R17 Drives Heparan Sulfate Clearance via Endolysosomal Trafficking

Heparan sulfate (HS) proteoglycans are information‐rich macromolecules that can orchestrate extracellular signaling and cargo uptake across diverse cellular contexts. Ligands that can engage cell surface HS possess significant potential as probes to investigate or manipulate cell–matrix interactions. Here, we show that R17, an HS‐binding protein from the rodent herpesvirus Peru, actively remodels the glycocalyx of pancreatic cancer cells. Recombinant R17 bound heparin in vitro and associated with the surface of pancreatic cancer cells to promote the dose‐dependent clearance of HS via trafficking to lysosomes. R17 also reduced wound closure without detectable cytotoxicity, indicative of the ability of R17 to suppress cellular migration. Notably, HS depletion persisted upon cation‐independent mannose‐6‐phosphate receptor knockdown, suggesting that clearance is independent of this mechanism. These findings identify R17 as an exogenous HS‐binding protein that can drive HS clearance via endolysosomal trafficking and suggest an alternative approach to modulate HS‐dependent functions.