Structure-based discovery of thiosemicarbazones as SARS-CoV-2 main protease inhibitors.

Aim: Discovery of novel SARS-CoV-2 main protease (Mpro) inhibitors using a structure-based drug discovery strategy. Materials & methods: Virtual screening employing covalent and noncovalent docking was performed to discover Mpro inhibitors, which were subsequently evaluated in biochemical and cellular assays. Results: 91 virtual hits were selected for biochemical assays, and four were confirmed as reversible inhibitors of SARS CoV-2 Mpro with IC50 values of 0.4-3 µM. They were also shown to inhibit SARS-CoV-1 Mpro and human cathepsin L. Molecular dynamics simulations indicated the stability of the Mpro inhibitor complexes and the interaction of ligands at the subsites. Conclusion: This approach led to the discovery of novel thiosemicarbazones as potent SARS-CoV-2 Mpro inhibitors.

Flau-A, a naphthoquinone derivative, is a promising therapeutic candidate against visceral leishmaniasis: A preliminary study.

Visceral leishmaniasis (VL) is a neglected tropical disease found in tropical and subtropical regions in the world. The therapeutics used for the treatment against disease presents problems, mainly related to drug toxicity, route of administration, high cost and/or by emergence of resistant strains. In this context, the search for alternative antileishmanial candidates is desirable. Recently, a naphthoquinone derivative namely 2-(2,3,4-tri-O-acetyl-6-deoxy-ß-L-galactopyranosyloxy)-1,4-naphthoquinone or Flau-A showed an effective in vitro biological action against Leishmania infantum. In the present study, the efficacy of this naphthoquinone derivative was evaluated in an in vivo infection model. BALB/c mice (n = 12 per group) were infected and later received saline or were treated with empty micelles (B/Mic), free Flau-A or it incorporated in Poloxamer 407-based micelles (Flau-A/Mic). The products were administered subcutaneously in the infected animals, which were then euthanized one (n = 6 per group) and 15 (n = 6 per group) days post-therapy, when immunological and parasitological evaluations were performed. Results showed that animals treated with Flau-A or Flau-A/Mic produced significantly higher levels of antileishmanial IFN-γ, IL-12, TNF-α, GM-CSF, nitrite and IgG2a isotype antibody, when compared to data found in the control (saline and B/Mic) groups; which showed significantly higher levels of parasite-specific IL-4, IL-10 and IgG1 antibody. In addition, animals receiving free Flau-A or Flau-A/Mic presented also significant reductions in the parasite load in their spleens, livers, bone marrows and draining lymph nodes, when compared to the controls. A low hepatic and renal toxicity was also found. Overall, Flau-A/Mic showed better immunological and parasitological results, when compared to the use of free molecule. In conclusion, preliminary data suggest that this composition could be considered in future studies as promising therapeutic candidate against VL.

Kinetic Properties of Microbial Exoenzymes Vary With Soil Depth but Have Similar Temperature Sensitivities Through the Soil Profile.

Current knowledge of the mechanisms driving soil organic matter (SOM) turnover and responses to warming is mainly limited to surface soils, although over 50% of global soil carbon is contained in subsoils. Deep soils have different physicochemical properties, nutrient inputs, and microbiomes, which may harbor distinct functional traits and lead to different SOM dynamics and temperature responses. We hypothesized that kinetic and thermal properties of soil exoenzymes, which mediate SOM depolymerization, vary with soil depth, reflecting microbial adaptation to distinct substrate and temperature regimes. We determined the Michaelis-Menten (MM) kinetics of three ubiquitous enzymes involved in carbon (C), nitrogen (N) and phosphorus (P) acquisition at six soil depths down to 90 cm at a temperate forest, and their temperature sensitivity based on Arrhenius/Q 10 and Macromolecular Rate Theory (MMRT) models over six temperatures between 4-50°C. Maximal enzyme velocity (V max) decreased strongly with depth for all enzymes, both on a dry soil mass and a microbial biomass C basis, whereas their affinities increased, indicating adaptation to lower substrate availability. Surprisingly, microbial biomass-specific catalytic efficiencies also decreased with depth, except for the P-acquiring enzyme, indicating distinct nutrient demands at depth relative to microbial abundance. These results suggested that deep soil microbiomes encode enzymes with intrinsically lower turnover and/or produce less enzymes per cell, reflecting distinct life strategies. The relative kinetics between different enzymes also varied with depth, suggesting an increase in relative P demand with depth, or that phosphatases may be involved in C acquisition. V max and catalytic efficiency increased consistently with temperature for all enzymes, leading to overall higher SOM-decomposition potential, but enzyme temperature sensitivity was similar at all depths and between enzymes, based on both Arrhenius/Q 10 and MMRT models. In a few cases, however, temperature affected differently the kinetic properties of distinct enzymes at discrete depths, suggesting that it may alter the relative depolymerization of different compounds. We show that soil exoenzyme kinetics may reflect intrinsic traits of microbiomes adapted to distinct soil depths, although their temperature sensitivity is remarkably uniform. These results improve our understanding of critical mechanisms underlying SOM dynamics and responses to changing temperatures through the soil profile.

