Virus adsorbent systems based on Amazon holocellulose and nanomaterials.

The environment preservation has been an important motivation to find alternative, functional, and biodegradable materials to replace polluting petrochemicals. The production of nonbiodegradable face masks increased the concentration of microplastics in the environment, highlighting the need for sustainable alternatives, such as the use of local by-products to create efficient and eco-friendly filtering materials. Furthermore, the use of smart materials can reduce the risk of contagion and virus transmission, especially in the face of possible mutations. The development of novel materials is necessary to ensure less risk of contagion and virus transmission, as well as to preserve the environment. Taking these factors into account, 16 systems were developed with different combinations of precursor materials (holocellulose, polyaniline [ES-PANI], graphene oxide [GO], silver nanoparticles [AgNPs], and activated carbon [AC]). Adsorption tests of the spike protein showed that the systems containing GO and AC were the most efficient in the adsorption process. Similarly, plate tests conducted using the VSV-IN strain cultured in HepG2 cells showed that the system containing all phases showed the greatest reduction in viral titer method. In agreement, the biocompatibility tests showed that the compounds extracted from the systems showed low cytotoxicity or no significant cytotoxic effect in human fibroblasts. As a result, the adsorption tests of the spike protein, viral titration, and biocompatibility tests showed that systems labeled as I and J were the most efficient. In this context, the present research has significantly contributed to the technological development of antiviral systems, with improved properties and increased adsorption efficiency, reducing the viral titer and contributing efficiently to public health. In this way, these alternative materials could be employed in sensors and devices for filtering and sanitization, thus assisting in mitigating the transmission of viruses and bacteria. RESEARCH HIGHLIGHTS: Sixteen virus adsorbent systems were developed with different combinations of precursor materials (holocellulose, polyaniline (ES-PANI), graphene oxide (GO), silver nanoparticles (AgNPs), and activated carbon (AC)). The system that included all of the nanocomposites holocellulose, PANI, GO, AgNPs, and AC showed the greatest reduction in viral titration. The biocompatibility tests revealed that all systems caused only mild or moderate cytotoxicity toward human fibroblasts.

Tackling Osimertinib Resistance in EGFR-Mutant Non-Small Cell Lung Cancer.

The current landscape of targeted therapies directed against oncogenic driver alterations in non-small cell lung cancer (NSCLC) is expanding. Patients with EGFR-mutant NSCLC can derive significant benefit from EGFR tyrosine kinase inhibitor (TKI) therapy, including the third-generation EGFR TKI osimertinib. However, invariably, all patients will experience disease progression with this therapy mainly due to the adaptation of cancer cells through primary or secondary molecular mechanisms of resistance. The comprehension and access to tissue and cell-free DNA next-generation sequencing have fueled the development of innovative therapeutic strategies to prevent and overcome resistance to osimertinib in the clinical setting. Herein, we review the biological and clinical implications of molecular mechanisms of osimertinib resistance and the ongoing development of therapeutic strategies to overcome or prevent resistance.

Genomic Landscape of Primary Resistance to Osimertinib Among Hispanic Patients with EGFR-Mutant Non-Small Cell Lung Cancer (NSCLC): Results of an Observational Longitudinal Cohort Study.

