Biochemical, cellular, and in vivo activity of novel ATP-competitive and selective inhibitors of the mammalian target of rapamycin.
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.
Yu
Ker
K
Discovery Oncology, Wyeth Research, Pearl River, New York 10965, USA. yuk@wyeth.com
Toral-Barza
Lourdes
L
Shi
Celine
C
Zhang
Wei-Guo
WG
Lucas
Judy
J
Shor
Boris
B
Kim
Jamie
J
Verheijen
Jeroen
J
Curran
Kevin
K
Malwitz
David J
DJ
Cole
Derek C
DC
Ellingboe
John
J
Ayral-Kaloustian
Semiramis
S
Mansour
Tarek S
TS
Gibbons
James J
JJ
Abraham
Robert T
RT
Nowak
Pawel
P
Zask
Arie
A
D000255:Adenosine Triphosphate / Q000378:metabolism*; D042501:Angiogenic Proteins / Q000037:antagonists & inhibitors; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D001667:Binding, Competitive; D045744:Cell Line, Tumor / Q000378:metabolism; D015536:Down-Regulation; D016193:G1 Phase / Q000187:drug effects; D045325:HCT116 Cells; D019073:HT29 Cells; D006801:Humans; D000076222:Mechanistic Target of Rapamycin Complex 1; D046912:Multiprotein Complexes; D047428:Protein Kinase Inhibitors / Q000378:metabolism / Q000494:pharmacology*; D011494:Protein Kinases; D011506:Proteins; D011687:Purines / Q000378:metabolism / Q000494:pharmacology*; D011720:Pyrazoles / Q000378:metabolism / Q000494:pharmacology*; D011743:Pyrimidines / Q000378:metabolism / Q000494:pharmacology*; D051381:Rats; D020123:Sirolimus / Q000494:pharmacology; D058570:TOR Serine-Threonine Kinases; D014157:Transcription Factors / Q000037:antagonists & inhibitors* / Q000378:metabolism
D042501:Angiogenic Proteins; C493245:CRTC2 protein, human; D046912:Multiprotein Complexes; D047428:Protein Kinase Inhibitors; D011506:Proteins; D011687:Purines; D011720:Pyrazoles; D011743:Pyrimidines; D014157:Transcription Factors; C000614249:WYE-354; D000255:Adenosine Triphosphate; D011494:Protein Kinases; C546842:MTOR protein, human; C546843:mTOR protein, mouse; D000076222:Mechanistic Target of Rapamycin Complex 1; D058570:TOR Serine-Threonine Kinases; D020123:Sirolimus