pH Programmed Optical Sensor Arrays for Cancer Plasma Straightforward Discrimination Based on Protein-Responsive Patterns.

Optical cross-reactive sensor arrays inspired by the mammalian olfactory system that can realize straightforward discrimination of plasma from cancer patients hold great potential for point-of-care diseases diagnostics. Herein, a pH programmed fluorescence sensor array based on protein-responsive patterns was designed for straightforward discrimination of different types of cancer plasma. It is worth noting that plasma discrimination can be realized only by programming one nanomaterial using different pH values, which greatly simplifies the programmable design of the sensor array, making it an important highlight of this work. In addition, the mechanism of the pH programmed fluorescence sensor array for protein responsiveness was systematically investigated through molecular docking simulation, fluorescence resonance energy transfer (FRET), and fluorescence lifetime experiments. Most importantly, not only can the differences between plasma from healthy people and and from patients with different cancer species including gastric cancer, liver cancer, breast cancer, and cervical cancer be discriminated by this pH programmed fluorescence sensor array, but also the blind test of unknown plasma samples can be well identified with 100% accuracy, indicating its promising prospect in clinical application.

Potency of the novel PolC DNA polymerase inhibitor CRS0540 in a disseminated Listeria monocytogenes intracellular hollow-fibre model.

BACKGROUND: Listeriosis is an orphan disease, which is nevertheless fatal in immunocompromised people. CRS0540 is a novel PolC DNA polymerase inhibitor that has demonstrated good in vitro and in vivo activity against Listeria monocytogenes. METHODS: Rodent-to-human allometry projection-based human population pharmacokinetics of CRS0540 were used for all studies. CRS0540 pharmacokinetics/pharmacodynamics studies in an intracellular hollow-fibre system model of disseminated listeriosis (HFS-Lister) examined the effect of eight treatment doses, administered daily over 7 days, in duplicate units. Total bacterial burden versus AUC/MIC exposures on each day were modelled using the inhibitory sigmoid Emax model, while CRS0540-resistant bacterial burden was modelled using a quadratic function. Ten thousand-subject Monte Carlo simulations were used to predict an optimal clinical dose for treatment. RESULTS: The mean CRS0540 intracellular/extracellular AUC0-24 ratio was 34.07 (standard error: 15.70) as measured in the HFS-Lister. CRS0540 demonstrated exposure-dependent bactericidal activity in the HFS-Lister, with the highest exposure killing approximately 5.0 log10 cfu/mL. The free drug AUC0-24/MIC associated with 80% of maximal kill (EC80) was 36.4. Resistance emergence versus AUC/MIC was described by a quadratic function, with resistance amplification at an AUC/MIC of 54.8 and resistance suppression at an AUC/MIC of 119. Monte Carlo simulations demonstrated that for the EC80 target, IV CRS0540 doses of 100 mg/kg achieved PTAs of >90% at MICs up to 1.0 mg/L. CONCLUSIONS: CRS0540 is a promising orphan drug candidate for listeriosis. Future PK/PD studies comparing it with penicillin, the standard of care, could lead to this drug as a new treatment in immunocompromised patients.

Discovery of benzothiazole amides as potent antimycobacterial agents.

From a high throughput screening of commercially available libraries against nontuberculous mycobacteria and Mycobacterium tuberculosis, numerous hits were identified with moderate activity. Extensive medicinal chemistry optimization has led to a series of potent benzothiazole amide antimycobacterial agents. Replacement of the adamantyl group with cyclohexyl derivatives and further development of this series resulted in an advanced lead compound, CRS400393, which demonstrated excellent potency and a mycobacteria-specific spectrum of activity. MIC values ranged from 0.03 to 0.12 µg/mL against Mycobacterium abscessus and other rapid-grower NTM, and 1-2 µg/mL against Mycobacterium avium complex. The preliminary mechanism of action studies suggested these agents may target MmpL3, a mycobacterial mycolic acid transporter. The series has demonstrated in vivo efficacy in a proof of concept mouse model of M. abscessus infection.

Pharmacologic inhibition of S-nitrosoglutathione reductase protects against experimental asthma in BALB/c mice through attenuation of both bronchoconstriction and inflammation.

