Alpha 1-antitrypsin (AAT) is normally secreted from hepatocytes and circulates in the plasma, protecting lung elastin from degradation by neutrophil elastase. Deficiency of AAT can lead to emphysema. In a subset of patients homozygous for the PI*Z mutation, liver disease may also develop, apparently related to accumulation and polymerization of the mutant protein within the endoplasmic reticulum of affected hepatocytes. Gene therapy for AAT deficiency-related emphysema requires producing high level of circulating AAT, which has been achieved by muscle and liver gene delivery in mouse models (Song et al 1998 PNAS, Song et al 2001 Gene Therapy). However, gene therapy for the liver disease requires decreasing mutant AAT and producing sufficient wt AAT. It will be beneficial for both liver and lung diseases from AAT deficiency, if we can turn the mutant mRNA into the wt mRNA of AAT. Recent studies have shown the correction of inherited skin diseases, and phenotype correction of hemophilia A mice by spliceosome-mediated RNA trans-splicing (Chao et al 2003 Nature Medicine). In this proposal, we will test the feasibility of correction of mutant AAT RNA by spliceosome-mediated RNA trans-splicing in vitro and in vivo. In the specific aim 1, we will optimize the efficiency of mutant hAAT correction in vitro. 4 pre-trans-splicing RNA molecules (PTMs) will be designed for targeting introns (1, 2, 3 or 4) of hAAT (PiZ) pre-mRNA and evaluated by plasmid transfection into transformed liver cells from PiZ transgenic mouse. In the specific aim 2, we will evaluate the effect of recombinant adeno-associated virus serotype 8 (rAAV8) mediated PTM-gene therapy in PiZ transgenic mouse model. Serum levels of wt hAAT and Z-form hAAT will be monitored. The reduction of Z-form AAT accumulation in the liver will be evaluated. Liver pathology will be examined. We anticipate achieving the therapeutic correction in PiZ transgenic mouse model. The success of this project may lead to a novel gene therapy for clinical treatment of human AAT deficiency.
| Li, Hong; Zhang, Bin; Lu, Yuanqing et al. (2011) Adipose tissue-derived mesenchymal stem cell-based liver gene delivery. J Hepatol 54:930-8 |
| Ma, H; Lu, Y; Li, H et al. (2010) Intradermal alpha1-antitrypsin therapy avoids fatal anaphylaxis, prevents type 1 diabetes and reverses hyperglycaemia in the NOD mouse model of the disease. Diabetologia 53:2198-204 |
| Li, Hong; Lu, Yuanqing; Witek, Rafal P et al. (2010) Ex vivo transduction and transplantation of bone marrow cells for liver gene delivery of alpha1-antitrypsin. Mol Ther 18:1553-8 |
| Lu, Yuanqing; Song, Sihong (2009) Distinct immune responses to transgene products from rAAV1 and rAAV8 vectors. Proc Natl Acad Sci U S A 106:17158-62 |
