Microneedle-Mediated Delivery of Immunomodulators Restores Immune Privilege in Hair Follicles and Reverses Immune-Mediated Alopecia.

Disorders in the regulatory arm of the adaptive immune system result in autoimmune-mediated diseases. While systemic immunosuppression is the prevailing approach to manage them, it fails to achieve long-lasting remission due to concomitant suppression of the regulatory arm and carries the risk of heightened susceptibility to infections and malignancies. Alopecia areata is a condition characterized by localized hair loss due to autoimmunity. The accessibility of the skin allows local rather than systemic intervention to avoid broad immunosuppression. It is hypothesized that the expansion of endogenous regulatory T cells (Tregs) at the site of antigen encounter can restore the immune balance and generate a long-lasting tolerogenic response. A hydrogel microneedle (MN) patch is therefore utilized for delivery of CCL22, a Treg-chemoattractant, and IL-2, a Treg survival factor to amplify them. In an immune-mediated murine model of alopecia, local bolstering of Treg numbers is shown, leading to sustained hair regrowth and attenuation of inflammatory pathways. In a humanized skin transplant mouse model, expansion of Tregs within human skin is confirmed without engendering peripheral immunosuppression. The patch offers high-loading capacity and shelf-life stability for prospective clinical translation. By harmonizing immune responses locally, the aim is to reshape the landscape of autoimmune skin disease management.

Transmembrane protein TMEM97 and epigenetic reader BAHCC1 constitute an axis that supports pro-inflammatory cytokine expression.

Pro-inflammatory cytokine production by the retinal pigment epithelium (RPE) is a key etiology in retinal degenerative diseases, yet the underlying mechanisms are not well understood. TMEM97 is a scarcely studied transmembrane protein recently implicated in retinal degeneration. BAH domain coiled coil 1 (BAHCC1) is a newly discovered histone code reader involved in oncogenesis. A role for TMEM97 and BAHCC1 in RPE inflammation was not known. Here we found that they constitute a novel axis regulating pro-inflammatory cytokine expression in RPE cells. Transcriptomic analysis using a TMEM97-/- ARPE19 human cell line and the validation via TMEM97 loss- and gain-of-function revealed a profound role of TMEM97 in promoting the expression of pro-inflammatory cytokines, notably IL1ß and CCL2, and unexpectedly BAHCC1 as well. Moreover, co-immunoprecipitation indicated an association between the TMEM97 and BAHCC1 proteins. While TMEM97 ablation decreased and its overexpression increased NFκB (p50, p52, p65), the master transcription factor for pro-inflammatory cytokines, silencing BAHCC1 down-regulated NFκB and downstream pro-inflammatory cytokines. Furthermore, in an RPE-damage retinal degeneration mouse model, immunofluorescence illustrated down-regulation of IL1ß and CCL2 total proteins and suppression of glial activation in the retina of Tmem97-/- mice compared to Tmem97+/+ mice. Thus, TMEM97 is a novel determinant of pro-inflammatory cytokine expression acting via a previously unknown TMEM97- > BAHCC1- > NFκB cascade. SYNOPSIS: Retinal pigment epithelium (RPE) inflammation can lead to blindness. We identify here a previously uncharacterized cascade that underlies RPE cell production of pro-inflammatory cytokines. Specifically, transmembrane protein TMEM97 positively regulates the recently discovered histone code reader BAHCC1, which in turn enhances pro-inflammatory cytokine expression via the transcription factor NFκB.

Bent π-Conjugation within a Macrocycle: Asymmetric Total Syntheses of Spirohexenolides A and B.