Do Drug-likeness Rules Apply to Oral Prodrugs?

This paper describes a comparative analysis of the physicochemical and structural properties of prodrugs and their corresponding drugs with regard to drug-likeness rules. The dataset used in this work was obtained from the DrugBank. Sixty-five pairs of prodrugs/drugs were retrieved and divided into the following categories: carrier-linked to increase hydrophilic character, carrier-linked to increase absorption, and bioprecursors. We compared the physicochemical properties related to drug-likeness between prodrugs and drugs. Our results show that prodrugs do not always follow Lipinski's Rule of 5, especially as we observed 15 prodrugs with more than 10 hydrogen bond acceptors and 18 with a molecular weight greater than 500 Da. This fact highlights the importance of extending Lipinski's rules to encompass other parameters as both strategies (filtering of drug-like chemical libraries and prodrug design) aim to improve the bioavailability of compounds. Therefore, critical reasoning is fundamental to determine whether a structure has drug-like properties or could be considered a potential orally active compound in the drug-design pipeline.

Nitrogen Isotope Fractionation During Archaeal Ammonia Oxidation: Coupled Estimates From Measurements of Residual Ammonium and Accumulated Nitrite.

The naturally occurring nitrogen (N) isotopes, 15N and 14N, exhibit different reaction rates during many microbial N transformation processes, which results in N isotope fractionation. Such isotope effects are critical parameters for interpreting natural stable isotope abundances as proxies for biological process rates in the environment across scales. The kinetic isotope effect of ammonia oxidation (AO) to nitrite (NO2 -), performed by ammonia-oxidizing archaea (AOA) and ammonia-oxidizing bacteria (AOB), is generally ascribed to the enzyme ammonia monooxygenase (AMO), which catalyzes the first step in this process. However, the kinetic isotope effect of AMO, or ε A M O , has been typically determined based on isotope kinetics during product formation (cumulative product, NO2 -) alone, which may have overestimated ε A M O due to possible accumulation of chemical intermediates and alternative sinks of ammonia/ammonium (NH3/NH4 +). Here, we analyzed 15N isotope fractionation during archaeal ammonia oxidation based on both isotopic changes in residual substrate (RS, NH4 +) and cumulative product (CP, NO2 -) pools in pure cultures of the soil strain Nitrososphaera viennensis EN76 and in highly enriched cultures of the marine strain Nitrosopumilus adriaticus NF5, under non-limiting substrate conditions. We obtained ε A M O values of 31.9-33.1‰ for both strains based on RS (δ15NH4 +) and showed that estimates based on CP (δ15NO2 -) give larger isotope fractionation factors by 6-8‰. Complementary analyses showed that, at the end of the growth period, microbial biomass was 15N-enriched (10.1‰), whereas nitrous oxide (N2O) was highly 15N depleted (-38.1‰) relative to the initial substrate. Although we did not determine the isotope effect of NH4 + assimilation (biomass formation) and N2O production by AOA, our results nevertheless show that the discrepancy between ε A M O estimates based on RS and CP might have derived from the incorporation of 15N-enriched residual NH4 + after AMO reaction into microbial biomass and that N2O production did not affect isotope fractionation estimates significantly.

