Short-fibre modification facilitates adenovirus-mediated gene delivery in rheumatoid arthritis synoviocytes and synovium: role of RGD and non-RGD binding integrins

Objective

Adenovirus 5 (Ad5) receptors other than Cocksackie-ad receptor have not been well explored in rheumatoid arthritis (RA) human fibroblast-like synoviocytes (FLS). Defining these receptors may enable development of improved Ad5 vectors to specific synovial targets. As Cocksackie-ad receptor is absent in RA FLS, we investigated the efficacy and receptor usage of fibre-modified Ad5 vectors for gene delivery in vitro and ex vivo.

Methods

Short fibre modified vectors with seven fibre repeats (Ad5GFP-R7-knob, Ad5GFP-R7-RGD) were compared with wild-type (WT) 22 repeats (Ad5GFP-FiWT) and GFP expression measured in RA human FLS and a synovium explant model. Integrin expression was determined by flow cytometry. Receptor usage was examined by competition binding assays with viral penton base (PB) WT, RGD and LDV-mutant peptides, and antibodies against PB and integrins. Viral neutralization by synovial fluid was examined.

Results

Ad5GFP-R7-knob gene transduction was 25-fold higher than Ad5GFP-R7-RGD and 40-fold to 50-fold higher than Ad5GFP-FiWT in FLS. Ad5GFP-R7-knob gene transduction was reduced with PB, RGD and LDV, and antibodies against PB and αv, αvβ3, αvβ5, and β1 integrins. Ad5GFP-R7-knob transduction was increased by tumour necrosis factor and IL-1, remained 30-fold to 40-fold higher than Ad5GFP-FiWT in the presence of synovial fluid and preferentially transduced synovium explant outgrowth/tissue.

Conclusion

Ad5GFP-R7-knob has broadened viral tropism superior to RGD-modified and WT vectors, mediated by integrin-binding RGD and non-RGD motifs in viral PB capsomers in RA FLS. Ad5 short fibre modifications utilize tropism of the native PB to optimise integrin usage as both attachment and endocytotic receptors. This may be a useful strategy of enhancing gene delivery in RA, reducing vector dose and induction of inflammation and toxicity.