Macrocycles with bent π-conjugation motif are extremely rare in nature and synthetically daunting and anticancer haouamines and spirohexenolides were representative of such rare natural products with synthetically challenging bent π-conjugation within a macrocycle. While the total synthesis of haouamines has been elegantly achieved, spirohexenolides remains an unmet synthetic challenge due to the highly strained bent 1,3,5-triene conjugation within C15 macrocycle. Inspired by the chemical synthesis of cycloparaphenylenes (CPPs) and haouamines, herein we devise a synthetic strategy to overcome the highly strained bent 1,3,5-triene conjugation within the macrocycle and achieve the first, asymmetric total synthesis of spirohexenolides A (>20 mg) and B (>50 mg). Our synthesis features strategic design of ring-closing metathesis (RCM) macrocyclization followed by double dehydration to achieve the C15 macrocycle with the deformed nonplanar 1,3,5-triene conjugation. In addition, we have developed a new enantioselective construction of highly functionalized spirotetronate fragment (northeast moiety) through RCM and Ireland-Claisen rearrangement. Our in vitro bioassay studies reveal that both spirohexenolides are cytotoxic against a panel of human cancer cells with IC50 1.2-13.3 µM and spirohexenolide A is consistently more potent (up to 3 times) than spirohexenolide B, suggesting the importance of alcohol for their bioactivity and for medicinal chemistry development.

Follicle-stimulating hormone orchestrates glucose-stimulated insulin secretion of pancreatic islets.

Follicle-stimulating hormone (FSH) is involved in mammalian reproduction via binding to FSH receptor (FSHR). However, several studies have found that FSH and FSHR play important roles in extragonadal tissue. Here, we identified the expression of FSHR in human and mouse pancreatic islet ß-cells. Blocking FSH signaling by Fshr knock-out led to impaired glucose tolerance owing to decreased insulin secretion, while high FSH levels caused insufficient insulin secretion as well. In vitro, we found that FSH orchestrated glucose-stimulated insulin secretion (GSIS) in a bell curve manner. Mechanistically, FSH primarily activates Gαs via FSHR, promoting the cAMP/protein kinase A (PKA) and calcium pathways to stimulate GSIS, whereas high FSH levels could activate Gαi to inhibit the cAMP/PKA pathway and the amplified effect on GSIS. Our results reveal the role of FSH in regulating pancreatic islet insulin secretion and provide avenues for future clinical investigation and therapeutic strategies for postmenopausal diabetes.

Long non-coding RNA BRE-AS1 inhibits proliferation, migration and invasion of clear cell renal cell carcinoma by downregulating miR-106b-5p.

OBJECTIVE: The objective of this study was to investigate the involvement of the long non-coding RNA (lncRNA) BRE-AS1 in clear cell renal cell carcinoma (ccRCC) and to explore its potential therapeutic role. METHODS: The expression of BRE-AS1 and miR-106b-5p was determined by qRT-PCR. Overexpression of BRE-AS1 and miR-106b-5p were performed to study their relationship. Transwell assays were used to evaluate cell movement. Methylation-specific PCR (MSP) was performed to explore the role of BRE-AS1 in the methylation of the miR-106b-5p gene. RESULTS: The results showed that the expression levels of BRE-AS1 were decreased, while those of miR-106b-5p were increased in ccRCC tissues. BRE-AS1 was found to be closely associated with the prognosis of patients with ccRCC. The expression of BRE-AS1 was inversely correlated with that of miR-106b-5p in tumor tissues. Overexpression of BRE-AS1 led to decreased expression levels of miR-106b-5p and increased methylation of the miR-106b-5p gene, whereas miR-106b-5p did not affect the expression of BRE-AS1. BRE-AS1 inhibited the movement and proliferation of ccRCC cell lines, while miR-106b-5p suppressed the role of BRE-AS1. CONCLUSION: BRE-AS1 may suppress ccRCC by downregulating the expression of miR-106b-5p.

Aerobic exercises regulate the epididymal anion homeostasis of high-fat diet-induced obese rats through TRPA1-mediated Cl- and HCO3- secretion†.

