Transient receptor potential melastatin 2 (TRPM2)
ion channel has an essential function in modulating cell survival following
oxidant injury and is highly expressed in many
cancers including
neuroblastoma. Here, in xenografts generated from
neuroblastoma cells in which TRPM2 was depleted with CRISPR/Cas9 technology and in in vitro experiments,
tumor growth was significantly inhibited and
doxorubicin sensitivity increased. The
hypoxia-inducible
transcription factor 1/2α (HIF-1/2α) signaling cascade including
proteins involved in
oxidant stress, glycolysis, and mitochondrial function was suppressed by TRPM2 depletion. TRPM2-depleted SH-SY5Y
neuroblastoma cells demonstrated reduced oxygen consumption and
ATP production after
doxorubicin, confirming impaired cellular bioenergetics. In cells in which TRPM2 was depleted, mitochondrial
superoxide production was significantly increased, particularly following
doxorubicin. Ectopic expression of
superoxide dismutase 2 (SOD2) reduced ROS and preserved viability of TRPM2-depleted cells, however, failed to restore
ATP levels. Mitochondrial
reactive oxygen species (ROS) were also significantly increased in cells in which TRPM2 function was inhibited by TRPM2-S, and pretreatment of these cells with the
antioxidant MitoTEMPO significantly reduced ROS levels in response to
doxorubicin and protected cell viability. Expression of the TRPM2 pore mutant E960D, in which
calcium entry through TRPM2 is abolished, also resulted in significantly increased mitochondrial ROS following
doxorubicin treatment, showing the critical role of TRPM2-mediated
calcium entry. These findings demonstrate the important function of TRPM2 in modulation of cell survival through mitochondrial ROS, and the potential of targeted inhibition of TRPM2 as a therapeutic approach to reduce cellular bioenergetics,
tumor growth, and enhance susceptibility to chemotherapeutic agents.