Therapeutic high affinity T cell receptor targeting a KRASG12D cancer neoantigen

Andrew Poole; Vijaykumar Karuppiah; Annabelle M Hartt; Marc W Van der Kamp; Andrew D. Whale; Chandramouli Chillakuri

doi:10.1038/s41467-022-32811-1

Therapeutic high affinity T cell receptor targeting a KRASG12D cancer neoantigen

Andrew Poole, Vijaykumar Karuppiah, Annabelle M Hartt, Marc W Van der Kamp, Andrew D. Whale^*, Chandramouli Chillakuri^*

^*Corresponding author for this work

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Abstract

Neoantigens derived from somatic mutations are specific to cancer cells and are ideal targets for cancer immunotherapy. KRAS is the most frequently mutated oncogene and drives the pathogenesis of several cancers. Here we show the identification and development of an affinity-enhanced T cell receptor (TCR) that recognizes a peptide derived from the most common KRAS mutant, KRASG12D, presented in the context of HLA-A*11:01. The affinity of the engineered TCR is increased by over one million-fold yet fully able to distinguish KRASG12D over KRASWT. While crystal structures reveal few discernible differences in TCR interactions with KRASWT versus KRASG12D, thermodynamic analysis and molecular dynamics simulations reveal that TCR specificity is driven by differences in indirect electrostatic interactions. The affinity enhanced TCR, fused to a humanized anti-CD3 scFv, enables selective killing of cancer cells expressing KRASG12D. Our work thus reveals a molecular mechanism that drives TCR selectivity and describes a soluble bispecific molecule with therapeutic potential against cancers harboring a common shared neoantigen.

Original language	English
Article number	5333
Journal	Nature Communications
Volume	13
Issue number	1
DOIs	https://doi.org/10.1038/s41467-022-32811-1 https://doi.org/10.1038/s41467-022-32811-1
Publication status	Published - 10 Sept 2022

Bibliographical note

Funding Information:
We would like to thank Michelle McCully, Laure Humbert, Peter Molloy, and Ita O’Kelly for critical reading and help preparing this manuscript. We also wish to thank: Vanessa Clarke, Max Beckmann, Sarah Bailey, Nicole Mai, Tein Foong-Leong, and Peter James for discussions and supporting identification of the KRASG12Dspecific TCR and affinity maturation; Kathryn Lamming, Alex Powlesland, and Ricardo Carreira for cell line characterisation; Hemza Ghadbane for design of CRISPR guides, repair templates and editing approach used to edit HLA and KRAS alleles, and Carmine Carpenito and JoAnn Suzich for their scientific input and insights. We would like to thank Diamond Light Source and staff of beamlines for access to beamtime (proposal mx22870). A.H. and M.v.d.K thank the Engineering and Physical Sciences Research Council [grant number EP/T517872/1] and the Biotechnology and Biological Sciences Research Council [grant number BB/M026280/1] for support. Computer simulations were conducted using the computational facilities of the Advanced Computing Research Centre of the University of Bristol.

Funding Information:
We would like to thank Michelle McCully, Laure Humbert, Peter Molloy, and Ita O’Kelly for critical reading and help preparing this manuscript. We also wish to thank: Vanessa Clarke, Max Beckmann, Sarah Bailey, Nicole Mai, Tein Foong-Leong, and Peter James for discussions and supporting identification of the KRAS specific TCR and affinity maturation; Kathryn Lamming, Alex Powlesland, and Ricardo Carreira for cell line characterisation; Hemza Ghadbane for design of CRISPR guides, repair templates and editing approach used to edit HLA and KRAS alleles, and Carmine Carpenito and JoAnn Suzich for their scientific input and insights. We would like to thank Diamond Light Source and staff of beamlines for access to beamtime (proposal mx22870). A.H. and M.v.d.K thank the Engineering and Physical Sciences Research Council [grant number EP/T517872/1] and the Biotechnology and Biological Sciences Research Council [grant number BB/M026280/1] for support. Computer simulations were conducted using the computational facilities of the Advanced Computing Research Centre of the University of Bristol. G12D

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© 2022, The Author(s).

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Cite this

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title = "Therapeutic high affinity T cell receptor targeting a KRASG12D cancer neoantigen",

abstract = "Neoantigens derived from somatic mutations are specific to cancer cells and are ideal targets for cancer immunotherapy. KRAS is the most frequently mutated oncogene and drives the pathogenesis of several cancers. Here we show the identification and development of an affinity-enhanced T cell receptor (TCR) that recognizes a peptide derived from the most common KRAS mutant, KRASG12D, presented in the context of HLA-A*11:01. The affinity of the engineered TCR is increased by over one million-fold yet fully able to distinguish KRASG12D over KRASWT. While crystal structures reveal few discernible differences in TCR interactions with KRASWT versus KRASG12D, thermodynamic analysis and molecular dynamics simulations reveal that TCR specificity is driven by differences in indirect electrostatic interactions. The affinity enhanced TCR, fused to a humanized anti-CD3 scFv, enables selective killing of cancer cells expressing KRASG12D. Our work thus reveals a molecular mechanism that drives TCR selectivity and describes a soluble bispecific molecule with therapeutic potential against cancers harboring a common shared neoantigen.",

