5-(4-amino-1-propan-2-yl-3-pyrazolo[3-4-d]pyrimidinyl)-1-3-benzoxazol-2-amine and dactolisib

Glioblastomas (GBs) frequently display activation of the epidermal growth factor receptor (EGFR) and mammalian target of rapamycin (mTOR). mTOR exists as part of two multiprotein complexes, mTOR complex 1 (mTORC1) and 2 (mTORC2). In GBs, mTORC1 inhibitors such as rapamycin have performed poorly in clinical trials, and in vitro protect GB cells from nutrient and oxygen deprivation. Next generation ATP-competitive mTOR inhibitors with affinity for both mTOR complexes have been developed, but data exploring their effects on GB metabolism are scarce. In this study, we compared the ATP-competitive mTORC1/2 inhibitors torin2, INK-128 and NVP-Bez235 to the allosteric mTORC1 inhibitor rapamycin under conditions that mimic the glioma microenvironment. In addition to inhibiting mTORC2 signaling, INK-128 and NVP-Bez235 more effectively blocked mTORC1 signaling and prompted a stronger cell growth inhibition, partly by inducing cell cycle arrest. However, under hypoxic and nutrient-poor conditions mTORC1/2 inhibitors displayed even stronger cytoprotective effects than rapamycin by reducing oxygen and glucose consumption. Thus, therapies that arrest proliferation and inhibit anabolic metabolism must be expected to improve energy homeostasis of tumor cells. These results mandate caution when treating physiologically or therapeutically induced hypoxic GBs with mTOR inhibitors.

Topics: Apoptosis; Benzoxazoles; Cell Cycle Checkpoints; Cell Line, Tumor; Cell Proliferation; Glioma; Humans; Imidazoles; Mechanistic Target of Rapamycin Complex 1; Mechanistic Target of Rapamycin Complex 2; Naphthyridines; Protein Kinase Inhibitors; Pyrimidines; Quinolines; Signal Transduction; Sirolimus; TOR Serine-Threonine Kinases

The mammalian target of rapamycin (mTOR) is a serine/threonine kinase that integrates nutrients to execute cell growth. We hypothesized that mTOR is influential in the intervertebral disc-largest avascular, low-nutrient organ. Our objective was to identify the optimal mTOR inhibitor for treating human degenerative disc disease.. mTOR complex 1 (mTORC1) regulates p70/ribosomal S6 kinase (p70/S6K), negatively regulates autophagy, and is controlled by Akt. Akt is controlled by phosphatidylinositol 3-kinase (PI3K) and mTOR complex 2 (mTORC2). mTORC1 inhibitors-rapamycin, temsirolimus, everolimus, and curcumin, mTORC1&mTORC2 inhibitor-INK-128, PI3K&mTOR inhibitor-NVP-BEZ235, and Akt inhibitor-MK-2206-were applied to human disc nucleus pulposus (NP) cells. mTOR signaling, autophagy, apoptosis, senescence, and matrix metabolism were evaluated.. mTORC1 inhibitors decreased p70/S6K but increased Akt phosphorylation, promoted autophagy with light chain 3 (LC3)-II increases and p62/sequestosome 1 (p62/SQSTM1) decreases, and suppressed pro-inflammatory interleukin-1 beta (IL-1β)-induced apoptotic terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) positivity (versus rapamycin, 95% confidence interval (CI) -0.431 to -0.194; temsirolimus, 95% CI -0.529 to -0.292; everolimus, 95% CI -0.477 to -0.241; curcumin, 95% CI -0.248 to -0.011) and poly (ADP-ribose) polymerase (PARP) and caspase-9 cleavage, senescent senescence-associated beta-galactosidase (SA-β-gal) positivity (versus rapamycin, 95% CI -0.437 to -0.230; temsirolimus, 95% CI -0.534 to -0.327; everolimus, 95% CI -0.485 to -0.278; curcumin, 95% CI -0.210 to -0.003) and p16/INK4A expression, and catabolic matrix metalloproteinase (MMP) release and activation. Meanwhile, dual mTOR inhibitors decreased p70/S6K and Akt phosphorylation without enhanced autophagy and suppressed apoptosis, senescence, and matrix catabolism. MK-2206 counteracted protective effects of temsirolimus. Additional disc-tissue analysis found relevance of mTOR signaling to degeneration grades.. mTORC1 inhibitors-notably temsirolimus with an improved water solubility-but not dual mTOR inhibitors protect against inflammation-induced apoptosis, senescence, and matrix catabolism in human disc cells, which depends on Akt and autophagy induction.

Topics: Adult; Aged; Aged, 80 and over; Apoptosis; Autophagy; Benzoxazoles; beta-Galactosidase; Cellular Senescence; Curcumin; Everolimus; Extracellular Matrix; Female; Heterocyclic Compounds, 3-Ring; Humans; Imidazoles; Inflammation; Male; Matrix Metalloproteinases; Mechanistic Target of Rapamycin Complex 1; Mechanistic Target of Rapamycin Complex 2; Microtubule-Associated Proteins; Middle Aged; Nucleus Pulposus; Protein Kinase Inhibitors; Proto-Oncogene Proteins c-akt; Pyrimidines; Quinolines; Ribosomal Protein S6 Kinases, 70-kDa; Sequestosome-1 Protein; Sirolimus