ANALYSIS OF REPLICATION ERROR (RER(-CARCINOMA()) PHENOTYPES IN CERVICAL)

Infection of epithelial cells with human papillomavirus is an importan t early event in the development of cervical dysplasia. However, progr ession to overt malignancy appears dependent upon further genetic and/ or epigenetic events. We have recently developed methodologies for the simultaneous analysis of loss of heterozygosity (LOH) at multiple PCR -based microsatellite loci using semiautomated fluorescent DNA sequenc ing technology to determine the locations of tumor suppressor genes wh ich are inactivated during tumor progression. While examining 30 micro satellite loci for LOH on chromosomes 3p, 4, and 11q, we detected nove l tumor-specific alleles indicative of microsatellite instability (MI) . The methodology allowed rapid and accurate comparison of over 3000 g enotypes from 89 primary tumors and 10 cervical carcinoma-derived cell lines and showed that five tumors (5.6%) and one human papillomavirus -negative cell line, C33A, had genetic features consistent with a repl ication error (RER(+)) phenotype as defined by MI at two or more loci. In each of the RER(+) tumors, LOH was also observed at one or more lo ci on each of the three chromosomes examined. These findings suggest t hat defects in DNA repair-associated genes are rarely acquired and do not supersede allelic loss during cervical carcinogenesis, In addition , the semiautomated multiplex approach has proven unequivocal in the d etection and interpretation of MI and should greatly accelerate the ra pidity and accuracy of analyses of such defects in tumors. Moreover, t he number of loci that can be relatively easily examined in this way w ill also allow a detailed statistical consideration of the importance of such events.