Discovery and Characterization of RGH-122, a Potent, Selective, and Orally Bioavailable V1a Receptor Antagonist.

The V1a receptor is a major contributor in mediating the social and emotional effects of arginine-vasopressin (AVP); therefore it represents a promising target in the treatment of several neuropsychiatric conditions. The aim of this research was to design and synthesize novel and selective V1a antagonists with improved in vitro and in vivo profiles. Through optimization and detailed SAR studies, we developed low nanomolar antagonists, and further characterizations led to the discovery of the clinical candidate compound 43 (RGH-122). The CNS activity of the compound was determined in a 3-chamber social preference test of autism in which RGH-122 successfully enhanced social preference with the lowest effective dose of 1.5 mg/kg.

Inequalities in Diabetes Mortality Between Microregions in Hungary.

Objectives: Regional differences in diabetes mortality are high in Hungary. In our cross-sectional study, we aim to reveal the drivers of the inequalities in diabetes mortality across the 197 microregions of Hungary. To account for the influence of changes in healthcare and social conditions, we compared two periods (2009-12 and 2013-16). Methods: Traditional and re-conceptualized deprivation- and healthcare provison measures were used in OLS regression models. Results: Microregions with a high proportion of population living in "service deserts," especially in regard to the lack of grocery stores, suffer the highest rates of diabetes mortality. Alcohol-related mortality has been proven to be a similarly and surprisingly strong predictor of diabetes mortality. Conclusion: Food provision should be supported in areas characterized by low service density, and alcohol policy should be strengthened and targeted.

Classification-based deep neural network vs mixture density network models for insulin sensitivity prediction problem.

Model-based glycemic control (GC) protocols are used to treat stress-induced hyperglycaemia in intensive care units (ICUs). The STAR (Stochastic-TARgeted) glycemic control protocol - used in clinical practice in several ICUs in New Zealand, Hungary, Belgium, and Malaysia - is a model-based GC protocol using a patient-specific, model-based insulin sensitivity to describe the patient's actual state. Two neural network based methods are defined in this study to predict the patient's insulin sensitivity parameter: a classification deep neural network and a Mixture Density Network based method. Treatment data from three different patient cohorts are used to train the network models. Accuracy of neural network predictions are compared with the current model- based predictions used to guide care. The prediction accuracy was found to be the same or better than the reference. The authors suggest that these methods may be a promising alternative in model-based clinical treatment for patient state prediction. Still, more research is needed to validate these findings, including in-silico simulations and clinical validation trials.

Engineering nanowires in bacteria to elucidate electron transport structural-functional relationships.

Bacterial pilin nanowires are protein complexes, suggested to possess electroactive capabilities forming part of the cells' bioenergetic programming. Their role is thought to be linked to facilitating electron transfer between cells and the external environment to permit metabolism and cell-to-cell communication. There is a significant debate, with varying hypotheses as to the nature of the proteins currently lying between type-IV pilin-based nanowires and polymerised cytochrome-based filaments. Importantly, to date, there is a very limited structure-function analysis of these structures within whole bacteria. In this work, we engineered Cupriavidus necator H16, a model autotrophic organism to express differing aromatic modifications of type-IV pilus proteins to establish structure-function relationships on conductivity and the effects this has on pili structure. This was achieved via a combination of high-resolution PeakForce tunnelling atomic force microscopy (PeakForce TUNA™) technology, alongside conventional electrochemical approaches enabling the elucidation of conductive nanowires emanating from whole bacterial cells. This work is the first example of functional type-IV pili protein nanowires produced under aerobic conditions using a Cupriavidus necator chassis. This work has far-reaching consequences in understanding the basis of bio-electrical communication between cells and with their external environment.

Metabolic engineering of Cupriavidus necator H16 for heterotrophic and autotrophic production of 3-hydroxypropionic acid.

