A new anti-CRISPR gene promotes the spread of drug-resistance plasmids in Klebsiella pneumoniae.

The Klebsiella pneumoniae (K. pneumoniae, Kp) populations carrying both resistance-encoding and virulence-encoding mobile genetic elements (MGEs) significantly threaten global health. In this study, we identified a new anti-CRISPR gene (acrIE10) on a conjugative plasmid with self-target sequence in K. pneumoniae with type I-E* CRISPR-Cas system. AcrIE10 interacts with the Cas7* subunit of K. pneumoniae I-E* CRISPR-Cas system. The crystal structure of the AcrIE10-KpCas7* complex suggests that AcrIE10 suppresses the I-E* CRISPR-Cas by binding directly to Cas7 to prevent its hexamerization, thereby preventing the surveillance complex assembly and crRNA loading. Bioinformatic and functional analyses revealed that AcrIE10 is functionally widespread across diverse species. Our study reports a novel anti-CRISPR and highlights its potential role in spreading resistance and virulence among pathogens.

Structural basis for enzymatic photocatalysis in chlorophyll biosynthesis.

The enzyme protochlorophyllide oxidoreductase (POR) catalyses a light-dependent step in chlorophyll biosynthesis that is essential to photosynthesis and, ultimately, all life on Earth1-3. POR, which is one of three known light-dependent enzymes4,5, catalyses reduction of the photosensitizer and substrate protochlorophyllide to form the pigment chlorophyllide. Despite its biological importance, the structural basis for POR photocatalysis has remained unknown. Here we report crystal structures of cyanobacterial PORs from Thermosynechococcus elongatus and Synechocystis sp. in their free forms, and in complex with the nicotinamide coenzyme. Our structural models and simulations of the ternary protochlorophyllide-NADPH-POR complex identify multiple interactions in the POR active site that are important for protochlorophyllide binding, photosensitization and photochemical conversion to chlorophyllide. We demonstrate the importance of active-site architecture and protochlorophyllide structure in driving POR photochemistry in experiments using POR variants and protochlorophyllide analogues. These studies reveal how the POR active site facilitates light-driven reduction of protochlorophyllide by localized hydride transfer from NADPH and long-range proton transfer along structurally defined proton-transfer pathways.

The efficacy and safety of short-course radiotherapy followed by sequential chemotherapy and Cadonilimab for locally advanced rectal cancer: a protocol of a phase II study.

BACKGROUND: For patients with locally advanced rectal cancer (LARC), total neoadjuvant therapy (TNT), namely, intensifying preoperative treatment through the integration of radiotherapy and systemic chemotherapy before surgery, was commonly recommended as the standard treatment. However, the risk of distant metastasis at 3 years remained higher than 20%, and the complete response (CR) rate was less than 30%. Several clinical trials had suggested a higher complete response rate when combining single-agent immunotherapy with short-course radiotherapy (SCRT). The CheckMate 142 study had shown encouraging outcomes of dual immunotherapy and seemingly comparable toxicity for CRC compared with single-agent immunotherapy in historical results. Therefore, dual immunotherapy might be more feasible in conjunction with the TNT paradigm of SCRT. We performed a phase II study to investigate whether the addition of a dual immune checkpoint inhibitor bispecific antibody, Cadonilimab, to SCRT combined with chemotherapy might further increase the clinical benefit and prognosis for LARC patients. METHODS: This single-arm, multicenter, prospective, phase II study included patients with pathologically confirmed cT3-T4N0 or cT2-4N + rectal adenocarcinoma with an ECOG performance score of 0 or 1. Bispecific antibody immunotherapy was added to SCRT combined with chemotherapy. Patients enrolled would be treated with SCRT (25 Gy in five fractions over 1 week) for the pelvic cavity, followed by 4 cycles of CAPOX or 6 cycles of mFOLFOX and Cadonilimab. The primary endpoint was the CR rate, which was the ratio of the pathological CR rate plus the clinical CR rate. The secondary endpoints included local-regional control, distant metastasis, disease-free survival, overall survival, toxicity profile, quality of life and functional outcome of the rectum. To detect an increase in the complete remission rate from 21.8% to 40% with 80% power, 50 patients were needed. DISCUSSION: This study would provide evidence on the efficacy and safety of SCRT plus bispecific antibody immunotherapy combined with chemotherapy as neoadjuvant therapy for patients with LARC, which might be used as a candidate potential therapy in the future. TRIAL REGISTRATION: This phase II trial was prospectively registered at ClinicalTrials.gov, under the identifier NCT05794750.