A clioquinol-containing Pluronic® F127 polymeric micelle system is effective in the treatment of visceral leishmaniasis in a murine model.

A clioquinol (ICHQ)-containing Pluronic® F127 polymeric micelle system (ICHQ/Mic) was recently shown to be effective against Leishmania amazonensis infection in a murine model. In the present study, ICHQ/Mic was tested against L. infantum infection. BALB/c mice (n = 12 per group) were infected with L. infantum stationary promastigotes through subcutaneous injection and, 45 days after challenge, received saline or were treated via the subcutaneous route with empty micelles, ICHQ or ICHQ/Mic. In addition, animals were treated with miltefosine by the oral route, as a drug control. Half of the animals were euthanized 1 and 15 days after treatment, aiming to evaluate two endpoints after therapy, when parasitological and immunological parameters were investigated. Results showed that the treatment using miltefosine, ICHQ or ICHQ/Mic induced significantly higher anti-parasite IFN-γ, IL-12, GM-CSF, nitrite and IgG2a isotype antibody levels, which were associated with low IL-4 and IL-10 production. In addition, a higher frequency of IFN-γ and TNF-α-producing CD4+ and CD8+ T-cells was found in these animals. The parasite load was evaluated in distinct organs, and results showed that the treatment using miltefosine, ICHQ or ICHQ/Mic induced significant reductions in organic parasitism in the treated and infected mice. A comparison between the treatments suggested that ICHQ/Mic was the most effective in inducing a highly polarized Th1-type response, as well as reducing the parasite load in significant levels in the treated and infected animals. Data obtained 15 days after treatment suggested maintenance of the immunological and parasitological responses. In conclusion, ICHQ/Mic could be considered in future studies for the treatment of visceral leishmaniasis.

TITLE: Un système à micelles polymériques Pluronic® F127 contenant du clioquinol est efficace pour le traitement de la leishmaniose viscérale dans un modèle murin. ABSTRACT: Un système à micelles polymériques Pluronic® F127 (ICHQ/Mic) contenant du clioquinol (ICHQ) s'est récemment révélé efficace contre l'infection à Leishmania amazonensis dans un modèle murin. Dans la présente étude, l'ICHQ/Mic a été testé contre l'infection à L. infantum. Les souris BALB/c (n = 12 par groupe) ont été infectées par des promastigotes stationnaires de L. infantum par injection sous-cutanée et ont reçu 45 jours après l'épreuve une solution saline ou ont été traitées par voie sous-cutanée avec des micelles vides, ICHQ ou ICHQ/Mic. De plus, les animaux ont été traités avec de la miltefosine par voie orale, comme contrôle médicamenteux. La moitié des animaux ont été euthanasiés 1 et 15 jours après le traitement, dans le but de mesurer deux critères d'évaluation après la thérapie, lorsque les paramètres parasitologiques et immunologiques ont été étudiés. Les résultats ont montré que le traitement par miltefosine, ICHQ ou ICHQ/Mic induisait des niveaux d'anticorps anti-parasite IFN-γ, IL-12, GM-CSF, nitrite et IgG2a significativement plus élevés, associés à de faibles productions d'IL-4 et IL-10. De plus, une fréquence plus élevée de cellules T CD4+ et CD8+ produisant de l'IFN-γ and TNF-α a été trouvée chez ces animaux. La charge parasitaire a été évaluée dans des organes distincts et les résultats ont montré que le traitement utilisant la miltefosine, ICHQ ou ICHQ/Mic induisait des réductions significatives du parasitisme des organes chez les souris traitées et infectées. Une comparaison entre les traitements a suggéré qu'ICHQ/Mic était le plus efficace pour induire une réponse de type Th1 polarisée, ainsi que pour réduire la charge parasitaire à des niveaux significatifs chez les animaux traités et infectés. Les données obtenues 15 jours après le traitement suggèrent le maintien des réponses immunologiques et parasitologiques. En conclusion, ICHQ/Mic pourrait être envisagé dans de futures études pour le traitement contre la leishmaniose viscérale.

