Synthetic corticosteroids as tryptophan hydroxylase stabilizers.

Background: Clinically, corticosteroids are used mainly for their immune-modulatory properties but are also known to influence mood. Despite evidence of a role in regulating tryptophan hydroxylases (TPH), key enzymes in serotonin biosynthesis, a direct action of corticosteroids on these enzymes has not been systematically investigated. Methodology & results: Corticosteroid effects on TPHs were tested using an in vitro assay. The compound with the strongest modulatory effect, beclomethasone dipropionate, activated TPH1 and TPH2 with low micromolar potency. Thermostability assays suggested a stabilizing mechanism, and computational docking indicated that beclomethasone dipropionate interacts with the TPH active site. Conclusion: Beclomethasone dipropionate is a stabilizer of TPHs, acting as a pharmacological chaperone. Our findings may inspire further development of steroid scaffolds as putative antidepressant drugs.

Potent analogues of etiprednol dicloacetate, a second generation of soft corticosteroids.

OBJECTIVES: Loteprednol etabonate (LE) is the first, highly successful soft corticosteroid (SC) designed using the 'inactive metabolite' approach, starting with ∆1 -cortienic acid (d-CA). The next generation of SCs based on d-CA was etiprednol dicloacetate (ED). The 17α-dichloroacetyl function serves both as a unique pharmacophore and as the source of the molecule's softness. Highly potent SCs were designed based on a combination of ED and LE, introducing 6, 9 and 16 substituents in the molecule. METHODS: The new 6α, 9α, 16α and ß 17α-dichloroacetyl 17ß-esters were synthesized from the correspondingly substituted ∆1 -cortienic acids. The anti-inflammatory activity was assessed using LPS-induced TNF α-release under various conditions to determine intrinsic activity vs. systemic biological stability. In vivo anti-inflammatory activity was studied in the widely used ovalbumin-sensitized and ovalbumin-challenged Brown Norway rat model. KEY FINDINGS: The 6α or 9α-fluoro substitution produced highly potent corticosteroids, but the 17α-dichloroacetyl substituent provided 'softness' in all cases. Local application of these steroids will significantly reduce systemic activity, due to the facile hydrolytic deactivation of these molecules. CONCLUSIONS: A 17α-dichloroacetyl derivative of fluticasone (FLU) is highly potent but much safer than the currently used propionate or furoate ester.

Chemical Pathways of Corticosteroids, Industrial Synthesis from Sapogenins.

Corticosteroids are products of high industrial and commercial importance. There are dozens of different synthesis published for all of them. Some are coming from academia and some from industry. Here, industrial processes for the synthesis of prednisone, prednisolone, hydrocortisone, dexamethasone, betamethasone, and methylprednisolone are described. The starting material is diosgenin and the desired molecules are reached due to a good combination of chemistry and biotechnology that was developed along the second part of the twentieth century.

Development of a concise synthesis of ouabagenin and hydroxylated corticosteroid analogues.

The natural product ouabagenin is a complex cardiotonic steroid with a highly oxygenated skeleton. This full account describes the development of a concise synthesis of ouabagenin, including the evolution of synthetic strategy to access hydroxylation at the C19 position of a steroid skeleton. In addition, approaches to install the requisite butenolide moiety at the C17 position are discussed. Lastly, methodology developed in this synthesis has been applied in the generation of novel analogues of corticosteroid drugs bearing a hydroxyl group at the C19 position.

Synthesis of multiply deuterated 3- and 21-monosulfates of allo-tetrahydrocorticosteroids as internal standards for mass spectrometry.

