Comparative biochemical and molecular responses of biotransformation and antioxidant systems in three species of Crassostrea (Sacco, 1897) oysters exposed to chrysene.

Chrysene (CHR) is among the most persistent polycyclic aromatic hydrocarbons (PAH) in water and a priority compound for pollutants monitoring, due to its carcinogenic, mutagenic and genotoxic potential. Aquatic animals exposed to CHR may present alterations of biomarkers involved in the biotransformation and oxidative stress-related parameters. The aim of this study was to investigate differences in antioxidant and biotransformation (phase I and II) systems of Crassostrea gigas, C. gasar and C. rhizophorae and its effects resulting from CHR exposure. Adult oysters of these species were exposed to 10 µg L-1 of CHR for 24 h and 96 h. In gills, the transcripts CYP1-like, CYP2-like, CYP2AU1-like, GSTO-like, MGST-like, SULT-like were evaluated after 24 h of exposure. The activity of SOD, CAT, GPx, GR and G6PDH were analyzed in gills and digestive glands after 96 h of exposure. CHR bioaccumulated in tissues. Differences in the remaining levels of CHR in water after 96 h were observed in aquaria containing C. gigas or C. gasar oysters and may be associated to the different filtration rates between these species. Downregulate of biotransformation genes were observed in gills of C. gasar (CYP2AU1-like and GSTO-like) and C. rhizophorae (CYP1-like1, CYP2-like, MGST-like and SULT-like), suggesting that biotransformation responses may be species-specific. Differential activity of antioxidant enzymes were observed in gills and digestive gland of oysters exposed to CHR. Biochemical responses suggested that C. gigas and C. gasar are more responsive to CHR. Differential responses observed among the three Crassostrea species can be related to evolutionary differences, ecological niches and adaptation to environment.

The exposure to polyvinyl chloride microplastics and chrysene induces multiple changes in the structure and functionality of marine meiobenthic communities.

The effects of microplastics and sorbed polycyclic aromatic hydrocarbons at community levels were rarely assessed in laboratory experiments, despite their obvious advantage in reflecting better the natural conditions compared to traditionally single species-focused toxicological experiments. In the current study, the multifaceted effects of polyvinyl chloride and chrysene, acting alone or combined, on general marine meiobenthos, but with a special focus on free-living marine nematode communities were tested in a laboratory experiment carried in microcosms. The meiobenthos was exposed to two polyvinyl chloride (5 and 10 mg.kg-1 Dry Weight 'DW') and chrysene (37.5 and 75 ng.g-1 DW) concentrations, respectively, as well as to a mixture of both compounds, for 30 days. The results highlighted a significant decrease in the abundance of all meiobenthic generic groups, including nematodes, directly with increasing dosages of these compounds when added alone. The addition of chrysene adheres to microplastics, making the sediment matrix glueyer, hence inducing greater mortality among generic meiobenthic groups. Moreover, the nematofauna went through a strong restructuring phase following the exposure to both compounds when added alone, leading to the disappearance of sensitive nematodes and their replacement with tolerant taxa. However, the similarity in nematofauna composition between control and polyvinyl chloride and chrysene mixtures suggests that the toxicity of the latter could be attenuated by its physical bonding to the former pollutant. Other changes in the functional traits within the nematode communities were a decline in the fertility of females and an increase of the pharyngeal pumping power following exposure to both pollutants for the dominant species. The latter results were also supported by additional toxicokinetics analyses and in silico modeling.

Are Myths and Preconceptions Preventing us from Applying Ionic Liquid Forms of Antiviral Medicines to the Current Health Crisis?

At the moment, there are no U.S. Food and Drug Administration (U.S. FDA)-approved drugs for the treatment of COVID-19, although several antiviral drugs are available for repurposing. Many of these drugs suffer from polymorphic transformations with changes in the drug's safety and efficacy; many are poorly soluble, poorly bioavailable drugs. Current tools to reformulate antiviral APIs into safer and more bioavailable forms include pharmaceutical salts and cocrystals, even though it is difficult to classify solid forms into these regulatory-wise mutually exclusive categories. Pure liquid salt forms of APIs, ionic liquids that incorporate APIs into their structures (API-ILs) present all the advantages that salt forms provide from a pharmaceutical standpoint, without being subject to solid-state matter problems. In this perspective article, the myths and the most voiced concerns holding back implementation of API-ILs are examined, and two case studies of API-ILs antivirals (the amphoteric acyclovir and GSK2838232) are presented in detail, with a focus on drug property improvement. We advocate that the industry should consider the advantages of API-ILs which could be the genesis of disruptive innovation and believe that in order for the industry to grow and develop, the industry should be comfortable with a certain element of risk because progress often only comes from trying something different.

