Differentiation of ROMK potency from hERG potency in the phenacetyl piperazine series through heterocycle incorporation.

Following the discovery of small molecule acyl piperazine ROMK inhibitors and their initial preclinical validation as a novel diuretic agent, our group set out to discover new ROMK inhibitors with reduced risk for QT effects, suitable for further pharmacological experiments in additional species. Several strategies for decreasing hERG affinity while maintaining ROMK inhibition were investigated and are described herein. The most promising candidate, derived from the newly discovered 4-N-heteroaryl acetyl series, improved functional hERG/ROMK ratio by >10× over the previous lead. In vivo evaluation demonstrated comparable diuretic effects in rat with no detectable QT effects at the doses evaluated in an in vivo dog model.

Discovery of a potent and selective ROMK inhibitor with improved pharmacokinetic properties based on an octahydropyrazino[2,1-c][1,4]oxazine scaffold.

Following the discovery of small molecule acyl piperazine ROMK inhibitors, the acyl octahydropyrazino[2,1-c][1,4]oxazine series was identified. This series displays improved ROMK/hERG selectivity, and as a consequence, the resulting ROMK inhibitors do not evoke QTc prolongation in an in vivo cardiovascular dog model. Further efforts in this series led to the discovery of analogs with improved pharmacokinetic profiles. This new series also retained comparable ROMK potency compared to earlier leads.

Design, synthesis, and biological evaluation of aminopyrazine derivatives as inhibitors of mitogen-activated protein kinase-activated protein kinase 2 (MK-2).

Several series of novel non-thiourea-containing aminopyrazine derivatives were designed based on the MK-2 inhibitors 1-(2-aminopyrazin-3-yl)methyl-2-thioureas. These compounds were synthesized and evaluated for their inhibitory activity against MK-2 enzyme in vitro. Compounds with low micromolar to sub-micromolar IC50 values were identified, and several compounds were also found to be active in suppressing the lipopolysaccharide (LPS)-stimulated TNFα production in THP-1 cells with minimum shift compared to their enzyme activity.

Soil Water Movement and Groundwater Recharge Under Different Land Uses in a Flood-Irrigated Area.

The water shortage in agriculture area in China requires to reduce the consumption of excessive water in flood irrigation. Therefore, the dynamics of soil water regime is needed to investigate and water-saving irrigation is necessary to alleviate water shortage. This study investigated the impact of flood irrigation on soil water movement and recharge to groundwater in the Yellow River irrigation area of Yinchuan Plain, China. Combining comprehensive field observation, stable isotopic techniques and water balance simulation, we described the soil water mechanism in vadose zone covered with bare soil in 2019 and planted with maize in 2020. The soil layers affected by precipitation infiltration and evaporation were mainly 0-50 cm, while the soil influenced by irrigation was the entire profile in the mode of piston flow. The maize root took up the soil water up to the depth of 100 cm during the tasseling period. The infiltration and capillary rise in 2020 were similar with those in 2019. However, the total deep percolation was 156.5 mm in 2020 which was about 50% of that in 2019 because of the maize root water uptake. The leakage of ditch water was the major recharge resource of groundwater for the fast water table rise. Precise irrigation is required to minimize deep percolation and leakage of ditch water and reduce excessive unproductive evapotranspiration. Therefore, understanding the soil water movement and groundwater recharge is critical for agricultural water management to improve irrigation efficiency and water use efficiency in arid regions.

Mtb-Selective 5-Aminomethyl Oxazolidinone Prodrugs: Robust Potency and Potential Liabilities.

