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Our ongoing efforts to explore the chemical space associated with marine cyanobacteria from coral reefs of Guam has yielded two new members of the anaenamide family of natural products, anaenamides C (3) and D (4). These compounds were isolated from a novel Hormoscilla sp. (VPG16-58). Our phylogenetic profiling (16S rDNA) of this cyanobacterium indicated that VPG16-58 is taxonomically distinct from the previously reported producer of the anaephenes, VPG16-59 (Hormoscilla sp.), and other previously documented species of the genus Hormoscilla. The planar structures of 3 and 4 were determined via spectroscopic methods, and absolute configurations of the α-hydroxy acids were assigned by enantioselective HPLC analysis. To address the requirement for sufficient material for testing, we first adapted our published linear synthetic approach for 1 and 2 to generate anaenoic acid (7) that served as a point for diversification, providing the primary amides 3 and 4 from synthetic intermediates 5 and 6, respectively. The compounds were then tested for effects on HCT116 colon cancer cell viability and in an ARE-luciferase reporter gene assay for Nrf2 modulation using HEK293 human embryonic kidney cells. Our findings indicate that, in contrast to cytotoxic methyl esters 1 and 2, the primary amides 3 and 4 activate the Nrf2 pathway at noncytotoxic concentrations. Overall, our data suggest that the anaenamide scaffold is tunable to produce differential biological outcomes.

Largazole is a potent class I selective histone deacetylase inhibitor prodrug with anticancer activity against solid tumors in preclinical models. Largazole possesses in vitro activity against glioblastoma multiforme (GBM) cells and sufficiently crosses the blood-brain barrier based on measurement of the active species, largazole thiol, to achieve therapeutically relevant concentrations in the mouse brain. The effective dose resulted in pronounced functional responses on the transcript level based on RNA sequencing and quantitative polymerase chain reaction after reverse transcription (RT-qPCR), revealing desirable expression changes of genes related to neuroprotection, including Bdnf and Pax6 upregulation, extending the applicability of largazole to the treatment of brain cancer and neurodegenerative disorders. The largazole-induced modulation of Pax6 unifies both activities, since Pax6 expression suppresses GBM proliferation and invasion and inversely correlates with GBM tumor grade, while it is also implicated in neurogenesis, neuronal plasticity, and cognitive ability. Our results suggest that largazole could be repurposed for diseases of the brain.

Ocular angiogenic diseases, characterized by abnormal blood vessel formation in the eye, are the leading cause of blindness. Although Anti-VEGF therapy is the first-line treatment in the market, a substantial number of patients are refractory to it or may develop resistance over time. As uncontrolled proliferation of vascular endothelial cells is one of the characteristic features of pathological neovascularization, we aimed to investigate the role of the class I histone deacetylase (HDAC) inhibitor Largazole, a cyclodepsipeptide from a marine cyanobacterium, in ocular angiogenesis. Our study showed that Largazole strongly inhibits retinal vascular endothelial cell viability, proliferation, and the ability to form tube-like structures. Largazole strongly inhibits the vessel outgrowth from choroidal explants in choroid sprouting assay while it does not affect the quiescent choroidal vasculature. Largazole also inhibits vessel outgrowth from metatarsal bones in metatarsal sprouting assay without affecting pericytes coverage. We further demonstrated a cooperative effect between Largazole and an approved anti-VEGF drug, Alflibercept. Mechanistically, Largazole strongly inhibits the expression of VEGFR2 and leads to an increased expression of cell cycle inhibitor, p21. Taken together, our study provides compelling evidence on the anti-angiogenic role of Largazole that exerts its function through mediating different signaling pathways. National Medical Research Council (NMRC) Published version This research was funded by the Duke-NUS Start-Up Grant to X.W., the Singapore National Medical Research Council DYNAMO NMRC/OFLCG/001/2017 and TAPP NMRC/OFLCG/004/2018 to X.W., and the Debbie and Sylvia DeSantis professorship to H.L.

