Mitochondrial Reprogramming Via ATP5H Loss Promotes Multimodal Cancer Therapy Resistance
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Mitochondrial reprogramming via ATP5H loss promotes multimodal cancer therapy resistance Kwon-Ho Song, … , T.C. Wu, Tae Woo Kim J Clin Invest. 2018;128(9):4098-4114. https://doi.org/10.1172/JCI96804. Research Article Immunology Oncology The host immune system plays a pivotal role in the emergence of tumor cells that are refractory to multiple clinical interventions including immunotherapy, chemotherapy, and radiotherapy. Here, we examined the molecular mechanisms by which the immune system triggers cross-resistance to these interventions. By examining the biological changes in murine and tumor cells subjected to sequential rounds of in vitro or in vivo immune selection via cognate cytotoxic T lymphocytes, we found that multimodality resistance arises through a core metabolic reprogramming pathway instigated by epigenetic loss of the ATP synthase subunit ATP5H, which leads to ROS accumulation and HIF-1α stabilization under normoxia. Furthermore, this pathway confers to tumor cells a stem-like and invasive phenotype. In vivo delivery of antioxidants reverses these phenotypic changes and resensitizes tumor cells to therapy. ATP5H loss in the tumor is strongly linked to failure of therapy, disease progression, and poor survival in patients with cancer. Collectively, our results reveal a mechanism underlying immune-driven multimodality resistance to cancer therapy and demonstrate that rational targeting of mitochondrial metabolic reprogramming in tumor cells may overcome this resistance. We believe these results hold important implications for the clinical management of cancer. Find the latest version: https://jci.me/96804/pdf RESEARCH ARTICLE The Journal of Clinical Investigation Mitochondrial reprogramming via ATP5H loss promotes multimodal cancer therapy resistance Kwon-Ho Song,1,2,3 Jae-Hoon Kim,4 Young-Ho Lee,1,2,3 Hyun Cheol Bae,5 Hyo-Jung Lee,1,2,3 Seon Rang Woo,1,2,3 Se Jin Oh,1,2,3 Kyung-Mi Lee,1,2 Cassian Yee,6 Bo Wook Kim,7 Hanbyoul Cho,4 Eun Joo Chung,8 Joon-Yong Chung,9 Stephen M. Hewitt,9 Tae-Wook Chung,10 Ki-Tae Ha,10 Young-Ki Bae,11 Chih-Ping Mao,12,13,14 Andrew Yang,14 T.C. Wu,14,15,16,17 and Tae Woo Kim1,2,3 1Department of Biochemistry and Molecular Biology, 2Department of Biomedical Science, College of Medicine, and 3Translational Research Institute for Incurable Diseases, Korea University College of Medicine, Seoul, South Korea. 4Department of Obstetrics and Gynecology, Gangnam Severance Hospital, Yonsei University College of Medicine, Seoul, South Korea. 5Department of Orthopedic Surgery, Seoul National University Hospital, Seoul, South Korea. 6Department of Gynecologic Oncology, The University of Texas MD Anderson Cancer Center, Houston, Texas, USA. 7Department of Obstetrics and Gynecology, International St. Mary’s Hospital, Catholic Kwandong University College of Medicine, Incheon, Seoul, South Korea. 8Radiation Oncology Branch and 9Experimental Pathology Laboratory, Laboratory of Pathology, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, Maryland, USA. 10Division of Applied Medicine, School of Korean Medicine, Pusan National University, Yangsan, South Korea. 11Comparative Biomedicine Research Branch, Research Institute, National Cancer Center, Goyang, South Korea. 12MD-PhD Program, 13Immunology Training Program, 14Department of Pathology, 15Department of Oncology, 16Department of Obstetrics and Gynecology, and 17Department of Molecular Microbiology and Immunology, Johns Hopkins School of Medicine, Baltimore, Maryland, USA. The host immune system plays a pivotal role in the emergence of tumor cells that are refractory to multiple clinical interventions including immunotherapy, chemotherapy, and radiotherapy. Here, we examined the molecular mechanisms by which the immune system triggers cross-resistance to these interventions. By examining the biological changes in murine and tumor cells subjected to sequential rounds of in vitro or in vivo immune selection via cognate cytotoxic T lymphocytes, we found that multimodality resistance arises through a core metabolic reprogramming pathway instigated by epigenetic loss of the ATP synthase subunit ATP5H, which leads to ROS accumulation and HIF-1α stabilization under normoxia. Furthermore, this pathway confers to tumor cells a stem-like and invasive phenotype. In vivo delivery of antioxidants reverses these phenotypic changes and resensitizes tumor cells to therapy. ATP5H loss in the tumor is strongly linked to failure of therapy, disease progression, and poor survival in patients with cancer. Collectively, our results reveal a mechanism underlying immune-driven multimodality resistance to cancer therapy and demonstrate that rational targeting of mitochondrial metabolic