Synthesis and cytotoxicity evaluation of glycosidic derivatives of lawsone against breast cancer cell lines.

Breast cancer is the most incident and mortal cancer type in women, with an estimated 2 million new cases expected by 2020 worldwide, with 600,000 deaths. As not all breast cancer types respond to the anti-hormonal therapy, the development of new antineoplastic drugs is necessary. Lawsone (2-hydroxy-1,4-naphtoquinone) is a natural bioactive naphtoquinone displaying a range of activities, with dozens of derivatives described in the literature, including some glycosides possessing antitumor activity. Here, a series of glycosides of lawsone are reported for the first time and all compounds displayed good activity against the SKBR-3 cell line, with IC50 below 10 µM. The most promising derivative was the glycosyl triazole derived from peracetylated d-glucose (11), which showed better cytotoxicity against SKBR-3 (IC50 = 0.78 µM), being the most selective toward this tumoral cell (SI > 20). All compounds described in this work were more active than lawsone, indicating the importance of the carbohydrate and glycosyl triazole moiety for activity.

The phthalimide analogues N-3-hydroxypropylphthalimide and N-carboxymethyl-3-nitrophthalimide exhibit activity in experimental models of inflammatory and neuropathic pain.

BACKGROUND: Phthalimide analogues devoid of the glutarimide moiety exhibit multiple biological activities, thus making them candidates for the treatment of patients with different diseases, including those with inflammatory and painful disorders. In the present study, the activities of five phthalimide analogues devoid of the glutarimide moiety (N-hydroxyphthalimide, N-hydroxymethylphthalimide, N-3-hydroxypropylphthalimide, N-carboxy-3-methylphthalimide, N-carboxymethyl-3-nitrophthalimide) were evaluated in experimental models of acute and chronic inflammatory and neuropathic pain. METHODS: The phthalimide analogues were administered per os (po) in Swiss mice or Wistar rats. Nociceptive response induced by formaldehyde and mechanical allodynia induced by chronic constriction injury (CCI) of the sciatic nerve or intraplantar (ipl) injection of complete Freund's adjuvant (CFA) were used as experimental models of pain. RESULTS: N-carboxymethyl-3-nitrophthalimide (700 mg/kg, -1 h) inhibited the second phase of the nociceptive response induced by the intraplantar injection of formaldehyde in mice. N-3-hidroxypropylphthalimide (546 mg/kg, -1 h) inhibited both phases of the nociceptive response induced by formaldehyde. Treatment of rats with N-carboxymethyl-3-nitrophthalimide (700 mg/kg) or N-3-hydroxypropylphthalimide (546 mg/kg) inhibited the mechanical allodynia induced by CCI of the sciatic nerve or ipl injection of CFA in rats. Intraperitoneal administration of the opioid antagonist naltrexone (10 mg/kg, -1.5 h) attenuated the antinociceptive activity of N-carboxymethyl-3-nitrophthalimide (700 mg/kg) in the model of nociceptive response induced by formaldehyde. CONCLUSIONS: N-3-hydroxypropylphthalimide and N-carboxymethyl-3-nitrophthalimide, two phthalimide analogues devoid of the glutarimide moiety, exhibited activities in different experimental models of pain, including models of chronic inflammatory and neuropathic pain.

Ammonia Oxidation by the Arctic Terrestrial Thaumarchaeote Candidatus Nitrosocosmicus arcticus Is Stimulated by Increasing Temperatures.

