Oxidative phosphorylation regulates B cell effector cytokines and promotes inflammation in multiple sclerosis.

Dysregulated B cell cytokine production contributes to pathogenesis of immune-mediated diseases including multiple sclerosis (MS); however, the underlying mechanisms are poorly understood. In this study we investigated how cytokine secretion by pro-inflammatory (GM-CSF-expressing) and anti-inflammatory (IL-10-expressing) B cells is regulated. Pro-inflammatory human B cells required increased oxidative phosphorylation (OXPHOS) compared with anti-inflammatory B cells. OXPHOS reciprocally modulated pro- and anti-inflammatory B cell cytokines through regulation of adenosine triphosphate (ATP) signaling. Partial inhibition of OXPHOS or ATP-signaling including with BTK inhibition resulted in an anti-inflammatory B cell cytokine shift, reversed the B cell cytokine imbalance in patients with MS, and ameliorated neuroinflammation in a myelin oligodendrocyte glycoprotein (MOG)-induced experimental autoimmune encephalitis mouse model. Our study identifies how pro- and anti-inflammatory cytokines are metabolically regulated in B cells and identifies ATP and its metabolites as a "fourth signal" that shapes B cell responses and is a potential target for restoring the B cell cytokine balance in autoimmune diseases.

Synergistic interaction between hyperlipidemia and obesity as a risk factor for stress urinary incontinence in Americans.

Urinary incontinence is a common disease among middle-aged and elderly women, which not only affects the physical and mental health of patients, but also brings a great medical burden to society. Obesity is a known risk factor for urinary incontinence and is the most common secondary cause of hyperlipidemia. Most obese patients also suffer from hyperlipidemia in the clinic. However, few studies have explored the role of hyperlipidemia in women with urinary incontinence. Using data from the 2005-2018 National Health and Nutrition Examination Survey (NHANES), we aimed to evaluated the independent associations of high body mass index and hyperlipidemia with urinary incontinence in Americans by conducting a weighted multivariate logistic regression model. Additive interactions were also assessed using the relative excess risk due to interaction (RERI), attributed proportion of interaction (AP) and synergy index (S). This study demonstrated that hyperlipidemia was associated with a higher risk of stress urinary incontinence among women with obesity (OR = 1.52, 95% CI = 1.03-2.25), and there was a significant synergistic effect of hyperlipidemia and obesity on stress urinary incontinence(adjusted RERI: 3.75, 95% CI 0.30-7.20; adjusted AP: 0.67, 95% CI 0.54-0.80; adjusted S: 5.49, 95% CI 4.15-7.27). Moreover, fasting serum triglyceride lipids were the most relevant blood lipid indicator for the risk of stress urinary incontinence, especially among obese women younger than 50 years old, which contributes to the development of more refined lipid control protocols for patients with urinary incontinence in different age groups.

Antitrypanosomal Chloronitrobenzamides.

Human African trypanosomiasis (HAT), a neglected tropical disease caused by Trypanosoma brucei gambiense (Tbg) or Trypanosoma brucei rhodesiense (Tbr), remains a significant public health concern with over 55 million people at risk of infection. Current treatments for HAT face the challenges of poor efficacy, drug resistance, and toxicity. This study presents the synthesis and evaluation of chloronitrobenzamides (CNBs) against Trypanosoma species, identifying previously reported compound 52 as a potent and selective orally bioavailable antitrypanosomal agent. 52 was well tolerated in vivo and demonstrated favorable oral pharmacokinetics, maintaining plasma concentrations surpassing the cellular EC50 for over 24 h and achieving peak brain concentrations exceeding 7 µM in rodents after single oral administration (50 mg/kg). Treatment with 52 significantly extended the lifespan of mice infected with Trypanosoma congolense and T. brucei rhodesiense. These results demonstrate that 52 is a strong antitrypanosomal lead with potential for developing treatments for both human and animal African trypanosomiasis.

The influence of biological sex in human skeletal muscle transcriptome during ageing.

