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TNF receptor agonists induce distinct receptor clusters to mediate differential agonistic activity

TNF receptor agonists induce distinct receptor clusters to mediate differential agonistic activity
TNF receptor agonists induce distinct receptor clusters to mediate differential agonistic activity

Monoclonal antibodies (mAb) and natural ligands targeting costimulatory tumor necrosis factor receptors (TNFR) exhibit a wide range of agonistic activities and antitumor responses. The mechanisms underlying these differential agonistic activities remain poorly understood. Here, we employ a panel of experimental and clinically-relevant molecules targeting human CD40, 4-1BB and OX40 to examine this issue. Confocal and STORM microscopy reveal that strongly agonistic reagents induce clusters characterized by small area and high receptor density. Using antibody pairs differing only in isotype we show that hIgG2 confers significantly more receptor clustering than hIgG1 across all three receptors, explaining its greater agonistic activity, with receptor clustering shielding the receptor-agonist complex from further molecular access. Nevertheless, discrete receptor clustering patterns are observed with different hIgG2 mAb, with a unique rod-shaped assembly observed with the most agonistic mAb. These findings dispel the notion that larger receptor clusters elicit greater agonism, and instead point to receptor density and subsequent super-structure as key determinants.

2399-3642
Yu, Xiaojie
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James, Sonya
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Felce, James H.
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Kellermayer, Blanka
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Johnston, David A.
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Chan, H. T.Claude
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Penfold, Christine A.
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Kim, Jinny
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Inzhelevskaya, Tatyana
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Mockridge, C. Ian
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Watanabe, Yasunori
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Crispin, Max
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French, Ruth R.
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Duriez, Patrick J.
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Douglas, Leon R.
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Glennie, Martin J.
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Cragg, Mark S.
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Yu, Xiaojie
44d52374-eacc-4e23-b7da-c881e6d3a5dd
James, Sonya
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Felce, James H.
53fa8b4e-b71b-4ba8-9c1b-2e8912e2cc0a
Kellermayer, Blanka
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Johnston, David A.
b41163c9-b9d2-425c-af99-2a357204014e
Chan, H. T.Claude
dd26be49-6344-432e-b458-8c644bf4828f
Penfold, Christine A.
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Kim, Jinny
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Inzhelevskaya, Tatyana
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Mockridge, C. Ian
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Watanabe, Yasunori
8c0ee4af-a293-4de5-9036-3ce2051b380c
Crispin, Max
cd980957-0943-4b89-b2b2-710f01f33bc9
French, Ruth R.
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Duriez, Patrick J.
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Douglas, Leon R.
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Glennie, Martin J.
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Cragg, Mark S.
ec97f80e-f3c8-49b7-a960-20dff648b78c

Yu, Xiaojie, James, Sonya, Felce, James H., Kellermayer, Blanka, Johnston, David A., Chan, H. T.Claude, Penfold, Christine A., Kim, Jinny, Inzhelevskaya, Tatyana, Mockridge, C. Ian, Watanabe, Yasunori, Crispin, Max, French, Ruth R., Duriez, Patrick J., Douglas, Leon R., Glennie, Martin J. and Cragg, Mark S. (2021) TNF receptor agonists induce distinct receptor clusters to mediate differential agonistic activity. Communications Biology, 4 (1), [772]. (doi:10.1038/s42003-021-02309-5).

Record type: Article

Abstract

Monoclonal antibodies (mAb) and natural ligands targeting costimulatory tumor necrosis factor receptors (TNFR) exhibit a wide range of agonistic activities and antitumor responses. The mechanisms underlying these differential agonistic activities remain poorly understood. Here, we employ a panel of experimental and clinically-relevant molecules targeting human CD40, 4-1BB and OX40 to examine this issue. Confocal and STORM microscopy reveal that strongly agonistic reagents induce clusters characterized by small area and high receptor density. Using antibody pairs differing only in isotype we show that hIgG2 confers significantly more receptor clustering than hIgG1 across all three receptors, explaining its greater agonistic activity, with receptor clustering shielding the receptor-agonist complex from further molecular access. Nevertheless, discrete receptor clustering patterns are observed with different hIgG2 mAb, with a unique rod-shaped assembly observed with the most agonistic mAb. These findings dispel the notion that larger receptor clusters elicit greater agonism, and instead point to receptor density and subsequent super-structure as key determinants.

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s42003-021-02309-5 - Version of Record
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e-pub ahead of print date: 23 June 2021
Additional Information: Funding Information: We would like to thank the members of the Antibody and Vaccine group for useful discussions and the pre-clinical unit staff for animal husbandry. We also thank Professors Simon Davis, Marion Brown and Neil Barclay for helpful discussions. Funding was provided by CRUK grants A10834, A20537, A18087, A25139 and A25169 as well as EU FP7 grant 602262-2. M.C. acknowledges funding by Against Breast Cancer. The STORM microscopy work was made possible through the generous funding of an ONI Nanoimager by the Mark Benevolent Fund. X.Y. is funded by a Careertrack Fellowship provided by the Faculty of Medicine in conjunction with the Centre for Cancer Immunology Talent fund. Funding Information: Mark S. Cragg acts as a consultant for a number of biotech companies, being retained as a consultant for BioInvent and has received research funding from BioInvent, GSK, UCB, iTeos, and Roche. Martin J. Glennie acts as a consultant to a number of biotech companies and receives institutional payments and royalties from antibody patents and licences. This work is related to patent Family WO 2015/145360 protecting antibodies containing modified hIgG2 domains which elicit agonist or antagonistic properties. Publisher Copyright: © 2021, The Author(s). Copyright: Copyright 2021 Elsevier B.V., All rights reserved.

Identifiers

Local EPrints ID: 450384
URI: http://eprints.soton.ac.uk/id/eprint/450384
ISSN: 2399-3642
PURE UUID: dca40235-525d-4a99-8904-08ff4a4f5dd6
ORCID for David A. Johnston: ORCID iD orcid.org/0000-0001-6703-6014
ORCID for Max Crispin: ORCID iD orcid.org/0000-0002-1072-2694
ORCID for Mark S. Cragg: ORCID iD orcid.org/0000-0003-2077-089X

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Date deposited: 27 Jul 2021 16:30
Last modified: 26 Nov 2021 03:11

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Contributors

Author: Xiaojie Yu
Author: Sonya James
Author: James H. Felce
Author: Blanka Kellermayer
Author: David A. Johnston ORCID iD
Author: H. T.Claude Chan
Author: Christine A. Penfold
Author: Jinny Kim
Author: Tatyana Inzhelevskaya
Author: C. Ian Mockridge
Author: Yasunori Watanabe
Author: Max Crispin ORCID iD
Author: Ruth R. French
Author: Patrick J. Duriez
Author: Leon R. Douglas
Author: Mark S. Cragg ORCID iD

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