Linezolid is a drug with proven human antitubercular activity whose use is limited to highly drug-resistant patients because of its toxicity. This toxicity is related to its mechanism of action─linezolid inhibits protein synthesis in both bacteria and eukaryotic mitochondria. A highly selective and potent series of oxazolidinones, bearing a 5-aminomethyl moiety (in place of the typical 5-acetamidomethyl moiety of linezolid), was identified. Linezolid-resistant mutants were cross-resistant to these molecules but not vice versa. Resistance to the 5-aminomethyl molecules mapped to an N-acetyl transferase (Rv0133) and these mutants remained fully linezolid susceptible. Purified Rv0133 was shown to catalyze the transformation of the 5-aminomethyl oxazolidinones to their corresponding N-acetylated metabolites, and this transformation was also observed in live cells of Mycobacterium tuberculosis. Mammalian mitochondria, which lack an appropriate N-acetyltransferase to activate these prodrugs, were not susceptible to inhibition with the 5-aminomethyl analogues. Several compounds that were more potent than linezolid were taken into C3HeB/FeJ mice and were shown to be highly efficacious, and one of these (9) was additionally taken into marmosets and found to be highly active. Penetration of these 5-aminomethyl oxazolidinone prodrugs into caseum was excellent. Unfortunately, these compounds were rapidly converted into the corresponding 5-alcohols by mammalian metabolism which retained antimycobacterial activity but resulted in substantial mitotoxicity.

Discovery of a novel sub-class of ROMK channel inhibitors typified by 5-(2-(4-(2-(4-(1H-Tetrazol-1-yl)phenyl)acetyl)piperazin-1-yl)ethyl)isobenzofuran-1(3H)-one.

A sub-class of distinct small molecule ROMK inhibitors were developed from the original lead 1. Medicinal chemistry endeavors led to novel ROMK inhibitors with good ROMK functional potency and improved hERG selectivity. Two of the described ROMK inhibitors were characterized for the first in vivo proof-of-concept biology studies, and results from an acute rat diuresis model confirmed the hypothesis that ROMK inhibitors represent new mechanism diuretic and natriuretic agents.

[Spatial-temporal Variation in Water Quality and Its Response to Precipitation and Land Use in Baiyangdian Lake in the Early Stage of the Construction of Xiong'an New Area].

To assess the water quality of Baiyangdian Lake in the early stage of construction of the Xiong'an New Area, the water quality data in Baiyangdian Lake from December 2020 to November 2021 were collected from five typical monitoring stations. Using the remote-sensing interpretation of land use types, Pearson correlation analysis, comprehensive pollution index, and principal component analysis, the spatial-temporal variation characteristics of the water quality and its response to precipitation and land use structure at different spatial scales were explored in Baiyangdian Lake. The results showed that:① the water quality of Baiyangdian Lake was the best in spring and the worst in summer. Nutrient pollution was the heaviest at Nanliuzhuang, which was greatly affected by the Fuhe River. Organic pollution was the heaviest at Caiputai. ② The comprehensive pollution index of Nanliuzhuang was the highest, followed by that of Shaochedian, Guangdianzhangzhuang, Caiputai, and Quantou. The main excessive pollutant was TN, and the pollution index of 1.55 reached the level of severe pollution. The water quality of Baiyangdian Lake was significantly better than that before the construction of the Xiong'an New Area. ③ The construction land and cropland within the 1.5 km buffer zone had the greatest impact on turbidity, NH4+-N, and TP. While the reed wetland acted as the "sink" for intercepting and absorbing pollutants, it also released nitrogen into the 800 m buffer zone, which was the "source" of the pollutants. ④ The correlation between water quality indicators and land use structure in the rainy season was lower than that in the dry season. This was due to the dilution effect of the upstream reservoir discharging in the rainy season, which weakened the impact of pollutants in the rainfall runoff on the water quality of Baiyangdian Lake.

Identifying surface water and groundwater interactions using multiple experimental methods in the riparian zone of the polluted and disturbed Shaying River, China.