New cyanobacteria-derived bifunctional analogues of doscadenamide A, a LasR-dependent quorum sensing (QS) activator in Pseudomonas aeruginosa, characterized by dual acylation of the pyrrolinone core structure and the pendant side chain primary amine to form an imide/amide hybrid are reported. The identities of doscadenamides B-J were confirmed through total synthesis and a strategic focused library with different acylation and unsaturation patterns was created. Key molecular interactions for binding with LasR and a functional response through mutation studies coupled with molecular docking were identified. The structure-activity relationships (SARs) were probed in various Gram-negative bacteria, including P. aeruginosa and Vibrio harveyi, indicating that the pyrrolinone-N acyl chain is critical for full agonist activity, while the other acyl chain is dispensable or can result in antagonist activity, depending on the bacterial system. Since homoserine lactone (HSL) quorum sensing activators have been shown to act in synergy with TRAIL to induce apoptosis in cancer cells, selected doscadenamides were tested in orthogonal eukaryotic screening systems. The most potent QS agonists, doscadenamides S10-S12, along with doscadenamides F and S4 with partial or complete saturation of the acyl side chains, exhibited the most pronounced synergistic effects with TRAIL in triple negative MDA-MB-231 breast cancer cells. The overall correlation of the SAR with respect to prokaryotic and eukaryotic targets may hint at coevolutionary processes and intriguing host-bacteria relationships. The doscadenamide scaffold represents a non-HSL template for combination therapy with TRAIL pathway stimulators.

35 p.-5 fig.-1 tab., Tubulin-targeted chemotherapy has proven to be a successful and wide spectrum strategy against solid and liquid malignancies. Therefore, new ways to modulate this essential protein could lead to new antitumoral pharmacological approaches. Currently known tubulin agents bind to six distinct sites at α/β-tubulin either promoting microtubule stabilization or depolymerization. We have discovered a seventh binding site at the tubulin intradimer interface where a novel microtubule-destabilizing cyclodepsipeptide, termed gatorbulin-1 (GB1), binds. GB1 has a unique chemotype produced by a marine cyanobacterium. We have elucidated this dual, chemical and mechanistic, novelty through multidimensional characterization, starting with bioactivity-guided natural product isolation and multinuclei NMR-based structure determination, revealing the modified pentapeptide with a functionally critical hydroxamate group; and validation by total synthesis. We have investigated the pharmacology using isogenic cancer cell screening, cellular profiling, and complementary phenotypic assays, and unveiled the underlying molecular mechanism by in vitro biochemical studies and high-resolution structural determination of the α/β-tubulin−GB1 complex., This research was supported by the NIH, National Cancer Institute Grants R01CA172310 (to H.L.) and R50CA211487 (to R.R.) and National Institute of General Medical Sciences Grant P41GM086210 (to H.L. and V.J.P.), Commercialization Fund Award from University of Florida (UF) Innovate (UF Office of Technology Licensing), and Debbie and Sylvia DeSantis Chair professorship (H.L.). The biochemical and the crystal structure work was supported by grants Ministerio de Ciencia e Innovación PID2019-10454RB-I00/AEI/10.13039/501100011033, Fondo de Investigaciones Sanitarias and COV20/01007E Proyecto Intramural Especial 201920E111 from Consejo Superior de Investigaciones Científicas (to J.F.D.) and European Union H2020-MSCA-ITN-2019 860070 TUBINTRAIN grant (to J.F.D. and A.E.P.).

We describe the total synthesis of tutuilamide A, a potent porcine pancreatic elastase (PPE) inhibitor and a representative member of the 3-amino-6-hydroxy-2-piperidone (Ahp) cyclodepsipeptide family, isolated from marine cyanobacteria. The Ahp unit serves as a pharmacophore and the adjacent 2-amino-2-butenoic acid (Abu) is a main driver of the selectivity among serine proteases. We adapted our previous convergent strategy to generate the macrocycle, common with lyngbyastatin 7 and related elastase inhibitors, and then appended the tutuilamide A-specific side chain bearing a vinyl chloride. Tutuilamide A and lyngbyastatin 7 were evaluated side by side for the inhibition of the disease-relevant human neutrophil elastase (HNE). Tutuilamide A and lyngbyastatin 7 were approximately equipotent against HNE, while tutuilamide A was previously shown to be more active against PPE compared with lyngbyastatin 7, further demonstrating that the side chain provides opportunities to not only modulate potency but also selectivity among proteases of the same function from different organisms. Profiling of tutuilamide A against mainly human serine proteases revealed high selectivity for HNE (IC50 0.73 nM) and pleiotropic activity against kallikrein 7 (KLK7, IC50 5.0 nM), without affecting other kallikreins, similarly to lyngbyastatin 7 (IC50 0.85 nM for HNE and 3.1 nM for KLK7). A comprehensive molecular docking study for elastases and KLK7 afforded deeper insight into the intricate differences between inhibitor interactions with HNE and PPE, accounting for the differential activities for both compounds. The synthesis and molecular studies serve as a proof-of-concept that the macrocyclic scaffold can be diversified to fine-tune the activity of serine protease inhibitors.