3-Hydroxypropionate (3-HP) is a versatile compound for chemical synthesis and a potential building block for biodegradable polymers. Cupriavidus necator H16, a facultative chemolithoautotroph, is an attractive production chassis and has been extensively studied as a model organism for biopolymer production. Here, we engineered C. necator H16 for 3-HP biosynthesis from its central metabolism. Wild type C. necator H16 can use 3-HP as a carbon source, a highly undesirable trait for a 3-HP production chassis. However, deletion of its three (methyl-)malonate semialdehyde dehydrogenases (mmsA1, mmsA2 and mmsA3) resulted in a strain that cannot grow on 3-HP as the sole carbon source, and this strain was selected as our production host. A stepwise approach was used to construct pathways for 3-HP production via ß-alanine. Two additional gene deletion targets were identified during the pathway construction process. Deletion of the 3-hydroxypropionate dehydrogenase, encoded by hpdH, prevented the re-consumption of the 3-HP produced by our engineered strains, while deletion of gdhA1, annotated as a glutamate dehydrogenase, prevented the utilization of aspartate as a carbon source, one of the key pathway intermediates. The final strain carrying these deletions was able to produce up to 8 mM 3-HP heterotrophically. Furthermore, an engineered strain was able to produce 0.5 mM 3-HP under autotrophic conditions, using CO2 as sole carbon source. These results form the basis for establishing C. necator H16 as an efficient platform for the production of 3-HP and 3-HP-containing polymers.

OGG1 Inhibition Reduces Acinar Cell Injury in a Mouse Model of Acute Pancreatitis.

Acute pancreatitis (AP) is a potentially life-threatening gastrointestinal disease with a complex pathology including oxidative stress. Oxidative stress triggers oxidative DNA lesions such as formation of 7,8-dihydro-8-oxo-2'-oxoguanine (8-oxoG) and also causes DNA strand breaks. DNA breaks can activate the nuclear enzyme poly(ADP-ribose) polymerase 1 (PARP1) which contributes to AP pathology. 8-oxoG is recognized by 8-oxoG glycosylase 1 (OGG1) resulting in the removal of 8-oxoG from DNA as an initial step of base excision repair. Since OGG1 also possesses a DNA nicking activity, OGG1 activation may also trigger PARP1 activation. In the present study we investigated the role played by OGG1 in AP. We found that the OGG1 inhibitor compound TH5487 reduced edema formation, inflammatory cell migration and necrosis in a cerulein-induced AP model in mice. Moreover, TH5487 caused 8-oxoG accumulation and reduced tissue poly(ADP-ribose) levels. Consistent with the indirect PARP inhibitory effect, TH5487 shifted necrotic cell death (LDH release and Sytox green uptake) towards apoptosis (caspase activity) in isolated pancreatic acinar cells. In the in vivo AP model, TH5487 treatment suppressed the expression of various cytokine and chemokine mRNAs such as those of TNF, IL-1ß, IL1ra, IL6, IL16, IL23, CSF, CCL2, CCL4, CCL12, IL10 and TREM as measured with a cytokine array and verified by RT-qPCR. As a potential mechanism underlying the transcriptional inhibitory effect of the OGG1 inhibitor we showed that while 8-oxoG accumulation in the DNA facilitates NF-κB binding to its consensus sequence, when OGG1 is inhibited, target site occupancy of NF-κB is impaired. In summary, OGG1 inhibition provides protection from tissue injury in AP and these effects are likely due to interference with the PARP1 and NF-κB activation pathways.

Synthetic Biology Toolbox, Including a Single-Plasmid CRISPR-Cas9 System to Biologically Engineer the Electrogenic, Metal-Resistant Bacterium Cupriavidus metallidurans CH34.

