Selective and noncovalent targeting of RAS mutants for inhibition and degradation

Kai Wen Teng; Steven T. Tsai; Takamitsu Hattori; Carmine Fedele; Akiko Koide; Chao Yang; Xuben Hou; Yingkai Zhang; Benjamin G. Neel; John P. O’Bryan; Shohei Koide

doi:10.1038/s41467-021-22969-5

Selective and noncovalent targeting of RAS mutants for inhibition and degradation

Kai Wen Teng, Steven T. Tsai, Takamitsu Hattori, Carmine Fedele, Akiko Koide, Chao Yang, Xuben Hou, Yingkai Zhang, Benjamin G. Neel, John P. O’Bryan, Shohei Koide

Chemistry

Research output: Contribution to journal › Article › peer-review

Abstract

Activating mutants of RAS are commonly found in human cancers, but to date selective targeting of RAS in the clinic has been limited to KRAS(G12C) through covalent inhibitors. Here, we report a monobody, termed 12VC1, that recognizes the active state of both KRAS(G12V) and KRAS(G12C) up to 400-times more tightly than wild-type KRAS. The crystal structures reveal that 12VC1 recognizes the mutations through a shallow pocket, and 12VC1 competes against RAS-effector interaction. When expressed intracellularly, 12VC1 potently inhibits ERK activation and the proliferation of RAS-driven cancer cell lines in vitro and in mouse xenograft models. 12VC1 fused to VHL selectively degrades the KRAS mutants and provides more extended suppression of mutant RAS activity than inhibition by 12VC1 alone. These results demonstrate the feasibility of selective targeting and degradation of KRAS mutants in the active state with noncovalent reagents and provide a starting point for designing noncovalent therapeutics against oncogenic RAS mutants.

Original language	English (US)
Article number	2656
Journal	Nature communications
Volume	12
Issue number	1
DOIs	https://doi.org/10.1038/s41467-021-22969-5
State	Published - Dec 1 2021

ASJC Scopus subject areas

Chemistry(all)
Biochemistry, Genetics and Molecular Biology(all)
General
Physics and Astronomy(all)

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10.1038/s41467-021-22969-5

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@article{3e54733d27bd43c38a2427b4dbcffc39,

title = "Selective and noncovalent targeting of RAS mutants for inhibition and degradation",

abstract = "Activating mutants of RAS are commonly found in human cancers, but to date selective targeting of RAS in the clinic has been limited to KRAS(G12C) through covalent inhibitors. Here, we report a monobody, termed 12VC1, that recognizes the active state of both KRAS(G12V) and KRAS(G12C) up to 400-times more tightly than wild-type KRAS. The crystal structures reveal that 12VC1 recognizes the mutations through a shallow pocket, and 12VC1 competes against RAS-effector interaction. When expressed intracellularly, 12VC1 potently inhibits ERK activation and the proliferation of RAS-driven cancer cell lines in vitro and in mouse xenograft models. 12VC1 fused to VHL selectively degrades the KRAS mutants and provides more extended suppression of mutant RAS activity than inhibition by 12VC1 alone. These results demonstrate the feasibility of selective targeting and degradation of KRAS mutants in the active state with noncovalent reagents and provide a starting point for designing noncovalent therapeutics against oncogenic RAS mutants.",

author = "Teng, {Kai Wen} and Tsai, {Steven T.} and Takamitsu Hattori and Carmine Fedele and Akiko Koide and Chao Yang and Xuben Hou and Yingkai Zhang and Neel, {Benjamin G.} and O{\textquoteright}Bryan, {John P.} and Shohei Koide",

note = "Funding Information: K.W.T., A.K., T.H., and S.K. are listed as inventors on a patent application on RAS-targeting monobodies filed by New York University (Application No. 63/121,903). A.K. and S.K. are listed as inventors on issued and pending patents on the monobody technology filed by The University of Chicago (US Patent 9512199 B2 and related pending applications). B.G.N. is a co-founder, chair of the Scientific Advisory Board (SAB), and holds equity in Navire Pharmaceuticals; a co-founder and SAB member and holds equity in and receives consulting fees from Northern Biologics; an SAB member and holds equity in and receives consulting fees from Arvinas; an SAB member and holds equity in Recursion Pharma; a co-founder and holds equity in and receives consulting fees from Jengu Therapeutics; serves as an expert witness for Johnson and Johnson; his spouse owns shares in Amgen. S.K. is an SAB member and holds equity in and receives consulting fees from Black Diamond Therapeutics; receives research funding from Puretech Health and Argenx BVBA. The other authors declare no competing interests. Funding Information: The authors would like to acknowledge Beatrix Ueberheide and Jackeline Ponce of the Proteomics Laboratory at NYU Grossman School of Medicine for their technical assistance and helpful discussion; Michael Cammer of the Microscopy Laboratory and the staff of the Cytometry and Cell Sorting Laboratory at NYU Grossman School of Medicine for assistance: Ankit Gupta and Kohei Kurosawa for assistance in cellular studies; Angel Sing for assistance with the mouse experiments; Mark Philips for the pEGFP-KRAS(WT) vector. The X-ray crystallography data were acquired at Beamline 19ID of the Advanced Photon Source (APS) at Argonne National Lab with the support of beamline scientists Jurek Opsipiuk and Kemin Tan. APS is a U.S. Department of Energy (DOE) Office of Science User Facility operated for the DOE Office of Science by Argonne National Laboratory under Contract No. DE-AC02-06CH11357. This work was supported by the National Institutes of Health grants R35 GM127040 (Y.Z.), R01 CA194864 (S.K.), R21 CA201717 (J.P.O.), and R01 CA212608 (J.P.O. and S.K.). K.W.T. was supported in part by NIH fellowship (F32 CA225131) and ACS fellowship (PF-18-180-01-TBE). The core facilities of NYU Grossman School of Medicine were partially supported by the Cancer Center Support Grant P30CA016087. Publisher Copyright: {\textcopyright} 2021, The Author(s).",

year = "2021",

month = dec,

day = "1",

doi = "10.1038/s41467-021-22969-5",

language = "English (US)",

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journal = "Nature Communications",

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T1 - Selective and noncovalent targeting of RAS mutants for inhibition and degradation

AU - Teng, Kai Wen

AU - Tsai, Steven T.