RNA N6-methyladenosine demethylase FTO inhibits glucocorticoid-induced osteoblast differentiation and function in bone marrow mesenchymal stem cells.

Excess glucocorticoids (GCs) have been reported as key factors that impair osteoblast (OB) differentiation and function. However, the role of RNA N6-methyladenosine (m6 A) in this process has not yet been elucidated. In this study, we report that both the mRNA and protein expression of fat mass and obesity-associated gene (FTO), a key m6 A demethylase, were dose-dependently downregulated during OB differentiation by dexamethasone (DEX) in bone marrow mesenchymal stem cells (BMSCs), and FTO was gradually increased during OB differentiation. Meanwhile, FTO knockdown suppressed OB differentiation and mineralization, whereas overexpression of wide-type FTO, but not mutant FTO (mutated m6 A demethylase active site), reversed DEX-induced osteogenesis impairment. Interfering with FTO inhibited proliferation and the expression of Ki67 and Pcna in BMSCs during OB differentiation, whereas forced expression of wide-type FTO improved DEX-induced inhibition of BMSCs proliferation. Moreover, FTO knockdown reduced the mRNA stability of the OB marker genes Alpl and Col1a1, and FTO-modulated OB differentiation via YTHDF1 and YTHDF2. In conclusion, our results suggest that FTO inhibits the GCs-induced OB differentiation and function of BMSCs.

Correlation between AI-based CT organ features and normal lung dose in adjuvant radiotherapy following breast-conserving surgery: a multicenter prospective study.

BACKGROUND: Radiation pneumonitis (RP) is one of the common side effects after adjuvant radiotherapy in breast cancer. Irradiation dose to normal lung was related to RP. We aimed to propose an organ features based on deep learning (DL) model and to evaluate the correlation between normal lung dose and organ features. METHODS: Patients with pathology-confirmed invasive breast cancer treated with adjuvant radiotherapy following breast-conserving surgery in four centers were included. From 2019 to 2020, a total of 230 patients from four nationwide centers in China were screened, of whom 208 were enrolled for DL modeling, and 22 patients from another three centers formed the external testing cohort. The subset of the internal testing cohort (n = 42) formed the internal correlation testing cohort for correlation analysis. The outline of the ipsilateral breast was marked with a lead wire before the scanning. Then, a DL model based on the High-Resolution Net was developed to detect the lead wire marker in each slice of the CT images automatically, and an in-house model was applied to segment the ipsilateral lung region. The mean and standard deviation of the distance error, the average precision, and average recall were used to measure the performance of the lead wire marker detection model. Based on these DL model results, we proposed an organ feature, and the Pearson correlation coefficient was calculated between the proposed organ feature and ipsilateral lung volume receiving 20 Gray (Gy) or more (V20). RESULTS: For the lead wire marker detection model, the mean and standard deviation of the distance error, AP (5 mm) and AR (5 mm) reached 3.415 ± 4.529, 0.860, 0.883, and 4.189 ± 8.390, 0.848, 0.830 in the internal testing cohort and external testing cohort, respectively. The proposed organ feature calculated from the detected marker correlated with ipsilateral lung V20 (Pearson correlation coefficient, 0.542 with p < 0.001 in the internal correlation testing cohort and 0.554 with p = 0.008 in the external testing cohort). CONCLUSIONS: The proposed artificial Intelligence-based CT organ feature was correlated with normal lung dose in adjuvant radiotherapy following breast-conserving surgery in patients with invasive breast cancer. TRIAL REGISTRATION: NCT05609058 (08/11/2022).

Rapid preparation of high efficiency hydrogen evolution catalyst with hydrophilicity.

The electrolytic water method is an outstanding hydrogen production process because of its high stability and no restriction. A low-priced and efficient catalyst for electro-deposition of Ni-Co microspheres and nanoclusters on carbon steel (Ni-Co/CS) has been prepared by the dynamic hydrogen bubble template. In the 6 M KOH solution, Ni-Co/CS only requires an overpotential of 48 mV to provide a current density of 50 mA cm-2. At the same time, it also has a large electrochemically active specific surface area (ECSA) and a hydrophilic surface. In addition, the study about the influence of carbon steel (CS) on Ni-Co coatings and the comparison experiment for different base materials has been completed. The results prove that CS is an excellent base material for hydrogen production. It can help the Ni-Co catalyst to have a stable electrolysis in 6 M KOH for 500 h. The above properties of Ni-Co/CS catalyst make it a new choice of hydrogen production by electrolysis of water in practical applications.