Indeno[1,2,3-cd]pyrene and picene mediate actions via estrogen receptor α signaling pathway in in vitro cell systems, altering gene expression.

Currently, the environmental impact of ubiquitous plastic debris triggered quite some public attention. However, the global impact of microplastic on human health is by and large either unknown or neglected. By looking at the underlying biochemical mechanisms leading to the global health threat microplastic was discovered to carry persistent organic pollutants, such as polycyclic aromatic hydrocarbons (PAH), to marine life. The effect of microplastic-ingestion in the human body remains unfortunately somewhat elusive as of yet. For this reason, we screened for compounds binding to the human estrogen receptor α (ERα) and identified the PAH compounds indeno[1,2,3-cd]pyrene (Indpy) and picene (Pice) with a high binding affinity. We applied next generation sequencing to analyze the differentially expressed genes in MCF-7 cells after treatment with Indpy and Pice. We found 8 upregulated genes: ABCC5, CCNG2, CYP1A1, DDIT4, IER3, RUNX2, STC2, and SLC7A5 and 14 downregulated genes: ADORA1, CEBPB, CELSR2, CTSD, CXCL12, KRT19, PGR, PKIB, RARA, RET, SEMA3B, SIAH2, TFAP2C, and XBP1 induced by both ligands and associated with ESR1-regulation. The altered gene expression may influence cell proliferation and metastasis, favoring cancer development with a poor response to therapy. In addition, we confirmed the binding of Indpy and Pice to ERα using molecular docking and microscale thermophoresis. ERα activation was measured with ESR1-overexpressing HEK293 (HEK-ESR1) cells and confirmed for Indpy. In conclusion, we showed an ESR1-mediated influence of the PAH compounds Indpy and Pice on the gene expression pattern of MCF-7 cells, possibly also promoting breast cancer development in patients.

Second generation of diazachrysenes: Protection of Ebola virus infected mice and mechanism of action.

Ebola virus (EBOV) causes a deadly hemorrhagic fever in humans and non-human primates. There is currently no FDA-approved vaccine or medication to counter this disease. Here, we report on the design, synthesis and anti-viral activities of two classes of compounds which show high potency against EBOV in both in vitro cell culture assays and in vivo mouse models Ebola viral disease. These compounds incorporate the structural features of cationic amphiphilic drugs (CAD), i.e they possess both a hydrophobic domain and a hydrophilic domain consisting of an ionizable amine functional group. These structural features enable easily diffusion into cells but once inside an acidic compartment their amine groups became protonated, ionized and remain trapped inside the acidic compartments such as late endosomes and lysosomes. These compounds, by virtue of their lysomotrophic functions, blocked EBOV entry. However, unlike other drugs containing a CAD moiety including chloroquine and amodiaquine, compounds reported in this study display faster kinetics of accumulation in the lysosomes, robust expansion of late endosome/lysosomes, relatively more potent suppression of lysosome fusion with other vesicular compartments and inhibition of cathepsins activities, all of which play a vital role in anti-EBOV activity. Furthermore, the diazachrysene 2 (ZSML08) that showed most potent activity against EBOV in in vitro cell culture assays also showed significant survival benefit with 100% protection in mouse models of Ebola virus disease, at a low dose of 10 mg/kg/day. Lastly, toxicity studies in vivo using zebrafish models suggest no developmental defects or toxicity associated with these compounds. Overall, these studies describe two new pharmacophores that by virtue of being potent lysosomotrophs, display potent anti-EBOV activities both in vitro and in vivo animal models of EBOV disease.

Design, Synthesis, and SAR of C-3 Benzoic Acid, C-17 Triterpenoid Derivatives. Identification of the HIV-1 Maturation Inhibitor 4-((1 R,3a S,5a R,5b R,7a R,11a S,11b R,13a R,13b R)-3a-((2-(1,1-Dioxidothiomorpholino)ethyl)amino)-5a,5b,8,8,11a-pentamethyl-1-(prop-1-en-2-yl)-2,3,3a,4,5,5a,5b,6,7,7a,8,11,11a,11b,12,13,13a,13b-octadecahydro-1 H-cyclopenta[ a]chrysen-9-yl)benzoic Acid (GSK3532795, BMS-955176).

