Glycosylated Cell‐Penetrating Poly(disulfide)s: Multifunctional Cellular Uptake at High Solubility

The glycosylation of cell‐penetrating poly(disulfide)s ( CPD s) is introduced to increase the solubility of classical CPD s and to achieve multifunctional cellular uptake. With the recently developed sidechain engineering, CPD s decorated with α ‐ d ‐glucose (Glu), β ‐ d ‐galactose (Gal), d ‐trehalose (Tre), and triethyleneglycol ( TEG ) were readily accessible. Confocal laser scanning microscopy images of HeLa Kyoto cells incubated with the new CPD s at 2.5 μ m revealed efficient uptake into cytosol and nucleoli of all glycosylated CPD s, whereas the original CPD s and TEG ylated CPD s showed much precipitation into fluorescent aggregates at these high concentrations. Flow cytometry analysis identified Glu‐ CPD s as most active, closely followed by Gal‐ CPD s and Tre‐ CPD s, and all clearly more active than non‐glycosylated CPD s. In the MTT assay, all glyco‐ CPD s were non‐toxic at concentrations as high as 2.5 μ m . Consistent with thiol‐mediated uptake, glycosylated CPD s remained dependent on thiols on the cell surface for dynamic covalent exchange, their removal with Ellman 's reagent DTNB efficiently inhibited uptake. Multifunctionality was demonstrated by inhibition of Glu‐ CPD s with d ‐glucose ( IC 50 ca . 20 m m ). Insensitivity toward l ‐glucose and d ‐galactose and insensitivity of conventional CPD s toward d ‐glucose supported that glucose‐mediated uptake of the multifunctional Glu‐ CPD s involves selective recognition by glucose receptors at the cell surface. Weaker but significant sensitivity of Gal‐ CPD s toward d ‐galactose but not d ‐glucose was noted ( IC 50 ca . 110 m m ). Biotinylation of Glu‐ CPD s resulted in the efficient delivery of streptavidin together with a fluorescent model substrate. Protein delivery with Glu‐ CPD s was more efficient than with conventional CPD s and remained sensitive to DTNB and d ‐glucose, i.e ., multifunctional.