N-Ethyl-N-Nitrosourea Induced Leukaemia in a Mouse Model: Protective Effect of Icaritin via Inhibition of IL-6/JAK2/STAT3 Pathway Causes Apoptosis.

Background: The present study aimed to investigate the protective effect of icaritin (ICT) on ENU-induced leukemia in male mice. Methods: The mice received intraperitoneal injections of 80 mg/kg ENU twice a week for three months for induction of leukemia. Blood smears from these mice showed blast cells, confirming the presence of leukemia. After confirming leukemia, mice were divided into control, ENU-induced leukemia, and leukemia groups (10 mg/kg bw and 20 mg/kg bw) were treated with ICT for 35 days. Blood, spleen, and liver samples were collected for analysis. The expression of IL-6, JAK2, STAT3, as well as inflammatory, pro-apoptotic (Bax), and anti-apoptotic (Bcl-2) proteins were evaluated using qPCR, immunohistochemistry, and immunofluorescence techniques. Results: The study found that ICT inhibited inflammation and the IL-6/JAK2/STAT3 pathway in ENU-induced mice. ICT treatment induced apoptosis in the spleen and liver by activating Bax and downregulating Bcl-2. The findings provide novel evidence that ICT acts as a dual inhibitor of IL-6/JAK2/STAT3 signaling, promoting apoptosis and playing an essential role in anti-leukemic activity. Conclusion: These results suggest that ICT has potential as a therapeutic target for treating leukemia, offering a novel approach to leukemia treatment through inhibiting the IL-6/JAK2/STAT3 pathway and induction of apoptosis.

Probing the chemical 'reactome' with high-throughput experimentation data.

High-throughput experimentation (HTE) has the potential to improve our understanding of organic chemistry by systematically interrogating reactivity across diverse chemical spaces. Notable bottlenecks include few publicly available large-scale datasets and the need for facile interpretation of these data's hidden chemical insights. Here we report the development of a high-throughput experimentation analyser, a robust and statistically rigorous framework, which is applicable to any HTE dataset regardless of size, scope or target reaction outcome, which yields interpretable correlations between starting material(s), reagents and outcomes. We improve the HTE data landscape with the disclosure of 39,000+ previously proprietary HTE reactions that cover a breadth of chemistry, including cross-coupling reactions and chiral salt resolutions. The high-throughput experimentation analyser was validated on cross-coupling and hydrogenation datasets, showcasing the elucidation of statistically significant hidden relationships between reaction components and outcomes, as well as highlighting areas of dataset bias and the specific reaction spaces that necessitate further investigation.

Attenuation of neurovirulence of chikungunya virus by a single amino acid mutation in viral E2 envelope protein.

BACKGROUND: Chikungunya virus (CHIKV) has reemerged as a major public health concern, causing chikungunya fever with increasing cases and neurological complications. METHODS: In the present study, we investigated a low-passage human isolate of the East/ Central/South African (ECSA) lineage of CHIKV strain LK(EH)CH6708, which exhibited a mix of small and large viral plaques. The small and large plaque variants were isolated and designated as CHIKV-SP and CHIKV-BP, respectively. CHIKV-SP and CHIKV-BP were characterized in vitro and in vivo to compare their virus production and virulence. Additionally, whole viral genome analysis and reverse genetics were employed to identify genomic virulence factors. RESULTS: CHIKV-SP demonstrated lower virus production in mammalian cells and attenuated virulence in a murine model. On the other hand, CHIKV-BP induced higher pro-inflammatory cytokine levels, compromised the integrity of the blood-brain barrier, and led to astrocyte infection in mouse brains. Furthermore, the CHIKV-SP variant had limited transmission potential in Aedes albopictus mosquitoes, likely due to restricted dissemination. Whole viral genome analysis revealed multiple genetic mutations in the CHIKV-SP variant, including a Glycine (G) to Arginine (R) mutation at position 55 in the viral E2 glycoprotein. Reverse genetics experiments confirmed that the E2-G55R mutation alone was sufficient to reduce virus production in vitro and virulence in mice. CONCLUSIONS: These findings highlight the attenuating effects of the E2-G55R mutation on CHIKV pathogenicity and neurovirulence and emphasize the importance of monitoring this mutation in natural infections.

