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Results for “Pharmacology and Toxicology”

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At least 199 records · Page 11Linked to original sources

bioRxiv2018

Hydrophobic organic contaminants are not linked to microplastic uptake in Baltic Sea herring

It is commonly accepted that microplastic (MP) ingestion can lead to lower food intake and bioaccumulation of hydrophobic organic contaminants (HOCs) in aquatic organisms. However, causal links between MP and contaminant levels in biota are poorly understood and in situ data are very limited. Here, we investigated whether HOC concentrations in herring muscle tissue (Clupea harengus membras) are related to MP ingestion using fish caught along the West coast of the Baltic Sea. The MP occurrence exhibited a large geographic variability, with MP found in 22.3% of the fish examined. The population average was 1.0 MP ind-1; however, when only individuals containing MP were considered, the average MP burden was 4.4 MP ind-1. We also found that MP burden decreased with reproductive stage of the fish but increased with its body size. To predict MP abundance in fish guts, we constructed a mass-balance model using literature data on MP in the water column and physiological rates on ingestion and gut evacuation for clupeids of a similar size. The model output was in agreement with the observed values, thus supporting the validity of the results. Contaminant concentrations in the muscle tissue were unrelated to the MP levels in fish, suggesting a lack of direct links between the levels of HOCs and MP ingestion. Thus, despite their ubiquity, MP are unlikely to have a measurable impact on food intake or the total body burden of hydrophobic contaminants in Baltic herring. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=80 SRC="FIGDIR/small/363127v3_ufig1.gif" ALT="Figure 1"> View larger version (22K): org.highwire.dtl.DTLVardef@4cb5b6org.highwire.dtl.DTLVardef@2de3c8org.highwire.dtl.DTLVardef@c58baborg.highwire.dtl.DTLVardef@154b828_HPS_FORMAT_FIGEXP M_FIG C_FIG

pharmacology and toxicology↗

bioRxiv2018

Transcriptional regulatory mechanisms of fibrosis development in mouse lung tissue exposed to carbon nanotubes

BackgroundCarbon nanotubes (CNTs) usage has rapidly increased in the last few decades due to their unique properties, exploited in various industrial and commercial products. Certain types of CNTs cause adverse health effects, including chronic inflammation and fibrosis. Despite the large number of in vitro and in vivo studies evaluating these effects, many important questions remain unanswered due to a lack of mechanistic understanding of how CNTs induce cellular stress responses. In order to predict CNT toxicity, it is important to understand which transcriptional programs are specifically activated in response to CNTs, and what similarities and differences exist in relation to other toxic inducers exerting similar adverse effects.\n\nResultsA systems biology approach was applied to reveal complex interactions at the molecular level in mouse lung tissue in response to different fibrosis inducers: two types of multi-walled CNTs, NM-401 and NRCWE-26, and bleomycin (BLM). Based on mRNA gene expression profiles, we inferred gene regulatory networks (GRNs) to capture functional hierarchical regulatory structures between genes and their regulators. We found that activities of the transcription factors (TFs) Myc, Arid5a and Mxd1 were associated with the regulation of cytokine transcription in response to CNTs, while in response to BLM treatment, Myc was associated with p53 signaling. TF Litaf was identified as the essential regulator for noncanonical signaling of TLR2/4 driven by CNTs. Despite the different nature of the lung injury caused by CNTs and BLM, we identified common stress response modules, that included DNA damage (TFs: E2f8, E2f1, Foxm1), M1/M2 macrophage polarization (TF: Mafb), Interferon response (TFs: Irf7, Stat2 and Irf9) for all agents.\n\nConclusionsThese results suggest that the reconstruction and analysis of TF-centric gene interaction networks can reveal key targets and regulators of cellular stress responses to toxic agents.

pharmacology and toxicology↗

bioRxiv2018

Drug adaptation influences cardiotoxicity caused by tyrosine kinase inhibitors in iPSC-derived human cardiomyocytes

