Knockdown of Z mutant alpha-1 antitrypsin in vivo using modified DNA antisense oligonucleotides

Abstract

Alpha-1 antitrypsin (AAT) is a serum protease inhibitor, mainly expressed in and secreted from hepatocytes, important for regulating neutrophil elastase activity among other proteases. Various mutations in AAT cause alpha-1 antitrypsin deficiency (AATD), a rare hereditary disorder that results in liver disease due to accumulation of AAT aggregates and lung disease from excessive neutrophil elastase activity. PiZ transgenic mice contain the human AAT genomic region harboring the most common AATD mutation, the Glu342Lys (Z) point mutation. These mice effectively recapitulate the liver disease exhibited in AATD patients, including AAT protein aggregates, hepatocyte death, and eventual liver fibrosis. Previously, we demonstrated that modified antisense oligonucleotides (ASOs) can dramatically reduce Z-AAT RNA and protein levels in PiZ mice enabling inhibition, prevention, and reversal of the associated liver disease. Here, we describe in detail usage of AAT-ASOs to knock down Z-AAT in PiZ mice with a focus on preparation and in vivo delivery of ASOs, as well as detailed workflows pertaining to the analysis of Z-AAT mRNA, plasma protein, and soluble/insoluble liver protein levels following ASO administration.