Climate change is causing arctic regions to warm disproportionally faster than those at lower latitudes, leading to alterations in carbon and nitrogen cycling, and potentially higher greenhouse gas emissions. It is thus increasingly important to better characterize the microorganisms driving arctic biogeochemical processes and their potential responses to changing conditions. Here, we describe a novel thaumarchaeon enriched from an arctic soil, Candidatus Nitrosocosmicus arcticus strain Kfb, which has been maintained for seven years in stable laboratory enrichment cultures as an aerobic ammonia oxidizer, with ammonium or urea as substrates. Genomic analyses show that this organism harbors all genes involved in ammonia oxidation and in carbon fixation via the 3-hydroxypropionate/4-hydroxybutyrate cycle, characteristic of all AOA, as well as the capability for urea utilization and potentially also for heterotrophic metabolism, similar to other AOA. Ca. N. arcticus oxidizes ammonia optimally between 20 and 28°C, well above average temperatures in its native high arctic environment (-13-4°C). Ammonia oxidation rates were nevertheless much lower than those of most cultivated mesophilic AOA (20-45°C). Intriguingly, we repeatedly observed apparent faster growth rates (based on marker gene counts) at lower temperatures (4-8°C) but without detectable nitrite production. Together with potential metabolisms predicted from its genome content, these observations indicate that Ca. N. arcticus is not a strict chemolithotrophic ammonia oxidizer and add to cumulating evidence for a greater metabolic and physiological versatility of AOA. The physiology of Ca. N. arcticus suggests that increasing temperatures might drastically affect nitrification in arctic soils by stimulating archaeal ammonia oxidation.

Antibacterial activity of synthetic 1,3-bis(aryloxy)propan-2-amines against Gram-positive bacteria.

Synthetic 1,3-bis(aryloxy)propan-2-amines have been shown in previous studies to possess several biological activities, such as antifungal and antiprotozoal. In the present study, we describe the antibacterial activity of new synthetic 1,3-bis(aryloxy)propan-2-amines against Gram-positive pathogens (Streptococcus pyogenes, Enterococcus faecalis and Staphylococcus aureus) including Methicillin-resistant S. aureus strains. Our compounds showed minimal inhibitory concentrations (MIC) in the range of 2.5-10 µg/ml (5.99-28.58 µM), against different bacterial strains. The minimal bactericidal concentrations found were similar to MIC, suggesting a bactericidal mechanism of action of these compounds. Furthermore, possible molecular targets were suggested by chemical similarity search followed by docking approaches. Our compounds are similar to known ligands targeting the cell division protein FtsZ, Quinolone resistance protein norA and the Enoyl-[acyl-carrier-protein] reductase FabI. Taken together, our data show that synthetic 1,3-bis(aryloxy)propan-2-amines are active against Gram-positive bacteria, including multidrug-resistant strains and can be a promising lead in the development of new antibacterial compounds for the treatment of these infections.

New 99mTc-Labeled Digitoxigenin Derivative for Cancer Cell Identification.

In recent years, cardiac glycosides (CGs) have been investigated as potential antiviral and anticancer drugs. Digitoxigenin (DIG) and other CGs have been shown to bind and inhibit Na+/K+-adenosinetriphosphatase (ATPase). Tumor cells show a higher expression rate of the Na+/K+-ATPase protein or a stronger affinity towards the binding of CGs and are therefore more prone to CGs than non-tumor cells. Cancer imaging techniques using radiotracers targeted at specific receptors have yielded successful results. Technetium-99m (99mTc) is one of the radionuclides of choice to radiolabel pharmaceuticals because of its favorable physical and chemical properties along with reasonable costs. Herein, we describe a new Na+/K+-ATPase targeting radiotracer consisting of digitoxigenin and diethylenetriaminepentaacetic acid (DTPA), a bifunctional chelating ligand used to prepare 99mTc-labeled complexes, and its evaluation as an imaging probe. We report the synthesis and characterization of the radiolabeled compound including stability tests, blood clearance, and biodistribution in healthy mice. Additionally, we investigated the binding of the compound to A549 human non-small-cell lung cancer cells and the inhibition of the Na+/K+-ATPase by the labeled compound in vitro. The 99mTc-labeled DTPA-digitoxigenin (99mTc-DTPA-DIG) compound displayed high stability in vitro and in vivo, a fast renal excretion, and a specific binding towards A549 cancer cells in comparison to non-tumor cells. Therefore, 99mTc-DTPA-DIG could potentially be used for non-invasive visualization of tumor lesions by means of scintigraphic imaging.

