ANALYSIS OF HLA CLASS I ANTIGEN EXPRESSION IN BREAST CANCER BY IMMUNOHISTOCHEMISTRY, BIOCHEMISTRY AND MOLECULAR BIOLOGY.
MP Pistillo, P Capanni, G Nicolò°, S Salvi°, L Perdelli°, GL Palmisano and GB Ferrara; Immunogenetics Lab. and °Pathology Lab, , National Cancer Institute c/o Advanced Biotechnology Center, Genova , Italy.
Downregulation of HLA class I antigen expression has been reported in a significant proportion of primary breast carcinomas suggesting an escape mechanism from CTL mediated lysis leading to tumor dissemination and metastasis. In our study we investigated the expression of HLA total class I, alfa-chain and beta2-m subunits in 60 primary breast carcinomas by conventional immunohistochemistry (IHC) and by biochemistry on a selected number of samples.
Three different class I expression patterns could be observed by IHC: High downregulation pattern, found in 43% of tumors, mostly associated with high downregulation of the alfa-chain; Low downregulation pattern, found in 20% of tumors, associated with similar amount of alfa- and beta-chains downregulation; Absent downregulation pattern, found in 37% of tumors, with few cases of alfa-chain low downregulation not interfering with class I expression. The frequency of class I downregulation of 8 primary tumors was compared with that of autologous lymphonode metastasis resulting in a higher incidence of HLA class I downregulation in metastasis.
Twentyfive out of the 60 samples were selected for biochemical studies on the basis of their minimal non neoplastic cell contamination. Quantitative western blot (WB) was performed on the class I alfa- and beta-chains from the same tumor tissues used for IHC and the corresponding autologous PBLs were used as a basis for comparison. Again three different downrregulation patterns could be observed by WB which were compared with the IHC downregulation patterns.
By using molecular techniques (Northern blot, RT-PCR and PCR amplifications with locus specific primers for beta2-m, HLA-A and -B, TAP1 and TAP2 ) we are investigating the molecular mechanisms underlying the observed HLA class I downregulations.