AU - Hattori, Takamitsu

AU - Fedele, Carmine

AU - Koide, Akiko

AU - Yang, Chao

AU - Hou, Xuben

AU - Zhang, Yingkai

AU - Neel, Benjamin G.

AU - O’Bryan, John P.

AU - Koide, Shohei

N1 - Funding Information: K.W.T., A.K., T.H., and S.K. are listed as inventors on a patent application on RAS-targeting monobodies filed by New York University (Application No. 63/121,903). A.K. and S.K. are listed as inventors on issued and pending patents on the monobody technology filed by The University of Chicago (US Patent 9512199 B2 and related pending applications). B.G.N. is a co-founder, chair of the Scientific Advisory Board (SAB), and holds equity in Navire Pharmaceuticals; a co-founder and SAB member and holds equity in and receives consulting fees from Northern Biologics; an SAB member and holds equity in and receives consulting fees from Arvinas; an SAB member and holds equity in Recursion Pharma; a co-founder and holds equity in and receives consulting fees from Jengu Therapeutics; serves as an expert witness for Johnson and Johnson; his spouse owns shares in Amgen. S.K. is an SAB member and holds equity in and receives consulting fees from Black Diamond Therapeutics; receives research funding from Puretech Health and Argenx BVBA. The other authors declare no competing interests. Funding Information: The authors would like to acknowledge Beatrix Ueberheide and Jackeline Ponce of the Proteomics Laboratory at NYU Grossman School of Medicine for their technical assistance and helpful discussion; Michael Cammer of the Microscopy Laboratory and the staff of the Cytometry and Cell Sorting Laboratory at NYU Grossman School of Medicine for assistance: Ankit Gupta and Kohei Kurosawa for assistance in cellular studies; Angel Sing for assistance with the mouse experiments; Mark Philips for the pEGFP-KRAS(WT) vector. The X-ray crystallography data were acquired at Beamline 19ID of the Advanced Photon Source (APS) at Argonne National Lab with the support of beamline scientists Jurek Opsipiuk and Kemin Tan. APS is a U.S. Department of Energy (DOE) Office of Science User Facility operated for the DOE Office of Science by Argonne National Laboratory under Contract No. DE-AC02-06CH11357. This work was supported by the National Institutes of Health grants R35 GM127040 (Y.Z.), R01 CA194864 (S.K.), R21 CA201717 (J.P.O.), and R01 CA212608 (J.P.O. and S.K.). K.W.T. was supported in part by NIH fellowship (F32 CA225131) and ACS fellowship (PF-18-180-01-TBE). The core facilities of NYU Grossman School of Medicine were partially supported by the Cancer Center Support Grant P30CA016087. Publisher Copyright: © 2021, The Author(s).

PY - 2021/12/1

Y1 - 2021/12/1

N2 - Activating mutants of RAS are commonly found in human cancers, but to date selective targeting of RAS in the clinic has been limited to KRAS(G12C) through covalent inhibitors. Here, we report a monobody, termed 12VC1, that recognizes the active state of both KRAS(G12V) and KRAS(G12C) up to 400-times more tightly than wild-type KRAS. The crystal structures reveal that 12VC1 recognizes the mutations through a shallow pocket, and 12VC1 competes against RAS-effector interaction. When expressed intracellularly, 12VC1 potently inhibits ERK activation and the proliferation of RAS-driven cancer cell lines in vitro and in mouse xenograft models. 12VC1 fused to VHL selectively degrades the KRAS mutants and provides more extended suppression of mutant RAS activity than inhibition by 12VC1 alone. These results demonstrate the feasibility of selective targeting and degradation of KRAS mutants in the active state with noncovalent reagents and provide a starting point for designing noncovalent therapeutics against oncogenic RAS mutants.

AB - Activating mutants of RAS are commonly found in human cancers, but to date selective targeting of RAS in the clinic has been limited to KRAS(G12C) through covalent inhibitors. Here, we report a monobody, termed 12VC1, that recognizes the active state of both KRAS(G12V) and KRAS(G12C) up to 400-times more tightly than wild-type KRAS. The crystal structures reveal that 12VC1 recognizes the mutations through a shallow pocket, and 12VC1 competes against RAS-effector interaction. When expressed intracellularly, 12VC1 potently inhibits ERK activation and the proliferation of RAS-driven cancer cell lines in vitro and in mouse xenograft models. 12VC1 fused to VHL selectively degrades the KRAS mutants and provides more extended suppression of mutant RAS activity than inhibition by 12VC1 alone. These results demonstrate the feasibility of selective targeting and degradation of KRAS mutants in the active state with noncovalent reagents and provide a starting point for designing noncovalent therapeutics against oncogenic RAS mutants.

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Selective and noncovalent targeting of RAS mutants for inhibition and degradation

Abstract

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