Cupriavidus metallidurans CH34 exhibits extraordinary metabolic versatility, including chemolithoautotrophic growth; degradation of BTEX (benzene, toluene, ethylbenzene, xylene); high resistance to numerous metals; biomineralization of gold, platinum, silver, and uranium; and accumulation of polyhydroxybutyrate (PHB). These qualities make it a valuable host for biotechnological applications such as bioremediation, bioprocessing, and the generation of bioelectricity in microbial fuel cells (MFCs). However, the lack of genetic tools for strain development and studying its fundamental physiology represents a bottleneck to boosting its commercial applications. In this study, inducible and constitutive promoter libraries were built and characterized, providing the first comprehensive list of biological parts that can be used to regulate protein expression and optimize the CRISPR-Cas9 genome editing tools for this host. A single-plasmid CRISPR-Cas9 system that can be delivered by both conjugation and electroporation was developed, and its efficiency was demonstrated by successfully targeting the pyrE locus. The CRISPR-Cas9 system was next used to target candidate genes encoding type IV pili, hypothesized by us to be involved in extracellular electron transfer (EET) in this organism. Single and double deletion strains (ΔpilA, ΔpilE, and ΔpilAE) were successfully generated. Additionally, the CRISPR-Cas9 tool was validated for constructing genomic insertions (ΔpilAE::gfp and ΔpilAE::λPrgfp). Finally, as type IV pili are believed to play an important role in extracellular electron transfer to solid surfaces, C. metallidurans CH34 ΔpilAE was further studied by means of cyclic voltammetry using disposable screen-printed carbon electrodes. Under these conditions, we demonstrated that C. metallidurans CH34 could generate extracellular currents; however, no difference in the intensity of the current peaks was found in the ΔpilAE double deletion strain when compared to the wild type. This finding suggests that the deleted type IV pili candidate genes are not involved in extracellular electron transfer under these conditions. Nevertheless, these experiments revealed the presence of different redox centers likely to be involved in both mediated electron transfer (MET) and direct electron transfer (DET), the first interpretation of extracellular electron transfer mechanisms in C. metallidurans CH34.

Significance of autoantibody assays in systemic lupus erythematosus / Az autoantitest-vizsgálatok jelentosége szisztémás lupus erythematosusban

Systemic lupus erythematosus (SLE) is a systemic autoimmune disease characterized by pathological immune complexes and various autoantibodies. Detectable pathological autoantibodies produced by plasma cells differentiated from B cells play a significant role in the establishment of clinical diagnosis, classification, and differential diagnosis as well as assessing the disease activity during patients' follow-up. Autoantibody determination is very important in

The hEag1 K+ Channel Inhibitor Astemizole Stimulates Ca2+ Deposition in SaOS-2 and MG-63 Osteosarcoma Cultures.

The hEag1 (Kv10.1) K+ channel is normally found in the brain, but it is ectopically expressed in tumor cells, including osteosarcoma. Based on the pivotal role of ion channels in osteogenesis, we tested whether pharmacological modulation of hEag1 may affect osteogenic differentiation of osteosarcoma cell lines. Using molecular biology (RT-PCR), electrophysiology (patch-clamp) and pharmacology (astemizole sensitivity, IC50 = 0.135 µM) we demonstrated that SaOS-2 osteosarcoma cells also express hEag1 channels. SaOS-2 cells also express to KCa1.1 K+ channels as shown by mRNA expression and paxilline sensitivity of the current. The inhibition of hEag1 (2 µM astemizole) or KCa1.1 (1 mM TEA) alone did not induce Ca2+ deposition in SaOS-2 cultures, however, these inhibitors, at identical concentrations, increased Ca2+ deposition evoked by the classical or pathological (inorganic phosphate, Pi) induction pathway without causing cytotoxicity, as reported by three completer assays (LDH release, MTT assay and SRB protein assay). We observed a similar effect of astemizole on Ca2+ deposition in MG-63 osteosarcoma cultures as well. We propose that the increase in the osteogenic stimuli-induced mineral matrix formation of osteosarcoma cell lines by inhibiting hEag1 may be a useful tool to drive terminal differentiation of osteosarcoma.

Low α2-Plasmin Inhibitor Antigen Levels on Admission Are Associated With More Severe Stroke and Unfavorable Outcomes in Acute Ischemic Stroke Patients Treated With Intravenous Thrombolysis.

