Transduction of ferret airway epithelia using a pre-treatment and lentiviral gene vector

BMC Pulmonary Medicine

David Parsons¹  Tim Kuchel³  Jahan Penny-Dimri¹  Chantelle McIntyre¹  Martin Donnelley¹  Nigel Farrow¹  Patricia Cmielewski² 

[1] School of Paediatrics and Reproductive Health, University of Adelaide, Adelaide, SA 5001, Australia;Centre for Stem Cell Research, University of Adelaide, Adelaide, SA 5001, Australia;South Australian Health and Medical Research Institute, Gilles Plains, SA 5086, Australia

关键词: Gene therapy;    Cystic fibrosis;    Lentivirus;    Lung;    Ferret;   

Others  :  1091268
DOI  :  10.1186/1471-2466-14-183

 received in 2014-09-19, accepted in 2014-11-10,  发布年份 2014

【 摘 要 】

Background

The safety and efficiency of gene therapies for cystic fibrosis (CF) need to be assessed in pre-clinical models. Using the normal ferret, this study sought to determine whether ferret airway epithelia could be transduced with a lysophosphatidylcholine (LPC) pre-treatment followed by a VSV-G pseudotyped HIV-1 based lentiviral (LV) vector, in preparation for future studies in CF ferrets.

Methods

Six normal ferrets (7 -8 weeks old) were treated with a 150 μL LPC pre-treatment, followed one hour later by a 500 μL LV vector dose containing the LacZ transgene. LacZ gene expression in the conducting airways and lung was assessed by X-gal staining after 7 days. The presence of transduction in the lung, as well as off-target transduction in the liver, spleen and gonads, were assessed by qPCR. The levels of LV vector p24 protein bio-distribution in blood sera were assessed by ELISA at 0, 1, 3, 5 and 7 days.

Results

The dosing protocol was well tolerated. LacZ gene expression was observed en face in the trachea of all animals. Histology showed that ciliated and basal cells were transduced in the trachea, with rare LacZ transduced single cells noted in lung. p24 levels was not detectable in the sera of 5 of the 6 animals. The LacZ gene was not detected in the lung tissue and no off-target transduction was detected by qPCR.

Conclusions

This study shows that ferret airway epithelia are transducible using our unique two-step pre-treatment and LV vector dosing protocol. We have identified a number of unusual anatomical factors that are likely to influence the level of transduction that can be achieved in ferret airways. The ability to transduce ferret airway epithelium is a promising step towards therapeutic LV-CFTR testing in a CF ferret model.

【 授权许可】

   
2014 Cmielewski et al.; licensee BioMed Central Ltd.

【 图 表 】

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