Pathogenesis of disorders caused by human P450 oxidoreductase mutations

Pathogenesis of disorders caused by human P450 oxidoreductase mutations Patients with POR deficiency have disordered steroidogenesis due to malfunctioning of several key enzymes in the steroidogenesis pathway. Patients with genital abnormalities and an abnormal urinary steroid profile that suggested combined defects in CYP17A1 (17a-hydroxylase/17,20 lyase) and CYP21A2 (21-hydroxylase) were reported that suggested a role of POR, however, when the POR knockout was found to be embryonically lethal in mice POR was excluded as a candidate gene. Almost two decades later in a 2004 we reported (Flück et al, Nature Genetics, March 2004) the first four patients with mutations in POR gene. Subsequent studies confirmed the presence of POR mutations in patients with similar patterns of steroid abnormalities with and without ABS (Huang, Pandey et al. Am. J. Hum. Genetics 2005). In the patients of European descent the A287P mutation is most common; while Japanese patients often have the R457H mutation. Role of POR in steroid metabolism: In the initial report the POR mutants A287P, R457H, V492E, C569Y and V608F were tested with 17a-hydroxylase and 17,20 lyase activities of CYP17A1. A good correlation between the clinical features and the CYP17A1 activities was observed. Later on we tested the effects of some POR variants on CYP19A1 (aromatase) activity. Mutations R457H and V492E located at FAD binding site of POR caused a complete loss of CYP19A1 activity, confirming that POR mutations disrupting the FAD binding and electron transfer will severely affect all P450s. We have recently reported a novel POR mutation P399_E401del found in two unrelated Turkish families. The novel POR mutation P399_E401del had a clinical phenotype of ABS and DSD but only subclinical cortisol deficiency. In vitro functional testing of mutant POR P399_E401del on single enzymes showed a loss 68-85% activity for different P450s. Diagnosis: PORD can be diagnosed prenatally from the amniotic fluid or urine of the mother. The urine steroid profile of a mother carrying a fetus with PORD is characteristic for low levels of estriol but increased epiallopregnanediol disulfate (metabolite of pregnenolone), and increased androsterone and its precursor 17a-hydroxyallopregnanolone (5a reduced products coming from the alternative backdoor pathway leading to dihydrotestosterone production. The diagnosis of POR deficiency may be suggested from clinical and hormonal analysis but it requires molecular genetic analysis for confirmation and to know the specific defect as treatment may depend on severity of the effect on different interaction partners of POR. Role in Drug metabolism: We have found that POR mutants Y181D, A457H, Y459H, V492E and R616X lost more than 99% of drug metabolizing CYP3A4 activity, while 60-85% activity loss was observed for POR mutations A287P, C569Y and V608F [24]. Loss of CYP3A4 activity may result in increased risk of drug toxicities and adverse drug reactions in patients with severe POR mutations. Among non P450 partner proteins we have observed that POR mutants Y181D, A457H, Y459H, V492E and R616X result in total loss of heme oxygenase 1 (HO1) activity, while POR mutations A287P, C569Y and V608F lost 50-70% of HO1 activity Therapeutic options: Treatment may include replacement of glucocorticoids and sex steroids (and rarely mineralocorticoids) as assessed by low serum hormone levels. The skeletal malformations caused by POR deficiency require orthopedic management, and often reported mortality is due to skeletal abnormalities leading to respiratory problems (e.g. choanal