Sex is a crucial biological variable, and influence of biological sex on the change of gene expression in ageing skeletal muscle has not yet been fully revealed. In this study, the mRNA expression profiles were obtained from the Gene Expression Omnibus database. Key genes were identified by differential expression analysis and weighted gene co-expression network analysis. The gene set enrichment analysis software and Molecular Signatures Database were used for functional and enrichment analysis. A protein-protein interaction network was constructed using STRING and visualized in Cytoscape. The results were compared between female and male subgroups. Differentially expressed genes and enriched pathways in different sex subgroups shared only limited similarities. The pathways enriched in the female subgroup were more similar to the pathways enriched in the older groups without taking sex difference into consideration. The pathways enriched in the female subgroup were more similar to the pathways enriched in the older groups without taking sex difference into consideration. The muscle myosin filament pathways were downregulated in the both aged female and male samples whereas transforming growth factor beta pathway and extracellular matrix-related pathways were upregulated. With muscle ageing, the metabolism-related pathways, protein synthesis and degradation pathways, results of predicted immune cell infiltration, and gene cluster associated with slow-type myofibers drastically different between the female and male subgroups. This finding may indicate that changes in muscle type with ageing may differ between the sexes in vastus lateralis muscle.

Acupuncture at ST36 ameliorates experimental autoimmune encephalomyelitis via affecting the function of B cells.

Acupuncture at ST36 can alleviate a variety of autoimmune diseases, including experimental autoimmune encephalomyelitis (EAE), while the specific mechanism for the treatment of EAE is not clear. In this study, we found that acupuncture at ST36 can significantly increase the excitability of splenic sympathetic nerve, and promote the differentiation of peripheral B and CD4+T cells in the anti-inflammatory direction. After blocking the splenic sympathetic nerve with 6-OHDA, this anti-inflammatory effect of acupuncture is partially reversed. In addition, the results of western blot and qPCR showed that acupuncture at ST36 simultaneously activated the ß2-AR-cAMP signaling pathway in the splenic B and CD4+T cells, and this activation was more significant in B cells. In vitro, when CD4+T cells were cultured alone, norepinephrine (NE) had no significant effect on their differentiation. While in the presence of B cells, NE significantly promotes the anti-inflammatory differentiation of B and CD4+T cells. Therefore, the above results reveal that acupuncture can relieve EAE by stimulating the sympathetic nerves of spleen, mainly through acting on B cells to mediate anti-inflammatory effects, and indirectly affecting the function of CD4+T cells.

Sex difference and risk factors in burden of urogenital congenital anomalies from 1990 to 2019.

Urogenital congenital anomalies (UCAs) is defined as "any live-birth with a urinary or genital condition" and affects millions of men and women worldwide. However, sex differences and related environmental risk factors in UCAs burden on a global scale have not been assessed. Using data from the Global Burden of Diseases, Injuries, and Risk Factors Study (GBD) 2019, we estimated prevalence, incidence, mortality and disability-adjusted life years (DALYs) of UCAs from 1990 to 2019 by sex, region, and socio-demographic Index (SDI) in 204 countries and territories. The disease burden of UCAs was also estimated attributable to each risk factor were estimated according to risk exposure. In 2019, UCAs caused 10,200 all-ages deaths (95% UI 7550-13,400). The combined global incidence rate was 8.38 per 1000 (95% UI 5.88-12.0) live births. The ASIR increased slightly, while the ASDR decreased from 1990 to 2019.The UCAs burden varies greatly depending on the development level and geographical location. The UCAs burden was significantly higher in men than in women, and the sex differences showed an enlarging trend. Health risks and issues, including pollution, child and maternal malnutrition, diet habits, unsafe sanitation and water source, were detected to be positively related to UCAs burden. Albeit the age-standardised prevalence, mortality, incidence, and DALYs of UCAs have decreased, they still cause a public health challenge worldwide. The high deaths and DALYs rates in low and low-middle SDI countries highlight the urgent need for improved preventive, diagnostic, and therapeutic measures. Global strategies for enhancing water safety, reducing pollution, and healthy diets are crucial steps in reducing the burden of UCAs.

Effects of Acupuncture on Adverse Events in Colonoscopy: A Systematic Review and Meta-analysis of Randomized Controlled Trials.

