Tracing novel androgen pathways: deciphering the role of 16α-hydroxylation in human fetal biology

Informations projet

SHOXY

N° de convention de subvention: 101023999

DOI

10.3030/101023999

Projet clôturé

Date de signature de la CE 22 Mars 2021

Date de début 1 Septembre 2021

Date de fin 31 Août 2023

Financé au titre de

• EXCELLENT SCIENCE - Marie Skłodowska-Curie Actions

Coût total

€ 191 149,44

Contribution de l’UE

€ 191 149,44

Coordonné par

UNIVERSITAET BERN
Switzerland

Periodic Reporting for period 1 - SHOXY (Tracing novel androgen pathways: deciphering the role of 16α-hydroxylation in human fetal biology)

Période du rapport: 2021-09-01 au 2023-08-31

Résumé du contexte et des objectifs généraux du projet

Steroid hormones play crucial roles in human fetal development as they bind and activate receptors which regulate key biological processes. These steroid hormones are also important after birth. During fetal development, a specific class of steroid hormones, termed androgens, are produced by the fetal adrenal and metabolised by the fetal liver and placenta. This biological route from the fetal adrenal through the fetal liver to the placenta is especially important, as an imbalance in androgen levels are implicated in endocrine-related disorders and diseases that present in the fetal and neonatal stages.

In the fetal liver, an enzyme termed cytochrome P450 3A7, CYP3A7, catalyses the production of unique hydroxylated androgens. This same process is also mediated by CYP3A4 in neonates, as a switch from the predominant CYP3A7 to CYP3A4 activity occurs after birth, and while both hepatic enzymes metabolise androgens, only a few downstream hydroxylated androgen metabolites are currently known.

The project “Tracing novel androgen pathways in human fetal biology: SHOXY” aimed to identify novel hydroxylated androgens and to characterise their metabolic pathway in human fetal biology. The characterization of these androgens and their downstream metabolites could prove integral to our understanding of the transition from the fetal to the neonatal stage and endocrine-related disorders that present at these stages. Ultimately, this project permitted an in-depth investigation into the hepatic metabolism of androgens specific to human fetal development, with an expected far-reaching impact on fetal and neonatal endocrinology.

The scientific objectives of this Marie Skłodowska-Curie Action (MSCA) were to investigate (a) the biosynthesis of hydroxylated androgens by hepatic CYP3A enzymes; (b) the efficiency of these conversions by CYP3A7 compared to CYP3A4; and (c) the conversions in in vitro cell models and systems. The overarching goal of this action was to combine steroid science, protein chemistry and analytical chemistry to describe the role of androgens in fetal development!

The training objectives of this MSCA were to (a) enhance the bioinformatic and analytical skills of the fellow, together with the networking skills of the fellow through the establishment of national and international collaborations; and (b) transfer the knowledge of adrenal steroidogenesis and mass spectrometric analysis of steroid metabolites from the fellow to the host institution, in the form of establishing analytical methods.

Travail effectué depuis le début du projet jusqu’à la fin de la période considérée dans le rapport et principaux résultats atteints jusqu’à présent

SHOXY utilised recombinant proteins, transiently and stably transfected immortalised cells, in vitro cell models and human liver microsomes to investigate the metabolism of adrenal 11Oxy-androgens. 11Oxy-androgens are present in placental tissue, fetal cord blood and neonatal serum, and could have a particular role during fetal development. The metabolism of the 11Oxy-androgens in the fetal unit, therefore, presents as a unique group of androgens which were investigated in SHOXY.

State-of-the-art liquid chromatography high resolution- and gas chromatography-mass spectrometry and nuclear magnetic resonance platforms were employed to quantify and identify steroid products. Tailor-made steroid conversion assays and analytical methods were developed to investigate the conversion of 11Oxy-androgens and classical androgens catalysed by CYP3A enzymes.

Work was conducted via 7 work packages (WPs). WP1 to WP3 comprised the production and identification of novel androgen metabolites, which were biosynthesised in conversion assays and identified using mass spectrometry. These WPs yielded 3 conference publications, 3 publications and 2 literature review manuscripts to date, with additional conference publications and original research articles underway. The fellow surpassed these WPs by also including in silico molecular docking in the investigations conducted. WP4 sought to validate the findings in WP1 to WP3, by investigating steroid conversions in in vitro cell models and systems, which will also contribute to the forthcoming manuscripts, and the project was managed under WP5. Researcher training and transfer-of-knowledge were conduced under WP6, during which the fellow attended numerous training workshops and conferences, and contributed her knowledge to research and publications of national and international research groups, which also included the mentoring of early career researchers and master and doctoral students. She was appointed an Early Career Reviewer for the Endocrinology journal, earned a certificate of advanced studies in Applied Data Science and assisted in securing a European Society for Pediatric Endocrinology (ESPE) Research Unit Grant (as co-investigator) between Switzerland, Denmark and Finland. In WP7, results and findings from this project were presented at 3 international conferences, 2 research days and 2 invited guest lectures to date, while public engagement activities were also fulfilled.

Progrès au-delà de l’état des connaissances et impact potentiel prévu (y compris l’impact socio-économique et les conséquences sociétales plus larges du projet jusqu’à présent)

SHOXY has expanded our knowledge on 11Oxy-androgen metabolism in human fetal biology as this project characterised the metabolism of 11Oxy-androgens by hepatic CYP3A enzymes and identified novel hydroxylated 11Oxy-androgen metabolites. Importantly, the data show the metabolism of 11Oxy-androgens by CYP3As is less efficient compared to classical androgens. Therefore, if overproduced, 11Oxy-androgens would not be efficiently metabolized by CYP3A7 in the fetal stage nor by CYP3A4 in the neonatal stage, underscoring the bioavailability of the 11Oxy-androgens in hyperandrogenic endocrine disorders.

Hyperandrogenic endocrine disorders (excess androgens), presents as congenital adrenal hyperplasia and/or disorders/differences of sex development at birth, with devastating consequences also later in life. Classical androgens have long been understood as the only androgens to which these disorders can be linked, however, the 11Oxy-androgens, now have a specific role to play. The research conducted during this project have and will shed new light on androgen metabolism during fetal development, and clearly underscores the 11Oxy-androgens as exceptional signalling molecules in human fetal biology.

In addition, the analytical methods that were developed during this project lay the blueprint for future steroidomic studies and will contribute to multiple collaborative research endeavours in the future, with an anticipated impact on the understanding and explanation of various endocrine-linked diseases and disorders, including premature adrenarche, polycystic ovary syndrome and 21-hydroxylase deficiency.

Moreover, this project provided research independence to the fellow and has generated numerous follow-up research questions that will be pursued in the near future with far reaching benefits on the research career of the fellow.

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