Transient receptor potential ion channel TRPM2 promotes AML proliferation and survival through modulation of mitochondrial function, ROS, and autophagy

Shu-Jen Chen,Barbara A Miller,Lei Bao,Kerry Keefer,Iwona Hirschler-Laszkiewicz,Salim Merali,John Lee,Muniswamy Madesh,Santhanam Shanmughapriya,Longgui Chen,Xue-Qian Zhang,Sinisa Dovat,Hong-Gang Wang,Joseph Y Cheung,Jufang Wang,Yuka Imamura,Carmen Merali

doi:10.1038/s41419-020-2454-8

Abstract

Transient receptor potential melastatin 2 (TRPM2) ion channel has an essential function in maintaining cell survival following oxidant injury. Here, we show that TRPM2 is highly expressed in acute myeloid leukemia (AML). The role of TRPM2 in AML was studied following depletion with CRISPR/Cas9 technology in U937 cells. In in vitro experiments and in xenografts, depletion of TRPM2 in AML inhibited leukemia proliferation, and doxorubicin sensitivity was increased. Mitochondrial function including oxygen consumption rate and ATP production was reduced, impairing cellular bioenergetics. Mitochondrial membrane potential and mitochondrial calcium uptake were significantly decreased in depleted cells. Mitochondrial reactive oxygen species (ROS) were significantly increased, and Nrf2 was decreased, reducing the antioxidant response. In TRPM2-depleted cells, ULK1, Atg7, and Atg5 protein levels were decreased, leading to autophagy inhibition. Consistently, ATF4 and CREB, two master transcription factors for autophagosome biogenesis, were reduced in TRPM2-depleted cells. In addition, Atg13 and FIP200, which are known to stabilize ULK1 protein, were decreased. Reconstitution with TRPM2 fully restored proliferation, viability, and autophagy; ATF4 and CREB fully restored proliferation and viability but only partially restored autophagy. TRPM2 expression reduced the elevated ROS found in depleted cells. These data show that TRPM2 has an important role in AML proliferation and survival through regulation of key transcription factors and target genes involved in mitochondrial function, bioenergetics, the antioxidant response, and autophagy. Targeting TRPM2 may represent a novel therapeutic approach to inhibit myeloid leukemia growth and enhance susceptibility to chemotherapeutic agents through multiple pathways.

Highlights

Increased reactive oxygen species (ROS) are found in acute myeloid leukemia (AML)[1,2]
Major findings are as follows: (1) Transient receptor potential melastatin 2 (TRPM2) is highly expressed in AML and depletion of TRPM2 inhibits leukemia proliferation and survival in vitro and in xenografts; (2) mitochondrial function and bioenergetics are reduced and mitochondrial ROS levels elevated in TRPM2-depleted leukemia cells; (3) multiple transcription factors including CREB, activating transcription factor 4 (ATF4), and Nrf[2] are reduced in TRPM2 depletion, which contributes to increased ROS; and (4) autophagy is impaired through modulation of transcription factors CREB and ATF4, which are master transcription factors for autophagosome biogenesis, resulting in decreased ULK1, Atg[7], and Atg[5] and autophagocytic flux. These findings demonstrate that inhibition of TRPM2 reduces leukemia growth and increases cytotoxicity in myeloid leukemia through mitochondrial dysfunction, increased mitochondrial ROS production, reduced antioxidant response and bioenergetics, and significantly impaired autophagy
Expression of TRPM2 in AML samples from patients with normal karyotypes and all major AML mutational subgroups including aberrant complex karyotypes was significantly greater than that measured in normal hematopoietic stem cells (HSC) or common myeloid progenitors (CMP) (Fig. 1b)

Summary

Introduction

Increased reactive oxygen species (ROS) are found in acute myeloid leukemia (AML)[1,2]. Mitochondria are a may promote proliferation and metastasis by aberrantly affecting proliferative or survival pathways, whereas an excessive increase results in cell death[4]. Use of pro-oxidants or inhibition of intracellular antioxidants to increase ROS above the cytotoxic threshold has been proposed as a novel approach to optimize anti-cancer drugs[4,8,9]. Myeloid leukemia stem cell have increased sensitivity to ROS, which could be utilized in their eradication[10]. Melastatin subfamily (TRPM) members have important roles in cell proliferation and survival[12]

Methods

Results

Conclusion