AFP-specific T cell receptors with fine-tuning affinity induce durable tumor remission in mice and acquire...

T cell receptor-engineered T cell (TCR-T cell) therapies enable targeting of intracellular antigens presented by HLA molecules, addressing the limitations of solid tumors lacking surface antigens. We developed a novel alpha-fetoprotein (AFP)-specific TCR-T cell for hepatocellular carcinoma. Starting from a previously cloned AFP-specific TCR (A01) restricted by HLA-A^∗02:03, we enhanced its affinity through structure-guided optimization. A single amino acid substitution in the CDR3 region of the TCR Vβ chain significantly enhanced the antigen-specific activity. This modified TCR (A01-m14) triggered robust responses in both CD4⁺ and CD8⁺ TCR-T cells, demonstrating potent efficacy against patient-derived organoids and in murine models. Notably, CD4⁺ and CD8⁺ TCR-T cells acted synergistically to sustain antitumor effects. A01-m14 TCR exhibited a favorable safety profile with no detectable off-target reactivity against the human proteome or unrelated HLA molecules. The further engineered TCR (A01-m58) was able to recognize most HLA-A^∗02 subtypes, significantly expanding the eligible patient population. Meanwhile, the pan-A^∗02-TCR-T cells also showed enhanced cytokine production, superior tumor cell killing, and achieved complete tumor eradication in murine models. This study developed an AFP-specific TCR-T cell with fine-tuning affinity, demonstrating remarkable preclinical efficacy and safety, alongside a sequence-optimized version with pan-HLA-A^∗02 applicability, offering broader patient access and therapeutic versatility.