title: Recombinant mouse/human chimeric anti-colorectal carcinoma antibody cACT19.
abstract: A mouse/human chimeric antibody cACT19 derived from the murine ACT19 antibody was constructed; it recognizes an epitope different from the sialosyl-Tn on the TAG72 antigen as defined by the B72.3 antibody. This chimeric cACT19 antibody was constructed by using two expression vectors, the heavy chain expression vector mpSV2neo-EP1-VHC gamma 1 and the light chain expression vector mpSV2gpt-EP1-VKCK. These vectors contain the following: (i) the neo or gpt gene as a selection marker, (ii) the murine immunoglobulin promoter and enhancer (EP1), (iii) the genomic DNA fragments of human immunoglobulin constant region (CK and C gamma 1) and (iv) the murine cDNA fragments of VH or VK region cloned from the murine ACT19 cDNA library. These two vector DNAs were sequentially transfected into the SP2/0Ag14 cell line. Transfectants were selected in media containing both G418 and mycophenolic acid. The chimeric cACT19 antibody was purified from the transfectant supernate by Protein A Sepharose chromatography. We confirmed that the chimeric cACT19 antibody was reactive for an epitope that differed from the sialosyl-Tn on the TAG72 antigen. This was achieved by using the TAG72-binding inhibition ELISA utilizing various monosaccharides and disialyllato-N-tetraose with the latter containing the NeuAc2-6 alpha Ga1NAc structure. The immunohistochemical double-staining technique provided further evidence of the difference between the epitope defined by the chimeric cACT19 antibody and the sialosyl-Tn epitope by illustrating complementary as well as noncomplementary expression of these two epitopes in different areas of colon carcinoma tissues. We also demonstrated that the chimeric cACT19 antibody displayed much more effective ADCC and CDC for the human OVAR3 tumor cells than the murine ACT19 antibody. Therefore, the mouse/human chimeric anti-colorectal carcinoma cACT19 antibody may prove to be useful in cancer immunotherapy in its own right, or especially when used in combination with the chimeric B72.3 antibody.
authors.affiliation: Saskatoon Cancer Center, Department of Microbiology, University of Saskatchewan, Canada.
authors.forename: J
authors.initials: J
authors.lastname: Xiang
authors.forename: T
authors.initials: T
authors.lastname: Moyana
authors.forename: E
authors.initials: E
authors.lastname: Liu
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D000912:Antibodies, Neoplasm / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D001483:Base Sequence; D003001:Cloning, Molecular; D015179:Colorectal Neoplasms / Q000276:immunology*; D003165:Complement System Proteins / Q000502:physiology; D003602:Cytotoxicity, Immunologic; D018076:DNA, Complementary; D005260:Female; D006801:Humans; D006825:Hybridomas; D051379:Mice; D008969:Molecular Sequence Data; D010051:Ovarian Neoplasms / Q000276:immunology; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification
chemical_list: D000911:Antibodies, Monoclonal; D000912:Antibodies, Neoplasm; D018076:DNA, Complementary; D011993:Recombinant Fusion Proteins; D003165:Complement System Proteins