Identifying groundwater (GW)-surface water (SW) interactions in riparian zones is important for assessing the transport pathways of pollutants and all potential biochemical processes, particularly in rivers with artificially controlled water levels. In this study, we constructed two monitoring transects along the nitrogen-polluted Shaying River, China. The GW-SW interactions were qualitatively and quantitatively characterized through an intensive 2-y monitoring program. The monitoring indices included water level, hydrochemical parameters, isotopes (δ18O, δD, and 222Rn) and microbial community structures. The results showed that the sluice altered the GW-SW interactions in the riparian zone. A decrease in river level occurs during the flood season owing to sluice regulation, resulting in discharge of riparian GW into the river. The water level, hydrochemistry, isotopes, and microbial community structures in near-river wells were similar to those in the river, indicating mixing of the river water with the riparian GW. As the distance from the river increased, the percentage of river water in the riparian GW decreased, whereas the GW residence time increased. We found that nitrogen may be easily transported through the GW-SW interactions, acting as a sluice regulator. Nitrogen stored in river water may be removed or diluted by mixing GW and rainwater during the flood season. As the residence time of the infiltrated river in the riparian aquifer increased, nitrate removal increased. Identifying the GW-SW interactions is crucial for water resource regulation and for further tracing the transport of contaminants such as nitrogen in the historically polluted Shaying River.

[Occurrence Characteristics of Microplastics in Baiyangdian Lake Water and Sediments].

In order to explore the occurrence characteristics of microplastics in the freshwater environment of Baiyangdian Lake in China, ten overlying water samples and ten sediment samples were collected in Baiyangdian Lake of Hebei Province in October 2021, and the abundance distribution, shape, particle size, and polymer type of microplastics in the samples were identified using laboratory pretreatment, microscope observation, and laser infrared spectroscopy. The sedimentation law of microplastics at the overlying water-sediment interface was studied using the Stokes sedimentation formula, and their pollution characteristics and potential sources were analyzed. The abundances of microplastics in the overlying water and sediments in Baiyangdian Lake ranged from 474-19382 n·m-3 and 95.3-29542.5 n·kg-1, respectively, with an average value of 6255.4 n·m-3 and 11088 n·kg-1. The main polymer of the microplastics in the overlying water was polyethylene terephthalate[PET, (17.20±0.26)%], and the microplastics in the sediments were mainly chlorinated polyethylene[CPE, (46.11±1.30)%]. The sedimentation velocities of microplastics in the sedimentation zone ranged from 0.0793-111.7547 mm·s-1. The particles with larger particle size had higher sedimentation velocity and easily settled and remained in the sediments. The main sources of microplastic pollution in the study area were the discharge of textile fibers from washing wastewater and the wear and tear of ship paint, ship rubber, and building materials.

Sources and hydrogeochemical processes of groundwater under multiple water source recharge condition.

Ecological water replenishment (EWR) is an essential approach for improving the quantity and quality of regional water. The Chaobai River is a major river in Beijing that is replenished with water from multiple sources, including reclaimed water (RW), the South-North Water Transfer Project (SNTP), reservoir discharge (RD). The effects of multiple water source recharge (MWSR) on groundwater quality remain unclear. In this study, hydrochemical ions, isotopes (δ2H-H2O, δ18O-H2O, δ15N-NO3-, and δ18O-NO3-), mixing stable isotope analysis in R (MixSIAR), and hydrogeochemical modeling were used to quantify the contributions and impacts of different water sources on groundwater and to propose a conceptual model. The results showed that during the period before reservoir discharge, RW and SNTP accounted for 38 %-41 % and 54 % of the groundwater in their corresponding recharge areas, respectively. The groundwater in the RW recharge area contained high levels of Na+ and Cl- leading to the precipitation of halite, and was the main factor for the spatial variation in groundwater hydrochemical components. The surface water changed from Na·K - Cl·SO4 type to Ca·Mg - HCO3 type which was similar to groundwater after reservoir discharge. RD accounted for 30 % of the groundwater; however, it did not change the hydrochemical type of groundwater. Dual nitrate stable isotopes and MixSIAR demonstrated that RW was the primary source of NO3- in groundwater, contributing up to 76-89 %, and reservoir discharge effectively reduced the contribution of RW. δ15N-NO3- or δ18O-NO3- in relation to NO3-N suggests that denitrification is the main biogeochemical process of nitrogen in groundwater, whereas water recharge from the SNTP and RD reduces denitrification and dilutes NO3-. This study provides insights into the impact of anthropogenically controlled ecological water replenishment from different water sources on groundwater and guides the reasonable allocation of water resources.

