New and improved glucocorticoid receptor ligands

The therapeutic and prophylactic use of glucocorticoids is widespread due to their powerful anti-inflammatory, antiproliferative and immunomodulatory activity. However, long-term use of these drugs can result in severe dose-limiting side effects. One of the most critical and debilitating side effects is osteoporosis, which leads to increased risk of fractures. Glucocorticoids damage bone through several different mechanisms. The search for novel glucocorticoids that have reduced side effects in bone and other tissues is being driven by the identification of new mechanisms of action of the glucocorticoid receptor. This may facilitate the detection of new, safer therapies with efficacies equivalent to currently prescribed steroids.     

Enhanced immune‐modulatory effects of thalidomide and dexamethasone co‐treatment on T cell subsets

Thalidomide (TM) has been reported to have anti‐cancer and anti‐inflammatory properties, and dexamethasone (DX) is known to reduce inflammation and inhibit production of inflammatory cytokines. Many studies have reported that combinatorial therapy with TM and DX is clinically used to treat multiple myeloma and lupus nephritis, but the mechanism responsible for its effects has not been elucidated. In this study, we determined that TM and DX co‐treatment had an enhanced immune‐modulatory effect on T cells through regulating the expression of co‐stimulatory molecules. Splenic naive T cells from C57BL/6 mice were sort‐purified and cultured for CD4⁺ T cell proliferation and regulatory T (Treg) cell conversion in the presence of TM and/or DX. Following incubation with the drugs, cells were collected and OX40, 4‐1BB, and glucocorticoid‐induced tumour necrosis factor receptor‐related protein (GITR) expression was quantified by flow cytometry. TM (1 or 10 μm ) decreased CD4⁺ T cell proliferation in a dose‐dependent manner, whereas TM/DX (0·1 or 1 nm ) co‐treatment further decreased proliferation. Treg cell populations were preserved following drug treatment. Furthermore, expression of co‐stimulatory molecules decreased upon TM/DX co‐treatment in effector T (Teff) cells and was preserved in Treg cells. Splenic CD4⁺ T cells isolated from TM‐ and DX‐treated mice exhibited the same patterns of Teff and Treg cell populations as observed in vitro. Considering the selective effect of TM on different T cell subsets, we suggest that TM may play an immunomodulatory role and that TM/DX combinatorial treatment could further enhance these immunomodulatory effects by regulating GITR, OX40, and 4‐1BB expression in CD4⁺ T cells.     

支气管哮喘患者外周白细胞糖皮质激素的受体变化

用放射配基结合分析法测定了外源性及内源性哮喘患者外周白细胞糖皮质激素受体（ＧＣＲ）水平，同时测定其血浆皮质醇水平。结果发现哮喘患者ＧＣＲ水平较正常对照组明显降低（Ｐ＜０．０１），其降低程度与疾病初发年龄、病程密切相关，而哮喘患者血浆皮质醇水平与正常对照组无显著差异性。提示哮喘患者肾上腺皮质功能尚未受到明显损害，但其ＧＣＲ功能存在异常。关于哮喘患者ＧＣＲ功能异常原因，我们推测与哮喘患者遗传倾向有关，同时与环境因素反复刺激、疾病反复发作有关。     

Novel glucocorticoid receptor agonists in the treatment of asthma

Introduction: Inhaled corticosteroids are the only drugs that effectively suppress the airway inflammation, but they can induce considerable systemic and adverse effects when they are administered chronically at high doses. Consequently, the pharmaceutical industry is still searching for newer entities with an improved therapeutic index.

Areas covered: Herein, the authors review the research in the glucocorticoid field to identify ligands of the glucocorticoid receptor (GR). These ligands preferentially induce transrepression with little or no transactivating activity, in order to have a potent anti-inflammatory action and a low side-effects profile.

