Effects of TORC1/2 inhibitor MLN0128 alone and in combination with MEK inhibition in BRAF mutated glioma cells.


April 21, 2015
Mueller S, Dik R, Yang X, Aleksandra O, Gustafson C, DuBois S, Weiss W, Waanders A, Resnick A, Haas-Kogan D


INTRODUCTION: First generation allosteric inhibitors of mTORC1 have shown clinical benefit for the treatment of pediatric low-grade gliomas (PLGG). MEK inhibition has shown great promise in early phase clinical trials for PLGG with the hypothesis that tumors carrying the KIAA1549:BRAF are particularly sensitive to such treatment. MLN0128, a second-generation, ATP-competitive, pan-mTOR kinase inhibitor, acts on both mTORC1 and mTORC2. Herein we investigate the effects of MLN0128 monotherapy as well as in combination with MEK inhibition in models of PLGG. METHODS: We used human glioma cell lines containing BRAFV600E (AM38), wild-type BRAF (LN229, TN98, SF188) and isogenic systems of KIAA1549:BRAF-expressing NIH3T3 cells. Signaling inhibitors included MLN0128, everolimus, BRAFV600E specific inhibitor PLX4720 and MEK specific inhibitors AZD6244 and GSK1120212. Cell proliferation was determined using an ATP-based assay. Biochemical effects were assessed using western blot analysis. In vivo activity of these inhibitors are ongoing using the BT40 PLGG xenograft mouse model. RESULTS: MLN0128 monotherapy shows stronger anti-proliferative effects compared to everolimus across cell lines independent of BRAF status. As expected, treatment with MLN0128 leads to down-regulation of p-AKT and p-4EBP1, whereas treatment with everolimus only reduces expression of p-S6. MEK inhibition with AZD6244 effectively decreases p-ERK expression in all lines; however, the addition of MLN0128 to AZD6244 causes a rebound in p-ERK expression in BRAFWT gliomas but not in gliomas expression BRAFV600E or KIAA1549:BRAF alterations. In cells carrying either the BRAFV600Eor KIAA1549:BRAF alteration, combined treatment with MLN0128 and GSK1120212 or AZD6244 leads to synergistic anti-proliferative effects whereas such synergy is not present in BRAFWT gliomas. In vivo studies in the PLGG xenograft model BT40 are ongoing. CONCLUSION: MLN0128 has synergistic effects with MEK inhibition in gliomas bearing BRAFV600E or KIAA1549:BRAF. In BRAF wild-type cells such synergy is not present, perhaps due to rebound in p-ERK when MLN0128 is combined with AZD6244.