Total syntheses of Tetrodotoxin and 9-epiTetrodotoxin.

Tetrodotoxin and congeners are specific voltage-gated sodium channel blockers that exhibit remarkable anesthetic and analgesic effects. Here, we present a scalable asymmetric syntheses of Tetrodotoxin and 9-epiTetrodotoxin from the abundant chemical feedstock furfuryl alcohol. The optically pure cyclohexane skeleton is assembled via a stereoselective Diels-Alder reaction. The dense heteroatom substituents are established sequentially by a series of functional group interconversions on highly oxygenated cyclohexane frameworks, including a chemoselective cyclic anhydride opening, and a decarboxylative hydroxylation. An innovative SmI2-mediated concurrent fragmentation, an oxo-bridge ring opening and ester reduction followed by an Upjohn dihydroxylation deliver the highly oxidized skeleton. Ruthenium-catalyzed oxidative alkyne cleavage and formation of the hemiaminal and orthoester under acidic conditions enable the rapid assembly of Tetrodotoxin, anhydro-Tetrodotoxin, 9-epiTetrodotoxin, and 9-epi lactone-Tetrodotoxin.

Experience of individualized nursing in patients with laryngeal squamous cell carcinoma combined with Helicobacter pylori infection after surgery.

OBJECTIVE: To explore the effect of personalized nursing on the therapeutic effect on patients with laryngeal squamous cell carcinoma (LSCC) combined with Helicobacter pylori (HP) infection after surgery. METHODS: This study enrolled 46 patients with LSCC as research subjects. The patients were divided into an experimental group and a control group using the random number table method. Routine nursing was provided in the control group, and individualized nursing intervention was conducted in the experimental group based on routine nursing. The incidence of adverse reactions (ARs) and the patients' nursing satisfaction were compared between the two groups. RESULTS: Compared with the control group, the individualized nursing intervention reduced the incidence of ARs of esophageal reflux (P < 0.05) and improved nursing satisfaction (P < 0.05). CONCLUSION: Individualized nursing intervention reduced the incidence of esophageal reflux ARs in patients with LSCC and HP infection after tracheotomy and improved both the therapeutic effect of the treatment and the satisfaction of the patients.

Associations of Dietary Zinc-Vitamin B6 Ratio with All-Cause Mortality and Cardiovascular Disease Mortality Based on National Health and Nutrition Examination Survey 1999-2016.

Previous studies have suggested a possible association among dietary zinc and vitamin B6 intake and CVD mortality and all-cause mortality. However, evidence on the association of dietary zinc and vitamin B6 intake and their interactions with CVD mortality and all-cause mortality remains unclear. This prospective study utilized data from the US National Health and Nutrition Examination Survey (NHANES) from 1999 to 2016. After a median follow-up of 10.4 years, 4757 deaths were recorded among 36,081 participants. Higher dietary zinc intake levels (≥9.87 mg/day) were associated with lower CVD mortality (hazard ratio (HR) = 0.85, 95% confidence interval (CI): 0.83−0.87). Vitamin B6 intake levels (≥1.73 mg/day) were associated with lower CVD mortality (HR = 0.91, 95% CI: 0.86−0.96) and all-cause mortality (HR = 0.91, 95% CI: 0.90−0.93). Higher dietary zinc intake and higher vitamin B6 intake were associated with a lower risk of CVD mortality, with an interaction between dietary zinc intake levels and vitamin B intake levels (LZLV group (HR, CI): 1.21,1.12−1.29; LZHV group (HR, CI): 1.42, 1.34−1.50; LZHV group (HR, CI): 1.28, 1.14−1.45; HZHV group (HR, CI): ref). There was also a J-type association (p for nonlinear < 0.001) between the dietary zinc−vitamin B6 ratio and CVD mortality, with a high dietary zinc−vitamin B6 ratio increasing the risk of CVD mortality (HR = 1.27, 95% CI: 1.19−1.35), whereas a moderate dietary zinc−vitamin B6 ratio appeared to be beneficial for CVD mortality. These results suggest that increasing the appropriate proportion of dietary zinc and vitamin B6 intake is associated with a lower risk of CVD mortality. Furthermore, precise and representative studies are needed to verify our findings.

An improved auxin-inducible degron system for fission yeast.