Cardiotoxicity induced by anti-cancer drugs is of increasing concern as the durability of therapeutic responses increases. The molecular basis of cardiotoxicity remains poorly understood, particularly when due to drug classes that do not inhibit the hERG potassium channel or cause the arrhythmias associated with long QT syndrome. This paper describes systematic molecular profiling of one such class of drugs, tyrosine kinase inhibitors (TKIs), which are widely used to treat solid tumors. Human cardiomyocytes differentiated from induced pluripotent stem cells (hiPSC-CMs) were exposed to one of four TKIs (Sunitinib, Sorafenib, Lapatinib and Erlotinib) observed to cause different levels of human cardiotoxicity and profiled by RNA sequencing (RNA-Seq) and mass spectroscopy-based proteomic analysis. We find that TKIs have diverse effects on hiPSC-CMs but genes involved in cardiac metabolism are particularly sensitive. In the case of Sorafenib, many genes involved in oxidative phosphorylation are down regulated resulting in a profound defect in mitochondrial metabolism. Cells adapt to this by upregulating aerobic glycolysis. Metabolic remodeling makes cells less acutely sensitive to Sorafenib and the effect is reversible upon drug withdrawal. Thus, the response of cardiomyocytes to Sorafenib is characterized by adaptive drug resistance previously described in tumor cells.

pharmacology and toxicology↗

bioRxiv2018

Anticlastogenic Effect of Ulva Fasciata, Against Cyclophosphamide and Mitomycin C Induced Chromosomal Damage in Swiss Albino Mice

Chemoprevention is a strategy to reduce the incidence of human cancer either by inhibiting initiation of carcinogenesis or by preventing exposure to carcinogens, by the use of plant or animal derived ingredients. In the present study we investigated the anticlastogenic effect of ethanol extract of Ulva fasciata, a green seaweed, against the chromosomal aberration and micronucleus induced by the anticancer drugs cyclophosphamide and mitomycin C. Three doses of extract (25, 50 and 75 mg/kg.b.w) was given by oral gavage for 5 days at 24 hr. intervals and on the 5th day, CP (25 and 50mg/kg.b.w) or MMC (1 and 2mg/kg.b.w) were intraperitoneally injected and 24hrs. later micronucleus and chromosomal aberrations assays were performed. Our results show that Ulva extract gave significant protection against the CP and MMC induced damages by reducing micronucleus and chromosomal aberrations. The protection imparted by Ulva could be due to the synergistic and/or additive effects of biologically active ingredients present in the seaweed.

pharmacology and toxicology↗

bioRxiv2018

Moderate developmental alcohol exposure reduces repetitive alternation in a zebrafish model of fetal alcohol spectrum disorders.

The damaging effects of alcohol on a developing fetus are well known and cause a range of conditions known as fetal alcohol spectrum disorder (FASD). High levels of alcohol exposure lead to physical deformity and severe cognitive deficits, but more moderate exposure leads to a range of subtle cognitive effects such as reduced social behavior, higher propensity to develop addictions, and reduced spatial working memory. Previous studies have demonstrated that following exposure to relatively low levels of ethanol during early brain development (equivalent in humans to moderate exposure) zebrafish display a range of social and behavioral differences. Here, our aim was to test the hypothesis that moderate developmental ethanol exposure would affect aspects of learning and memory in zebrafish. In order to do this, we exposed zebrafish embryos to 20mM [0.12% v/v] ethanol from 2 to 9 dpf to model the effects of moderate prenatal ethanol (MPE) exposure. At 3 months old, adult fish were tested for appetitive and aversive learning, and for spatial alternation in a novel unconditioned y-maze protocol. We found that MPE did not affect appetitive or aversive learning, but exposed-fish showed a robust reduction in repetitive alternations in the y-maze when compared to age matched controls. This study confirms that moderate levels of ethanol exposure to developing embryos have subtle effects on spatial working memory in adulthood. Our data thus suggest that zebrafish may be a promising model system for studying the effects of alcohol on learning and decision-making, but also for developing treatments and interventions to reduce the negative effects of prenatal alcohol.

