Spatiotemporally regulated mitochondrial genome editing via enzyme and NIR-activated CRISPR/Cas9 nanoplatform.

Mitochondrial DNA (mtDNA) mutations play critical roles in tumor progression and metabolic reprogramming. Controllable gene editing within tumor cell mitochondria remains a challenge due to the double-membrane barrier and the lack of tumor-selective activation. Herein, we report a dual-responsive CRISPR/Cas delivery platform (UCRP-TPP) that enables spatiotemporally regulated mtDNA editing for targeted tumor therapy. This nanoplatform integrates near infrared light-responsive upconversion nanoparticle (UCNP), an apurinic endonuclease 1 (APE-1)-responsive DNA complex, and a mitochondrial-targeting ligand (TPP), ensuring selective activation and mitochondrial release of Cas9/sgRNA complexes. Upon activation by endogenous APE-1 enzyme and exogenous NIR light, UCRP-TPP induces mtDNA editing by CRISPR/Cas, which leads to mtDNA copy number reduction, mitochondrial membrane depolarization, reactive oxygen species generation, and tumor cell apoptosis. In vivo studies further confirm the robust antitumor efficacy of the UCRP-TPP-based nanoplatform. This work presents a versatile and controllable mitochondrial gene-editing strategy.