Background T cell engagers (TCEs) are a promising class of targeted therapies and recent clinical successes have shown the strong potential of TCEs to drive robust response rates and increase T cell infiltration in solid tumours.¹ Intracellular targets presented by HLA provide an opportunity to build next generation TCEs. Among these intracellular targets, PRAME is a highly expressed target presented by HLA-A*0201 or HLA-A*2402 across multiple cancers. We here present our discovery and development of a half-life extended TCE against HLA-A2- and HLA-A24-restricted PRAME peptides. We have developed a novel TCE drug format, based on a solubilised TCR that is engineered as an asymmetric Fc-based bispecific, showing high potency and safety in pre-clinical evaluation.

Methods We have performed T cell repertoire analysis of 100s of donors to identify the most potent TCRs from CD8 T cells against PRAME A2 and PRAME A24. This has resulted in the isolation of PRAME-specific TCRs. We have then taken the most potent TCRs (low micromolar affinity) through affinity engineering using a proprietary approach that combines machine learning-guided mutagenesis with high-throughput functional screening. A library of novel asymmetric TCE formats with engineered single chain TCRs and an anti-CD3 engager arm were tested for stability and function using T cell:target cell co-culture assays. Finally, lead TCEs were assessed for efficacy, safety, longevity & biodistribution in ex vivo primary human cell assays and humanised xenograft models.

Results We have created a half-life extended TCE in a novel format with specific picomolar binding affinity that results in the activation of T cells and the subsequent killing of cancer cells in primary T cell assays. We observed no T cell activation or killing of primary normal cell panels. Our machine learning-guided affinity engineering process took an unprecedented 5 months to go from uM to pM binding TCRs with an affinity enhancement of >100,000 fold. We have identified a novel TCE lead candidate which is potent, specific and safe in ex vivo human samples with a large therapeutic window. Furthermore, the TCE showed long circulating properties and efficacy in humanised xenograft models.

Conclusions We have developed a novel half-life extended TCE format utilising unique TCR engineering that allows for the formation of asymmetric Fc-based anti-CD3 T cell engagers targeting PRAME A2 and PRAME A24. Currently, we are finalising the preclinical evaluation and expect to be able to enter IND enabling studies in early 2025.

Reference

1. Paz-Ares L, Champiat S, Lai WV, Hiroki I, Govindan R, Boyer M, Hummel H, Hossein B, Johnson M, Steeghs N, Blackhall F, Dowlati A, Reguart N, Yoshida T, He K, Gadgeel S, Felip E, Zhang Y, Pati A, Minocha M, Mukherjee S, Goldrick A, Nagorsen D, Sadraei NH, Owonikoko TK. Tarlatamab, a First-in-Class DLL3-Targeted bispecific T-Cell Engager, in recurrent small-cell lung cancer: an open-label, phase I study. J Clin Oncol. 2023;41:2893–2903.

Ethics Approval This study was approved by the South Central - Berkshire Research Ethics Committee; approval number 22/SC/0025 and the NHSBT; approval number NCI 2367.