Chimeric (mouse/human) anti-colon cancer antibody c30.6 inhibits the growth of human colorectal cancer xenografts in scid/scid mice.
The mouse monoclonal antibody, m30.6 (IgG2b), detects an antigenic determinant expressed predominantly on the surface of colorectal adenocarcinoma cells and has been shown previously to be a potentially useful therapeutic and diagnostic reagent for human colon cancer. We report the production and characterization of a mouse/human chimeric antibody, c30.6, with potent in vitro and in vivo antitumor activity. The genes encoding the variable domains for heavy and light chains were amplified by thermal cycling using degenerate oligonucleotide primers complementary to conserved immunoglobulin framework sequences. The gene segments were sequenced, subcloned into eukaryotic expression vectors containing human constant region genes (IgG1 and kappa), and cotransfected into nonsecreting Sp2/0 mouse myeloma cells. There were significant differences in the biological activities of the murine and chimeric antibodies. The i.p. administration of c30.6 but not of m30.6 produced a marked growth inhibition of s.c. 30.6+ COLO 205 tumors in scid/scid mice (approximately 40% reduction in tumor size, measured 21 days after tumor inoculation). Reduced tumor growth was not due to altered binding characteristics of c30.6 because: (a) the chimeric antibody was shown by flow cytometry to bind exclusively to cell lines that expressed the 30.6 determinant; (b) c30.6 was able to completely inhibit the binding of m30.6 on 30.6+ cells; and (c) the affinity of binding of the two antibodies was the same (Ka, approximately 1.50 x 10(8)). Up to 15% of the total injected antibody dose/g tissue was localized in 30.6+ tumors at 24 h, approximately 13% was present in the tumors at 48 h, and approximately 10% was present at 72 h. Furthermore, c30.6 demonstrated a shorter circulating half-life (53 h; m30.6, 72 h) when given i.p. to C57BL6 x BALB/cF1 mice. Unlike m30.6, c30.6 was also strongly active in antibody-dependent cell-mediated cytotoxicity against a range of 30.6+ tumor target cells in vitro. Up to 80% specific 51Cr release was achieved using either freshly isolated human peripheral blood mononuclear cells or 2-day-old interleukin 2-stimulated human peripheral blood mononuclear cells as effectors. The enhanced antitumor activity of c30.6 suggests that it might be a useful immunotherapeutic reagent for colorectal carcinoma.
Austin Research Institute, Austin Hospital, Heidelberg, Australia.
P F
PF
Mount
V R
VR
Sutton
W
W
Li
J
J
Burgess
I F
IF
McKEnzie
G A
GA
Pietersz
J A
JA
Trapani
D000230:Adenocarcinoma / Q000276:immunology / Q000628:therapy*; D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000235:genetics / Q000378:metabolism / Q000627:therapeutic use*; D000912:Antibodies, Neoplasm / Q000235:genetics / Q000378:metabolism / Q000627:therapeutic use*; D000915:Antibody Affinity; D000920:Antibody-Dependent Cell Cytotoxicity; D001483:Base Sequence; D015179:Colorectal Neoplasms / Q000276:immunology / Q000628:therapy*; D006801:Humans; D007135:Immunoglobulin Variable Region / Q000235:genetics; D051379:Mice; D008807:Mice, Inbred BALB C; D008808:Mice, Inbred CBA; D016513:Mice, SCID; D008969:Molecular Sequence Data; D009368:Neoplasm Transplantation; D011993:Recombinant Fusion Proteins / Q000378:metabolism / Q000627:therapeutic use*; D014183:Transplantation, Heterologous
D000911:Antibodies, Monoclonal; D000912:Antibodies, Neoplasm; D007135:Immunoglobulin Variable Region; D011993:Recombinant Fusion Proteins