ONCOGENE AMPLIFICATION IN ARCHIVAL OVARIAN-CARCINOMA DETECTED BY FLUORESCENT DIFFERENTIAL POLYMERASE CHAIN-REACTION - A ROUTINE ANALYTICAL APPROACH

For quantitative determination of gene amplification of oncogenes invo lved in ovarian cancer development we have established a rapid, nonrad ioactive approach. Determination of c-erbB-2 and c-myc gene amplificat ion in archival ovarian carcinoma specimens was based on differential polymerase chain reaction (dPCR) and fluorescent DNA technique. c-erbB -2 or c-myc sequences were amplified by PCR, in which one of each prim er pair was fluorescently labeled, simultaneously with a reference gen e (gamma-IFN) as internal single copy gene control. PCR products were separated by polyacrylamide gel electrophoresis with an automated DNA sequencer (A.L.F.(TM), Pharmacia, Freiburg, Germany) and directly quan tified after laser activation and emission scanning using appropriate software (Fragment Manager(TM), Pharmacia, Freiburg, Germany). This fl uorescent differential PCR (fdPCR) method was used for quantitative de termination of c-erbB-2 and c-myc gene amplification in 79 formalin-fi xed, paraffin-embedded epithelial ovarian carcinoma tissues and 15 ben ign ovarian tissues. c-erbB-2 gene amplification was found in 18 (22%) and c-myc gene amplification in 13 (17%) of these carcinomas, but cou ld not be detected in benign ovarian tissues. Single oncogene amplific ation correlated significantly with higher stage and higher grade (P<0 .05). Patients with either c-erbB-2 or c-myc gene amplification or bot h had significantly shorter relapse-free survival and overall survival . The fdPCR assay is a valuable tool for determination of oncogene amp lification even in archival tumor tissue. More detailed information ab out individual tumor biology of the primary tumor and metastasis as we ll as criteria for additional therapeutic decisions may be acquired by this analytical approach.