P53 MUTATIONS AND OVEREXPRESSION IN LOCALLY ADVANCED BREAST CANCERS

Alterations in the p53 gene were analysed in 39 patients with locally advanced breast cancers (LABCs) (stage III-IV) with inflammatory signs in most cases (UICC stage T4d = 32 patients) by molecular and immunoh istochemical (IHC) approaches. All patients were included in the same therapy protocol. Using polymerase chain reaction (PCR) and a single-s trand conformational polymorphism migration technique (SSCP), the pres ence of mutations in exons 2-11, covering the entire coding sequence o f the p53 gene, was evaluated. Using the mouse specific anti-human p53 monoclonal antibody (PAb 1801), we also looked for overexpression of the p53 protein in tissue sections. In 16 cases shifted bands were rep roducibly identified by PCR-SSCP, and all but one (localised to exon 1 0) were in exons 5-8, the usual mutational hotspots. Fifteen of these 16 samples were sequenced and 14 of the suspected mutations (36%) were confirmed. Most of them (12) were single nucleotide substitutions, an d transitions were more frequent (eight cases) than transversions (fou r cases). Fourteen of the tumour samples were positively stained with the monoclonal antibody PAb 1801, 11 with nuclear staining only, two w ith mixed cytoplasmic and nuclear staining and one with cytoplasmic st aining only. Staining patterns were very heterogeneous in terms of the percentage of positive cells (10-75%) and their distribution in the t issue section (isolated foci or dispersed cells). In 11 of the 14 muta ted cases a positive immunostaining was observed. The presence of a p5 3 mutation was significantly associated with larger tumour diameter (c hi(2) = 7.490, P = 0.0062) and the presence of clinical metastases (st age IV) (chi(2) = 10.113, P = 0.0015). A non-statistically significant trend of association was observed between p53 mutation, negative oest rogen receptors and lower response rate to therapy. Our results in thi s group of patients and the heterogeneity of the staining of tumour ce lls in tissue sections suggest that p53 mutations could be a late even t in this non-familiar form of breast cancer.