New modified depsipeptides and geometric isomers, termed anaenamides A (1a) and B (1b), along with the presumptive biosynthetic intermediate, anaenoic acid (2), were discovered from a marine cyanobacterium from Guam. Structures were confirmed by total synthesis. The alkyl-salicylic acid fragment and the C-terminal α-chlorinated α,β-unsaturated ester are novelties in cyanobacterial natural products. Cancer cell viability assays indicated that the C-terminal unit serves as the pharmacophore and that the double bond geometry impacts the cytotoxicity.

Renal, hepatocellular, and neuroendocrine carcinomas are known as highly vascularized tumors. Although vascular endothelial growth factor A (VEGF-A)-targeted therapies have shown efficacy in the treatment of these cancers, drug resistance is a major concern and might be mediated by interleukin 6 (IL-6). Furthermore, upon antiangiogenic drug exposure, tumor cells may adapt to survive in a vascular-independent manner. Apratoxins are potent marine-derived cytotoxic in vivo-active agents, preventing cotranslational translocation in the secretory pathway, and show promise to overcome resistance by targeting angiogenesis and tumor growth simultaneously. We designed and synthesized a novel apratoxin analogue, apratoxin S10, with a balanced potency and stability as well as synthetic accessibility and scalability. We showed that apratoxin S10 potently inhibits both angiogenesis in vitro and growth of cancer cells from vascularized tumors. Apratoxin S10 down-regulated vascular endothelial growth factor receptor 2 (VEGFR2) on endothelial cells and blocked the secretion of VEGF-A and IL-6 from cancer cells. It inhibited cancer cell growth through down-regulation of multiple receptor tyrosine kinases (RTKs) and compares favorably to currently approved RTK inhibitors in both angiogenesis and cancer cell growth.

Seaweeds are an important component of human diets, especially in Asia and the Pacific islands, and have shown chemopreventive as well as anti-inflammatory properties. However, structural characterization and mechanistic insight of seaweed components responsible for their biological activities are lacking. We isolated cymopol and related natural products from the marine green alga Cymopolia barbata and demonstrated their function as activators of transcription factor Nrf2-mediated antioxidant response to increase the cellular antioxidant status. We probed the reactivity of the bioactivation product of cymopol, cymopol quinone, which was able to modify various cysteine residues of Nrf2's cytoplasmic repressor protein Keap1. The observed adducts are reflective of the polypharmacology at the level of natural product, due to multiple electrophilic centers, and at the amino acid level of the cysteine-rich target protein Keap1. The non-polar C. barbata extract and its major active component cymopol, reduced inflammatory gene transcription in vitro in macrophages and mouse embryonic fibroblasts in an Nrf2-dependent manner. Cymopol-containing extracts attenuated neutrophil migration in a zebrafish tail wound model. RNA-seq analysis of colonic tissues of mice exposed to non-polar extract or cymopol showed an antioxidant and anti-inflammatory response, with more pronounced effects exhibited by the extract. Cymopolia extract reduced DSS-induced colitis as measured by fecal lipocalin concentration. RNA-seq showed that mucosal-associated bacterial composition and transcriptional profile in large intestines were beneficially altered to varying degrees in mice treated with either the extract or cymopol. We conclude that seaweed-derived compounds, especially cymopol, alter Nrf2-mediated host and microbial gene expression, thereby providing polypharmacological effects.

