ABSTRACT
Mechanistic target of rapamycin (mTOR) forms two distinct complexes, mTOR complex 1 (mTORC1) and mTORC2. Here we investigated the antitumor effect of dual mTORC1/2 inhibitor AZD2014 on epithelial ovarian cancer (EOC) and its potential effect on immunosuppressive myeloid-derived suppressor cells (MDSCs). Immunohistochemical analysis of mTORC1 and mTORC2 was performed on a human ovarian cancer tissue microarray. High mTORC2 expression level was associated with shorter survival in EOC, whereas mTORC1 was not correlate with patients' prognosis. AZD2014 suppressed mTOR signaling pathway in ovarian cancer cells, inhibited proliferation and induced G1-phase cell cycle arrest and apoptosis. In tumor-bearing mice, AZD2014 treatment limited tumor growth, reduced peritoneal ascites, and prolonged survival. AZD2014 specifically reduced MDSCs migration and accumulation in EOC peritoneal fluid but not in the spleen. Moreover, subsequent AZD2014 treatment after cisplatin chemotherapy delayed EOC recurrence. Collectively, we observed that high mTORC2 expression level in EOC indicated a poor prognosis. Remarkably, in tumor-bearing mice, AZD2014 diminished MDSC accumulation and delayed tumor growth and recurrence.
Subject(s)
Benzamides/pharmacology , Carcinoma, Ovarian Epithelial/drug therapy , Mechanistic Target of Rapamycin Complex 1/antagonists & inhibitors , Mechanistic Target of Rapamycin Complex 2/antagonists & inhibitors , Morpholines/pharmacology , Myeloid-Derived Suppressor Cells/drug effects , Ovarian Neoplasms/drug therapy , Pyrimidines/pharmacology , Animals , Apoptosis , Benzamides/adverse effects , Benzamides/therapeutic use , Carcinoma, Ovarian Epithelial/mortality , Carcinoma, Ovarian Epithelial/pathology , Cell Line, Tumor , Female , Humans , Mechanistic Target of Rapamycin Complex 1/analysis , Mechanistic Target of Rapamycin Complex 2/analysis , Mice , Mice, Inbred C57BL , Morpholines/adverse effects , Morpholines/therapeutic use , Myeloid-Derived Suppressor Cells/physiology , Ovarian Neoplasms/mortality , Ovarian Neoplasms/pathology , Pyrimidines/adverse effects , Pyrimidines/therapeutic use , TOR Serine-Threonine Kinases/physiology , Xenograft Model Antitumor AssaysABSTRACT
The evolutionarily conserved Target of Rapamycin (TOR) complex-2 (TORC2) is an essential regulator of plasma membrane homeostasis in budding yeast (Saccharomyces cerevisiae). In this yeast, TORC2 phosphorylates and activates the effector protein kinase Ypk1 and its paralog Ypk2. These protein kinases, in turn, carry out all the crucial functions of TORC2 by phosphorylating and thereby controlling the activity of at least a dozen downstream substrates. A previously uncharacterized interplay between the Rab5 GTPases and TORC2 signaling was uncovered through analysis of a newly suspected Ypk1 target. Muk1, one of two guanine nucleotide exchange factors for the Rab5 GTPases, was found to be a physiologically relevant Ypk1 substrate; and, genetic analysis indicates that Ypk1-mediated phosphorylation activates the guanine nucleotide exchange activity of Muk1. Second, it was demonstrated both in vivo and in vitro that the GTP-bound state of the Rab5 GTPase Vps21/Ypt51 physically associates with TORC2 and acts as a direct positive effector required for full TORC2 activity. These interrelationships provide a self-reinforcing control circuit for sustained up-regulation of TORC2-Ypk1 signaling. In this overview, we summarize the experimental basis of these findings, their implications, and speculate as to the molecular basis for Rab5-mediated TORC2 activation.
Subject(s)
Mechanistic Target of Rapamycin Complex 2/metabolism , Saccharomyces cerevisiae/metabolism , rab5 GTP-Binding Proteins/physiology , Glycogen Synthase Kinase 3/metabolism , Guanine Nucleotide Exchange Factors/metabolism , Guanine Nucleotide Exchange Factors/physiology , Humans , Mechanistic Target of Rapamycin Complex 2/analysis , Models, Molecular , Saccharomyces cerevisiae/genetics , Saccharomyces cerevisiae Proteins/metabolism , Saccharomyces cerevisiae Proteins/physiology , Signal Transduction , Up-Regulation , rab5 GTP-Binding Proteins/analysis , rab5 GTP-Binding Proteins/metabolismABSTRACT
Lymphangioleiomyomatosis (LAM) is a rare progressive cystic lung disease with features of a low-grade neoplasm. It is primarily caused by mutations in TSC1 or TSC2 genes. Sirolimus, an inhibitor of mTOR complex 1 (mTORC1), slows down disease progression in some, but not all patients. Hitherto, other potential therapeutic targets such as mTOR complex 2 (mTORC2) and various metabolic pathways have not been investigated in human LAM tissues. The aim of this study was to assess activities of mTORC1, mTORC2 and various metabolic pathways in human LAM tissues through analysis of protein expression. Immunohistochemical analysis of p-S6 (mTORC1 downstream protein), Rictor (mTORC2 scaffold protein) as well as GLUT1, GAPDH, ATPB, GLS, MCT1, ACSS2 and CPT1A (metabolic pathway markers) were performed on lung tissue from 11 patients with sporadic LAM. Immunoreactivity was assessed in LAM cells with bronchial smooth muscle cells as controls. Expression of p-S6, Rictor, GAPDH, GLS, MCT1, ACSS2 and CPT1A was significantly higher in LAM cells than in bronchial smooth muscle cells (P<.01). No significant differences were found between LAM cells and normal bronchial smooth muscle cells in GLUT1 and ATPB expression. The results are uniquely derived from human tissue and indicate that, in addition to mTORC1, mTORC2 may also play an important role in the pathobiology of LAM. Furthermore, glutaminolysis, acetate utilization and fatty acid ß-oxidation appear to be the preferred bioenergetic pathways in LAM cells. mTORC2 and these preferred bioenergetic pathways appear worthy of further study as they may represent possible therapeutic targets in the treatment of LAM.
