Browsing by Author "Gent, Rachelle"

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    The metabolism and receptor interactions of C11-oxy steroids: implications for hormonal homeostasis and pathophysiology
    (Stellenbosch : Stellenbosch University, 2024-03) Gent, Rachelle; Swart, Amanda C. ; Snoep, Jacky L. ; Kaschula, Catherine H.; Stellenbosch University. Faculty of Science. Dept. of Biochemistry.
    ENGLISH ABSTRACT: This study describes: • the determination of the kinetic parameters of the 11,B-hydroxysteroid dehydrogenase type 1 (11,BHSD1) and type 2 (11,BHSD2) isozyme conversion of C11-oxy C21 steroids, 11,B- hydroxyprogesterone (11,BOHP4) and 11-ketoprogesterone (11KP4), and C11-oxy C19 steroids, 11,B-hydroxyandrostenedione (11OHA4) and 11-ketoandrostenedione (11KA4), and 11,B- hydroxytestosterone (11OHT) and 11-ketotestosterone (11KT). • the in vitro conversion of C11-oxy C21 steroids to C11-oxy C19 steroids by cytochrome P450 17a-hydroxylase (CYP17A1) present novel mechanisms whereby metabolites of the C11-oxy C21 backdoor pathway may contribute to androgen excess. The conversion of 11,BOHP4 to 21- deoxycortisol (21dF) and 11OHA4 as well as the conversion of 11KP4 to 21-deoxycortisone (21dE) and 11KA4 are catalysed by the 17a-hydroxylation and 17,20-lyase activity of CYP17A1 in vitro. • the utilisation of cell models (PNT2, VCaP, LNCaP and C4-2B) to demonstrate the possible role of C11-oxy steroids in prostate cancer (PCa) pathophysiology. Assays showed an increase in 11,BHSD2 activity in LNCaP (androgen-dependent castration resistant prostate cancer (CRPC)) and C4-2B (metastatic CRPC) cells compared to PNT2 (normal prostate) and VCaP (PCa model) cell lines. This demonstrates the role of 11,BHSD2 in PCa directing flux to the biosynthesis of 11KT and 11KDHT, potentially contributing to CRPC. In addition, an increase in 11KT production in LNCaP cells is directly associated with an increase in prostate specific antigen. • the in vitro metabolism of the 11,BHSD2 inhibitor, 11a-hydroxyprogesterone (11aOHP4), by CYP17A1 which resulted in the biosynthesis of novel C11a-metabolites, 11a,17a- dihydroxyprogesterone (11a,17a-diOHP4) and 11a-hydroxyandrostenedione (11aOHA4), suggesting the potential presence of novel in vivo C11-oxy pathways. • the pre-receptor regulation of the androgen receptor (AR), progesterone receptor A (PRA) and progesterone receptor B (PRB) by the 11,BHSD catalysed interconversion of the C11-oxy steroids. Specific C11-oxy metabolites act as potential agonists or antagonists of these steroid receptors while also suggesting regulatory roles for 5a-reductase (SRD5A), 17,B-hydroxysteroid dehydrogenase (17,BHSD), 3a-hydroxysteroid dehydrogenase (3aHSD) as well as CYP17A1. • the development and validation of three novel ultra-high performance supercritical fluid chromatography tandem mass spectrometry (UHPSFC-MS/MS) methods for the separation and quantification of C11-oxy C19, C11-oxy C21 and adrenal steroids. The application of these methods in the comprehensive steroid analyses of southern white rhinoceros (Ceratotherium simum simum) faecal samples and of type 2 diabetes (T2D) patient serum samples implicate homeostatic dysregulation of the C11-oxy steroid metabolites in pathophysiologies.