A Pluronic® F127-based polymeric micelle system containing an antileishmanial molecule is immunotherapeutic and effective in the treatment against Leishmania amazonensis infection.

Clioquinol (5-chloro-7-iodoquinolin-8-ol or ICHQ) was recently showed to presents an in vitro effective antileishmanial action, causing changes in membrane permeability, mitochondrial functionality, and parasite morphology. In the present study, ICHQ was incorporated into a Poloxamer 407-based polymeric micelles system (ICHQ/M), and its antileishmanial activity was in vivo evaluated in L. amazonensis-infected BALB/c mice. Amphotericin B (AmpB) and its liposomal formulation (Ambisome®) were used as controls. Parasitological and immunological evaluations were performed 30 days after the treatment. Results indicated more significant reductions in the average lesion diameter and parasite burden in ICHQ or ICHQ/M-treated mice, which were associated with the development of a polarized Th1 immune response, based on production of high levels of IFN-γ, IL-12, TNF-α, GM-CSF, and antileishmanial IgG2a antibody. Control groups´ mice produced high levels of IL-4, IL-10, and IgG1 isotype antibody. No organic toxicity was found by using ICHQ or ICHQ/M to treat the animals, although those receiving AmpB and Ambisome® have presented higher levels of renal and hepatic damage markers. In conclusion, results suggested that the ICHQ/M composition can be considered as an antileishmanial candidate to be tested against human leishmaniasis.

In vivo antileishmanial efficacy of a naphthoquinone derivate incorporated into a Pluronic® F127-based polymeric micelle system against Leishmania amazonensis infection.

New therapeutic strategies against leishmaniasis are desirable, since the treatment against disease presents problems, such as the toxicity, high cost and/or parasite resistance. As consequence, new antileishmanial compounds are necessary to be identified, as presenting high activity against Leishmania parasites, but low toxicity in mammalian hosts. Flau-A is a naphthoquinone derivative recently showed to presents an in vitro effective action against Leishmania amazonensis and L. infantum species. In the present work, the in vivo efficacy of Flau-A, which was incorporated into a Poloxamer 407-based micelle system, was evaluated in a murine model against L. amazonensis infection. Amphotericin B (AmB) and Ambisome® were used as controls. The animals were infected and later treated with the compounds. Thirty days after the treatment, parasitological and immunological parameters were evaluated. Results showed that AmB, Ambisome®, Flau-A or Flau-A/M-treated animals presented significantly lower average lesion diameter and parasite burden in tissue and organs evaluated, when compared to the control (saline and micelle) groups. Flau-A or Flau-A/M-treated mice were those presenting the most significant reductions in the parasite burden, when compared to the others. These animals developed also a more polarized antileishmanial Th1 immune response, which was based on significantly higher levels of IFN-γ, IL-12, TNF-α, GM-CSF, and parasite-specific IgG2a isotype; associated with low levels of IL-4, IL-10, and IgG1 antibody. The absence of toxicity was found in these animals, although mice receiving AmB have showed high levels of renal and hepatic damage markers. In conclusion, results suggested that the Flau-A/M compound may be considered as a possible therapeutic target to be evaluated against human leishmaniasis.

Drying and Rainfall Shape the Structure and Functioning of Nitrifying Microbial Communities in Riverbed Sediments.