The accurate analysis of trace components in complex biological matrices requires the use of reliable internal standards. For liquid chromatography/mass spectrometry analysis, the stable isotope-labeled analogues of the analyte molecules are the most appropriate internal standards. In this paper the synthesis of the 3- and 21-monosulfates of allo-tetrahydrocorticosteroids labeled with four or five deuterium atoms is described. The principal reactions used were (1) hydrogen-deuterium exchange reaction of active methylene groups adjacent to 3- and 11-oxo group of 17,20;20,21-bismethylenedioxy derivatives of 5α-3-ketosteroids and/or 5α-11-ketosteroids with NaOD in CH(3)OD followed by reduction with NaBD(4), (2) epimerization of the 3ß-hydroxy group into a 3α configuration, (3) sulfation of hydroxy groups at C-3 or C-21 in the resulting substrates with sulfur trioxide-trimethylamine complex, and (4) removal of 17,20;20,21-bismethylenedioxy groups with hydrogen fluoride in ethanol. Isotopic purity was found to be satisfactory by MS, and NMR properties of the new compounds were tabulated. The labeled compounds can be used as internal standards in liquid chromatography/mass spectrometry assays for clinical and biochemical studies.

New applications for bismuth(III) salts in organic synthesis: from bulk chemicals to steroid and terpene chemistry.

Bismuth(III) salts are currently considered efficient and "ecofriendly" reagents and catalysts for the development of new applications in organic synthesis. The preparation of bismuth(III) triflate and its analogues is reviewed as well as some of their applications to the synthesis of bulk chemicals via electrophilic addition and cyclization reactions. The use of bismuth(III) salts in the development of new chemical processes involving steroids and terpenes as substrates is also discussed.

Design and synthesis of long acting inhaled corticosteroids for the treatment of asthma.

In this Letter we present data for a novel series of ICS for the treatment of asthma. 'Inhalation by design' principles have been applied to a series of highly potent steroidal GR agonists, with a focus on optimising the potential therapeutic index in human. Pharmacokinetic properties were tuned with high intrinsic clearance and low oral bioavailability in mind, to minimise systemic exposure and reduce systemically driven adverse events. High CYP mediated clearance as well as glucuronidation were targeted to achieve high intrinsic clearance coupled with multiple routes of clearance to minimise drug-drug interactions. Furthermore, pharmaceutical properties such as stability, crystallinity and solubility were considered to ensure compatibility with a dry powder inhaler. This work culminated in the identification of the clinical candidate 15, which demonstrates preclinically the desired efficacy and safety profiles confirming its potential as an inhaled agent for the treatment of asthma.

Synthesis of 3- and 21-monosulfates of [2,2,3ß,4,4-²H5]-tetrahydrocorticosteroids in the 5ß-series as internal standards for mass spectrometry.

The 3- and 21-monosulfates of pentadeuterated 5ß-tetrahydrocorticosteroides were synthesized, starting from cortisol and 11-deoxycotisol. The principal reactions used were (1) perdeuteration of the methylene groups adjacent to the 3-oxo group of 17,20:20,21-bismethylendioxy-5ß-3-ketosteroids with NaOD in CH(3)OD followed by stereoselective reduction with NaBD(4), (2) sulfation of hydroxy groups with sulfur trioxide-trimethylamine complex, and (3) removal of the 17,20:20,21-bismethylendioxy group with hydrogen fluoride. The labeled compounds can be used as internal standards in liquid chromatography/mass spectrometry assays for clinical and biochemical studies.

Mecanismos celulares e moleculares de ação dos glicocorticóides endógenos sobre a mobilização de neutrófilos / Cellular and molecular mechanisms of action of endogenous glucocorticoids on the neutrophil mobilization