Background: Intravenous administration of recombinant tissue plasminogen activator (rt-PA) fails to succeed in a subset of acute ischemic stroke (AIS) patients, while in approximately 6-8% of cases intracerebral hemorrhage (ICH) occurs as side effect. Objective: Here, we aimed to investigate α2-plasmin inhibitor (α2-PI) levels during thrombolysis and to find out whether they predict therapy outcomes in AIS patients. Patients/Methods: In this prospective, observational study, blood samples of 421 AIS patients, all undergoing i.v. thrombolysis by rt-PA within 4.5 h of their symptom onset, were taken before and 24 h after thrombolysis. In a subset of patients (n = 131), blood was also obtained immediately post-lysis. α2-PI activity and antigen levels were measured by chromogenic assay and an in-house ELISA detecting all forms of α2-PI. α2-PI Arg6Trp polymorphism was identified in all patients. Stroke severity was determined by NIHSS on admission and day 7. Therapy-associated ICH was classified according to ECASSII. Long-term outcomes were defined at 3 months post-event by the modified Rankin Scale (mRS). Results: Median α2-PI activity and antigen levels showed a significant drop immediately post-lysis and increased to subnormal levels at 24 h post-event. Admission α2-PI levels showed a significant negative stepwise association with stroke severity. Patients with favorable long-term outcomes (mRS 0-1) had significantly higher admission α2-PI antigen levels (median:61.6 [IQR:55.9-70.5] mg/L) as compared to patients with poor outcomes (mRS 2-5: median:59.7 [IQR:54.5-69.1] and mRS 6: median:56.0 [IQR:48.5-61.0] mg/L, p < 0.001). In a Kaplan-Meier survival analysis, patients with an α2-PI antigen in the highest quartile on admission showed significantly better long-term survival as compared to those with α2-PI antigen in the lowest quartile (HR: 4.54; 95%CI:1.92-10.8, p < 0.001); however, in a multivariate analysis, a low admission α2-PI antigen did not prove to be an independent risk factor of poor long-term outcomes. In patients with therapy-related ICH (n = 34), admission α2-PI antigen levels were significantly, but only marginally, lower as compared to those without hemorrhage. Conclusions: Low α2-PI antigen levels on admission were associated with more severe strokes and poor long-term outcomes in this cohort. Our results suggest that in case of more severe strokes, α2-PI may be involved in the limited efficacy of rt-PA thrombolysis.

Oxygen-Tolerant RAFT Polymerization Initiated by Living Bacteria.

Living organisms can synthesize a wide range of macromolecules from a small set of natural building blocks, yet there is potential for even greater materials diversity by exploiting biochemical processes to convert unnatural feedstocks into new abiotic polymers. Ultimately, the synthesis of these polymers in situ might aid the coupling of organisms with synthetic matrices, and the generation of biohybrids or engineered living materials. The key step in biohybrid materials preparation is to harness the relevant biological pathways to produce synthetic polymers with predictable molar masses and defined architectures under ambient conditions. Accordingly, we report an aqueous, oxygen-tolerant RAFT polymerization platform based on a modified Fenton reaction, which is initiated by Cupriavidus metallidurans CH34, a bacterial species with iron-reducing capabilities. We show the synthesis of a range of water-soluble polymers under normoxic conditions, with control over the molar mass distribution, and also the production of block copolymer nanoparticles via polymerization-induced self-assembly. Finally, we highlight the benefits of using a bacterial initiation system by recycling the cells for multiple polymerizations. Overall, our method represents a highly versatile approach to producing well-defined polymeric materials within a hybrid natural-synthetic polymerization platform and in engineered living materials with properties beyond those of biotic macromolecules.

Tricetin Reduces Inflammation and Acinar Cell Injury in Cerulein-Induced Acute Pancreatitis: The Role of Oxidative Stress-Induced DNA Damage Signaling.

Acute pancreatitis (AP) poses a worldwide challenge due to the growing incidence and its potentially life-threatening course and complications. Specific targeted therapies are not available, prompting the identification of new pathways and novel therapeutic approaches. Flavonoids comprise several groups of biologically active compounds with wide-ranging effects. The flavone compound, tricetin (TCT), has not yet been investigated in detail but sporadic reports indicate diverse biological activities. In the current study, we evaluated the potential protective effects of TCT in AP. TCT (30 µM) protected isolated primary murine acinar cells from the cytotoxic effects of cerulein, a cholecystokinin analog peptide. The protective effects of TCT were observed in a general viability assay (calcein ester hydrolysis), in an apoptosis assay (caspase activity), and in necrosis assays (propidium iodide uptake and lactate dehydrogenase release). The effects of TCT were not related to its potential antioxidant effects, as TCT did not protect against H2O2-induced acinar cell death despite possessing radical scavenging activity. Cerulein-induced expression of IL1ß, IL6, and matrix metalloproteinase 2 and activation of nuclear factor-κB (NFκB) were reduced by 30 µM TCT. In vivo experiments confirmed the protective effect of TCT in a mouse model of cerulein-induced AP. TCT suppressed edema formation and apoptosis in the pancreas and reduced lipase and amylase levels in the serum. Moreover, TCT inhibited interleukin-1ß (IL1ß), interleukin-6 (IL6), and tumor necrosis factor-α (TNFα) expression in the pancreas and reduced the activation of the oxidative DNA damage sensor enzyme poly(ADP-ribose) polymerase-1 (PARP-1). Our data indicate that TCT can be a potential treatment option for AP.