Expert opinion: Several agents have been synthesized, but few have been tested in experimental models of asthma. Furthermore, only three (BI-54903, GW870086X and AZD5423) have entered clinical development, although the development of at least one of them (BI-54903) was discontinued. The reason for the limited success so far obtained is that the model of transactivation versus transrepression is a too simplistic representation of GR activity. It is difficult to uncouple the therapeutic and harmful effects mediated by GR, but some useful information that might change the current perspective is appearing in the literature. The generation of gene expression ‘fingerprints’ produced by different GR agonists in target and off-target human tissues could be useful in identifying drug candidates with an improved therapeutic ratio.     

Responses of serum corticosterone and corticosteroid-binding globulin to acute and prolonged stress in the rat

Responses of serum corticosterone (B) and corticosteroid-binding globulin (CBG) to ether anesthesia (a “classic” acute stress) and to a number of stressors influencing metabolic homeostasis—fasting, physical exercise, cold exposure, and water deprivation—were studied in male and female rats. Metabolic stressors included placing in an ice bath, physical exercise (swimming), fasting for 2 d, swimming after fasting for 2 d, cold-room (4°C) exposure for 2 d, fasting in combination with cold-room exposure for 1 d, and water deprivation for 2 d. The study demonstrated clear differences between males and females in basal B levels and B responses to some stressors. Only ether anesthesia and fasting resulted in similar B levels in males and females whereas in control and all other groups serum B levels were higher in females. Serum CBG was considerably higher in females. In females, ether, swimming, swimming after fasting, fasting, and fasting during cold exposure resulted in a decrease in circulating CBG. Ice bathing and cold exposure did not influence CBG, and water deprivation elevated serum CBG. In males, animals subjected to fasting and fasting during cold exposure had CBG levels lower than control animals. Other groups did not differ from the control. Higher CBG levels in females counterbalanced higher total B in setting circulating free B: significant sex differences in free B were observed only after swimming or fasting during cold exposure. Stress-responsive changes in CBG levels seem to contribute little to changes in free B; the main contributing factor is the rise in total B. However, CBG may play a special role, independent of the functions of corticosteroids. It is proposed that the need for substantial mobilization of spare fuel (as it takes place during physical exercise or fasting) is critical in involving CBG in the stress response.     

A novel mutation of sgk-1 gene in central serous chorioretinopathy

AIM

To investigate the association of serum glucocorticoid kinase gene-1 (SGK-1) DNA variants with chronic central serous chorioretinopathy (CSC).

METHODS

We enrolled 32 eyes of 32 patients who were diagnosed with chronic CSC and composed 32 normal eyes as a control group. Peripheral blood was used for DNA extraction and polymerase chain reaction (PCR) amplification. SGK1 gene was sequenced by using BigDye^® Terminator v3.1 cycle sequencing KIT (Applied Biosystems, Foster City, CA, USA). The SGK1 gene and its variants were investigated in CSC patient group and control group.

RESULTS

We identified a new polymorphism M32V in two person in the patient group (Minor allele frequency (MAF)=0.009) on the region of 1-60 amino acids. The rs1057293 was located in the encoder region of the SGK 1 gene but not associated with CSC (P=0.68). An intrinsic rs1743966 is also not associated (P=0.28).

CONCLUSIONS

The new polymorphism M32V is located on the region of 1-60 amino acids which is necessary for localization to the mitochondria in CSC patient. This mutation is probably important for the energy metabolism and plays an important role in the cellular response to hyperosmotic stress and other stress stimuli. Both rs1057293 and rs1743966 are not associated with CSC.     

