Targeting Mtor Pathways Using Novel Inhibitors in the Treatment of Glioblastoma

Introduction: Mechanistic Target of Rapamycin (mTOR), which functions via two distinct
multiprotein complexes, mTORC1 and mTORC2, is a downstream signaling kinase in the
PI3K/AKT pathway. mTOR is often deregulated in glioblastoma (GBM), the most aggressive
primary brain tumor in adults, due to the frequent loss of tumor suppressor, PTEN. mTORC1
and mTORC2 control cell growth, proliferation, migration, survival, and stem cell regulation in
response to nutrients and growth factors. mTORC1 regulates protein synthesis through the
downstream substrate, 4EBP1, p70S6K1/2. The substrate of activated AKT, PRAS40 controls
mTORC1, is a surrogate marker of activated AKT after mTOR inhibition therapy in GBM
patients. We aim to study the efficacy of first (Rapamycin (RAPA)), second (Torin1, Torin2,
and XL388), and third (Rapalink-1) mTOR inhibitors to suppress cell growth, dissemination,
overcome drug resistance of GBM cells.

Methods: GBM tumors were evaluated for the expression of pAKTSer473 and pmTORSer2448 via
immunohistochemistry. Effect of Novel mTOR inhibitors was assessed using Western-blotting and
functional assays, including cell proliferation, migration, cell cycle, and drug resistance.

Results: A significant number of tumors expressed pAKTSer473 and pmTORSer2448 . Torin2, but not Torin1 or
XL388, suppressed mTORC1 activity completely as shown by the reduced pS6K Ser235/236 expression in a
dose-dependent manner; Torin1, Torin2, and XL388 inhibited the phosphorylation of 4E-BP1 and
PRAS40; Torin1, Torin2, and XL388 suppressed dephosphorylation of pAKTSer473 . Functional analysis
revealed that RAPA, Torin1, Torin2, XL388, and Rapalink-1 effectively suppressed GBM cell
proliferation and S-phase entry of; RAPA, Torin1, Torin2 and Rapalink-1 inhibited GB cell migration;
drug resistance analysis revealed Torin2 was most effective in eradicating GBM tumor cells.

Conclusion : Results suggested that RAPA, Torin1, Torin2, and Rapalink-1, but not XL388, are useful in
suppressing mTORC1 and mTORC2 activities, thereby inhibiting GBM cell proliferation, dissemination
and drug resistance. Therefore, use of novel mTOR inhibitors present a promising treatment strategy for
Glioblastoma.