3-Substituted 3-(4-aryloxyaryl)-propanoic acids as GPR40 agonists.

The design, synthesis, and structure-activity relationship (SAR) for a series of ß-substituted 3-(4-aryloxyaryl)propanoic acid GPR40 agonists is described. Systematic replacement of the pendant aryloxy group led to identification of potent GPR40 agonists. In order to identify candidates suitable for in vivo validation of the target, serum shifted potency and pharmacokinetic properties were determined for several compounds. Finally, further profiling of compound 7 is presented, including demonstration of enhanced glucose tolerance in an in vivo mouse model.

Design and synthesis of a new class of malonyl-CoA decarboxylase inhibitors with anti-obesity and anti-diabetic activities.

A new series of thiazole-substituted 1,1,1,3,3,3-hexafluoro-2-propanols were prepared and evaluated as malonyl-CoA decarboxylase (MCD) inhibitors. Key analogs caused dose-dependent decreases in food intake and body weight in obese mice. Acute treatment with these compounds also led to a drop in elevated blood glucose in a murine model of type II diabetes.

Discovery of 5-aryloxy-2,4-thiazolidinediones as potent GPR40 agonists.

Systematic structure-activity relationship (SAR) studies of a screening lead led to the discovery of a series of thiazolidinediones (TZDs) as potent GPR40 agonists. Among them, compound C demonstrated an acute mechanism-based glucose-lowering in an intraperitoneal glucose tolerance test (IPGTT) in lean mice, while no effects were observed in GPR40 knock-out mice.

Dual targeting of CCR2 and CX3CR1 in an arterial injury model of vascular inflammation.

OBJECTIVES: The chemokine receptors CCR2 and CX3CR1 are important in the development of coronary artery disease. The purpose of this study is to analyze the effect of a novel CCR2 inhibitor in conjunction with CX3CR1 deletion on vascular inflammation. METHODS: The novel CCR2 antagonist MRL-677 was characterized using an in vivo model of monocyte migration. To determine the relative roles of CCR2 and CX3CR1 in vascular remodeling, normal or CX3CR1 deficient mice were treated with MRL-677. After 14 days, the level of intimal hyperplasia in the artery was visualized by paraffin sectioning and histology of the hind limbs. RESULTS: MRL-677 is a CCR2 antagonist that is effective in blocking macrophage trafficking in a peritoneal thioglycollate model. Intimal hyperplasia resulting from vascular injury was also assessed in mice. Based on the whole-blood potency of MRL-677, sufficient drug levels were maintained for the entire 14 day experimental period to afford good coverage of mCCR2 with MRL-677. Blocking CCR2 with MRL-677 resulted in a 56% decrease in the vascular injury response (n = 9, p < 0.05) in normal animals. Mice in which both CCR2 and CX3CR1 pathways were targeted (CX3CR1 KO mice given MRL-677) had an 88% decrease in the injury response (n = 6, p = 0.009). CONCLUSION: In this study we have shown that blocking CCR2 with a low molecular weight antagonist ameliorates the inflammatory response to vascular injury. The protective effect of CCR2 blockade is increased in the presence of CX3CR1 deficiency suggesting that CX3CR1 and CCR2 have non-redundant functions in the progression of vascular inflammation.

Discovery and optimization of novel 4-[(aminocarbonyl)amino]-N-[4-(2-aminoethyl)phenyl]benzenesulfonamide ghrelin receptor antagonists.

This Letter describes optimization of ghrelin receptor antagonists and inverse agonists starting from a screening hit.

Identification of novel and orally active spiroindoline NPY Y5 receptor antagonists.

A series of spiroindoline-3,4'-piperidine derivatives were synthesized and evaluated for their binding affinities and antagonistic activities at Y5 receptors. Potent Y5 antagonists were tested for their oral bioavailabilities and brain penetration in rats. Some of the antagonists showed good oral bioavailability and/or good brain penetration. In particular, compound 6e was orally bioavailable and brain penetrant, and oral administration of 6e inhibited bPP-induced food intake in rats with a minimum effective dose of 10mg/kg.