Purpose Abnormal blood vessel formation is a defining feature of many blinding eye diseases. Targeting abnormal angiogenesis by inhibiting VEGF has revolutionized the treatment of many ocular angiogenic diseases over the last decade. However, a substantial number of patients are refractory to anti-VEGF treatment or may develop resistance over time. The objective of this study was to determine the efficacy and the mechanism of action of Apratoxin S4 in ocular angiogenesis. Methods Retinal vascular cell proliferation, migration, and the ability to form tube-like structure were studied in vitro. Ex vivo aortic ring, choroid, and metatarsal assays were used to study Apratoxin S4's impact on vessel outgrowth in a multicellular environment. Apratoxin S4 was also tested in mouse models of oxygen-induced retinopathy (OIR) and laser-induced choroidal neovascularization (CNV), and in a rabbit model of persistent retinal neovascularization (PRNV). Western blot and ELISA were used to determine the expression of key angiogenic regulators after Apratoxin S4 treatment. Results Apratoxin S4 strongly inhibits retinal vascular cell activation by suppressing multiple angiogenic pathways. VEGF-activated vascular cells and angiogenic vessels are more susceptible to Apratoxin S4 treatment than quiescent vascular cells and vessels. Both intraperitoneal and intravitreal delivery of Apratoxin S4 are able to impede ocular neovascularization in vivo. Apratoxin S4 specifically attenuates pathological ocular angiogenesis and exhibits a combinatorial inhibitory effect with standard-of-care VEGF inhibitor drug (aflibercept). Conclusions Apratoxin S4 is a potent antiangiogenic drug that inhibits the activation of retinal endothelial cells and pericytes through mediating multiple angiogenic pathways.

Apratoxin E provided the inspiration for the design of apratoxin A/E hybrids under preclinical development. Through total synthesis using two different strategies, it was determined that the originally proposed configuration of the thiazoline at C30 is opposite from that in apratoxin A, in contrast to previous assumptions on biosynthetic grounds. The epimer and true natural apratoxin E were synthesized, and the biological activities were evaluated.

Two new cyclic lipopeptides termed laxaphycins B4 (1) and A2 (2) were discovered from a collection of the marine cyanobacterium Hormothamnion enteromorphoides, along with the known compound laxaphycin A. The planar structures were solved based on a combined interpretation of 1D and 2D NMR data and mass spectral data. The absolute configurations of the subunits were determined by chiral LC-MS analysis of the hydrolysates, advanced Marfey’s analysis and 1D and 2D ROESY experiments. Consistent with similar findings on other laxaphycin A- and B-type peptides, laxaphycin B4 (1) showed antiproliferative effects against human colon cancer HCT116 cells with IC50 of 1.7 µM, while laxaphycins A and A2 (2) exhibited weak activities. The two major compounds isolated from the sample, laxaphycins A and B4, were shown to act synergistically to inhibit the growth of HCT116 colorectal cancer cells.

Herein we describe the research and development of the process for the scale-up total synthesis of largazole, a potent class I selective histone deacetylase (HDAC) inhibitor, a potential anticancer agent and also useful for the treatment of other disorders where transcriptional reprogramming might be beneficial. The synthetic route and conditions for each fragment and final product were modified and optimized to make them suitable for larger-scale synthesis. With the process we developed, hundreds of grams of each fragment and decagrams of final product largazole were synthesized in good to excellent yields. The final target largazole was obtained in 21% overall yield over eight steps based on the longest sequence with over 95% HPLC purity.

A novel linear depsipeptide enriched with tyrosine-derived moieties, termed apratyramide, was isolated from an apratoxin-producing cyanobacterium. The structure was determined using a combination of NMR spectroscopy, mass spectrometry, and chiral analysis of the acid hydrolyzate and confirmed by total synthesis. Apratyramide up-regulated multiple growth factors at the transcript level in human keratinocyte (HaCaT) cells and induced the secretion of vascular endothelial growth factor A (VEGF-A) from HaCaT cells, suggesting the compound's potential wound-healing properties through growth factor induction. Transcriptome analysis and sequential validation supported the hypothesis and indicated its mode of action (MOA) through the unfolded protein response (UPR) pathway, which is functionally related to wound healing and angiogenesis. The conditioned medium of HaCaT cells treated with apratyramide induced angiogenesis in vitro. An ex vivo rabbit corneal epithelial model was applied to confirm the VEGF-A induction in this wound-healing model.

Largazole is a potent class I selective histone deacetylase (HDAC) inhibitor. The majority of largazole analogues to date have modified the thiazole-thiazoline and the warhead moiety. In order to elucidate class I-specific structure-activity relationships, a series of analogues with modifications in the valine or the linker region were prepared and evaluated for their class I isoform selectivity. The inhibition profile showed that the C2 position of largazole has an optimal steric requirement for efficient HDAC inhibition and that substitution of the trans-alkene in the linker with an aromatic group results in complete loss of activity. This data will aid the design of class I isoform selective HDAC inhibitors.