INTRODUCTION: Acupuncture has gradually penetrated into many disciplines in clinical medicine, such as surgery, anesthesia, and outpatient examinations. Although a number of clinical trials have investigated the effects of acupuncture on colonoscopy, the results were inconsistent. In this meta-analysis, we analyzed the effects of acupuncture on colonoscopy to provide evidence for subsequent research and clinical application of acupuncture in colonoscopy. METHODS: This meta-analysis was performed using Review Manager version 5.4 and Stata version 16 software. The primary outcome was the incidence of adverse events, and the secondary outcomes included patients' anxiety score before colonoscopy, time to insert the colonoscope, total detection time, propofol consumption, patients' pain score, and patient satisfaction rate. RESULTS: The results showed that the incidence of adverse events (odds ratio [OR] 0.27, 95% confidence interval [CI] 0.16-0.43, P = 0.00, I2 = 25%), patients' pain score (mean difference [MD] - 1.03, 95% CI - 1.45 to - 0.62, P = 0.00, I2 = 94%), and time to insert the colonoscope (MD = - 2.54, 95% CI - 4.96 to - 0.13, P = 0.04, I2 = 0%) were significantly lower in the treatment group than in the control group. Compared with the control group, the satisfaction rate of patients (OR 2.53, 95% CI 1.56-4.10, P = 0.00, I2 = 47%) in the treatment group was significantly improved. There was no significant between-group difference in patients' anxiety score, the total detection time, and propofol dosage. CONCLUSIONS: During colonoscopy, acupuncture can significantly reduce the incidence of adverse events, relieve patients' pain, and improve patient satisfaction. REGISTRATION: PROSPERO registration number CRD42022324428.

Therapeutic Effect and Mechanism of Acupuncture in Autoimmune Diseases.

Autoimmune diseases (AIDs) are conditions arising from abnormal immune reactions to autoantigens, which can be defined as the loss of immune tolerance to autoantigens, causing the production of autoantibodies and subsequent inflammation and tissue injury. The etiology of AIDs remains elusive, which may involve both genetic and environmental factors, such as diet, drugs, and infections. Despite rapid progress in the treatment of autoimmune diseases over the past few decades, there is still no approach that can cure AIDs. As an alternative approach, traditional Chinese medicine (TCM) such as acupuncture has been used in an attempt to treat AIDs including multiple sclerosis (MS), rheumatoid arthritis (RA), and inflammatory bowel disease (IBD), and the results have proven to be quite promising, despite the fact that its mechanism is still not fully understood. In this review, the present knowledge regarding mechanisms of acupuncture in the treatment of AIDs has been summarized, and deeper insights will be provided in order to better understand how acupuncture may regulate immune responses during AIDs.

Selecting an anti-malarial clinical candidate from two potent dihydroisoquinolones.

BACKGROUND: The ongoing global malaria eradication campaign requires development of potent, safe, and cost-effective drugs lacking cross-resistance with existing chemotherapies. One critical step in drug development is selecting a suitable clinical candidate from late leads. The process used to select the clinical candidate SJ733 from two potent dihydroisoquinolone (DHIQ) late leads, SJ733 and SJ311, based on their physicochemical, pharmacokinetic (PK), and toxicity profiles is described. METHODS: The compounds were tested to define their physicochemical properties including kinetic and thermodynamic solubility, partition coefficient, permeability, ionization constant, and binding to plasma proteins. Metabolic stability was assessed in both microsomes and hepatocytes derived from mice, rats, dogs, and humans. Cytochrome P450 inhibition was assessed using recombinant human cytochrome enzymes. The pharmacokinetic profiles of single intravenous or oral doses were investigated in mice, rats, and dogs. RESULTS: Although both compounds displayed similar physicochemical properties, SJ733 was more permeable but metabolically less stable than SJ311 in vitro. Single dose PK studies of SJ733 in mice, rats, and dogs demonstrated appreciable oral bioavailability (60-100%), whereas SJ311 had lower oral bioavailability (mice 23%, rats 40%) and higher renal clearance (10-30 fold higher than SJ733 in rats and dogs), suggesting less favorable exposure in humans. SJ311 also displayed a narrower range of dose-proportional exposure, with plasma exposure flattening at doses above 200 mg/kg. CONCLUSION: SJ733 was chosen as the candidate based on a more favorable dose proportionality of exposure and stronger expectation of the ability to justify a strong therapeutic index to regulators.

