Title Promoter Affiliations Abstract "Tissue specific modulation of the effects of glucocorticoids by androgens and the androgen receptor: revealing the mechanisms of cross tal" "Frank Claessens" "Laboratory of Molecular Endocrinology, Clinical and Experimental Endocrinology, Universiteit Leiden" "Androgens and glucocorticoids are steroid hormones with very different roles in (patho-)physiology. While androgens are known as the male steroids with anabolic effects, glucocorticoids are catabolic hormones with a key role in metabolic processes that govern whole-body energy metabolism. Clinically, long term use of androgens results in increased bone and muscle formation, while glucocorticoids induce osteoporosis and muscle loss. By contrast, our recent data indicate that androgens enhance the effects of glucocorticoids on metabolism in liver and adipous tissue.  Androgens and glucocorticoids act via the activation of the androgen receptor (AR) or the glucocorticoid receptor (GR), respectively. (AR, GR). The latter are ligand-controlled transcription factors with very similar mechanisms of action, even having overlapping sets of target genes. This leaves their differential physiological effects unexplained. Combining our knowhow on androgen and glucocorticoid action mechanisms with state-of-the-art genetic models, we will first determine the role of androgens and AR in the modulation of glucocorticoid effects in muscle (where androgens have opposing effects) and in liver (where they have potentiating effects). Secondly, at the level of these target organs, we want to zoom in at the transcriptome and chromatin level to discover how AR and GR interfere with each other’s mechanisms of actions. This is a fundamental research topic, but deeper knowledge on how androgens and glucocorticoids interfere will inspire translational work, for example on how to mitigate the effects of long term glucocorticoid and antiandrogen treatments on metabolism.   " "Differential effects of androgens and estrogens ont he hyperthrophy and atrophy signals in the aging male skeletal muscle" "Wim Derave" "Department of Movement and Sports Sciences" "This project aims to investigate the effects of androgens and estrogens on the atrophy and hypertrophy signalling pathways in skeletal muscle of young and aging males. According to the hypothesis, the decline in adrogens in the aging male activates atrophy signals leading to sarcopenia, whereas increased androgen levels will activate the hypertrophy pathways." "Classical and non-classical actions of androgens and their receptor" "Frank Claessens" "Laboratory of Molecular Endocrinology, Nephrology and Renal Transplantation Research Group, Urogenital, Abdominal and Plastic Surgery, Clinical and Experimental Endocrinology, Woman and Child" "Androgens have their well-known classical role in the development and homeostasis of the classical androgen target organs like the male reproductive organs and accessory sex glands. They also have many pleiotropic effects on non-classical organs like bone, muscle, kidney and brain. This C1 application brings together molecular biologists, endocrinologists, nephrologists and urologists as an interdisciplinary study group, with special interests in both classical and non-classical target effects. The focus of this grant application is on the molecular biology of the androgen receptor and androgens (WP1), their actions on the musculoskeletal system and kidney (WP2), their possible role in the calcification paradox in kidney disease and the molecular effects of androgen deprivation in high-risk prostate cancer (WP3)." "On the function of maternal yolk androgens in birds: from early adjustments to post-hatching conditions to long-term effects." "Wendt Müller" "Behavioural Ecology & Ecophysiology" "The effects of maternal hormones in birds are thought to represent examples of so-called maternal effects and are thought to have evolved to translate the environmental conditions experienced by the mother into adaptive phenotypic variation of the offspring. Their adaptive significance is therefore likely to depend on the post-hatching conditions, but long-lasting changes in offspring phenotype need to be considered too." "Androgen Actions: from Bench to Clinic (A2B2C)." "Frank Claessens" "Laboratory of Molecular Endocrinology, Woman and Child, Clinical and Experimental Endocrinology, Urogenital, Abdominal and Plastic Surgery" "Androgens and estrogens are crucial for maintaining sexual function, germ cell development and accessory sex glands, but they also affect many aspects of skeletal muscle, the skeleton, adipose tissue, hematopoiesis, metabolism etc. At the intracellular level, androgens act via binding to the nuclear androgen receptor (AR) or act as precursors for estrogens which bind estrogen receptors. We study three intertwined aspects of sex hormone action: - Sex steroids are carried to their target tissues via the bloodstream where they can bind sex hormone binding globulin (SHBG). We are defining the role of free and SHBG bound hormone levels in preclinical models as well as in endocrine disease. - Anti-androgens are widely used in the clinic to treat metastatic castration-resistant forms of prostate cancer. We study the AR in preclinical models and clinical stages of prostate cancer that are resistant against the latest AR targeting drugs. - We also investigate the direct and indirect effects of androgens on the musculoskeletal system by the use of cell-specific AR knock out models. For all three aspects, we translate basic and preclinical findings to human situations