Development and Comprehensive Benchmark of a High-Quality AMBER-Consistent Small Molecule Force Field with Broad Chemical Space Coverage for Molecular Modeling and Free Energy Calculation.

Biomolecular simulations have become an essential tool in contemporary drug discovery, and molecular mechanics force fields (FFs) constitute its cornerstone. Developing a high quality and broad coverage general FF is a significant undertaking that requires substantial expert knowledge and computing resources, which is beyond the scope of general practitioners. Existing FFs originate from only a limited number of groups and organizations, and they either suffer from limited numbers of training sets, lower than desired quality because of oversimplified representations, or are costly for the molecular modeling community to access. To address these issues, in this work, we developed an AMBER-consistent small molecule FF with extensive chemical space coverage, and we provide Open Access parameters for the entire modeling community. To validate our FF, we carried out benchmarks of quantum mechanics (QM)/molecular mechanics conformer comparison and free energy perturbation calculations on several benchmark data sets. Our FF achieves a higher level of performance at reproducing QM energies and geometries than two popular open-source FFs, OpenFF2 and GAFF2. In relative binding free energy calculations for 31 protein-ligand data sets, comprising 1079 pairs of ligands, the new FF achieves an overall root-mean-square error of 1.19 kcal/mol for ΔΔG and 0.92 kcal/mol for ΔG on a subset of 463 ligands without bespoke fitting to the data sets. The results are on par with those of the leading commercial series of OPLS FFs.

Different mechanisms of X-ray irradiation-induced male and female sterility in Aedes aegypti.

BACKGROUND: Aedes aegypti (Ae. aegypti) is the major vector that transmits many diseases including dengue, Zika, and filariasis in tropical and subtropical regions. Due to the growing resistance to chemical-based insecticides, biological control methods have become an emerging direction to control mosquito populations. The sterile insect technique (SIT) deploys high doses of ionizing radiation to sterilize male mosquitoes before the release. The Wolbachia-based population suppression method of the incompatible insect technique (IIT) involves the release of Wolbachia-infected males to sterilize uninfected field females. Due to the lack of perfect sex separation tools, a low percentage of female contamination is detected in the male population. To prevent the unintentional release of these Wolbachia-infected females which might result in population replacement, a low dose of X-ray irradiation is deployed to sterilize any female escapees. However, it remains unclear whether these irradiation-induced male and female sterilizations share common mechanisms. RESULTS: In this work, we set out to define the minimum dose of X-ray radiation required for complete female sterilization in Ae. aegypti (NEA-EHI strain). Further results showed that this minimum dose of X-ray irradiation for female sterilization significantly reduced male fertility. Similar results have been reported previously in several operational trials. By addressing the underlying causes of the sterility, our results showed that male sterility is likely due to chromosomal damage in the germ cells induced by irradiation. In contrast, female sterility appears to differ and is likely initiated by the elimination of the somatic supporting cells, which results in the blockage of the ovariole maturation. Building upon these findings, we identified the minimum dose of X-ray irradiation on the Wolbachia-infected NEA-EHI (wAlbB-SG) strain, which is currently being used in the IIT-SIT field trial. Compared to the uninfected parental strain, a lower irradiation dose could fully sterilize wAlbB-SG females. This suggests that Wolbachia-carrying mosquitoes are more sensitive to irradiation, consistent with a previous report showing that a lower irradiation dose fully sterilized Wolbachia-infected Ae. aegypti females (Brazil and Mexican strains) compared to those uninfected controls. CONCLUSIONS: Our findings thus reveal the distinct mechanisms of ionizing X-ray irradiation-induced male or female sterility in Ae. aegypti mosquitoes, which may help the design of X-ray irradiation-based vector control methods.

Harnessing Preclinical Data as a Predictive Tool for Human Brain Tissue Targeting.