Drug consumption of suspected drug-influenced drivers in Hungary (2016-2018).

The hazard caused by driving under the influence of drugs (DUID) is determined by the time of consumption, dose and biological effects of a substance, as well as by synergistic drug interactions after multi-drug use. The aim of this work was to investigate the prevalence and pattern of psychoactive substance use of suspected DUID drivers and to present the advantages and disadvantages of the system currently used for determination of impairment in Hungary. Blood and urine samples, collected between 2016 and 2018, were taken from 2369 drivers with a positivity rate of 95% for at least one substance. Classical illicit drugs were detected in 76-87%, prescription medications in 9-15%, stimulant New Psychoactive Substances (sNPS) in 3-8%, and synthetic cannabinoids (SCs) in 20-22% of the positive samples. The most frequent substances according to substance groups were: classical illicit drugs: cannabis (n = 1240), amphetamine and methamphetamine (AM/MA) (n = 753), MDMA (n = 196), and cocaine (n = 180), medicines: alprazolam (n = 188) and clonazepam (n = 83), sNPS: N-ethyl-hexedrone (n = 115), SCs: 5 F-MDMB-PINACA (n = 267), AMB-FUBINACA (n = 92) and ADB-FUBINACA (n = 90). The median age of classical illicit drugs users was 29 years, prescription medicine users were 33 years old, sNPS users were 28 years, and SC users were 26 years old. Compared to the previous two years, we found pronounced changes in the ratio of sNPS (14% decrease) and SC users (10% increase), and in the pattern of NPS consumption. The ratio of multi-drug use varied between 38% and 50%. 69% of drivers tested positive were deemed impaired. Impairment was determined according to impairment limits (80-82%), multi-drug use (12-13%), and the result of medical investigation when a single active substance with no set impairment limit was detected in the blood (6-8%). The results of medical investigations may be uncertain due to the long time delay between arrest and clinical examination and to the structure of medical investigations created for determination of alcoholic impairment. In conclusion, a revision of the current medical investigation protocol is warranted to standardize clinical symptom scores that better correlate with driving impairment.

Epigallocatechine-3-gallate Inhibits the Adipogenesis of Human Mesenchymal Stem Cells via the Regulation of Protein Phosphatase-2A and Myosin Phosphatase.

Epigallocatechin-3-gallate (EGCG) has widespread effects on adipocyte development. However, the molecular mechanisms of EGCG are not fully understood. We investigate the adipogenic differentiation of human-derived mesenchymal stem cells, including lipid deposition and changes in the expression and phosphorylation of key transcription factors, myosin, protein phosphatase-2A (PP2A), and myosin phosphatase (MP). On day 6 of adipogenic differentiation, EGCG (1-20 µM) suppressed lipid droplet formation, which was counteracted by an EGCG-binding peptide for the 67 kDa laminin receptor (67LR), suggesting that EGCG acts via 67LR. EGCG decreased the phosphorylation of CCAAT-enhancer-binding protein beta via the activation of PP2A in a protein kinase A (PKA)-dependent manner, leading to the partial suppression of peroxisome proliferator-activated receptor gamma (PPARγ) and adiponectin expression. Differentiated cells exhibited a rounded shape, cortical actin filaments, and lipid accumulation. The EGCG treatment induced cell elongation, stress fiber formation, and less lipid accumulation. These effects were accompanied by the degradation of the MP target subunit-1 and increased the phosphorylation of the 20 kDa myosin light chain. Our results suggest that EGCG acts as an agonist of 67LR to inhibit adipogenesis via the activation of PP2A and suppression of MP. These events are coupled with the decreased phosphorylation and expression levels of adipogenic transcription factors and changes in cell shape, culminating in curtailed adipogenesis.

