ABSTRACT
Glucocorticoids (GCs) are widely used to treat a variety of autoimmune and inflammatory diseases; however, systemic delivery of GCs is associated with side effects that affect essentially every organ system, reflecting the nearly ubiquitous expression of the glucocorticoid receptor (GR). Targeted delivery of GCs to diseased tissues using antibody-glucocorticoid conjugates (GC-ADCs) offers a therapeutic alternative to overcome these adverse effects. Herein, we describe novel classes of GCs that exhibited greater potency than dexamethasone and budesonide, a 100-fold selectivity toward the GR over other nuclear receptors, and no in vitro safety liability in pharmacology assays (hERG, AMES) and that demonstrated a substantial reduction in tumor necrosis factor-α (TNF-α) release in mice challenged with lipopolysaccharide (LPS). The site-specific conjugated GC-ADCs via cathepsin-cleavable linkers were highly stable in plasma and specifically released GCs in antigen-positive cells, suggesting that these novel GCs can serve as ADC payloads to treat autoimmune and inflammatory diseases.
Subject(s)
Budesonide/analogs & derivatives , Budesonide/therapeutic use , Glucocorticoids/therapeutic use , Immunoconjugates/therapeutic use , Inflammation/drug therapy , Animals , Budesonide/metabolism , Budesonide/pharmacokinetics , Cathepsin B/metabolism , Glucocorticoids/chemical synthesis , Glucocorticoids/metabolism , Glucocorticoids/pharmacokinetics , Humans , Immunoconjugates/chemistry , Immunoconjugates/immunology , Immunoconjugates/metabolism , Inflammation/chemically induced , Inflammation/immunology , Lipopolysaccharides , Male , Mice, Inbred C57BL , Molecular Docking Simulation , Molecular Structure , Receptors, Glucocorticoid/metabolism , Receptors, Prolactin/immunology , Structure-Activity RelationshipABSTRACT
Glucocorticoid receptor (GR) belongs to the superfamily of steroid hormone receptors. The dissociated or selective GR modulators (SEGRMs), preferring the transrepression rather than the transactivation, might exhibit anti-inflammatory activities with fewer side effects. This work presents a review of the molecular mechanism of GR involved in regulation of inflammation. As complementary or alternative therapeutic agents, the botanical compounds have been extensively used in the treatment of various diseases. Hence, this work reviews the botanical compounds as well as the synthetic compounds currently known to be potential SEGRMs. High-throughput virtual screening of SEGRMs from natural products has also been summarized.
Subject(s)
Anti-Inflammatory Agents/pharmacology , Glucocorticoids/pharmacology , Phytochemicals/pharmacology , Receptors, Glucocorticoid/agonists , Animals , Anti-Inflammatory Agents/chemical synthesis , Glucocorticoids/chemical synthesis , High-Throughput Screening Assays , Humans , Ligands , Molecular Structure , Phytochemicals/chemistry , Receptors, Glucocorticoid/metabolism , Signal Transduction , Structure-Activity RelationshipABSTRACT
Member States of the EU are required to monitor the use of pharmacologically active substances in food-producing animals. There is evidence, however, that the target-based approach currently applied in official monitoring plans might under-estimate the real incidence of growth promoter abuse in livestock. As demonstrated for sex hormones, the association of effect-oriented biological screening with chemical confirmatory techniques could be the best strategy in revealing the abuse of veterinary drugs. Here we demonstrate the reliability of a cell-based assay to screen calf urine samples for synthetic glucocorticoids. The validation included the most widely used synthetic drugs (flumethasone, dexamethasone, betamethasone, methylprednisolone and prednisolone) and was developed according to the Commission Decision 2002/657/EC, thus including the verification of cut-off level, the ß error, the specificity, ruggedness and stability. The study was carried out using prednisolone as representative substance at 5 ng mL-1 concentration. All blank and spiked urine fulfilled the EU criteria, moreover the method resulted in being specific and sound, and the analytes in urine were stable for at least 30 days. The assay results indicated its suitability for a qualitative analysis of calf urine samples. This method enabled the detection of low doses of synthetic glucocorticoids (GCs) in matrix (<2 ng mL-1 for flumethasone, dexamethasone, betamethasone; < 4 ng mL-1 for methylprednisolone; 5 ng mL-1 for prednisolone), with the possibility of detecting new or unknown molecules and cumulative effects of low-level mixtures with glucocorticoid bioactivity.