BACKGROUND: S-nitrosoglutathione (GSNO) serves as a reservoir for nitric oxide (NO) and thus is a key homeostatic regulator of airway smooth muscle tone and inflammation. Decreased levels of GSNO in the lungs of asthmatics have been attributed to increased GSNO catabolism via GSNO reductase (GSNOR) leading to loss of GSNO- and NO- mediated bronchodilatory and anti-inflammatory actions. GSNOR inhibition with the novel small molecule, N6022, was explored as a therapeutic approach in an experimental model of asthma. METHODS: Female BALB/c mice were sensitized and subsequently challenged with ovalbumin (OVA). Efficacy was determined by measuring both airway hyper-responsiveness (AHR) upon methacholine (MCh) challenge using whole body plethysmography and pulmonary eosinophilia by quantifying the numbers of these cells in the bronchoalveolar lavage fluid (BALF). Several other potential biomarkers of GSNOR inhibition were measured including levels of nitrite, cyclic guanosine monophosphate (cGMP), and inflammatory cytokines, as well as DNA binding activity of nuclear factor kappa B (NFκB). The dose response, onset of action, and duration of action of a single intravenous dose of N6022 given from 30 min to 48 h prior to MCh challenge were determined and compared to effects in mice not sensitized to OVA. The direct effect of N6022 on airway smooth muscle tone also was assessed in isolated rat tracheal rings. RESULTS: N6022 attenuated AHR (ED50 of 0.015 ± 0.002 mg/kg; Mean ± SEM) and eosinophilia. Effects were observed from 30 min to 48 h after treatment and were comparable to those achieved with three inhaled doses of ipratropium plus albuterol used as the positive control. N6022 increased BALF nitrite and plasma cGMP, while restoring BALF and plasma inflammatory markers toward baseline values. N6022 treatment also attenuated the OVA-induced increase in NFκB activation. In rat tracheal rings, N6022 decreased contractile responses to MCh. CONCLUSIONS: The significant bronchodilatory and anti-inflammatory actions of N6022 in the airways are consistent with restoration of GSNO levels through GSNOR inhibition. GSNOR inhibition may offer a therapeutic approach for the treatment of asthma and other inflammatory lung diseases. N6022 is currently being evaluated in clinical trials for the treatment of inflammatory lung disease.

Long-term outcomes and risk factors for early bacterial infection after pediatric liver transplantation: a prospective cohort study.

BACKGROUND: Liver transplantation (LT) is the most efficient treatment for pediatric patients with end-stage liver diseases, while bacterial infection is the leading reason for posttransplant mortality. The present study is to explore the outcomes and risk factors of early bacterial infection (within 1 mo) after pediatric LT. METHODS: In this prospective cohort study, 1316 pediatric recipients (median [IQR] age: 9.1 [6.3-28.0] months; male: 48.0%; median [IQR] follow-up time: 40.6 [29.1-51.4] months) who received LT from September 2018 to April 2022 were included. Bacterial culture samples such as sputum, abdominal drainage, blood and so on were collected when recipients were presented with infective symptoms. Kaplan-Meier analysis was applied to estimate the long-term survival rates and logistic regression was used to identify independent risk factors. To explore the role of pretransplant rectal swab culture (RSC) in reducing posttransplant bacterial infection rate, 188 infant LT recipients (median [IQR] age: 6.8 [5.5-8.1] months; male: 50.5%) from May 2022 to September 2023 were included. Log-binomial regression was used to measure the association of pretransplant RSC screening and posttransplant bacterial infection. The "Expectation Maximization" algorithm was used to impute the missing data. RESULTS: Bacterial infection was the primary cause for early (38.9%) and overall mortality (35.6%) after pediatric LT. Kaplan-Meier analysis revealed inferior 1- and 5-year survival rates for recipients with posttransplant bacterial infection (92.6% vs. 97.1%, 91.8% vs. 96.4% respectively; P<0.001). Among all detected bacteria, Staphylococcus spp. (34.3%) and methicillin-resistant coagulase-negative Staphylococci (43.2%) were the dominant species and multi-drug resistant organisms, respectively. Multivariable analysis revealed that infant recipients (adjusted odds ratio [aOR], 1.49; 95% CI, 1.01-2.20), male recipients (aOR, 1.43; 95% CI, 1.08-1.89), high graft-to-recipient weight ratio (aOR, 1.64; 95% CI, 1.17-2.30), positive posttransplant RSC (aOR, 1.45; 95% CI, 1.04-2.02) and nasopharyngeal swab culture (aOR 2.46; 95% CI, 1.72-3.52) were independent risk factors for early bacterial infection. Furthermore, RSC screening and antibiotic prophylaxis before transplantation could result in a relatively lower posttransplant infection rate, albeit without statistical significance (adjusted RR, 0.53; 95% CI, 0.25-1.16). CONCLUSION: In this cohort study, posttransplant bacterial infection resulted in an inferior long-term patient survival rate. The five identified independent risk factors for posttransplant bacterial infection could guide the prophylaxis strategy of posttransplant bacterial infection in the future. Additionally, pretransplant RSC might decrease posttransplant bacterial infection rate.

