QUANTIFICATION AND CHARACTERIZATION OF TOTAL CELLULAR P53 PROTEIN IN COLORECTAL-CANCER

Immunochemical methods mere developed for the optimal detection and ch aracterization of total cellular p53 protein expression, both in the n uclear attached, and soluble fractions of colorectal cancers, in order to improve the correlation between protein deregulation and gene stat us. Seventy colorectal carcinomas mere studied using 3 monoclonal anti bodies in a sensitive analyzing system combining flow cytometry (nucle ar-bound fraction) and enzyme-linked immunosorbent assay (ELISA; solub le fraction). DNA indices ere calculated on the DNA histograms and mut ations of the p53 gene mere searched for in a subset of 41 cases. Thre e p53 expression patterns were found: 35 tumors were classified. as pa ttern ''A,'' characterized by high p53 expression including, ''mutant' ' conformation and missense mutations of the gene (16/17 cases tested) , pattern ''B'' consisted of 15 tumors with total absence of p53 expre ssion corresponding to nonsense mutations of the gene (8/9 cases teste d), and pattern ''C'' of 20 tumors presenting low or undetectable nucl ear-bound p53 but intermediate p53 protein content (pAb (1801+) in tbe soluble fraction. The latter pattern was associated with wild-type ge nes (14/15 cases tested), and with tumors that were often localized in the right colon compared to pattern ''A'' and ''B'' tumors (45% versu s 8%, P< 0.009) and were frequently near-diploid (80% versus 29%, P < 0.0002). Ne correlation was found between tumor stage and the patterns of p53 expression, The results indicate that both flow cytometry (FCM ) and ELISA seem necessary for the proper characterization of the p53 expression pattern, thus achieving a high degree of concordance with m olecular analysis of gene mutations, (C) 1998 Wiley-Liss, Inc.