Abstract 5413: Intracellular ATP reduction by an artificial protein as a novel approach to cancer therapy

Abstract

Introduction: The use of chemotherapeutic agents that target key metabolic processes in cancer cells is an important therapeutic strategy. As a result of reduced mitochondrial activity, adenine triphosphate (ATP) is produced mainly by aerobic glycolysis and lactic acid fermentation in the cytosol of cancer cells (Warburg effect). Since ATP is a vital energy carrier that is crucial for various biochemical reactions in cancerous cells, interfering with the production/availability of ATP offers a new approach to cancer therapy. To assess this idea, a man-made synthetic ATP-binding protein, DX, was tested as a novel cancer therapy. Hypothesis: Reduced ATP availability, due to the expression of DX in HeLa cells, will negatively affect cellular metabolism and viability. Methods: For controlled expression, DX was cloned into the pcDNA™5/TO expression vector (with an N-terminal FLAG tag), whose expression is driven by a tetracycline-regulated cytomegalovirus promoter. Cultured T-REx™ HeLa cells (which stably express the tetracycline repressor protein) were transfected with pcDNA™5/TO::DX. As a control for transfection and DX expression, T-REx™ HeLa cells were transfected with pcDNA™5/TO/lacZ and cultured under identical conditions. The impact of DX expression on cell viability and morphology was assessed using a tetrazolium-based colorimetric cell viability assay and by light and fluorescent microscopy. To assess the impact of ATP depletion in combination with currently used anti-cancer agents on HeLa cell viability, toxicity assays were performed comparing Doxorubicin, Paclitaxel, or Gemcitabine treatment alone or in combination with DX. Results: HeLa cells expressing DX experienced reduced viability and altered morphology compared to cells expressing lacZ alone. These results suggest that ATP depletion potentiate cell death in HeLa cells. HeLa cells treated singly with DX do not possess the same level of toxicity as compared with the tested chemotherapy agents. Of the chemotherapy agents assayed, DX displayed a synergistic effect with Doxorubicin to reduce cell viability. Conclusion: Reduced intracellular ATP levels, mediated by a synthetic ATP-binding protein, negatively affects the viability of cervical cancer cells. The reduction in ATP, and the consequence of this on biochemical pathways and cell viability requires further investigation. We suggest that artificial proteins, such as DX, could be used to control and regulate specific targets in metabolic pathways and represents a new approach for cancer chemotherapy.

Citation Format: Dennis Datuin, Matt Hamada, Bernardo Chavira, Andrew Fontes, Nagaraj Vinay Janthakahalli, Shaleen Korch. Intracellular ATP reduction by an artificial protein as a novel approach to cancer therapy. [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 5413. doi:10.1158/1538-7445.AM2015-5413