[Background] Restriction landmark genomic scanning (RLGS) is a type of two-dimensional electrophoresis which has been used to study genetic and epigenetic changes in normal tissues, primary tumors and cancer cell lines. In this study, we examined the genetic changes in breast cancer by RLGS.
[Material and methods] Genomic DNAs were extracted from breast cancer samples and their noncancerous counterparts of 14 patients who had undergone curative operation at Keio University hospital. Genomic DNAs were cleaved by restriction enzyme Not I, radiolabeled at cleavage sites, and then size-fractionated by two-dimensional electrophoresis. Comparing RLGS profiles between breast cancer tissues and noncancerous counterparts, significant change of spot was marked and nucleotide sequence and homology search analysis of the spot was performed. The RNA expression of the detected gene was examined in one breast cancer cell line, MCF7, two other cell lines (WiDr and A431) and seven breast cancer specimens by real-time PCR method.
[Results] We found one spot which was observed in four cancerous profiles but not observed in any noncancerous profiles. Nucleotide sequence and homology search analysis showed that the spot represented voltage-dependent calcium channel α 1H subunit gene (CACNA1H). Expression of CACNA1H was found in MCF-7 (Estrogen receptor (ER) positive and HER2 negative) and two breast cancer tissue samples but not found in WiDr (ER and HER2 negative), A431 (ER negative, EGFR and HER2 positive) and the other five tissue samples. The two samples which the expression of the gene was found consisted of one estrogen receptor positive and the other estrogen negative sample. [Conclusion] These results suggest that CACNA1H is a breast cancer related gene which is independent from ER and HER2 expression.
98th AACR Annual Meeting-- Apr 14-18, 2007; Los Angeles, CA