Dynamic Covalent Bond-Assisted Programmed and Traceless Protein Release: High Loading Nanogel for Systemic and Cytosolic Delivery

Systemic and cytosolic delivery represents a grand challenge preventing many therapeutic proteins from clinical applications. Despite tremendous progresses in the past decade, most approaches generally lack the ability of triggered traceless protein release, require complicated formulation, and/or yield low protein loading. By using the protein as a crosslinker, here, a simple and general formulation affording protein nanogels (NG) with uniformed sizes and exceptionally high protein loading (>50%) is reported. By using the fine-tuned bis- and monosubstituted maleamic anhydride-amine chemistry for the crosslinking of the protein and a 4-armed PEG-MA(4), the NG is implemented with a tandem pH-programmed and traceless release character. The final NG, CDM-MA-NG, is stable under normal physiological conditions and effectively protects the crosslinked cargo cytochrome C from serum fouling, proteolytic and thermal degradation. In vitro, CDM-MA-NG exhibits a high level of cellular uptake and potent cancer cell killing only when incubated at pH 6.5, but not 7.4. Systemic administration of CDM-MA-NG leads to significantly inhibited tumor growth and extended survival rate. Given the abundance of the amine groups on protein surface, this work describes a universal platform for therapeutic protein formulation, and opens up enormous opportunities for the systemic, cytosolic, and traceless delivery of protein-based nanomedicines.