Temos mostrado que os glicocorticóides endógenos (GE) modulam o rolling e a aderência de neutrófilos in vivo, mediando a expressão de moléculas de adesão no leucócito e no endotélio. Adicionalmente, os GE controlam a maturação neutrofílica na medula e a sua mobilização para o sangue periférico. O presente trabalho visou investigar os mecanismos moleculares e celulares envolvidos na modulação exercida pelos GE neste processo. Utilizando ratos Wistar submetidos à adrenalectomia bilateral, tratados com RU 38486 ou controles (falso-operados, tratados com veículo ou não manipulados), foi demonstrado que: 1) os GE controlam, negativamente, a expressão de L-selectina em neutrófilos circulantes e ICAM-1, VCAM-1, PECAM-1, VAP-1 na célula endotelial e, positivamente, a expressão de L-selectina em granulócitos da medula óssea via seu receptor citosólico (GCR); 2) o mecanismo envolvido no controle dos GE sobre a expressão de L-selectina é independente de ação sobre sua expressão gênica ou da atividade de NFkB, mas dependente da expressão de anexina-A1, como verificado em camundongos knockouts (KO) para esta proteína 3) o controle da expressão de moléculas de adesão endotelial é dependente de ações sobre a expressão gênica, via translocação nuclear do NFkB; 4) a neutrofilia detectada em animais adrenalectomizados (ADR) é mediada pelo GCR, e dependente de anexina- A1; 5) a neutrofilia parece ser dependente da ação da anexina-A1 sobre a secreção de SDF-1 na medula óssea e expressão de CXCR-4 em neutrófilos circulantes e da medula; 6) concentrações circulantes elevadas de GE induzidas pela administração de ACTH confirmaram o controle dos GE, via anexina A-1, sobre o tráfego de neutrófilos da medula óssea para o sangue, mas sugerem um controle diferencial dos GE e anexina A-1 sobre a expressão de L-selectina em células da medula e do sangue circulante. Estes dados mostram mecanismos inéditos do controle dos GE sobre o tráfego de neutrófilos, que diferem em cada microambiente...

We have shown that endogenous glucocorticoids (GE) modulate the rolling and adhesion of neutrophils in vivo, mediating the expression of adhesion molecules on leukocytes and the endothelium. Additionally, the GE control neutrophil maturation in bone marrow and mobilization to peripheral blood. This work aimed to investigate the molecular and cellular mechanisms involved in the modulation exerted by GE in this process. Using male Wistar rats, submitted to bilateral adrenalectomy, treatment with RU 38 486 or controls (sham operated, vehicle or non manipulated), it was shown that: 1) GE control, negatively, L-selectin expression on circulating neutrophils and ICAM-1, VCAM-1, PECAM-1, VAP-1 on endothelial cell and, positively, L-selectin expression on bone marrow granulocytes via their cytosolic receptor (GCR); 2) the mechanism involved in the control of GE on the L-selectin expression is independent of its action on gene expression or NFkB activity, but dependent on the expression of anexina-A1, as observed in mice knockouts for this protein; 3) the control of endothelial adhesion molecules is dependent on gene expression, via NFkB translocation; 4) the neutrophilia detected in adrenalectomized animals (ADR) is mediated by GCR, and dependent on anexina-A1; 5) the neutrophilia seems to be dependent on the action of annexin A-1 on SDF-1á secretion in bone marrow and expression of CXCR-4 in peripheral blood and bone marrow; 6) high circulating concentrations of GE induced by administration of ACTH confirmed the control of GE, via the annexin-1, on the traffic of neutrophils from the bone marrow to the blood, but suggest a differential control of GE and annexin A-1 on the L-selectin expression in the bone marrow and circulating blood. These data indicate unpublished mechanisms of control of GE on the traffic of neutrophils, which differ in each microenvironment and cell type involved in this complex phenomenon.

Corticosteroids 21-glucuronides: synthesis and complete characterization by (1)H and (13)C NMR.

Despite the important physiological role of the corticosteroids glucuronides, very poor NMR data for this class of compounds are reported. For this reason we prepared a set of corticosteroids and submitted them to a detailed NMR study. A complete assignment of (1)H and (13)C signals was accomplished arranging mono- and two-dimensional NMR techniques.

Corticosteroid design for the treatment of asthma: structural insights and the therapeutic potential of soft corticosteroids.

