Summary Forty-two patients with clinical stage IIIA or IIIB breast cancer were treated with neoadjuvant chemotherapy followed by mastectomy and radiotherapy. The median follow-up was 32 months (range 10-72 months) and the median time to progression was 17 months (range 10-30 months). A multivariate analysis showed that a longer disease-free survival (DFS) was related to more chemotherapy cycles given (P = 0.003), a better pathological response to chemotherapy (P = 0.04) and fewer positive axillary lymph nodes (P = 0.05). A better overall survival (OS) was related to more chemotherapy cycles given (P = 0.03) and better pathological response to chemotherapy (P= 0.04). In patients with residual tumour after neoadjuvant chemotherapy, high levels of staining for Ki-67 was correlated with a worse DFS (P = 0.008). Other biological characteristics, including oestrogen receptor status, microvessel density (CD31 staining), P-glycoprotein (P-gp) staining and nuclear accumulation of p53, were not independent prognostic factors for either DFS or OS. If both P-gp and p53 were expressed, DFS and OS were worse in the uni-and multivariate analysis. The preliminary results of this phase 11 study suggest that coexpression of P-gp/p53 and a high level of staining for Ki-67 after chemotherapy are associated with a worse prognosis, and that prolonged neoadjuvant chemotherapy and the attainment of a pathological complete remission are important factors in determining outcome for patients with this disease.

Summary Expression of both P-glycoprotein (P-gp) and mutant p53 have recently been reported to be associated with poor prognosis of breast cancer. The expression of P-gp is associated in vitro and in vivo with cross-resistance to several anti-cancer drugs. p53 plays a regulatory role in apoptosis, and mutant p53 has been suggested to be involved in drug resistance. Interestingly, in vitro experiments have shown that mutant p53 can activate the promoter of the MDRI gene, which encodes P-gp. We investigated whether p53 and P-gp are simultaneously expressed in primary breast cancer cells and analysed the impact of the co-expression on patients' prognosis. Immunohistochemistry was used to investigate P-gp expression (JSB-1, C219) and nuclear p53 accumulation (DO-7) in 20 operable chemotherapy untreated and 30 locally advanced breast cancers undergoing neoadjuvant chemotherapy with doxorubicin and cyclophosphamide. Double immunostaining showed that P-gp expression and nuclear p53 accumulation often occur concomitantly in the same tumour cells. A correlation between p53 and P-gp expression was found in all 50 breast cancers (P=0.003; Fisher's exact test). P-gp expression, nuclear p53 accumulation, and co-expression of p53 and P-gp were more frequently observed in locally advanced breast cancers than in operable breast cancers (P=0.0004; P=0.048; P=0.002 respectively, Fisher's exact test). Co-expression of p53 and P-gp was the strongest prognostic factor for shorter survival by multivariate analysis (P=0.004) in the group of locally advanced breast cancers (univariate analysis: P=0.0007). Only 3 out of 13 samples sequentially taken before and after chemotherapy displayed a change in P-gp or p53 staining. In conclusion, nuclear p53 accumulation is often associated with Pgp expression in primary breast cancer, and simultaneous expression of p53 and P-gp is associated with shorter survival in locally advanced breast cancer patients. Co-expression of P-gp and mutant p53 belong to a series of molecular events resulting in a more aggressive phenotype, drug resistance and poor prognosis.

In 42 patients with locally advanced breast cancer treated with neoadjuvant chemotherapy followed by surgery and radiation therapy, the effects of chemotherapy on tumor architecture, morphometric nuclear and nucleolar characteristics, DNA ploidy, proliferation index measured by mitotic activity index, expression of differentiation antigens, and microvessel density were studied. Pretreatment biopsy specimens were available to compare with mastectomy specimens for 24 patients, and subclavicular biopsy specimens taken before chemotherapy were available for 9 patients. In the remaining patients, fine-needle aspiration was performed before chemotherapy, and morphologic and biologic features of the tumors could be studied only after chemotherapy. In 23 patients, only microscopic tumor or no tumor was left after chemotherapy, and in these patients we observed a characteristic pattern of relatively cellular fibrous tissue with lymphocytic infiltrate, ironloaded macrophages, and, when present, scattered foci of tumor cells in between. We found a reduction in mitotic activity index and in global microvessel density over all the tumors as a group. There was, however, no consistent pattern of changes in nuclear and nucleolar morphometric characteristics, DNA ploidy, and expression of differentiation antigens, and no pathologic or biologic features were predictive for response to chemotherapy.

Pgp expression in tumor cells, and especially when accompanied by Pgp expression in fibroblasts in desmoplastic stroma, has prognostic value in primary breast cancer patients and is likely to be a marker of a more malignant phenotype. Pgp expression of tumor cells might play a role in tamoxifen resistance. These findings may have important implications for teh treatment of breast cancer patients, and warrant further prospective investigation in a larger patient population.

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