GSK3532795, formerly known as BMS-955176 (1), is a potent, orally active, second-generation HIV-1 maturation inhibitor (MI) that advanced through phase IIb clinical trials. The careful design, selection, and evaluation of substituents appended to the C-3 and C-17 positions of the natural product betulinic acid (3) was critical in attaining a molecule with the desired virological and pharmacokinetic profile. Herein, we highlight the key insights made in the discovery program and detail the evolution of the structure-activity relationships (SARs) that led to the design of the specific C-17 amine moiety in 1. These modifications ultimately enabled the discovery of 1 as a second-generation MI that combines broad coverage of polymorphic viruses (EC50 <15 nM toward a panel of common polymorphisms representative of 96.5% HIV-1 subtype B virus) with a favorable pharmacokinetic profile in preclinical species.

Novel flexible heteroarotinoid, SL-1-18, promotes ERα degradation to inhibit breast cancer cell growth.

SL-1-18 (1-(chrysen-6-yl)-3-(4-nitrophenyl)thiourea) is new flexible heteroarotinoid (Flex-Het) analog derived from the parent compound, SHetA2, and our previous study showed comparable activity to SHetA2 in terms of inhibiting ER+ breast cancer cell growth. This current study aims to determine the molecular mechanism underlying SL-1-18's effect on breast cancer cell growth. Our results indicate that SL-1-18 inhibits cell proliferation of ER+ breast cancer cells (MCF-7 and T-47D) by preventing cell cycle progression. SL-1-18 treatment correlated positively with decreased expression of key cell-cycle regulators, such as cyclin D1, as well as other ERα-target genes at both the transcript and protein levels. Interestingly, decreased expression of ERα was also observed, with a significant reduction at the protein level within 2 h of SL-1-18 treatment, while the decrease in mRNA occurred at a later time point. ERα degradation was shown to be mediated by the ubiquitination-proteasome pathway. In summary, this is the first study to show that a Flex-Het- SL-1-18- can promote the degradation of ERα via the ubiquitin-proteasome pathway and should be further developed as a therapeutic option for ER+ breast cancer.

An unlikely DNA cleaving agent: A photo-active trinuclear Cu(II) complex based on hexaazatriphenylene.

This paper describes the synthesis of a trinuclear Cu(II) complex (4) containing a central 1,4,5,8,9,12-hexaazatriphenylene-hexacarboxylate (hat) core (3). Low, micromolar concentrations of the negatively charged parent ligand 3 and the neutral trinuclear complex 4 were found to photocleave negatively charged pUC19 plasmid DNA with high efficiency at neutral pH (350nm, 50min, 22°C). The interactions of complex 4 with double-helical DNA were studied in detail. Scavenger and colorimetric assays pointed to the formation of Cu(I), superoxide anion radicals, hydrogen peroxide, and hydroxyl radicals during photocleavage reactions. UV-visible absorption, circular dichroism, DNA thermal denaturation, and fluorescence data suggested that the Cu(II) complex contacts double-stranded DNA in an external fashion. The persistent association of ligand 3 and complex 4 with Na(I) and/or other cations in aqueous solution might facilitate electrostatic DNA interactions.

Inhibition of dengue virus replication by diisopropyl chrysin-7-yl phosphate.

Dengue fever is a tropical disease and caused by dengue virus (DENV), which is transmitted by mosquitoes and infects about 400 million people annually. With the development of international trade and travel, China is facing a growing threat. Over 40 thousands of people were infected during the 2014 DENV outbreak in Guangdong. Neither licensed vaccine nor therapeutic drug has been available. In this report, we isolated two clinical DENV strains. The full-length genome was sequenced and characterized. We also applied a flavonoid, CPI, into an anti-DENV assay. Replication of viral RNA and expression of viral protein was all strongly inhibited. These results indicated that CPI may serve as potential protective agents in the treatment of patients with chronic DENV infection.

n-3 Fatty acids regulate the inflammatory-state related genes in the lung epithelial cells exposed to polycyclic aromatic hydrocarbons.

