P2.02-032 Interplaying between Gamma-H2AX and Autophagy in A549 Cells Treated with Cisplatin and Etoposide

Abstract

DNA damage-repair and autophagy may contribute to the efficacy of anticancer therapy. The gamma-H2AX is phosphorylated to repair DNA after the double stranded breakdown. Thus, this study was aimed to check the expression of gamma-H2AX and autophagy in cisplatin- and etoposide-treated lung cancer cell line. A549 cells were cultured within Dulbecco’s Modified Essential Medium with 10% of fetal bovine serum. Cisplatin and etoposide were treated into the cells with time and dose dependent manners. MTT assay and light microscopy were performed to determine the LD50 of these drugs. Western blotting was performed to define the expression of proteins related to LC3A/B-I/II, NBR1 and DNA damage (gamma-HA2X). Based on MTT assay, LC50’s were determined as the followings: LC50 of CDDP was 14.4 micromole/L and LC50 of etoposide was approximately 25 micromole/L. Chemoresistant cancer cell islets were defined the surviving cells under light microscopy with the treatment of cisplatin and etoposide. In 0-, 5-, 10- and 20-h time points, gamma HA2X showed increasing expressional pattern. This pattern was also similar in etoposide with the peak time point of 10 hour. LC3A/B-I/II and NBR1 expressions were increased by 25 micromole/L of CDDP and 100 micromole/L of etoposide whose patterns were also similar to gamma H2AX. Similar expressional patterns of DNA breakdowns (gamma H2AX) and autophagy (LC3A/B-I/II, NBR1) by cisplatin and etoposide may suggest the linkage between autophagy and DNA breakdown. Therefore, this tentative conclusion must be furthermore defined by the research for molecular networks in the chemoresistant lung cancer cells.