Validation and analysis of 12,000 AI-driven CAR-T designs in the Bits to Binders competition.

Abstract

Artificial intelligence (AI) methods for proteins have advanced rapidly, improving structure prediction and design, particularly for de novo binders. However, most evaluations emphasize binding affinity rather than higher-order biological function. We present Bits to Binders, a global competition evaluating de novo binder design in the context of chimeric antigen receptor (CAR) T cells. Teams from 42 countries submitted 12,000 designs of 80-amino acid de novo minibinders targeting human CD20 as CAR binding domains. Designs were screened by pooled CAR-T proliferation, identifying 707 designs exhibiting significant CD20-specific enrichment, with team hit rates from 0.6% to 38.4%. Top-performing candidates were validated as individual constructs, measuring CD20-specific proliferation, expansion, cytokine production, and targeted cell lysis. We identified common design methodologies and factors correlated with DNA synthesis, expression, and target-specific T cell activation which nearly double the success rates when applied as a retrospective filter. We release this dataset as an open resource, with practical recommendations to support more effective AI-driven binder design.