Confinement of Therapeutic Enzymes in Selectively Permeable Polymer Vesicles by Polymerization-Induced Self-Assembly (PISA) Reduces Antibody Binding and Proteolytic Susceptibility.

Covalent PEGylation of biologics has been widely employed to reduce immunogenicity, while improving stability and half-life . This approach requires covalent protein modification, creating a new entity. An alternative approach is stabilization by encapsulation into polymersomes; however this typically requires multiple steps, and the segregation requires the vesicles to be permeable to retain function. Herein, we demonstrate the one-pot synthesis of therapeutic enzyme-loaded vesicles with size-selective permeability using polymerization-induced self-assembly (PISA) enabling the encapsulated enzyme to function from within a confined domain. This strategy increased the proteolytic stability and reduced antibody recognition compared to the free protein or a PEGylated conjugate, thereby reducing potential dose frequency and the risk of immune response. Finally, the efficacy of encapsulated l-asparaginase (clinically used for leukemia treatment) against a cancer line was demonstrated, and its biodistribution and circulation behavior was compared to the free enzyme, highlighting this methodology as an attractive alternative to the covalent PEGylation of enzymes.