BACKGROUND: Epidermal growth factor receptor (EGFR) mutations (EGFRm) represent one of the most common genomic alterations identified among patients with non-small cell lung cancer (NSCLC). Several targeted agents for patients with EGFRm have been proven safe and effective, including the third-generation tyrosine kinase inhibitor (TKI) osimertinib. Nonetheless, some patients will present with or develop EGFR-TKI resistance mechanisms. OBJECTIVE: We characterized the genomic landscape of primary resistance to osimertinib among Hispanic patients with EGFR-mutant NSCLC. METHODS: An observational longitudinal cohort study was conducted with two groups of patients, those with intrinsic resistance (cohort A) and those with long-term survival (cohort B). All patients were treated and followed between January 2018 and May 2022. All patients were assessed for Programmed Cell Death Ligand 1 (PD-L1) expression and Bcl-2-like protein 11 (BIM)/AXL mRNA expression before starting TKI. After 8 weeks of treatment, a liquid biopsy was performed to determine the presence of circulating free DNA (cfDNA), and next-generation sequencing (NGS) was used to identify mutations at the time of progression. In both cohorts, overall response rate (ORR), progression-free survival (PFS), and overall survival (OS) were evaluated. RESULTS: We found a homogeneous distribution of EGFR-sensitizing mutations in both cohorts. For cohort A, exon 21 mutations were more common than exon 19 deletions (ex19dels) for cohort B (P = 0.0001). The reported ORR for osimertinib was 6.3% and 100% for cohorts A and B, respectively (P = 0.0001). PFS was significantly higher in cohort B (27.4 months vs. 3.1 months; P = 0.0001) and ex19del patients versus L858R (24.5 months, 95% confidence interval [CI] 18.2-NR), vs. 7.6 months, 95% CI 4.8-21.1; P = 0.001). OS was considerably lower for cohort A (20.1 months vs. 36.0 months; P = 0.0001) and was better for patients with ex19del, no brain metastasis, and low tumor mutation burden. At the time of progression, more mutations were found in cohort A, identifying off-target alterations more frequently, including TP53, RAS, and RB1. CONCLUSION: EGFR-independent alterations are common among patients with primary resistance to osimertinib and significantly impact PFS and OS. Our results suggest that among Hispanic patients, other variables associated with intrinsic resistance include the number of commutations, high levels AXL mRNA, and low levels of BIM mRNA, T790M de novo, EGFR p.L858R presence, and a high tumoral mutational burden.

Follow-up of patients with R/R FLT3-mutation-positive AML treated with gilteritinib in the phase 3 ADMIRAL trial.

The phase 3 ADMIRAL (NCT02421939; Study ID: 2215-CL-0301) trial showed superior overall survival in patients with relapsed/refractory FLT3-mutation-positive acute myeloid leukemia (AML) randomized 2:1 to receive the oral FMS-like tyrosine kinase 3 inhibitor gilteritinib vs those randomized to receive salvage chemotherapy (SC). Here we provide a follow-up of the ADMIRAL trial 2 years after the primary analysis to clarify the long-term treatment effects and safety of gilteritinib in these patients with AML. At the time of this analysis, the median survival follow-up was 37.1 months, with deaths in 203 of 247 and 97 of 124 patients in the gilteritinib and SC arms, respectively; 16 gilteritinib-treated patients remained on treatment. The median overall survival for the gilteritinib and SC arms was 9.3 and 5.6 months, respectively (hazard ratio, 0.665; 95% confidence interval [CI], 0.518, 0.853; two-sided P = .0013); 2-year estimated survival rates were 20.6% (95% CI, 15.8, 26.0) and 14.2% (95% CI, 8.3, 21.6). The gilteritinib-arm 2-year cumulative incidence of relapse after composite complete remission was 75.7%, with few relapses occurring after 18 months. Overall, 49 of 247 patients in the gilteritinib arm and 14 of 124 patients in the SC arm were alive for ≥2 years. Twenty-six gilteritinib-treated patients remained alive for ≥2 years without relapse; 18 of these patients underwent transplantation (hematopoietic stem cell transplantation [HSCT]) and 16 restarted gilteritinib as post-HSCT maintenance therapy. The most common adverse events of interest during years 1 and 2 of gilteritinib therapy were increased liver transaminase levels; adverse event incidence decreased in year 2. Thus, continued and post-HSCT gilteritinib maintenance treatment sustained remission with a stable safety profile. These findings confirm that prolonged gilteritinib therapy is safe and is associated with superior survival vs SC. This trial was registered at www.clinicaltrials.gov as #NCT02421939.

Identification and evaluation of 4-anilinoquin(az)olines as potent inhibitors of both dengue virus (DENV) and Venezuelan equine encephalitis virus (VEEV).

