Solution structure and synaptic analyses reveal determinants of bispecific T cell engager potency

Bispecific T cell engagers (TcEs) link T cell receptors to tumor-associated antigens on cancer cells, forming cytotoxic immunological synapses (IS). Close membrane-to-membrane contact (≤13 nm) has been proposed as a key mechanism of TcE function. To investigate this and identify potential additional mechanisms, we compared four immunoglobulin G1-based (IgG1) TcE Formats (A-D) targeting CD3ε and Her2, designed to create varying intermembrane distances (A < B < C < D). Small-angle X-ray scattering (SAXS) and modeling of the conformational states of isolated TcEs and TcE-antigen complexes predicted close contacts (≤13 nm) for Formats A and B and far contacts (≥18 nm) for Formats C and D. In supported lipid bilayer (SLB) model interfaces, Formats A and B recruited, whereas Formats C and D repelled, CD2-CD58 interactions. Formats A and B also excluded bulky Quantum dots more effectively. SAXS also revealed that TcE-antigen complexes formed by Formats A and C were less flexible than complexes formed by Formats B and D. Functional data with Her2-expressing tumor cells showed cytotoxicity, surface marker expression, and cytokine release following the order A > B = C > D. In a minimal system for IS formation on SLBs, TcE performance followed the trend A = B = C > D. Addition of close contact requiring CD58 costimulation revealed phospholipase C-γ activation matching cytotoxicity with A > B = C > D. Our findings suggest that when adhesion is equivalent, TcE potency is determined by two parameters: contact distance and flexibility. Both the close/far-contact formation axis and the low/high flexibility axis significantly impact TcE potency, explaining the similar potency of Format B (close contact/high flexibility) and C (far contact/low flexibility).

Authors

Institutions

Publication Details

Journal
Proceedings of the National Academy of Sciences
Published
2025-05-30
DOI
https://doi.org/10.1073/pnas.2425781122
Citations
4
Primary Topic
Monoclonal and Polyclonal Antibodies Research
Type
article
Field-Weighted Citation Impact
5.44

Funders

Controls
|||
ALL TIME
JAN
FEB
MAR
APR
MAY
JUN
JUL
AUG
SEP
OCT
article

Solution structure and synaptic analyses reveal determinants of bispecific T cell engager potency

Felix Schiele, Andrew E. Nixon, S. Hoerer, Hannah Dada et al.
4 citations
Proceedings of the National Academy of Sciences
Monoclonal and Polyclonal Antibodies Research
5.44
article

Solution structure and synaptic analyses reveal determinants of bispecific T cell engager potency

Felix Schiele, Andrew E. Nixon, S. Hoerer, Hannah Dada, Oskar Staufer, Omer Dushek, Maureen Crames, Salvatore Valvo, Edward M. Jenkins, Mikhail A. Kutuzov, Stephen R. Comeau, Michael Loran Dustin, David A. Klenerman, Alexander Leithner, Herbert Nar, Sophia Reindl, Simon J. Davis, Noah Pefaur, Sarah Löw, Falk Liberta, Srinath Kasturirangan, Tanmay Mitra, Weijie Zhou, David S. Young, David Wyatt, J.‐M. Spaeth
article en
4 citations

Abstract

Bispecific T cell engagers (TcEs) link T cell receptors to tumor-associated antigens on cancer cells, forming cytotoxic immunological synapses (IS). Close membrane-to-membrane contact (≤13 nm) has been proposed as a key mechanism of TcE function. To investigate this and identify potential additional mechanisms, we compared four immunoglobulin G1-based (IgG1) TcE Formats (A-D) targeting CD3ε and Her2, designed to create varying intermembrane distances (A < B < C < D). Small-angle X-ray scattering (SAXS) and modeling of the conformational states of isolated TcEs and TcE-antigen complexes predicted close contacts (≤13 nm) for Formats A and B and far contacts (≥18 nm) for Formats C and D. In supported lipid bilayer (SLB) model interfaces, Formats A and B recruited, whereas Formats C and D repelled, CD2-CD58 interactions. Formats A and B also excluded bulky Quantum dots more effectively. SAXS also revealed that TcE-antigen complexes formed by Formats A and C were less flexible than complexes formed by Formats B and D. Functional data with Her2-expressing tumor cells showed cytotoxicity, surface marker expression, and cytokine release following the order A > B = C > D. In a minimal system for IS formation on SLBs, TcE performance followed the trend A = B = C > D. Addition of close contact requiring CD58 costimulation revealed phospholipase C-γ activation matching cytotoxicity with A > B = C > D. Our findings suggest that when adhesion is equivalent, TcE potency is determined by two parameters: contact distance and flexibility. Both the close/far-contact formation axis and the low/high flexibility axis significantly impact TcE potency, explaining the similar potency of Format B (close contact/high flexibility) and C (far contact/low flexibility).

Proceedings of the National Academy of SciencesVol. 122(22)
Academy of Medical Sciences (GB), Boehringer Ingelheim (Germany) (DE), Chinese Academy of Medical Sciences & Peking Union Medical College (CN), University of Cambridge (GB), Nuffield Orthopaedic Centre (GB), John Radcliffe Hospital (GB), University of Oxford (GB), Leibniz-Institute for New Materials (DE), Institute of Rheumatology (CZ), Kennedy Center (DK), MRC Human Immunology Unit (GB), Institute of Molecular Medicine (IN), Boehringer Ingelheim (Australia) (AU), Boehringer Ingelheim (United States) (US), Boehringer Ingelheim (Spain) (ES), Medical Research Council (GB)
Joachim Herz Stiftung, Wellcome Trust, Diamond Light Source, Kennedy Trust for Rheumatology Research, University of Oxford, Austrian Science Fund, Chinese Academy of Medical Sciences, Directorate for Biological Sciences, Medical Research Council, Biotechnology and Biological Sciences Research Council, Leibniz-Gemeinschaft
Openalex Percentile: Top 4%
Monoclonal and Polyclonal Antibodies Research
5.44
DOIOpen Access PDFOpenAlex
AI Navigator

Ask Laika to Summarize, Analyze, and Connect papers live on the map.

Summarize Papers & Methodologies

Extract key findings, datasets, and comparative methods across publications.

Benchmark Rankings & Visual Analytics

Rank top research institutions, authors, funders, topics, and journals by Field-Weighted Citation Impact (FWCI) and paper volume with instant charts.

Connect Distant Disciplines

Bridge topological clusters on the map to find hidden collaborative intersections.