Piperidinyl Ureas Chemically Control Defective in Cullin Neddylation 1 (DCN1)-Mediated Cullin Neddylation.

We previously discovered and validated a class of piperidinyl ureas that regulate defective in cullin neddylation 1 (DCN1)-dependent neddylation of cullins. Here, we report preliminary structure-activity relationship studies aimed at advancing our high-throughput screen hit into a tractable tool compound for dissecting the effects of acute DCN1-UBE2M inhibition on the NEDD8/cullin pathway. Structure-enabled optimization led to a 100-fold increase in biochemical potency and modestly increased solubility and permeability as compared to our initial hit. The optimized compounds inhibit the DCN1-UBE2M protein-protein interaction in our TR-FRET binding assay and inhibit cullin neddylation in our pulse-chase NEDD8 transfer assay. The optimized compounds bind to DCN1 and selectively reduce steady-state levels of neddylated CUL1 and CUL3 in a squamous cell carcinoma cell line. Ultimately, we anticipate that these studies will identify early lead compounds for clinical development for the treatment of lung squamous cell carcinomas and other cancers.

Discovery of novel, orally bioavailable, antileishmanial compounds using phenotypic screening.

Leishmaniasis is a parasitic infection that afflicts approximately 12 million people worldwide. There are several limitations to the approved drug therapies for leishmaniasis, including moderate to severe toxicity, growing drug resistance, and the need for extended dosing. Moreover, miltefosine is currently the only orally available drug therapy for this infection. We addressed the pressing need for new therapies by pursuing a two-step phenotypic screen to discover novel, potent, and orally bioavailable antileishmanials. First, we conducted a high-throughput screen (HTS) of roughly 600,000 small molecules for growth inhibition against the promastigote form of the parasite life cycle using the nucleic acid binding dye SYBR Green I. This screen identified approximately 2,700 compounds that inhibited growth by over 65% at a single point concentration of 10 µM. We next used this 2700 compound focused library to identify compounds that were highly potent against the disease-causing intra-macrophage amastigote form and exhibited limited toxicity toward the host macrophages. This two-step screening strategy uncovered nine unique chemical scaffolds within our collection, including two previously described antileishmanials. We further profiled two of the novel compounds for in vitro absorption, distribution, metabolism, excretion, and in vivo pharmacokinetics. Both compounds proved orally bioavailable, affording plasma exposures above the half-maximal effective concentration (EC50) concentration for at least 12 hours. Both compounds were efficacious when administered orally in a murine model of cutaneous leishmaniasis. One of the two compounds exerted potent activity against trypanosomes, which are kinetoplastid parasites related to Leishmania species. Therefore, this compound could help control multiple parasitic diseases. The promising pharmacokinetic profile and significant in vivo efficacy observed from our HTS hits highlight the utility of our two-step phenotypic screening strategy and strongly suggest that medicinal chemistry optimization of these newly identified scaffolds will lead to promising candidates for an orally available anti-parasitic drug.

Hit-to-Lead Studies for the Antimalarial Tetrahydroisoquinolone Carboxanilides.

Phenotypic whole-cell screening in erythrocytic cocultures of Plasmodium falciparum identified a series of dihydroisoquinolones that possessed potent antimalarial activity against multiple resistant strains of P. falciparum in vitro and show no cytotoxicity to mammalian cells. Systematic structure-activity studies revealed relationships between potency and modifications at N-2, C-3, and C-4. Careful structure-property relationship studies, coupled with studies of metabolism, addressed the poor aqueous solubility and metabolic vulnerability, as well as potential toxicological effects, inherent in the more potent primary screening hits such as 10b. Analogues 13h and 13i, with structural modifications at each site, were shown to possess excellent antimalarial activity in vivo. The (+)-(3S,4S) enantiomer of 13i and similar analogues were identified as the more potent. On the basis of these studies, we have selected (+)-13i for further study as a preclinical candidate.

