Search

Several drugs were found after their market approval to unexpectedly inhibit adrenal 11β-hydroxylase (CYP11B1)-dependent cortisol synthesis. Known side-effects of CYP11B1 inhibition include hypertension and hypokalemia, due to a feedback activation of adrenal steroidogenesis, leading to supraphysiological concentrations of 11-deoxycortisol and 11-deoxycorticosterone that can activate the mineralocorticoid receptor. This results in potassium excretion and sodium and water retention, ultimately causing hypertension. With the risk known but usually not addressed in preclinical evaluation, this study aimed to identify drugs and drug candidates inhibiting CYP11B1. Two conceptually different virtual screening methods were combined, a pharmacophore based and an induced fit docking approach. Cell-free and cell-based CYP11B1 activity measurements revealed several inhibitors with IC 50 values in the nanomolar range. Inhibitors include retinoic acid metabolism blocking agents (RAMBAs), azole antifungals, α 2 -adrenoceptor ligands, and a farnesyltransferase inhibitor. The active compounds share a nitrogen atom embedded in an aromatic ring system. Structure activity analysis identified the free electron pair of the nitrogen atom as a prerequisite for the drug-enzyme interaction, with its pK a value as an indicator of inhibitory potency. Another important parameter is drug lipophilicity, exemplified by etomidate. Changing its ethyl ester moiety to a more hydrophilic carboxylic acid group dramatically decreased the inhibitory potential, most likely due to less efficient cellular uptake. The presented work successfully combined different in silico and in vitro methods to identify several previously unknown CYP11B1 inhibitors. This workflow facilitates the identification of compounds that inhibit CYP11B1 and therefore pose a risk for inducing hypertension and hypokalemia., Competing Interests: Declaration of Competing Interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Alex Odermatt reports financial support was provided by Swiss Centre for Applied Human Toxicology., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)

11β-Hydroxysteroid dehydrogenase type 1 (11β-HSD1) converts inactive 11-keto-glucocorticoids to their active 11β-hydroxylated forms. It also catalyzes the oxoreduction of other endogenous and exogenous substrates. The ubiquitously expressed 11β-HSD1 shows high levels in liver and other metabolically active tissues such as brain and adipose tissue. Pharmacological inhibition of 11β-HSD1 was found to ameliorate adverse metabolic effects of elevated glucocorticoids in rodents and humans, improve wound healing and delay skin aging, and enhance memory and cognition in rodent Alzheimer's disease models. Thus, there is an interest to develop 11β-HSD1 inhibitors for therapeutic purposes. This chapter describes in vitro methods to assess 11β-HSD1 enzyme activity for different purposes, be it in disease models, for the assessment of the kinetics of novel substrates or for the screening and characterization of inhibitors. 11β-HSD1 protein expression and preparations of the different biological samples are discussed first, followed by a description of a well-established and easily adaptable 11β-HSD1 enzyme activity assay. Finally, different readout methods are shortly described. This chapter should provide the reader with a toolbox of methods to assess 11β-HSD1 activity with instructions in the form of a decision tree for the choice and implementation of an appropriate enzyme activity assay., (Copyright © 2023. Published by Elsevier Inc.)

11β-Hydroxysteroid dehydrogenase type 2 (11β-HSD2) converts active 11β-hydroxyglucocorticoids to their inactive 11-keto forms, fine-tuning the activation of mineralocorticoid and glucocorticoid receptors. 11β-HSD2 is expressed in mineralocorticoid target tissues such as renal distal tubules and cortical collecting ducts, and distal colon, but also in placenta where it acts as a barrier to reduce the amount of maternal glucocorticoids that reach the fetus. Disruption of 11β-HSD2 activity by genetic defects or inhibitors causes the syndrome of apparent mineralocorticoid excess (AME), characterized by hypernatremia, hypokalemia and hypertension. Secondary hypertension due to 11β-HSD2 inhibition has been observed upon consumption of excessive amounts of licorice and in patients treated with the azole fungicides posaconazole and itraconazole. Furthermore, inhibition of 11β-HSD2 during pregnancy with elevated exposure of the fetus to cortisol can cause neurological complications with a lower intelligence quotient, higher odds of attention deficit and hyperactivity disorder as well as metabolic reprogramming with an increased risk of cardio-metabolic disease in adulthood. This chapter describes in vitro methods for the determination of 11β-HSD2 activity that can be applied to identify inhibitors that may cause secondary hypertension and characterize the enzyme's activity in disease models. The included decision tree and the list of methods with their advantages and disadvantages aim to enable the reader to select and apply an in vitro method suitable for the scientific question and the equipment available in the respective laboratory., (Copyright © 2023. Published by Elsevier Inc.)