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A portrait of rare RAS mutants

Mutations in the RAS family of proteins promote various cancers, of which mutations at Gln61 (also referred to as Q61) are relatively and unexpectedly rare in KRAS. Huynh et al. compared the structural, biochemical, and biological effects of Gln61 mutations in KRAS (see also Sealover and Kortum). The Q61E mutant had weaker metabolic activity and induced greater cell adhesion and less motility than the other mutants, thus potentially explaining the relative rarity of this mutant. Nonetheless, other aspects made Gln61 mutant KRAS pancreatic cancer cells susceptible to the same inhibitors that are currently in trials to treat tumors dependent on the more common KRAS mutants. The findings may both broaden and differentiate treatment strategies for these rarer mutant KRAS–driven tumors.

Abstract

Missense mutations at the three hotspots in the guanosine triphosphatase (GTPase) RAS—Gly12, Gly13, and Gln61 (commonly known as G12, G13, and Q61, respectively)—occur differentially among the three RAS isoforms. Q61 mutations in KRAS are infrequent and differ markedly in occurrence. Q61H is the predominant mutant (at 57%), followed by Q61R/L/K (collectively 40%), and Q61P and Q61E are the rarest (2 and 1%, respectively). Probability analysis suggested that mutational susceptibility to different DNA base changes cannot account for this distribution. Therefore, we investigated whether these frequencies might be explained by differences in the biochemical, structural, and biological properties of KRASQ61 mutants. Expression of KRASQ61 mutants in NIH 3T3 fibroblasts and RIE-1 epithelial cells caused various alterations in morphology, growth transformation, effector signaling, and metabolism. The relatively rare KRASQ61E mutant stimulated actin stress fiber formation, a phenotype distinct from that of KRASQ61H/R/L/P, which disrupted actin cytoskeletal organization. The crystal structure of KRASQ61E was unexpectedly similar to that of wild-type KRAS, a potential basis for its weak oncogenicity. KRASQ61H/L/R-mutant pancreatic ductal adenocarcinoma (PDAC) cell lines exhibited KRAS-dependent growth and, as observed with KRASG12-mutant PDAC, were susceptible to concurrent inhibition of ERK-MAPK signaling and of autophagy. Our results uncover phenotypic heterogeneity among KRASQ61 mutants and support the potential utility of therapeutic strategies that target KRASQ61 mutant–specific signaling and cellular output.

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MDAR Reproducibility Checklist

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Science Signaling
Volume 15 | Issue 746
August 2022

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Received: 16 November 2021
Accepted: 15 July 2022

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Acknowledgments

We thank A. Maitra and J. Fleming for PDAC cell lines and E. Baldelli for assistance with RPPA analyses.
Funding: M.V.H. was supported by the Slomo and Cindy Silvian Foundation. G.A.H. was supported by National Institute of General Medical Sciences grant P20GM130457. L.M.C. was supported by NCI T32CA009156 and American Cancer Society PF-20-140-01–CDD. J.N.D. was supported by the Slomo and Cindy Silvian Foundation, NCI T32CA071341, and F30CA243253. A.M.W. was supported by a fellowship from the American Cancer Society (F-18-061). R.D.T. was supported by NCI T32CA009156. S.L.C. and E.F.P. were supported by NCI CA203657. Support was provided by grants from the NCI to A.D.C. and/or C.J.D. (CA42978, R01CA175747, R01CA223775, P50CA196510, U01CA199235, P01CA203657, and R35CA232113) and from the Pancreatic Cancer Action Network/AACR (15-90-25-DER).
Author contributions: M.V.H. and C.J.D. designed the experiments. M.V.H., G.A.H., A.M.W., J.H.C., K.M.H., A.D.C., and C.J.D. wrote the manuscript. M.V.H., G.A.H., A.S., M.P., L.M.C., J.M.D., R.D.T., L.L., and A.R.C. performed the experiments. J.N.D., C.M.G., J.H.C., and K.M.H. provided computational analyses. L.L., C.A.L., and D.M.S. provided key experimental reagents. E.F.P., S.L.C., K.M.H., D.M.S., N.U.R., C.A.L., A.D.C., and C.J.D. provided scientific guidance.
Competing interests: C.J.D. is an advisory board member for Anchiano Therapeutics, Boragen, Deciphera Pharmaceuticals, Mirati Therapeutics, Revolution Medicines, and SHY Therapeutics; has received research funding support from Boragen, Deciphera Pharmaceuticals, Mirati Therapeutics, and SpringWorks Therapeutics; and has consulted for Day One Biotherapeutics, Eli Lilly, Jazz Therapeutics, Ribometrix, Sanofi, and Turning Point Therapeutics. A.D.C. has consulted for Eli Lilly and Mirati Therapeutics. E.F.P. and M.P. are consultants for and receive royalties from Avant Diagnostics and TheraLink Technologies Inc. E.F.P. is a consultant to and shareholder of Avant Diagnostics Inc. and Perthera Inc. and received funding support from Mirati Therapeutics, Genentech Inc., and AbbVie Inc. The other authors declare they have no competing interests.
Data and materials availability: Plasmids generated for this study will be deposited into Addgene. The crystal structure of KRASQ61E has been deposited into the Protein Data Bank (PDB accession code 7LZ5). All other data needed to evaluate the conclusions in the paper are present in the paper or the Supplementary Materials.

