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

• 成果类型: Article
• 署名作者: Leithner, Alexander; Staufer, Oskar; Mitra, Tanmay; Liberta, Falk; Valvo, Salvatore; Kutuzov, Mikhail; Dada, Hannah; Spaeth, Jacob; Zhou, Weijie; Schiele, Felix; Reindl, Sophia; Nar, Herbert; Hoerer, Stefan; Crames, Maureen; Comeau, Stephen; Young, David; Low, Sarah; Jenkins, Edward; Davis, Simon J.; Klenerman, David; Nixon, Andrew; Pefaur, Noah; Wyatt, David; Dushek, Omer; Kasturirangan, Srinath; Dustin, Michael L.
• 署名单位: University of Oxford; Kennedy Institute for Rheumatology; Leibniz Association; Leibniz Institut fur Neue Materialien (INM); University of Oxford; Chinese Academy of Medical Sciences - Peking Union Medical College; Boehringer Ingelheim; University of Oxford; Boehringer Ingelheim; University of Oxford; UK Research & Innovation (UKRI); Medical Research Council UK (MRC); University of Oxford; University of Cambridge
• 刊物名称: PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
• ISSN/ISSBN: 0027-13818
• DOI: 10.1073/pnas.2425781122
• 发表日期: 2025-06-03
• 关键词: ligand interactions; antibody; activation; complex; tcr; accumulation; adhesion; saxs; cd2

摘要

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 CD3 epsilon 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-gamma 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).