There is an urgent need for novel strategies for the treatment of emerging arthropod-borne viral infections, including those caused by dengue virus (DENV) and Venezuelan equine encephalitis virus (VEEV). We prepared and screened focused libraries of 4-anilinoquinolines and 4-anilinoquinazolines for antiviral activity and identified three potent compounds. N-(2,5-dimethoxyphenyl)-6-(trifluoromethyl)quinolin-4-amine (10) inhibited DENV infection with an EC50 = 0.25 µM, N-(3,4-dichlorophenyl)-6-(trifluoromethyl)quinolin-4-amine (27) inhibited VEEV with an EC50 = 0.50 µM, while N-(3-ethynyl-4-fluorophenyl)-6,7-dimethoxyquinazolin-4-amine (54) inhibited VEEV with an EC50 = 0.60 µM. These series of compounds demonstrated nearly no toxicity with CC50 values greater than 10 µM in all cases. These promising results provide a future prospective to develop a clinical compound against these emerging viral threats.

Discovery of a Potent Dual SLK/STK10 Inhibitor Based on a Maleimide Scaffold.

SLK (STE20-like kinase) and STK10 (serine/threonine kinase 10) are closely related kinases whose enzymatic activity is linked to the regulation of ezrin, radixin, and moesin function and to the regulation of lymphocyte migration and the cell cycle. We identified a series of 3-anilino-4-arylmaleimides as dual inhibitors of SLK and STK10 with good kinome-wide selectivity. Optimization of this series led to multiple SLK/STK10 inhibitors with nanomolar potency. Crystal structures of exemplar inhibitors bound to SLK and STK10 demonstrated the binding mode of the inhibitors and rationalized their selectivity. Cellular target engagement assays demonstrated the binding of the inhibitors to SLK and STK10 in cells. Further selectivity analyses, including analysis of activity of the reported inhibitors against off-targets in cells, identified compound 31 as the most potent and selective inhibitor of SLK and STK10 yet reported.

A novel substrate for arrhythmias in Chagas disease.

BACKGROUND: Chagas disease (CD) is a neglected disease that induces heart failure and arrhythmias in approximately 30% of patients during the chronic phase of the disease. Despite major efforts to understand the cellular pathophysiology of CD there are still relevant open questions to be addressed. In the present investigation we aimed to evaluate the contribution of the Na+/Ca2+ exchanger (NCX) in the electrical remodeling of isolated cardiomyocytes from an experimental murine model of chronic CD. METHODOLOGY/PRINCIPAL FINDINGS: Male C57BL/6 mice were infected with Colombian strain of Trypanosoma cruzi. Experiments were conducted in isolated left ventricular cardiomyocytes from mice 180-200 days post-infection and with age-matched controls. Whole-cell patch-clamp technique was used to measure cellular excitability and Real-time PCR for parasite detection. In current-clamp experiments, we found that action potential (AP) repolarization was prolonged in cardiomyocytes from chagasic mice paced at 0.2 and 1 Hz. After-depolarizations, both subthreshold and with spontaneous APs events, were more evident in the chronic phase of experimental CD. In voltage-clamp experiments, pause-induced spontaneous activity with the presence of diastolic transient inward current was enhanced in chagasic cardiomyocytes. AP waveform disturbances and diastolic transient inward current were largely attenuated in chagasic cardiomyocytes exposed to Ni2+ or SEA0400. CONCLUSIONS/SIGNIFICANCE: The present study is the first to describe NCX as a cellular arrhythmogenic substrate in chagasic cardiomyocytes. Our data suggest that NCX could be relevant to further understanding of arrhythmogenesis in the chronic phase of experimental CD and blocking NCX may be a new therapeutic strategy to treat arrhythmias in this condition.

In vitro biocompatibility screening of a colloidal gum Arabic-polyaniline conducting nanocomposite.