Subject(s)
Biological Assay , Glucocorticoids/urine , Animals , Cattle , Chromatography, Liquid , Glucocorticoids/chemical synthesis , Glucocorticoids/metabolism , Luciferases/genetics , Luciferases/metabolism , Spectrometry, Mass, Electrospray IonizationABSTRACT
Synthetic glucocorticoids (sGC) are administered to women at risk for pre-term delivery to reduce respiratory distress syndrome in the newborn. The prefrontal cortex (PFC) is important in regulating stress responses and related behaviours and expresses high levels of glucocorticoid receptors (GR). Further, antenatal exposure to sGC results in a hyperactive phenotype in first generation (F1) juvenile male and female offspring, as well as F2 and F3 juvenile females from the paternal lineage. We hypothesized that multiple courses of antenatal sGC modify gene expression in the PFC, that these effects are sex-specific and maintained across multiple generations, and that the gene sets affected relate to modified locomotor activity. We performed RNA sequencing on PFC of F1 juvenile males and females, as well as F2 and F3 juvenile females from the paternal lineage and used regression modelling to relate gene expression and behavior. Antenatal sGC resulted in sex-specific and generation-specific changes in gene expression. Further, the expression of 4 genes (C9orf116, Calb1, Glra3, and Gpr52) explained 20-29% of the observed variability in locomotor activity. Antenatal exposure to sGC profoundly influences the developing PFC; effects are evident across multiple generations and may drive altered behavioural phenotypes.
Subject(s)
Glucocorticoids/administration & dosage , Locomotion/drug effects , Prefrontal Cortex/drug effects , Premature Birth/drug therapy , Respiratory Distress Syndrome, Newborn/drug therapy , Adult , Calbindin 1/genetics , Female , Gene Expression Regulation, Developmental/drug effects , Glucocorticoids/adverse effects , Glucocorticoids/chemical synthesis , Humans , Infant, Newborn , Locomotion/genetics , Male , Prefrontal Cortex/metabolism , Premature Birth/genetics , Premature Birth/pathology , Premature Birth/prevention & control , Receptors, G-Protein-Coupled/genetics , Receptors, Glycine/genetics , Respiratory Distress Syndrome, Newborn/genetics , Respiratory Distress Syndrome, Newborn/pathology , Respiratory Distress Syndrome, Newborn/prevention & control , Sex Characteristics , Stress, Physiological/drug effects , Transcription, Genetic/drug effectsABSTRACT
A short and efficient synthesis, based on a one-step double elimination, of a key intermediate in the synthesis of various glucocorticosteroids has been developed. This method can be carried out on large scale for further industrial applications. The synthesis allowed us to identify a novel prednisolone derivative 10 and its anti-inflammatory activity was determined in an in vivo model of inflammation. In order to understand the regioselectivity of the double elimination under various conditions, mechanistic studies were undertaken and confirmed the experimental results. We also propose a mechanism for the formation of the new steroid 10 studied by molecular modeling.