pharmacology and toxicology↗

bioRxiv2018

Costing ‘the’ MTD … in 2-D

BackgroundI have previously evaluated the efficiency of one-size-fits-all dosing for single agents in oncology (Norris 2017b). By means of a generic argument based on an Emax-type dose-response model, I showed that one-size-fits-all dosing may roughly halve a drugs value to society. Since much of the past decades innovation in oncology dose-finding methodology has involved the development of special methods for combination therapies, a generalization of my earlier investigations to combination dosing seems called-for.\n\nMethodsFundamental to my earlier work was the premise that optimal dose is a characteristic of each individual patient, distributed across the population like any other physiologic characteristic such as height. I generalize that principle here to the 2-dimensional setting of combination dosing with drugs A and B, using a copula to build a bivariate joint distribution of (MTDi,A, MTDi,B) from single-agent marginal densities of MTDi,A and MTDi,B, and interpolating toxicity isocontours in the (a, b)-plane between the respective monotherapy intercepts. Within this framework, three distinct notional toxicities are elaborated: one specific to drug A, a second specific to drug B, and a third nonspecific toxicity clinically attributable to either drug. The dose-response model of (Norris 2017b) is also generalized to this 2-D scenario, with the addition of an interaction term to provide for a complementary effect from combination dosing. A population of 1,000 patients is simulated, and used as a basis to evaluate population-level efficacy of two pragmatic dose-finding designs: a dose-titration method that maximizes dose-intensity subject to tolerability, and the well-known POCRM method for 1-size-fits-all combination-dose finding. Hypothetical oracular methods are also evaluated, to define theoretical upper limits of performance for individualized and 1-size-fits-all dosing respectively.\n\nResultsIn our simulation, pragmatic titration attains 89% efficiency relative to theoretically optimal individualized dosing, whereas POCRM attains only 55% efficiency. The passage from oracular individualized dosing to oracular 1-size-fits-all dosing incurs an efficiency loss of 33%, while the parallel passage (within the pragmatic realm) from titration to POCRM incurs a loss of 38%.\n\nConclusionsIn light of the 33% figure above, the greater part of POCRMs 38% efficiency loss relative to titration appears attributable to POCRMs 1-size-fits-all nature, rather than to any pragmatic difficulties it confronts. Thus, appeals to pragmatic considerations would seem neither to justify the decision to use 1-size-fits-all dose-finding designs, nor to excuse their inefficiencies

pharmacology and toxicology↗

bioRxiv2018

Apigenin relaxes rat intrarenal arteries: involvement of Cl- channels and K+ channels

The vasodilator effect of apigenin (API) was demonstrated in a number of vascular beds. We aimed to characterize the vasospasmolytic and electrophysiological effects of apigenin (API) in intrarenal arteries (IRAs). The vascular tone of male rat isolated IRAs was recorded with a myograph. Transmembrane Cl- currents through Ca2+-activated Cl- channels (CaCCs), K+ currents through voltage-gated K+ (Kv) channels and inwardly rectifier K+ (Kir) channels were recorded with patch clamp in the freshly isolated arterial smooth muscle cells (ASMCs). Preincubation with API (10-100 M) concentration-dependently depressed the contractions induced by KCl, 9,11-dideoxy-9,11-methanoepoxy prostaglandin F2 (U46619), phenylephrine and vasopressin without significant preference and the IC50 values were 13.27-26.26 M. Acute application of API elicited instant relaxations in the IRAs precontracted with these vasoconstrictors and the RC50 values were 5.80-24.33 M. API relaxation was attenuated by chloride deprivation, CaCC blockers, Kv blocker and nitric oxide synthase inhibitor, but not by Kir blocker and cyclooxygenase inhibitor. At 10-100 M, API depressed CaCC currents and Kir currents while enhanced Kv currents of IRA ASMCs. The present results demonstrate that API counteracts various vasoconstrictors noncompetitively and nonspecifically and suggest that modulation of CaCCs, Kv and Kir channels of IRA ASMCs is involved in its vasospasmolytic effects.

pharmacology and toxicology↗

bioRxiv2018

Development of an imaging toolbox to assess the therapeutic potential and biodistribution of macrophages in a mouse model of multiple organ dysfunction

Cell-based regenerative medicine therapies require robust preclinical safety, efficacy, biodistribution and engraftment data prior to clinical testing. To address these challenges, we have developed an imaging toolbox comprising multi-spectral optoacoustic tomography and ultrasonography, which allows the degree of kidney, liver and cardiac injury and the extent of functional recovery to be assessed non-invasively in a mouse model of multi-organ dysfunction. This toolbox allowed us to determine the therapeutic effects of adoptively transferred M2 macrophages. Using bioluminescence imaging, we could then investigate the association between amelioration and biodistribution. Macrophage therapy improved kidney and liver function to a limited extent, but did not ameliorate histological damage. No improvement in cardiac function was observed. Biodistribution analysis showed that macrophages homed and persisted in the injured kidneys and liver, but did not populate the heart. Our data suggest that the limited improvement observed in kidney and liver function could be mediated by M2 macrophages.