Liver organoids: established tools for disease modeling and drug development.

In the past decade, liver organoids have evolved rapidly as valuable research tools, providing novel insights into almost all types of liver diseases, including monogenic liver diseases, alcohol-associated liver disease, metabolic-associated fatty liver disease, various types of (viral) hepatitis, and liver cancers. Liver organoids in part mimic the microphysiology of the human liver and fill a gap in high-fidelity liver disease models to a certain extent. They hold great promise to elucidate the pathogenic mechanism of a diversity of liver diseases and play a crucial role in drug development. Moreover, it is challenging but opportunistic to apply liver organoids for tailored therapies of various liver diseases. The establishment, applications, and challenges of different types of liver organoids, for example, derived from embryonic, adult, or induced pluripotent stem cells, to model different liver diseases, are presented in this review.

HRG inhibits liver cancer lung metastasis by suppressing neutrophil extracellular trap formation.

BACKGROUND: Distant metastasis is a sign of poor prognosis for cancer patients. Extrahepatic liver cancer metastases commonly spread to the lung. Remodelling of the metastatic microenvironment is essential for tumour metastasis. Neutrophil-associated metastatic microenvironment contributes to the early metastatic colonisation of cancer cells in the lung. METHOD: The lung metastasis models were constructed via treated cancer cells by tail vein injection into mice. And samples of lung were harvested at the indicated time to analyze tumor growth and immune cells in the microenvironment. Tumors and lung metastasis specimens were obtained via surgical operations for research purposes. Neutrophils were obtained from peripheral blood of patients with liver cancer or healthy donors (HD). RESULTS: Hepatocellular carcinoma cells reduce the secretion of histidine-rich glycoprotein (HRG), regulate the recruitment and activation of neutrophils in the metastatic microenvironment and promote the production of neutrophil extracellular traps (NETs), thereby promoting liver cancer lung metastasis. HRG binds to FCγR1 on the neutrophil membrane while inhibiting PI3K and NF-κB activation, thereby reducing IL-8 secretion to reduce neutrophil recruitment. Meanwhile, HRG inhibited IL8-MAPK and NF-κB pathway activation and ROS production, resulting in reduced NETs formation. CONCLUSIONS: Our study reveals that liver cancer regulates neutrophil recruitment and NETs formation in the metastatic microenvironment by reducing HRG secretion, thereby promoting tumour lung metastasis. The results of this study will contribute to the development of possible strategies for treating metastases.

Programmable DNA Hydrogel Assisting Microcrystal Formulations for Sustained Locoregional Drug Delivery in Surgical Residual Tumor Lesions and Lymph Node Metastasis.