Largazole is a potent and class I-selective histone deacetylase (HDAC) inhibitor purified from marine cyanobacteria and was demonstrated to possess antitumor activity. Largazole employs a unique prodrug strategy, via a thioester moiety, to liberate the bioactive species largazole thiol. Here we report alternate prodrug strategies to modulate the pharmacokinetic and pharmacodynamics profiles of new largazole-based compounds. The in vitro effects of largazole analogues on cancer cell proliferation and enzymatic activities of purified HDACs were comparable to the natural product. However, in vitro and in vivo histone hyperacetylation in HCT116 cells and implanted tumors, respectively, showed differences, particularly in the onset of action and oral bioavailability. These results indicate that, by employing a different approach to disguise the "warhead" moiety, the functional consequence of these prodrugs can be significantly modulated. Our data corroborate the role of the pharmacokinetic properties of this class of compounds to elicit the desired and timely functional response.

Apratoxins are cytotoxic natural products originally isolated from marine cyanobacteria that act by preventing cotranslational translocation early in the secretory pathway to downregulate receptor levels and inhibit growth factor secretion, leading to potent antiproliferative activity. Through rational design and total synthesis of an apratoxin A/E hybrid, apratoxin S4 (1a), we have previously improved the antitumor activity and tolerability in vivo. Compound 1a and newly designed analogues apratoxins S7–S9 (1b–d), with various degrees of methylation at C34 (1b,c) or epimeric configuration at C30 (1d), were efficiently synthesized utilizing improved procedures. Optimizations have been applied to the synthesis of key intermediate aldehyde 7 and further include the application of Leighton’s silanes and modifications of Kelly’s methods to induce thiazoline ring formation in other crucial steps of the apratoxin synthesis. Apratoxin S9 (1d) exhibited increased activity with subnanomolar potency. Apratoxin S8 (1c) lacks the propensity to be deactivated by dehydration and showed efficacy in a human HCT116 xenograft mouse model.

Apratoxins are cytotoxic marine natural products that prevent cotranslational translocation early in the secretory pathway. We showed that apratoxins downregulate receptors and growth factor ligands, giving a one-two punch to cancer cells, particularly those that rely on autocrine loops. Through total synthesis, we tested the effects of amino acid substitutions, including alanine scanning, on the downregulation of receptor tyrosine kinases and vascular endothelial growth factor A (VEGF-A) and probed the stereospecificity of target engagement by epimerization of selected chiral centers. Differential effects on two types of secretory molecules suggest that the apratoxins' substrate selectivity with respect to inhibition of secretion may be tuned through structural modifications to provide tailored therapy. Our structure-activity relationship studies and medicinal chemistry efforts led to a potent inhibitor with in vivo efficacy in a colorectal tumor xenograft model without irreversible toxicity exerted by apratoxin A, demonstrating that this novel mechanism of action has therapeutic potential.

OBJECTIVES To evaluate the safety and efficacy of performing ultrasound-guided minimally invasive percutaneous nephrolithotomy (MPCNL) in the flank position for the management of complex renal calculi. Percutaneous nephrolithotomy is usually performed with the patient in the prone position under fluoroscopic guidance; however, this position, and guidance method have some limitations. METHODS From January 2007 to December 2009, 93 patients (101 kidneys) with complex renal calculi underwent ultrasound-guided MPCNL in the flank position. RESULTS The mean age of the patients was 45.3 years (range 29-71). The calculi-free rate in the patients who underwent a single procedure was 78.2% (79 of 101 kidneys). The average operative duration was 82.6 minutes (range 45-190). Although the perioperative blood loss was not significantly different between single-tract and double-tract MPCNL (P = .087, F = 2.981), the calculi-free rate was significantly greater in the patients who underwent double-tract MPCNL than in those who underwent single-tract MPCNL (P = .027, chi-square = 4.873). Perioperative blood transfusions were not required in any patient. Similarly, ureteral calculi due to percutaneous nephrolithotomy were not observed. Secondary renal hemorrhage occurred in 3 patients who had undergone single-tract MPCNL and 1 underwent nephrectomy. CONCLUSIONS The results of our study have shown that ultrasound-guided MPCNL with the patient in the flank position is safe and effective for treating complex renal calculi, without the side effects of radiation to the patient and surgeon. Double-tract MPCNL is suitable for complex renal calculi and, in some cases, is required to increase the calculi-free rate. The insertion of twin ureteral catheters before lithotripsy might be helpful in avoiding residual ureteral calculi after percutaneous nephrolithotomy.