Design, synthesis, and structure-activity relationship of novel CCR2 antagonists.

A series of novel 1-aminocyclopentyl-3-carboxyamides incorporating substituted tetrahydropyran moieties have been synthesized and subsequently evaluated for their antagonistic activity against the human CCR2 receptor. Among them analog 59 was found to posses potent antagonistic activity.

Novel 1-(2-aminopyrazin-3-yl)methyl-2-thioureas as potent inhibitors of mitogen-activated protein kinase-activated protein kinase 2 (MK-2).

Novel 1-(2-aminopyrazin-3-yl)methyl-2-thioureas are described as inhibitors of mitogen-activated protein kinase-activated protein kinase 2 (MK-2). These compounds demonstrate potent in vitro activity against the enzyme with IC(50) values as low as 15 nM, and suppress expression of TNFalpha in THP-1 cells and in vivo in an acute inflammation model in mice. The synthesis, structure-activity relationship (SAR), and biological evaluation of these compounds are discussed.

[Effects of Astragalus heteropolysaccharides on erythrocyte immune adherence function of mice with adjuvant-induced arthritis].

Astragalus heteropolysaccharides (AHPS) is obtained from the dried roots of Astragalus membranaceus (Fisch.) Bunge var. mongholious (Bunge) Hsiao. In the present study, we observed its effects on erythrocyte immune adherence function in mice with adjuvant-induced arthritis (AA). The mice were treated intragastrically with AHPS of 1 000, 500, and 250 mg x kg(-1) x d(-1) separately and treated with tripterygium glycosides (TG) of 60 mg x kg(-1) x d(-1) as positive control. The number of complement receptor type 1 (CR1) on erythrocyte, the concentration of circulating immune complex (CIC) in serum and the amount of immune complex (IC) deposition in synovium of knee joint were determined by flow cytometry, polyethylene glycol (PEG-6000) precipitation and ponceau S (P-S) staining and fluorescent immunohistochemistry respectively. The pathological change of knee joint was evaluated by histological section. The results showed that both AHPS and TG improved significantly the primary and secondary local or systemic symptoms of the mice with AA and reduced the synovium hyperplasia, inflammatory cell infiltrate, pannus and cartilage demolish of knee joint, and AHPS of 1 000, 500, and 250 mg x kg(-1) x d(-1) could significantly increase the number of CR1 on erythrocyte, improve the elimination of CIC in the peripheral blood and reduce the deposition of IC in joint synovium in a dose-dependent manner (P < 0.01 or P < 0.05). The results indicate that one of the therapeutic effective mechanisms of AHPS on mice with AA could be to increase gene expression of CR1 of mice with AA.

A fluorescence-based thiol quantification assay for ultra-high-throughput screening for inhibitors of coenzyme A production.

Here we report the development and miniaturization of a cell-free enzyme assay for ultra-high-throughput screening (uHTS) for inhibitors of two potential drug targets for obesity and cancer: fatty acid synthase (FAS) and acetyl-coenzyme A (CoA) carboxylase (ACC) 2. This assay detects CoA, a product of the FAS-catalyzed condensation of malonyl-CoA and acetyl-CoA. The free thiol of CoA can react with 7-diethylamino-3-(4'-maleimidylphenyl)-4-methylcoumarin (CPM), a profluorescent coumarin maleimide derivative that becomes fluorescent upon reaction with thiols. FAS produces long-chain fatty acid and CoA from the condensation of malonyl-CoA and acetyl-CoA. In our FAS assay, CoA released in the FAS reaction forms a fluorescence adduct with CPM that emits at 530 nm when excited at 405 nm. Using this detection method for CoA, we measured the activity of sequential enzymes in the fatty acid synthesis pathway to develop an ACC2/FAS-coupled assay where ACC2 produces malonyl-CoA from acetyl-CoA. We miniaturized the FAS and ACC2/FAS assays to 3,456- and 1,536-well plate format, respectively, and completed uHTSs for small molecule inhibitors of this enzyme system. This report shows the results of assay development, miniaturization, and inhibitor screening for these potential drug targets.