Aerobic exercises could improve the sperm motility of obese individuals. However, the underlying mechanism has not been fully elucidated, especially the possible involvement of the epididymis in which sperm acquire their fertilizing capacity. This study aims to investigate the benefit effect of aerobic exercises on the epididymal luminal milieu of obese rats. Sprague-Dawley male rats were fed on a normal or high-fat diet (HFD) for 10 weeks and then subjected to aerobic exercises for 12 weeks. We verified that TRPA1 was located in the epididymal epithelium. Notably, aerobic exercises reversed the downregulated TRPA1 in the epididymis of HFD-induced obese rats, thus improving sperm fertilizing capacity and Cl- concentration in epididymal milieu. Ussing chamber experiments showed that cinnamaldehyd (CIN), agonist of TRPA1, stimulated an increase of the short-circuit current (ISC) in rat cauda epididymal epithelium, which was subsequently abolished by removing the ambient Cl- and HCO3-. In vivo data revealed that aerobic exercises increased the CIN-stimulated Cl- secretion rate of epididymal epithelium in obese rats. Pharmacological experiments revealed that blocking cystic fibrosis transmembrane regulator (CFTR) and Ca2+-activated Cl- channel (CaCC) suppressed the CIN-stimulated anion secretion. Moreover, CIN application in rat cauda epididymal epithelial cells elevated intracellular Ca2+ level, and thus activate CACC. Interfering with the PGHS2-PGE2-EP2/EP4-cAMP pathway suppressed CFTR-mediated anion secretion. This study demonstrates that TRPA1 activation can stimulate anion secretion via CFTR and CaCC, which potentially forming an appropriate microenvironment essential for sperm maturation, and aerobic exercises can reverse the downregulation of TRPA1 in the epididymal epithelium of obese rats.

Environmental epigenetic interaction of gametes and early embryos†.

In recent years, the developmental origins of diseases have been increasingly recognized and accepted. As such, it has been suggested that most adulthood chronic diseases such as diabetes, obesity, cardiovascular disease, and even tumors may develop at a very early stage. In addition to intrauterine environmental exposure, germ cells carry an important inheritance role as the primary link between the two generations. Adverse external influences during differentiation and development can cause damage to germ cells, which may then increase the risk of chronic disease development later in life. Here, we further elucidate and clarify the concept of gamete and embryo origins of adult diseases by focusing on the environmental insults on germ cells, from differentiation to maturation and fertilization.

Integrated Multi-Omics Analysis Reveals the Effect of Maternal Gestational Diabetes on Fetal Mouse Hippocampi.

Growing evidence suggests that adverse intrauterine environments could affect the long-term health of offspring. Recent evidence indicates that gestational diabetes mellitus (GDM) is associated with neurocognitive changes in offspring. However, the mechanism remains unclear. Using a GDM mouse model, we collected hippocampi, the structure critical to cognitive processes, for electron microscopy, methylome and transcriptome analyses. Reduced representation bisulfite sequencing (RRBS) and RNA-seq in the GDM fetal hippocampi showed altered methylated modification and differentially expressed genes enriched in common pathways involved in neural synapse organization and signal transmission. We further collected fetal mice brains for metabolome analysis and found that in GDM fetal brains, the metabolites displayed significant changes, in addition to directly inducing cognitive dysfunction, some of which are important to methylation status such as betaine, fumaric acid, L-methionine, succinic acid, 5-methyltetrahydrofolic acid, and S-adenosylmethionine (SAM). These results suggest that GDM affects metabolites in fetal mice brains and further affects hippocampal DNA methylation and gene regulation involved in cognition, which is a potential mechanism for the adverse neurocognitive effects of GDM in offspring.

Mammalian hybrid pre-autophagosomal structure HyPAS generates autophagosomes.

