Development of DARPin T cell engagers for specific targeting of tumor-associated HLA/peptide complexes

The balance between affinity and specificity in T cell receptor (TCR)-dependent targeting of HLA-restricted tumor-associated antigens presents a significant challenge for immunotherapy development. T cell engagers that circumvent these limitations are therefore of particular interest. We established a process to generate bispecific designed ankyrin repeat proteins (DARPins) that simultaneously target HLA-I/peptide complexes and CD3e. These high-affinity T cell engagers elicited CD8<sup>+</sup> T cell activation against tumor targets with strong peptide specificity, as confirmed by X-scanning mutagenesis and functional killing assays. A cryo-EM structure of the ternary DARPin/HLA-A∗0201/NY-ESO1<sub>157-165</sub> complex revealed a rigid, concave DARPin surface spanning the full length of the peptide-binding cleft, contacting both α-helices and the peptide. The present findings reveal promising immuno-oncotherapeutic approaches and demonstrate the feasibility of rapidly developing DARPins with high affinity and specificity for HLA/peptide targets, which can be readily combined with a new generation of anti-CD3e-specific DARPins.

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Publication Details

Journal
iScience
Published
2025-11-05
DOI
https://doi.org/10.1016/j.isci.2025.113926
Primary Topic
Monoclonal and Polyclonal Antibodies Research
Type
article
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article

Development of DARPin T cell engagers for specific targeting of tumor-associated HLA/peptide complexes

Sandra Bruckmaier, Marcel Walser, Denis Villemagne, Tim Schulte et al.
iScience
Monoclonal and Polyclonal Antibodies Research
article

Development of DARPin T cell engagers for specific targeting of tumor-associated HLA/peptide complexes

Sandra Bruckmaier, Marcel Walser, Denis Villemagne, Tim Schulte, Stefanie Fischer, Nicole Pina, Adnane Achour, Filip Radom, Evren Alici, Nadir Kadri, Xiao Han, Tanja Hospodarsch, Tatyana Sandalova, Benedict J. Chambers, Andreas Cornelius, María Emilia Paladino, Marta Carroni, Sandra Müller, Victor Levitsky, R. Sun, Karin Wallden, Hans‐Gustaf Ljunggren, Tom Resink, Natalia Venetz-Arenas
article en

Abstract

The balance between affinity and specificity in T cell receptor (TCR)-dependent targeting of HLA-restricted tumor-associated antigens presents a significant challenge for immunotherapy development. T cell engagers that circumvent these limitations are therefore of particular interest. We established a process to generate bispecific designed ankyrin repeat proteins (DARPins) that simultaneously target HLA-I/peptide complexes and CD3e. These high-affinity T cell engagers elicited CD8<sup>+</sup> T cell activation against tumor targets with strong peptide specificity, as confirmed by X-scanning mutagenesis and functional killing assays. A cryo-EM structure of the ternary DARPin/HLA-A∗0201/NY-ESO1<sub>157-165</sub> complex revealed a rigid, concave DARPin surface spanning the full length of the peptide-binding cleft, contacting both α-helices and the peptide. The present findings reveal promising immuno-oncotherapeutic approaches and demonstrate the feasibility of rapidly developing DARPins with high affinity and specificity for HLA/peptide targets, which can be readily combined with a new generation of anti-CD3e-specific DARPins.

iScienceVol. 28(12)
Karolinska University Hospital (SE), Stockholm University (SE), Science for Life Laboratory (SE), Karolinska Institutet (SE), Molecular Partners (Switzerland) (CH), Integrated Cardio Metabolic Centre (SE)
Openalex Percentile: Top 53%
Monoclonal and Polyclonal Antibodies Research
DOIOpen Access PDFOpenAlex
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