Design, synthesis and mechanism research of novel algicide based on bioactive fragments synthesis strategy.

The frequency and intensity of harmful cyanobacterial blooms (HCBs) are increasing all over the world, their prevention and control have become a great challenge. In this paper, a series of 1,3,4-thiadiazole thioacetamides (T series) were designed and synthesized as potential algaecides. Among them, the compound T3 showed its best algacidal activity against Synechocystis sp. PCC 6803 (PCC 6803, EC50 = 1.51 µM) and Microcystis aeruginosa FACHB 905 (FACHB905, EC50 = 4.88 µM), which was more effective than the lead compound L1 (PCC6803, EC50 = 7.7 µM; FACHB905, EC50 = 8.8 µM) and the commercially available herbicide prometryn (PCC6803, EC50 = 4.64 µM；FACHB905, EC50 = 6.52 µM). Meanwhile, T3 showed a lower inhibitory activity (EC50 = 12.76 µM) than prometryn (EC50 = 7.98 µM) to Chlorella FACHB1227, indicating that T3 had selective inhibition to prokaryotic algae (PCC6803, FACHB905) and eukaryotic algae (FACHB1227). Furthermore, the algacidal and anti-algae activities of T3 were significantly better than those of prometryn, while the toxicity of zebrafish and human cells was less than prometryn. Electron microscope, physiological, biochemical and metabonomic analysis showed that T3 interfered with light absorption and light conversion during photosynthesis by significantly reducing chlorophyll content, thus inhibited metabolic pathways such as the Calvin cycle and TCA cycle, and eventually led to the cell rupture of cyanobacteria. These results afforded further development of effective and safe algaecides.

Circular RNA circKIF2A Contributes to the Progression of Neuroblastoma Through Regulating PRPS1 Expression by Sponging miR-377-3p.

Neuroblastoma is a malignant tumor originating from the primitive neural crest. Circular RNA (circRNA) Kinesin Superfamily Protein 2A (circKIF2A, also known as hsa_circ_0129276) has been reported to be upregulated in neuroblastoma. However, the molecular mechanism of circKIF2A participated in neuroblastoma is poorly defined. We analyzed the expression levels of circKIF2A, microRNA-377-3p (miR-377-3p), and phosphoribosyl pyrophosphate synthetase 1 (PRPS1) in neuroblastoma tissues and cell lines (SK-N-AS and LAN-6) and explored their roles. The expression levels of CircKIF2A and PRPS1 were increased and that of miR-377-3p were decreased in 21 neuroblastoma tissues and cells. Functionally, the silencing of circKIF2A inhibited cell proliferation, migration, invasion, and glycolysis, boosted apoptosis in neuroblastoma cells in vitro, and blocked the growth of subcutaneously transplanted tumors in nude mice. Mechanically, circKIF2A could work as a sponge of miR-377-3p to enhance PRPS1 expression. CircKIF2A knockdown impedes cell proliferation, metastasis, and glycolysis partly by regulating the miR-377-3p/PRPS1 axis, suggesting that targeting circKIF2A can be a feasible therapeutic strategy for neuroblastoma.

Design, synthesis, high algicidal potency, and putative mode of action of new 2-cyclopropyl-4-aminopyrimidine hydrazones.

Control of cyanobacteria harmful algal blooms remains a global challenge. In the present study, a series of novel 2-cyclopropyl-4-aminopyrimidine hydrazones were designed and synthesized as potential algicides. Compounds 4a, 4b, 4h, 4j, 4k, 4l, and 4m showed potent inhibition against Synechocystis sp. PCC6803 (median effective concentration, EC50 = 1.1 to 1.7 µM) and Microcystis aeruginosa FACHB905 (EC50 = 1.2 to 2.0 µM), more potent than, or comparably with, copper sulfate (PCC6803, EC50 = 1.8 µM; FACHB905, EC50 = 2.2 µM) and prometryne (PCC6803, EC50 = 12.3 µM; FACHB905, EC50 = 7.2 µM). Compound 4k exhibited algicidal activity in an expanded culture system, and was less toxic than copper sulfate to zebrafish. Electron microscope analyses showed that 4k damaged cyanobacterial cells and decreased the number of thylakoid lamellae. Transcriptomic and qPCR analyses suggest that 4k interfered photosynthesis-related pathways. Treatment with 4k significantly decreased the maximum quantum yield of photosystem II and the photosynthetic electron transfer rate, and the resulting reactive oxygen species damaged thylakoid membranes and photosystem I. The results suggest that 4k is a potential lead for further development of effective and safe algicides.