Although polyaniline (PANI) is a widely investigated conductive polymer for biological applications, studies addressing the biocompatibility of colloidal PANI dispersions are scarcely found in the literature of the area. Therefore, PANI nanoparticles stabilized by the natural polysaccharide gum Arabic (GA) were screened for their biocompatibility. The GA successfully stabilized the colloidal PANI-GA dispersions when exposed to a protein-rich medium, showing compatibility with the biological environment. The results obtained from a series of in vitro assays showed that, after up to 48 h of exposure to a range of PANI-GA concentrations (1-50 µg/mL), both mouse BALB/3T3 fibroblasts and RAW 264.7 macrophages showed no evidence of change in cellular proliferation, viability and metabolic activity. An increase in macrophage granularity poses as evidence of phagocytic uptake of PANI-GA, without resulting activation of this cell type. Additionally, the PANI-GA nanoparticles modulated the cell morphology changes induced on fibroblasts by GA in a concentration-dependent manner. Thus, this unprecedented biocompatibility study of PANI nanoparticles stabilized by a plant gum exudate polysaccharide showed promising results. This simple biomaterial might be further developed into colloidal formulations for biological and biomedical applications, taking advantage of its versatility, biocompatibility, and conductive properties.

The MPO system participates actively in the formation of an oxidative environment produced by neutrophils and activates the antioxidant mechanism of Naegleria fowleri.

Naegleria fowleri produces a fatal disease called primary amebic meningoencephalitis (PAM), which is characterized by an extensive inflammatory reaction in the CNS. It is known that the immune response is orchestrated mainly by neutrophils, which activate several defense mechanisms in the host, including phagocytosis, the release of different enzymes such as myeloperoxidase (MPO), and the production of neutrophil extracellular traps. However, the mechanisms by which amoebas evade the neutrophil response are still unknown. In this study, we analyzed the ability of N. fowleri to respond to the stress exerted by MPO. Interestingly, after the interaction of trophozoites with neutrophils, the amoeba viability was not altered; however, ultrastructural changes were observed. To analyze the influence of MPO against N. fowleri and its participation in free radical production, we evaluated its enzymatic activity, expression, and localization with and without the specific 4-aminobenzoic acid hydrazide inhibitor. The production of oxidizing molecules is the principal mechanism used by neutrophils to eliminate pathogens. In this context, we demonstrated an increase in the production of NO, superoxide anion, and reactive oxygen species; in addition, the overexpression of several antioxidant enzymes present in the trophozoites was quantified. The findings strongly suggest that N. fowleri possesses antioxidant machinery that is activated in response to an oxidative environment, allowing it to evade the neutrophil-mediated immune response, which may contribute to the establishment of PAM.

Evaluation of [18F]-N-Methyl lansoprazole as a Tau PET Imaging Agent in First-in-Human Studies.

Development of positron emission tomography (PET) imaging agents capable of quantifying tau aggregates in neurodegenerative disorders such as Alzheimer's disease (AD) is of enormous importance in the field of dementia research. The aim of the present study was to conduct first-in-man imaging studies with the potential novel tau imaging agent [18F]N-methyl lansoprazole ([18F]NML). Herein we report validation of the synthesis of [18F]NML for clinical use by labeling the trifluoromethyl group via radiofluorination of the corresponding gem-difluoro enol ether precursor. This is the first use of this method for clinical production of PET radiotracers and confirmed that it can be readily implemented at multiple production facilities to provide [18F]NML in good noncorrected radiochemical yield (3.4 ± 1.5 GBq, 4.6% ± 2.6%) and molar activity (120.1 ± 186.3 GBq/µmol), excellent radiochemical purity (>97%), and suitable for human use (n = 15). With [18F]NML in hand, we conducted rodent biodistribution, estimates of human dosimetry, and preliminary evaluation of [18F]NML in human subjects at two imaging sites. Healthy controls (n = 4) and mildly cognitively impaired (MCI) AD patients (n = 6) received [18F]NML (tau), [18F]AV1451 (tau), and [18F]florbetaben or [18F]florbetapir (amyloid) PET scans. A single progressive supranuclear palsy (PSP) patient also received [18F]NML and [18F]AV1451 PET scans. [18F]NML showed good brain uptake, reasonable pharmacokinetics, and appropriate imaging characteristics in healthy controls. The mean ± SD of the administered mass of [18F/19F]NML was 2.01 ± 2.17 µg (range, 0.16-8.27 µg) and the mean administered activity was 350 ± 62 MBq (range, 199-403 MBq). There were no adverse or clinically detectable pharmacologic effects in any of the 11 subjects, and no significant changes in vital signs were observed. However, despite high affinity for tau in vitro, brain retention in MCI/AD and PSP patients was low, and there was no evidence of specific signals in vivo that corresponded to tau. Although it is still unclear why clinical translation of the radiotracer was unsuccessful, we nevertheless conclude that further development of [18F]NML as a tau PET imaging agent is not warranted at this time.

