The mTOR Kinase Inhibitors, CC214-1 and CC214-2, Preferentially Block the Growth of EGFRvIII-Activated Glioblastomas

Abstract

Purpose mTOR pathway hyperactivation occurs in nearly 90% of glioblastomas, but the allosteric mTOR inhibitor rapamycin has failed in the clinic. Here we examine the efficacy of the newly discovered ATP-competitive mTOR kinase inhibitors CC214-1 and CC214-2 in glioblastoma, identifying molecular determinants of response and mechanisms of resistance, and develop a pharmacological strategy to overcome it. Experimental design We performed in vitro and in vivo studies in glioblastoma cell lines and an intracranial model to: determine the potential efficacy of the recently reported mTOR kinase inhibitors CC214-1 (in vitro use) and CC214-2 (in vivo use) at inhibiting rapamycin resistant signaling and blocking GBM growth and a novel single cell technology, DNA Encoded Antibody Libraries, was used to identify mechanisms of resistance. Results Here we demonstrate that CC214-1 and CC214-2 suppress rapamycin-resistant mTORC1 signaling; block mTORC2 signaling and significantly inhibit the growth of glioblastomas in vitro and in vivo. EGFRvIII expression and PTEN loss enhance sensitivity to CC214 compounds, consistent with enhanced efficacy in strongly mTOR-activated tumors. Importantly, CC214 compounds potently induce autophagy, preventing tumor cell death. Genetic or pharmacologic inhibition of autophagy greatly sensitizes GBM cells and orthotopic xenografts to CC214-1 and CC214-2 induced cell death. Conclusions These results identify CC214-1 and CC214-2 as potentially efficacious mTOR kinase inhibitors in GBM and suggest a strategy for identifying patients most likely to benefit from mTOR inhibition. This study also demonstrates a central role for autophagy in preventing mTOR-kinase inhibitor-mediated tumor cell death, and suggests a pharmacological strategy for overcoming it.

Dual mTORC1/2 Blockade Inhibits Glioblastoma Brain Tumor Initiating Cells In Vitro and In Vivo and Synergizes with Temozolomide to Increase Orthotopic Xenograft Survival

“…In agreement with our findings, the dual mTORC1/2 inhibitors CC214-1 and CC214-2 have been recently reported to effectively inhibit growth of GBM cell lines and xenografts resistant to mTORC1 blockade with rapamycin (26). These results support the idea that inhibition of mTORC2 must be achieved for effective blockade of PI3K signaling (Supplementary Fig.…”

“…Although Gini and colleagues (26) reported that ectopic expression of the EGFRvIII mutation or PTEN deficiency was necessary to sensitize U87 cells to dual mTORC1/2 inhibition with CC214-2, we found that all BTICs we tested were sensitive to AZD regardless of their underlying endogenous EGFR and PTEN mutational status. Therefore, mTORC1/2 signaling may be a key signaling hub, common to the biology of molecularly diverse GBMs, and provides support for clinical use of mTORC1/2 inhibitors.…”

Compensatory glutamine metabolism promotes glioblastoma resistance to mTOR inhibitor treatment

“…Using gas chromatography-mass spectroscopy (GC/MS) of U87 and U87/EGFRvIII cells treated with mTOR inhibitors (rapamycin or PP242) for 48 hours, we identified 91 metabolites whose levels significantly changed in response to the allosteric mTOR inhibitor rapamycin or the ATPcompetitive mTOR inhibitor PP242 ( Figure 2 and Supplemental Table 1). We have previously shown that rapamycin has minimal activity against mTORC2 signaling in GBM cell lines in in vivo models and patients treated with the drug, whereas PP242 blocks both mTORC1 and mTORC2 signaling in GBM cells (12,13,22). The principal component analysis (PCA) of variation in the metabolites for each treatment group demonstrated distinct clustering or a clear separation of the 3 groups (Supplemental Figure 2, A and B).…”

Mechanistic Target of Rapamycin (mTOR) Inhibition Synergizes with Reduced Internal Ribosome Entry Site (IRES)-mediated Translation of Cyclin D1 and c-MYC mRNAs to Treat Glioblastoma

“…As a consequence of EGFR amplification or activating mutation and phosphatase and tensin homolog (PTEN) loss (3,4), hyperactivation of the PI3K pathway is seen in nearly 90% of all GBMs (5,6). As a result, the mechanistic target of rapamycin (mTOR) kinases, a downstream effector, are often constantly hyperactivated (7). mTOR is a central regulator of mRNA translation, metabolism, and autophagy in the cell and, consequently, controls tumor cell growth, survival, and drug resistance (8,9).…”