| Monoclonal antibody therapy for pancreas cancer -> Monitor Keywords |
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Monoclonal antibody therapy for pancreas cancerRelated Patent Categories: Drug, Bio-affecting And Body Treating Compositions, Immunoglobulin, Antiserum, Antibody, Or Antibody Fragment, Except Conjugate Or Complex Of The Same With Nonimmunoglobulin Material, Monoclonal Antibody Or Fragment Thereof (i.e., Produced By Any Cloning Technology), Binds Eukaryotic Cell Or Component Thereof Or Substance Produced By Said Eukaryotic Cell, Cancer CellMonoclonal antibody therapy for pancreas cancer description/claimsThe Patent Description & Claims data below is from USPTO Patent Application 20060228363, Monoclonal antibody therapy for pancreas cancer. Brief Patent Description - Full Patent Description - Patent Application Claims
1. INTRODUCTION [0001] The present invention relates to the use of monoclonal antibody 31.1 and its equivalents and co-specific antibodies in the treatment of pancreas cancer. It is based, at least in part, on the discovery that monoclonal antibody 31.1 is reactive with malignant, but not non-malignant, pancreatic cells. The present invention further provides polynucleotide and amino acid sequences comprising the light chain variable region and heavy chain variable region of Mu-31.1 as set forth in FIG. 2 and FIG. 4, respectively. Such polynucleotide sequences may be used to recombinantly express 31.1 equivalent antibodies for use in the methods of the invention. 2. BACKGROUND OF THE INVENTION 2.1. Pancreas Cancer [0002] Pancreas cancer is the fifth leading cause of cancer death in the United States, with approximately 28,000 Americans expected to die from the disease this year Pancreas Cancer Web, The Johns Hopkins Medical Institutions, http://162.129.103.69:80/PANCREAS_INTRO). At present, the only potentially curative treatment is surgical removal of the cancer, in the context of an extensive and complex procedure which removes the head, neck and uncinate process of the pancreas as well as the majority of the duodenum (the "Whipple operation"). Without treatment, the overall 5 year survival rate is only 3 percent (Id.). [0003] Chemotherapy (often using gemcitabine (Gemzar.RTM.)) and radiation therapy are the main treatments offered to patients with unresectable tumors (Id.). An experimental immunotherapy is currently being studied in which a patient's own cells are genetically modified to express the immune stimulatory protein, granulocyte-macrophage colony stimulating factor, irradiated to prevent tumor growth, and then reintroduced into the patient, where they will hopefully stimulate an immune response (1997, Cancer Res. 57:1537-1546; Pancreas Cancer Web, The Johns Hopkins Medical Institutions, http://162.129.103.69:80/PANCREAS_MEDICAL_TX)). 2.2. Monoclonal Antibody 31.1 [0004] Antibody 31.1 represents a protein-directed monoclonal antibody derived by immunizing BALB(c) mice with a preparation of membrane obtained from pooled (human) allogeneic colon carcinoma specimens. The cells used to prepare the antigen were fragmented using a nitrogen (Parr) bomb and then subjected to ultracentrifugation. Membrane material was initially tested by electron microscopy to guarantee consistency from batch to batch, ruling out cytoplasmic and nuclear components. It was then sonicated and fractionated with sephadex G200. Discontinuous polyacrylamide gel electrophoresis was used for the initial partial purification (approximately 80%) and 30 .mu.gm tested for delayed cutaneous hypersensitivity (DHR), (3). BALB mice were immunized by intraperitoneal injection of 50 micrograms of colon carcinoma associated antigen. A second injection was given 10 days later and the mice then sacrificed to obtain spleen cells for fusion. Fusion was performed by incubating 5.times.10.sup.7 mouse spleen cells with 10.sup.7 sp2/0-AG 14 myeloma cells in 40% PEG. The antigen defined by the monoclonal antibody 31.1 has been shown to have M.W. of 72,000. Studies using immunoperoxidase have suggested that the antigen recognized by 31.1 is seen with greater frequency in the higher grade colon tumors. Specificity for the antibody is high, so that in a study of shed colonocytes at the Mayo Clinic, sensitivity and specificity were superior when compared with anti-CEA, anti-MUC1 and B72.3. [0005] Several candidate antibodies were isolated and tested from the 1st generation TAA. All proved to be protein derived and relatively specific for colon carcinoma. Antibody 31.1 corresponded to one of the two antigens that have been shown to migrate closely on gel-electrophoresis and related to the immunogenic glycoprotein inducing the DHR. The murine version is of IgG2a isotype which converts to an IgG1 isotype on chimerization. 