BACKGROUND: Chronic airway inflammation is coordinated by a complex of inflammatory mediators, including eicosanoids. The aim of this study was to evaluate the impact of polycyclic aromatic hydrocarbons (PAHs) on the human lung epithelial carcinoma A549 cells supplemented with docosahexaenoic (DHA) and eicosapentaenoic (EPA) acids. METHODS: We analyzed the influence of DHA, EPA and/or benzo(a)pyrene (BaP), chrysene (Chr), fluoranthene (Flu) and benzo(a)anthracene (Baa) treatment on the fatty acids (FAs) profile and the formation of isoprostanes. We studied the cyclooxygenase-2, FP-receptor, peroxisome proliferator-activated receptors PPARδ and PPARγ, transcription factor NF-кB p50 and p65 expression by Western blot, phospholipase A2 (cPLA2) activity, as well as aryl hydrocarbon receptor (AHR), cytochrome P450 (CYP1A1), phospholipase A2 (PLA2G4A) and prostaglandin synthase 2 (PTGS2) gene expression by qRT-PCR. RESULTS: DHA or EPA supplementation and BaP or Baa treatment resulted in a higher level of PGF3α. COX-2 expression was decreased while PPARδ expression and cPLA2 activity was increased after fatty acid supplementation and PAHs treatment. DHA and EPA up-regulated AHR and PLA2G4A genes. CONCLUSIONS: Supplementation with n-3 FAs resulted in changes of inflammatory-state related genes in the lung epithelial cells exposed to PAHs. The altered profile of lipid mediators from n-3 FA as well as repression of the COX-2 protein by n-3 PUFAs in A549 cells incubated with PAHs suggests anti-inflammatory and pro-resolving properties of DHA and EPA. It remains to be shown whether these pleiotropic and protective actions of n-3 FAs contribute to fish oil's therapeutic effect in asthma.

Anti-enterovirus 71 effects of chrysin and its phosphate ester.

Enterovirus 71 (EV71) can cause severe disease and even lead to death in children, and an effective antiviral drug is currently unavailable. The anti-EV71 effect of chrysin (5,7-dihydroxyflavone), a natural flavonoid commonly found in many plants, was tested in this report. By using the predicting program Autodock 4.0 and an in vitro protease inhibition assay, we found that chrysin could suppress viral 3Cpro activity. Replication of viral RNA and production of viral capsid protein and the infectious virion were strongly inhibited by chrysin, without noticeable cytotoxicity. Cytopathic effects on cells were also prevented. Diisopropyl chrysin-7-yl phosphate (CPI), the phosphate ester for chrysin, was generated through a simplified Atheron-Todd reaction to achieve stronger anti-viral activity. CPI was also able to bind with and inhibit viral 3Cpro activity in vitro. As expected, CPI demonstrated more potent antiviral activity against EV71.

Mutagenicity and tumorigenicity of the four enantiopure bay-region 3,4-diol-1,2-epoxide isomers of dibenz[a,h]anthracene.

Each enantiomer of the diastereomeric pair of bay-region dibenz[a,h]anthracene 3,4-diol-1,2-epoxides in which the benzylic 4-hydroxyl group and epoxide oxygen are either cis (isomer 1) or trans (isomer 2) were evaluated for mutagenic activity. In strains TA 98 and TA 100 of Salmonella typhimurium, the diol epoxide with (1S,2R,3S,4R) absolute configuration [(-)-diol epoxide-1] had the highest mutagenic activity. In Chinese hamster V-79 cells, the diol epoxide with (1R,2S,3S,4R) absolute configuration [(+)-diol epoxide-2] had the highest mutagenic activity. The (1R,2S,3R,4S) diol epoxide [(+)-diol epoxide-1] also had appreciable activity, whereas the other two bay-region diol epoxide enantiomers had very low activity. In tumor studies, the (1R,2S,3S,4R) enantiomer was the only diol epoxide isomer tested that had strong activity as a tumor initiator on mouse skin and in causing lung and liver tumors when injected into newborn mice. This stereoisomer was about one-third as active as the parent hydrocarbon, dibenz[a,h]anthracene as a tumor initiator on mouse skin; it was several-fold more active than dibenz[a,h]anthracene as a lung and liver carcinogen when injected into newborn mice. (-)-(3R,4R)-3ß,4α-dihydroxy-3,4-dihydro-dibenz[a,h]anthracene [(-)-3,4-dihydrodiol] was slightly more active than dibenz[a,h]anthracene as a tumor initiator on mouse skin, whereas (+)-(3S,4S)-3α,4ß-dihydroxy-3,4-dihydro-dibenz[a,h]anthracene [(+)-3,4-dihydrodiol] had only very weak activity. The present investigation and previous studies with the corresponding four possible enantiopure bay-region diol epoxide enantiomers/diastereomers of benzo[a]pyrene, benz[a]anthracene, chrysene, benzo[c]phenanthrene, dibenz[c,h]acridine, dibenz[a,h]acridine and dibenz[a,h]anthracene indicate that the bay-region diol epoxide enantiomer with [R,S,S,R] absolute stereochemistry has high tumorigenic activity on mouse skin and in newborn mice.