One of the objectives within the medicinal chemistry discipline is to design tissue targeting molecules. The objective of tissue specificity can be either to gain drug access to the compartment of interest (e.g., the CNS) for Neuroscience targets or to restrict drug access to the CNS for all other therapeutic areas. Both neuroscience and non-neuroscience therapeutic areas have struggled to quantitatively estimate brain penetration or the lack thereof with compounds that are substrates of efflux transport proteins such as P-glycoprotein (P-gp) and breast cancer resistant protein (BCRP) that are key components of the blood-brain barrier (BBB). It has been well established that drug candidates with high efflux ratios (ER) of these transporters have poor penetration into brain tissue. In the current work, we outline a parallel analysis to previously published models for the prediction of brain penetration that utilize an alternate MDR1-MDCK cell line as a better predictor of brain penetration and whether a correlation between in vitro, rodent data, non-human primate (NHP), and human in vivo brain penetration data could be established. Analysis of structural and physicochemical properties in conjunction with in vitro parameters and preclinical in vivo data has been highlighted in this manuscript as a continuation of the previously published work.

Current and Future Roles of Artificial Intelligence in Medicinal Chemistry Synthesis.

Artificial intelligence and machine learning have demonstrated their potential role in predictive chemistry and synthetic planning of small molecules; there are at least a few reports of companies employing in silico synthetic planning into their overall approach to accessing target molecules. A data-driven synthesis planning program is one component being developed and evaluated by the Machine Learning for Pharmaceutical Discovery and Synthesis (MLPDS) consortium, comprising MIT and 13 chemical and pharmaceutical company members. Together, we wrote this perspective to share how we think predictive models can be integrated into medicinal chemistry synthesis workflows, how they are currently used within MLPDS member companies, and the outlook for this field.

Correction to: Description of Wenzhouxiangella salilacus sp. nov., a moderate halophilic bacterium isolated from a salt lake in Xinjiang Province, China.

In the original publication of the article, the deposit accession numbers of strain 15181T in the acknowledgment section were incorrectly provided as "KCTC 62172T and MCCC 1K03442T".

Optimal designs for pairwise calculation: An application to free energy perturbation in minimizing prediction variability.

Pairwise-based methods such as the free energy perturbation (FEP) method have been widely deployed to compute the binding free energy differences between two similar host-guest complexes. The calculated pairwise free energy difference is either directly adopted or transformed to absolute binding free energy for molecule rank ordering. We investigated, through both analytic derivations and simulations, how the selection of pairs in the experiment could impact the overall prediction precision. Our studies showed that (1) the estimated absolute binding free energy ( ΔG^ ) derived from calculated pairwise differences (ΔΔG) through weighted least squares fitting is more precise in prediction than the pairwise difference values when the number of pairs is more than the number of ligands and (2) prediction precision is influenced by both the total number of pairs and the specifically selected pairs, the latter being critically important when the number of calculated pairs is limited. Furthermore, we applied optimal experimental design in pair selection and found that the optimally selected pairs can outperform randomly selected pairs in prediction precision. In an illustrative example, we showed that, upon weighing ligand structure similarity into design optimization, the weighted optimal designs are more efficient than the literature reported designs. This work provides a new approach to assess retrospective pairwise-based prediction results, and a method to design new prospective pairwise-based experiments for molecular lead optimization. © 2019 Wiley Periodicals, Inc.

Molecular Transformer unifies reaction prediction and retrosynthesis across pharma chemical space.

Predicting how a complex molecule reacts with different reagents, and how to synthesise complex molecules from simpler starting materials, are fundamental to organic chemistry. We show that an attention-based machine translation model - Molecular Transformer - tackles both reaction prediction and retrosynthesis by learning from the same dataset. Reagents, reactants and products are represented as SMILES text strings. For reaction prediction, the model "translates" the SMILES of reactants and reagents to product SMILES, and the converse for retrosynthesis. Moreover, a model trained on publicly available data is able to make accurate predictions on proprietary molecules extracted from pharma electronic lab notebooks, demonstrating generalisability across chemical space. We expect our versatile framework to be broadly applicable to problems such as reaction condition prediction, reagent prediction and yield prediction.

Author Correction: Dopamine D3 receptor antagonist reveals a cryptic pocket in aminergic GPCRs.

A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has not been fixed in the paper.

Alkalilacustris brevis gen. nov., sp. nov., isolated from a soda lake.