31.1 was found to have strong localization indices. As such, this antibody was the first to be chimerized. [0006] For chimerization of monoclonal antibody 31.1, the protein coding exon of 31.1 heavy chain variable region gene was spliced to the protein coding exons of human gamma 1 chain constant region. PCR was employed. The 31.1 VH cDNA was amplified by the PCR using the degenerate backward primers synthesized based on the consensus first framework (FR1) region DNA sequences and a forward primer synthesized according to the consensus J-C junction region DNA sequences. The amplified 31.1 V.sub.H DNA was cloned into the pBluescript vector and sequenced. Chimeric 31.1 was produced by transfecting SP2/0 AG14 cells with the vector. [0007] Monoclonal antibody 31.1 and a chimeric (humanized) version of that antibody are described in U.S. Pat. No. 5,688,657 issued Nov. 18, 1997, now the subject of a reissue application, the contents of which are hereby incorporated by reference in its entirety herein. Furthermore, monoclonal antibody 31.1 has been deposited with the American Type Culture Collection ("ATCC"), having an address at 10801 University Blvd., Manassas, Va., 20110-2209 and assigned accession number ATCC PTA-2497 and the chimerized version has been deposited with the ATCC and assigned accession number 12316. 3. SUMMARY OF THE INVENTION [0008] The present invention relates to the use of binding equivalents of monoclonal antibody 31.1, including chimerized and/or humanized versions thereof, antibody fragments and competitively binding antibodies and antibody fragments, as well as co-specific antibodies, derivatives and fragments in the treatment of pancreas cancer. [0009] It is based, at least in part, on in vitro studies using both murine and chimeric versions of 31.1 which compared the ADCC activites of the 31.1 antibodies with D6-12 and 17.1a (Panorex). While the murine version of 31.1 can induce a 35% ADCC response, the chimeric version has been shown to result in 80% of tumor cells being destroyed every three hours, using a chromium release assay. This compares with a 30% rate of destruction associated with D6-12 and a 15% rate for Panorex. Using xenograft models with human colon cancer cell lines LC-174T and Colo205, chimeric 31.1 was found to cause regression of established tumor lines (well defined molecules) after inoculating two million tumor cells into the hind legs of nude mice and administering intra-peritoneal antibody at 10 days along with human effector cells. At 30 days the volume of tumor in the treated animals when compared to controls was reduced by more than 95%. Similar results may be expected when the antibody is directed toward pancreatic cancer cells, as the 31.1 antibody has been shown to bind to antigen present in pancreatic cancer cells, but not non-malignant pancreatic tissues. [0010] The present invention provides polynucleotide and amino acid sequences comprising the light chain variable region and heavy chain variable region of Mu-31.1 which may be used to express chimerized 31.1 antibodies. The nucleotide sequences of the invention include: (a) the nucleotides sequences shown in FIG. 2 or FIG. 4; (b) a nucleotide sequence that encodes the amino acid sequence shown in FIG. 2 or FIG. 4; and (c) any nucleotide sequence that (i) binds to the nucleotide sequence of FIG. 2 or FIG. 4 under stringent hybridization conditions, e.g., hybridization to filter boun DNA in 0.5 M NaBPO.sub.4, 7% sodium dodecyl sulfate (SDS), 1 mM EDTA at 65.degree., and washing in 0.1.times.SSC/0.1% SDS at 68.degree. and (ii) encodes for a light and heavy chain variable region capable of binding with the same immunospecificity as the chimeric 31.1 monoclonal antibody. [0011] The invention further provides for a new expression construct of chimerized 31.1 antibody, termed pRc/CMV 31.1 which has been depostied with the ATCC and assigned accession no. ATCC [ ]. This plasmid carries a dihydrofolate reductase ("dhfr") expression unit driven by an enhancer-deficient SV40 early promoter that allows expresion at greater than 200 mg/liter in dihyfrofolate reductase deficient Chinese hamster ovary cells. 4. BRIEF DESCRIPTION OF THE DRAWINGS [0012] FIG. 1A-F. Series of plasmids used to construct pRc/CMV 31.1 vector by inserting the Chi31.1-1 light chain and heavy chain genes into plasmid pDCM-dhfr (FIG. 1F). [0013] FIG. 2. Nucleic acid sequence (double stranded) and possible amino acid sequences (depending on reading frame) of the 31.1 light chain variable region, showing restriction enzyme cleavage sites. [0014] FIG. 3. List of non-cutting enzymes of the light chain variable region nucleic acid sequence shown in FIG. 2. [0015] FIG. 4. Nucleic acid sequence (double stranded) and possible amino acid sequences (depending on reading frame) of the 31.1 heavy chain variable region, showing restriction enzyme cleavage sites. [0016] FIG. 5. List of non-cutting enzymes of the heavy chain variable region nucleic acid sequence shown in FIG. 4. [0017] FIG. 6A-B. An antibody-dependent cellular cytotoxicity (ADCC) assay was conducted to test the effector funtion of the CHO chi 31.1 antibody against target cells SW1643 and PANC-1. As cell lysis occurs in the presence of 31.1 antibody (FIG. 6A) but not in the presence of control antibody (FIG. 6B). 5. DETAILED DESCRIPTION OF THE INVENTION Continue reading about Monoclonal antibody therapy for pancreas cancer... Full patent description for Monoclonal antibody therapy for pancreas cancer Brief Patent Description - Full Patent Description - Patent Application Claims Click on the above for other options relating to this Monoclonal antibody therapy for pancreas cancer 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. 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