Development of an RNAi therapeutic for alpha-1-antitrypsin liver disease
Christine I. Wooddell, Keith Blomenkamp, Ryan M. Peterson, Vladimir M. Subbotin, Christian Schwabe, James Hamilton, Qili Chu, Dawn R. Christianson, Julia O. Hegge, John Kolbe, Holly L. Hamilton, Maria F. Branca-Afrazi, Bruce D. Given, David L. Lewis, Edward Gane, Steven B. Kanner, Jeffrey H. Teckman
Christine I. Wooddell, Keith Blomenkamp, Ryan M. Peterson, Vladimir M. Subbotin, Christian Schwabe, James Hamilton, Qili Chu, Dawn R. Christianson, Julia O. Hegge, John Kolbe, Holly L. Hamilton, Maria F. Branca-Afrazi, Bruce D. Given, David L. Lewis, Edward Gane, Steven B. Kanner, Jeffrey H. Teckman
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Research Article Hepatology Therapeutics

Development of an RNAi therapeutic for alpha-1-antitrypsin liver disease

Abstract

The autosomal codominant genetic disorder alpha-1 antitrypsin (AAT) deficiency (AATD) causes pulmonary and liver disease. Individuals homozygous for the mutant Z allele accumulate polymers of Z-AAT protein in hepatocytes, where AAT is primarily produced. This accumulation causes endoplasmic reticulum (ER) stress, oxidative stress, damage to mitochondria, and inflammation, leading to fibrosis, cirrhosis, and hepatocellular carcinoma. The magnitude of AAT reduction and duration of response from first-generation intravenously administered RNA interference (RNAi) therapeutic ARC-AAT and then with next-generation subcutaneously administered ARO-AAT were assessed by measuring AAT protein in serum of the PiZ transgenic mouse model and human volunteers. The impact of Z-AAT reduction by RNAi on liver disease phenotypes was evaluated in PiZ mice by measuring polymeric Z-AAT in the liver; expression of genes associated with fibrosis, autophagy, apoptosis, and redox regulation; inflammation; Z-AAT globule parameters; and tumor formation. Ultrastructure of the ER, mitochondria, and autophagosomes in hepatocytes was evaluated by electron microscopy. In mice, sustained RNAi treatment reduced hepatic Z-AAT polymer, restored ER and mitochondrial health, normalized expression of disease-associated genes, reduced inflammation, and prevented tumor formation. RNAi therapy holds promise for the treatment of patients with AATD-associated liver disease. ARO-AAT is currently in phase II/III clinical trials.

Authors

Christine I. Wooddell, Keith Blomenkamp, Ryan M. Peterson, Vladimir M. Subbotin, Christian Schwabe, James Hamilton, Qili Chu, Dawn R. Christianson, Julia O. Hegge, John Kolbe, Holly L. Hamilton, Maria F. Branca-Afrazi, Bruce D. Given, David L. Lewis, Edward Gane, Steven B. Kanner, Jeffrey H. Teckman

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Figure 2

Sustained RNAi-mediated reduction of Z-AAT prevented liver disease and reversed polymer accumulation in adult PiZ mice.

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Sustained RNAi-mediated reduction of Z-AAT prevented liver disease and r...
Male PiZ mice, 11–17 weeks old, were given IV injections Q2W of 8 mg/kg ARC-AAT (8 mg/kg ARC-AAT API + 4 mg/kg EX1), 12 mg/kg RNAi-Control plus 6 mg/kg EX1, 12 mg/kg ARC-AAT API alone, or saline (n = 4–12) for 30–31 weeks and were euthanized for evaluation 2 weeks after the final injection. Untreated baseline mice were euthanized at the start of the study. (A) Plasma was collected at the indicated times and Z-AAT protein measured, shown as the group mean relative to pretreatment. (B and C) The amounts of monomeric and polymeric Z-AAT protein in liver lysates were measured by semiquantitative Western blotting, shown relative to baseline for individual animals and as the group means ± SEM. (D) Representative H&E liver sections from PiZ mice at baseline or following 33 weeks of saline (age-matched control) or ARC-AAT treatment. Asterisks indicate globules; arrowheads point to foci of inflammatory cells. Scale bar: 20 µm. (E–G) Z-AAT globules in PAS-D–stained liver sections were quantified in 3 fields of view for each animal (n = 8–12) for the number of globules/field of view (E), the size of globules (F), and the area of the specimens containing globules (G). (H and I) The number of inflammatory foci and area of liver specimens containing inflammatory foci are compared for baseline, saline-injected, and ARC-AAT–treated mice. Means are shown with SEM. Comparisons between groups (B, C, and E–I) were performed using nonparametric Wilcoxon’s rank sum test.