Surgical residual tumor lesions (R1 resection of surgical procedures (e.g., liver cancer infiltrating the diaphragm, surgical residual breast cancer, postoperative residual ovarian cancer) or boundary residual after ablation) and lymph node metastasis that cannot be surgically resected (retroperitoneal lymph nodes) significantly affect postoperative survival of tumor patients. This clinical conundrum poses three challenges for local drug delivery systems: stable and continuous delivery, good biocompatibility, and the ability to package new targeted drugs that can synergize with other treatments. Here, a drug-laden hydrogel generated from pure DNA strands and highly programmable in adjusting its mesh size is reported. Meanwhile, the DNA hydrogel can assist the microcrystallization of novel radiosensitizing drugs, ataxia telangiectasia and rad3-related protein (ATR) inhibitor (Elimusertib), further facilitating its long-term release. When applied to the tumor site, the hydrogel system demonstrates significant antitumor activity, minimized systemic toxicity, and has a modulatory effect on the tumor-immune cell interface. This drug-loaded DNA-hydrogel platform represents a novel modality for adjuvant therapy in patients with surgical residual tumor lesions and lymph node metastasis.

Microbiological profile, preclinical pharmacokinetics and efficacy of CRS0393, a novel antimycobacterial agent targeting MmpL3.

The benzothiazole amide CRS0393 demonstrated excellent in vitro activity against nontuberculous mycobacteria (NTM), including M. abscessus isolates from cystic fibrosis (CF) patients, with minimum inhibitory concentrations (MICs) of ≤0.03-0.5 µg/mL. The essential transport protein MmpL3 was confirmed as the target via analysis of spontaneous resistant mutants and further biological profiling. In mouse pharmacokinetic studies, intratracheal instillation of a single dose of CRS0393 resulted in high concentrations of drug in epithelial lining fluid (ELF) and lung tissue, which remained above the M. abscessus MIC for at least 9 hours post-dose. This exposure resulted in a penetration ratio of 261 for ELF and 54 for lung tissue relative to plasma. CRS0393 showed good oral bioavailability, particularly when formulated in kolliphor oil, with a lung-to-plasma penetration ratio ranging from 0.5 to 4. CRS0393 demonstrated concentration-dependent reduction of intracellular M. abscessus in a THP-1 macrophage infection model. CRS0393 was well tolerated following intranasal administration (8 mg/kg) or oral dosing (25 mg/kg) once daily for 28 days in dexamethasone-treated C3HeB/FeJ mice. Efficacy against M. abscessus strain 103 was achieved via the intranasal route, while oral dosing will need further optimization. CRS0393 holds promise for development as a novel agent with broad antimycobacterial activity.

Mechanism of inhibition for N6022, a first-in-class drug targeting S-nitrosoglutathione reductase.

N6022 is a novel, first-in-class drug with potent inhibitory activity against S-nitrosoglutathione reductase (GSNOR), an enzyme important in the metabolism of S-nitrosoglutathione (GSNO) and in the maintenance of nitric oxide (NO) homeostasis. Inhibition of GSNOR by N6022 and related compounds has shown safety and efficacy in animal models of asthma, chronic obstructive pulmonary disease, and inflammatory bowel disease [Sun, X., et al. (2011) ACS Med. Chem. Lett. 2, 402-406]. N6022 is currently in early phase clinical studies in humans. We show here that N6022 is a tight-binding, specific, and fully reversible inhibitor of GSNOR with an IC(50) of 8 nM and a K(i) of 2.5 nM. We accounted for the fact that the NAD(+)- and NADH-dependent oxidation and reduction reactions, catalyzed by GSNOR are bisubstrate in nature in our calculations. N6022 binds in the GSNO substrate binding pocket like a competitive inhibitor, although in kinetic assays it behaves with a mixed uncompetitive mode of inhibition (MOI) toward the GSNO substrate and a mixed competitive MOI toward the formaldehyde adduct, S-hydroxymethylglutathione (HMGSH). N6022 is uncompetitive with cofactors NAD(+) and NADH. The potency, specificity, and MOI of related GSNOR inhibitor compounds are also reported.

Structure-activity relationship of pyrrole based S-nitrosoglutathione reductase inhibitors: carboxamide modification.