Strepyrazinone, a tricyclic diketopiperazine derivative with cytotoxicity from a marine-derived actinobacterium.

Strepyrazinone (1), a tricyclic diketopiperazine derivative with a carbon skeleton unprecedented in natural products, was isolated from the marine-derived Streptomyces sp. B223. Its structure was elucidated by spectroscopic analyses and electronic circular dichroism calculation. Compound 1 showed cytotoxic activity against HCT-116 cancer cell lines with an IC50 value of 0.34 µM.

Simultaneous Microcystin Degradation and Microcystis aeruginosa Inhibition with the Single Enzyme Microcystinase A.

Harmful Microcystis blooms (HMBs) seriously threaten the ecology of environments and human health. Microcystins (MCs) produced by Microcystis are powerful mediators of HMB induction and maintenance. In this study, microcystinase A (MlrA), an enzyme with MC-degrading ability, was successfully obtained at over 90% purity for the first time through overexpression in Escherichia coli K12 TB1. The obtained MlrA exhibited high stability at high temperature and under alkaline conditions, while also exhibiting a long half-life. MlrA selectively inhibited MC-producing Microcystis cultures, but had no effect on MC-nonproducing Synechocystis cultures. The inhibition mechanism of MlrA against Microcystis was investigated by evaluating the morphological and physiological characteristics of cultures. MlrA effectively degraded extracellular MCs and decreased the synthesis of intracellular MCs by causing downregulation of genes involved in the microcystin biosynthesis pathway. Concomitantly, MlrA inhibited Microcystis photosynthesis by causing the downregulated expression of important photosynthesis pathway genes and interrupting electron transport chain activities and pigment synthesis. Thus, MlrA achieved the inhibition of Microcystis growth by reducing its photosynthetic capacity and intracellular MC contents, while also degrading extracellular MCs. On the basis of these results, we propose a new paradigm to achieve the simultaneous removal of MCs and HMBs using the single enzyme characterized here.

Characterization of PPIB interaction in the P3H1 ternary complex and implications for its pathological mutations.

The P3H1/CRTAP/PPIB complex is essential for prolyl 3-hydroxylation and folding of procollagens in the endoplasmic reticulum (ER). Deficiency in any component of this ternary complex is associated with the misfolding of collagen and the onset of osteogenesis imperfecta. However, little structure information is available about how this ternary complex is assembled and retained in the ER. Here, we assessed the role of the KDEL sequence of P3H1 and probed the spatial interactions of PPIB in the complex. We show that the KDEL sequence is essential for retaining the P3H1 complex in the ER. Its removal resulted in co-secretion of P3H1 and CRTAP out of the cell, which was mediated by the binding of P3H1 N-terminal domain with CRTAP. The secreted P3H1/CRTAP can readily bind PPIB with their C-termini close to PPIB in the ternary complex. Cysteine modification, crosslinking, and mass spectrometry experiments identified PPIB surface residues involved in the complex formation, and showed that the surface of PPIB is extensively covered by the binding of P3H1 and CRTAP. Most importantly, we demonstrated that one disease-associated pathological PPIB mutation on the binding interface did not affect the PPIB prolyl-isomerase activity, but disrupted the formation of P3H1/CRTAP/PPIB ternary complex. This suggests that defects in the integrity of the P3H1 ternary complex are associated with pathological collagen misfolding. Taken together, these results provide novel structural information on how PPIB interacts with other components of the P3H1 complex and indicate that the integrity of P3H1 complex is required for proper collagen formation.

Design, synthesis and algicides activities of thiourea derivatives as the novel scaffold aldolase inhibitors.