pharmacology and toxicology↗

bioRxiv2018

Atropselective Oxidation of 2,2′,3,3′,4,6′-Hexachlorobiphenyl (PCB 132) to Hydroxylated Metabolites by Human Liver Microsomes: Involvement of Arene Oxide Intermediates

PCBs and their hydroxylated metabolites have been associated with neurodevelopmental disorders. Several neurotoxic congeners display axial chirality and atropselectively affect cellular targets implicated in PCB developmental neurotoxicity; however, only limited information is available regarding the metabolism of these congeners in humans. We hypothesize that the oxidation of 2,2',3,3',4,6'-hexachlorobiphenyl (PCB 132) by human liver microsomes (HLMs) is atropselective and displays inter-individual variability. To test this hypothesis, PCB 132 (50 M) was incubated with pooled or single donor HLMs for 10, 30 or 120 min at 37 {degrees}C, and levels and enantiomeric fractions of PCB 132 and its metabolites were determined gas chromatographically. The major metabolite formed by different HLM preparations was either 2,2',3,4,4',6'-hexachlorobiphenyl-3'-ol (3'-140) or 2,2',3,3',4,6'-hexachlorobiphenyl-5'-ol (5'-132). 2,2',3,3',4,6'-Hexachlorobiphenyl-4'-ol (4'-132) and 2,2',3,3',4,6'-hexachlorobiphenyl-4',5'-diol (4',5'-132) were minor metabolites. The second eluting atropisomer of PCB 132 was slightly enriched in some HLM incubations. The formation of the first eluting atropisomer of 3'-140 was nearly enantiospecific (EF > 0.8). The second eluting atropisomer of 5'-132 was enriched in all microsomal preparations investigated. EF values differed slightly between single donor HLM preparations (EF = 0.84 to 0.96 for 3'-140; EF = 0.12 to 0.19 for 5'-132). These findings suggest that there are inter-individual differences in the atropselective biotransformation of PCB 132 to OH-PCBs in humans that may affect neurotoxic outcomes.

pharmacology and toxicology↗

bioRxiv2018

Development and Application of a High-Content Virion Display Human GPCR Array

G protein-coupled receptors (GPCRs) comprise the largest membrane protein family in humans and can respond to a wide variety of ligands and stimuli. Like other multi-pass membrane proteins, the biochemical properties of GPCRs are notoriously difficult to study because they must be embedded in lipid bilayers to maintain their native conformation and function. To enable an unbiased, high-throughput platform to profile biochemical activities of GPCRs in native conformation, we individually displayed 315 human non-odorant GPCRs (>85% coverage) in the envelope of human herpes simplex virus-1 and immobilized on glass to form a high-content Virion Display (VirD) array. Using this array, we found that 50% of the tested commercial anti-GPCR antibodies (mAbs) is ultra-specific, and that the vast majority of those VirD-GPCRs, which failed to be recognized by the commercial mAbs, could bind to their canonical ligands, indicating that they were folded correctly. Next, we used the VirD-GPCR arrays to examine binding specificity of two known peptide ligands and recovered expected interactions, as well as new off-target interactions, three of which were confirmed with real-time kinetics measurements. Finally, we explored the possibility of discovering novel pathogen targets by probing VirD-GPCR arrays with live group B Streptococcus (GBS), a common Gram-positive bacterium causing neonatal meningitis. Using cell invasion assays and a mouse model of hematogenous meningitis, we showed that inhibition of one of the five newly identified GPCRs, CysLTR1, greatly reduced GBS penetration in brain-derived endothelial cells and in mouse brains. Therefore, our work demonstrated that the VirD-GPCR array holds great potential for high-throughput, unbiased screening for small molecule drugs, affinity reagents, and deorphanization.

