Volume 3 Supplement 1
Effect of anthracyclin-based neoadjuvant chemotherapy on disseminated tumor cells in breast cancer patients: an immunocytologic and molecular approach
© BioMed Central Ltd 2001
Received: 10 May 2001
Published: 31 May 2001
The leading cause of death from epithelial cancer is metastatic tumor relapse due to early dissemination of tumor cells. Cytokeratins are specific markers of epithelial cancer cells in bone marrow. As previously shown, these epithelial cells in bone marrow seem to be resting 'in dormancy'. This biological behaviour might be an explanation for the resistance to cytotoxic agents. In the present study, we evaluated whether primary chemotherapy in locally advanced, nonmetastatic breast cancer can eliminate cytokeratin-positive cells in bone marrow. Furthermore, we investigated the influence of primary chemotherapy on the tumor-associated gene expression.
Twenty-one breast cancer patients underwent bone marrow aspiration before and after neodjuvant chemotherapy. For immunocytologic tumor cell detection we used the monoclonal antibody 5D3 (Biogenex), which is directed against common epitope on cytokeratin polypeptides, including cytokeratin 8/18/19. For the molecular approach, after isolation of RNA, the reverse transcription with Superscript II Reverse Transkriptase(Gibco/BRL) and Oligo (dT)12-18 Primers (Gibco/BRL) was performed, followed by the amplification procedure with nested-RT-PCR for β2-microglobulin, muc-1, CK-20 and carcinoembryogenicantigen (CEA).
Fifteen patients met the inclusion criteria. Before chemotherapy, five out of 15 (30%) had cytokeratin-positive cells in bone marrow. Four out of five were still tested positive after finishing chemotherapy. With the RT-PCR procedure, all bone marrow aspirates showed a signal for β2-microglobulin (positive control).Expression of cytokeratin-20 was absent, whereas muc-1 was expressed in all aspirates. Two out of five showed expression of the CEA before chemotherapy, which was absent in one and only slight in the other case after chemotherapy. Before chemotherapy immunocytologic tumor cell detection and RT-PCR procedure for CEA was concurrently negative in 10 out of 15 patients.
(1) A negative result in immunocytochemistry of bone marrow of breast cancer patients seems to agree on the RT-PCR for CEA. (2) Tumor cells could develop a different pattern of gene expression under primary chemotherapy. (3) RT-PCR procedure for muc-1 and CK-20 seems not to be useful for investigating disseminated tumor cells in bone marrow.