Substitution of terminal amide with 1H-1,2,3-triazole: Identification of unexpected class of potent antibacterial agents

3-Methoxybenzamide (3-MBA) derivatives have been identified as novel class of potent antibacterial agents targeting the bacterial cell division protein FtsZ. As one of isosteres for the amide group, 1,2,3-triazole can mimic the topological and electronic features of the amide, which has gained increasing attention in drug discovery. Based on these considerations, we prepared a series of 1H-1,2,3-triazole-containing 3-MBA analogues via isosteric replacement of the terminal amide with triazole, which had increased antibacterial activity. This study demonstrated the possibility of developing the 1H-1,2,3-triazole group as a terminal amide-mimetic element which was capable of both keeping and modulating amide-related bioactivity. Surprisingly, a different action mode of these new 1H-1,2,3-triazole-containing analogues was observed, which could open new opportunities for the development of antibacterial agents.     

Zebularine Induces Long-Term Survival of Pancreatic Islet Allotransplants in Streptozotocin Treated Diabetic Rats

Background

Coping with the immune rejection of allotransplants or autologous cells in patients with an active sensitization towards their autoantigens and autoimmunity presently necessitates life-long immune suppressive therapy acting on the immune system as a whole, which makes the patients vulnerable to infections and increases their risk of developing cancer. New technologies to induce antigen selective long-lasting immunosuppression or immune tolerance are therefore much needed.

Methodology/Principal Findings

The DNA demethylating agent Zebularine, previously demonstrated to induce expression of the genes for the immunosuppressive enzymes indolamine-2,3-deoxygenase-1 (IDO1) and kynureninase of the kynurenine pathway, is tested for capacity to suppress rejection of allotransplants. Allogeneic pancreatic islets from Lewis rats were transplanted under the kidney capsule of Fischer rats previously made diabetic by a streptozotocin injection (40 mg/kg). One group was treated with Zebularine (225 mg/kg) daily for 14 days from day 6 or 8 after transplantation, and a control group received no further treatment. Survival of the transplants was monitored by blood sugar measurements. Rats, normoglycemic for 90 days after allografting, were subjected to transplant removal by nephrectomy to confirm whether normoglycemia was indeed due to a surviving insulin producing transplant, or alternatively was a result of recovery of pancreatic insulin production in some toxin-treated rats. Of 9 Zebularine treated rats, 4 were still normoglycemic after 90 days and became hyperglycemic after nephrectomy. The mean length of normoglycemia in the Zebularine group was 67±8 days as compared to 14±3 days in 9 controls. Seven rats (2 controls and 5 Zebularine treated) were normoglycemic at 90 days due to pancreatic recovery as demonstrated by failure of nephrectomy to induce hyperglycemia.

Conclusions/Significance

Zebularine treatment in vivo induces a long-lasting suppression of the immune destruction of allogeneic pancreatic islets resulting in protection of allograft function for more than 10 weeks after end of treatment.     

Simultaneous fMRI and EEG during the Multi-Source Interference Task

Background

fMRI and EEG are two non-invasive functional imaging techniques within cognitive neuroscience that have complementary advantages to obtain both temporal and spatial information. The multi-source interference task (MSIT) has been shown to generate robust activations of the dorsal anterior cingulate cortex (dACC) on both a single-subject level and in group averages, in fMRI studies. We have now simultaneously acquired fMRI and EEG during a cognitive interference task.

Materials and Methods

Healthy volunteers were tested in an MRI scanner with simultaneous EEG and fMRI recordings during the MSIT.

Results

The interference condition significantly increased the reaction time in the task. The fMRI analyses revealed activation of dACC as expected, in all subjects at the individual level and in group analyses. The posterior cingulate cortex was de-activated. Simultaneous EEG showed the expected anterior distribution of the interference effect, as it was restricted to frontal sites within a time frame of 80–120 ms post response.

Conclusion

The MSIT task is a reliable task for interference evaluation. fMRI shows robust activation of dACC and by adding EEG, an interference effect can be noticed within a temporal interval of 80–120 ms after the response, as a CRN (correct response negativity). This means that EEG could add a more detailed temporal aspect to the fMRI data from an interference task, and that despite the hostile environment within an MRI scanner, EEG data could be used.     

Deforestation and Benthic Indicators: How Much Vegetation Cover Is Needed to Sustain Healthy Andean Streams?

Deforestation in the tropical Andes is affecting ecological conditions of streams, and determination of how much forest should be retained is a pressing task for conservation, restoration and management strategies. We calculated and analyzed eight benthic metrics (structural, compositional and water quality indices) and a physical-chemical composite index with gradients of vegetation cover to assess the effects of deforestation on macroinvertebrate communities and water quality of 23 streams in southern Ecuadorian Andes. Using a geographical information system (GIS), we quantified vegetation cover at three spatial scales: the entire catchment, the riparian buffer of 30 m width extending the entire stream length, and the local scale defined for a stream reach of 100 m in length and similar buffer width. Macroinvertebrate and water quality metrics had the strongest relationships with vegetation cover at catchment and riparian scales, while vegetation cover did not show any association with the macroinvertebrate metrics at local scale. At catchment scale, the water quality metrics indicate that ecological condition of Andean streams is good when vegetation cover is over 70%. Further, macroinvertebrate community assemblages were more diverse and related in catchments largely covered by native vegetation (>70%). Our results suggest that retaining an important quantity of native vegetation cover within the catchments and a linkage between headwater and riparian forests help to maintain and improve stream biodiversity and water quality in Andean streams affected by deforestation. This research proposes that a strong regulation focused to the management of riparian buffers can be successful when decision making is addressed to conservation/restoration of Andean catchments.     