Combinatory lung tumor inhibition by myo-inositol and iloprost/rapamycin: association with immunomodulation.

Although both preclinical and clinical studies have suggested that myo-inositol (MI) may be a safe and effective lung cancer chemopreventive agent, its efficacy is moderate. To test whether the chemopreventive agents iloprost (IL) or rapamycin enhance the lung tumor inhibitory effects of MI, A/J mice were treated with the tobacco smoke carcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) and, beginning one week after the end of NNK treatment, given MI, IL, rapamycin, MI + IL or MI + rapamycin for 17 weeks. Analyses of the number and size of tumors on the surface of the lung have indicated that MI, IL, rapamycin, MI + IL and MI + rapamycin reduced the multiplicity of NNK-induced lung tumors by 41, 34, 46, 79 and 67%, respectively, and larger tumors (lung tumors with a diameter of 1-2 or >2 mm) were absent in the MI + IL and MI + rapamycin groups. These results clearly indicated that MI + IL and MI + rapamycin are more effective than MI alone in inhibiting the formation and growth of lung tumors. Assessment of the immunomodulatory effects of the drugs showed that whereas MI + rapamycin and MI + IL increased the infiltration of lung tumors by CD4+ and CD8+ T cells, MI + rapamycin reduced the expression of the immune checkpoint protein programmed-death ligand-1 (PD-L1). Moreover, all treatments, except IL, increased apoptosis, whereas cell proliferation was markedly suppressed in all treated groups. In summary, these results suggest that IL and rapamycin could enhance the efficacy of MI in lung cancer chemoprevention trials.

The multitargeted receptor tyrosine kinase inhibitor sunitinib induces resistance of HER2 positive breast cancer cells to trastuzumab-mediated ADCC.

Despite recent advances in the development of novel personalized therapies, breast cancer continues to challenge physicians with resistance to various advanced therapies. The anticancer action of the anti-HER2 antibody, trastuzumab, involves antibody-dependent cell-mediated cytotoxicity (ADCC) by natural killer (NK) cells. Here, we report a repurposing screen of 774 clinically used compounds on NK-cell + trastuzumab-induced killing of JIMT-1 breast cancer cells. Using a calcein-based high-content screening (HCS) assay for the image-based quantitation of ADCC that we have developed and optimized for this purpose, we have found that the multitargeted tyrosine kinase inhibitor sunitinib inhibits ADCC in this model. The cytoprotective effect of sunitinib was also confirmed with two other assays (lactate dehydrogenase release, and electric cell substrate impedance sensing, ECIS). The drug suppressed NK cell activation as indicated by reduced granzyme B deposition on to the target cells and inhibition of interferon-γ production by the NK cells. Moreover, sunitinib induced downregulation of HER2 on the target cells' surface, changed the morphology and increased adherence of the target cells. Moreover, sunitinib also triggered the autophagy pathway (speckled LC3b) as an additional potential underlying mechanism of the cytoprotective effect of the drug. Sunitinib-induced ADCC resistance has been confirmed in a 3D tumor model revealing the prevention of apoptotic cell death (Annexin V staining) in JIMT-1 spheroids co-incubated with NK cells and trastuzumab. In summary, our HCS assay may be suitable for the facile identification of ADCC boosting compounds. Our data urge caution concerning potential combinations of ADCC-based immunotherapies and sunitinib.

Clinical symptoms and blood concentration of new psychoactive substances (NPS) in intoxicated and hospitalized patients in the Budapest region of Hungary (2018-19).

