Insulin-like Growth Factor 2 (IGF2)-Fused Lysosomal Targeting Chimeras for Degradation of Extracellular and Membrane Proteins

Targeted degradation of the cell-surface and extracellular proteins via the endogenous lysosomal degradation pathways, such as lysosome-targeting chimeras (LYTACs), has recently emerged as an attractive tool to expand the scope of extracellular chemical biology. Herein, we report a series of recombinant proteins genetically fused to insulin-like growth factor 2 (IGF2), which we termed iLYTACs, that can be conveniently obtained in high yield by standard cloning and bacterial expression in a matter of days. We showed that both type-I iLYTACs, in which IGF2 was fused to a suitable affibody or nanobody capable of binding to a specific protein target, and type-II iLYTAC (or IGF2-Z), in which IGF2 was fused to the IgG-binding Z domain that served as a universal antibody-binding adaptor, could be used for effective lysosomal targeting and degradation of various extracellular and membrane-bound proteins-of-interest. These heterobifunctional iLYTACs are fully genetically encoded and can be produced on a large scale from conventional E. coli expression systems without any form of chemical modification. In the current study, we showed that iLYTACs successfully facilitated the cell uptake, lysosomal localization, and efficient lysosomal degradation of various disease-relevant protein targets from different mammalian cell lines, including EGFR, PD-L1, CD20, and α-synuclein. The antitumor properties of iLYTACs were further validated in a mouse xenograft model. Overall, iLYTACs represent a general and modular strategy for convenient and selective targeted protein degradation, thus expanding the potential applications of current LYTACs and related techniques.