Inhibition of RAS function through targeting an allosteric regulatory site.
RAS GTPases are important mediators of oncogenesis in humans. However, pharmacological inhibition of RAS has proved challenging. Here we describe a functionally critical region, located outside the effector lobe of RAS, that can be targeted for inhibition. We developed NS1, a synthetic binding protein (monobody) that bound with high affinity to both GTP- and GDP-bound states of H-RAS and K-RAS but not N-RAS. NS1 potently inhibited growth factor signaling and oncogenic H-RAS- and K-RAS-mediated signaling and transformation but did not block oncogenic N-RAS, BRAF or MEK1. NS1 bound the α4-β6-α5 region of RAS, which disrupted RAS dimerization and nanoclustering and led to blocking of CRAF-BRAF heterodimerization and activation. These results establish the importance of the α4-β6-α5 interface in RAS-mediated signaling and define a previously unrecognized site in RAS for inhibiting RAS function.
Department of Pharmacology, University of Illinois at Chicago, Chicago, Illinois, USA.|University of Illinois Cancer Center, University of Illinois at Chicago, Chicago, Illinois, USA.|Jesse Brown VA Medical Center, Chicago, Illinois, USA.
Russell
R
Spencer-Smith
Department of Biochemistry and Molecular Biology, University of Chicago, Chicago, Illinois, USA.|Perlmutter Cancer Center, New York University Langone Medical Center, New York, New York, USA.|Department of Medicine, New York University Langone Medical Center, New York, New York, USA.
Akiko
A
Koide
Department of Integrative Biology and Pharmacology, University of Texas Medical School at Houston, Houston, Texas, USA.
Yong
Y
Zhou
Department of Biochemistry and Molecular Biology, University of Chicago, Chicago, Illinois, USA.
Raphael R
RR
Eguchi
Department of Biochemistry and Molecular Biology, University of Chicago, Chicago, Illinois, USA.
Fern
F
Sha
Department of Pharmacology, University of Illinois at Chicago, Chicago, Illinois, USA.|University of Illinois Cancer Center, University of Illinois at Chicago, Chicago, Illinois, USA.
Priyanka
P
Gajwani
Department of Pharmacology, University of Illinois at Chicago, Chicago, Illinois, USA.|University of Illinois Cancer Center, University of Illinois at Chicago, Chicago, Illinois, USA.
Dianicha
D
Santana
Department of Biochemistry and Molecular Biology, University of Chicago, Chicago, Illinois, USA.|Perlmutter Cancer Center, New York University Langone Medical Center, New York, New York, USA.
Ankit
A
Gupta
Department of Pharmacology, University of Illinois at Chicago, Chicago, Illinois, USA.|University of Illinois Cancer Center, University of Illinois at Chicago, Chicago, Illinois, USA.
Miranda
M
Jacobs
Department of Pharmacology, University of Illinois at Chicago, Chicago, Illinois, USA.|University of Illinois Cancer Center, University of Illinois at Chicago, Chicago, Illinois, USA.|Jesse Brown VA Medical Center, Chicago, Illinois, USA.
Erika
E
Herrero-Garcia
Department of Pharmacology, University of Illinois at Chicago, Chicago, Illinois, USA.|University of Illinois Cancer Center, University of Illinois at Chicago, Chicago, Illinois, USA.
Jacqueline
J
Cobbert
Institute for Research in Immunology and Cancer, Department of Pathology and Cell Biology, Université de Montréal, Montreal, Quebec, Canada.
Hugo
H
Lavoie
Department of Medical Biophysics, Campbell Family Cancer Research Institute, Princess Margaret Cancer Centre, University of Toronto, Toronto, Ontario, Canada.
Matthew
M
Smith
Centre for Systems Biology, Lunenfeld-Tanenbaum Research Institute, Toronto, Ontario, Canada.|Department of Molecular Genetics, University of Toronto, Toronto, Ontario, Canada.|Department of Biochemistry, University of Toronto, Toronto, Ontario, Canada.
Thanashan
T
Rajakulendran
Department of Biochemistry and Molecular Biology, University of Chicago, Chicago, Illinois, USA.
Evan
E
Dowdell
Department of Pharmacology, University of Illinois at Chicago, Chicago, Illinois, USA.|University of Illinois Cancer Center, University of Illinois at Chicago, Chicago, Illinois, USA.
Mustafa Nazir
MN
Okur
Department of Biochemistry and Molecular Biology, University of Chicago, Chicago, Illinois, USA.
Irina
I
Dementieva
Centre for Systems Biology, Lunenfeld-Tanenbaum Research Institute, Toronto, Ontario, Canada.|Department of Molecular Genetics, University of Toronto, Toronto, Ontario, Canada.|Department of Biochemistry, University of Toronto, Toronto, Ontario, Canada.
Frank
F
Sicheri
Institute for Research in Immunology and Cancer, Department of Pathology and Cell Biology, Université de Montréal, Montreal, Quebec, Canada.
Marc
M
Therrien
Department of Integrative Biology and Pharmacology, University of Texas Medical School at Houston, Houston, Texas, USA.
John F
JF
Hancock
Department of Medical Biophysics, Campbell Family Cancer Research Institute, Princess Margaret Cancer Centre, University of Toronto, Toronto, Ontario, Canada.
Mitsuhiko
M
Ikura
Department of Biochemistry and Molecular Biology, University of Chicago, Chicago, Illinois, USA.|Perlmutter Cancer Center, New York University Langone Medical Center, New York, New York, USA.|Department of Biochemistry and Molecular Pharmacology, New York University Langone Medical Center, New York, New York, USA.
Shohei
S
Koide
Department of Pharmacology, University of Illinois at Chicago, Chicago, Illinois, USA.|University of Illinois Cancer Center, University of Illinois at Chicago, Chicago, Illinois, USA.|Jesse Brown VA Medical Center, Chicago, Illinois, USA.
John P
JP
O'Bryan
D000495:Allosteric Site / Q000187:drug effects*; D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology*; D019556:COS Cells; D002478:Cells, Cultured; D002522:Chlorocebus aethiops; D057809:HEK293 Cells; D006801:Humans; D051379:Mice; D041681:NIH 3T3 Cells; D018631:ras Proteins / Q000037:antagonists & inhibitors* / Q000737:chemistry* / Q000378:metabolism
D000911:Antibodies, Monoclonal; D018631:ras Proteins