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NY-BR-1 is a differentiation antigen and a potential target for cancer immunotherapy. Its mRNA expression is restricted to breast, testis, prostate and breast cancer by RT-PCR. In this study, we correlated NY-BR-1 protein and mRNA expression on tissue microarrays of mammary, prostatic and testicular malignancies using immunohistochemisty andin situhybridization with probes for exon 4–7 and 30–33. NY-BR-1 mRNA was confined to primary spermatocytes, suggesting a role in spermatogenesis. Exon 4–7 and 30–33 were equally expressed this cell type. However, NY-BR-1 was absent in all germ cell tumours analyzed (n= 475) and present in one of 56 (2%) prostate carcinomas. In breast, NY-BR-1 mRNA expression was detected in 307 of 442 (70%) primary carcinomas, with strong correlation to its protein expression (p< 0.0001). mRNA expression was significantly stronger and more frequently detected by the exon 30–33 probe than by the exon 4–7 probe (70%vs.35%,p< 0.0001), indicating the presence of alternative splice variants that lack 5-prime sequences. A similar restricted mRNA pattern was also observed in the normal breast epithelium. NY-BR-1 protein and mRNA correlated significantly with estrogen receptor α (ERα) protein expression (p< 0.0001), with stronger association to NY-BR-1 mRNA than protein (odds ratio 7.7 compared to 4.6). We identified 4 estrogen response elements (ERE)-like sequences nearby the promoter region, suggesting thatNY-BR-1transcription might be controlled by ERα. Accordingly, analysis of matching pairs of primary tumors with their recurrences showed a marked decrease of NY-BR-1 expression in recurrences after tamoxifen treatment (p< 0.0001).