IMPE Abstracts

Background: Congenital adrenal hyperplasia (CAH) is due to defects of steroid synthetic enzymes, which includes microsomal and mitochondrial P450s. P450 enzymes need coenzymes respectively. Microsomal P450s include 21-hydroxylase need P450 oxidoreductase which coded by POR. 21-hydroxylase deficiency (21-OHD), in which CYP21A2 is mutated or deleted, is the most common cause of CAH and results in underproduction of glucocorticoids and mineralocorticoids, and overproduction of androgens. Patients with CAH are treated with oral steroid supplementation, but optimal control of blood steroid levels remains difficult. Thus, new therapeutic approaches are still needed. We succeeded gene therapy for model mice of CAH and iPS cell from CAH patients. Intravenous adeno-associated virus-mediated administration of human CYP21A2 into adult 21-OHD patients has been reported. In this study, we examined the effects of induction of coenzyme genes with an AAV vector into a model mouse.

Methods: 21OHD mice were made by breeding with heterozygous of H-2aw18 haplotype mice and treated with subcutaneous corticosteroid injection until 3 weeks of age. Type 9 of adenoassociated virus vector (AAV9) containing Cyp21a cDNA (AAV9-Cyp21a) was constructed with pAAV-CMV-shuttle. AAV9 containing Por cDNA (AAV9-Por) was also made with same package. These vectors were injected into limbs muscles of a 21OHD mouse. Serum progesterone (P4) and deoxycorticosterone (DOC) concentrations were measured before and every 4 weeks after injection. P4 and DOC concentrations were measured by LC-MS/MS.

Results: An AAV9-Cyp21a1 and AAV9-Por injected mouse showed improved steroid synthesis up to 7 months after AAV9 induction. Comparing with a mouse injected only AAV9-Cyp21a1, significant reduction of P4/DOC was observed after 4 weeks of injection but there was no difference in the effectiveness of Cyp21a1 and Por combined treatment at the end of observation.

Conclusion: These results indicate that extra-adrenal induction of Cyp21a1 and Por may ameliorate steroid metabolism in 21-OHD model mice. Although this coenzyme combined therapy showed some effects, but no long-term improvement of the efficacy exhibited. Coenzymes of P450s are necessary for their activity, but our results suggest that the coenzyme combined gene therapeutic strategies are needed to be improved.