Subject(s)
Glucocorticoids/chemistry , Glucocorticoids/chemical synthesis , Animals , Anti-Inflammatory Agents/chemical synthesis , Anti-Inflammatory Agents/chemistry , Anti-Inflammatory Agents/pharmacology , Anti-Inflammatory Agents/therapeutic use , Chemistry Techniques, Synthetic , Edema/drug therapy , Glucocorticoids/pharmacology , Glucocorticoids/therapeutic use , Male , Mice , Models, Molecular , Molecular Conformation , Prednisolone/chemical synthesis , Prednisolone/chemistry , Prednisolone/pharmacology , Prednisolone/therapeutic useABSTRACT
Chronic glucocorticoid (GC) treatment represents a widely-prescribed therapy for several diseases in consideration of both anti-inflammatory and immunosuppressive activity but, if used at high doses for prolonged periods, it can determine the systemic effects characteristic of Cushing's syndrome. In addition to signs and symptoms of hypercortisolism, patients on chronic GC therapy are at risk to develop tertiary adrenal insufficiency after the reduction or the withdrawal of corticosteroids or during acute stress. This effect is mediated by the negative feedback loop on the hypothalamus-pituitary-adrenal (HPA) axis, which mainly involves corticotropin-release hormone (CRH), which represents the most important driver of adrenocorticotropic hormone (ACTH) release. In fact, after withdrawal of chronic GC treatment, reactivation of CRH secretion is a necessary prerequisite for the recovery of the HPA axis. In addition to the well-known factors which regulate the degree of inhibition of the HPA during synthetic GC therapy (type of compound, method of administration, cumulative dose, duration of the treatment, concomitant drugs which can increase the bioavailability of GCs), there is a considerable variation in individual physiology, probably related to different genetic profiles which regulate GC receptor activity. This may represent an interesting basis for possible future research fields.
Subject(s)
Glucocorticoids/adverse effects , Hypothalamo-Hypophyseal System/drug effects , Pituitary-Adrenal System/drug effects , Adrenocorticotropic Hormone/metabolism , Animals , Corticotropin-Releasing Hormone/metabolism , Cushing Syndrome/chemically induced , Cushing Syndrome/metabolism , Feedback, Physiological , Glucocorticoids/chemical synthesis , Glucocorticoids/therapeutic use , HumansABSTRACT
The adipokine lipocalin 2 is linked to obesity and metabolic disorders. However, its role in human adipose tissue glucose and lipid metabolism is not explored. Here we show that the synthetic glucocorticoid dexamethasone dose-dependently increased lipocalin 2 gene expression in subcutaneous and omental adipose tissue from pre-menopausal females, while it had no effect in post-menopausal females or in males. Subcutaneous adipose tissue from both genders treated with recombinant human lipocalin 2 showed a reduction in protein levels of GLUT1 and GLUT4 and in glucose uptake in isolated adipocytes. In subcutaneous adipose tissue, lipocalin 2 increased IL-6 gene expression whereas expression of PPARγ and adiponectin was reduced. Our findings suggest that lipocalin 2 can contribute to insulin resistance in human adipose tissue. In pre-menopausal females, it may partly mediate adverse metabolic effects exerted by glucocorticoid excess.
Subject(s)
Adipose Tissue/metabolism , Glucocorticoids/physiology , Insulin Resistance , Lipocalin-2/physiology , Adiponectin/genetics , Adipose Tissue/drug effects , Adult , Aged , Dexamethasone/pharmacology , Female , Glucocorticoids/chemical synthesis , Glucose/metabolism , Humans , Lipocalin-2/metabolism , Male , Middle Aged , PPAR gamma/metabolism , Recombinant Proteins/pharmacology , Subcutaneous Fat/drug effects , Subcutaneous Fat/metabolism , Up-Regulation , Young AdultABSTRACT
A sensitive and comprehensive analytical method for glucocorticoids (GCs) in water samples was developed and applied to effluent of sewage treatment plants (STPs). In the present study, totally 10 natural and synthetic GCs, including cortisol, betamethasone valerate, clobetasol propionate, clobetasone butyrate, difluprednate, betamethasone, dexamethasone, betamethasone dipropionate, methylprednisolone, and prednisolone, were targeted. Analytes were extracted and concentrated using an OASIS HLB solid phase extraction cartridge. Chromatographic separation and quantification were achieved using an ultrahigh performance liquid chromatograph coupled with a tandem mass spectrometer (UHPLC-MS/MS). Method detection limits were 0.05 to 0.89 ng/L, which were 1-2 orders of magnitude more sensitive than in the previous reports. Cortisol was detected in more than half of (27 out of 50) analyzed effluent samples at concentrations in the range of ND-1.36 ng/L, indicating continuous discharge of natural GC via STP effluent. On the other hand, dexamethasone + betamethasone, prednisolone, betamethasone valerate, and clobetasol propionate were detected in 25, 8, 20, and 9 samples among 50 effluent samples, respectively, suggesting not extreme but significant administration of synthetic GCs.
