26. Prognostic value of c-erbB2 gene amplification detected in archival human breast carcinomas by quantitative differential PCR

s 167 A (40 years old or younger) = 47.3%, group B (41-55 years) = 26.9%, group C (over 55 years) = 23.8%. Pain was the most frequent complaint (58.8%) in group A whereas a breast lump was most commonly found in groups B and C. In only 49.5% of the patients a breast mass was really detected at clinical examination. Breast cancer was found in 240 (10.2%) patients (group A = 0.8%, group B = 12.8%, group C = 25.8%). The predominant finding in all groups was fibrocystic breast disease (FBD). A statistically significant correlation between malignancy and breast lump was evident only in groups B and C (P < 0.05) but no correlation between malignancy and pain was found in any group (P = NS). The occurrence of cancer in 541 patients with FNAB-confirmed FBD over a median follow-up of 56 months (range 12-140 months) was similar to that reported for the general population. Many patients with breast complaints do not have any breast disease and more than 90% of the examined patients do not have cancer. In women 40 years of age and under with no history of malignancy the cancer rate is very low (< 1%) and a significant correlation between breast complaints and risk of cancer does not seem to exist. There is a need for a better information as to the significance of breast complaints. 26. Prognostic value of c-erbB2 gene amplification detected in archival human breast carcinomas by quantitative differential PCR H G Schniirch, H X An, D Niederacher, S I Dominik, S Scharl, U J Gbhring, H G Bender and M W Beckmann Heinrich-Heine-Universittit, Diisseldorf; und *Universitiit zu Kiiln, Germany Gene amplification is a common mechanism of proto-oncogene activation and contributes to tumour progression. Analysis of such genetic alterations is relevant to our understanding of tumour genetics and can provide prognostic information for the patients. Standard analytical approaches using Southern blotting require a large amount of good quality DNA. Quantitative differential polymerase chain reaction (qdPCR) requires a small amount of material and can be used to analyse archival tissues. A rapid, non-radioactive approach based on qdPCR and fluorescent DNA technique was applied for determination of c-erbB2 gene amplification. c-erbB2 gene amplification was correlated with other prognostic factors in a series of 195 archival breast cancer tissues. Sequences from the c-erbB-2 gene and from a single-copy reference gene (g-IFN) were amplified simultaneously by PCR, in which one of each primer pair was fluorescently labelled (fd-PCR). fd-PCR products were separated by polyacrylamide gel electrophoresis in an automated DNA sequencer (ALFTM Pharmacia) and directly quantified after laser activation and emission scanning using appropriate software (Fragment Managerm, Pharmacia). cerbB2 gene amplification was found in 52 of these tumours and correlated significantly with tumour size, absence of ER and pS2 expression, and presence of HER-Yneu protein, but not with absence of PgR or presence of EGFR expression, lymph node metastases or grading. In univariate analysis, c-erbB-2 gene amplification showed no significant correlation with patients’ outcome. Only, node-negative patients with c-erbB2 gene amplification had significantly decreased relapse-free survival and overall survival (P < 0.05). The fd-PCR assay is a valuable tool for determination of amplification of c-erbB-2 gene as well as of further oncogenes. Therewith, more detailed information about individual tumour biology and outcome may be acquired by this routine assay and provide prognostic impact. 27. Ln breast cancer deletions on the short arm of chromosome 17 (17p13.3~ter) are associated with a worse prognosis D S Liscia, T Venesio, M Donadio, C Palenzona, R Morizio, P Ferrer0 and A P M Cappa Torino, Italy Disease-free survival (DFS) is a strong indicator of the biological behaviour of human breast cancer. We have demonstrated that tumours showing allele loss on the short arm of chromosome 17 (17p13-ter) have a significantly higher proliferation rate compared with tumours that have a normal status of that chromosome region. Since it is known that growth rate correlates with malignancy and, therefore, with a worst prognosis, we have analysed the DFS of a panel of 140 primary breast tumours in order to test the hypothesis that loss of heterozygosity (LOH) at 17~13 is associated with a more aggressive phenotype. The initial intent of this study was based on the idea that the real target for these LOH was the known tumour suppressor gene TP53. However, we now have consistent evidence that TP53 (17~13.1) is not the only gene involved in the deletions of regions mapping at 17p since most of the LOH are telomeric with respect to this tumour suppressor gene. In fact the observed frequency TP53 LOH or point mutations was around 20% while the LOH at 17p13.3-ter in our study was 43%. The 17pl3.3ter region was analysed with Southern blots using VNTR probes (D17S5, Dl7S28, D17S34) and by PCR using both primers amplifying VNTR sequences and microsatellites (ca-repeats). The rate of recurrence in the group of patients with tumours that had a normal status of chr. 17p was 17.5% while 35% of the patients with tumours that suffered a LOH at that region had a recurrence within 6 years. Comparison of the Kaplan-Meier curves of the two groups showed a significant difference in DFS (x2 5.7, P = 0.01).