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Vaccine-induced bovine antibodies to HIV-1 Env

© 2016 Dr. Behnaz HeydarchiAn important feature of a potential vaccine against human immunodeficiency virus (HIV) will be the production of broadly neutralising antibodies (BrNAbs) capable of blocking infectivity of a diverse array of HIV strains. BrNAbs naturally arise in some HIV infected individuals after several years of infection and their serum immunoglobulin G (IgG) can neutralise various HIV strains across different subtypes. We previously showed that vaccination of cows with HIV Glycoprotein (Gp) 140 AD8 trimers resulted in a high titre of serum IgG specific for HIV envelope (Env) that had strong BrNAb activity. These polyclonal BrNAbs concentrated into the colostrum during the late stage of pregnancy and can be harvested in vast quantities immediately after calving. In this study, we investigated the effect of prolonged vaccination of cows with HIV Gp140 on IgG with HIV Env-binding and BrNAb activity in the colostrum-derived IgG over two subsequent pregnancies for 3 colostrum batches over a 4 year period. Overall, long-term vaccination resulted in a sustained high level of Env-specific IgG titre in colostrum batches, however repeated vaccination led to a maintained high titre of HIV Env specific IgG in the colostrum batches, but this did not increase through repeated cycles. Colostrum IgG from all colostrum batches also strongly competed with soluble CD4 (sCD4) binding to Gp140 Env trimer and with human-derived monoclonal VRC01 and b12 BrNAbs that bind the CD4 binding site (CD4bs). This result showed that high IgG titre /avidity of anti-CD4bs antibodies against b12 and VRC01 epitopes was achieved during the first year of vaccination and thereafter sustained through ensuing years of repeated vaccinations in the cow tested. Although IgG of subsequent colostrum batches may have a higher avidity towards the CD4bs, the overall breadth in neutralisation was not enhanced. This implies that the boosting vaccinations over 4 years elicited a polyclonal antibody response that maintained the proportion of both neutralising and non-neutralising CD4bs antibodies. We then aimed to investigate the molecular characteristics of anti-HIV bovine monoclonal antibodies (mAbs). AD8 HIV specific memory B cells were single sorted by Flow cytometry and the variable (V) genes were cloned to construct human/bovine chimeric or fully bovine mAbs. HIV AD8 specific mAbs were characterised and genomic analysis of heavy V genes was performed. Alanine (Ala) mutagenesis was carried out to investigate the critical residues for antigen binding activity. Among 42 positive clones from the initial screen, only 2 mAbs (6A and 8C) showed high affinity binding to Gp140 Env. Characterisation of these 2 mAbs revealed that they are capable of binding to soluble AD8 trimeric Gp140 and SOS-IP Gp140 but not monomeric Gp120. 6A and 8C showed inhibition activity against sCD4 binding to Gp140 Env, however they did not display neutralising activity. The somatic mutation rate of the variable heavy (VH) region was 27% and 25% for 6A and 8C, respectively. The third heavy complementarity determining region (CDRH3) in 6A mAb was 21aa in length with 57% somatic mutations while 8C CDRH3 was 14aa long with 93% somatic mutations compared to their germline genes. These features are consistent with previous studies of bovine derived mAbs, and mirror some of the extraordinary features that exist in some human BrNAbs such as the presence of Cys and aromatic residues. The HIV-specific binding activities of both 6A and 8C mAbs were eliminated when CDRH3 cysteine (Cys) and tryptophan (Trp) amino acids were mutated to Ala. Our analyses thus showed that the bovine immune system can elicit anti-HIV antibodies with similar properties to human BrNAbs in terms of the CDRH3 size and the apparent level of somatic mutation during affinity maturation. Thus, ongoing vaccination of the cow successfully sustained production of HIV specific antibodies. However, the first vaccination round was sufficient to induce strong cross-clade neutralisation activity against a broad range of viral strains. The results also showed that bovine vaccination with soluble Gp140 was able to induce antibodies against epitopes that exist on the SOS-IP Env trimer that closely replicates the native unliganded form of Env present on infectious virion. However, using SOS-IP Gp140 as vaccine or during the mAb selection steps may increase the chance of eliciting and isolating neutralising mAbs. The results of this study showed that the bovine immune system is a powerful antibody production system that is capable of producing anti-HIV neutralising antibodies in response to vaccination with Env Gp140 trimer. These BrNAbs can be collected in large quantities in colostrum and in a sustained manner over multiple rounds of vaccination and pregnancy