A Gram-stain-negative, aerobic, non-pigmented and short-rod-shaped bacterium, designated 34079T, was isolated from a water sample of a soda lake in Jilin, a province of China. Strain 34079T grew at 10-50 °C (optimum, 35 °C), pH 7-10 (optimum, pH 8.0-8.5). NaCl was required for growth at the concentration range 1-10.0 % (w/v), with an optimum at 2.5-4 % (w/v). Chemotaxonomic analysis indicated that the sole respiratory quinone was Q-10. The predominant cellular fatty acids (>5 %) were summed feature 8 (C18 : 1 ω7c/C18 : 1 ω6c) and C16 : 0. The major polar lipids consisted of diphosphatidylglycerol, phosphatidylglycerol, phosphatidylcholine, three unidentified amino lipids, one unidentified amino phosphoglycolipid, one phosphoglycolipid, one unidentified glycolipid, three unidentified phospholipids and two unidentified lipids. The DNA G+C content was 65.6 mol%. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain 34079T formed a distinct lineage in the clade of the family 'Rhodobacteraceae' with the highest sequence similarity of 96.1 % to Pararhodobacter aggregans, followed by Rhodobaca bogoriensis DSM 18756T (95.7 %) and Roseibaca ekhonensis DSM 11469T (94.7 %). The distinct biochemical, chemotaxonomic and phylogenetic differences from the previously described taxa supported that strain 34079T represents a novel species of a new genus, for which the name Alkalilacustris brevis gen. nov., sp. nov. is proposed. The type strain is 34079T (=KCTC 62428T=MCCC 1K03493T).

Ligand biological activity predicted by cleaning positive and negative chemical correlations.

Predicting ligand biological activity is a key challenge in drug discovery. Ligand-based statistical approaches are often hampered by noise due to undersampling: The number of molecules known to be active or inactive is vastly less than the number of possible chemical features that might determine binding. We derive a statistical framework inspired by random matrix theory and combine the framework with high-quality negative data to discover important chemical differences between active and inactive molecules by disentangling undersampling noise. Our model outperforms standard benchmarks when tested against a set of challenging retrospective tests. We prospectively apply our model to the human muscarinic acetylcholine receptor M1, finding four experimentally confirmed agonists that are chemically dissimilar to all known ligands. The hit rate of our model is significantly higher than the state of the art. Our model can be interpreted and visualized to offer chemical insights about the molecular motifs that are synergistic or antagonistic to M1 agonism, which we have prospectively experimentally verified.

Validation of Human MDR1-MDCK and BCRP-MDCK Cell Lines to Improve the Prediction of Brain Penetration.

It is of great challenge to predict human brain penetration for substrates of multidrug resistance protein 1 (MDR1) and breast cancer resistance protein (BCRP), 2 major efflux transporters at blood-brain barrier. Thus, a physiologically based pharmacokinetic (PBPK) model with the incorporation of in vitro MDR1 and BCRP transporter function data and transporter protein expression levels has been developed. As such, it is crucial to generate MDR1 and BCRP substrate data with a high fidelity. In this study, 2 widely used human MDR1 cell lines from Borst and National Institutes of Health laboratories were evaluated using rodent brain penetration data, and the study suggested that the MDR1 expressed in Madin-Darby canine kidney (MDCK) cell line from National Institutes of Health laboratory predicted brain penetration better, particularly for compounds with a high passive permeability. In addition, human BCRP-MDCK cell line with 1 µM PSC833, a specific MDR1 inhibitor, demonstrated the ability to identify BCRP substrates without the confounding of endogenous canine Mdr1. Comparison of human BCRP and mouse Bcrp transporter functions revealed that the functional differences of BCRP between the 2 species is minimal. The incorporation of both the validated MDR1 and BCRP assays into our brain PBPK model has significantly improved the prediction for the brain penetration of MDR1 and BCRP substrates across species.

Description of Wenzhouxiangella salilacus sp. nov., a moderate halophilic bacterium isolated from a salt lake in Xinjiang Province, China.

