Disruption of GRM1‐mediated signalling using riluzole results in DNA damage in melanoma cells

Abstract

Summary Gain-of-function of the neuronal receptor, metabotropic glutamate receptor 1 (Grm1), was sufficient to induce melanocytic transformation in vitro and spontaneous melanoma development in vivo when ectopically expressed in melanocytes. The human form of this receptor, GRM1, has been shown to be ectopically expressed in a subset of human melanomas but not benign nevi or normal melanocytes, suggesting that misregulation of GRM1 is involved in the pathogenesis of certain human melanomas. Sustained stimulation of Grm1 by the ligand, glutamate, is required for the maintenance of transformed phenotypes in vitro and tumorigenicity in vivo. In this study we investigate the mechanism of an inhibitor of glutamate release, riluzole, on human melanoma cells that express metabotropic glutamate receptor 1 (GRM1). Various in vitro assays conducted show that inhibition of glutamate release in several human melanoma cell lines resulted in an increase of oxidative stress and DNA damage response markers.

Metabotropic glutamate receptors in cancer

“…53BP1, a checkpoint regulator that binds to p53 and translocates to sites of DNA breaks in response to DNA damage (Wall et al, 2015). Similar results were observed in isogenic vector control or mGluR1 clones derived from immortalized, non-tumorigenic human melanocytes (Wall et al, 2014). …”

“…In particular, mGluRs have been shown empirically to be the predominant mediators of glutamatergic signaling in many cancers (Khan et al, 2011; Koochekpour, 2013; Martino et al, 2013; Speyer et al, 2014; Teh and Chen, 2012; Zhang et al, 2015). The mechanisms by which mGluRs modulate peripheral cell transformation and tumor growth are postulated to be either ectopic expression of wild type mGluRs, increased proliferative signals arising from receptor overexpression, mutations, or expression of polymorphic variants (Ali et al, 2014; Brocke et al, 2010; Mehta et al, 2013; Namkoong et al, 2007; Wall et al, 2014; Zhang et al, 2015). …”

“…Further research led to the unexpected discovery that riluzole by itself led to DNA damage and that riluzole in association with irradiation produced additive DNA double-strand breaks in mGluR1-postive human melanoma cells in vitro and in vivo, as evident by the histological detection of γ-H2AX foci and phosphorylation of histone H2AX on Western immunoblots (Wall et al, 2014). These findings only manifest in mGluR1-positive human melanoma cells but not mGluR1-negative cells.…”

“…Using a genetic inducible siRNA (si-GRM1) approach in C8161 human melanoma cells, DNA damage was reduced in riluzole-treated induced si-GRM1 clones, compared to the same clones not subject to the induction of si-GRM1 (Wall et al, 2014). These results support mGluR1-dependent riluzole-mediated DNA damage as the mode of action.…”

“…These results support mGluR1-dependent riluzole-mediated DNA damage as the mode of action. In the same report, we proposed that in the presence of riluzole, synthesis of the antioxidant glutathione (GSH), may be interrupted by increased intracellular glutamate that could disrupt the bidirectional transport of glutamate and cysteine required for GSH synthesis (Wall et al, 2014). A decrease in GSH causes increased oxidative stress and reactive oxygen species (ROS) detectible by the ROS-specific marker dihydrorhodamine 123 (DHR123).…”