Detection of Mammaglobin A-mRNA-positive circulating tumor cells in peripheral blood of patients with operable breast cancer with nested RT-PCR

OBJECTIVES: The development and validation of a nested RT-PCR methodology for the detection of Mammaglobin A-mRNA-positive circulating tumor cells in peripheral blood of patients with operable breast cancer and evaluation of its prognostic significance. DESIGN AND METHODS: Different combinations of specific primers were in silico designed and selected, so that false positive results due to genomic DNA contamination were avoided. The specificity of the primers used was evaluated in 30 healthy individuals, 20 patients with colorectal cancer and 20 patients with non-small cell lung cancer. The method was applied in 101 patients with operable breast cancer before the administration of adjuvant chemotherapy and 39 patients with metastatic breast cancer. RESULTS: Mammaglobin A-mRNA-positive cells were detected in 14/101 (13.9%) of early breast cancer patients but not in the control population studied (0%); 9 of them (64.3%) relapsed during the follow-up period. Mammaglobin A was detected in 7/39 (17.9%) of patients with verified metastasis. Multivariate analysis revealed the detection of Mammaglobin A-mRNA-positive cells, as an independent risk factor for reduced DFI. CONCLUSIONS: Mammaglobin A is a highly specific molecular marker for the detection of circulating tumor cells in operable breast cancer, with important prognostic applications.