Carboranylanilinoquinazoline EGFR-inhibitors: toward 'lead-to-candidate' stage in the drug-development pipeline.

Background: Carboranylanilinoquinazoline-hybrids, developed for boron neutron capture therapy, have demonstrated cytotoxicity against murine-glioma cells with EGFR-inhibition ability. In addition, their adequate aqueous/metabolic stabilities and ability to cross blood-brain barrier make them good leads as to become antiglioma drugs. Aim: Analyze drug-like properties of representative carboranylanilinoquinazolines. Materials & methods: To expand carboranylanilinoquinazolines therapeutic spectrum, we studied their ability to act against glioma-mammal cells, U-87 MG and other tyrosine kinase-overexpress cells, HT-29. Additionally, we predicted theoretically and studied experimentally drug-like properties, in other words, organization for economic cooperation and development-recommended toxicity-studies and, due to some aqueous-solubility problems, and vehicularization for oral and intravenous administrations. Conclusion: We have identified a promising drug-candidate with broad activity spectrum, appropriate drug-like properties, adequate toxicological behavior and able ability to be loaded in suitable vehicles.

Potential Inhibitors of the Activity of the Cholesterol-Ester Transfer Protein.

The cholesterol-ester transfer protein (CETP) exchanges lipids between high-density lipoproteins (HDLs) and low-density lipoproteins (LDLs). The excessive transport of lipids from HDLs to LDLs mediated by this protein can cause an alteration in the deposition of lipoproteins onto the arterial walls, thus promoting the development of arteriosclerosis. Different CETP inhibitors have been tested in recent years, but none has been confirmed as being effectively palliative for the disease. We employed in silico databases and molecular docking as a computational method to predict how potential CETP inhibitors could interact with the active site of the CETP protein. Upon previously comparing two computer software packages to determine which generated a greater number of accurate CETP-inhibitor-complex structures, we chose the more appropriate program for our studies. We then abstracted a series of databases of known CETP inhibitors and noninhibitors exhibiting different 50% concentrations of CETP-inhibitory (INH) activity, to generate virtual structures for docking with different combinations of the CETP receptor. From this process, we obtained as the most suitable structure 4F2A_1OB_C_PCW-it accordingly having a greater area under the receiver operating characteristic curve. The molecular docking of known compounds in comparison with the respective conformation of this inhibitor enabled us to obtain ΔGs (in kcal/mol) from which data we made a first exploration of unknown compounds for CETP-INH activity. Thus, the 4F2A_1OB_C_PCW structure was docked with DrugBank-Approved commercial compounds in an extensive database, whose status had already been established from pharmacokinetics and toxicology. In this study, we present a group of potential compounds as CETP-inhibitor candidates.

Availability of FLT3 inhibitors: how do we use them?

The natural history of FLT3-mutated AML is changing after the approval of midostaurin for frontline therapy and gilteritinib for relapsed or refractory patients. Recently reported, positive randomized trials of the drugs gilteritinib, quizartinib, and sorafenib predict even wider use of FLT3 inhibitors going forward. FLT3 inhibitors now emerge as an important, if not indispensable, part of therapy for a large subset of high-risk patients.

In vitro antiplasmodial activity, pharmacokinetic profiles and interference in isoprenoid pathway of 2-aniline-3-hydroxy-1.4-naphthoquinone derivatives.