The enzyme S-nitrosoglutathione reductase (GSNOR) is a member of the alcohol dehydrogenase family (ADH) that regulates the levels of S-nitrosothiols (SNOs) through catabolism of S-nitrosoglutathione (GSNO). GSNO and SNOs are implicated in the pathogenesis of many diseases including those in respiratory, gastrointestinal, and cardiovascular systems. The pyrrole based N6022 was recently identified as a potent, selective, reversible, and efficacious GSNOR inhibitor which is currently in clinical development for acute asthma. We describe here the synthesis and structure-activity relationships (SAR) of novel pyrrole based analogs of N6022 focusing on carboxamide modifications on the pendant N-phenyl moiety. We have identified potent and novel GSNOR inhibitors that demonstrate efficacy in an ovalbumin (OVA) induced asthma model in mice.

Discovery of potent and novel S-nitrosoglutathione reductase inhibitors devoid of cytochrome P450 activities.

The pyrrole based N6022 was recently identified as a potent, selective, reversible, and efficacious S-nitrosoglutathione reductase (GSNOR) inhibitor and is currently undergoing clinical development for the treatment of acute asthma. GSNOR is a member of the alcohol dehydrogenase family (ADH) and regulates the levels of S-nitrosothiols (SNOs) through catabolism of S-nitrosoglutathione (GSNO). Reduced levels of GSNO, as well as other nitrosothiols (SNOs), have been implicated in the pathogenesis of many diseases including those of the respiratory, cardiovascular, and gastrointestinal systems. Preservation of endogenous SNOs through GSNOR inhibition presents a novel therapeutic approach with broad applicability. We describe here the synthesis and structure-activity relationships (SAR) of novel pyrrole based analogues of N6022 focusing on removal of cytochrome P450 inhibition activities. We identified potent and novel GSNOR inhibitors having reduced CYP inhibition activities and demonstrated efficacy in a mouse ovalbumin (OVA) model of asthma.

Structure-activity relationships of pyrrole based S-nitrosoglutathione reductase inhibitors: pyrrole regioisomers and propionic acid replacement.

S-Nitrosoglutathione reductase (GSNOR) is a member of the alcohol dehydrogenase family (ADH) that regulates the levels of S-nitrosothiols (SNOs) through catabolism of S-nitrosoglutathione (GSNO). GSNO and SNOs are implicated in the pathogenesis of many diseases including those in respiratory, cardiovascular, and gastrointestinal systems. The pyrrole based N6022 was recently identified as a potent, selective, reversible, and efficacious GSNOR inhibitor which is currently undergoing clinical development. We describe here the synthesis and structure-activity relationships (SAR) of novel pyrrole based analogues of N6022 focusing on scaffold modification and propionic acid replacement. We identified equally potent and novel GSNOR inhibitors having pyrrole regioisomers as scaffolds using a structure based approach.

Quinazolin-2-ylamino-quinazolin-4-ols as novel non-nucleoside inhibitors of bacterial DNA polymerase III.

High throughput screening led to the discovery of a novel series of quinazolin-2-ylamino-quinazolin-4-ols as a new class of DNA polymerase III inhibitors. The inhibition of chromosomal DNA replication results in bacterial cell death. The synthesis, structure-activity relationships and functional activity are described.

Optimization and Lead Selection of Benzothiazole Amide Analogs Toward a Novel Antimycobacterial Agent.

Mycobacteria remain an important problem worldwide, especially drug resistant human pathogens. Novel therapeutics are urgently needed to tackle both drug-resistant tuberculosis (TB) and difficult-to-treat infections with nontuberculous mycobacteria (NTM). Benzothiazole adamantyl amide had previously emerged as a high throughput screening hit against M. tuberculosis (Mtb) and was subsequently found to be active against NTM as well. For lead optimization, we applied an iterative process of design, synthesis and screening of several 100 analogs to improve antibacterial potency as well as physicochemical and pharmacological properties to ultimately achieve efficacy. Replacement of the adamantyl group with cyclohexyl derivatives, including bicyclic moieties, resulted in advanced lead compounds that showed excellent potency and a mycobacteria-specific spectrum of activity. MIC values ranged from 0.03 to 0.12 µg/mL against M. abscessus (Mabs) and other rapid- growing NTM, 1-2 µg/mL against M. avium complex (MAC), and 0.12-0.5 µg/mL against Mtb. No pre-existing resistance was found in a collection of n = 54 clinical isolates of rapid-growing NTM. Unlike many antibacterial agents commonly used to treat mycobacterial infections, benzothiazole amides demonstrated bactericidal effects against both Mtb and Mabs. Metabolic labeling provided evidence that the compounds affect the transfer of mycolic acids to their cell envelope acceptors in mycobacteria. Mapping of resistance mutations pointed to the trehalose monomycolate transporter (MmpL3) as the most likely target. In vivo efficacy and tolerability of a benzothiazole amide was demonstrated in a mouse model of chronic NTM lung infection with Mabs. Once daily dosing over 4 weeks by intrapulmonary microspray administration as 5% corn oil/saline emulsion achieved statistically significant CFU reductions compared to vehicle control and non-inferiority compared to azithromycin. The benzothiazole amides hold promise for development of a novel therapeutic agent with broad antimycobacterial activity, though further work is needed to develop drug formulations for direct intrapulmonary delivery via aerosol.

