patent apps like Antagonists of neuropilin receptor function and use thereof or other areas of interest. ### Previous Patent Application: Process for producing protein in cell-free protein synthesis system and reagent kit for protein synthesis Next Patent Application: Selective capture and enrichment of proteins expressed on the cell surface Industry Class: Chemistry: natural resins or derivatives; peptides or proteins; lignins or reaction products thereof ### FreshPatents.com Support Thank you for viewing the Antagonists of neuropilin receptor function and use thereof patent info. IP-related news and info Results in 0.3226 seconds Other interesting Feshpatents.com categories: Electronics: Semiconductor , Audio , Illumination , Connectors , Crypto , 174 |
 * Protect your Inventions * US Patent Office filing 
PATENT INFO |
|
