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Abstract

Recessive dystrophic epidermolysis bullosa (RDEB) has been defined as severe chronic skin fragility and caused by mutations in COL7A1, which encodes for the elastic structural protein type VII collagen (C7). The 8.9 Kb COL7A1 transcript is particularly a large sequence with many repeating units which makes it difficult to manipulate and package into viral systems. Therefore, the minicircle system is ideal for use with COL7A1, firstly to minimize the overall DNA construct size while secondly increasing the safety profile of the gene therapy. We successfully inserted COL7A1 into the parental plasmid MN512A1 and combined it with our highly efficient a non-viral vector (HPAE). HPAE-MC-COL7A1 polyplexes successfully produced significant levels of recombinant C7 with negligible cytotoxicity in RDEB-TA4 keratinocytes. Minimal effect was seen on primary keratinocyte metabolic health, even after multiple applications of HPAE polyplexes. Furthermore, in vivo transfection studies revealed that HPAE carrying MC-COL7A1 restores the expression of C7 along the basement membrane zone in a human RDEB graft mouse model after intradermal injection and topical application. Of the 7 animals treated, 5 were positive for recombinant C7, with all animals receiving 2 or more applications having strong positive signal. While further assessment is required to prove this approach is safe and well tolerated in the long term, HPAE-MC-COL7A1 polyplexes showed great promise as a potential therapeutic for RDEB.

Biography

Email: lara.cutlar@ucd.ie