Evaluation of a series of 2-napthamide derivatives as inhibitors of the drug efflux pump AcrB for the reversal of antimicrobial resistance

Drug efflux pumps confer multidrug resistance to dangerous pathogens which makes these pumps important drug targets. We have synthesised a novel series of compounds based on a 2-naphthamide pharmacore aimed at inhibiting the efflux pumps from Gram-negative bacteria. The archeatypical transporter AcrB from Escherichia coli was used as model efflux pump as AcrB is widely conserved throughout Gram-negative organisms. The compounds were tested for their antibacterial action, ability to potentiate the action of antibiotics and for their ability to inhibit Nile Red efflux by AcrB. None of the compounds were antimicrobial against E. coli wild type cells. Most of the compounds were able to inhibit Nile Red efflux indicating that they are substrates of the AcrB efflux pump. Three compounds were able to synergise with antibiotics and reverse resistance in the resistant phenotype. Compound A3, 4-(isopentyloxy)-2-naphthamide, reduced the MICs of erythromycin and chloramphenicol to the MIC levels of the drug sensitive strain that lacks an efflux pump. A3 had no effect on the MIC of the non-substrate rifampicin indicating that this compound acts specifically through the AcrB efflux pump. A3 also does not act through non-specific mechanisms such as outer membrane or inner membrane permeabilisation and is not cytotoxic against mammalian cell lines. Therefore, we have designed and synthesised a novel chemical compound with great potential to further optimisation as inhibitor of drug efflux pumps.     

Sequestration by IFIT1 Impairs Translation of 2′O-unmethylated Capped RNA

Viruses that generate capped RNA lacking 2′O methylation on the first ribose are severely affected by the antiviral activity of Type I interferons. We used proteome-wide affinity purification coupled to mass spectrometry to identify human and mouse proteins specifically binding to capped RNA with different methylation states. This analysis, complemented with functional validation experiments, revealed that IFIT1 is the sole interferon-induced protein displaying higher affinity for unmethylated than for methylated capped RNA. IFIT1 tethers a species-specific protein complex consisting of other IFITs to RNA. Pulsed stable isotope labelling with amino acids in cell culture coupled to mass spectrometry as well as in vitro competition assays indicate that IFIT1 sequesters 2′O-unmethylated capped RNA and thereby impairs binding of eukaryotic translation initiation factors to 2′O-unmethylated RNA template, which results in inhibition of translation. The specificity of IFIT1 for 2′O-unmethylated RNA serves as potent antiviral mechanism against viruses lacking 2′O-methyltransferase activity and at the same time allows unperturbed progression of the antiviral program in infected cells.     

Studies on the role of β-cell metabolism in the insulinotropic effect of α-ketoisocaproic acid

α-Ketoisocaproic acid has been shown to be apotent insulin secretagogue but the mechanism has not been elucidated. To define the role of β-cell metabolism in the insulinotropic activity of α-ketoisocaproic acid the utilization of glucose and the oxidation of α-ketoisocaproic and isovaleric acid by incubated islets of obese hyperglycemic mice were measured.Glucose metabolism was never enhanced by α-ketoisocaproic acid. The same ¹⁴CO₂ amounts were released from the non-secretagogue [1-¹⁴C]isovaleric acid (10 mM) or from α-keto [2-¹⁴C]isocaproic acid (5–20 mM). Pyruvate (20 mM) did not inhibit α-ketoisocaproic acid-induced insulin secretion in spite of reduction of decarboxylation of α-ketoisocaproic acid by more than 40%.The results indicate that stimulated insulin release in response to α-ketoisocaproic acid is not mediated by an indirect increase in glucose metabolism and further suggest that isovaleryl-CoA and following CoA-esters in α-ketoisocaproic acid degradation are not likely recognized as signals. The possibility, however, remains that enhanced intramitochondrial production of reducing equivalents elicits insulin secretion.     

Constructive Contrasts Between Modeled and Measured Climate Responses Over a Regional Scale

Reducing uncertainty in predictions of regional-scale models depends on meaningful contrasts with field measurements. This paper introduces a two-stage process that works from the premise that an appropriate goal for regional models is to produce reasonable behavior over dominant environmental gradients. We demonstrate two techniques for contrasting models with data, one based on the shape of modeled relationships (functional contrasts) and the other based on an examination of the residuals (residual contrasts) between the model and an empirically derived surface fit to field data. Functional contrasts evaluated the differences between the response of simulated net primary production (NPP) to climate variables and the response observed in field measurements of NPP. Residual contrasts compared deviations of NPP from the empirical surface to identify groupings (for example, vegetation classes, geographic regions) with model deviations different from those of the field data. In all model–data contrasts, we assigned sample weights to field measurements to ensure unbiased representation of the region, and we included both constructive comparisons and formal statistical tests. In general, we learned more from constructive methods designed to reveal structure or pattern in discrepancy than we did from statistical tests designed to falsify models. Although our constructive methods were more subjective and less concise, they succeeded in revealing gaps in our understanding of regional-scale processes that can guide future efforts to reduce scientific uncertainty. This was best illustrated by NPP predictions from the Biome-BGC model, which showed a stronger response to precipitation than apparently operates in the field. In another case, differences revealed in savanna and dry woodlands had insufficient field-data support, suggesting a need for future field studies to improve understanding in this, and other, poorly studied ecosystems. Received 24 September 1999; accepted 20 April 2000.