Non-flow periods in fluvial ecosystems are a global phenomenon. Streambed drying and rewetting by sporadic rainfalls could drive considerable changes in the microbial communities that govern stream nitrogen (N) availability at different temporal and spatial scales. We performed a microcosm-based experiment to investigate how dry period duration (DPD) (0, 3, 6, and 9 weeks) and magnitude of sporadic rewetting by rainfall (0, 4, and 21 mm applied at end of dry period) affected stocks of N in riverbed sediments, ammonia-oxidizing bacteria (AOB) and archaea (AOA) and rates of ammonia oxidation (AO), and emissions of nitrous oxide (N2O) to the atmosphere. While ammonium (NH4 +) pool size decreased, nitrate (NO3 -) pool size increased in sediments with progressive drying. Concomitantly, the relative and absolute abundance of AOB and, especially, AOA (assessed by 16S rRNA gene sequencing and quantitative PCR of ammonia monooxygenase genes) increased, despite an apparent decrease of AO rates with drying. An increase of N2O emissions occurred at early drying before substantially dropping until the end of the experiment. Strong rainfall of 21 mm increased AO rates and NH4 + in sediments, whereas modest rainfall of 4 mm triggered a notable increase of N2O fluxes. Interestingly, such responses were detected only after 6 and 9 weeks of drying. Our results demonstrate that progressive drying drives considerable changes in in-stream N cycling and the associated nitrifying microbial communities, and that sporadic rainfall can modulate these effects. Our findings are particularly relevant for N processing and transport in rivers with alternating dry and wet phases - a hydrological scenario expected to become more important in the future.

Unifying the global phylogeny and environmental distribution of ammonia-oxidising archaea based on amoA genes.

Ammonia-oxidising archaea (AOA) are ubiquitous and abundant in nature and play a major role in nitrogen cycling. AOA have been studied intensively based on the amoA gene (encoding ammonia monooxygenase subunit A), making it the most sequenced functional marker gene. Here, based on extensive phylogenetic and meta-data analyses of 33,378 curated archaeal amoA sequences, we define a highly resolved taxonomy and uncover global environmental patterns that challenge many earlier generalisations. Particularly, we show: (i) the global frequency of AOA is extremely uneven, with few clades dominating AOA diversity in most ecosystems; (ii) characterised AOA do not represent most predominant clades in nature, including soils and oceans; (iii) the functional role of the most prevalent environmental AOA clade remains unclear; and (iv) AOA harbour molecular signatures that possibly reflect phenotypic traits. Our work synthesises information from a decade of research and provides the first integrative framework to study AOA in a global context.

1,3-Bis(aryloxy)propan-2-ols as potential antileishmanial agents.

We describe herein the synthesis and antileishmanial activity of 1,3-bis(aryloxy)propan-2-ols. Five compounds (2, 3, 13, 17, and 18) exhibited an effective antileishmanial activity against stationary promastigote forms of Leishmania amazonensis (IC50  < 15.0 µm), and an influence of compound lipophilicity on activity was suggested. Most of the compounds were poorly selective, as they showed toxicity toward murine macrophages, except 17 and 18, which presented good selective indexes (SI ≥ 10.0). The five more active compounds (2, 3, 13, 17, and 18) were selected for the treatment of infected macrophages, and all of them were able to reduce the number of internalized parasites by more than 80%, as well as the number of infected macrophages by more than 70% in at least one of the tested concentrations. Altogether, these results demonstrate the potential of these compounds as new hits of antileishmanial agents and open future possibilities for them to be tested in in vivo studies.

Glycotriazole-peptides derived from the peptide HSP1: synergistic effect of triazole and saccharide rings on the antifungal activity.

This work proposes a strategy that uses solid-phase peptide synthesis associated with copper(I)-catalyzed azide alkyne cycloaddition reaction to promote the glycosylation of an antimicrobial peptide (HSP1) containing a carboxyamidated C-terminus (HSP1-NH2). Two glycotriazole-peptides, namely [p-Glc-trz-G1]HSP1-NH2 and [p-GlcNAc-trz-G1]HSP1-NH2, were prepared using per-O-acetylated azide derivatives of glucose and N-acetylglucosamine in the presence of copper(II) sulfate pentahydrate (CuSO4·5H2O) and sodium ascorbate as a reducing agent. In order to investigate the synergistic action of the carbohydrate motif linked to the triazole-peptide structure, a triazole derivative [trz-G1]HSP1-NH2 was also prepared. A set of biophysical approaches such as DLS, Zeta Potential, SPR and carboxyfluorescein leakage from phospholipid vesicles confirmed higher membrane disruption and lytic activities as well as stronger peptide-LUVs interactions for the glycotriazole-peptides when compared to HSP1-NH2 and to its triazole derivative, which is in accordance with the performed biological assays: whereas HSP1-NH2 presents relatively low and [trz-G1]HSP1-NH2 just moderate fungicidal activity, the glycotriazole-peptides are significantly more effective antifungal agents. In addition, the glycotriazole-peptides and the triazole derivative present strong inhibition effects on ergosterol biosynthesis in Candida albicans, when compared to HSP1-NH2 alone. In conclusion, the increased fungicidal activity of the glycotriazole-peptides seems to be the result of (A) more pronounced membrane-disruptive properties, which is related to the presence of a saccharide ring, together with (B) the inhibition of ergosterol biosynthesis, which seems to be related to the presence of both the monosaccharide and the triazole rings.