The biogenesis of mammalian autophagosomes remains to be fully defined. Here, we used cellular and in vitro membrane fusion analyses to show that autophagosomes are formed from a hitherto unappreciated hybrid membrane compartment. The autophagic precursors emerge through fusion of FIP200 vesicles, derived from the cis-Golgi, with endosomally derived ATG16L1 membranes to generate a hybrid pre-autophagosomal structure, HyPAS. A previously unrecognized apparatus defined here controls HyPAS biogenesis and mammalian autophagosomal precursor membranes. HyPAS can be modulated by pharmacological agents whereas its formation is inhibited upon severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection or by expression of SARS-CoV-2 nsp6. These findings reveal the origin of mammalian autophagosomal membranes, which emerge via convergence of secretory and endosomal pathways, and show that this process is targeted by microbial factors such as coronaviral membrane-modulating proteins.

Long non-coding RNA MALAT1 regulates cell proliferation, invasion and apoptosis by modulating the Wnt signaling pathway in squamous cell carcinoma.

OBJECTIVE: To explore the mechanisms by which long non-coding RNA, (lncRNA) metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) regulates cell proliferation, invasion and apoptosis through signaling axes in cutaneous squamous cell carcinoma (CSCC) cells. METHODS: A total of 60 CSCC samples and 15 normal skin tissue samples were collected. qRT-PCR was used to determine MALAT1 expression. After knockdown of MALAT1 expression in A431 cells, Transwell assay was performed to detect cell migration and invasion, CCK8 assay was used to detect cell proliferation, and Western blotting was used to detect EMT-related protein expression. RESULTS: Compared with the normal group, the MALAT1 positive expression rate was significantly higher in the low, moderate, and high differentiation groups (P < 0.05). The expression of MALATl in A431 cells in the siMALATl-1 and siMALATl-2 groups was lower than that in siNC group (P < 0.05). A431 cell proliferation, invasion and apoptosis at 24 h, 48 h and 72 h in the siMALATl-1 and siMALATl-2 groups were all lower and the apoptosis rate of A431 cells were all higher than that of the siNC group (P < 0.05). E-cadherin expression was higher while the expression of ß-catenin, vimentin, and Bcl-2 was lower in the siMALATl-1 and siMALATl-2 groups than those of the siNC group (P < 0.05). CONCLUSION: Down-regulation of lncRNA MALAT1 expression may promote apoptosis of CSCC cells and inhibit cell migration, invasion and proliferation by regulating the Wnt signaling pathway.

Intrauterine Hyperglycemia Alters the Metabolomic Profile in Fetal Mouse Pancreas in a Gender-Specific Manner.

Mounting evidence has shown that intrauterine hyperglycemia exposure during critical stages of development may be contributing to the increasing prevalence of diabetes. However, little is known about the mechanisms responsible for offspring metabolic disorder. In this present study, we explored intrauterine hyperglycemia exposure on fetal pancreatic metabolome, and its potential link to impaired glucose tolerance in adult offspring. Here, using a GDM mouse model, we found the metabolome profiling of pancreas from male and female fetus showing altered metabolites in several important pathways, including 5-methylcytosine, α-KG, branched-chain amino acids, and cystine, which are associated with epigenetic modification, insulin secretion, and intracellular redox status, respectively. This finding suggests that intrauterine exposure to hyperglycemia could cause altered metabolome in pancreas, which might be a metabolism-mediated mechanism for GDM-induced intergenerational diabetes predisposition.

Increased Susceptibility and Intrinsic Apoptotic Signaling in Neurons by Induced HDAC3 Expression.

