Young, Emily Rose; (2022) Delivery of disease responsive miniintronic plasmids with receptor targeted nanoparticles for rheumatoid arthritis. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

Rheumatoid arthritis (RA) is an autoimmune disorder of the joint characterised by immune cell infiltration, synovial hyperplasia and neovascularisation, leading to cartilage and bone degradation. Current treatments risk severe immunosuppression and full disease remission is rare. Local gene therapy promises to overcome this, providing high therapeutic concentrations at the disease site with lower systemic concentrations. Optimal vectors should be delivered systemically to target all affected joints simultaneously, but should be specific to the inflamed joint. Our approach optimised receptor targeted nanoparticles (RTN) for plasmid DNA delivery to fibroblast-like synoviocytes (FLS), particularly aggressive cells in RA pathogenesis. Inclusion of an integrin targeting peptide achieved optimal transfection efficiency in cultured FLS and the PEGylated cationic lipid, ME42, stabilised RTN formulations in solutions without compromising transfection efficiency. However, delivery in vivo did not result in transfection and further optimisation is necessary. Additionally, mini-intronic plasmids (MIPs) were investigated for their ability to provide high and sustained transgene expression. When delivered with RTNs, these vectors transfected FLS as or more efficiently than conventional plasmid DNA (pDNA), dependent on cell type and promoter. Further, MIPs encoding a luciferase transgene driven by an NF-kB responsive promoter could upregulate luciferase expression in response to inflammatory cues in vitro, which may increase the specificity of the therapy. This system could drive expression of the therapeutic dimeric TNF receptor in primary human FLS, highlighting its potential as a human gene therapy. Interestingly, RTN transfection in FLS activated inflammatory signalling, as measured by transgene expression from the NF-kB promoter and IL-6 expression. Both liposomes and pDNA were required to drive this response, but delivering either alone had no effect. Further characterisation of this response is required. Taken together, this work provides the basis for a targeted gene therapy for RA.

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