and back." "Optimizing an experimental approach to silence disease-related genes by interfering with their chromatin structure. Epigenetic silencing of the human androgen receptor." "Annemie Haelens" "Laboratory of Molecular Endocrinology" "In this CREA-project, we aim to develop methods to induce alterations in the chromatin structure at a specific locus in the genome, to shut down one singe gene. Three approaches will be testen: (1) expression of shRNAs, (2) expression of non-coding RNAs and (3) transfection of methylated oligonucleotides. Epigenetic silencing by these mechanisms has been described for other test organisms, but has not been testen thoroughly in human/mammalian cells. Such ""epigenetic therapy"" has the advantage that it acts before the gene is actually expressed, preventing the protein from appearing. The use of mRNA targeting siRNAs to experimentally control protein levels has only short term effects, since they target the mRNA which is continuously synthesized as long as the gene is active. In contrast, a transient exposure to an epigenetic modifying agent is expected to suffice for the stable shut down of the gene. The changed epigenome will even be inherited by the daughter cells. During the two years of this CREA project, we will test the possibility to shut down the expression of the androgen receptor gene in prostate cancer cells. Prostate cancer is the most common cancer excluding skin cancer, and the second leading cause of cancer-related dead. It is likely to become a more promiment and pressing problem as the percentage of elderly men increases. Current treatments have severe side effects and a high chance for recurrence. Therefore, the need for development of new highly effective therapies with a small impact on the quality-of-life and minimal side effects is obvious. The fact that the AR is crucial for both the initiation and maintenance of prostate cancer makes it the ideal target for therapy. It has been shown that the knock down of androgen receptor (AR) expression at the mRNA level, leads to cell death of prostate cancer cells. Our approach is innovative because silencing the AR promoter depletes the cell of AR protein. This may enhance the benefits of androgen ablation by reducing of preventing AR cross activation by other ligands and other signaling pathways. Our new epigenetic silencing protocol will be a major step forward in the treatment of prostate cancer. When our approach is successful, we plan to target other genes with a proven role in prostate and/or other cancers, such as oncogenes or tumor survival genes. Of course the same protocols will be useful in fundamental research in which shutting down specific genes (cfr. knock out) has provided many insights in protein functions." "Molecular biology of the androgen receptor and its role in the development and homeostasis of skeletal muscle." "Frank Claessens" "Laboratory of Molecular Endocrinology, Clinical and Experimental Endocrinology" "The gender differences in skeletal muscles are in a large part due to differences in sex steroids. Male hormones act by activating the androgen receptor, which is a transcription factor. Direct and indirect actions of the androgens will be studied at the molecular level to determine which signal transduction pathways are affected. This will be investigated by means of transgenic mouse models and in vitro cultures of skeletal muscle cells." "Synthesis of novel compounds with potential antiandrogen activity" "Wim Dehaen" "Sustainable Chemistry for Metals and Molecules, Laboratory of Molecular Endocrinology" "Prostate cancer (PCa) is a major public health problem worldwide. It is estimated that it is the second common malignant neoplasm among men representing 13.5% of all the cancers diagnosed in men worldwide. Several therapies of metastatic PCa are based on the inhibition of the androgen receptor (AR) which is the key regulator of the prostate cancer cell growth.The AR is activated by androgens like testosterone and 5α–dihydrotestosterone (DHT) and it is the key transcription factor both in healthy prostate function and neoplastic transformation. Antiandrogens or AR antagonists are compounds that interfere in the biological effects of androgens by binding to the AR and inhibiting its functioning. The antagonists that are currently used in the clinic have relatively low affinities for the AR and also evoke resistance. Therefore, it is of paramount importance to develop more potent AR antagonists and also antagonists useful in resistant PCa.In the research group of Prof. Claessens, as a result of screening a collection of commercially available compounds, the MEL-6 compound was identified as having potential antiandrogen activity and was earmarked for further investigation. Unfortunately, the commercial suppliers did not include information on the stereochemistry of MEL-6. Indeed, preliminary research in our group showed that the activity of the (resynthesized) MEL-6.2 analogue was due to only one of the 4 possible (dia)stereoisomers, the other ones being inactive. The synthesis of this compound was only possible at small scale and the absolute configuration still remains to be determined. Moreover, an in vitro assay using mouse liver microsomes, determined the half-life to be only 4 minutes Thus, the development of metabolically more stable derivatives is the major priority, before embarking in more laborious and expensive preclinical and clinical trials.For that reason, we planned and prepared three libraries of novel compounds which have been tested for their antiandrogen