Discovery of s-nitrosoglutathione reductase inhibitors: potential agents for the treatment of asthma and other inflammatory diseases.

S-Nitrosoglutathione reductase (GSNOR) regulates S-nitrosothiols (SNOs) and nitric oxide (NO) in vivo through catabolism of S-nitrosoglutathione (GSNO). GSNOR and the anti-inflammatory and smooth muscle relaxant activities of SNOs, GSNO, and NO play significant roles in pulmonary, cardiovascular, and gastrointestinal function. In GSNOR knockout mice, basal airway tone is reduced and the response to challenge with bronchoconstrictors or airway allergens is attenuated. Consequently, GSNOR has emerged as an attractive therapeutic target for several clinically important human diseases. As such, small molecule inhibitors of GSNOR were developed. These GSNOR inhibitors were potent, selective, and efficacious in animal models of inflammatory disease characterized by reduced levels of GSNO and bioavailable NO. N6022, a potent and reversible GSNOR inhibitor, reduced bronchoconstriction and pulmonary inflammation in a mouse model of asthma and demonstrated an acceptable safety profile. N6022 is currently in clinical development as a potential agent for the treatment of acute asthma.

New agents for Clostridium difficile-associated disease.

BACKGROUND: Clostridia-derived diseases, in particular C. difficile-associated disease (CDAD), have been increasing in incidence, severity, and morbidity. The mainstay of treatment options has relied upon metronidazole and vancomycin, but these treatments routinely result in high relapse rates (20%) and, in the case of metronidazole, decreasing efficacy. OBJECTIVE: Evaluate and compare the current clinical and preclinical therapies of CDAD. RESULTS/CONCLUSION: The new antibiotics in development and preclinical development reflect next-generation versions of older drugs or two new mechanism-of-action class drugs (OPT-80, REP3123). Based on the current preclinical and clinical data, the next-generation drugs impart only a subtle difference from the intrinsic weaknesses of their genre. In contrast, OPT-80 and REP3123 seem to be differentiated.

Aminoacyl-tRNA synthetases: essential and still promising targets for new anti-infective agents.

The emergence of resistance to existing antibiotics demands the development of novel antimicrobial agents directed against novel targets. Historically, bacterial cell wall synthesis, protein, and DNA and RNA synthesis have been major targets of very successful classes of antibiotics such as beta-lactams, glycopeptides, macrolides, aminoglycosides, tetracyclines, rifampicins and quinolones. Recently, efforts have been made to develop novel agents against validated targets in these pathways but also against new, previously unexploited targets. The era of genomics has provided insights into novel targets in microbial pathogens. Among the less exploited--but still promising--targets is the family of 20 aminoacyl-tRNA synthetases (aaRSs), which are essential for protein synthesis. These targets have been validated in nature as aaRS inhibition has been shown as the specific mode of action for many natural antimicrobial agents synthesized by bacteria and fungi. Therefore, aaRSs have the potential to be targeted by novel agents either from synthetic or natural sources to yield specific and selective anti-infectives. Numerous high-throughput screening programs aimed at identifying aaRS inhibitors have been performed over the last 20 years. A large number of promising lead compounds have been identified but only a few agents have moved forward into clinical development. This review provides an update on the present strategies to develop novel aaRS inhibitors as anti-infective drugs.