obstruction). Computational docking studies and functional assays suggest that the activity loss of some POR variants may be rescued by flavin supplementation We were able to restore activities of POR Y181D and A287P which is not near the cofactor binding sites. However, which POR variant with what partner P450 may respond to external FMN, and whether flavin can be used as a treatment for affected patients remains to be tested. References: 1. Flück CE, Tajima T, Pandey AV, Arlt W, Okuhara K, Verge CF, Jabs EW, Mendonca BB, Fujieda K and Miller WL. Mutant P450 oxidoreductase causes disordered steroidogenesis with and without Antley-Bixler syndrome. Nature Genetics 36: 228-30 (2004). 2. Pandey AV, Flück CE, Huang N, Tajima T, Fuijeda K and Miller WL. P450 oxidoreductase deficiency: a new disorder of steroidogenesis affecting all microsomal P450 enzymes. Endocrine Res 30: 881-888 (2004). 3. Huang N, Pandey AV, Agrawal V, Reardon W, Lapunzina PD, Mowatt D, Jabs EW, vanVliet G; Sack, J, Flück CE and Miller WL. Diversity and function of mutations in P450 oxidoreductase in patients with Antley-Bixler syndrome and disordered steroidogenesis. Am. J. Hum. Genet. 76: 729-749 (2005). 5. Pandey AV. Biochemical analysis of mutations in P450 oxidoreductase. Biochem Soc Trans 34: 1186-1191 (2006). 4. Flück CE, Nicolo C & Pandey AV. Clinical structural and functional implications of mutations and polymorphisms in human NADPH P450 oxidoreductase. Fund. Clin. Pharmacol. 21: 399-410 (2007). 6. Pandey AV, Kempna P, Hofer G, Mullis PE & Flück CE. Modulation of human CYP19A1 activity by mutant NADPH P450 oxidoreductase. Mol Endocrinol 21: 2579-2595 (2007). 16. Sim SC, Miller WL, Zhong XB, Arlt W, Ogata T, Ding X, Wolf CR, Flück CE, Pandey AV, Henderson CJ, Porter TD, Daly AK, Nebert DW, Ingelman-Sundberg M. Nomenclature for Alleles of the Cytochrome P450 Oxidoreductase (POR) Gene. Pharmacogenetics and Genomics 19: 565-566 (2009) 15. Flück CE, Mullis PE and Pandey AV. Modeling of human P450 oxidoreductase structure by In-silico mutagenesis and molecular dynamics simulations. Mol Cell Endocrinol 313: 17-22 (2009). 14. Fluck CE, Pandey AV, Huang N, Agrawal, V & Miller WL. P450 oxidoreductase deficiency – a new form of congenital adrenal hyperplasia. Endocr Dev 13: 67-81 (2008) 13. Nicolo C, Flück CE, Mullis PE and Pandey AV. Restoration of mutant cytochrome P450 reductase activity by external flavin. Mol Cell Endocrinol. 321: 245-252 (2010). 12. Pandey AV, Flück CE, Mullis PE. Altered heme catabolism by heme oxygenase-1 caused by mutations in human NADPH cytochrome P450 reductase. Biochem. Biophys. Res. Commun 400: 374-378 (2010). 11. Flück CE, Mullis PE and Pandey AV. Reduction in hepatic drug metabolizing CYP3A4 activities caused by P450 oxidoreductase mutations identified in patients with disordered steroid metabolism. Biochem. Biophys. Res. Commun 401: 149-153 (2010). 10. Flück CE and Pandey AV. Clinical and biochemical consequences of P450 oxidoreductase deficiency. Endo. Dev. 20: 63-79 (2011). 9. Flück CE, Pandey AV, Dick B, Camats N, Fernandez-Cancio M, Celmente M, Gussinye M, Carrascosa A, Mullis PE and Audi L. Characterization of novel StAR (Steroidogenic Acute Regulatory Protein) mutations causing non-classic lipoid adrenal hyperplasia. PLoS One 6(5):e20178 (2011). 8. Flück CE, Mallet D, Hofer G, Samara-Boustani D, Leger J, Polak M, Morel Y and Pandey AV. Biochem. Biophys. Res. Commun 412: 572-577 (2011). 7. Flück CE, Meyer-Böni M, Pandey AV, Kempná P, Miller WL, Schoenle EJ, and Biason-Lauber A. Why Boys will be Boys: Two Pathways of Fetal Testicular Androgen Biosynthesis Are Needed for Male Sexual Differentiation. Am J. Hum Genetics 89:201-218 (2011).

