Immunocamouflage: the biophysical basis of immunoprotection by grafted methoxypoly(ethylene glycol) (mPEG)

Development of novel approaches for the immunomodulation of donor cells would have significant utility in transfusion and transplantation medicine. Immunocamouflage of cell surfaces by covalently grafted methoxypoly(ethylene glycol) (mPEG) (PEGylation) has emerged as a promising approach. While previous studies demonstrated the in vitro and in vivo efficacy of immunocamouflaged allogeneic blood cells, the biophysical mechanisms of immunoprotection have not been well-defined due to the fragility of intact cells. To overcome this limitation, polystyrene beads (1.2 and 8.0 microm) were used to elucidate the biophysical effects of polymer size, density and linker chemistry on charge camouflage and protein adsorption. These findings were correlated with biological studies using red blood cells and lymphocytes. Charge camouflage of both beads and cells was best achieved with long polymers. However, protein adsorption studies demonstrated an unexpected effect of target size. For 1.2 microm beads, decreased protein adsorption was best achieved with short (2 kDa) polymers whereas long chain (20 kDa) polymers were optimal for 8.0 microm particles. The biophysical findings correlated well with biological immunocamouflage as measured by particle electrophoresis and the inhibition of antibody-antigen (CD3, CD4 and CD28) recognition. Moreover, it was observed that antigen topography (CD28 vs. CD4) was of significance in selecting the appropriate polymer size. The biophysical interactions of PEGylated surfaces and macromolecules involve complex mechanisms dependent on the molecular weight, grafting concentration, target size and surface complexity. Cellular PEGylation strategies must be customized to account for target cell size, membrane complexity and antigen density and height.