Preclinical analysis of MTOR complex 1/2 inhibition in diffuse intrinsic pontine glioma

Abstract

Abstract. Diffuse intrinsic pontine glioma (DIPG) is an incurable childhood brain tumor. The mechanistic target of rapamycin (MTOR), a key oncogene, functions as two distinct signaling complexes, MTORC1 and MTORC2. We set out to determine the preclinical efficacy and mechanism of action of MTOR inhibitors in DIPG. We evaluated the MTORC1 inhibitor everolimus and the MTORC1/2 inhibitor AZD2014 in three patient-derived DIPG cell lines using cell culture models. We created dose-response curves for both compounds. We measured phenotypic effects on cell selfrenewal, apoptosis, cell cycle, differentiation, senescence, and autophagy. We assessed the effects of each compound on the AKT pathway. Finally, we measured the efficacy of AZD2014 in combination with radiation therapy (RT) and a panel of FDA-approved chemotherapy drugs. While everolimus showed minimal antitumor efficacy, AZD2014 revealed IC 50 levels of 410-552 nM and IC 90 levels of 1.30-8.86 µM in the three cell lines. AZD2014 demonstrated increased inhibition of cell self-renewal compared to everolimus. AZD2014 decreased expression of phospho-AKT, while no such effect was noted with everolimus. Direct AKT inhibition showed similar efficacy to AZD2014, and induction of constitutive AKT activity rescued DIPG cells from the effects of AZD2014. AZD2014 exhibited synergistic relationships with both RT and various chemotherapy agents across classes, including the multikinase inhibitor ponatinib. MTORC1/2 inhibition shows antitumor activity in cell culture models of DIPG due to the effect of MTORC2 inhibition on AKT. This strategy should be further assessed for potential incorporation into combinatorial approaches to the treatment of DIPG. IntroductionDiffuse intrinsic pontine glioma (DIPG) is an incurable brain tumor that mostly affects young children (1). Treatment with radiation therapy (RT) is usually transiently effective, but median progression-free survival is only 7 months, and median overall survival is 11 months (2). Hundreds of clinical trials studying systemically delivered cytotoxic and targeted chemotherapy agents have been conducted without showing any clinical benefit, potentially due to inadequate tumor penetration (3). Investigators are therefore studying novel drug delivery methods for these patients (4,5), and some of these have reached clinical trials (6,7). As these studies advance, better targeted therapeutic options will be needed, especially as part of combination treatments.The mechanistic target of rapamycin (MTOR) is a serinethreonine kinase that has crucial roles in many cellular pathways dysregulated in cancer, including metabolism, growth, survival, and response to stress. It functions as part of two distinct signaling complexes, MTOR complex 1 (MTORC1), which also contains the regulatory associated protein of MTORC1 (RPTOR), and MTOR complex 2 (MTORC2), containing the RPTOR independent companion of MTORC2 (RICTOR). MTORC2 activates the V-Akt murine thymoma viral oncogene (AKT) through phosphorylation at S473, which mediates many of th...

Cell senescence in neuropathology: A focus on neurodegeneration and tumours

“…shown to result in a strong synergistic antitumour activity preclinically [197], suggesting that the use of senolytics or SASP modulators could be of therapeutic relevance. Likewise, a recent study has proposed a new model where senescence is induced in DMG tumour cells by inhibition of BMI1.…”

Recent Advances in Understanding the Role of Autophagy in Paediatric Brain Tumours

“…Tsoli et al have observed a significant increase in cytotoxic activity, mitochondrial dysfunction, and apoptotic cell death on patient-derived DIPG cells following treatment with dual pharmacological inhibition of adenine-nucleotide translocase (ANT) mitochondrial protein (using PENAO) and mTOR (using temsirolimus) [ 118 ]. Similarly, Asby et al demonstrated an anti-proliferative effect and a decrease in cell-viability after administration of temsirolimus, both alone and together with a CDK 4/6 inhibitor [ 119 ]. Another study, conducted by Flannery et al, evaluated the inhibition of mTORC1, both alone (everolimus) and together with mTORC2 inhibition (AZD2014) on DIPG cell lines, observing that the inhibition of both complexes via AZD2014 works better than everolimus in the elicited antitumour activity [ 120 ].…”

Dodging the bullet: therapeutic resistance mechanisms in pediatric cancers

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