The
mammalian target of rapamycin (mTOR) is centrally involved in cell growth, metabolism, and angiogenesis. While showing clinical efficacy in a subset of
tumors,
rapamycin and
rapalogs are specific and allosteric inhibitors of mTOR complex 1 (
mTORC1), but they do not directly inhibit mTOR complex 2 (
mTORC2), an emerging player in
cancer. Here, we report chemical structure and biological characterization of three pyrazolopyrimidine
ATP-competitive
mTOR inhibitors, WAY-600,
WYE-687, and
WYE-354 (IC(50), 5-9 nmol/L), with significant selectivity over
phosphatidylinositol 3-kinase (PI3K) isofoms (>100-fold). Unlike the
rapalogs, these inhibitors acutely blocked substrate phosphorylation by
mTORC1 and
mTORC2 in vitro and in cells in response to
growth factor,
amino acids, and hyperactive PI3K/AKT. Unlike the inhibitors of PI3K or dual-pan PI3K/mTOR, cellular inhibition of P-S6K1(T389) and P-AKT(S473) by the pyrazolopyrimidines occurred at significantly lower inhibitor concentrations than those of P-AKT(T308) (PI3K-PDK1 readout), showing mTOR selectivity in cellular setting. mTOR
kinase inhibitors reduced AKT downstream function and inhibited proliferation of diverse
cancer cell lines. These effects correlated with a strong G(1) cell cycle arrest in both the
rapamycin-sensitive and
rapamycin-resistant cells, selective induction of apoptosis, repression of global
protein synthesis, and down-regulation of angiogenic factors. When injected into
tumor-bearing mice,
WYE-354 inhibited
mTORC1 and
mTORC2 and displayed robust antitumor activity in PTEN-null
tumors. Together, our results highlight mechanistic differentiation between
rapalogs and mTOR
kinase inhibitors in targeting
cancer cell growth and survival and provide support for clinical development of mTOR
kinase inhibitors as new
cancer therapy.