BACKGROUND: Plasmodium falciparum has shown multidrug resistance, leading to the necessity for the development of new drugs with novel targets, such as the synthesis of isoprenic precursors, which are excellent targets because the pathway is different in several steps when compared with the human host. Naphthoquinone derivatives have been described as potentially promising for the development of anti-malarial leader molecules. In view of that, the focus in this work is twofold: first, evaluate the in vitro naphthoquinone antiplasmodial activity and cytotoxicity; secondly, investigate one possible action mechanism of two derivatives of hydroxy-naphthoquinones. RESULTS: The two hydroxy-naphthoquinones derivatives have been tested against P. falciparum in vitro, using strains of parasites chloroquine-sensitive (3D7) and chloroquine-resistant (Dd2), causing 50% inhibition of parasite growth with concentrations that varied from 7 to 44.5 µM. The cell viability in vitro against RAW Cell Line displayed IC50 = 483.5 and 714.9 µM, whereas, in primary culture tests using murine macrophages, IC50 were 315.8 and 532.6 µM for the two selected compounds, causing no haemolysis at the doses tested. The in vivo acute toxicity assays exhibited a significant safety margin indicated by a lack of systemic and behavioural toxicity up to 300 mg/kg. It is suggested that this drug seems to inhibit the biosynthesis of isoprenic compounds, particularly the menaquinone and tocopherol. CONCLUSIONS: These derivatives have a high potential for the development of new anti-malarial drugs since they showed low toxicity associated to a satisfactory antiplasmodial activity and possible inhibition of a metabolic pathway distinct from the pathways found in the mammalian host.

Usnic acid-loaded polyaniline/polyurethane foam wound dressing: preparation and bactericidal activity.

The improved bactericidal activity of new composites for wound dressing prototypes represents an important strategy for development of more efficient devices that make use of synergistic interaction between components. The doping level of polyaniline represents a critical parameter for its corresponding biologic activity. In this work, it is explored the doping effect of usnic acid on undoped polyaniline, that introduces important advantages namely, improved bactericidal activity of polyaniline and the anti-biofilm properties of lichen derivative. The deposition of the resulting material on polyurethane foam potentializes its applicability as wound dressing, characterizing a new platform for application against Escherichia coli and Staphylococcus aureus.

Preparation of Novel Homodimers Derived from Cytotoxic Isoquinolinequinones. A Twin Drug Approach.

The synthesis of five novel homodimers is reported based on the anilinoisoquinolinequinone scaffold. In these twin-drug derivatives, two units of the anilinoquinone pharmacophores are linked through a methylene spacer. The formation of dimers was achieved by reaction of isoquinolinequinones with 4, 4'-diaminodiphenylmethane via a sequence of two oxidative amination reactions. A preliminary in vitro screening of the homodimers reveals moderate to high cytotoxic activities against MDA-MB-21 breast adenocarcinoma and B16-F10 murine metastatic melanoma cell lines. The asymmetrical homodimer 15 stands out due to its cytotoxic potencies at submicromolar concentrations and high selectivity index (mean IC50 = 0.37 µM; SI = 6.97) compared to those of etoposide (mean IC50 = 3.67; SI = 0.32) and taxol (mean IC50 = 0.35; SI = 0.91) employed as reference anticancer drugs.

Novel piperonal 1,3,4-thiadiazolium-2-phenylamines mesoionic derivatives: Synthesis, tyrosinase inhibition evaluation and HSA binding study.

A novel series of piperonal mesoionic derivatives (PMI 1-6) was synthesized. Tyrosinase inhibition in the presence of PMI-1, -2, -3, -4, -5 and -6 as well as human serum albumin (HSA) binding studies with PMI-5 and PMI-6 were done by spectroscopic and theoretical methods. The mesoionic compound PMI-5 is the most promising tyrosinase inhibitor with a noncompetitive inhibitory mechanism and an IC50=124µmolL-1. In accordance with the kinetic profile, molecular docking results show that PMI-5 is able to interact favorably with the tyrosinase active site containing the substrate molecule, L-DOPA, interacting with Val-247, Phe-263 and Val-282 residues. The spectroscopic results for the interaction HSA:PMI-5 and HSA:PMI-6 indicated that these mesoionic compounds can associate with HSA in the ground state and energy transfer can occur with high probability. The binding was moderate, spontaneous and can perturb significantly the secondary structure of the albumin. The molecular docking results suggest that PMI-5 and PMI-6 are able to be accommodated inside the Sudlow's site I in HSA, interacting with hydrophobic and hydrophilic amino acid residues.