Purpose: Inhibition or targeted deletion of histone deacetylase 3 (HDAC3) is neuroprotective in a variety neurodegenerative conditions, including retinal ganglion cells (RGCs) after acute optic nerve damage. Consistent with this, induced HDAC3 expression in cultured cells shows selective toxicity to neurons. Despite an established role for HDAC3 in neuronal pathology, little is known regarding the mechanism of this pathology. Methods: Induced expression of an HDAC3-mCherry fusion protein in mouse RGCs was accomplished by transduction with AAV2/2-Pgk-HDAC3-mCherry. Increased susceptibility to optic nerve damage in HDAC3-mCherry expressing RGCs was evaluated in transduced mice that received acute optic nerve crush surgery. Expression of HDAC3-FLAG or HDAC3-mCherry was induced by nucleofection or transfection of plasmids into differentiated or undifferentiated 661W tissue culture cells. Immunostaining for cleaved caspase 3, localization of a GFP-BAX fusion protein, and quantitative RT-PCR was used to evaluate HDAC3-induced damage. Results: Induced expression of exogenous HDAC3 in RGCs by viral-mediated gene transfer resulted in modest levels of cell death but significantly increased the sensitivity of these neurons to axonal damage. Undifferentiated 661W retinal precursor cells were resilient to induced HDAC3 expression, but after differentiation, HDAC3 induced GFP-BAX recruitment to the mitochondria and BAX/BAK dependent activation of caspase 3. This was accompanied by an increase in accumulation of transcripts for the JNK2/3 kinases and the p53-regulated BH3-only gene Bbc3/Puma. Cell cycle arrest of undifferentiated 661W cells did not increase their sensitivity to HDAC3 expression. Conclusions: Collectively, these results indicate that HDAC3-induced toxicity to neurons is mediated by the intrinsic apoptotic pathway.

Diverse expression patterns of mucin 2 in colorectal cancer indicates its mechanism related to the intestinal mucosal barrier.

BACKGROUND: Abnormal expression patterns of mucin 2 (MUC2) have been reported in a variety of malignant tumors and precancerous lesions. Reduced MUC2 expression in the intestinal mucosa, caused by various pathogenic factors, is related to mechanical dysfunction of the intestinal mucosa barrier and increased intestinal mucosal permeability. However, the relationship between MUC2 and the intestinal mucosal barrier in patients with colorectal cancer (CRC) is not clear. AIM: To explore the relationship between MUC2 and intestinal mucosal barrier by characterizing the multiple expression patterns of MUC2 in CRC. METHODS: Immunohistochemical staining was performed on intestinal tissue specimens from 100 CRC patients, including both cancer tissues and adjacent normal tissues. Enzyme-linked immunosorbent assays were performed on preoperative sera from 66 CRC patients and 20 normal sera to detect the serum levels of MUC2, diamine oxide (DAO), and D-lactate (D-LAC). The relationship between MUC2 expression and clinical parameters was calculated by the χ 2 test or Fisher's exact test. Prognostic value of MUC2 was evaluated by Kaplan-Meier curve and log-rank tests. RESULTS: Immunohistochemical staining of 100 CRC tissues showed that the expression of MUC2 in cancer tissues was lower than that in normal tissues (54% vs 79%, P < 0.05), and it was correlated with tumor-node-metastasis (TNM) stage and lymph node metastasis in CRC patients (P < 0.05). However, the serum level of MUC2 in CRC patients was higher than that in normal controls, and was positively associated with serum levels of human DAO (χ 2 = 3.957, P < 0.05) and D-LAC (χ 2 = 7.236, P < 0.05), which are the biomarkers of the functional status of the intestinal mucosal barrier. And the serum level of MUC2 was correlated with TNM stage, tumor type, and distant metastasis in CRC patients (P < 0.05). Kaplan-Meier curves showed that decreased MUC2 expression in CRC tissues predicted a poor survival. CONCLUSION: MUC2 in tissues may play a protective role by participating in the intestinal mucosal barrier and can be used as an indicator to evaluate the prognosis of CRC patients.

Immune Checkpoint Inhibitors-Related Thyroid Dysfunction: Epidemiology, Clinical Presentation, Possible Pathogenesis, and Management.