Id102
Grant Value331000
Commencement Date / Completion Date1 July 2012 - UNSPECIFIED
Contributors PD Dr. Amit Vikram Pandey (Principle Investigator)
Funders [4] Swiss National Science Foundation
URIhttp://p3.snf.ch/Project-134926
KeywordsP450 oxidoreductase, POR, Steroid biosynthesis, disorders of sexual development, CYP3A4, CYP17A1, CYP19A1, CYP21A2, Heme oxygenase, Drug metabolism, Pharmacogenomics, Cytochrome P450
Publications Balmer, Cécile; Pandey, Amit Vikram; Rüfenacht, Véronique; Nuoffer, Jean-Marc; Fang, Ping; Wong, Lee-Jun; Häberle, Johannes (2014). Mutations and polymorphisms in the human argininosuccinate lyase (ASL) gene. Human mutation, 35(1), pp. 27-35. Wiley-Blackwell 10.1002/humu.22469
Hu, Liyan; Pandey, Amit Vikram; Eggimann, Sandra; Rüfenacht, Véronique; Möslinger, Dorothea; Nuoffer, Jean-Marc; Häberle, Johannes (2013). Understanding the role of argininosuccinate lyase transcript variants in the clinical and biochemical variability of the urea cycle disorder argininosuccinic aciduria. Journal of biological chemistry, 288(48), pp. 34599-611. American Society for Biochemistry and Molecular Biology 10.1074/jbc.M113.503128
Camats Tarruella, Núria; Pandey, Amit Vikram; Fernández-Cancio, Mónica; Fernández, Juan M.; Ortega, Ana M.; Udhane, Sameer Sopanrao; Andaluz, Pilar; Audí, Laura; Flück, Christa (2014). STAR splicing mutations cause the severe phenotype of lipoid congenital adrenal hyperplasia: insights from a novel splice mutation and review of reported cases. Clinical endocrinology, 80(2), pp. 191-199. Blackwell Scientific Publications 10.1111/cen.12293
Flück, Christa; Pandey, Amit Vikram (2014). P450 oxidoreductase deficiency (PORD). In: New, Maria I.; Lekarev, Oksana; Parsa, Alan; Yuen, Tony; O'Malley, Bert; Hammer, Gary (eds.) Genetic steroid disorders (pp. 125-143). Elsevier 10.1016/B978-0-12-416006-4.00010-7
Biason-Lauber, Anna; Pandey, Amit Vikram; Miller, Walter L.; Flück, Christa (2013). Marsupial pathway in humans. In: New, Maria I.; Lekarev, Oksana; Parsa, Alan; Yuen, Tony; O'Malley, Bert; Hammer, Gary (eds.) Genetic steroid disorders (pp. 215-224). New York: Elsevier 10.1016/B978-0-12-416006-4.00015-6
Bouchoucha, Nadia; Samara-Boustani, Dinane; Pandey, Amit Vikram; Bony-Trifunovic, Helene; Hofer, Gaby; Aigrain, Yves; Polak, Michel; Flück, Christa (2014). Characterization of a novel CYP19A1 (aromatase) R192H mutation causing virilization of a 46,XX newborn, undervirilization of the 46,XY brother, but no virilization of the mother during pregnancies. Molecular and cellular endocrinology, 390(1-2), pp. 8-17. Elsevier Ireland 10.1016/j.mce.2014.03.008
Hu, Liyan; Pandey, Amit Vikram; Balmer, Cécile; Eggimann, Sandra; Rüfenacht, Véronique; Nuoffer, Jean-Marc; Häberle, Johannes (2015). Unstable argininosuccinate lyase in variant forms of the urea cycle disorder argininosuccinic aciduria. Journal of inherited metabolic disease, 38(5), pp. 815-827. Springer 10.1007/s10545-014-9807-3
