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Establishment of functional antibodies for inhibition on adhesion between colon cancer and human liver sinusoidal endothelial cells
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Abstract:
Abstract Objective: To screen for functional monoclonal antibody which can inhibit the adhesion between colon cancer cells and human liver sinusoidal endothelial cells (LSECs), and to identify the specific antigen of the screened antibody. Methods: BALB/c mice were immunized with LSECs, the spleen cells of the immunized mice were fused with SP2/0 cells and cultured on methyl cellulose to establish antiLSECs monoclonal antibody library. Functional monoclonal antibodies which inhibited the adherence between colon cancer cells (Ls174 T cells) and LSECs were screened and identified by immunofluorescence, ELISA and adhesive assay. The membrane proteins were extracted from LSECs and the specific antigen recognized by monoclonal antibody was detected by Western blotting assay. Results:Totally 1 440 monoclonal antibody clones were obtained by the fusing spleen cells of immunized mice with SP2/0 cells, 199 of the monoclonal antibodies recognized LSECs; and 20 clones, including 15 IgM type, 3 IgG type, and 2 with the heavy strand undetectable, significantly suppressed the adherence between colon cancer cells and LSECs. One of these clones with the strongest antiadhesive ability (inhibitory rate 51%) significantly inhibited the adhesion between colon cancer cells and HLSECs at 15-200 μg/ml in a dosedependent manner and reacted with antigens of 46 kD. Conclusion: We have successfully established an antiHLSEC monoclonal antibody library and obtained 20 functional monoclonal antibodies which can suppress the adherence of colon cancer cells with LSECs. One adhesive molecule which might mediate colon cancer liver metastasis has been preliminarily identified. These antibodies may help to understand the mechanism of organspecific metastasis of colon cancer cells and shed light on antimetastasis therapy of colon cancer.
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Project Supported:
Supported by the National Natural Science Foundation of China（No.30570818）; the Major State Basic Research Development Program of China（No.2002CB513106）
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