Immune checkpoint inhibitors (ICIs) are a group of drugs employed in the treatment of various types of malignant tumors and improve the therapeutic effect. ICIs blocks negative co-stimulatory molecules, such as programmed cell death gene-1 (PD-1) and its ligand (PD-L1) and cytotoxic T-lymphocyte-associated antigen-4 (CTLA-4), reactivating the recognition and killing effect of the immune system on tumors. However, the reactivation of the immune system can also lead to the death of normal organs, tissues, and cells, eventually leading to immune-related adverse events (IRAEs). IRAEs involve various organs and tissues and also cause thyroid dysfunction. This article reviews the epidemiology, clinical manifestations, possible pathogenesis, and management of ICIs-related thyroid dysfunction.

A hierarchical and collaborative BRD4/CEBPD partnership governs vascular smooth muscle cell inflammation.

Bromodomain protein BRD4 reads histone acetylation (H3K27ac), an epigenomic mark of transcription enhancers. CCAAT enhancer binding protein delta (CEBPD) is a transcription factor typically studied in metabolism. While both are potent effectors and potential therapeutic targets, their relationship was previously unknown. Here we investigated their interplay in vascular smooth muscle cell (SMC) inflammation. Chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq) revealed H3K27ac/BRD4 enrichment at Cebpd in injured rat carotid arteries. While genomic deletion of BRD4-associated enhancer in SMCs in vitro decreased Cebpd transcripts, BRD4 gene silencing also diminished Cebpd mRNA and protein, indicative of a BRD4 control over CEBPD expression. Bromodomain-1, but not bromodomain-2, accounted for this BRD4 function. Moreover, endogenous BRD4 protein co-immunoprecipitated with CEBPD, and both proteins co-immunoprecipitated the Cebpd promoter and enhancer DNA fragments. These co-immunoprecipitations (coIPs) were all abolished by the BRD4-bromodomain blocker JQ1, suggesting a BRD4/CEBPD /promoter/enhancer complex. While BRD4 and CEBPD were both upregulated upon tumor necrosis factor alpha (TNF-α) stimulation of SMC inflammation (increased interleukin [IL]-1b, IL-6, and MCP-1), they mediated this stimulation via preferentially elevated expression of platelet-derived growth factor receptor alpha (PDGFRα, versus PDGFRß), as indicated by loss- and gain-of-function experiments. Taken together, our study unravels a hierarchical yet collaborative BRD4/CEBPD relationship, a previously unrecognized mechanism that prompts SMC inflammation and may underlie other pathophysiological processes as well.

Efficacy and safety of a selective URAT1 inhibitor SHR4640 in Chinese subjects with hyperuricaemia: a randomized controlled phase II study.

OBJECTIVE: To evaluate the efficacy and safety of SHR4640, a highly selective urate transporter 1 inhibitor, in Chinese subjects with hyperuricaemia. METHODS: This was a randomized double-blind dose-ranging phase II study. Subjects whose serum uric acid (sUA) levels were ≥480 µmol/l with gout, ≥480 µmol/l without gout but with comorbidities, or ≥540 µmol/l were enrolled. Subjects were randomly assigned (1:1:1:1:1) to receive once daily 2.5 mg, 5 mg, 10 mg of SHR4640, 50 mg of benzbromarone or placebo, respectively. The primary end point was the proportion of subjects who achieved target sUA level of ≤360 µmol/l at week 5. RESULTS: 99.5% of subjects (n = 197) were male and 95.9% of subjects had gout history. The proportions of subjects who achieved target sUA at week 5 were 32.5%, 72.5% and 61.5% in the 5 mg, 10 mg SHR4640 and benzbromarone groups, respectively, significantly higher than the placebo group (0%; P < 0.05 for 5 mg and 10 mg SHR4640 group). The sUA was reduced by 32.7%, 46.8% and 41.8% at week 5 with 5 mg, 10 mg SHR4640 and benzbromarone, respectively, vs placebo (5.9%; P < 0.001 for each comparison). The incidences of gout flares requiring intervention were similar among all groups. Occurrences of treatment-emergent adverse events (TEAEs) were comparable across all groups, and serious TEAEs were not reported. CONCLUSIONS: The present study indicated a superior sUA-lowering effect and well tolerated safety profile after 5-week treatment with once-daily 5 mg/10 mg of SHR4640 as compared with placebo in Chinese subjects with hyperuricaemia. TRIAL REGISTRATION: ClinicalTrials.gov number, NCT03185793.

