Membrane-Assisted Fugogenic Delivery of Ribonucleoprotein for Efficient CRISPR-Based Gene Therapy

Despite the great potential of gene therapy using CRISPR/Cas9 systems, the targeted delivery of CRISPR components such as Cas9 and sgRNA to the diseased cells, and specific intracellular localization the organs needed to provide optimal therapeutic outcome, still remains a significant challenge for clinical translation. Currently, viral vectors are the most common vehicle to transport those CRISPR components into the cell. However, they are limited in use for clinical application due to incremental risk of mutagenesis and several unknown side effects. Although the synthetic nanoparticle are promising candidates for the alternative deliverying vehicles for CRISPR-based gene therapy, a major obstacle to in vivo gene delivery is that the primary uptake pathway, cellular endocytosis, results in extracellular excretion and lysosomal degradation of genetic material. Herein, we developed a cellular membrane-derived delivery platform that securely and efficiently deliver ribonucleoprotein (RNP) complexes to target-specific cells in vitro and in vivo. Using fusogenic lipid and cancer cell-derived membrane for nanoparticle formulation as a the RNP host and tumor-targeting peptides for selective tissue homing, the RNP-loaded nanoparticles induce the membrane fusion efficiently, thus shows that the uptake mechanism can be engineered to be independent of common receptor-mediated endocytosis pathways. This platform is also demonstrated for intracellular delivery of common chemotherapeutics including cisplatin and doxorubicin. Two examples of the potential broad clinical applicability of this concept in a mouse xenograft model of colon cancer and Alzheimer’s’ disease are demonstrated and show exceptionally improved delivery efficiency and therapeutic index. Taken together, the combination of high payload capacity and fusogenic uptake yielded the promising gene editing efficacy and potential therapeutic means for clinical translation. The work presented here reports the an great advance of successful gene therapy against cancer and neurodegenerative disease.