Versatile hollow Ca²⁺-phenolic nanoparticles for intracellular delivery of diverse bioactive molecules and CRISPR-Cas9 genome editing.

Abstract

The development of versatile nanoplatforms capable of universally encapsulating diverse bioactive molecules holds significant promise in biomedicine. In this study, size-tunable hollow Ca²⁺-tannic acid (TA) nanoparticles (HCT NPs) are synthesized as universally applicable drug nanocarriers by simply adding TA into amorphous calcium carbonate nanoparticles. The formation of HCT NPs is identified as a surface-protected self-etching process. A wide range of hydrophobic and hydrophilic small-molecule drugs, metal ions, and biomacromolecules including proteins and nucleic acids can be encapsulated in HCT NPs for efficient intracellular delivery. HCT NPs show rapid and efficient endosomal escape, which is crucial for maintaining the bioactivity of biomacromolecules. Remarkably, a wide array of cargo proteins, spanning different molecular weights and isoelectric points can be delivered into the cytosol by HCT NPs without compromising their bioactivities. The therapeutic potential of HCT NPs for intracellular cargo delivery is exemplified by cytosolic delivery of Cas9 plasmids and Cas9 ribonucleoprotein (RNP) for CRISPR-Cas9 genome editing both in vitro and in vivo. The facile and ultrafast synthesis, versatile cargo encapsulation capabilities, efficient cell uptake and endosomal escape, and excellent biocompatibility make HCT NPs a prominent candidate for intracellular delivery of diverse bioactive molecules, particularly in therapeutic applications such as genome editing.