Abstract 3039: Targeting mitochondrial DNA polymerase gamma (POLG) as a novel therapeutic strategy for acute myeloid leukemia (AML)

Abstract

Abstract Mitochondria contain their own 16.6kb genome (mtDNA) which encodes 13 protein-coding genes essential for electron transport chain activity. In mammalian cells, it is replicated solely by the nuclear-encoded mitochondrial DNA polymerase gamma (POLG), which exists as a heterotrimer with POLG2 to perform replication and repair of mtDNA. Through bioinformatic analyses, we observed increased POLG expression in a subset of AML cell lines compared to other cancer cell lines. As a subset of AML patients also have unique mitochondrial characteristics, including increased mitochondrial mass and mtDNA content compared to normal hematopoietic stem cells (Skrtic et. al., Cancer Cell, 20:674, 2011), we addressed the effects of targeting POLG as a novel therapeutic strategy for AML. OCI-AML2 and TEX leukemia cells were treated with the anti-retroviral drug 2′,3′-dideoxycytidine(ddC) that exhibits off-target inhibition of POLG catalytic activity. At concentrations as low as 200nM, ddC depleted mtDNA, decreased mRNA expression of mtDNA transcripts, reduced the expression of mitochondrial encoded proteins COX 1 and 2 subunits that form the catalytic core of respiratory chain complex IV, decreased basal oxygen consumption rate (OCR), and reduced the proliferation and viability of AML cells. However, AML cells had large reserves in their mtDNA; as significant changes in mitochondrial proteins, metabolism, proliferation and viability were only observed with a threshold of >95% depletion of mtDNA. In contrast, lesser reductions in mtDNA content with ddC did not significantly affect levels of mitochondrial transcripts, metabolism, or cell viability. Next, we used a genetic approach and explored the impact of knocking down POLG with multiple independent shRNA in AML cells. POLG knockdown in OCI-AML2 cells depleted mtDNA content by 60% compared to controls. Consistent with the findings with ddC, mtDNA depletion to this extent produced only quantitatively minor changes in the protein expression of the mtDNA-encoded COX 1 and 2 and basal OCR. Despite the minimal effect on mitochondrial metabolism, POLG knockdown decreased the growth and viability of OCI-AML2 cells, and increased apoptosis, as measured by Annexin V staining. Results of electron microscopy analyses indicate that POLG knockdown disrupted mitochondrial cristae structure compared to controls. In summary, these results indicate that POLG plays a role in maintaining AML cell viability and mitochondrial structure and this function is independent of mtDNA replication and oxidative metabolism. Citation Format: Sanduni Liyanage, Rose Hurren, Rebecca Laposa, Aaron Schimmer. Targeting mitochondrial DNA polymerase gamma (POLG) as a novel therapeutic strategy for acute myeloid leukemia (AML). [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3039. doi:10.1158/1538-7445.AM2015-3039