Subject(s)
Chromatography, High Pressure Liquid/methods , Glucocorticoids/analysis , Sewage/chemistry , Tandem Mass Spectrometry/methods , Waste Management , Water Pollutants, Chemical/analysis , Glucocorticoids/chemical synthesis , Glucocorticoids/isolation & purification , Solid Phase Extraction , Water Pollutants, Chemical/chemical synthesis , Water Pollutants, Chemical/isolation & purificationABSTRACT
Osteoarthritis and rheumatoid arthritis are the two most common types of arthritis. Cartilage breakdown is a key feature of both diseases which contributes to the pain and joint deformity experienced by patients. Therefore, anti-arthritis drugs are of great importance. The aim of this review is to present recent progress in studies of various agents against osteoarthritis and rheumatoid arthritis. The structures and activities of anti-arthritic agents, which used in medical practice or are in development, are presented and discussed. The effects and mechanisms of action of opioids, glucocorticoids, non-steroidal anti-inflammatory drugs, disease-modifying anti-rheumatic drugs, natural products derived from plants, nutraceuticals, and a number of new and perspective agents are considered. Various perspective targets for the treatment of osteoarthritis and rheumatoid arthritis are also discussed. Trials of good quality are needed to draw solid conclusions regarding efficacy of many of the studied agents. Unfortunately, to date, there is no pharmacologic agent proven to prevent the progression of both diseases, and there is an urgent need for further development of better anti-arthritic agents.
Subject(s)
Anti-Inflammatory Agents, Non-Steroidal/therapeutic use , Antirheumatic Agents/therapeutic use , Arthritis, Rheumatoid/drug therapy , Biological Products/therapeutic use , Drugs, Investigational/therapeutic use , Osteoarthritis/drug therapy , Analgesics, Opioid/chemical synthesis , Analgesics, Opioid/therapeutic use , Anti-Inflammatory Agents, Non-Steroidal/chemical synthesis , Antirheumatic Agents/chemical synthesis , Arthritis, Rheumatoid/pathology , Arthritis, Rheumatoid/physiopathology , Biological Products/chemistry , Biological Products/isolation & purification , Cartilage/drug effects , Cartilage/pathology , Cartilage/physiopathology , Drugs, Investigational/chemical synthesis , Glucocorticoids/chemical synthesis , Glucocorticoids/therapeutic use , Humans , Joints/drug effects , Joints/pathology , Joints/physiopathology , Methotrexate/chemical synthesis , Methotrexate/therapeutic use , Osteoarthritis/pathology , Osteoarthritis/physiopathology , Pain/pathology , Pain/physiopathology , Pain/prevention & controlABSTRACT
Certain membrane-active cationic steroids are known to also possess both anti-inflammatory and antimicrobial properties. This combined functionality is particularly relevant for potential therapies of infections associated with elevated tissue damage, for example, cystic fibrosis airway disease, a condition characterized by chronic bacterial infections and ongoing inflammation. In this study, six novel cationic glucocorticoids were synthesized using beclomethasone, budesonide, and flumethasone. Products were either monosubstituted or disubstituted, containing one or two steroidal groups, respectively. In vitro evaluation of biological activities demonstrated dual anti-inflammatory and antimicrobial properties with limited cytotoxicity for all synthesized compounds. Budesonide-derived compounds showed the highest degree of both glucocorticoid and antimicrobial properties within their respective mono- and disubstituted categories. Structure-activity analyses revealed that activity was generally related to the potency of the parent glucocorticoid. Taken together, these data indicate that these types of dual acting cationic lipids can be synthesized with the appropriate starting steroid to tailor activities as desired.