A Gram-stain negative, non-motile, strictly aerobic and rod-shaped bacterium, designated as 15181T, was isolated from a salt lake in Xinjiang Province, China. Strain 15181T was able to grow at 10-40 °C (optimum 37 °C), pH 6.0-8.5 (optimum 7.0) and with 1-14% NaCl (optimum 4%, w/v). According to phylogenetic analysis based on 16S rRNA gene sequences, strain 15181T was assigned to the genus Wenzhouxiangella with high 16S rRNA gene sequence similarity of 97.4% to Wenzhouxiangella sediminis XDB06T, followed by Wenzhouxiangella marina KCTC 42284T (95.9%). Strain 15181T exhibited ANI values of 80.0% and 72.0% to W. sediminis XDB06T and W. marina KCTC 42284T, respectively. The in silico DDH analysis revealed that strain 15181T shared 19.1% and 18.7% DNA relatedness with W. sediminis XDB06T and W. marina KCTC 42284T, respectively. Chemotaxonomic analysis showed that the sole respiratory quinone was ubiquinone-8, the major fatty acids included iso-C15:0, iso-C16:0 and summed feature 9 (C16:0 10-methyl and/or iso-C17:1ω9c). The major polar lipids included diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine, two unidentified glycolipids, two unidentified phospholipids, two unidentified aminophospholipids and an unidentified lipid. On the basis of phenotypic, genotypic and chemotaxonomic characteristics presented in this study, strain 15181T is concluded to represent a novel species in the genus Wenzhouxiangella, for which the name Wenzhouxiangella salilacus sp. nov. is proposed. The type strain is 15181T (=KCTC 62172T=MCCC 1K03442T).

Confluentibacter flavum sp. nov., Isolated from the Saline Lake.

A Gram-stain-negative, rod-shaped, non-motile, bacterial isolate designated 3BT, was isolated from a saline lake, and subjected to a polyphasic taxonomic investigation. The phylogenetic analysis based on 16S rRNA gene sequence clearly showed an allocation to the genus Confluentibacter with similarity ranging from 95.1 to 98%. OrthoANI values between strain 3BT and related strains of Confluentibacter (< 90%) were lower than the threshold value of 95% ANI relatedness recommended for species demarcation. Strain 3BT grew at 4-35 °C and pH 6.0-8.0 (optimum, 28 °C and pH 6.5) and with 0-3% (w/v) NaCl (optimum, 0.5%). The predominant respiratory quinone was menaquinone-6 (MK-6) and the major fatty acids were iso-C15:0, iso-C15:1 G, iso-C15:0 3-OH, and iso-C17:0 3-OH. The polar lipid profile of strain 3BT comprised phosphatidylethanolamine, one unidentified aminolipid, one aminophospholipid, and three unidentified lipids (L1-3). The DNA G+C content was 33.1 mol%. On the basis of morphological, physiological, and chemotaxonomic characteristics, together with the results of phylogenetic analysis, strain 3BT is described as a novel species in genus Confluentibacter, for which the name Confluentibacter flavum sp. nov. (type strain 3BT = CGMCC115960T = KCTC52969T) is proposed.

Rhodohalobacter barkolensis sp. nov., isolated from a saline lake and emended description of the genus Rhodohalobacter.

A Gram-stain-negative, non-motile, aerobic, rod-shaped bacterium, designated 15182T, was isolated from a saline lake in China. The novel strain 15182T was able to grow at 10-40 °C (optimum, 37 °C), pH 7.0-8.0 (optimum, 7.5) and with 0.5-4 % NaCl (optimum, 2-3 %, w/v). The phylogenetic analysis based on 16S rRNA gene sequences revealed that strain 15182T was most closely related to the genus Rhodohalobacter by sharing the highest sequence similarity of 97.0 % with Rhodohalobacter halophilus JZ3C29T. Chemotaxonomic analysis showed that the sole respiratory quinone was menaquinone 7, the major fatty acids included C16 : 0 N alcohol and C16 : 1ω11c. The major polar lipids included diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine, four uncharacterized glycolipids, one uncharacterized phospholipid and two uncharacterized lipids. The genomic DNA G+C content of the strain 15182T was 42.4 mol%. The average nucleotide identity value between 15182T and R. halophilus JZ3C29T was 75.4 %, and the in silico DNA-DNA hybridization value of the two strains was 19.1 %. On the basis of its phenotypic, chemotaxonomic, genotypic and genomic characteristics presented in this study, strain 15182T is suggested to represent a novel species in the genus Rhodohalobacter, for which the name Rhodohalobacter barkolensis sp. nov. is proposed. The type strain is 15182T (=KCTC 62172T=MCCC 1K03442T). An emended description of the genus Rhodohalobacter is also presented.

