English abstract
Breast and ovarian cancers are both devastating heterogeneous diseases. Breast cancer accounts for 25% of cancers among women worldwide. In Oman, breast cancer accounts for 26% of cancer cases among women. Although, research has identified and developed potential predictive markers, none is a reliable in predicting outcome causing high mortality, particularly in triple negative breast cancer subtype. This study aimed at identifying potential biomarkers differentially expressed in the molecular subtypes of breast cancer involved in breast cancer pathogenesis. Microarray gene expression profiling was conducted on 24 clinical samples covering Luminal A, Luminal B, Her2+ and Triple negative subtypes of breast cancer with their corresponding normal tissues. Data analysis revealed three significantly up-regulated (BRIPI, HOXB6 and EYAI) and two down-regulated (DSP and RHPN2) candidate genes. RT-PCR and western blot analyses validated the differential gene expression profiles of the genes selected. Furthermore, localization of the identified genes in-situ was determined by immunohistochemistry. Kaplan Meier survival analysis showed that BRIPI, HOXB6, EYAI, DSP and RHPN2 were associated with poor survival.
On the other hand, ovarian cancer is rare comprising 3,5% of cancer cases in women. In Oman, ovarian cancer accounts for 4.5% cancer cases among women. To identify early stage biomarkers involved in the pathogenesis of epithelial ovarian cancer (EOC), chromatin immunoprecipitation (CHIP) was performed using the low (MCAS) and high grade (OVSAHO) ovarian cancer cell lines and a specific antibody raised against E2Fs, a transcription factor involved in early stages of EOC pathogenesis to identify downstream genes that might have a role in EOC. Data from ChIP analysis revealed a list of 118 genes under the regulation of the transcription factor, E2F5. While, 36 genes were found in MCAS, 68 genes were identified in OVSAHO. Analysis between MCAS and OVSAHO in-vitro models identified 14 common genes. Among the short-listed genes, FBXW7 was identified as one of the potential gene with an important role in ovarian cancer pathogenesis. Gene panel exome sequencing on 6 EOC samples against 5 normal/benign ovarian tissues revealed infrequent mutations of FBXW7 gene in ovarian cancer, FBXW7 gene expression was assessed in ovarian cancer cell lines (MCAS, OVSAHO and OV2009) by RT-PCR and western blotting. Expression of FBXW7 was high in MCAS and low in OV2008 and OVSAHO cells. FBXW7 expression was restored in OVSAHO and OV2008 cells by treatment with the demethylating agent, 5' aza-2' deoxycitidine. Western blotting results showed an upregulation of apoptotic proteins, indicating that cells underwent apoptosis. Analysis of FBXW7 gene methylation status showed OVSAHO and OV2008 as well as five out of eight (62.5%) EOC patients displayed methylation. Response of OV2008 and OVSAHO cells to drugs showed that OVSAHO was sensitive to platinum-drugs, while, OV2008 was highly resistant. Interestingly, resistant OV2008 cells treated with 5-azacitidine, showed an enhanced sensitivity to platinum, confirming the tumor suppressor role of FBXW7 gene in ovarian cancer. In conclusion, this research study identified genes with a putative role as biological markers in a population characterized by an early onset of both breast as well as ovarian cancer, which might contribute to the development of targeted therapies for breast and ovarian cancer.