BACKGROUND: New Psychoactive Substances (NPS) impose a new challenge on the legal and health care system, yet, there is little information available about how new substances spread based on hospitalization of intoxicated patients. The aims of this study were: (i) to investigate the frequency of NPS among suspected drug intoxicated patients, (ii) to study the connection between blood concentration and clinical symptoms, (iii) to determine their half-life with a time-series blood sampling protocol. METHODS: During the observation period, 116 suspected drug intoxicated patients were sampled. The samples were analyzed for alcohol, 20 classical illicit and licit drugs, and for 78 NPS. Clinical symptoms were registered on-site (by the Emergency Medical Services) and (also) at hospital admittance. RESULTS: NPS were detected in 51 patients of which cathinones were found in 4, the synthetic cannabinoids (SCs) 5 F-MDMB-PINACA and 5 F-MDMB-PICA in 23-23, and CUMYL-CH-MEGACLONE in 2 cases. Poison severity scores (PSS) showed mild to moderate intoxications overall. Connection between blood concentration and severity of clinical symptoms were inconclusive. The calculated half-life of 5 F-MDMB-PINACA and 5 F-MDMB-PICA was 2.50 and 2.68 h, respectively. CONCLUSION: The ratio of SCs among the selected intoxicated patients was higher than expected from seizure data which could be the consequence of targeted patient selection. The clinical symptoms and the severity of intoxication cannot be characterized simply by NPS blood levels. The short half-life of SCs can explain the relatively rapid consolidation of intoxication symptoms.HighlightsIn the Budapest region, the majority of hospitalized NPS intoxications was caused by the synthetic cannabinoids 5F-MDMB-PINACA and 5F-MDMB-PICA in 2018-19.No correlation between blood concentration and symptoms severity could be established.The clinical symptoms of synthetic cannabinoid users improved quickly and no ICU treatment was necessary.The half-life of 5F-MDMB-PINACA and 5F-MDMB-PICA was proved to be 2.50 hours and 2.68 hours, respectively.

Biosynthesis of Poly(3HB-co-3HP) with Variable Monomer Composition in Recombinant Cupriavidus necator H16.

Polyhydroxyalkanoates are attractive alternatives to traditional plastics. However, although polyhydroxybutyrate (PHB) is produced in large quantities by Cupriavidus necator H16, its properties are far from ideal for the manufacture of plastic products. These properties may be improved through its coproduction with 3-hydroxypropionate (3HP), which leads to the formation of the copolymer poly(3-hydroxybutyrate-co-3-hydroxypropionate) (poly(3HB-co-3HP). To achieve this, a pathway was designed to enable C. necator H16 to convert ß-alanine to 3HP. The initial low levels of incorporation of 3HP into the copolymer were overcome by the overproduction of the native propionyl-CoA transferase together with PHA synthase from Chromobacterium sp. USM2. Following optimization of 3HP incorporation into the copolymer, the molar fraction of 3HP could be controlled by cultivation in medium containing different concentrations of ß-alanine. Between 0 and 80 mol % 3HP could be achieved. Further supplementation with 2 mM cysteine increased the maximum 3HP molar fraction to 89%. Additionally, the effect of deletions of the phaA and phaB1 genes of the phaCAB operon on 3HP molar fraction were investigated. A phaAB1 double knockout resulted in a copolymer containing 91 mol % 3HP without the need for cysteine supplementation.

The pMTL70000 modular, plasmid vector series for strain engineering in Cupriavidus necator H16.

Cupriavidus necator H16 can convert CO2 into industrial chemicals and fuels. To facilitate its engineering, we designed, built and tested the pMTL70000 modular plasmids comprising standardised Cupriavidus and E. coli replicons, selectable markers and application specific modules. Plasmids were characterised in terms of transmissibility, stability, copy number and compatibility.

Synthesis and Characterization of New V1A Antagonist Compounds: The Separation of Four Atropisomeric Stereoisomers.

A new class of selective vasopressin receptor 1A (V1A) antagonists was identified, where "methyl-scan" was performed around the benzene ring of the 5-hydroxy-triazolobenzazepine core. This led to the synthesis of two 10-methyl derivatives, each possessing a chiral axis and a stereogenic center. The four atropisomeric stereoisomers (involving two enantiomer pairs and atropisomeric diastereomers) could be successfully isolated and spectroscopically characterized. According to the in vitro pharmacological profiles of the compounds, the human V1A receptor has a strong preference toward the isomers having an aR axial chirality, the most active isomer being the aR,5S isomer. Furthermore, the structure-activity relationships obtained for the isomers and for the newly synthesized analogues could be tentatively explained by an in silico study.