MYCN-Induced Calcium Signaling Promotes the Malignant Progression of Neuroblastoma
Neuroblastoma (NB) is an extra-cranial solid tumor that occurs mainly in infants and children. Refractory disease (~10% of patients) and relapse of high-risk NB (~40-50% of patients) signifies poor prognosis, and are difficult to treat due to the lack of response to current therapies and aggressive disease progression. Amplification of the MYCN gene is associated with advanced stage, high risk neuroblastoma (NB) and poor prognosis. Novel drugs and alternative treatments are being investigated for patients with advanced NB with MYCN amplification. However, finding an effective treatment strategy for these patients continues to be a major challenge due to aggressive phenotypes of MYCN amplified NB and a myriad of complex mechanisms that promote resistance to treatments and malignant progression. Therefore, my long-term goals are to elucidate the mechanisms that promote the progression of NB, and identify novel targets for the development of more effective chemotherapeutic agents and treatment strategies for advanced stage NB. In order to address this limitation, this project focused on elucidating the role of the transient receptor potential channel melastatin family member 7 (TRPM7) channel and kinase in cell proliferation, apoptosis, migration and invasion of NB tumor cells with different MycN status and different sensitivities to chemotherapeutic drugs. Previous studies and our preliminary data show that MYCN amplification promotes TRPM7 expression, and TRPM7 plays a key role in regulating NB cell migration, invasion and proliferation. In addition to the channel activity of TRPM7, there is also a ser-thr alpha-like kinase domain. The kinase is active when it is attached to the channel, and may also be cleaved from the channel domain and translocate to other areas of the cell where it phosphorylates other target proteins (e.g. nuclear proteins such as histones). The regulation and role of the TRPM7 channel and kinase are dependent on cell type, and varies in NB cells with different MYCN status and with different sensitivities to anti-cancer drugs. This project examines the mechanism by which MycN regulates TRPM7 expression, as well as the role of the channel and kinase of full-length TRPM7 and cleaved TRPM7 in NB progression. The data suggest that MycN regulates TRPM7 expression indirectly through a mechanism that involves intracellular polyamines, as inhibition or down-regulation of polyamine biosynthesis enzymes Ornithine Decarboxylase (ODC1) or Adenosylmethionine Decarboxylase (AMD1) inhibited TRPM7 expression, which was reversed by spermidine. MycN and polyamines also regulated TRPM7 channel activity, increased intracellular Ca2+ signaling, and promoted cell migration in NB cells by promoting the phosphorylation of myosin IIA. In addition, clinical correlation and expression analyses in tumors from NB patients showed that TRPM7 expression correlates with MYCN expression and lower survival probability. Finally, in vivo analysis showed that DFMO decreases tumor volume and inhibits TRPM7 expression. Overall, the data suggest that MycN promotes NB progression through a process that involves polyamine-mediated increase in TRPM7 expression, as well as increased TRPM7 channel and kinase activities.