Abstract
The Z variant (E342K) of α1-antitrypsin (AT) polymerizes and accumulates in hepatocyte endoplasmic reticulum (ER) resulting in neonatal hepatitis and liver cirrhosis. The secretory defect leaves lungs vulnerable to elastolysis and early-onset emphysema. Prevention of Z-AT polymerization can be achieved in vitro by targeting strand 4a of the AT molecule. Here we evaluate whether Ac-TTAI-NH2 (4M) would inhibit Z-AT polymerization in vivo. HEK293 cells transfected with Z-AT exhibited a significantly reduced AT secretion compared to M-AT cells; p=<0.001, due to retention of Z-AT in inclusion bodies. Electron microscopy demonstrated distension of rough ER in Z-AT cells in keeping with aggregation of Z-AT polymers. ELISA and Western blot with an anti-polymer antibody detected Z-AT polymers exclusively in inclusion bodies of Z-AT cells, 2468±145ng/ml, p=<0.001. 4M significantly reduced Z-AT polymers in inclusions in a dose-dependent manner; 5μg, 2181.7±26.2ng/ml p=0.006; 10μg, 1576±164.7ng/ml p=0.001 and 20 and 30μg completely prevented polymer formation, p=<0.001. Supernatant AT concentrations and anti-elastase activity from Z-AT cells was restored by 20μg 4M compared to Z-AT, p=<0.001. Functional activity of secreted AT after treatment with 4M was confirmed by its ability to form an SDS-stable complex with elastase as shown by immunoblot. These findings are the first evidence that inhibitors of Z-AT polymerization can prevent its cellular accumulation and restore plasma levels in vivo, and as such is a major step towards treatment of patients with Z-AT related lung and liver disease.
- © 2011 ERS
