Abstract 436: Folate cycle represents a new metabolic vulnerability for hepatocellular carcinoma treatment
Authors: Lee Derek , Xu Iris Ming-Jing , Chiu David Kung-Chun , Lai Robin Kit-Ho , Wong Chun-Ming , Ng Irene Oi-Lin , Wong Carmen Chak-Lui 
Published: July 2017

Abstract
Hepatocellular carcinoma (HCC), primary liver cancer, ranks the third most lethal cancer worldwide due to late symptom presentation and lack of promising curative therapy. Metabolic reprogramming has been recognized as a major and new hallmark of cancer in recent years. Better understanding of its underpinning molecular mechanisms favoring cancer growth will be crucial for the development of effective HCC therapeutics. The folate cycle fuels metabolic processes and the production of metabolites essential to cell growth and tumorigenesis maintenance. Through the shuttling of a single carbon unit by a folate derivative through the tetrahydrofolate (THF) backbone in the cytoplasmic and mitochondrial compartments, metabolites like NADPH - a major cellular antioxidant for redox balance, s-adenosyl methionine (SAM) - precursor of DNA and histone methylation, and pyrimidine and purine - the building blocks of DNA are produced. We found folate to be indispensable for HCC cell growth. Furthermore, methylene-THF dehydrogenase 1-like (MTHFD1L), a key enzyme facilitating the folate cycle from the mitochondria, was found to be significantly up-regulated in HCC with association to poorer clinical features for patients. Genetic inhibition of MTHFD1L by knockdown and knockout by shRNA and CRISPR-Cas9 systems, respectively, blocked NADPH production. Rapid elevation in oxidative stress induced DNA damage and cell cycle delay; ultimately inhibiting HCC proliferation. Binding of the transcription factor NRF2, a potent protector of oxidative stress, and MTHFD1L was confirmed by ChIP assay. NRF2 over-expression using the CRISPR-activating system in HCC cells further highlighted the dependent relationship between NRF2 and MTHFD1L. Metabolomics analysis showed that MTHFD1L knockdown caused a disruption to the folate cycle and accumulation of serine. Surprisingly, MTHFD1L knockdown did not reduce the levels of SAM and nucleotides. Knockdown of MTHFD1L in HCC cells significantly inhibited primary liver tumor growth and lung metastasis in orthotopic liver implantation model. Therapeutically, the administration of methotrexate, an anti-folate agent, sensitized HCC cells towards Sorafenib treatment both in vitro and vivo. The folate cycle represents a metabolic vulnerability and attractive therapeutic target for HCC. Inhibition of MTHFD1L disrupts the folate cycle and sensitizes HCC cells towards its convention treatment agent, Sorafenib in various HCC mouse models. Our investigation unravels a metabolic vulnerability in cancer which contributes to better understanding and is beneficial for the development of precise inhibitors specifically targeting associated pathways.
Citation Format: Derek Lee, Iris Ming-Jing Xu, David Kung-Chun Chiu, Robin Kit-Ho Lai, Chun-Ming Wong, Irene Oi-Lin Ng, Carmen Chak-Lui Wong. Folate cycle represents a new metabolic vulnerability for hepatocellular carcinoma treatment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 436. doi:10.1158/1538-7445.AM2017-436