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Am J Physiol Regul Integr Comp Physiol 271: R797-R805, 1996;
0363-6119/96 $5.00
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AJP - Regulatory, Integrative and Comparative Physiology, Vol 271, Issue 3 797-R805, Copyright © 1996 by American Physiological Society

ARTICLES

Characterization of the CCK-B/gastrin-like receptor in human colon cancer

J. P. Smith, E. A. Stock, M. G. Wotring, P. J. McLaughlin and I. S. Zagon
Department of Medicine, Milton S. Hershey Medical Center, Pennsylvania State University, Hershey, 17033, USA.

The gastrointestinal peptide, gastrin, tonically stimulates growth of human colon cancer cells in vivo and in vitro, and does so in a receptor-mediated fashion. This study defined the nature of gastrin binding in human colon cancer using [3H]L-365,260, a specific cholecystokinin B (CCK-B)/gastrin antagonist found to block gastrin's effects on growth. Following elucidation of optimal binding conditions (e.g., pH, time, and temperature) in log phase HT-29 human colon cancer cells, specific and saturable binding with a dissociation constant of 4.8 +/- 0.7 nM and a maximal binding capacity (Bmax) of 320 +/- 120 fmol/mg protein, consistent with a single binding site, was recorded. Binding was localized to the membrane fraction. Exposure to gastrin or receptor antagonist decreased and increased, respectively, the Bmax. Competition experiments indicated that L-365,260 was 25- and 200-fold more effective at displacing radiolabeled L-365,260 than gastrin and cholecystokinin, respectively. In contrast to log phase cells, the Bmax was decreased by 67 to 76% in confluent and postconfluent cultures. Binding activity was observed in other cell lines examined, as well as in xenografts and colon cancers obtained at surgery. Binding in normal human colonic mucosa was 10-fold less than in colon cancer. These results provide the first comprehensive identification and characterization of a CCK-B/gastrin-like receptor in human colon cancer.


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