Kordiimonas pumila sp. nov., isolated from coastal sediment.

A novel Gram-stain-negative, translucent-white, aerobic, motile and rod-shaped strain, designated N18T, was isolated from a coastal sediment sample collected in Zhoushan, Zhejiang Province, China. 16S rRNA gene similarity analysis revealed that strain N18T demonstrated highest similarity to the genus Kordiimonas(95.3-97.2 %). Phylogenetic analysis of 16S rRNA gene sequence showed that strain N18T represented a distinct lineage in the clade consisting of the genus Kordiimonas. Strain N18T was found to grow at 10-37 °C (optimum 28 °C), pH 6.0-8.0 (optimum 7.0) and with 1.0-4.0 % (w/v) NaCl (optimum 2.5 %). The G+C content of the genomic DNA was 55.3 mol%. The major cellular fatty acids were identified as summed feature 3 (comprising iso-C15 : 0 2-OH/C16 : 1ω7c), iso-C17 : 1ω9c and iso-C15 : 0. The polar lipid profile of N18T consisted of phosphatidylglycerol, phosphatidylethanolamine, diphosphatidylglycerol, an unidentified glycolipid, an unidentified aminoglycolipid, an unidentified aminophospholipid and five unidentified lipids. The respiratory quinone was Q-10. Based on chemotaxonomic, morphological and physiological properties, strain N18T could be distinguished from its closest phylogenetic neighbours. Thus, we propose Kordiimonas pumila sp. nov., the type strain is N18T (=MCCC 1K03436T=KCTC 62164T).

Design and Synthesis of Clinical Candidate PF-06751979: A Potent, Brain Penetrant, ß-Site Amyloid Precursor Protein Cleaving Enzyme 1 (BACE1) Inhibitor Lacking Hypopigmentation.

A major challenge in the development of ß-site amyloid precursor protein cleaving enzyme 1 (BACE1) inhibitors for the treatment of Alzheimer's disease is the alignment of potency, drug-like properties, and selectivity over related aspartyl proteases such as Cathepsin D (CatD) and BACE2. The potential liabilities of inhibiting BACE2 chronically have only recently begun to emerge as BACE2 impacts the processing of the premelanosome protein (PMEL17) and disrupts melanosome morphology resulting in a depigmentation phenotype. Herein, we describe the identification of clinical candidate PF-06751979 (64), which displays excellent brain penetration, potent in vivo efficacy, and broad selectivity over related aspartyl proteases including BACE2. Chronic dosing of 64 for up to 9 months in dog did not reveal any observation of hair coat color (pigmentation) changes and suggests a key differentiator over current BACE1 inhibitors that are nonselective against BACE2 in later stage clinical development.

Dopamine D3 receptor antagonist reveals a cryptic pocket in aminergic GPCRs.

The recent increase in the number of X-ray crystal structures of G-protein coupled receptors (GPCRs) has been enabling for structure-based drug design (SBDD) efforts. These structures have revealed that GPCRs are highly dynamic macromolecules whose function is dependent on their intrinsic flexibility. Unfortunately, the use of static structures to understand ligand binding can potentially be misleading, especially in systems with an inherently high degree of conformational flexibility. Here, we show that docking a set of dopamine D3 receptor compounds into the existing eticlopride-bound dopamine D3 receptor (D3R) X-ray crystal structure resulted in poses that were not consistent with results obtained from site-directed mutagenesis experiments. We overcame the limitations of static docking by using large-scale high-throughput molecular dynamics (MD) simulations and Markov state models (MSMs) to determine an alternative pose consistent with the mutation data. The new pose maintains critical interactions observed in the D3R/eticlopride X-ray crystal structure and suggests that a cryptic pocket forms due to the shift of a highly conserved residue, F6.52. Our study highlights the importance of GPCR dynamics to understand ligand binding and provides new opportunities for drug discovery.