Lyngbyastatin 7 (1) is a marine cyanobacteria-derived lariat-type cyclic depsipeptide, of which the macrocyclic core possesses modified amino acids, including a featured 3-amino-6-hydroxy-2-piperidone (Ahp) moiety and a (Z)-2-amino-2-butenoic acid (Abu) moiety. The first total synthesis of 1 was successfully established via 31 steps, and the conditions of several crucial steps were optimized to ensure smooth operations. The previously reported structural assignments and elastase inhibitory activity of the isolated natural product were confirmed. According to the extensive in vitro biological evaluation, compound 1 displayed low nanomolar IC50 in blocking elastase activity, strong ability in protecting bronchial epithelial cells against elastase-induced antiproliferation and abrogating the elastase-triggered induction of pro-inflammatory cytokine expression. Its overall performance was superior over sivelestat, the only approved small molecule drug targeting elastase, which indicated its potential in developing as a pharmacotherapeutic against elastase-mediated pathologies. The success in total synthesis, designed with a novel convergent strategy, not only overcame the supply issue for thorough preclinical studies, but also paved the way for convenient synthesis of analogues with improved potency and drug-like properties.

A new neutral fluorescent chemosensor 5 based on a calix[4]arene tetraamide derivative has been synthesized. It exhibits high selectivity for H 2 PO 4 - over a wide range of anions, and the selectivity for H 2 PO 4 - is more than 2700-fold higher than for F - . In acetonitrile, the fluorescence intensity of 5 is efficiently enhanced about 130% [(I - I 0 )/I 0 ] upon the addition of 5 equiv. of H 2 PO 4 - . From the fluorescence titration experiments, the association constant for 5/H 2 PO 4 - (1:2) was calculated to be 5.48×10 9 M - 2 . Furthermore, the 1 H NMR spectral and X-ray crystal studies suggested that multiple hydrogen-bonding interactions occur between 5 and H 2 PO 4 - .

A new fluorescent as well as chromogenic anion sensor, 1,8-anthradiol bis(N-phenylcarbamate) 2 , was synthesized. It exhibits new selective red-shifted absorption and fluorescence bands with F− and AcO−, which could be attributed to the anthracene moiety directly involved in the bonding interaction with the anions.

A new fluorescent chemosensor based on an acyclic tetra-sulfonamide derivative linked to two dansyl groups has been conveniently synthesized. Its high selective binding ability to fluoride ions over other halide ions was demonstrated by using fluorescence as well as 1H NMR spectra.

To determine the hemodynamic status, fluid-electrolyte changes and complications associated with irrigation time in percutaneous nephrolithotripsy.A total of 68 renal calculi patients (31 males and 37 females) were recruited. The lateral recumbent percutaneous nephrolithotripsy was operated with Ho laser under ultrasonic guidance. 0.9% NaCI was used as perfusion fluid. The following items were recorded: mean arterial blood pressure (MAP), heart rate, central venous pressure (CVP), hemoglobin, sodium, potassium and chloride; perfusion time during operation; peri-operative and post-operative complications.(1) Peri-operative and post-operative conditions: the average operative time was 83.1 +/- 22.21 minutes. Two cases stopped because of bleeding after puncture and the tube of stoma was placed for stone clearance of the second time. There was more bleeding in 11 patients, but the operations were continued with blood transfusion and close monitoring. Two operations ceased because of a premunition of congestive heart failure. Nine patients needed post-operative blood transfusion and 18 had a post-operative fever. One patient bled in and around the tube and had a peri-renal infection a week later. (2) Changes of observation parameters: there was no significant difference in CVP, heart rate, hemoglobin, sodium, potassium and chloride (P0.05). The post-perfusion value of MAP increased (P0.05) especially in the cases of more bleeding and long time of irrigation. Peri-operative and post-operative injection of furosemide could reduce the CVP value. The average irrigation time in the fever group was longer than the non-fever group (P0.05) and the CVP value of the fever group was higher than the non-fever group (P0.05).Low pressure and short time of perfusion are safe in clinical practice. Congestive heart failure after the perfusion and the occurrence of post-operative infections are difficult to avoid when there are a long time of irrigation and more bleeding during operation.