reprogramming in tumor cells may overcome this resistance. We believe these results hold important implications for the clinical management of cancer. Introduction may contribute to drug resistance. In this study, we investigated The adaptation of tumor cells to immune surveillance as well as this concept and its underlying mechanisms. the emergence of refractory tumor cells after initial therapy are To explore this issue, we developed a system to simulate responsible for eventual disease relapse, progression, and metas- the evolution of cancer cells in a live host and to dissect the tasis. Multiple tumor-intrinsic mechanisms have been proposed mechanisms responsible for escape from immune defens- to cause tumor drug resistance (1, 2), including the overexpres- es mounted by CD8+ cytotoxic T lymphocytes (CTLs), the sion of antiapoptotic factors (3), the ejection of pharmacologic predominant arm of the cancer immune surveillance net- agents from tumor cells via drug transporters (4), the biochem- work (16–18). Through this analysis, we report here that loss ical or structural modification of the drug target (5–8), and the of the D subunit of mitochondrial ATP synthase (ATP5H) adoption of detoxification or bypass routes (9–14). Furthermore, constitutes an important pathway through which can- a recent study provides evidence that preferential selection and cer evolution occurs in response to immune editing. Mito- subsequent expansion of a subset of stem-like tumor cells with chondrial ATP5H is a critical component of the energy- an undifferentiated phenotype contribute to resistance (15). producing apparatus in eukaryotic cells; thus, disruptions in ATP However, the potential link between host immune-intrinsic synthase expression in cancer cells would be expected to influ- mechanisms (i.e., cancer immune editing) and the development ence tumor metabolism. In fact, altered metabolism is a funda- of resistance to therapy is not well understood. We hypothesized mental feature of cancer and has been shown to contribute to that selection pressures imposed by host immune surveillance chemo- and radiotherapy resistance (19–21). Tumor mitochon- drial dysfunction promotes the development of chemoresis- tance and has increasingly been recognized as an integral target Authorship note: KHS, JHK, YHL, and HCB contributed equally to this work. for new cancer therapy approaches (22, 23). Likewise, downreg- Conflict of interest: The authors have declared that no conflict of interest exists. ulation of ATP synthase components has been documented in Submitted: August 9, 2017; Accepted: June 28, 2018. Reference information: J Clin Invest. 2018;128(9):4098–4114. a majority of carcinomas (24, 25) and has been associated with https://doi.org/10.1172/JCI96804. chemotherapy resistance (26). However, the underlying mech- 4098 jci.org Volume 128 Number 9 September 2018 The Journal of Clinical Investigation RESEARCH ARTICLE Figure 1. Immune editing triggers epigenetic loss of ATP synthase in tumor cells. (A) ATP5H protein levels in TC-1 or CaSki tumor cells at various stages of immune editing were determined by Western blot analysis (numbers below each blot are densitometric values). (B) mRNA levels of Atp5h (TC-1 cells) or ATP5H (CaSki cells) in TC-1 or CaSki tumor cells at various stages of immune editing were determined by qRT-PCR. (A and B) N1, N2, and N3 were generated through serial selection by irrelevant antigen specific CTLs and were used as negative controls. (C) TC-1 or CaSki P0 and P3 tumor cells were treated with DMSO, 5-AzaC (5 μM), or TSA (100 nM). Atp5h (TC-1 cells) or ATP5H (CaSki cells) mRNA levels in these cells were probed by qRT-PCR. (D) The histone acetylation status of TC-1 or CaSki P0 and P3 cells was determined by ChIP using anti–histone H3 and H4 antibodies, fol- lowed by qPCR for the Atp5h or ATP5H promoter locus. (E) TC-1 or CaSki P3 cells were treated with or without TSA, and histone H4 acetylation at the Atp5h or ATP5H promoter was determined by ChIP-qPCR. (F–H) TC-1 or CaSki P3 cells were transfected with the indicated siRNAs. (F) Atp5h or ATP5H mRNA levels were determined by qPCR. (G) Protein levels of HDAC1 and ATP5H were determined by Western blot analysis (numbers below each blot are densitometric values). (H) Relative levels of AcH4 at the Atp5h or ATP5H promoter were assessed by qChIP. All experiments were performed in triplicate. *P < 0.05 and **P < 0.01, by 1-way ANOVA (B and C) or 2-tailed Student’s t test (D–F and H). Data represent the mean ± SD. anisms responsible for linking ATP synthase loss to drug resis- Results tance have remained unclear. Immune editing triggers multimodal resistance to chemotherapy and In this study, we focused on: (a) the mechanisms by which radiotherapy. We first asked whether tumor cells acquire multimodal immune editing leads to silencing