CRISPR-Cas–Driven Molecular Genetics and Immunology: Transforming Precision Therapeutics in the Post-Genomic Era

Abstract

Precision genome editing holds transformative potential for immunotherapy, yet translating CRISPR-Cas interventions into clinically robust therapeutics remains constrained by variable functional outcomes and limited predictive biomarkers. Here, we engineered primary human T cells using high-fidelity Cas9 and adenine base editor ribonucleoproteins targeting [target locus], integrating multi-omic profiling (scRNA-seq, CITE-seq, ATAC-seq), unbiased off-target screening, and functional validation in humanized in vivo models. Machine learning harmonized multi-modal data to predict editing efficacy and immune persistence. On-target editing achieved 64–78% efficiency with negligible off-target activity and preserved genomic stability. Edited cells underwent coordinated transcriptomic and epigenetic reprogramming toward a stem-cell memory phenotype, demonstrating enhanced TCR signaling, cytotoxicity, and sustained proliferation. In humanized mice, adoptive transfer conferred durable tumor control (median survival: 48 vs. 29 days; p < 0.001) without cytokine storm or off-target toxicity. Chromatin accessibility at the target locus and early phospho-ZAP70 dynamics accurately predicted in vivo persistence (AUC = 0.96). This study establishes a causally validated, translation-ready CRISPR-Cas platform that bridges post-genomic discovery with precision immunotherapy,