Metabolic disposition of [14 C]-abivertinib, an epidermal growth factor receptor tyrosine kinase inhibitor: Role of glutathione conjugation.

AIMS: To determine the absorption, distribution, metabolism and excretion of abivertinib, a third-generation epidermal growth factor receptor tyrosine kinase inhibitor, in patients with advanced non-small cell lung cancer (NSCLC). METHODS: Seven patients with advanced NSCLC were given a single 200 mg/83 µCi oral suspension of [14 C]-abivertinib. Blood, urine and faeces were collected. Mass balance of radioactivity, the pharmacokinetics of abivertinib, and the total radioactivity were determined. Metabolite profiling and characterisation were performed. RESULTS: The mean recovery was 82.16%, with 2.38 and 79.78% of the radioactive dose excreted in urine and faeces, respectively. The unchanged abivertinib was the major radioactive component detected in plasma within the first 24 hours after dosing, accounting for 59.17% of the total drug-related radioactivity. Abivertinib in urine accounted for only 0.96% of the administered dose, whereas in faeces it accounted for 33.36%. Eight metabolites were detected and characterised in plasma, among which MII-7, a product of cysteine glycine conjugate, was the only circulating metabolite, accounting for approximate 10.6% of the total drug-related exposure. MII-2 (an abivertinib cysteine-glycine adduct) and M7 (a reduced product of abivertinib) were the 2 major metabolites in the excreta, accounting for 20.0 and 12.4%, respectively, of the drug-related radioactivity in faeces. CONCLUSION: Following a single oral administration, the unchanged abivertinib was the predominant drug-related material in plasma, urine and faeces. The drug-related materials were primarily eliminated via the faecal route. Direct glutathione conjugation of abivertinib played a significant role in the metabolic clearance and metabolite exposure of abivertinib.

BRD2 regulation of sigma-2 receptor upon cholesterol deprivation.

The sigma-2 receptor (S2R) has long been pharmacologically targeted for antipsychotic treatment and tumor imaging. Only recently was it known for its coding gene and for its role implicated in cholesterol homeostasis. Here, we have investigated the transcriptional control of S2R by the Bromo/ExtraTerminal epigenetic reader family (BETs, including BRD2, 3, and 4) upon cholesterol perturbation. Cholesterol deprivation was induced in ARPE19 cells using a blocker of lysosomal cholesterol export. This condition up-regulated S2R mRNA and protein, and also SREBP2 but not SREBP1, both transcription factors key to cholesterol/fatty acid metabolism. Silencing BRD2 but not BRD3 or BRD4 (though widely deemed a master regulator) averted S2R up-regulation that was induced by cholesterol deprivation. Silencing SREBP2 but not SREBP1 diminished S2R expression. Furthermore, endogenous BRD2 co-immunoprecipitated with the transcription-active N-terminal half of SREBP2, and chromatin immunoprecipitation-qPCR signified co-occupancy of BRD2, H3K27ac (histone acetylation), and SREBP2Nterm at the S2R gene promoter. In summary, this study reveals a previously unrecognized BRD2/SREBP2 cooperative regulation of S2R transcription, thus shedding new light on signaling in response to cholesterol deprivation.

Mass balance, metabolic disposition, and pharmacokinetics of [14C]ensartinib, a novel potent anaplastic lymphoma kinase (ALK) inhibitor, in healthy subjects following oral administration.

