In Vitro Fabrication of Microscale Secretory Granules

Advanced medical treatments involving drug delivery require fully biocompatible materials with the ability to release functional drugs in a time-prolonged way. Ideally, the delivered molecules should be self-contained as chemically homogenous entities to prevent the use of potentially toxic scaffolds or hold matrices. In nature, peptidic hormones are self-stored in protein-only secretory granules formed by the reversible coordination of Zn ²⁺ and histidine residues. Inspired by this concept, an in vitro transversal procedure is developed, analyzed, and comparatively applied for the fabrication of protein-only secretory granules at the microscale. These materials can be produced from any polyhistidine-tagged protein using physiological concentrations of Zn ²⁺ as a potent and versatile glue-like agent. The screening of granules formed by 12 engineered and nonengineered proteins at different Zn ²⁺ concentrations revealed optimal fabrication conditions and the consequent release profiles. Moreover, the functional and structural properties of the delivered protein are fully validated using a drug-targeting protein platform in a mouse model of human colorectal cancer. In summary, short histidine tags allow the packaging of structurally and functionally dissimilar polypeptides, which supports the proposed fabrication method as a powerful protocol extensible to diverse clinical scenarios in which slow protein drug delivery is required.