Subject(s)
Anti-Infective Agents/chemical synthesis , Anti-Inflammatory Agents/chemistry , Anti-Inflammatory Agents/chemical synthesis , Lipids/chemistry , Anti-Infective Agents/chemistry , Anti-Infective Agents/pharmacology , Anti-Inflammatory Agents/pharmacology , Cations/chemistry , Cell Line, Tumor , Cell Survival/drug effects , Erythrocytes/cytology , Erythrocytes/drug effects , Erythrocytes/metabolism , Escherichia coli/drug effects , Glucocorticoids/chemical synthesis , Glucocorticoids/chemistry , Hemolysis/drug effects , Humans , Lipids/chemical synthesis , Lipids/pharmacology , Methicillin-Resistant Staphylococcus aureus/drug effects , Microbial Sensitivity Tests , Pseudomonas aeruginosa/drug effects , Structure-Activity RelationshipABSTRACT
Challenges to testing for the illicit use of anabolic substances in meat-producing animals stem from the production of new synthetic compounds and the administration of low-dose cocktails to circumvent detection by the surveillance schemes of European Union member states. This work evaluated for the first time GR-CALUX, a highly sensitive reporter gene assay, as a screening tool for the detection of synthetic glucocorticoids in bovine urine. In order to verify the effect of natural corticosteroids on the method, the bioassay was tested first using blank urine samples collected at the farm and the slaughterhouse. Next, the dose-response curves were measured for the most commonly used synthetic glucocorticoids. The bioassay's ability to detect them in spiked and incurred samples of bovine urine was then evaluated. Finally, its performance was compared against a commercially available ELISA kit ordinarily used in screening activities. GR-CALUX performance did not appear to be influenced by physiological levels of endogenous corticosteroids in the farm samples, whereas an increase in these hormones might invalidate the analysis in samples obtained at the slaughterhouse. Using pure compounds, GR-CALUX showed a high sensitivity toward the synthetic glucocorticosteroids tested in order of relative potencies: flumethasone â« dexamethasone > betamethasone > methylprednisolone > prednisolone. As expected, the bioassay failed to detect the prohormone prednisone. The results obtained from analysis of the spiked and incurred specimens reproduced those of the blank samples and the pure compounds. GR-CALUX is a promising screening tool for the detection of illicit treatments in meat-producing bovines. Its ability to detect the most commonly used synthetic glucocorticoids was comparable with the ELISA test. Importantly, it appeared to be less susceptible to matrix effects than ELISA.
Subject(s)
Glucocorticoids/urine , Animals , Cattle , Enzyme-Linked Immunosorbent Assay , Glucocorticoids/chemical synthesis , Glucocorticoids/chemistryABSTRACT
Molecular docking studies were performed on 18 17ß-carboxamide steroids in order to select compounds with potential local anti-inflammatory activity. These derivatives are amides of cortienic acids (obtained from hydrocortisone, prednisolone, and methylprednisolone) with methyl or ethyl esters of six amino acids. Interactions with the glucocorticoid receptor (GR), binding energies and ligand efficiency values of these compounds were compared with dexamethasone and cortienic acid obtained from prednisolone (inactive metabolite). On the basis of molecular docking studies, seven compounds were selected and their binding affinities for the GR were predicted by use of the exponential model created in this study. Subsequently, selected compounds were synthesized in good yields by use of modified N,N'-dicyclohexylcarbodiimide (DCC)/1-hydroxybenzotriazole (HOBt) coupling procedure. Finally, the local anti-inflammatory activity of the synthesized compounds was examined by use of the croton oil-induced ear edema test. In vivo evaluation of systemic side effects as well as in silico prediction of metabolism were performed on the derivative with the best local anti-inflammatory activity. The combination of molecular docking studies and the exponential model for the GR binding affinity prediction could be used as an in silico tool for the rational design of novel 17ß-carboxamide steroids with potentially better biological profile than dexamethasone.