Authors

Affiliations

Department of Biochemistry and Biophysics, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
Roles: Conceptualization, Formal analysis, Investigation, Methodology, Validation, Visualization, and Writing - original draft.
Department of Cell and Molecular Pharmacology and Experimental Therapeutics, Medical University of South Carolina, Charleston, SC 29425, USA.
Roles: Conceptualization, Investigation, Methodology, Visualization, Writing - original draft, and Writing - review & editing.
Department of Pharmacology, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
Roles: Investigation, Methodology, and Validation.
Mariaelena Pierobon
Center for Applied Proteomics and Molecular Medicine, George Mason University, Manassas, VA 20110, USA.
Roles: Investigation, Methodology, Project administration, and Writing - review & editing.
Department of Biochemistry and Biophysics, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
Roles: Investigation, Validation, Visualization, Writing - original draft, and Writing - review & editing.
Curriculum in Genetics and Molecular Biology, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
Roles: Formal analysis, Software, Visualization, and Writing - review & editing.
Jonathan M. DeLiberty
Department of Pharmacology, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
Role: Investigation.
Ryan D. Thurman
Department of Biochemistry and Biophysics, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
Roles: Conceptualization, Formal analysis, and Methodology.
Present address: University at Albany, SUNY, Albany, NY 12222, USA.
Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
Role: Investigation.
Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
Roles: Formal analysis, Investigation, Software, and Visualization.
Department of Cancer Biology, Dana-Farber Cancer Institute, Boston, MA 02215, USA.
Department of Medicine, Brigham and Women’s Hospital, Harvard Medical School, Boston, MA 02115, USA.
Department of Biomedical Informatics, Harvard Medical School, Boston, MA 02115, USA.
Roles: Data curation, Formal analysis, Investigation, Resources, Software, Visualization, and Writing - review & editing.
Department of Molecular and Integrative Physiology, University of Michigan Health System, Ann Arbor, MI 48109, USA.
Roles: Methodology and Resources.
Department of Pharmacology, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
Roles: Formal analysis, Investigation, Methodology, Resources, Validation, Visualization, and Writing - review & editing.
Naim U. Rashid
Department of Biostatistics, University of North Carolina at Chapel Hill, NC 27955, USA.
Role: Visualization.
Emanuel F. Petricoin III
Center for Applied Proteomics and Molecular Medicine, George Mason University, Manassas, VA 20110, USA.
Roles: Conceptualization, Investigation, Methodology, Resources, and Writing - review & editing.
Department of Biochemistry and Biophysics, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
Roles: Conceptualization, Funding acquisition, Methodology, Project administration, Resources, Supervision, Validation, Visualization, and Writing - review & editing.
Kevin M. Haigis
Department of Cancer Biology, Dana-Farber Cancer Institute, Boston, MA 02215, USA.
Department of Medicine, Brigham and Women’s Hospital, Harvard Medical School, Boston, MA 02115, USA.
Broad Institute, Cambridge, MA 02142, USA.
Harvard Digestive Disease Center, Harvard Medical School, Boston, MA 02115, USA.
Roles: Conceptualization, Project administration, and Supervision.
Perlmutter Cancer Center, New York University, New York, NY 10016, USA.
Roles: Conceptualization, Data curation, Formal analysis, Investigation, Methodology, Project administration, Resources, Supervision, Validation, Visualization, Writing - original draft, and Writing - review & editing.
Department of Molecular and Integrative Physiology, University of Michigan Health System, Ann Arbor, MI 48109, USA.
Division of Gastroenterology, Department of Internal Medicine, University of Michigan, Ann Arbor, MI 48198, USA.
University of Michigan Comprehensive Cancer Center, Ann Arbor, MI 48109, USA.
Roles: Methodology, Resources, Validation, and Writing - review & editing.
Department of Pharmacology, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
Department of Radiation Oncology, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
Roles: Conceptualization, Funding acquisition, Project administration, Supervision, Visualization, Writing - original draft, and Writing - review & editing.
Department of Pharmacology, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
Curriculum in Genetics and Molecular Biology, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
Roles: Conceptualization, Funding acquisition, Methodology, Project administration, Supervision, Validation, Visualization, Writing - original draft, and Writing - review & editing.

Funding Information

American Cancer Society: PF-20-140-01 – CDD
American Cancer Society: NCI T32CA009156
Slomo and Cindy Silvian Foundation
Slomo and Cindy Silvian Foundation

Notes

*
Corresponding author. Email: [email protected] (C.J.D.); [email protected] (G.A.H.); [email protected] (S.L.C.)

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  1. Heterogeneity in RAS mutations: One size does not fit all, Science Signaling, 15, 746, (2022)./doi/10.1126/scisignal.adc9816
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