High throughput screening of 7-methylpicene-1,2-diol as arylamine N-acetyltransferase (NAT) inhibitor to establish a isoniazid supplement in anti-tubercular therapy.

Mycobacterium tuberculosis (Mtb), due to its unusual organization crosses different immune barriers and causes tuberculosis. The advent of multidrug resistance tuberculosis (MDR-TB) has attained alarming situation. Hence, computational drug design has been performed in this work to find potent molecules for this purpose. Isoniazid is a widely used frontline drug against tuberculosis. But reports justified the inactivity of isoniazid on acetylation by Arylamine N-acetyltransferase (NAT). 35 countries were highlighted to have isoniazid resistance from survey in 1998. Hence, Mtb NAT has been selected as the target in the present case and hundred compounds were screened in order to find potent NAT inhibitor to raise the efficacy of isoniazid. Molecular docking with Biosolveit LeadIT and Autodock 4.2 simulation was performed. The result showed 7- methylpicene-1, 2-diol to have -26.77 and -8.26 kcal/mol score in LeadIT and Autodock 4.2. The work validated 7- methylpicene-1, 2-diol to be a potent NAT inhibitor to supplement isoniazid.

Mesoscale DNA structural changes on binding and photoreaction with Ru[(TAP)2PHEHAT]2+.

We used scanning force microscopy (SFM) to study the binding and excited state reactions of the intercalating photoreagent Ru[(TAP)(2)PHEHAT](2+) (TAP = 1,4,5,8-tetraazaphenanthrene; PHEHAT = 1,10-phenanthrolino[5,6-b]1,4,5,8,9,12-hexaazatriphenylene) with DNA. In the ground state, this ruthenium complex combines a strong intercalative binding mode via the PHEHAT ligand, with TAP-mediated hydrogen bonding capabilities. After visible irradiation, SFM imaging of the photoproducts revealed both the structural implications of photocleavages and photoadduct formation. It is found that the rate of photocleaving is strongly increased when the complex can interact with DNA via hydrogen bonding. We demonstrated that the photoadduct increases DNA rigidity, and that the photo-biadduct can crosslink two separate DNA segments in supercoiled DNA. These mechanical and topological effects might have important implications in future therapeutic applications of this type of compounds.

The effect of selective oestrogen receptor antagonists in an in vitro model of growth plate chondrogenesis.

While oestrogen is recognized to play a key role in regulating growth, particularly in relation to epiphyseal fusion, the mechanisms that mediate its effects are still unclear. We utilized an in vitro model of chondrogenesis, the RCJ3.1C5.18 cell line, to explore the effect of oestrogen on this process. We demonstrated the presence of oestrogen receptors (ER) α and ß in these cells, with increased abundance of both receptor sub-types evident as the cells differentiated. ERα localized to the nucleus, suggesting it was signalling by genomic pathways, while ERß was seen predominantly in the cytoplasm, suggesting it may be utilizing non-genomic signalling. While exogenous oestrogen had no effect on proliferation or differentiation, we found some evidence for the endogenous production of oestrogen (intracrinology), as suggested by the expression of aromatase in these cells. Selective ERα blockade with methyl piperidinopyrazole (MPP) led to a significant reduction in both proliferation and differentiation, while ERß blockade with R,R tetrahydrochrysene (THC) led to an increase in these parameters. This is in keeping with results from mouse knockout models suggesting that unopposed ERß signalling leads to an inhibition of skeletal growth. Our results are further evidence for the importance of differential ER signalling in regulating chondrogenesis. Future studies examining in vivo effects of these agents are required to extrapolate these findings to a mammalian model.

A chemotype that inhibits three unrelated pathogenic targets: the botulinum neurotoxin serotype A light chain, P. falciparum malaria, and the Ebola filovirus.