Inhaled and intranasal corticosteroids (ICSs) still are the most effective treatment available for allergic airway diseases and are likely to remain the cornerstone of managing persistent asthma/allergic rhinitis in the foreseeable future. Even if the therapeutic index of this class increased significantly with the introduction of newer corticosteroids, and even if new therapeutic potentials are beginning to emerge with our increasing understanding of the mechanisms of asthma, chronic obstructive pulmonary disease, and rhinitis, corticosteroid development still remains a very important field for drug designers. After a brief review of issues related to the structure-activity relationships of glucocorticoids and the main determinants of their receptor-binding affinity at the glucocorticoid receptor, the main focus of the present article will be on the development of soft corticosteroids, as they are particularly well suited to separate local activity from systemic side effects, which still is an important issue for ICSs. Design consideration required in the search for safe and effective soft drugs on one hand, and safe and effective ICSs on the other hand, will be briefly discussed and illustrated with a number of cases, in particular, with that of loteprednol etabonate and etiprednol dicloacetate, soft corticosteroids that are being developed for a full spectrum of therapeutic applications including asthma and allergic rhinitis.

Microwave-enhanced nucleophilic fluorination in the synthesis of fluoropyridyl derivatives of [3,2-c]pyrazolo-corticosteroids, potential glucocorticoid receptor-mediated imaging agents.

Fluoropyridyl derivatives of [3,2-c]pyrazolo-corticosteroids have high affinity for the glucocorticoid receptor (GR) and are highly active glucocorticoids. They are thus considered to be excellent candidates for PET imaging of GR containing tissues when labeled with fluorine-18 (t(1/2)=110 min). Previously reported syntheses of these fluorinated glucocorticoids were accomplished by conventional thermal nucleophilic halogen exchange reactions with chloropyridyl precursors. These reactions were found to proceed at rates too slow for feasible application to radiosynthesis using [(18)F]fluoride. We have applied microwave-heating methods to these reactions and found that significant rate enhancements can be realized. Kinetic experiments showed an average relative rate ratio of 3/1 for microwave versus conventional heating and preparative experiments showed an average relative conversion ratio of 4.5/1 during the initial 120 min, a period approximating one half-life of the isotope. The microwave method described was used to prepare previously unreported 2'-(2-fluoro-4-pyridyl)-11beta,17,21-trihydroxy-16alpha-methyl-20-oxo-pregn-4-eno-[3,2-c]-pyrazole, which was evaluated for biological activity.

Programação pré-natal de hipertensão arterial na vida adulta / Prenatal programming of adult arterial hypertension

Na busca da melhor compreensão da origem das doenças cardiovasculares, uma importante linha de investigação surgiu a partir da demonstração da associação entre o baixo peso ao nascer e o desenvolvimento de hipertensão foi a base para a formulação da hipótese de que doenças cardiovasculares manifestadas na idade adulta podem ser programadas a partir de insultos ocorridos no período pré natal.Diversos modelos experimentais apóiam esta teoria. Em animais de laboratório, estudos com desnutrição induzida durante a gestação resultaram em animais adultos com níveis mais elevados de pressão arterial do que os controles...

Preparation of polymeric nanoparticles containing corticosteroid by a novel aerosol flow reactor method.

Polymeric drug-containing nanoparticles were prepared using a novel aerosol flow reactor method. The polymeric drug-containing nanoparticles prepared consist of a poorly water soluble corticosteroid, beclomethasone dipropionate, and polymeric materials Eudragit E 100 or Eudragit L 100. The novel method used in this study allows synthesis of nanoparticles directly as dry powders. The nanoparticles can contain various ratios of drug and polymer, and the use of any additional stabilisation materials is avoided. In this study, nanoparticles with different drug-to-polymer ratios were prepared. Particle size and morphology, crystallinity, and thermal behaviour were determined as a function of particle composition. It was found that all the nanoparticles produced, regardless of particle composition, had geometric number mean diameters of approximately 90 nm, and were spherical showing smooth surfaces. The drug was molecularly dispersed in the amorphous polymeric matrix of the nanoparticles, and drug crystallisation was not observed when the ambient temperature was below the glass transition temperature of the polymer.

