Methods and compositions for immunization against virus
Immunogenic compositions comprising partially glycosylated viral glycoproteins for use as vaccines against viruses are provided. Vaccines formulated using mono-, di-, or tri-glycosylated viral surface glycoproteins and polypeptides provide potent and broad protection against viruses, even across strains. Pharmaceutical compositions comprising monoglycosylated hemagglutinin polypeptides and vaccines generated therefrom and methods of their use for prophylaxis or treatment of viral infections are disclosed. Methods and compositions are disclosed for influenza virus HA, NA and M2, RSV proteins F, G and SH, Dengue virus glycoproteins M or E, hepatitis C virus glycoprotein E1 or E2 and HIV glycoproteins gp120 and gp41.
1. A method of manufacturing an immunogenic composition the method comprising:
(i) providing a viral glycoprotein, or immunologically active fragment thereof, which comprises a glycan bound to a glycosylation site of the viral glycoprotein, or the immunologically active fragment thereof;
(ii) removing a portion of the glycan to form a truncated glycan, which consists of one, two, or three sugar residues, to produce the partially glycosylated viral glycoprotein comprising the truncated glycan; and
(iii) formulating the partially glycosylated viral glycoprotein or immunologically active fragment thereof produced in step (ii) into an immunogenic composition, which further comprises an adjuvant.
2. The method of claim 1 , wherein the viral glycoprotein or the immunologically active fragment thereof is produced in and isolated from a eukaryotic host cell.
3. The method of claim 2 , wherein the eukaryotic host cell comprises a construct suitable for expressing the viral glycoprotein or the immunologically active fragment thereof.
4. The method of claim 2 , wherein the eukaryotic host cell is selected from the group consisting of a yeast cell, an insect cell, a mammalian cell, and a human cell.
5. The method of claim 4 , wherein the eukaryotic host cell is a HEK293E cell.
6. The method of claim 1 , wherein step (ii) is performed by incubating the viral glycoprotein or the immunologically active fragment thereof set forth in (i) with a glycosidase, a neuraminidase, an alpha-1-mannosidase, an endo F glycanase, an endo N glycanase, an endoglycosidase H (Endo H), an enzyme that selectively digests the glycan at an N- or O-linked glycosylation site, or a combination thereof.
7. The method of claim 1 , wherein step (ii) is performed by a chemical method that selectively digests the glycan at an N- or O-linked glycosylation site.
8. The method of claim 1 , wherein the viral glycoprotein is selected from the group consisting of:
influenza virus neuraminidase, influenza virus hemagglutinin, RSV F glycoprotein, RSV G glycoprotein, herpes simplex virus glycoproteins gB, gC, gD, and gE, Chlamydia MOMP and PorB antigens, Dengue virus core protein, Dengue virus matrix protein measles virus hemagglutinin, herpes simplex virus type 2 glycoprotein gB, poliovirus I VP1, envelope glycoproteins of HIV 1, hepatitis B surface antigen, pseudorabies virus g50 (gpD), pseudorabies virus II (gpB), pseudorabies virus III (gpC), pseudorabies virus glycoprotein H, pseudorabies virus glycoprotein E, transmissible gastroenteritis glycoprotein 195, transmissible gastroenteritis matrix protein, swine rotavirus glycoprotein 38, swine parvovirus capsid protein, bovine viral diarrhea glycoprotein 55, Newcastle disease virus hemagglutinin-neuraminidase, swine flu hemagglutinin, swine flu neuraminidase, infectious bovine rhinotracheitis virus glycoprotein E, infectious bovine rhinotracheitis virus glycoprotein G, infectious laryngotracheitis virus glycoprotein G, infectious laryngotracheitis virus glycoprotein I, hepatitis B virus core protein, equine influenza virus type A/Alaska 91 neuraminidase, equine influenza virus type A/Miami 63 neuraminidase, equine influenza virus type A/Kentucky 81 neuraminidase, equine herpes virus type 1 glycoprotein B, equine herpes virus type 1 glycoprotein D, bovine respiratory syncytial virus attachment protein (BRSV G), bovine respiratory syncytial virus fusion protein (BRSV F), bovine respiratory syncytial virus nucleocapsid protein (BRSVN), bovine parainfluenza virus type 3 fusion protein, bovine parainfluenza virus type 3 hemagglutinin neuraminidase, bovine viral diarrhea virus glycoprotein 48, bovine viral diarrhea virus glycoprotein 53, glycoprotein E of Dengue virus, glycoprotein E1 of human hepatitis C virus, and glycoprotein E2 of human hepatitis C virus.
9. The method of claim 1 , wherein the adjuvant is selected from the group consisting of aluminum hydroxide, aluminum phosphate, both aluminum hydroxide and aluminum phosphate, incomplete Freund's adjuvant (IFA), squalene, squalane, alum, and MF59.