Design, synthesis and structure-activity relationship studies of a novel focused library of 2,3,4-substituted oxazolidines with antiproliferative activity against cancer cell lines.

In the present work we describe the synthesis and antiproliferative evaluation of a focused library of 30 novel oxazolidines designed by modification of N-substituent, by ring variation, by alkyl variation or by extension of the structure. It was noted that carbamate and N,O-aminal groups were essential for activity. In general, replacement of the phenyl ring with pyridinyl was not tolerated. However, the introduction of a second phenyl ring with an appropriate spacer at the 3- or 4-position of the first phenyl ring generally enhanced the cytotoxic profile. Among all the prepared compounds, 24 was the most potent compound found in this class, being active on four of five cancer cell lines and it was 5-fold and 10-fold more potent than the lead compounds against HL60 and JURKAT cells, respectively. In addition, it showed relevant activity against MCF-7 and HCT-116 cells, which were resistant to lead. Moreover, 24 showed little antiproliferative activity against VERO, indicating low toxicity to normal cells. Thus, this compound has the potential to be developed as an anticancer agent.

Production of the Cytotoxic Cardenolide Glucoevatromonoside by Semisynthesis and Biotransformation of Evatromonoside by a Digitalis lanata Cell Culture.

Recent studies demonstrate that cardiac glycosides, known to inhibit Na+/K+-ATPase in humans, have increased susceptibility to cancer cells that can be used in tumor therapy. One of the most promising candidates identified so far is glucoevatromonoside, which can be isolated from the endangered species Digitalis mariana ssp. heywoodii. Due to its complex structure, glucoevatromonoside cannot be obtained economically by total chemical synthesis. Here we describe two methods for glucoevatromonoside production, both using evatromonoside obtained by chemical degradation of digitoxin as the precursor. 1) Catalyst-controlled, regioselective glycosylation of evatromonoside to glucoevatromonoside using 2,3,4,6-tetra-O-acetyl-α-D-glucopyranosyl bromide as the sugar donor and 2-aminoethyldiphenylborinate as the catalyst resulted in an overall 30 % yield. 2) Biotransformation of evatromonoside using Digitalis lanata plant cell suspension cultures was less efficient and resulted only in overall 18 % pure product. Structural proof of products has been provided by extensive NMR data. Glucoevatromonoside and its non-natural 1-3 linked isomer neo-glucoevatromonoside obtained by semisynthesis were evaluated against renal cell carcinoma and prostate cancer cell lines.

Leishmanicidal Activity and Structure-Activity Relationships of Essential Oil Constituents.

Several constituents of essential oils have been shown to be active against pathogens such as bacteria, fungi, and protozoa. This study demonstrated the in vitro action of ten compounds present in essential oils against Leishmania amazonensis promastigotes. With the exception of p-cymene, all evaluated compounds presented leishmanicidal activity, exhibiting IC50 between 25.4 and 568.1 µg mL-1. Compounds with the best leishmanicidal activity presented a phenolic moiety (IC50 between 25.4 and 82.9 µg mL-1). Alicyclic alcohols ((-)-menthol and isoborneol) and ketones ((-)-carvone) promoted similar activity against the parasite (IC50 between 190.2 and 198.9 µg mL-1). Most of the compounds showed low cytotoxicity in L929 fibroblasts. Analysis of the structure-activity relationship of these compounds showed the importance of the phenolic structure for the biological action against the promastigote forms of the parasite.