Synthesis of novel arylpyrazolo corticosteroids as potential ligands for imaging brain glucocorticoid receptors.

Corticosteroids regulate a variety of essential physiological functions, such as mineral balance and stress. The great interest in these steroids, especially the glucocorticoids, stems from roles they are thought to play in neuropsychiatric disorders, such as severe depression and anxiety.The development of glucocorticoid receptor (GR) ligands which are appropriately labeled with short-lived positron-emitting radioisotopes would allow the non-invasive in-vivo imaging and mapping of brain GRs by means of positron emission tomography (PET). In this context we have synthesized a series of novel arylpyrazolo steroids exhibiting different substitution patterns at the D-ring of the steroid skeleton, as ligands for brain GRs. Special attention was given to 4-fluorophenyl pyrazolo steroids, which are known to display high binding affinity toward the GR. The compounds were evaluated in a competitive radiometric receptor binding assay to determine their relative binding affinities (RBA) to the GR. Some compounds show good binding affinities of up to 56% in comparison to dexamethasone (100%). In initial experiments, selected candidates were labeled with the positron emitter fluorine-18 and in one case with the gamma-emitter iodine-131.

Two new C-nor/D-homo corticosteroids formed by dehydration rearrangement of a 12beta-hydroxy corticoid.

Two new C-nor/D-homo corticosteroids were isolated from the filtrate of the triene carbonate intermediate during the production of betamethasone. Their structures were elucidated based on NMR spectroscopic studies and mass spectral analyses. The mechanism of formation of each of these two steroids was postulated as a Wagner-Meerwein rearrangement of the corresponding 12beta-hydroxy steroid. The new 12beta-hydroxy corticosteroid was also isolated from the fermentation process of betamethasone synthesis. Its structure was identified via NMR and mass spectroscopic studies. Treatment of the new 12beta-hydroxy corticoid with PCl(5) at low temperature provided two C-nor/D-homo compounds in a ratio of 1:1. Conversion of 12beta-hydroxy corticoid into a corresponding 12beta-mesylate followed by heating in HOAc to 110 degrees C in the presence of NaOAc produced a mixture of the rearranged products in a 1:4 ratio.

Synthesis and structure-activity relationships in a series of antiinflammatory corticosteroid analogues, halomethyl androstane-17 beta-carbothioates and -17 beta-carboselenoates.

The preparation and topical antiinflammatory potencies of a series of halomethyl 17 alpha-(acyloxy)- 11 beta-hydroxy-3-oxoandrosta-1,4-diene-17 beta-carbothioates, carrying combinations of 6 alpha-fluoro, 9 alpha-fluoro, 16-methyl, and 16-methylene substituents, are described. Key synthetic stages were the preparation of carbothioic acids and their reaction with dihalomethanes. The carbothioic acids were formed from 17 beta-carboxylic acids by initial reaction with dimethylthiocarbamoyl chloride followed by aminolysis of the resulting rearranged mixed anhydride with diethylamine, or by carboxyl activation with 1,1'-carbonyldiimidazole (CDI) or 2-fluoro-N-methylpyridinium tosylate (FMPT) and reaction with hydrogen sulfide, the choice of reagent being governed by the 17 alpha-substituent. Carboxyl activation with FMPT and reaction with sodium hydrogen selenide led to the halomethyl 16-methyleneandrostane-17 beta-carboselenoate analogues. Anti-inflammatory potencies were measured in humans using the vasoconstriction assay and in rats and mice by a modification the Tonelli croton oil ear assay. Best activities were shown by fluoromethyl and chloromethyl carbothioates with a 17 alpha-propionyloxy group. S-Fluoromethyl 6 alpha, 9 alpha-difluoro-11 beta-hydroxy-16 alpha-methyl-3-oxo-17 alpha- (propionyloxy)androsta-1,4-diene-17 beta-carbothioate (fluticasone propionate, FP) was selected for clinical study as it showed high topical antiinflammatory activity but caused little hypothalamic-pituitary-adrenal suppression after topical or oral administration to rodents.