Subject(s)
Anti-Inflammatory Agents/chemical synthesis , Anti-Inflammatory Agents/pharmacology , Drug Design , Edema/prevention & control , Glucocorticoids/chemical synthesis , Glucocorticoids/pharmacology , Inflammation/prevention & control , Animals , Anti-Inflammatory Agents/metabolism , Biotransformation , Croton Oil , Disease Models, Animal , Dose-Response Relationship, Drug , Edema/chemically induced , Glucocorticoids/metabolism , Hydrocortisone/analogs & derivatives , Hydrocortisone/chemical synthesis , Hydrocortisone/metabolism , Hydrocortisone/pharmacology , Inflammation/chemically induced , Ligands , Methylprednisolone/analogs & derivatives , Methylprednisolone/chemical synthesis , Methylprednisolone/metabolism , Methylprednisolone/pharmacology , Models, Biological , Molecular Docking Simulation , Molecular Structure , Prednisolone/analogs & derivatives , Prednisolone/chemical synthesis , Prednisolone/metabolism , Prednisolone/pharmacology , Rats , Receptors, Glucocorticoid/drug effects , Receptors, Glucocorticoid/metabolism , Structure-Activity RelationshipABSTRACT
Synthesis and structure-activity relationship (SAR) of a series of alkyl and cycloalkyl containing non-steroidal dissociated glucocorticoid receptor (GR) agonists is reported. This series of compounds was identified as part of an effort to replace the CF3 group in a scaffold represented by 1a. The study culminated in the identification of compound 14, a t-butyl containing derivative, which has shown potent activity for GR, selectivity against the progesterone receptor (PR) and the mineralocorticoid receptor (MR), in vitro anti-inflammatory activity in an IL-6 transrepression assay, and dissociation in a MMTV transactivation counter-screen. In a collagen-induced arthritis mouse model, 14 displayed prednisolone-like efficacy, and lower impact on body fat and free fatty acids than prednisolone at an equivalent anti-inflammatory dose.
Subject(s)
Drug Discovery , Glucocorticoids/chemical synthesis , Methanol/chemistry , Receptors, Glucocorticoid/agonists , Animals , Anti-Inflammatory Agents/chemical synthesis , Anti-Inflammatory Agents/chemistry , Anti-Inflammatory Agents/pharmacology , Arthritis/drug therapy , Binding Sites , Disease Models, Animal , Dose-Response Relationship, Drug , Glucocorticoids/chemistry , Glucocorticoids/pharmacology , Humans , Inhibitory Concentration 50 , Methanol/chemical synthesis , Methanol/pharmacology , Mice , Models, Molecular , Molecular Structure , Prednisolone/chemistry , Prednisolone/pharmacology , Protein Binding/drug effects , Rats , Rats, Sprague-DawleySubject(s)
Fluorine/chemistry , Pharmaceutical Preparations/chemistry , Anti-Infective Agents/chemical synthesis , Anti-Infective Agents/chemistry , Antineoplastic Agents/chemical synthesis , Antineoplastic Agents/chemistry , Antiviral Agents/chemical synthesis , Antiviral Agents/chemistry , Drug Industry/economics , Glucocorticoids/chemical synthesis , Glucocorticoids/chemistry , Hypoglycemic Agents/chemical synthesis , Hypoglycemic Agents/chemistry , Hypolipidemic Agents/chemical synthesis , Hypolipidemic Agents/chemistry , Legislation, Drug , Neurotransmitter Agents/chemical synthesis , Neurotransmitter Agents/chemistry , Pharmaceutical Preparations/chemical synthesisABSTRACT
The Hedgehog (Hh) pathway is a developmental signaling pathway that plays multiple roles during embryonic development and in adult tissues. Constitutive Hh signaling has been linked to the development and progression of several forms of cancer, and the application of small-molecule pathway inhibitors as anticancer chemotherapeutics is well studied and clearly defined. Activation of the Hh pathway as a therapeutic strategy for a variety of degenerative or ischemic disorders has also been proposed; however, the development of small-molecule Hh agonists has received less attention. The goal of this review is to highlight the recent evidence supporting the therapeutic potential of Hh pathway activators and to provide a comprehensive overview of small-molecule pathway agonists.