pharmacology and toxicology↗

bioRxiv2018

Predicting negative control drugs to support research in drug safety

The lack of high-quality reference data is a major limitation in drug safety and drug discovery science. Unreliable standards prohibit the use of supervised learning methods and make evaluation of algorithms difficult. While some data is available for positive examples (e.g. which drugs are associated with a side effect), there are no systematic resources of negative controls. To solve this issue, we introduced SIDERctrl, a computational method that ranks drugs based on the likelihood of not causing a side effect. We applied SIDERctrl to predict negative controls from unreported drugs of 890 side effects in SIDER. Our predictions decreased the false negative rate by one-third according to a validation study using AEOLUS data. Three sets of predicted negative controls by different thresholds of precision were provided, and can be accessed at http://tatonettilab.org/resources/negative-drugs.html. This new reference standard will be important in chemical biology, drug development, and pharmacoepidemiology.\n\nKEY POINTSO_LIThe lack of systematic resources providing negative control drugs limits the performance of existing research in drug safety.\nC_LIO_LIWe developed a novel method that integrated chemical and biological properties a drug and the target proteins to calculate the likelihood of the drug being negative control.\nC_LIO_LIWe applied our method to 890 side effects, and showed that our method significantly decreased the false negative rate of predictions.\nC_LI

pharmacology and toxicology↗

bioRxiv2018

In silico prioritization of transporter-drug relationships from drug sensitivity screens

The interplay between drugs and cell metabolism is a key factor in determining both compound potency and toxicity. In particular, how and to what extent transmembrane transporters affect drug uptake and disposition is currently only partially understood. Most transporter proteins belong to two protein families: the ATP-Binding Cassette (ABC) transporter family, whose members are often involved in xenobiotic efflux and drug resistance, and the large and heterogeneous family of Solute carriers (SLCs). We recently argued that SLCs are collectively a rather neglected gene group, with most of its members still poorly characterized, and thus likely to include many yet-to-be-discovered associations with drugs. We searched publicly available resources and literature to define the currently known set of drugs transported by ABCs or SLCs, which involved ~500 drugs and more than 100 transporters. In order to extend this set, we then mined the largest publicly available pharmacogenomics dataset, which involves approximately 1000 molecularly annotated cancer cell lines and their response to 265 chemical compounds, and used regularized linear regression models (Elastic Net, LASSO) to predict drug responses based on SLC and ABC data (expression levels, SNVs, CNVs). The most predictive models included both known and previously unidentified associations between drugs and transporters. To our knowledge, this represents the first application of regularized linear regression to this set of genes, providing an extensive prioritization of potentially pharmacologically interesting interactions.

pharmacology and toxicology↗

bioRxiv2018

Isoform-specific NADPH oxidase inhibition for pharmacological target validation

Unphysiological reactive oxygen species (ROS) formation is considered an important pathomechanism for several disease phenotypes with high unmet medical need. Therapeutically, antioxidants have failed multiple times. Instead, focusing on only disease-relevant, enzymatic sources of ROS appears to be a more promising and highly validated approach. Here the family of five NADPH oxidases (NOX) stands out as drug targets. Validation has been restricted, however, mainly to genetically modified rodents and is lacking in other species including human. It is thus unclear whether the different NOX isoforms are sufficiently distinct to allow selective pharmacological modulation. Here we show for five of the most advanced NOX inhibitors that indeed isoform selectivity can be achieved. NOX1 was most potently (IC50) targeted by ML171 (0.1 M); NOX2, by VAS2870 (0.7 M); NOX4, by M13 (0.01 M) and NOX5, by ML090 (0.01 M). Conditions need to be carefully controlled though as previously unrecognized non-specific antioxidant and assay artefacts may limit the interpretation of data and this included, surprisingly, one of the most advanced NOX inhibitors, GKT136901. As proof-of-principle that now also pharmacological and non-rodent target validation of different NOX isoforms is possible, we used a human blood-brain barrier model and NOX inhibitor panel at IC50 concentrations. The protective efficacy pattern of this panel confirmed the predominant role of NOX4 in stroke from previous genetic models. Our findings strongly encourage further lead optimization efforts for isoform-selective NOX inhibitors and clinical development and provide an experimental alternative when genetic validation of a NOX isoform is not an option.\n\nO_FIG O_LINKSMALLFIG WIDTH=193 HEIGHT=200 SRC=\"FIGDIR/small/382226v2_ufig1.gif\" ALT=\"Figure 1\">\nView larger version (26K):\norg.highwire.dtl.DTLVardef@1708996org.highwire.dtl.DTLVardef@15909daorg.highwire.dtl.DTLVardef@960159org.highwire.dtl.DTLVardef@328083_HPS_FORMAT_FIGEXP M_FIG Graphical abstract C_FIG

pharmacology and toxicology↗

bioRxiv2018

A standardised framework to identify optimal animal models for efficacy assessment in drug development

