Abstract
Since the isolation of the HIV-1 virus in 1983 our growing knowledge of basic virology has lead to multiple treatment strategies, turning a previously rapid terminal infection into a chronic manageable disease. A protective vaccine is urgently required to stem the rate of new HIV infections globally, however an immunogen that induces such a response has not been identified.Anti-HIV-1 broadly neutralising antibodies (bnAbs) are produced by 10-30% of HIV-1 infected individuals. These antibodies target conserved epitopes found across multiple virus clades and have shown to be protective when passively transferred to macaques in challenge studies. Utilising the longitudinal biobanked plasma samples from the SPARTAC trial cohort, I present a systematic screening of 50 patients to identify patients producing HIV-1 bnAbs and epitope mapping to determine the site on the viral envelope that the antibodies are binding. In depth analysis of two patients that were determined to be targeting the N-linked glycosylation site at position 332 and V3 loop (N332/V3) was performed.
Over 200 functional env genes from single RNA genomes were amplified and cloned into expression vectors from multiple time points throughout the course of infection. Cloned env genes were sequenced and aligned to look for mutation accumulations overtime from their most recent common ancestors. Previously characterised bnAbs and autologous plasma samples were used to identify which epitopes were expressed on patient Env’s overtime. Through this analysis we determined that patient 22 was producing a contemporaneous neutralisation immune response, whereby circulating viruses were expressing Env’s that had failed to escape from the neutralising antibodies circulating in sera. Patient 22 was subsequently superinfected around 133 weeks post infection despite the presence of bnAbs, offering a unique case study for vaccine design/strategy. Upon superinfection the patient’s viral load increases considerably, however we were unable to isolate any recombination events between the two viral vpu/env genes.
Patient 29 followed a more textbook case of viral escape to circulating sera antibodies; nonetheless the Env’s isolated were extremely resistant to multiple bnAbs that target multiple different epitopes on the Env despite expressing their known epitopes. This shows the need for immunity against a wide range of neutralizing epitopes across the HIV-1 Env for a vaccine to be effective. In this patient we identified a small subset of env that appears from 185 weeks post infection that utilise the CXCR4 co-receptor for viral entry instead of CCR5. This is achieved through deletion of N-linked glycosylation sites and amino acid mutations in the V3 loop. Upon the isolation of Env’s that utilise CXCR4 the patients circulating CD4+ T cell count declines rapidly and the patient progresses to AIDS. We also utilised these patient derived Env’s to look at the sensitivity to Interferon-induced transmembrane (IFITM) proteins 1, 2 and 3. We see that Env’s that show the highest resistance to these interferon induced restriction factors co-inside with peaks in circulating viral load implying that they may play a role in regulating viral load. Through these two patient case studies we have identified that neutralising antibodies that target the N332/V3 epitope can have very distinctive consequences on viral env evolution. They also provide great case studies for vaccine design strategies whilst highlighting the difficulties that still have to be addressed by the field.
| Date of Award | 1 Aug 2019 |
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| Original language | English |
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| Supervisor | Katherine Doores (Supervisor) |