Abstract
Using whole viable human colon carcinoma HT29 cells as immunogen, we produced a monoclonal antibody (mAb) termed 69-6-5. The antibody was functionally selected on its anti-cell-spreading activity. By immunoprecipitation of surface radiolabeled cell lysates from HT29-D4 cells (an HT29 cell clone), mAb 69-6-5 recognized a molecular complex resembling integrin heterodimers. Sequential immunodepletions with mAb to the integrin αv subunit demonstrated that this complex was composed of αv-containing integrins. Accordingly, mAb 69-6-5 reacted with integrin αvβ3 immunopurified from melanoma cells and integrins αvβ5 and αvβ6 immunopurified from pancreatic carcinoma cells. In cell adhesion assays, the 69-6-5 mAb was able to inhibit strongly in a dose-dependent manner arginine-glycine-aspartic acid-mediated adhesion of HT29-D4 cells to vitronectin, fibronectin, or ProNectin F but not to laminin or collagen. Immunoprecipitations with β chain-specific antisera indicated that these cells express integrins αvβ5 (receptor for vitronectin) and αvβ6 (receptor for fibronectin) but neither αvβ1 nor αvβ3.
In summary, these results indicated that mAb 69-6-5 reacts with several αv integrins and that it can effectively interfere with the adhesive functions of at least αvβ5 and αvβ6, which represent the major receptors on HT29-D4 cells responsible for their adhesion on vitronectin and fibronectin.
Footnotes
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↵1 This work was supported by Institut National de la Santé et de la Recherche Médicale Grant CRE 920 205, the Fédération Nationale des Centres de Lutte Contre le Cancer, and the Association de Recherche sur le Cancer.
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↵2 To whom requests for reprints should be addressed.
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↵3 These two authors contributed equally to the completion of the biochemical part of this work.
- Received May 21, 1993.
- Accepted February 18, 1994.
- ©1994 American Association for Cancer Research.
Volume 54, Issue 8