IntroductionPoor translation of efficacy data derived from animal models is a potential contributor to costly and unnecessary attrition in clinical trials.\n\nObjectivesTo develop a tool to assess, validate and compare the clinical translatability of animal models used for the preliminary assessment of efficacy.\n\nDesign and ResultsWe conducted an exploratory literature search to identify the key aspects to validate animal models. Eight aspects (Epidemiology, Pathophysiology, Genetic, Biochemistry, Aetiology, Histology, Pharmacology and Endpoints) were identified for which questions were drafted to evaluate the different faces of the human disease simulation. Features of the framework include standardised instructions, a weighting and scoring system to compare models as well as contextualising factors regarding model similarity and evidence uncertainty. We included a quality assessment of the internal validity of drug intervention studies included in the Pharmacological validation section for both effective and ineffective drugs in humans. A web-based survey was conducted with experts from different stakeholders to gather input on the framework. Finally, we present a case study of a preliminary validation and comparison of two animal models for Duchenne Muscular Dystrophy (mdx mouse and GRMD dog) and Diabetes Type 2 (ZDF rat and db/db mouse). We show that there are significant differences between the mdx mouse and the GRMD dog, the latter mimicking the human condition to a greater extent than the mouse despite the considerable lack of published data. In DT2, both the ZDF rat and the db/db mouse are comparable with minor differences in pathophysiology.\n\nConclusionsFIMD facilitates drug development by serving as the basis to select the most relevant model that can provide meaningful and translatable results to progress drug candidates to the clinic.

pharmacology and toxicology↗

bioRxiv2018

A non-canonical chemical feedback self-limits nitric oxide-cyclic GMP signaling in health and disease

Nitric oxide (NO)-cyclic GMP (cGMP) signaling is a vasoprotective pathway therapeutically targeted for example in pulmonary hypertension. Its dysregulation in disease is incompletely understood. Here we show in pulmonary artery endothelial cells that feedback inhibition by NO of the NO receptor, the cGMP forming soluble guanylate cyclase (sGC), may contribute to this. Both endogenous NO from endothelial NO synthase or exogenous NO from NO donor compounds decreased sGC protein and activity. This was not mediated by cGMP as the NO-independent sGC stimulator or direct activation of cGMP-dependent protein kinase did not mimic it. Thiol-sensitive mechanisms were also not involved as the thiol-reducing agent, N-acetyl-L-cysteine did not prevent this feedback. Instead, both in-vitro and in-vivo and in health and acute respiratory lung disease, chronically elevated NO led to the inactivation and degradation of sGC whilst leaving the heme-free isoform, apo-sGC, intact or even increasing its levels. Thus, NO regulates sGC in a bimodal manner, acutely stimulating and chronically inhibiting, as part of self-limiting direct feedback that is cGMP-independent. In high NO disease conditions, this is aggravated but can be functionally recovered in a mechanism-based manner by apo-sGC activators that re-establish cGMP formation.

pharmacology and toxicology↗

bioRxiv2018

In vitro determination of the CB1 efficacy of illicit synthetic cannabinoids

BACKGROUND AND PURPOSEThe morbidity and mortality associated with recreational use of synthetic cannabinoid receptor agonists (SCRAs) is a major health concern, and may involve over-activation of CB1 receptors. Thus, we sought to determine the efficacy of 13 SCRAs at CB1 using receptor depletion with the irreversible CB1 antagonist AM6544 followed by fitting the curve with the Black and Leff operational model to calculate efficacy.\n\nEXPERIMENTAL APPROACHReceptor depletion in mouse AtT-20 neuroblastoma cells stably expressing human CB1 was achieved by pre-treatment of cells with AM6544 (10 {micro}M, 60 mins). The CB1-mediated hyperpolarisation of AtT20 cells was measured using membrane potential dye. From data fit to the operational model, the efficacy (tau) and affinity (KA) parameters were obtained for each drug.\n\nKEY RESULTSAM6544 did not affect the potency or maximal effect of native somatostatin receptor-induced hyperpolarisation (Control, pEC50 9.13 {+/-} 0.05, Emax 38 {+/-} 1%; AM6544 treated pEC50 9.18 {+/-} 0.04, Emax 39 {+/-} 0.7%). The tau value of {triangleup}9-THC was 70-fold less than the reference CB-agonist CP55940, and 240-fold less than the highest efficacy SCRA, 5F-MDMB-PICA. Most of the SCRAs had about 50% of the efficacy of CP55940. There was no correlation between the tau and KA values for any SCRA.\n\nCONCLUSION AND IMPLICATIONSAll the SCRA tested showed substantially higher agonist activity at CB1 than {triangleup}9-THC, which may contribute to the adverse effects seen with these drugs but not {triangleup}9-THC, although the mechanisms underlying SCRA toxicity are still poorly defined.

