Immunoaffinity adsorption for extracorporeal removal of xenoreactive IgM
Hyperacute rejection following xenogeneic transplantation of pig organs to primates is mediated by naturally occurring anti-Gal IgM antibodies. Removal of these anti-αGal IgM antibodies by plasmapheresis followed by immunoaffinity adsorption on bead columns containing synthetic Gal epitopes, prevents hyperacute rejection but requires two pieces of complex equipment. In this study, we explored the potential use of immunoaffinity adsorption with hollow fiber microporous or dialysis membranes to which a synthetic αGal trisaccharide ligand was covalently attached. Ligand was immobilized on the entire accessible area of the microfiltration membrane (inner, outer, and internal surface), and all surfaces of the fiber were capable of selectively binding the xenoreactive antibody. Blood was pumped through the lumen of the hollow fibers, and plasma flowed under the action of the imposed hydrostatic pressure difference through the walls of the membrane where the xenoreactive antibody was bound. The purified plasma could then be recombined with the outlet blood. These devices would allow the use of whole blood and high flow rates. Covalent attachment of ligand directly to the surface of the membrane produced negligible binding, but use of long-chain polyamines as reactive spacers yielded binding densities as high as 80 mg/mL membrane volume of anti-Gal antibodies in break-through curve experiments with microporous nylon membranes having an internal surface area of 4.2 m2/ml membrane volume. Operation in crossflow with whole blood produced a plasma filtrate flux of 0.035 ml/(min cm2). Thus, a crossflow microfilter fabricated from the membranes would be able to carry out plasma separation and immunoadsorption in a single device with a large excess of binding capacity to ensure that all plasma that filters across the device and is returned to the patient is essentially free of anti-αGal IgM. This procedure has been successfully used in baboon in vivo studies for the first time. In the studies the levels of both xenoreactive IgM and IgG were reduced close to zero in vivo with no adverse reactions (such as complement activation) in the experimental animals. We conclude that immunoaffinity removal of xenoreactive antibodies using microfiltration hollow fiber membranes is feasible and has potential advantages of efficiency and simplicity for clinical application. Hollow-fiber affinity technology shows great promise as a platform technology for xenotransplantation, and possibly for other applications as well, such as treatment of autoimmune disease, removal of β2 microglobulin and other toxic molecules from whole blood.