Subject(s)
Hedgehog Proteins/metabolism , Small Molecule Libraries/chemistry , Diabetes Mellitus/metabolism , Diabetes Mellitus/pathology , Glucocorticoids/chemical synthesis , Glucocorticoids/chemistry , Glucocorticoids/pharmacology , Hedgehog Proteins/agonists , Humans , Ischemia/metabolism , Ischemia/pathology , Neurodegenerative Diseases/metabolism , Neurodegenerative Diseases/pathology , Obesity/metabolism , Obesity/pathology , Signal Transduction/drug effects , Small Molecule Libraries/chemical synthesis , Small Molecule Libraries/pharmacology , Sterols/chemical synthesis , Sterols/chemistry , Sterols/pharmacologyABSTRACT
A six step transformation of prednisolone to 17α,21-dihydroxy-9ß,11ß-epoxy-16α-amethylpregna-1,4-diene-3,20-dione 21-acetate has been achieved in 13% unoptimised yield. Novel conditions for effecting a Mattox rearrangement and double dehydration of prednisolone were identified. Enhanced knowledge on the oxidation of silyl Δ(19,20)-enol ethers and structural factors that impact the success of the oxidation are also presented.
Subject(s)
Glucocorticoids/chemical synthesis , Prednisolone/analogs & derivatives , Prednisolone/chemistry , Oxidation-Reduction , Prednisolone/chemical synthesisABSTRACT
This study was aimed at synthesizing and evaluating a prednisolone-glucose derivative conjugate (PDG) that was expected to increase renal biodistribution without affecting pharmacological action and to decrease the systemic side effects of prednisolone. The PDG was designed and synthesized by tethering 6-amino-6-deoxy-D-glucose (a D-glucose derivative) to prednisolone and its chemical structure was confirmed by (1) H NMR, (13) C NMR, and LC-MS. This conjugate was then subjected to in vitro and in vivo evaluation like stability studies, biological distribution, pharmacodynamics, and systemic side effects studies. In these studies, PDG not only showed significant enhancement of renal target efficiency with high values of relative uptake efficiency (RE, 24.1), concentration efficiency (CE, 8.6), and kidney targeting index (KTI, 16.3), but retained the curative potency against minimal change nephrosis (MCN). In the systemic side effects study, no osteoporosis was observed in rats after the administration of PDG for 20 days, which exhibited limited side effects. Conclusively, our findings showed a pharmacologically active conjugate with the characteristics of renal targeting and limited systemic side effects. The results implied the potential of PDG as a promising therapeutic in the treatment of renal diseases.