pharmacology and toxicology↗

bioRxiv2018

Using selective lung injury to improve murine models of spatially heterogeneous lung diseases

Many lung diseases, such as acute respiratory distress syndrome (ARDS), display significant regional heterogeneity, with patches of severely injured tissue adjacent to apparently healthy tissue. Current mouse models that aim to mimic ARDS generally produce diffuse injuries that cannot reproducibly generate ARDSs regional heterogeneity. This deficiency prevents the evaluation of how well therapeutic agents reach the most injured regions, and precludes many regenerative medicine studies, since it is not possible to know which apparently healing regions suffered severe injury initially. Finally, these diffuse injury models must be mild to allow for survival, as their diffuse nature does not allow for residual healthy lung to keep an animal alive long enough for many drug and regenerative medicine studies. To solve all of these deficiencies of current animal models, we have created a simple and reproducible technique to selectively induce lung injury in specific areas of the lung. Our technique, catheter-in-catheter selective lung injury (CICSLI), involves guiding an inner catheter to a particular area of the lung and delivering an injurious agent mixed with nanoparticles (fluorescently and/or radioactively labeled) that can be used to track the location and extent of where the initial injury was, days later. Further, we demonstrate that CICSLI can produce a more severe injury than diffuse models, yet has much higher survival since CICSLI intentionally leaves undamaged lung regions. Collectively, these attributes of CICSLI will allow better study of how drugs act within heterogeneous lung pathologies and how regeneration occurs in severely damaged lung tissue, thereby aiding the development of new therapies for ARDS and other lung diseases.

pharmacology and toxicology↗

bioRxiv2018

LRRK2 kinase inhibitors induce a reversible effect in the lungs of non-human primates with no measurable pulmonary deficits

Putative gain-of-function mutations in leucine-rich repeat kinase 2 (LRRK2), resulting in increased kinase activity and cellular toxicity, are a leading genetic cause of Parkinsons disease (PD). Hence, there is strong interest in developing LRRK2 kinase inhibitors as a disease-modifying therapy. Published reports that repeat dosing with two LRRK2 kinase inhibitors (GNE-7915 and GNE-0877) induce histopathological changes in the lung of non-human primates Fuji et al. 2015 (1) raised concerns about potential safety liability of LRRK2 kinase inhibitors. In the present study, we sought to determine whether previously observed effects in the lung: (a) represent on-target pharmacology, but with the potential for margin of safety, (b) are reversible upon drug withdrawal, and (c) are associated with pulmonary function deficits. To this end, we evaluated the histopathological effects, toxicokinetics and target inhibition of three structurally diverse LRRK2 kinase inhibitors, GNE-7915 (30 mg/kg, BID, as a positive control), MLi-2 (15 and 50 mg/kg, QD) and PFE-360 (3 and 6 mg/kg, QD) following 2 weeks of dosing in non-human primates. Subsets of animals dosed with GNE-7915 or MLi-2 were evaluated after 2-week dose-free periods. All three LRRK2 kinase inhibitors induced mild cytoplasmic vacuolation of type II pneumocytes, as reported previously, confirming an on-target effect of these compounds. Interestingly, despite lower doses of both PFE-360 and MLi-2 producing nearly complete inhibition of LRRK2 kinase activity in the brain as assessed by levels of pS935-LRRK2, histopathological changes in lung were absent in animals treated with low-dose PFE-360 and observed only sporadically in the low-dose MLi-2 group. The lung effect was fully reversible at 2 weeks post-dosing of GNE-7915. In a second study of identical dosing with MLi-2 and GNE-7915, no deficits were observed in a battery of translational pulmonary functional tests. In aggregate, these results do not preclude the development of LRRK2 kinase inhibitors for clinical investigation in Parkinsons disease.

pharmacology and toxicology↗