Antiviral activity of A771726, the active metabolite of leflunomide, against Junín virus.

The aim of this study was to investigate the effect of A771726, the active metabolite of leflunomide, (CONICET-UBA), Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Ciudad against the infection with Junín virus (JUNV), agent of Argentine hemorrhagic fever (AHF). The treatment with non-cytotoxic concentrations of A771726 of Vero and A549 cells infected with JUNV inhibited virus replication in a dose-dependent manner, as determined by virus yield reduction assay. The antiviral effectiveness of A771726 was not importantly affected by the multiplicity of infection and the virus strain. Moreover, the combination of A771726 and ribavirin had a significantly more potent antiviral activity than each single drug treatment. Mechanistic studies showed that the main action of A771726 is exerted before 6 h of JUNV infection. Accordingly, inhibition of viral RNA synthesis was detected in treated infected cells by real time RT-PCR. The exogenous addition of uridine or orotic acid produced a partial reversal of the inhibitory effect of A771726 on infective virus production whereas a total reversion was detected on JUNV RNA synthesis, probably by restoration of the enzymatic activity of dihydroorotate dehydrogenase (DHODH) and the intracellular pyrimidine pools. In conclusion, these results suggest that the antiviral target would be viral RNA synthesis through pyrimidine depletion, but any other effect of the compound on JUNV infection cannot be excluded. This study opens the possibility of the therapeutic application of a wide spectrum host-targeted compound alone or in combination with ribavirin to combat AHF as well as other human pathogenic arenaviruses.

Tetrodotoxin-insensitive electrical field stimulation-induced contractions on Crotalus durissus terrificus corpus cavernosum.

Reptiles are the first amniotes to develop an intromitent penis, however until now the mechanisms involved in the electrical field stimulation-induced contraction on corpora cavernosa isolated from Crotalus durissus terrificus were not investigated. Crotalus and rabbit corpora cavernosa were mounted in 10 mL organ baths for isometric tension recording. Electrical field stimulation (EFS)-induced contractions were performed in presence/absence of phentolamine (10 µM), guanethidine (30 µM), tetrodotoxin (1 µM and 1mM), A-803467 (10 µM), 3-iodo-L-Tyrosine (1 mM), salsolinol (3 µM) and a modified Krebs solution (equimolar substitution of NaCl by N-methyl-D-glucamine). Immuno-histochemistry for tyrosine hydroxylase was also performed. Electrical field stimulation (EFS; 8 Hz and 16 Hz) caused contractions in both Crotalus and rabbit corpora cavernosa. The contractions were abolished by previous incubation with either phentolamine or guanethidine. Tetrodotoxin (1 µM) also abolished the EFS-induced contractions of rabbit CC, but did not affect EFS-induced contractions of Crotalus CC. Addition of A-803467 (10 µM) did not change the EFS-induced contractions of Crotalus CC but abolished rabbit CC contractions. 3-iodo-L-Tyrosine and salsolinol had no effect on EFS-induced contractions of Crotalus CC and Rabbit CC. Replacement of NaCl by N- Methyl-D-glucamine (NMDG) abolished EFS-induced contractions of rabbit CC, but did not affect Crotalus CC. The presence of tyrosine hydroxylase was identified in endothelial cells only of Crotalus CC. Since the EFS-induced contractions of Crotalus CC is dependent on catecholamine release, insensitive to TTX, insensitive to A803467 and to NaCl replacement, it indicates that the source of cathecolamine is unlikely to be from adrenergic terminals. The finding that tyrosine hydroxylase is present in endothelial cells suggests that these cells can modulate Crotalus CC tone.