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Abstract
Breast cancer is one of the most common malignancies in women. It continues to be a major burden and cause of death among women worldwide. Molecular oncology is now one of the most promising fields that may contribute considerably to diagnosis of breast cancer and its metastases addressing major problems with early detection, accurate staging, and monitoring of breast cancer patients. The overall objective of these feasibility studies was to contribute to improved diagnosis, prognosis, and prediction of breast cancer disease through the development of reagents and protocols for the use of molecular biological advances and the assessment of the relative potential of these diagnostic procedures for the detection and quantification of multiple specific mRNA tumor markers. Newest molecular technologies such as real-time quantitative TaqMan RT-PCR assays, microarray analyses, and production of “in-house” arrays were included in the study. Tissue, blood, and bone marrow samples were obtained from surgeries of confirmed and suspected breast cancer patients. TaqMan assays were performed for six mRNA markers: MAGE 3, HER2/NEU, MGB 1, CK 20, PSA, and HPR. Low-density nylon arrays with 265 immobilized genes included in cell to cell interactions were used for microarray analyses. Three highly overexpressed genes from microarray analyses and negative controls were selected for custom spotting on nylon membranes to produce “in-house” arrays. It was concluded that TaqMan assays can be easily designed and implemented for the screening of a large number of clinical specimens when including carefully selected controls, high purity RNA from samples, and a set of mRNA markers. Custom arrays can be produced incorporating multiple selected mRNA markers. It is suggested that the initial screening of biological samples could be done by microarray analyses and individual positive samples could be confirmed by additional tests using real-time quantitative TaqMan assays.