To explore the optimal method for protein expression in rhES (recombinant human endostatin) and study the anti-tumor activities of rhES in solid tumor and established cell line.Different IPTG concentrations, the timing of adding IPTG into the culture medium and the different time of expression were employed to explore the optimized conditions of protein expression. Activity examination: (1) animal experiment: nude mice bearing subcutaneous cancer in treated group and controlled group were observed. (2) cellular experiment: the inhibitory effect of rhES in T-24 established cell line were observed by MTT assay and cancer cell growth curve.The expression of rhES protein was 58.65%. Of all the E. coli body proteins, the protein purity after purification was 96.22%. Activity examination indicated that rhES could inhibit the growth of solid tumor and the established cell line. In animal experiment, the tumor inhibition rate was 66.8%. Cell experiment: the 50% inhibitory concentration (IC(50)) was 22 microg/ml. The cell population decreased 58.75% than the control group at Day 7 in the tumor cell growth curve.A high expression and activity of rhES protein can be obtained by the optimized expression conditions. rhES can inhibit the cellular growth in both solid tumor and established cell line of bladder cancer.

We previously identified two small molecules targeting the HIV-1 gp41, N-(4-carboxy-3-hydroxy)phenyl-2,5-dimethylpyrrole 12 (NB-2) and N-(3-carboxy-4-chloro)phenylpyrrole 13 (NB-64), that inhibit HIV-1 infection at low micromolar levels. On the basis of molecular docking analysis, we designed a series of 2-aryl 5-(4-oxo-3-phenethyl-2-thioxothiazolidinylidenemethyl)furans. Compared with 12 and 13, these compounds have bigger molecular size (437-515 Da) and could occupy more space in the deep hydrophobic pocket on the gp41 NHR trimer. Fifteen 2-aryl 5-(4-oxo-3-phenethyl-2-thioxothiazolidinylidenemethyl)furans (11a-o) were synthesized by Suzuki-Miyaura cross-coupling followed by a Knoevenagel condensation and tested for their anti-HIV-1 activity and cytotoxicity on MT-2 cells. We found that all 15 compounds had improved anti-HIV-1 activity and 3 of them (11a, 11b, and 11d) exhibited inhibitory activity against replication of HIV-1(IIIB) and 94UG103 at

N-(4-Arylazobenzoyl)-1H-benzotriazoles 15a, 15b react with dipeptides 12a-d, (12d+12d') and tripeptides 14a, 14b to give azodye labeled-dipeptides (16a-e), (16d+16d'), (16e+16e') and -tripeptides 16f, 16g in high yields (73-93%) with retention of chirality.

To investigate the clinical value and safety of holmium: YAG laser endoureterotomy in the treatment of ureteral obstruction.Holmium: YAG laser endoureterotomy, with the laser optic fiber 550 microm in diameter and the output power of 3.5 Watt, via ureteroscopy, was performed on 18 patients ureteral obstruction, 8 males and 10 females, aged 52.1 (34-67), 11 with the stricture in the upper segment (complete obstruction in 4 cases), 5 in the middle segment, and 2 in lower segment; and 6 cases complicated with ureteral calculus. Postoperatively, an orthopedic ureteral stent (a 6-Fr double-J ureteral stent with a movable 5 cm length 9-Fr orthopedic cannula) was remained indwelling for 3-6 months. Follow-up was conducted for 10.7 (2-14) months.The operative duration was 32 (25-70) minutes. One patient underwent failed endoureterotomy and was turned to percutaneous nephroscopy. Success was achieved in 16 patients. The glomerular filtration rate (GFR) of these affected kidneys increased from 16.4+/-6.9 ml/min ante-operatively to 24.9+/-8.2 ml/min (P0.01) postoperatively. One kidney was resected because of non-function, with GFR of 2 ml/min and intractable pyelitis. No recurrence of ureteral stricture was observed.Holmium: YAG laser endoureterotomy with insertion of orthopedic ureteral stent is an efficient and safe treatment for ureteral strictures with minimal invasion, less complications and easy recovery. This operation should be performed with a thorough preparation and severely restricted indication.