Optimization of a Novel Series of Ataxia-Telangiectasia Mutated Kinase Inhibitors as Potential Radiosensitizing Agents.

We previously reported a novel inhibitor of the ataxia-telangiectasia mutated (ATM) kinase, which is a target for novel radiosensitizing drugs. While our initial lead, compound 4, was relatively potent and nontoxic, it exhibited poor stability to oxidative metabolism and relatively poor selectivity against other kinases. The current study focused on balancing potency and selectivity with metabolic stability through structural modification to the metabolized site on the quinazoline core. We performed extensive structure-activity and structure-property relationship studies on this quinazoline ATM kinase inhibitor in order to identify structural variants with enhanced selectivity and metabolic stability. We show that, while the C-7-methoxy group is essential for potency, replacing the C-6-methoxy group considerably improves metabolic stability without affecting potency. Promising analogues 20, 27g, and 27n were selected based on in vitro pharmacology and evaluated in murine pharmacokinetic and tolerability studies. Compound 27g possessed significantly improve pharmacokinetics relative to that of 4. Compound 27g was also significantly more selective against other kinases than 4. Therefore, 27g is a good candidate for further development as a potential radiosensitizer.

(+)-SJ733, a clinical candidate for malaria that acts through ATP4 to induce rapid host-mediated clearance of Plasmodium.

Drug discovery for malaria has been transformed in the last 5 years by the discovery of many new lead compounds identified by phenotypic screening. The process of developing these compounds as drug leads and studying the cellular responses they induce is revealing new targets that regulate key processes in the Plasmodium parasites that cause malaria. We disclose herein that the clinical candidate (+)-SJ733 acts upon one of these targets, ATP4. ATP4 is thought to be a cation-transporting ATPase responsible for maintaining low intracellular Na(+) levels in the parasite. Treatment of parasitized erythrocytes with (+)-SJ733 in vitro caused a rapid perturbation of Na(+) homeostasis in the parasite. This perturbation was followed by profound physical changes in the infected cells, including increased membrane rigidity and externalization of phosphatidylserine, consistent with eryptosis (erythrocyte suicide) or senescence. These changes are proposed to underpin the rapid (+)-SJ733-induced clearance of parasites seen in vivo. Plasmodium falciparum ATPase 4 (pfatp4) mutations that confer resistance to (+)-SJ733 carry a high fitness cost. The speed with which (+)-SJ733 kills parasites and the high fitness cost associated with resistance-conferring mutations appear to slow and suppress the selection of highly drug-resistant mutants in vivo. Together, our data suggest that inhibitors of PfATP4 have highly attractive features for fast-acting antimalarials to be used in the global eradication campaign.

Pharmacokinetics and efficacy of the spleen tyrosine kinase inhibitor r406 after ocular delivery for retinoblastoma.

PURPOSE: Retinoblastoma is a childhood cancer of the retina. Clinical trials have shown that local delivery of broad spectrum chemotherapeutic agents is efficacious. Recent studies characterizing the genomic and epigenomic landscape of retinoblastoma identified spleen tyrosine kinase (SYK) as a promising candidate for targeted therapy. The purpose of this study was to conduct preclinical testing of the SYK antagonist R406 to evaluate it as a candidate for retinoblastoma treatment. METHODS: The efficacy of the SYK antagonist R406 delivered locally in a human orthotopic xenograft mouse model of retinoblastoma was tested. Intraocular exposure of R406 was determined for various routes and formulations. RESULTS: There was no evidence of efficacy for subconjunctival. R406. Maximal vitreal concentration was 10-fold lower than the minimal concentration required to kill retinoblastoma cells in vitro. Dosage of R406 subconjunctivally from emulsion or suspension formulations, direct intravitreal injection of the soluble prodrug of R406 (R788), and repeated topical administration of R406 all increased vitreal exposure, but failed to reach the exposure required for retinoblastoma cell death in culture. CONCLUSION: Taken together, these data suggest that R406 is not a viable clinical candidate for the treatment of retinoblastoma. This study highlights the importance of pharmacokinetic testing of molecular targeted retinoblastoma therapeutics.

