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Gene therapy for pyoderma gangrenosum: optimal transfection conditions and effect of drugs on gene delivery in the HaCaT cell line using cationic liposomes

Teagle, Alexandra R., Birchall, James Caradoc and Hargest, Rachel 2016. Gene therapy for pyoderma gangrenosum: optimal transfection conditions and effect of drugs on gene delivery in the HaCaT cell line using cationic liposomes. Skin Pharmacology and Physiology 29 , pp. 119-129. 10.1159/000444859

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Abstract

Background/Aims: Pyoderma gangrenosum (PG) is a rare ulcerative skin disease, currently treated empirically with immunosuppression. PG is a good target for gene therapy since the skin is easily accessible. This study used the FDA-approved vector Lipofectamine® 2000 to investigate in vitro transfection of skin keratinocytes. The aim was to determine an optimum transfection protocol, including the effect of drugs currently used to treat PG on the efficiency of gene transfer, since gene therapy is unlikely to be used as monotherapy. Methods: Cells of the HaCaT line were transfected with the lacZ reporter gene, and transgene expression was measured after a given time period. Conditions tested were: relative concentrations of DNA and Lipofectamine®, time from transfection to measurement of expression, pH, and exposure to clinically relevant drugs (hydrocortisone, methotrexate, infliximab). Results: The greatest levels of β-galactosidase expression were observed using a DNA:Lipofectamine® ratio of 1:5 (μg/μl) on day 3 after transfection, using culture medium at pH 7, and in the presence of hydrocortisone. Transfection efficiency was reduced by the presence of methotrexate and not significantly affected by infliximab. Conclusion: Gene therapy is a potential future strategy for the management of PG; this study is a step towards the development of a topical gene-based agent.

Item Type: Article
Date Type: Published Online
Status: Published
Schools: Pharmacy
Subjects: R Medicine > RM Therapeutics. Pharmacology
Publisher: Karger
ISSN: 1660-5527
Date of First Compliant Deposit: 29 May 2016
Date of Acceptance: 19 February 2016
Last Modified: 04 Jun 2017 09:10
URI: http://orca-mwe.cf.ac.uk/id/eprint/91327

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