Characterisation of llama antibody fragments able to act as HIV-1 entry inhibitors.
Doctoral thesis, UCL (University College London).
Human immunodeficiency virus type 1 (HIV-1) entry into cells is mediated by the functional envelope spike, which consists of trimers of gp120 bound to gp41. Most variants of HIV-1 enter cells through attachment of the envelope spike to the main cellular receptor CD4, allowing interaction with a co-receptor and eventually fusion of viral and cellular membranes. Neutralising antibodies inhibit HIV-1 entry by targeting epitopes on the functional spike. HIV-1 has, however, evolved several ways to evade recognition by antibodies, including variable regions, carbohydrates, and conformational masking. As a result, the neutralising antibody response in HIV-1 infection and post-immunisation is generally narrow, and only a handful of broadly neutralising monoclonal antibodies have been reported. In this thesis, the isolation and characterisation of novel, broadly neutralising antibody fragments derived from llamas is described. Llamas produce antibodies devoid of light chains, which have their antigen-binding properties confined to a single fragment, the VHH, and a preference for cleftrecognition. VHH were isolated from llamas immunised with recombinant gp120 using phage display-based methods. In order increase the chances of isolating neutralising VHH, a functional selection strategy was employed, involving a competitive elution with soluble CD4. Three VHH able to neutralise HIV-1 primary isolates of subtype B and C were characterised. These VHH bound to gp120 with high affinities and competed with soluble CD4 and antibodies to the CD4-binding site for this binding, indicating that their mechanism of neutralisation involves interacting with the functional envelope spike prior to binding to CD4. These results indicate that llama VHH can be potent HIV-1 entry inhibitors. Since VHH are stable and can be produced at a relatively low cost, they may be considered for HIV-1 microbicide development. Anti-gp120 VHH might also prove useful in defining neutralising and non-neutralising epitopes on HIV-1 envelope proteins, with implications for HIV-1 vaccine design.
|Title:||Characterisation of llama antibody fragments able to act as HIV-1 entry inhibitors|
|Open access status:||An open access version is available from UCL Discovery|
|Additional information:||Ablynx N.V., Ghent, Belgium, share the intellectual property rights to the llama antibody fragments described in this thesis, together with UCL|
|UCL classification:||UCL > School of Life and Medical Sciences > Faculty of Medical Sciences > Infection and Immunity (Division of) > Research Department of Infection|
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