Polymer-mediated immunocamouflage of red blood cells: Effects of polymer size on antigenic and immunogenic recognition of allogeneic donor blood cells

Abstract

Developing a practical means of reducing alloimmunization in chronically transfused patients would be of significant clinical benefit. Immunocamouflaging red blood cells (RBCs) by membrane grafting of methoxypoly(ethylene glycol) (mPEG) may reduce the risk of allo-immunization. The results of this study showed that antibody recognition of non-ABO antigens was significantly reduced in an mPEG-dose-and polymer size-dependent manner, with higher molecular weight mPEGs providing better immunoprotection. Furthermore, in vivo immunogenicity was significantly reduced in mice serially transfused with mPEG-modified xenogeneic (sheep; sRBCs), allogeneic (C57Bl/6), or syngeneic (Balb/c) RBCs. Following a primary transfusion of sRBCs, mice receiving mPEG-sRBCs showed a >90% reduction in anti-sRBC IgG antibody levels. After two transfusions, mice receiving mPEG-sRBCs showed reductions of >80% in anti-sRBC IgG levels. Importantly, mPEG-modified autologous cells did not induce neoantigens or an immune (IgG or IgM) response. These data suggest that the global immunocamouflage of RBCs by polymer grafting may provide a safe and cost-effective means of reducing the risk of alloimmunization. Safe and effective blood transfusions are arguably one of the most important medical advances made in the past century. These procedures are now common place and have saved millions of lives; however, until recently, this process has been of only limited interest to the research community.There is a substantial body of evidence showing that clinical problems can occur in chronically transfused patients because of alloimmunization [1][2][3]. These problems are compounded by the fact that it is difficult, time-consuming and expensive to phenotype blood cells for mismatched antigens at non-ABO/RhD epitopes. Yet these mismatches can, and do, have a significant impact on patient care--especially for chronically transfused sickle cell and thalassemic patients. Indeed, by some estimates, up to 35% of sickle patients show clinically significant levels of alloimmunization (i.e., shortened RBC survival and mild fever) to non-ABO/RhD antigens [4,5]. Of these alloimmunized patients, some become so reactive to donor RBCs that even the most closely matched samples cause a transfusion reaction, thus endangering the life of the patient.

Microfluidic analysis of cellular deformability of normal and oxidatively damaged red blood cells

“…For the in vivo survival studies, murine RBC were labeled using a fluorescent, membrane anchored marker, PKH-26 (Sigma, St. Louis, MO) [45,46]. Equal numbers of control or oxidized RBC (40% hematocrit) were transfused as previously described [46,47]. Blood samples from the recipient mice were followed until the labeled RBC were cleared from circulation (approximately 40-50 days for allotransfusions).…”

Simultaneous Camouflage of Major and Minor Antigens on Red Blood Cell Surface With Activated mPEGs

“…Ongoing work on the modification of RBC suggests that PEGylation may be a practical method for decreasing the risk of transfusional alloimmunization, and treatment of alloimmunized patients (5,8).…”

“…However, it is difficult to identify appropriate blood donors (3)(4)(5). The chronically transfused patients, who suffer from thalassemia and sickle cell disease encounter the problems of alloimmunization (6,7).…”

“…immune cells) to them (8). Several reactive derivatives of mPEG, such as mPEG-succinimidyl valerate (mPEG-SVA) and mPEG-succinimidyl carbonate (mPEG-SC), have been employed to covalently attach mPEG to the surface of RBCs (5,8,(10)(11)(12)(13)(14)(15).…”

“…Recently, Wang et al have investigated the effect of PEGylation using mPEG-SVA on reducing the antigenic rec-ognition of blood group antigens expressed by normal human RBCs (5). Lee and Scott used mPEG-SC and mPEG-SVA for PEGylation of polystyrene beads to evaluate the charge camouflage and protein adsorption (8).…”