Sustained miRNA-mediated Knockdown of Mutant AAT With Simultaneous Augmentation of Wild-type AAT Has Minimal Effect on Global Liver miRNA Profiles

Department of Pediatrics and Gene Therapy Center, UMass Medical School, Worcester, Massachusetts 01605, USA.
Molecular Therapy (Impact Factor: 6.23). 03/2012; 20(3):590-600. DOI: 10.1038/mt.2011.292
Source: PubMed


α-1 antitrypsin (AAT) deficiency can exhibit two pathologic states: a lung disease that is primarily due to the loss of AAT's antiprotease function, and a liver disease resulting from a toxic gain-of-function of the PiZ-AAT (Z-AAT) mutant protein. We have developed several recombinant adeno-associated virus (rAAV) vectors that incorporate microRNA (miRNA) sequences targeting the AAT gene while also driving the expression of miRNA-resistant wild-type AAT-PiM (M-AAT) gene, thus achieving concomitant Z-AAT knockdown in the liver and increased expression of M-AAT. Transgenic mice expressing the human PiZ allele treated with dual-function rAAV9 vectors showed that serum PiZ was stably and persistently reduced by an average of 80%. Treated animals showed knockdown of Z-AAT in liver and serum with concomitant increased serum M-AAT as determined by allele-specific enzyme-linked immunosorbent assays (ELISAs). In addition, decreased globular accumulation of misfolded Z-AAT in hepatocytes and a reduction in inflammatory infiltrates in the liver was observed. Results from microarray studies demonstrate that endogenous miRNAs were minimally affected by this treatment. These data suggests that miRNA mediated knockdown does not saturate the miRNA pathway as has been seen with viral vector expression of short hairpin RNAs (shRNAs). This safe dual-therapy approach can be applied to other disorders such as amyotrophic lateral sclerosis, Huntington disease, cerebral ataxia, and optic atrophies.

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    • "ess - ing of the miRNA from the RHO transcript is likely to reduce the level of mature mRNA either by interfering with splicing or polyadenylation . Nevertheless , delivery of artificial miRNAs with a resistant replacement gene has been success - fully employed in a mouse model of liver failure associated with the PiZ allele of a - 1 antitrypsin ( Mueller et al . 2012 ) . O ' Reilly and colleagues attempted the RNA replacement approach in transgenic mice bear - ing a P23H RHO gene with limited success ( O ' Reilly et al . 2007 ) . Their AAV serotype 5 vec - tor contained a pol III promoter driving the expression of an shRNA that they showed to be effective in suppressing human rhodopsin mRNA in trans"
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