LAUSR

CRISPR/Cas9-based gene therapy and photodynamic therapy both show promise for cancer treatment but still have their drawbacks limited by tumor microenvironment and long treatment duration. Herein, CRISPR/Cas9 genome editing and photodynamic strategy for a synergistic anti-tumor therapeutic modality is merged. Chlorophyll (Chl) extracted from natural green vegetables is encapsulated in Pluronic F127 (F127) micelles and Histidine-tagged Cas9 can be effectively chelated onto micelles via metal coordination by simple incubation, affording Cas9-Chl@F127 micelles. Mg2+ acts as an enzyme cofactor to correlatively enhance Cas9 gene-editing activity. Upon laser irradiation, Chl as an effective photosensitizer generates reactive oxygen species (ROS) to kill tumor cells. Meanwhile, CRISPR/Cas9, mediated by dual deliberately designed gRNAs of APE1 and NRF2, can reprogram the tumor microenvironment by increasing the intracellular oxygen accumulation and impairing the oxidative defense system of tumor cells. Cas9-Chl@F127 micelles can responsively release Cas9 in the presence of abundant ATP or low pH in tumor cells. In a murine tumor model, Cas9-Chl@F127 complexed with dual gRNAs including APE1 and NRF2 significantly inhibits the tumor growth. Taken together, Cas9-Chl@F127 micelles, representing the first Chl-based green biomaterial for the delivery of Cas9, show great promise for the synergistic anti-tumor treatment by PDT and gene editing.