P53 MUTATIONS IN HUMAN BLADDER-CANCER

Mutations in the tumor suppressor gene p53 play an important role in c arcinogenesis and tumor progression. To assess the status of p53 from genomic DNA from bladder cancer samples a two stage polymerase chain r eaction was employed. The technique provided material for subsequent d etection of mutations by Single Strand Conformation Polymorphism (SSCP ) analysis followed by DNA sequence analysis. SSCP analysis of exons 5 to 9 of p53 was performed using fragments from PCR end-labeled with P -32 followed by autoradiography using an electrophoresis system with t emperature control. This SSCP method improved resolution of mutations in exons 5, 7, and 8 and the sharpness of bands in exons 6 and 9. Bond s with altered migration patterns were excised from the dried SSCP gel s, reamplified by PCR, and sequenced. Mutations in conserved exons 5, 6, 7, 8, and 9 of the p53 gene were analyzed from bladder tumor biopsi es. Our results are consistent with the literature in that mutations i n p53 are predominantly found in high grade bladder cancer (Odds Ratio = 4.05, Fisher Exact P = 0.104); however, the results were not statis tically significant due to small numbers. Eight of 35 (23%) tumor samp les examined showed mutations in p53 (including two double mutations). Six of 13 (46%) grade III and IV tumors had p53 mutations vs. 2 of 17 (12%) grade I and II tumors. Normal individuals carried no p53 mutati ons. We found no correlation between pock years of smoking and mutatio n in p53. The spectrum of mutations confirmed a high proportion of G:C C:G transversions as well as the occurrence of double mutations. (C) 1994 Wiley-Liss, Inc.