Subject(s)
Glucocorticoids/chemical synthesis , Glucose/analogs & derivatives , Kidney/drug effects , Prednisolone/analogs & derivatives , Animals , Drug Delivery Systems , Drug Stability , Glucocorticoids/adverse effects , Glucocorticoids/pharmacokinetics , Glucocorticoids/pharmacology , Glucose/chemical synthesis , Glucose/pharmacokinetics , Glucose/pharmacology , Kidney/metabolism , Male , Molecular Structure , Nephrosis/blood , Nephrosis/drug therapy , Nephrosis/urine , Organ Specificity , Prednisolone/chemical synthesis , Prednisolone/pharmacokinetics , Prednisolone/pharmacology , Rats , Rats, Sprague-Dawley , Tissue DistributionABSTRACT
Aerosol glucocorticoid medications have become more and more important in treating BA (bronchial asthma). Although these agents are dosed to directly target airway inflammation, adrenocortical suppression and other systematic effects are still seen. To tackle this problem in a novel way, two L-carnitine ester derivatives of prednisolone (as the model drug), namely, PDC and PDSC, were synthesized to increase the absorption of prednisolone across the human bronchial epithelial BEAS-2B cells by the organic cation/carnitine transporter OCTN2 (SLC22A5) and then to slowly and intracellularly release prednisolone. The transport of prednisolone, PDC and PDSC into the human bronchial epithelial BEAS-2B cells was in the order PDSC > prednisolone > PDC at 37 °C. It was found that PDSC displayed 1.79-fold increase of uptake compared to prednisolone. Transport of PDSC by BEAS-2B was temperature-, time-, and Na(+)-dependent and saturable, with an apparent K(m) value of 329.74 µM, suggesting the involvement of carrier-mediated uptake. An RT-PCR study showed that organic cation/carnitine transporters OCTN1 and OCTN2 are expressed in BEAS-2B cells, but little in HEK293T cells. The order of uptake by HEK293T was prednisolone > PDC > PDSC. In addition, the inhibitory effects of organic cations such as L-carnitine, ergothioneine, TEA(+) and ipratropium on PDSC uptake in BEAS-2B cells were in the order L-carnitine > ipratropium > TEA(+) > ergothioneine, whereas their inhibitory effects on PDSC uptake in HEK293T cells were negligible. Finally, in vitro LPS-induced IL-6 production from BEAS-2B was more and longer suppressed by PDSC than prednisolone and PDC. All of these results suggested PDSC may be an attractive candidate for asthma treatment.
Subject(s)
Carnitine/analogs & derivatives , Glucocorticoids/chemical synthesis , Glucocorticoids/pharmacology , Prednisolone/analogs & derivatives , Prodrugs/chemical synthesis , Prodrugs/pharmacology , Animals , Anti-Inflammatory Agents/blood , Anti-Inflammatory Agents/chemical synthesis , Anti-Inflammatory Agents/pharmacokinetics , Anti-Inflammatory Agents/pharmacology , Biological Transport , Biotransformation , Bronchi/drug effects , Bronchi/immunology , Bronchi/metabolism , Carnitine/blood , Carnitine/chemical synthesis , Carnitine/pharmacology , Cell Line , Drug Stability , Gene Expression Regulation , Glucocorticoids/blood , Glucocorticoids/pharmacokinetics , HEK293 Cells , Humans , Interleukin-6/metabolism , Mice , Organic Cation Transport Proteins/antagonists & inhibitors , Organic Cation Transport Proteins/genetics , Organic Cation Transport Proteins/metabolism , Prednisolone/blood , Prednisolone/chemical synthesis , Prednisolone/pharmacology , Prodrugs/analysis , Prodrugs/pharmacokinetics , RNA, Messenger/metabolism , Respiratory Mucosa/drug effects , Respiratory Mucosa/immunology , Respiratory Mucosa/metabolism , Solute Carrier Family 22 Member 5 , Substrate Specificity , Succinates/blood , Succinates/chemical synthesis , Succinates/pharmacokinetics , Succinates/pharmacology , SymportersABSTRACT
Glucocorticoids (GCs) are routinely believed to take effect through genomic mechanisms, which are also largely responsible for GCs' side effects. Beneficial non-genomic effects of GCs have been reported as being independent of the genomic pathway. Here, we synthesized a new type of GCs, which took effect mainly via non-genomic mechanisms. Hydrocortisone was conjugated with glycine, lysine and phenylalanine to get a bigger molecular structure, which could hardly go through the cell membrane. Evaluation of the anti-inflammatory efficacy showed that hydrocortisone-conjugated glycine (HG) and lysine could inhibit neutrophil degranulation within 15 min. HG could inhibit IgE-mediated histamine release from mast cells via a non-genomic pathway, and rapidly alleviate allergic reaction. Luciferase reporter assay showed that HG would not activate the glucocorticoid response element within 30 min, which verified the rapid effects independent of the genomic pathway. The work proposes a novel insight into the development of novel GCs, and provides new tools for experimental study on non-genomic mechanisms.