Conditional degron technologies, which allow a protein of interest to be degraded in an inducible manner, are important tools for biological research, and are especially useful for creating conditional loss-of-function mutants of essential genes. The auxin-inducible degron (AID) technology, which utilizes plant auxin signaling components to control protein degradation in nonplant species, is a widely used small-molecular-controlled degradation method in yeasts and animals. However, the currently available AID systems still have room for further optimization. Here, we have improved the AID system for the fission yeast Schizosaccharomyces pombe by optimizing all three components: the AID degron, the small-molecule inducer, and the inducer-responsive F-box protein. We chose a 36-amino-acid sequence of the Arabidopsis IAA17 protein as the degron and employed three tandem copies of it to enhance efficiency. To minimize undesirable side effects of the inducer, we adopted a bulky analog of auxin, 5-adamantyl-IAA, and paired it with the F-box protein OsTIR1 that harbors a mutation (F74A) at the auxin-binding pocket. 5-adamantyl-IAA, when utilized with OsTIR1-F74A, is effective at concentrations thousands of times lower than auxin used in combination with wild-type OsTIR1. We tested our improved AID system on 10 essential genes and achieved inducible lethality for all of them, including ones that could not be effectively inactivated using a previously published AID system. Our improved AID system should facilitate the construction of conditional loss-of-function mutants in fission yeast.

Structure of PDE3A-SLFN12 complex and structure-based design for a potent apoptosis inducer of tumor cells.

Molecular glues are a class of small molecular drugs that mediate protein-protein interactions, that induce either the degradation or stabilization of target protein. A structurally diverse group of chemicals, including 17-ß-estradiol (E2), anagrelide, nauclefine, and DNMDP, induces apoptosis by forming complexes with phosphodiesterase 3A (PDE3A) and Schlafen 12 protein (SLFN12). They do so by binding to the PDE3A enzymatic pocket that allows the compound-bound PDE3A to recruit and stabilize SLFN12, which in turn blocks protein translation, leading to apoptosis. In this work, we report the high-resolution cryo-electron microscopy structure of PDE3A-SLFN12 complexes isolated from cultured HeLa cells pre-treated with either anagrelide, or nauclefine, or DNMDP. The PDE3A-SLFN12 complexes exhibit a butterfly-like shape, forming a heterotetramer with these small molecules, which are packed in a shallow pocket in the catalytic domain of PDE3A. The resulting small molecule-modified interface binds to the short helix (E552-I558) of SLFN12 through hydrophobic interactions, thus "gluing" the two proteins together. Based on the complex structure, we designed and synthesized analogs of anagrelide, a known drug used for the treatment of thrombocytosis, to enhance their interactions with SLFN12, and achieved superior efficacy in inducing apoptosis in cultured cells as well as in tumor xenografts.

5-IP7 is a GPCR messenger mediating neural control of synaptotagmin-dependent insulin exocytosis and glucose homeostasis.

5-diphosphoinositol pentakisphosphate (5-IP7) is a signalling metabolite linked to various cellular processes. How extracellular stimuli elicit 5-IP7 signalling remains unclear. Here we show that 5-IP7 in ß cells mediates parasympathetic stimulation of synaptotagmin-7 (Syt7)-dependent insulin release. Mechanistically, vagal stimulation and activation of muscarinic acetylcholine receptors triggers Gαq-PLC-PKC-PKD-dependent signalling and activates IP6K1, the 5-IP7 synthase. Whereas both 5-IP7 and its precursor IP6 compete with PIP2 for binding to Syt7, Ca2+ selectively binds 5-IP7 with high affinity, freeing Syt7 to enable fusion of insulin-containing vesicles with the cell membrane. ß-cell-specific IP6K1 deletion diminishes insulin secretion and glucose clearance elicited by muscarinic stimulation, whereas mice carrying a phosphorylation-mimicking, hyperactive IP6K1 mutant display augmented insulin release, congenital hyperinsulinaemia and obesity. These phenotypes are absent in mice lacking Syt7. Our study proposes a new conceptual framework for inositol pyrophosphate physiology in which 5-IP7 acts as a GPCR second messenger at the interface between peripheral nervous system and metabolic organs, transmitting Gq-coupled GPCR stimulation to unclamp Syt7-dependent, and perhaps other, exocytotic events.

Design of Dimeric Bile Acid Derivatives as Potent and Selective Human NTCP Inhibitors.

Dimeric bile acid derivatives (DBADs) were developed and tested for their anti-HBV and anti-HDV activities as sodium taurocholate cotransporting polypeptide (NTCP) inhibitors. DBADs exhibited strong and persistent potency of NTCP inhibition, whereas diverse linkers and constitutions showed distinct inhibition features. Motif aa157-165 on NTCP was shown to be a possible binding site of DBADs; therefore, we determined DBADs' selectivity among NTCPs from different species. A cyclized DBAD scaffold DBA-41 exhibited a high affinity to human NTCP (hNTCP). Intraperitoneal administration of DBA-41 to hNTCP-tg mice induced serum total bile acid elevation. DBA-41 may serve as a biological tool to study NTCP physiological function.

