Immunogenicity of Influenza Virus Vaccine Is Increased by Anti-Gal-Mediated Targeting to Antigen-Presenting Cells

Abstract

This study describes a method for increasing the immunogenicity of influenza virus vaccines by exploiting the natural anti-Gal antibody to effectively target vaccines to antigen-presenting cells (APC). This method is based on enzymatic engineering of carbohydrate chains on virus envelope hemagglutinin to carry the ␣-Gal epitope (Gal␣1-3Gal␤1-4GlcNAc-R). This epitope interacts with anti-Gal, the most abundant antibody in humans (1% of immunoglobulins). Influenza virus vaccine expressing ␣-Gal epitopes is opsonized in situ by anti-Gal immunoglobulin G. The Fc portion of opsonizing anti-Gal interacts with Fc␥ receptors on APC and induces effective uptake of the vaccine virus by APC. APC internalizes the opsonized virus to transport it to draining lymph nodes for stimulation of influenza virus-specific T cells, thereby eliciting a protective immune response. The efficacy of such an influenza vaccine was demonstrated in ␣1,3galactosyltransferase (␣1,3GT) knockout mice, which produce anti-Gal, using the influenza virus strain A/Puerto Rico/8/34-H1N1 (PR8). Synthesis of ␣-Gal epitopes on carbohydrate chains of PR8 virus (PR8 ␣gal ) was catalyzed by recombinant ␣1,3GT, the glycosylation enzyme that synthesizes ␣-Gal epitopes in cells of nonprimate mammals. Mice immunized with PR8 ␣gal displayed much higher numbers of PR8-specific CD8 ؉ and CD4؉ T cells (determined by intracellular cytokine staining and enzyme-linked immunospot assay) and produced anti-PR8 antibodies with much higher titers than mice immunized with PR8 lacking ␣-Gal epitopes. Mice immunized with PR8 ␣gal also displayed a much higher level of protection than PR8 immunized mice after being challenged with lethal doses of live PR8 virus. We suggest that a similar method for increasing immunogenicity may be applicable to avian influenza vaccines.

“Lost sugars” — reality of their biological and medical applications

“…Similar results were obtained for the influenza virus [68]. The inactivated virus was treated in vitro with recombinant α1,3GalT to synthesize α-gal epitopes.…”

“…Antigenic peptides are then presented on MHC class I and MHC class II that lead to the activation of CD8 + and CD4 + T cells, respectively. CD4 + T cells help B cells to produce the corresponding antibodies and also help activated CD8 + T cells to become cytotoxic T lymphocytes [68,69]. The immunogenicity of vaccines is increased by administering them in the form of immune complexes with the relevant IgG antibody because the interaction of FcγR (receptor) on APCs with the Fc of the IgG molecule induces internalisation.…”

“…The acceptor substrate for the enzyme was primarily LacNAc on the envelope glycoprotein hemagglutinin. For this reason, it was speculated that recombinant hemagglutinin expressing α-gal could substitute for vaccines containing inactivated influenza virus [68].…”

The Galα1,3Galβ1,4GlcNAc-R (α-Gal) epitope: A carbohydrate of unique evolution and clinical relevance

“…After being immunized with xenograft tissue such as pig kidney membrane homogenate, KO mice produce anti-Gal in titers comparable to those found in human serum. Immunization of KO mice with 1μg of inactivated flu virus vaccine expressing α-gal epitopes, administered twice in 2 weeks interval, resulted in ∼100 fold higher anti-flu virus antibody and T cell response in comparison to the immune response in KO mice immunized with the original inactivated flu virus lacking α-gal epitopes [114]. Moreover, intranasal infection (challenge) of the immunized KO mice with live virus and monitoring their survival for one month, demonstrated 11% mortality in mice immunized with the vaccine expressing α-gal epitopes, vs. 89% mortality in KO mice immunized with vaccine lacking this epitope [114].…”

“…Thus, α-gal epitopes could be added to all carbohydrate chains of HA using recombinant α1,3GT [113,114]. To illustrated that this is possible, we cloned the α1,3GT gene from a New World monkey cell line [67], produced the recombinant enzyme in the yeast Pichia pastoris expression system [115], and used it to glycosylate inactivated flu virus.…”