ESPE Abstracts

Background: The adrenal cortex undergoes constant cellular renewal and any effect from recombinant adeno-associated vectors (rAAV) targeting differentiated cells will be transient as extra-chromosomal DNA will be lost by dividing cells. A novel option is to use rAAV to express ectopic adrenocortical enzymes in a stable organ outside the adrenal gland. Our aim was to correct the biochemical phenotype in a mouse model for congenital adrenal hyperplasia (CAH) with 21-hydroxylase expression in the liver.

Method: A 21-hydroxylase deficient (-/-) mouse model was used. An AAV8-CYP21A2 vector was produced using established protocols and administered at 5e11 vector genomes/mouse (n=10). After 4 weeks steroid profiles were measured using liquid chromatography tandem mass spectrometry. Hepatic vector copy number (vgc/diploid cell) was determined using digital droplet PCR and transgene expression was determined as a ratio of CYP21A2:albumin transcripts using quantitative PCR. Statistical analysis used the Mann-Whitney U test.

Results: The total and relative adrenal mass improved in the treated -/- mice, suggesting reduced chronic ACTH stimulation (table 1). Serum aldosterone normalised and serum corticosterone improved in the treated. Serum progesterone improved in the females but not in the males. The median vgc/diploid cell was 12.34 (males) and 13.06 (females). Despite more pronounced effect on progesterone in females, males had higher transgene expression: median 0.79 vs 0.20.

Conclusion: In a mouse model of CAH, rAAV-induced expression of 21-hydroxylase in hepatocytes completely recovered aldosterone, improved corticosterone, improved progesterone in females and reduced adrenal mass. This validates the potential of the liver as a site for ectopic adrenal enzyme expression, serving as a pre-clinical model for a potential treatment for CAH. This also demonstrates that some effect on phenotype seen by adrenal-targeting gene therapy may be due to ectopic 21-hydroxylation. Experiments are ongoing to extend this study using simultaneous dual AAV8-CYP21A2 and AAV8-CYP11B1 vectors to boost corticosterone production.