mTORC1 inhibition in pediatric low-grade glioma depletes glutathione and therapeutically synergizes with carboplatin

Abstract

Background

Pediatric low-grade glioma (pLGG) often initially respond to front-line therapies such as carboplatin, but more than 50% of treated tumors eventually progress and require additional therapy. The discovery that pLGG often contain mammalian Target of Rapamycin (mTOR) activation, new treatment modalities and combinations are now possible for patients. The purpose of this study was to determine if carboplatin is synergistic with the mTORC1 inhibitor everolimus in pLGG.

Methods

We treated four pLGG cell lines and one patient-derived xenograft line representing various pLGG genotypes, including NF1 loss, BRAF-KIAA1549 fusion, and BRAF ^V600E mutation, with carboplatin and/or everolimus and performed assays for growth, cell proliferation, and cell death. Immunohistochemistry as well as in vivo and in vitro metabolomics studies were also performed.

Results

Carboplatin synergized with everolimus in all of our four pLGG cell lines (combination index< 1 at Fa 0.5). Combination therapy was superior at inhibiting tumor growth in vivo. Combination treatment increased levels of apoptosis as well as gamma-H2AX phosphorylation compared to either agent alone. Everolimus treatment suppressed the conversion of glutamine and glutamate into glutathione both in vitro and in vivo. Exogenous glutathione reversed the effects of carboplatin and everolimus.

Conclusions

The combination of carboplatin and everolimus was effective at inducing cell death and slowing tumor growth in pLGG models. Everolimus decreased the amount of available glutathione inside the cell, preventing the detoxification of carboplatin and inducing increased DNA damage and apoptosis.