Int J Biol Sci 2022; 18(7):2684-2702. doi:10.7150/ijbs.71041 This issue

Research Paper

Toosendanin, a novel potent vacuolar-type H+-translocating ATPase inhibitor, sensitizes cancer cells to chemotherapy by blocking protective autophagy

Yu Dong1,2#, Guoyuan Zhu3#, Sheng-Fang Wang1, Kristine A. Keon4,5, John L. Rubinstein4,5,6, Si-Xin Zeng1, Shuang Zhang1, Qiu-Ling Chen1, Jing Fu3, Min Li7, Han-Ming Shen8, Jin-Jian Lu1,2, Xiu-Ping Chen1, Jia-Hong Lu1,2✉

1. State Key Laboratory of Quality Research in Chinese Medicine, Institute of Chinese Medical Sciences, University of Macau, Macau SAR, China.
2. Guangdong-Hong Kong-Macau Joint Lab on Chinese Medicine and Immune Disease Research, University of Macau, Macau SAR, China.
3. State Key Laboratory of Quality Research in Chinese Medicine, Guangdong-Hong Kong-Macao Joint Laboratory of Respiratory Infectious Disease, Macau Institute for Applied Research in Medicine and Health, Macau University of Science and Technology, Macau SAR, China.
4. Molecular Medicine Program, The Hospital for Sick Children, 686 Bay Street, Toronto, Ontario, Canada M5G0A4.
5. Department of Medical Biophysics, The University of Toronto, 101 College Street Suite 15-701, Toronto, Ontario Canada M5G 1L7.
6. Department of Biochemistry, The University of Toronto, 1 King's College Circle, Toronto Ontario, Canada M5S 1A8.
7. Mr. & Mrs. Ko Chi-Ming Centre for Parkinson's Disease Research, School of Chinese Medicine, Hong Kong Baptist University, Hong Kong SAR, China.
8. Faculty of Health Sciences, University of Macau, Macau, China.
#These authors contributed equally to this work

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Citation:
Dong Y, Zhu G, Wang SF, Keon KA, Rubinstein JL, Zeng SX, Zhang S, Chen QL, Fu J, Li M, Shen HM, Lu JJ, Chen XP, Lu JH. Toosendanin, a novel potent vacuolar-type H+-translocating ATPase inhibitor, sensitizes cancer cells to chemotherapy by blocking protective autophagy. Int J Biol Sci 2022; 18(7):2684-2702. doi:10.7150/ijbs.71041. Available from https://www.ijbs.com/v18p2684.htm

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Abstract

Graphic abstract

Macroautophagy/autophagy is the process of self-digestion through the lysosomes; it disassembles unnecessary or dysfunctional long-lived proteins and damaged organelles for the recycling of biomacromolecules. Unfortunately, cancer cells can hijack this mechanism to survive under metabolic stress or develop drug resistance during chemotherapy. Increasing evidence indicates that the combination of autophagy inhibition and chemotherapy is a promising cancer treatment strategy. However, effective autophagy inhibitors with satisfied potency, bioavailability, and clearly-defined drug targets are still rare. Here, we report the identification of a potent autophagy inhibitor toosendanin which can effectively block autophagosome maturation, causing the accumulation of autophagy substrates in multiple cancer cells. Toosendanin did not inhibit the fusion process between autophagosome and lysosome but elevated lysosomal pH and impaired lysosomal enzymes activity. Using rat liver lysosome fraction and purified yeast V-ATPase, we found that toosendanin directly inhibited V-ATPase activity. By applying cellular thermal shift assay (CETSA), immunoprecipitation-coupled LC-MS/MS analysis, and biotin-toosendanin pull-down assay, we confirmed the direct binding between toosendanin and V-ATPase. Furthermore, toosendanin blocked chemotherapy-induced protective autophagy in cultured cancer cells and xenograft tumor tissues to significantly enhance anti-cancer activity. These results suggest that toosendanin has the potential to be developed into an anti-cancer drug by blocking chemotherapy-induced protective autophagy.

Keywords: toosendanin, V-ATPase inhibitor, autophagy inhibitor, protective autophagy, anti-cancer effect