By using a new Fragment-Based Virtual Screen strategy, two series of novel FBA-II inhibitors (thiourea derivatives) were de novo discovered based on the active site of fructose-1, 6-bisphosphate aldolase from Cyanobacterial (CyFBA). In comparison, most of the N-(2-benzoylhydrazine-1-carbonothioyl) benzamide derivatives (L14∼L22) exhibit higher CyFBA-II inhibitory activities compared to N-(phenylcarbamothioyl) benzamide derivatives (L1∼L13). Especially, compound L14 not only shows higher CyFBA-II activity (Ki = 0.65 µM), but also exhibits most potent in vivo activity against Synechocystis sp. PCC 6803 (EC50 = 0.09 ppm), higher (7-fold) than that of our previous inhibitor (EC50 = 0.6 ppm). The binding modes of compound L14 and CyFBA-II were further elucidated by jointly using DOX computational protocol, MM-PBSA and site-directed mutagenesis assays. The positive results suggest that strategy adopted in this study was promising to rapidly discovery the potent inhibitors with novel scaffolds. The satisfactory algicide activities suggest that the thiourea derivatives is very likely to be a promising lead for the development of novel specific algicides to solve Cyanobacterial harmful algal blooms (CHABs).

Structure optimization and bioactivity evaluation of ThDP analogs targeting cyanobacterial pyruvate dehydrogenase E1.

Harmful cyanobacteria bloom (HCB) has occurred frequently in recent years and it is urgent to develop novel algicides to deal with this problem. In this paper, a series of novel thiamin diphosphate (ThDP) analogs 5a-5g were designed and synthesized targeting cyanobacterial pyruvate dehydrogenase complex E1 (Cy-PDHc E1). Our results showed that compounds 5a-5g have higher inhibitory activities against Cy-PDHc E1 (IC50 9.56-3.48 µM) and higher inhibitory activities against two model cyanobacteria strains Synechocystis sp PCC6803 (EC50 2.03-1.58 µM) and Microcystis aeruginosa FACHB905 (EC50 1.86-0.95 µM). Especially, compound 5b displayed highest inhibitory activities (IC50 = 3.48 µM) against Cy-PDHc E1 and powerful inhibitory activities against cyanobacteria Synechocystis sp PCC6803 (EC50 = 1.58 µM) and Microcystis aeruginosa FACHB905 (EC50 = 1.04 µM). Moreover, the inhibitory activities of compound 5b were even higher than those of copper sulfate (EC50 = 2.02 and 1.71 µM separately) which has been widely used as algicide against cyanobacteria PCC6803 and FACHB905. The more important was that compound 5b display much higher inhibitory selectivity between Cy-PDHc E1 (Inhibitory rate 97.4%) and porcine PDHc E1 (Inhibitory rate 11.8%) under the same concentration (100 µM). The inhibition kinetic experiment and molecular docking research showed that compound 5b can inhibit Cy-PDHc E1 by occupying the ThDP-binding pocket and then blocking Cy-PDHc E1 bound to ThDP as competitive inhibitor. The imagines of SEM and TEM showed that cellular microstructures were heavily destroyed under compound 5b stress. Our results demonstrated compound 5b could be taken as a potential lead compound targeting Cy-PDHc E1 to obtain environment-friendly algicide for harmful cyanobacterial blooms control.

Design, synthesis and biological evaluation of novel inhibitors against cyanobacterial pyruvate dehydrogenase multienzyme complex E1.

Cyanobacterial pyruvate dehydrogenase multienzyme complex E1 (PDHc E1) is a potential target enzyme for finding inhibitors to control harmful cyanobacterial blooms. In this study, a series of novel triazole thiamin diphosphate (ThDP) analogs were designed and synthesized by modifying the substituent group of triazole ring and optimizing triazole-benzene linker as potential cyanobacterial PDHc E1 (Cy-PDHc E1) inhibitors. Their inhibitory activities against Cy-PDHc E1 in vitro and algicide activities in vivo were further examined. Most of these compounds exhibited prominent inhibitory activities against Cy-PDHc E1 (IC50 1.48-4.48 µM) and good algicide activities against Synechocystis PCC6803 (EC50 0.84-2.44 µM) and Microcystis aeruginosa FACHB905 (EC50 0.74-1.77 µM). Especially, compound 8d showed not only the highest inhibitory activity against Cy-PDHc E1 (IC50 1.48 µM), but also the most powerful inhibitory selectivity between Cy-PDHc E1 (inhibitory rate 98.90%) and porcine PDHc E1 (inhibitory rate only 9.54%). Furthermore, the potential interaction between compound 8d and Cy-PDHc E1 was analyzed by a molecular docking method and site-directed mutagenesis and enzymatic analysis and fluorescence spectral analysis. These results indicated that compound 8d could be used as a hit compound for further optimization and might have potential to be developed as a new algicide.

