Expression of neutralizing recombinant human antibodies against Varicella Zoster virus for use as a potential prophylactic.
Chickenpox is a highly infectious disease that can be life-threatening to certain groups such as the newborn of nonimmune mothers and immunocompromised patients. At present, prophylactic treatment of individuals at risk involves the use of a polyclonal antibody preparation derived from the pooled sera of hyperimmune donors. While this product is effective, there are problems associated with maintaining supply, which depends on the availability of donors, and the variation of potency between batches. An effective human monoclonal preparation would be of value by providing a well-characterized and standardized preparation available on demand. In this study recombinant human anti-varicella zoster virus (VZV) monoclonals were generated from the mRNA of unstable anti-VZV secreting heterohybridoma cell lines, and characterized according to their molecular weight, isoelectric point, glycosylation, binding to C1q, and efficacy at neutralizing VZV in vitro. In one antibody (AEVZ 5.3) the VH region was grafted from the IgG1 parent antibody onto an IgG3 backbone to determine the effect of isotype on neutralization in vitro. Antibodies were expressed from NSO cells at concentrations of 3-24 microg/mL and contained the expected heavy and light chain fragments and N-linked glycan structures. Both AEVZ 5.1 and AVEZ 4 antibodies were IgG1 and recognized the viral coat protein glycoprotein E; both showed complement-independent and complement-enhanced neutralization. Changing the isotype of AEVZ 5.1 from IgG1 to IgG3 (AEVZ 5.3) further enhanced VZV neutralization in the presence of complement, but reduced its neutralization capacity in the absence of complement. Complement enhancement was consistent with our findings that the IgG3 form could bind more molecules of C1q. The results demonstrate the successful use of recombinant methods to generate stable, functional monoclonal antibodies. Modifications of the original antibodies were made with the aim of improving functionality. The resulting cell lines could be used for large-scale production of well-characterized antibodies for therapeutic use.
Lloyd-Evans
P
P
International Blood Group Reference Laboratory, Bristol, United Kingdom.
Gilmour
J E
JE
D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000235:genetics* / Q000276:immunology*; D000956:Antigens, Viral / Q000276:immunology; D002213:Capsid / Q000276:immunology; D002460:Cell Line; D002644:Chickenpox / Q000517:prevention & control*; D015922:Complement C1q / Q000276:immunology; D006031:Glycosylation; D014645:Herpesvirus 3, Human / Q000276:immunology*; D006801:Humans; D006825:Hybridomas / Q000737:chemistry; D007074:Immunoglobulin G / Q000235:genetics / Q000276:immunology; D007132:Immunoglobulin Isotypes; D007135:Immunoglobulin Variable Region / Q000235:genetics / Q000276:immunology; D007526:Isoelectric Point; D008970:Molecular Weight; D011993:Recombinant Fusion Proteins / Q000276:immunology; D014162:Transfection; D010948:Viral Plaque Assay
D000911:Antibodies, Monoclonal; D000956:Antigens, Viral; D007074:Immunoglobulin G; D007132:Immunoglobulin Isotypes; D007135:Immunoglobulin Variable Region; D011993:Recombinant Fusion Proteins; D015922:Complement C1q