Synthesis of two potent glucocorticoid receptor agonists labeled with carbon‐14 and stable isotopes

Two potent glucocorticoid receptor agonists were prepared labeled with carbon‐14 and with stable isotopes to perform drug metabolism, pharmacokinetics, and bioanalytical studies. Carbon‐14 labeled (1) was obtained from an enantiopure alkyne (5) via a Sonogashira coupling to a previously reported 5‐amino‐4‐iodo‐[2‐¹⁴C]pyrimidine [¹⁴C]‐(6), followed by a base‐mediated cyclization (1) in 72% overall radiochemical yield. Carbon‐14 labeled (2) was prepared in five steps employing a key benzoic acid intermediate [¹⁴C]‐(13), which was synthesized in one pot from enolization of trifluoromethylketone (12), followed by bromine–magnesium exchange and then electrophile trapping reaction with [¹⁴C]‐carbon dioxide. A chiral auxiliary (S)‐1‐(4‐methoxyphenyl)ethylamine was then coupled to this acid to give [¹⁴C]‐(15). Propargylation and separation of diastereoisomers by crystallizations gave the desired diastereomer [¹⁴C]‐(17) in 34% yield. Sonogashira coupling to iodopyridine (10) followed by cyclization to the azaindole [¹⁴C]‐(18) and finally removal of the chiral auxiliary gave [¹⁴C]‐(2) in 7% overall yield. For stable isotope syntheses, [¹³C₆]‐(1) was obtained in three steps using [¹³C₄]‐(6) and trimethylsilylacetylene‐[¹³C₂] in 26% yield, while [²H₅]‐(2) was obtained by first preparing the iodopyridine [²H₅]‐(10) in five steps. Then, Sonogashira coupling to chiral alkyne (24) and cyclization gave [²H₅]‐(2) in 42% overall yield.     

山楂酸对糖皮质激素诱发骨质疏松大鼠的干预作用研究

目的：研究山楂酸对糖皮质激素造成大鼠骨质疏松的干预作用及其作用机制。方法60只SD大鼠分为6组,正常组、模型组、对照组（灌胃给予碳酸钙375 mg? kg－1? d－1及维生素D 350 U? kg－1? d－1）及3个剂量实验组（灌胃给予山楂酸100,50,25 mg? kg－1? d－1）,共给药12周。生化试剂盒测定血清中的钙、磷的含量、骨钙素含量和抗酒石酸酸性磷酸酶活性,双能X线骨密度测定仪测定骨密度。结果与正常组相比,泼尼松可导致骨无机质钙盐和有机质羟脯胺酸下降,骨密度下降。山楂酸能对抗由泼尼松引起的上述异常,在增加骨密度方面优于对照组。结论山楂酸能有效预防糖皮质激素造成的大鼠骨质疏松。     

An altered composition of the microbiome in microscopic colitis is driven towards the composition in healthy controls by treatment with budesonide

Background and aim: Microscopic colitis (MC) is an inflammatory disease of the bowel, hypothetically induced by an immunologic response to a luminal microbial agent. We aimed to characterize the microbiome composition in MC and subtypes collagenous colitis (CC) and lymphocytic colitis (LC) and to identify a possible microbial effect of treatment.

Method: Stool samples were collected from MC patients prior to treatment, at 8 weeks (during treatment) and at 16 weeks (after treatment), and from healthy controls, not receiving treatment, at matched time-points. Microbiome composition was analyzed by sequencing of the 16S and 18S genes. Differences between patients and controls were analyzed by Shannon’s diversity index (mean, standard deviation (SD)) and principal coordinate analysis (PCoA) complemented with a permanova test of UniFrac distances.

Results: Ten LC patients, 10?CC patients and 10 controls were included. By PCoA, the bacterial composition in MC patients differed from controls at baseline (p?=?.02), but not during and after treatment (p?=?.09 and p?=?.33, respectively). At baseline, bacterial diversity was lower in MC patients compared to controls (2.5, SD: 0.5 vs 3.5, SD: 0.3, p?<?.05). Diversity in MC patients increased during (3.0, SD: 0.6) and after treatment and (2.9, SD: 0.5) compared with baseline (p?<?.01). Eukaryotes were detected in fewer samples from MC patients compared with controls (11/20 (55%) vs. 9/10 (90%), p?=?.06) with no effect of treatment.

Conclusion: Microbiome composition is altered in MC patients. During and after treatment with budesonide the microbiome composition in MC patients was driven towards the composition in healthy controls.