Evaluation of Mesenteric Lymph Nodes in a Pediatric Population with Mesenteric Lymphadenitis Using Superb Microvascular Imaging.

BACKGROUND This study aimed to evaluate superb microvascular imaging (SMI) as an adjunctive imaging method to evaluate mesenteric lymph nodes in children with mesenteric lymphadenitis compared with healthy children. MATERIAL AND METHODS A retrospective study compared children with mesenteric lymphadenitis (n=27) and healthy children (n=30). Lymph node size was determined using grayscale ultrasonography and parameters of lymph node vascularity were compared using color Doppler flow imaging (CDFI) and SMI. The diagnostic performance of ultrasound (US), US combined with SMI, and US combined with CDFI were compared. RESULTS Lymph nodes from children with mesenteric lymphadenitis (n=77) and normal lymph nodes (n=84) were evaluated by SMI, which showed that the least diameter of lymph nodes in cases of mesenteric lymphadenitis was 0.58±0.15 mm and of normal mesenteric lymph nodes was 0.47±0.08 mm (p<0.001). SMI identified 92.6% of abnormal mesenteric lymph nodes while CDFI detected 85.2%. US combined with SMI had the highest sensitivity (81.5%), and specificity (78.9%) compared with US alone (sensitivity, 63.0%; specificity, 64.9%), and compared with US combined with CDFI (sensitivity, 74.1%; specificity, 75.4%). US combined with SMI and US combined with CDFI achieved the same specificity (76.7%), which was higher than that of US alone (66.7%). CONCLUSIONS SMI was superior to color Doppler flow imaging in evaluating the microvasculature in lymphadenopathy in mesenteric lymphadenitis. SMI may be used as an adjunct to grayscale ultrasonography to assist in identifying mesenteric lymphadenopathy in pediatric patients.

1-α,25-dihydroxyvitamin D3 potentiates avian osteoclast activation by increasing the formation of zipper-like structure via Src/Rac1 signaling.

Avian bone metabolism diseases affect the development and production of chickens, and many of these diseases can be prevented and controlled by balanced nutrition and hormone medicine. The steroid hormone 1α,25-dihydroxyvitamin D3 plays a key role in maintaining the balance of avian bone metabolism. Clinically, 1α,25-(OH)2D3 has been used to treat several bone diseases. Although several previous studies have investigated the effects of 1α,25-(OH)2D3 on osteoclastogenesis, the mechanisms underpinning osteoclast (OC) activity remain largely unknown. Herein, we used molecular and cell biology approaches to demonstrate that 1α,25-(OH)2D3 increases avian OC formation and activity, and upregulates bone resorption-related genes. Moreover, 1α,25-(OH)2D3 regulates the OC cytoskeleton by increasing the formation of zipper-like structure in OC precursor cells to potentiate OC activity via the Src/Rac1 signaling pathway. These findings provide new insight into the role of 1α,25-(OH)2D3 in OC activity.

Design and synthesis of highly selective pyruvate dehydrogenase complex E1 inhibitors as bactericides.

In order to obtain PDHc-E1 inhibitors with high selectivity and efficacy, four series (7, 12, 15, and 19) of 35 novel 4-aminopyrimidine derivatives were rationally designed and synthesized based on the binding site of ThDP in E. coli PDHc-E1. 12, 15, and 19 were confirmed to be potent inhibitors against E. coli PDHc-E1. Selected compounds 12g, 12i, 15f, and 19a showed negligible inhibition against porcine PDHc-E1. To understand their selectivity, the interaction of inhibitor and E. coli PDHc-E1 or porcine PDHc-E1 was studied by molecular docking. The newly introduced acylhydrazone and N-phenylbenzamide moieties could form stronger interaction by hydrogen bond at the active site of E. coli PDHc-E1 compared with that of porcine PDHc-E1. A part of title compounds as potent PDHc-E1 inhibitors also exhibited notable antibacterial activity. In particular, 12e, 12f, 12g, 12o, and 19a exhibited 72-92% inhibition against Xanthomonas oryzae pv. Oryzae and Ralstonia solanacearum at 100 µg/mL, which was better than thiodiazole-copper (34 and 29%, respectively) and bismerthiazol (56 and 55%, respectively). The results proved that we could obtain effective bactericidal compounds as highly selective PDHc inhibitors by rational molecular design utilizing the binding model of active site of E. coli PDHc-E1.