To investigate the effects of 4- hydroxytamoxifen (OHT) on the proliferation and apoptosis of prostate smooth muscle cells and the expression of estrogen receptor (ER) and androgen receptor (AR).Prostate smooth muscle cells were isolated from the resected specimens of prostate glands of 10 patients with benign prostatic hypertrophy (BPH), cultured, and exposed to estradiol (E(2)), diethylstilbestrol (DES), and OHT of different concentrations (1 x 10(-8) - 1 x 10(-5) mol/L) or mixture of E(2) (1 x 10(-8) - 1 x 10(-6) mol/L) with OHT (1 x 10(-7) mol/L). Flow cytometry was used to test the proliferation and apoptosis of the cells, and immunocytochemistry was used to test the expression of estrogen and androgen receptors.E(2) and DES promoted the proliferation of the prostate smooth muscle cells in a certain concentration range, but not dose-dependently, and OHT at the concentration of 1 x 10(-8) mol/L slightly increased the G(2)-M peak rate of the prostate smooth muscle cells, but suppressed the G(2)-M peak rate dose-dependently when its concentration wasor= 1 x 10(-7) mol/L (P0.05) and this suppression effect was dose-dependently (r = -0.312, P = 0.011). E(2) at the concentrationor= 1 x 10(-5) mol/L and DES at the concentrationor= 1 x 10(-6) mol/L slightly promoted the apoptosis of the prostate smooth muscle cells, but not dose-dependently, and OHT at the concentrations from 1 x 10(-8) mol/L to 1 x 10(-5) mol/L promoted the apoptosis of the prostate smooth muscle cells dose-dependently (r = 0.363, P = 0.021) and this effect could not be reversed by administration of E(2) at the concentration 1 x 10(-8) - 1 x 10(-6) mol/L (P0.05). E(2), DES, and OHT of different concentrations all increased the ERalpha and AR positive staining rates of the prostate smooth muscle cells (all P0.05).OHT suppresses the proliferation and promotes the apoptosis of prostate smooth muscle cells, and these functions do not depend on the estrogen receptor pathway. Low blood OHT concentration after oral administration of TAM and up-regulation of estrogen receptors by OHT may be the caused of the inefficiency of TAM for treatment of BPH.

N-(4-Arylazobenzoyl)-1H-benzotriazoles 3 react with amino acids 4 and amines 6 to give azo-dye labeled amino acids (5a–m) and amines (7a–n) in high yields (74–100%) with retention of chirality.

trans-tert-Butyldimethylsiloxy-L-proline displays greater catalytic activity and affords higher enantioselectivity than the parent proline in the alpha-amination reaction of carbonyl compounds with azodicarboxylate. A quantum mechanical calculation reveals the structure of the transition state. In the presence of a catalytic amount of siloxyproline and water (3-9 equiv), alpha-amino carbonyl derivatives, which are important synthetic intermediates, are obtained in good yield and with excellent enantioselectivity.

trans-tert-Butyldimethylsiloxy-L-proline displays greater catalytic activity and affords higher enantioselectivity than the parent proline in the alpha-amination reaction of carbonyl compounds with azodicarboxylate. A quantum mechanical calculation reveals the structure of the transition state. In the presence of a catalytic amount of siloxyproline and water (3-9 equiv), alpha-amino carbonyl derivatives, which are important synthetic intermediates, are obtained in good yield and with excellent enantioselectivity.

Methoxy-9,10-anthraquinones with mono-, di- and tetraether groups at different positions la-h can be directly reduced to the corresponding methoxyanthracenes 3a-h in moderate to good yields by zinc in refluxing acetic acid. Under similar conditions, ethyl I'-anthracenoxyacetate (3i) with the ester group unaffected and 1,8-oxybis(ethyleneoxyethyleneoxy)anthracene (5) were also conveniently synthesized in 65 and 70% yields, respectively.

N-(4-Arylazobenzoyl)-1H-benzotriazoles 3 react with amino acids 4 and amines 6 to give azo-dye labeled amino acids (5a-m) and amines (7a-n) in high yields (74-100%) with retention of chirality.

Azocalix[4]arene derivatives 1–5, with one or two azophenol chromophores and different binding sites, were found to allow not only for high selective and sensitive colorimetric detection but also easy colorimetric differentiation of F−, AcO− and H2PO4−, of similar basicity, depending upon the azocalix[4]arene structure, guest basicity and conformational complementarity between the host and guest.