Search for New Multidrug Resistant Genes by Methylation Sensitive Representational Difference Analysis (MS-RDA) in Breast Cancer

摘要

The WTH3 gene was obtained by its N-terminal sequence that was isolated by the Methylation Sensitive-Representational Difference Analysis (MS- RDA) technique due to its hypermethylation in the human multidrug resistant (MDR) breast cancer cell line MCF7/AdrR. The WTR3 gene product is 89% and 91% identical to the human Rab6 and Rab6c proteins, but possesses an elongated C- terminal region that contains 46 extra amino acids. Nevertheless, we consider WTH3 a new member of the RabS gene family. Semi-Quantitative RT-PCR results showed that WTH3 was 15 and 4 times down regulated in MCF7/AdrR and MES-SA- Dx5 MDR cell lines. Permanent expression of the WTH3 transgene in MDR cell lines increased to varying degrees their sensitivity to several anti-cancer drugs, which include doxorubicin (DOX), taxol, vinblastine, vincristine, and etoposide, as compared to the control sublines transfected with the empty vector. Flow cytometry and fluorescence microscope experiments suggest that the WTH3 transgene stimulates the host's uptake and retention of DOX. These results imply that the WTH3 gene plays a role(s) in MDR phenotype development in breast cancer cells.