Preparation, Characterization, Pharmacokinetics and Biodistribution of Baicalin-Loaded Liposome on Cerebral Ischemia-Reperfusion after i.v. Administration in Rats.

The dry root of Scutellaria baicalensis, has traditionally been applied in the treatment of cerebral ischemia in Chinese clinics. Baicalin (BA) is considered the key ingredient in it for the brain protection effects. The bioavailability of BA is very low because of its poor lipid and water solubility, which limits the therapeutic effects and clinical application. The aim of the present study was to develop a novel BA-loaded liposome (BA-LP) formulation to improve the drug lipophilicity and further to enhance the drug-concentration in the brain tissues. This study is also designed to investigate the pharmacokinetics of BA in the pathological conditions of stroke and evaluate the pharmacokinetic differences of BA caused by stroke after intravenous administration with BA and BA-LP. In this study, the novel BA-LP prepared in early stage were characterized by morphology, size, zeta potential, encapsulation rate and the in vitro release. The pharmacokinetics and biodistribution of BA and BA-LP were investigated by intravenous administration in rats with middle cerebral artery occlusion (MCAO) model and normal group respectively. BA-LP had a mean particle size of 160⁻190 nm, zeta potential of -5.7 mV, and encapsulation efficiency of 42 ± 1%. The BA-LP showed a sustained-release behavior, the in vitro drug-release kinetic model of BA-LP fit well with the biphasic dynamic model equation: Q = 1 - (60.12e0.56t - 59.08e0.0014t). Pharmacokinetic behavior in MCAO rats is not consistent with that of normal rats. The middle cerebral artery occlusion rats got higher Cmax and AUC0⁻t, which were about 1.5⁻2 times to normal rats both in BA and liposome groups. In addition, it got especially higher distribution in brain, while BA were not detected in brain tissues on normal rats. The Cmax and AUC0⁻t values were significantly greater with liposome than BA on both normal and MCAO rats. The tissue distribution behavior was significantly altered in the case of liposome administrated in comparison with BA, which the concentrations in the heart, liver, spleen, lungs and brain were all increased after administrated liposome, but decreased in kidneys. The TI values showed that the target of liposome was improved especially to heart, spleen and brain, and the brain's target was higher in striatum and cerebellum. In conclusion, BA-LP might be a potential drug delivery system to improve the therapeutic efficacy of BA. In addition, these results also suggest that the pathological damages of ischemia-reperfusion have a significant impact on the pharmacokinetic traits of BA.

Design, Synthesis, and Potency of Pyruvate Dehydrogenase Complex E1 Inhibitors against Cyanobacteria.

Safe and effective algaecides are needed to control agriculturally and environmentally significant algal species. Four series (6, 10, 17, and 21) of 29 novel 4-aminopyrimidine derivatives were rationally designed and synthesized. A part of 10, 17, and 21 displayed potent inhibition of Escherichia coli pyruvate dehydrogenase complex E1 (E. coli PDHc-E1) (IC50 = 2.12-18.06 µM) and good inhibition of Synechocystis sp. PCC 6803 (EC50 = 0.7-7.1 µM) and Microcystis sp. FACH 905 (EC50 = 3.7-7.6 µM). The algaecidal activity of these compounds positively correlated with their inhibition of E. coli PDHc-E1. In particular, 21l and 10b exhibited potent algaecidal activity against PCC 6803 (EC50 = 0.7 and 0.8 µM, respectively), values that were 2-fold increased compared to that of copper sulfate (EC50 = 1.8 µM), and showed the best inhibition of cyanobacterium PDHc-E1 (IC50 = 5.10 and 6.06 µM, respectively). 17h and 21e, the best inhibitors of E. coli PDHc-E1, were studied by molecular docking, site-directed mutagenesis, and enzymatic assays. These results revealed that the improved inhibition of novel inhibitors compared with that of the lead compound I was due to the formation of a new hydrogen bond with Leu264 at the active site of E. coli PDHc-E1. The results proved the great potential to obtain effective algaecides via the rational design of PDHc-E1 inhibitors.