Abstract 852: Novel role and mechanism of the LDHA protein as a survival factor for cancer cells.

Abstract

Abstract Rationale: Lactate dehydrogenase A (LDHA), which catalyzes the conversion of pyruvate to lactate, serves as a poor prognostic serum marker for many different types of human cancers; however, it is not known whether this is simply the result of LDHA being released from dying cancer cells in patients with high tumor burden or whether LDHA could also be a signaling molecule that portends poor clinical outcome. Based on the observation that extracellular LDHA was identified as the key survival factor for cardiomyocytes exposed to oxidative stress, we sought to determine whether extracellular LDHA can act as a cancer cell survival factor and hence contributes to poor clinical outcome. We reasoned that LDHA released from the damaged cells could activate intracellular signaling pathways in surrounding tumor cells by means of membrane receptors, ultimately protecting those surrounding tumor cells from oxidative stress-induced cell death. Methods: To investigate our hypothesis, recombinant LDHA and its isoform LDHB was produced by transforming E. coli strain BL21(DE3) with pET-SUMO expression vector (Clontech) containing full-length human LDHA or LDHB cDNA. The resulting recombinant proteins were purified using TALON Metal Affinity Resin and used for 16h pre-treatment of Ramos human Burkitt's lymphoma and A6L human pancreatic cancer cells cultured in full serum containing medium. The amount of recombinant proteins relative to serum albumin is estimated to be less 1% and hence any effects of LDHA could not be simply due to non-specific protein concentration effect. Following the pre-treatment procedure, these cancer cells were subjected to oxidative stress induced by hydrogen peroxide (H2O2) at 100 μM for 24h. Cells were then counted in a hemacytometer using trypan blue dye to exclude dead cells. Results: We observed that while LDHB had no effect on cell death induced by H2O2 as compared to the non-pretreatment group, LDHA can act as a protective factor against oxidative stress induced by H2O2, and ultimately cell death. We also investigated the mechanism involved in the protective role of LDHA by using recombinant LDHA as a molecular probe against membrane proteins that were extracted from cancer cells. Conclusion: Our study defines the novel role and mechanism of LDHA protein as a survival factor in cancer cells, in addition to its enzymatic activity, which may explain the prognostic significance of extracellular LDHA. Furthermore, this study can lead to the identification of new membrane receptor(s) for LDHA which may provide new targets for cancer therapy. Citation Format: Brad Poore, Joshua Kee Park, Iman Afif, Laura Shingleton, Chi Van Dang, Anne Le. Novel role and mechanism of the LDHA protein as a survival factor for cancer cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 852. doi:10.1158/1538-7445.AM2013-852