Generalizable design parameters for soluble T cell receptor-based T cell engagers

Area of Science:

• Immunology
• Oncology
• Biotechnology

Background:

• T cell receptor (TCR) therapeutics target intracellular antigens, offering a unique advantage over extracellularly-focused immunotherapies.
• Recombinant TCR (rTCR)-based therapies are advancing clinically, with recent positive Phase III trials for uveal melanoma.
• The optimal molecular format for rTCR bifunctional therapeutics remains underexplored compared to antibody-based engagers.

Purpose of the Study:

• To evaluate the efficacy of various molecular formats for rTCR/anti-CD3 bifunctional therapeutics targeting intracellular tumor antigens.
• To compare the redirected lysis activity of different rTCR bifunctional formats against NY-ESO-1 and MAGE-A3 expressing tumors.

Main Methods:

• Generation of rTCR/anti-CD3 bifunctional proteins in diverse molecular formats (Fab, single-chain variable fragment, IgG-like, IgG-Fc-containing).
• Assessment of redirected T cell lysis activity against tumor cells expressing NY-ESO-1 or MAGE-A3 antigens.

Main Results:

• Potent tumor cell lysis was observed with small, tandem binding formats, specifically rTCR/anti-CD3 Fab and single-chain variable fragments.
• Larger formats, including IgG-like and IgG-Fc-containing molecules, exhibited significantly reduced or poor activity.
• The efficacy of rTCR bifunctionals appears format-dependent, with smaller constructs being superior.

Conclusions:

• Small, tandem molecular formats are optimal for achieving potent redirected T cell lysis with rTCR bifunctional therapeutics.
• The findings suggest a universal principle for rTCR bifunctional design, influenced by the TCR/human leukocyte antigen structural paradigm.
• This study provides critical insights for the rational design of next-generation TCR-based cancer immunotherapies.