Optimization of the electrophile of chloronitrobenzamide leads active against Trypanosoma brucei.

We previously reported the phenylchloronitrobenzamides (PCNBs), a novel class of compounds active against the species of trypanosomes that cause Human African Trypanosomiasis (HAT). Herein, we explored the potential to adjust the reactivity of the electrophilic chloronitrobenzamide core. These studies identified compound 7d that potently inhibited the growth of trypanosomes (EC50=120nM for Trypanosoma b. brucei, 18nM for Trypanosoma b. rhodesiense, and 38nM for Trypanosoma b. gambiense) without significant cytotoxicity against mammalian cell lines (EC50>25µM for HepG2, HEK293, Raji, and BJ cell lines) and also had good stability in microsomal models (t1/2>4h in both human and mouse). Overall these properties indicate the compound 7d and its analogs are worth further exploration as potential leads for HAT.

Optimization of chloronitrobenzamides (CNBs) as therapeutic leads for human African trypanosomiasis (HAT).

We previously reported the discovery of the activity of chloronitrobenzamides (CNBs) against bloodstream forms of Trypanosoma brucei . Herein we disclose extensive structure-activity relationship and structure-property relationship studies aimed at identification of tractable early leads for clinical development. These studies revealed a promising lead compound, 17b, that exhibited nanomolar potency against T. brucei (EC50 = 27 nM for T. b. brucei, 7 nM for T. b. rhodesiense, and 2 nM for T. b. gambiense ) with excellent selectivity for parasite cells relative to mammalian cell lines (EC50 > 25 µM). In addition compound 17b displayed suitable physiochemical characteristics and microsomal stability (t1/2 > 4 h for human and mouse) to justify pursuing in vivo studies.

Lead optimization of antimalarial propafenone analogues.

Previously reported studies identified analogues of propafenone that had potent antimalarial activity, reduced cardiac ion channel activity, and properties that suggested the potential for clinical development for malaria. Careful examination of the bioavailability, pharmacokinetics, toxicology, and efficacy of this series of compounds using rodent models revealed orally bioavailable compounds that are nontoxic and suppress parasitemia in vivo. Although these compounds possess potential for further preclinical development, they also carry some significant challenges.

Lead optimization of 3-carboxyl-4(1H)-quinolones to deliver orally bioavailable antimalarials.

Malaria is a protozoal parasitic disease that is widespread in tropical and subtropical regions of Africa, Asia, and the Americas and causes more than 800,000 deaths per year. The continuing emergence of multidrug-resistant Plasmodium falciparum drives the ongoing need for the development of new and effective antimalarial drugs. Our previous work has explored the preliminary structural optimization of 4(1H)-quinolone ester derivatives, a new series of antimalarials related to the endochins. Herein, we report the lead optimization of 4(1H)-quinolones with a focus on improving both antimalarial potency and bioavailability. These studies led to the development of orally efficacious antimalarials including quinolone analogue 20g, a promising candidate for further optimization.

Synthesis and evaluation of sulfonylnitrophenylthiazoles (SNPTs) as thyroid hormone receptor-coactivator interaction inhibitors.

We previously identified a series of methylsulfonylnitrobenzoates (MSNBs) that block the interaction of the thyroid hormone receptor with its coactivators. MSNBs inhibit coactivator binding through irreversible modification of cysteine 298 of the thyroid hormone receptor (TR). Although MSNBs have better pharmacological features than our first generation inhibitors (ß-aminoketones), they contain a potentially unstable ester linkage. Here we report the bioisosteric replacement of the ester linkage with a thiazole moiety, yielding sulfonylnitrophenylthiazoles (SNPTs). An array of SNPTs representing optimal side chains from the MSNB series was constructed using parallel chemistry and evaluated to test their antagonism of the TR-coactivator interaction. Selected active compounds were evaluated in secondary confirmatory assays including regulation of thyroid response element driven transcription in reporter constructs and native genes. In addition the selected SNPTs were shown to be selective for TR relative to other nuclear hormone receptors (NRs).