General high-performance liquid chromatographic procedures for the rapid screening of natural and synthetic corticosteroids.

A study of a multiple high-performance liquid chromatographic procedure for the separation of 30 different corticosteroids is described. Normal- and reversed-phase general methods operating in a wide polarity range have been developed for the rapid screening of multicomponent mixtures. The complementary application of both normal- and reversed-phase methods could permit clarification of uncertainty deriving from analyses performed by only one method.

A history of significant steroid discoveries and developments originating at the Schering Corporation (USA) since 1948.

A firsthand historical account of some of the significant contributions of the steroid research group at the Schering Corporation (Bloomfield, NJ, USA) to the discovery and/or development of important therapeutic agents is presented. These include the discovery of the antiinflammatory corticosteroid drugs prednisone, prednisolone, and betamethasone, all of which, more than 30 years after their introduction, continue to enjoy wide use in human and animal medical practice throughout the world.

Steroid hormones and the Luftwaffe. A venture into fundamental strategic research and some of its consequences: the Birch reduction becomes a birth reduction.

I recount my involvement in the development of biologically active, totally synthetic steroid hormones since 1941. The reasons for my approach to the first total synthesis of a potent androgen are given. I touch on the even more important general synthetic outcomes from ideas generated by the search for useful amounts of a cortically active hormone, such as novel angular methylations, partial hydrogenations of aromatic systems (Birch reductions), and novel usages in synthesis based on the unique structures so generated. The Birch process was critical for synthesis of the first oral contraceptives. A need to explain the structures of Birch reduction products and the experimental requirements resulted in further elucidations of the mechanism of reduction processes, notably for aromatic compounds. The first theoretical and practical distinction between structures of products as determined by a reaction rate or an equilibrium position was first exploited experimentally in the deconjugation of cholest-4-en-3-one in the final step of the first facile total synthesis of cholesterol. A knowledge gained of the biosynthesis of steroids and its specific enzymes helped to initiate my general polyketide theory of biosynthesis and also my idea of outdoing enzyme achievements with organometallic complexes ("inorganic enzyme chemistry"). I assert the high historic importance of steroids in promoting the advance of general organic chemistry within many fields.

PIP: Arthur J. Birch, chemistry professor at Australian National University, reviewed his role in developing the biologically active, totally synthetic steroid hormones beginning in 1941 after graduating from Oxford University in the UK. His supervisor asked him to serve at the Dyson Perrins Laboratory at Oxford, affiliated with ICI which was affiliated with the UK Government, to produce cortically active hormones for RAF pilots, since it was rumored that German Luftwaffe pilots used them. His research and ideas led him to develop new angular methylations, partial hydrogenations of aromatic systems (Birch reductions), and new synthesis practices based on the unique structures so generated. The Birch reductions were a key element to synthesis of the first oral contraceptives. Specifically, Gregory Pincus orally administered the synthetic 19-nor ethinyl derivatives obtained from Birch reduction and found them to be active progestagens, leading to the development of oral contraceptives. His research into revealing the structures of chemicals produced by Birch reductions and the experimental requirements brought about more clarity of the mechanism of reduction processes, especially for aromatic compounds. His first attempt to differentiate between structures of products as determined by a reaction rate or an equilibrium position was deconjugation of cholest-4-en-3-one in the last step of the first elementary total synthesis of cholesterol. The knowledge acquired from biosynthesis of steroids and of specific enzymes contributed to his general polyketide theory of biosynthesis and to his theory of outdoing enzyme achievements with organometallic complexes, which he called inorganic enzyme chemistry. Professor Birch's research has maintained the very historic significance of steroids in furthering the progress of general organic chemistry within many fields.