Graham, Carl

English

Understanding how viruses and antibodies evolve in response to one another can help inform future vaccines and therapies. This project investigated longitudinal samples from individuals infected with HIV-1 and SARS-CoV-2, the causative viruses of AIDS and COVID-19 respectively.Multiple lineages of antibodies that can neutralise a wide range of HIV-1 variants have been previously isolated. They target conserved epitopes on the HIV-1 env glycoprotein, the only virally encoded antigen presented on the viral surface. Understanding how bnAbs develop during natural infection could inform future vaccination strategies. Initially, this project sought to isolate bnAbs from a HIV-1 infected individual across multiple timepoints, which required the optimization of numerous methods. These included biotinylated antigenic bait production, PBMC staining, FACS sorting and a function monoclonal antibody library generation pipeline. During this project, antibodies capable of cross-binding the gp120s of two HIV-1 global panel viruses were isolated. Through site-directed mutagenesis and other molecular cloning techniques, insights were gained into the role of the V3 and V1 loops in viral escape in donor SJU.With the arrival of the COVID-19 pandemic, the focus of this project was shifted to the antibody response to SARS-CoV-2. As such, many of the aforementioned methods were updated for work with the novel virus. Next, the polyclonal neutralising antibody responses of subjects infected with SARS-CoV-2 was investigated longitudinally. This revealed that neutralising titres sharply declined throughout the first 3-months following infection and that disease severity had an impact on the magnitude of the response. To understand these responses at a monoclonal level, B-cells were sorted from three SARS-CoV-2 infected donors and a library of 108 monoclonal antibodies was generated. Characterization of this library revealed that Spike specific antibodies had low levels of somatic hypermutation and that neutralisers could be separated into 7 competition groups. Further, it was shown that particular competition groups were unable to neutralise the B.1.1.7 variant of concern as a result of specific mutations within the receptor binding domain and NTD. To ascertain whether vaccination with ChAdOx1-nCoV resulted in a similar antibody response to natural infection, 44 SARS-CoV-2 specific monoclonal antibodies were isolated from a single donor. All 7 competition groups were found to be present and an additional NTD-specific group was discovered. Despite low levels of plasma neutralisation, the monoclonal antibodies isolated form this subject showed higher levels of SHM, increased potency and better breadth (against a panel of 5 variants of concern) compared to natural infection monoclonal antibodies. IgG spike specific B-cells were still measurable in this donor 9-months post-boost, at which time plasma neutralisation was undetectable. The conclusions of this work have highlighted the importance monitoring antibody and B-cell decline following SARS-CoV-2 infection and vaccination. Furthermore, it has highlighted sites of vulnerability on the SARS-CoV-2 spike and how these are impacted by viral escape mutations

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Neutralising antibody responses against HIV-1 and SARS-CoV-2

https://kclpure.kcl.ac.uk/portal/files/230647983/2023_Graham_Carl_1313273_ethesis.pdf

Neutralising antibody responses against HIV-1 and SARS-CoV-2

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