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    Research Grants Awarded

    Identifying New Tumor Suppressor Genes Involved In Human Breast Cancer

    Study Section:
    Tumor Cell Biology V

    Scientific Abstract:
    Cancers of the breast are amongst the most common neoplasms in women, with half of the patients dying of the disease. Cancer is caused by the accumulation of genetic mutations, among others in tumor suppressor genes. Few tumor suppressor genes have been identified that are frequently mutated in breast cancers, suggesting that there are still several major breast cancer tumor suppressor genes to be discovered. By allelotyping, we have identified a 1-Mb chromosomal region on 17q that was lost in 70% of 41 human breast cancer cell lines. Because only 10% of the cell lines had a mutation of the BRCA1 gene in this region, we hypothesize that there is yet another tumor suppressor gene located in its vicinity. This project aims to identify this putative 17q tumor suppressor gene. One hundred positional candidate genes from 17q will be screened for truncating mutations through culture of the cell lines with the drug Emetine, which inhibits the cellular mechanism of nonsense mediated decay of mRNA. Candidates will be selected for their Emetine-induced expression, based on microarray expression profiles and real-time RT-PCR. The selected candidates will be screened for mismatches in the gene sequence by LCGreen heteroduplex melting and subsequent sequencing. When at least two cell lines carry a truncating gene mutation, the gene will be sequenced in all 41 breast cancer cell lines. Candidates with inactivating gene mutations in at least 10% of the cell lines will be considered valid. The identified tumor suppressor gene will be validated by mutation analysis of primary breast cancer specimens. The sequence of a new tumor suppressor gene immediately provides a basis for the development of diagnostic assays. Identification of mutations of the gene and subsequent correlation with patient's prognosis may guide physicians in the treatment of patients. Insight in the biological implications of the gene mutations will aid the development of more rational treatment modalities.

    Lay Abstract:
    Breast cancer is the most common cause of death by cancer for women. Cancer develops due to an accumulation of mutations in so-called tumor suppressor genes. Only few tumor suppressor genes have been identified that are mutated at high frequency in breast cancers, and it is therefore likely that there are still several major breast cancer tumor suppressor genes to be discovered. We previously identified a region on chromosome 17q that was lost in almost three-quarters of 41 human breast cancer cell lines. Such regions of deletion suggest the location of a tumor suppressor gene. This project aims to identify the 17q tumor suppressor gene. The one hundred genes that are located in the deletion region are all potential candidates for the 17q tumor suppressor gene. We will screen all of these candidates for aberrant characteristics. We will first determine which of the candidates are differentially expressed by the breast cancer cell lines upon culturing of the cells with Emetine, a drug that re-expresses dysfunctional genes. Thus selected candidates will be evaluated for mutations in the gene, through mismatch screening and sequencing. If more than 10% of the breast cancer cell lines carry mutations that inactivate the gene, we conclude that we have identified the 17q tumor suppressor gene. The tumor suppressor gene will be validated in primary breast cancer tissues. Once we have identified the 17q tumor suppressor gene, a diagnostic assay for breast cancer can be developed. Correlation of the presence of gene mutations in breast cancers with the prognosis of the patients will generate guidelines for physicians in deciding on treatment options for breast cancer patients. Finally, studying the biological consequences of the gene mutations will allow the development of new treatments for breast cancer patients.