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note = "Funding Information: We would like to thank Michelle McCully, Laure Humbert, Peter Molloy, and Ita O{\textquoteright}Kelly for critical reading and help preparing this manuscript. We also wish to thank: Vanessa Clarke, Max Beckmann, Sarah Bailey, Nicole Mai, Tein Foong-Leong, and Peter James for discussions and supporting identification of the KRASG12Dspecific TCR and affinity maturation; Kathryn Lamming, Alex Powlesland, and Ricardo Carreira for cell line characterisation; Hemza Ghadbane for design of CRISPR guides, repair templates and editing approach used to edit HLA and KRAS alleles, and Carmine Carpenito and JoAnn Suzich for their scientific input and insights. We would like to thank Diamond Light Source and staff of beamlines for access to beamtime (proposal mx22870). A.H. and M.v.d.K thank the Engineering and Physical Sciences Research Council [grant number EP/T517872/1] and the Biotechnology and Biological Sciences Research Council [grant number BB/M026280/1] for support. Computer simulations were conducted using the computational facilities of the Advanced Computing Research Centre of the University of Bristol. Funding Information: We would like to thank Michelle McCully, Laure Humbert, Peter Molloy, and Ita O{\textquoteright}Kelly for critical reading and help preparing this manuscript. We also wish to thank: Vanessa Clarke, Max Beckmann, Sarah Bailey, Nicole Mai, Tein Foong-Leong, and Peter James for discussions and supporting identification of the KRAS specific TCR and affinity maturation; Kathryn Lamming, Alex Powlesland, and Ricardo Carreira for cell line characterisation; Hemza Ghadbane for design of CRISPR guides, repair templates and editing approach used to edit HLA and KRAS alleles, and Carmine Carpenito and JoAnn Suzich for their scientific input and insights. We would like to thank Diamond Light Source and staff of beamlines for access to beamtime (proposal mx22870). A.H. and M.v.d.K thank the Engineering and Physical Sciences Research Council [grant number EP/T517872/1] and the Biotechnology and Biological Sciences Research Council [grant number BB/M026280/1] for support. Computer simulations were conducted using the computational facilities of the Advanced Computing Research Centre of the University of Bristol. G12D Publisher Copyright: {\textcopyright} 2022, The Author(s).",

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AU - Van der Kamp, Marc W

AU - Whale, Andrew D.

AU - Chillakuri, Chandramouli

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N2 - Neoantigens derived from somatic mutations are specific to cancer cells and are ideal targets for cancer immunotherapy. KRAS is the most frequently mutated oncogene and drives the pathogenesis of several cancers. Here we show the identification and development of an affinity-enhanced T cell receptor (TCR) that recognizes a peptide derived from the most common KRAS mutant, KRASG12D, presented in the context of HLA-A*11:01. The affinity of the engineered TCR is increased by over one million-fold yet fully able to distinguish KRASG12D over KRASWT. While crystal structures reveal few discernible differences in TCR interactions with KRASWT versus KRASG12D, thermodynamic analysis and molecular dynamics simulations reveal that TCR specificity is driven by differences in indirect electrostatic interactions. The affinity enhanced TCR, fused to a humanized anti-CD3 scFv, enables selective killing of cancer cells expressing KRASG12D. Our work thus reveals a molecular mechanism that drives TCR selectivity and describes a soluble bispecific molecule with therapeutic potential against cancers harboring a common shared neoantigen.

AB - Neoantigens derived from somatic mutations are specific to cancer cells and are ideal targets for cancer immunotherapy. KRAS is the most frequently mutated oncogene and drives the pathogenesis of several cancers. Here we show the identification and development of an affinity-enhanced T cell receptor (TCR) that recognizes a peptide derived from the most common KRAS mutant, KRASG12D, presented in the context of HLA-A*11:01. The affinity of the engineered TCR is increased by over one million-fold yet fully able to distinguish KRASG12D over KRASWT. While crystal structures reveal few discernible differences in TCR interactions with KRASWT versus KRASG12D, thermodynamic analysis and molecular dynamics simulations reveal that TCR specificity is driven by differences in indirect electrostatic interactions. The affinity enhanced TCR, fused to a humanized anti-CD3 scFv, enables selective killing of cancer cells expressing KRASG12D. Our work thus reveals a molecular mechanism that drives TCR selectivity and describes a soluble bispecific molecule with therapeutic potential against cancers harboring a common shared neoantigen.

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DO - 10.1038/s41467-022-32811-1

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JO - Nature Communications

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Therapeutic high affinity T cell receptor targeting a KRASG12D cancer neoantigen

Abstract

Bibliographical note

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