Blood glutamate scavengers increase pro-apoptotic signaling and reduce metastatic melanoma growth in-vivo

Yona Goldshmit,Evgeni Banyas,Moshe Shalom,Angela Ruban,Alex Yakovchuk,Pablo Blinder,Amit Benbenishty,Rita Perelroizen,Lior Mayo,Sari David

doi:10.1038/s41598-021-94183-8

Abstract

Inhibition of extracellular glutamate (Glu) release decreases proliferation and invasion, induces apoptosis, and inhibits melanoma metastatic abilities. Previous studies have shown that Blood-glutamate scavenging (BGS), a novel treatment approach, has been found to be beneficial in attenuating glioblastoma progression by reducing brain Glu levels. Therefore, in this study we evaluated the ability of BGS treatment to inhibit brain metastatic melanoma progression in-vivo. RET melanoma cells were implanted in C56BL/6J mice to induce brain melanoma tumors followed by treatment with BGS or vehicle administered for fourteen days. Bioluminescent imaging was conducted to evaluate tumor growth, and plasma/CSF Glu levels were monitored throughout. Immunofluorescence staining of Ki67 and 53BP1 was used to analyze tumor cell proliferation and DNA double-strand breaks. In addition, we analyzed CD8, CD68, CD206, p-STAT1 and iNOS expression to evaluate alterations in tumor micro-environment and anti-tumor immune response due to treatment. Our results show that BGS treatment reduces CSF Glu concentration and consequently melanoma growth in-vivo by decreasing tumor cell proliferation and increasing pro-apoptotic signaling in C56BL/6J mice. Furthermore, BGS treatment supported CD8+ cell recruitment and CD68+ macrophage invasion. These findings suggest that BGS can be of potential therapeutic relevance in the treatment of metastatic melanoma.

Highlights

Inhibition of extracellular glutamate (Glu) release decreases proliferation and invasion, induces apoptosis, and inhibits melanoma metastatic abilities
We examined the anti-tumor effects of such a Bloodglutamate scavenging (BGS) treatment in a brain metastatic melanoma mice model
The main findings of this study were: (1) Glu concentrations in the CSF consistently increased along with tumor growth, and was significantly higher in implanted animals as compared to healthy mice; (2) The BGS treatment significantly decreased plasma and CSF Glu concentrations; (3) The BGS treatment reduced melanoma growth in vivo by decreasing tumor cell proliferation and by increasing pro-apoptotic signaling; (4) The BGS treatment mediated CD8+ T cell recruitment into the tumor; and (5) The BGS treatment enhanced the pro-inflammatory response by decreasing CD206 positive cells, and mediated phagocytic CD68+ macrophage invasion into the tumor

Summary

Introduction

Inhibition of extracellular glutamate (Glu) release decreases proliferation and invasion, induces apoptosis, and inhibits melanoma metastatic abilities. The abundance of research suggesting that extracellular Glu accumulation is involved in glioblastoma progression has prompted us to hypothesize the existence of similar mechanisms in other brain tumors, such as brain melanoma metastases Supporting this hypothesis, it has been shown that the presence of ectopic metabotropic glutamate receptor 1 (mGluR1) in murine melanocytes results in the formation of melanoma. It is possible that more advanced tumors with stage III and IV metastases have a higher frequency of mGluR1 expression and of ectopic mGluR119–21 In such cases, the reduction of extracellular brain glutamate could be effective in the management of brain metastatic melanomas. This unique ability of Glu, to induce divergent effects on activated T cells in a concentration-dependent manner, may be important in ongoing anti-tumor immune responses

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Results

Conclusion