Parween, Shaheena; Boulez, Florence Roucher; Flück Pandey, Christa Emma; Lienhardt-Roussie, Anne; Mallet, Delphine; Morel, Yves; Pandey, Amit Vikram (2016). P450 Oxidoreductase Deficiency: loss of activity caused by protein instability from a novel L374H mutation. The Journal of clinical endocrinology and metabolism, 101(12), jc.2016-1928. The Endocrine Society 10.1210/jc.2016-1928
Zalewski, Adam; Ma, Nina S; Legeza, Balazs; Renthal, Nora; Flück Pandey, Christa Emma; Pandey, Amit Vikram (2016). Vitamin D-Dependent Rickets Type 1 Caused by Mutations in CYP27B1 Affecting Protein Interactions With Adrenodoxin. The Journal of clinical endocrinology and metabolism, 101(9), pp. 3409-3418. Endocrine Society 10.1210/jc.2016-2124
Udhane, Sameer S; Dick, Bernhard; Hu, Qingzhong; Hartmann, Rolf W; Pandey, Amit V (2016). Specificity of anti-prostate cancer CYP17A1 inhibitors on androgen biosynthesis. Biochemical and biophysical research communications, 477(4), pp. 1005-1010. Academic Press 10.1016/j.bbrc.2016.07.019
Burkhard, Fabian Z; Parween, Shaheena; Udhane, Sameer Sopanrao; Flück Pandey, Christa Emma; Pandey, Amit Vikram (2017). P450 Oxidoreductase deficiency: Analysis of mutations and polymorphisms. Journal of steroid biochemistry and molecular biology, 165(Pt A), pp. 38-50. Elsevier 10.1016/j.jsbmb.2016.04.003
Flück Pandey, Christa Emma; Pandey, Amit Vikram (2017). Impact on CYP19A1 activity by mutations in NADPH cytochrome P450 oxidoreductase. Journal of steroid biochemistry and molecular biology, 165(Pt A), pp. 64-70. Elsevier 10.1016/j.jsbmb.2016.03.031
Udhane, Sameer Sopanrao; Parween, Shaheena; Kagawa, Norio; Pandey, Amit Vikram (2017). Altered CYP19A1 and CYP3A4 Activities Due to Mutations A115V, T142A, Q153R and P284L in the Human P450 Oxidoreductase. Frontiers in Pharmacology, 8(580), p. 580. Frontiers 10.3389/fphar.2017.00580
Pandey, Amit Vikram; Henderson, Colin J; Ishii, Yuji; Kranendonk, Michel; Backes, Wayne L; Zanger, Ulrich M (2017). Editorial: Role of Protein-Protein Interactions in Metabolism: Genetics, Structure, Function. Frontiers in Pharmacology, 8(881), p. 881. Frontiers 10.3389/fphar.2017.00881
Pandey, Amit Vikram; Henderson, Colin J.; Ishii, Yuji; Kranendonk, Michel; Backes, Wayne L.; Zanger, Ulrich M. (eds.) (2018). Role of Protein-Protein Interactions in Metabolism: Genetics, Structure, Function. Frontiers in Pharmacology. Lausanne: Frontiers Media 10.3389/978-2-88945-385-6
Pandey, Amit Vikram; Flück Pandey, Christa Emma (1 April 2015). Altered Drug and Steroid Metabolism by Mutations in Human NADPH Cytochrome P450 Reductase. FASEB journal, 29(Suppl 1), p. 522. Federation of American Societies for Experimental Biology
Udhane, Sameer S; Parween, Shaheena; Pandey, Amit Vikram (2016). Aromatase Activity is Disrupted by Mutations in P450 Oxidoreductase. Hormone research in paediatrics, 86(Suppl. 1), p. 267. Karger 10.1159/000449142
Udhane, Sameer S; Pandey, Amit Vikram (1 September 2015). Effect of CYP17A1 Inhibitors Orteronel and Galeterone on Adrenal Androgen Biosynthesis (Unpublished). In: Annual Meeting of European Society for Paediatric Endocrinology ESPE.

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