Discovery of a molecular glue promoting CDK12-DDB1 interaction to trigger cyclin K degradation.

Molecular-glue degraders mediate interactions between target proteins and components of the ubiquitin-proteasome system to cause selective protein degradation. Here, we report a new molecular glue HQ461 discovered by high-throughput screening. Using loss-of-function and gain-of-function genetic screening in human cancer cells followed by biochemical reconstitution, we show that HQ461 acts by promoting an interaction between CDK12 and DDB1-CUL4-RBX1 E3 ubiquitin ligase, leading to polyubiquitination and degradation of CDK12-interacting protein Cyclin K (CCNK). Degradation of CCNK mediated by HQ461 compromised CDK12 function, leading to reduced phosphorylation of a CDK12 substrate, downregulation of DNA damage response genes, and cell death. Structure-activity relationship analysis of HQ461 revealed the importance of a 5-methylthiazol-2-amine pharmacophore and resulted in an HQ461 derivate with improved potency. Our studies reveal a new molecular glue that recruits its target protein directly to DDB1 to bypass the requirement of a substrate-specific receptor, presenting a new strategy for targeted protein degradation.

Anal swab findings in an infant with COVID-19.

INTRODUCTION: The transmission pathways of coronavirus disease 2019 (COVID-19) remain not completely clear. In this case study the test for the novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in pharyngeal swab and anal swab were compared. CASE PRESENTATION: A 3-month-old girl was admitted to our hospital with COVID-19. Her parents had both been diagnosed with COVID-19. The results of pharyngeal swab and anal swab of the little girl were recorded and compared during the course of the disease. The oropharyngeal specimen showed negative result for SARS-CoV-2 on the 14th day after onset of the illness. However, the anal swab was still positive for SARS-CoV-2 on the 28th day after the onset of the illness. CONCLUSION: The possibility of fecal-oral transmission of COVID-19 should be assessed. Personal hygiene during home quarantine merits considerable attention.

Alpha-kinase 1 is a cytosolic innate immune receptor for bacterial ADP-heptose.

Immune recognition of pathogen-associated molecular patterns (PAMPs) by pattern recognition receptors often activates proinflammatory NF-κB signalling1. Recent studies indicate that the bacterial metabolite D-glycero-ß-D-manno-heptose 1,7-bisphosphate (HBP) can activate NF-κB signalling in host cytosol2-4, but it is unclear whether HBP is a genuine PAMP and the cognate pattern recognition receptor has not been identified. Here we combined a transposon screen in Yersinia pseudotuberculosis with biochemical analyses and identified ADP-ß-D-manno-heptose (ADP-Hep), which mediates type III secretion system-dependent NF-κB activation and cytokine expression. ADP-Hep, but not other heptose metabolites, could enter host cytosol to activate NF-κB. A CRISPR-Cas9 screen showed that activation of NF-κB by ADP-Hep involves an ALPK1 (alpha-kinase 1)-TIFA (TRAF-interacting protein with forkhead-associated domain) axis. ADP-Hep directly binds the N-terminal domain of ALPK1, stimulating its kinase domain to phosphorylate and activate TIFA. The crystal structure of the N-terminal domain of ALPK1 and ADP-Hep in complex revealed the atomic mechanism of this ligand-receptor recognition process. HBP was transformed by host adenylyltransferases into ADP-heptose 7-P, which could activate ALPK1 to a lesser extent than ADP-Hep. ADP-Hep (but not HBP) alone or during bacterial infection induced Alpk1-dependent inflammation in mice. Our findings identify ALPK1 and ADP-Hep as a pattern recognition receptor and an effective immunomodulator, respectively.

Diaqua-[(E)-2-(2-oxidobenzyl-idene-amino)-2-phenyl-acetato]zinc(II) dimethyl sulfoxide monosolvate.

In the title compound, [Zn(C(15)H(11)NO(3))(H(2)O)(2)]·C(2)H(6)OS, the Zn(II) ion is coordinated by two O atoms and one N atom of the deprotonated chelate ligand and two water mol-ecules in a distorted trigonal bipyramidal coordination environment. A linear supra-molecular structure built from O-H⋯O hydrogen bonds runs parallel to [100].