PURPOSE: Ensartinib is a novel, potent and highly selective inhibitor of anaplastic lymphoma kinase (ALK) that has promising clinical activity and low toxicity in patients with ALK-positive non-small cell lung cancer. This study was conducted to investigate the pharmacokinetics, metabolism and excretion of ensartinib following a single 200 mg/100 µCi oral dose of radiolabeled ensartinib to healthy subjects. METHODS: Six healthy male subjects were enrolled and administrated an oral suspension in a fasted state. Blood, urine and feces were collected. Radioactivity concentrations were measured by liquid scintillation counting and plasma concentrations of ensartinib by liquid chromatography-tandem mass spectrometry. Both techniques were applied for metabolite profiling and characterization. RESULTS: The mean total recovery was 101.21% of the radiolabeled dose with 91.00% and 10.21% excreted in feces and urine, respectively. Unchanged ensartinib was the predominant drug-related component in urine and feces, representing 4.39% and 38.12% of the administered dose, respectively. Unchanged ensartinib and its metabolite M465 were the major circulating components, accounting for the same 27.45% of the plasma total radioactivity (AUC0-24h pool), while other circulating metabolites were minor, accounting for less than 10%. Mean Cmax, AUC0-∞, T1/2 and Tmax values for ensartinib in plasma were 185 ng/mL, 3827 h ng/mL, 18.3 h and 3.25 h, respectively. The total radioactivity in plasma was cleared with terminal half-life of 27.2 h. Treatment with ensartinib was well tolerated, and no serious adverse events were reported. CONCLUSION: It was well tolerated in the six healthy male subjects following a single oral administration of 200 mg/100 µCi dose of ensartinib. Besides unchanged ensartinib, metabolite of M465 was the predominant circulating drug-related component. The drug was primarily eliminated in feces. CLINICAL TRIAL REGISTRATION: ClinicalTrials.gov NCT03804541.

[Effect of new simple breathing apparatus on oxygen therapy in patients with severe and critical coronavirus disease 2019].

OBJECTIVE: To make a new simple respirator and observe the oxygen therapy effect of the respirator on patients with severe and critical coronavirus disease 2019 (COVID-19). METHODS: Based on the infectivity and hospital requirements of COVID-19, a new simple respirator was designed by the medical staff of the Department of Anesthesiology of the Second Affiliated Hospital of Nanchang University, which was applied on the 22 patients with severe and critical COVID-19 who needed oxygen therapy admitted to the Cancer Center of Tongji Medical College of Huazhong University of Science and Technology from February 15th to March 15th in 2020. The new simple respirator contained two National Utility Model Patents (a respirator: ZL 2015 2 0410623.6, a fluid switch and oxygen suction device: ZL 2017 2 0873509.6), which was mainly composed of anesthesia mask and filter, L-shaped connecting tube, soft breathing bladder, connecting tube and elastic fixing belt. When in use, the anesthesia mask was fixed to the patient's mouth and nose with elastic straps, the connecting tube was inserted into the oxygen meter interface, the oxygen flow was adjusted to 6-10 L/min, and the L-shaped connecting tube was opened immediately after the soft breathing bag was full. The carbon dioxide and excess oxygen in the body was discharged from exhaust port. The oxygen flow was lowered to 2-3 L/min, the patient's respiratory rate (RR) was observed through the soft breathing bag fluctuations, and the oxygen flow was adjusted at any time. The changes of pulse oxygen saturation (SpO2), RR and heart rate (HR) before and after application of new simple respirator were observed, and the blood gas test results of part of the patients were collected. RESULTS: Twenty-two patients with severe and critical COVID-19 had significantly higher SpO2 at 10 minutes after application of the new simple ventilator than before application (0.994±0.007 vs. 0.952±0.017, P < 0.01), and RR was significantly lower than that before application (times/min: 27.59±3.63 vs. 29.64±3.81, P < 0.01); after 1 day of application, each index was further improved. All 13 patients who received blood gas analysis indicated no carbon dioxide accumulation. CONCLUSIONS: The new simple respirator can significantly improve the oxygen therapy effect of patients with severe and critical COVID-19. At the same time, 2019 novel coronavirus (2019-nCoV) can be filtered through the filter to reduce the formation of aerosol and protect the medical staff and patients.