Newton, Lydia Sarah;
(2024)
Design of PROTACs to counteract cyclophilin proviral activity.
Doctoral thesis (Ph.D), UCL (University College London).
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Abstract
The cyclophilin (Cyp) family of human proteins are cofactors for many viruses including human immunodeficiency virus (HIV-1) and hepatitis C virus (HCV). In the case of HIV-1, cyclophilin A (CypA) binds an exposed proline-rich loop on the viral capsid, regulating capsid surface dynamics and protein interactions including shielding the virus from restriction factor tripartite motif-containing protein 5α (TRIM5α). For HCV, CypA interacts with viral non-structural protein 5A (NS5A), facilitating evasion of immune responses and enhancing viral RNA replication by a poorly understood mechanism. We designed and characterised two fully synthetic proteolysis targeting chimeras (PROTACs), CG167 and RJS308, targeting CypA. These Cyp-PROTACs are based on our novel sanglifehrin A (SfA)-derived Cyp inhibitor TWH106, which is structurally distinct to the Cyp inhibitor cyclosporine A (CsA). I demonstrate that these Cyp-PROTACs degrade CypA in a dose-dependent manner via a PROTAC mechanism. They are selective for CypA over other closely related Cyps, including CypB, illustrated using proteomics experiments. Selectivity may be explained by differing subcellular locations of Cyps and/or kinetics of ternary complex formation governed by Cyp sequence. I demonstrate that Cyp-PROTACs are antiviral against HIV-1 spreading infection in primary human T cells and HCV replicon replication in Huh7 cells. Importantly, CypA degradation facilitates more potent antiviral activity compared to the non-PROTAC parental inhibitor TWH106, at limiting inhibitor concentrations. This host-targeting approach promises high antiviral potency, reduced evolution of viral resistance, and broad efficacy against unrelated viruses. Finally, I utilise the PROTACs to investigate the role of CypA in inflammatory signalling using a lipopolysaccharide (LPS) model of inflammation. These Cyp-PROTACs provide a proof of concept that targeting host proteins using PROTAC technology is a viable antiviral strategy. Furthermore, they provide useful tools to probe cyclophilin functions in health and disease.
| Type: | Thesis (Doctoral) |
|---|---|
| Qualification: | Ph.D |
| Title: | Design of PROTACs to counteract cyclophilin proviral activity |
| Open access status: | An open access version is available from UCL Discovery |
| Language: | English |
| Additional information: | Copyright © The Author 2024. Original content in this thesis is licensed under the terms of the Creative Commons Attribution-NonCommercial 4.0 International (CC BY-NC 4.0) Licence (https://creativecommons.org/licenses/by-nc/4.0/). Any third-party copyright material present remains the property of its respective owner(s) and is licensed under its existing terms. Access may initially be restricted at the author’s request. |
| UCL classification: | UCL UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Medical Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Medical Sciences > Div of Infection and Immunity |
| URI: | https://discovery.ucl.ac.uk/id/eprint/10200989 |
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