Optimization and Stability of Cell–Polymer Hybrids Obtained by “Clicking” Synthetic Polymers to Metabolically Labeled Cell Surface Glycans

Abstract

Re-engineering of mammalian cell surfaces with polymers enables the introduction of functionality including imaging agents, drug cargoes or antibodies for cell-based therapies, without resorting to genetic techniques. Glycan metabolic labeling has been reported as a tool for engineering cell surface glycans with synthetic polymers through the installation of biorthogonal handles, such as azides. Quantitative assessment of this approach and the robustness of the engineered coatings has yet to be explored. Here, we graft poly(hydroxyethyl acrylamide) onto azido-labeled cell surface glycans using strain-promoted azide–alkyne “click” cycloaddition and, using a combination of flow cytometry and confocal microscopy, evaluate the various parameters controlling the outcome of this “grafting to” process. In all cases, homogeneous cell coatings were formed with >95% of the treated cells being covalently modified, superior to nonspecific “grafting to” approaches. Controllable grafting densities could be achieved through modulation of polymer chain length and/or concentration, with longer polymers having lower densities. Cell surface bound polymers were retained for at least 72 h, persisting through several mitotic divisions during this period. Furthermore, we postulate that glycan/membrane recycling is slowed by the steric bulk of the polymers, demonstrating robustness and stability even during normal biological processes. This cytocompatible, versatile and simple approach shows potential for re-engineering of cell surfaces with new functionality for future use in cell tracking or cell-based therapies.

Developing immune-regulatory materials using immobilized monosaccharides with immune-instructive properties

“…To make combinatorial libraries of different monosaccharides, a ‘grafting to’ approach was used as we have previously described [ [28] , [29] , [30] , [31] , [32] ] with minor adjustments. The method involves synthesizing a polymer to enable anchoring the monosaccharides of interest to the epoxide-functionalized polystyrene plate.…”

A Decade of Advances in Single‐Cell Nanocoating for Mammalian Cells

“…For example, after incubation of cells in an N ‐azidoacetylmannosamine (Ac 4 ManNAz)‐added culture medium, which promoted the introduction of azide groups on the glycocalyx via cellular metabolism, the cells were conjugated with alkyne‐functionalized polymers via click reaction; resulting in above 95% coating efficiency without cell damage. [ 73,74 ] Rarely, azide‐functionalized amino acid (i.e., l ‐azidohomoalanine) has been introduced to cells for metabolic protein labeling. [ 75 ] Liposome fusion could be a possible alternative to introduce tailored functional groups on the cell surface.…”

Engineering the Mammalian Cell Surface with Synthetic Polymers: Strategies and Applications

“…In a follow‐up report, it was shown that synthetic polymers installed at the surface of azide‐labeled A549 remained attached for over 72 h, even after multiple cell division cycles. [ 74 ] Grafting density was also evaluated in this study, with longer polymer chains (degree of polymerization higher than 50) showing lower grafting density as a consequence of steric hindrance.…”