activity.A first class of compounds similar to MEL-6 was obtained via two distinct synthetic pathways. In a first approach, the nitroalkene derivative underwent Michael addition from the N-substituted piperazine followed by reduction of the nitro group and acylation to yield the MEL-6 analogue. In a second approach, we proposed the oxidative amination of substituted phenylacetone with N-substituted piperazine and then conversion of the carbonyl group to the oxime moiety followed by reduction and acylation. The two different strategies led to the formation of the active and inactive against prostate cancer MEL-6 analogues which were subjected to in vitro and metabolic studies.As an alternative class of compounds, we wanted to introduce the 1,2,3-triazole ring which is considered a bioisoster of the amide group. We used our newly developed multicomponent strategy to access fused bicyclic 1,2,3-triazole derivatives from enolizable ketone, primary amines and 4-nitrophenyl azide. The compounds have been tested for their antiandrogen activity. However, in vitro evaluation on ClARE cells showed that none but one compound exhibits moderate AR inhibitory activity. Nevertheless, the triazolo fused derivatives were submitted for general antiviral and anticancer screening. Indeed, some compounds show promising antiviral properties against Coronavirus 229E.As a variant, we wanted to prepare N-connected triazoles from a homochiral propane-1,3-diamine building block based on the organocatalytic reactions developed in our group. We succeeded in obtaining a small library of bis-1,2,3-tiazoles which were submitted for biological evaluation." "Use of dried blood spots and microsampling technologies for anti-doping analysis" "Peter Van Eenoo" "Department of Diagnostic Sciences" "Historically, urine has been used for the detection of prohibited substances in sports. Since 2022, the use of dried blood spots and blood microsampling technologies has been approved. This sampling technique offers advantages (fast, non-intrussive sample collection; cheap and stable matrix, farmacokinetics). However, to allow the implementation new methods will need to be developed to allow for an adequate switch in sample matrix.This project will focus on the development and validation of a wide-range of methods for doping substances at concentrations that will allow adequate detection. High performance chromatography-mass spectrometric methods will be developed and evaluated. Sveeral sample preparation (proteine precipitation, solid phase and liquid-liquid extraction) will be used and optimized. The influence of the hematocrit effect will be investigated and correction factors will be determined.Once developed, the methods will be used in a diverse range of research projects as well as transferred to routine applications within and outside the doping context (e.g. endocrinology for the endogenous steroids, stimulants/narcotics to toxicology, etc). Hence, research networks will be set-up.In a first phase, the new LC-MS will be used to develop methods in DBS and microsamples for a full steroid profile in blood. The developed methods will focus also on phase 2 metabolites and therefore offer a full range of target substances for doping and clinical applications. The methods would aim at the detection of the following steroids in adequate concentration ranges: Free steroids: testosterone, epitestosterone, androstenedione, dehydroepiandrosterone, dihydrotestosterone, androsterone, etiocholanolone, progesterone, 17a-OH-progesterone, cortisol, corticosterone, deoxycorticosterone, 11-deoxycorticosterone, 21-deoxycortisol, estradiol, estrone and intact conjugates: testosterone glucuronide, epitestosterone glucuronide, 5bab-androstanediol-3-glucuronide, 5bab-androstanediol-17-glucuronide, epiandrosterone glucuronide, testosterone sulfate, epitestosterone sulfate, androsterone sulfate, etiocholanolone sulfate, dehydroepiandrosterone sulfate Indeed, one of the great advantages of DBS/microsampling offers the opportunity to apply the developed methods in other research fields, by focussing not only on androgens, but also estrogens, and other endogenous steroids in a clinically/pharmacokinetically relevant matrix. The additional steroids will make the methods suitable for applications for clinical applications. While certainly this will require additional effort, the scientific return can be tremendous. In the clinical field, the determination of steroid levels is critical for diagnosis, treatment and follow-up of androgen and estrogen levels in children and adults and follow-up of sexual development disorders and fertility. Improvements in the analytical field  facilitating easy sample collection and an improved pre-analytical phase are therefore important.To achieve a full impact the cooperation with the department of endocrinology will be intensified and the method transferred. The joint-expertise of DoCoLab and the Department of endocrinology will allow to set-up the world-leading collaborative unit." "No pain, just gain: new dried blood microsample-based avenues for steroid testing" "Christophe Stove" "Department of Diagnostic Sciences, Department of Bio-analysis" "For decades, anabolic androgenic steroids have been the most commonly abused substances in sports. This project aims at optimizing fully validated LC-MS/MS methods for steroid determination in dried blood microsamples, including dried blood spots (DBS) and samples collected by volumetric absorptive microsampling (VAMS). A fully automated DBS method will be developed, allowing analysis of a steroid panel."