A 1,7-bis(alkylamino)diazachrysene-based small molecule was previously identified as an inhibitor of the botulinum neurotoxin serotype A light chain metalloprotease. Subsequently, a variety of derivatives of this chemotype were synthesized to develop structure-activity relationships, and all are inhibitors of the BoNT/A LC. Three-dimensional analyses indicated that half of the originally discovered 1,7-DAAC structure superimposed well with 4-amino-7-chloroquinoline-based antimalarial agents. This observation led to the discovery that several of the 1,7-DAAC derivatives are potent in vitro inhibitors of Plasmodium falciparum and, in general, are more efficacious against CQ-resistant strains than against CQ-susceptible strains. In addition, by inhibiting ß-hematin formation, the most efficacious 1,7-DAAC-based antimalarials employ a mechanism of action analogous to that of 4,7-ACQ-based antimalarials and are well tolerated by normal cells. One candidate was also effective when administered orally in a rodent-based malaria model. Finally, the 1,7-DAAC-based derivatives were examined for Ebola filovirus inhibition in an assay employing Vero76 cells, and three provided promising antiviral activities and acceptably low toxicities.

Novel 6,12-disubstituted chrysene as potent anticancer agent: synthesis, in vitro and in vivo study.

We describe herein the synthesis of novel 6,12-distributed chrysene as potent anticancer agents. In vitro and in vivo studies are also reported here.

Neuroprotective effects of R,R-tetrahydrochrysene against glutamate-induced cell death through anti-excitotoxic and antioxidant actions involving estrogen receptor-dependent and -independent pathways.

Glutamate-induced neural cell death is mediated by excitotoxicity and oxidative stress. Treatment of glutamate toxicity with estrogen and its related compounds for neuroprotection remains controversial. In this study, we examined the effects of selective estrogen receptor (ER) ligands on glutamate toxicity and found that R,R-tetrahydrochrysene (R,R-THC), an antagonist of ERbeta and agonist of ERalpha, has neuroprotective effects against glutamate-induced death in primary rat cortical cells and mouse N29/4 hypothalamic cells. The protective effect of R,R-THC was dose-dependent and was maintained even when added several hours after the initial glutamate exposure. R,R-THC blocked glutamate-induced depletion of intracellular glutathione, increased superoxide dismutase activity, and protected cells from hydrogen peroxide-induced death. R,R-THC also prevented glutamate-induced nuclear translocation of apoptotic inducing factor and release of mitochondrial cytochrome c. The protective effect of R,R-THC was blocked by methyl-piperidino-pyrazole (MPP; an ERalpha antagonist) in glutamate-treated cortical cells, and pretreatment with MK-801 (an NMDA receptor antagonist) but not CNQX (an AMPA/kainate receptor antagonist) increased cell survival. On the other hand, MPP did not block the protective effect of R,R-THC in glutamate-treated N29/4 cells, and neither MK-801 nor CNQX conferred protection. Activation of ERalpha and/or ERbeta with 17beta-estradiol (E2), propyl-pyrazole-triol or diarylpropionitrile did not provide effective neuroprotection, and pretreatment with ICI 182,780 did not inhibit the protective effect of R,R-THC in either type of cell. These results suggest that the use of ER agonists (including E2) has limited beneficial effects when both excitotoxicity and oxidative stress occur. In contrast to agonists of ERs, R,R-THC, which possesses anti-excitotoxic and antioxidant actions via ER-dependent and -independent pathways, provides significant neuroprotection.

A comparative study on the uptake of polycyclic aromatic hydrocarbons by Anodonta californiensis.

Uptake of polycyclic aromatic hydrocarbons (PAHs) by the freshwater bivalve mollusc Anodonta californiensis was examined in the presence and absence of surfactant in order to gain further insight into mixture toxicity and to predict whether certain mixtures have negative and/or positive effects on aquatic organisms. In the presence of surfactant, the uptake of anthracene or chrysene was higher than that of naphthalene, given the same concentration in the solution. In the absence of surfactant, the trend was similar, but the uptakes were increased by approximately 100% compared to those in the presence of surfactant. On the uptake of naphthalene, the presence of anthracene showed only minor influence. The uptake of anthracene was affected by both naphthalene and chrysene. The uptake of chrysene was influenced by neither naphthalene nor anthracene. There was no observable displacement of divalent cations from the surface of the gill membrane by any of the PAHs studied.

Three-dimensional database mining identifies a unique chemotype that unites structurally diverse botulinum neurotoxin serotype A inhibitors in a three-zone pharmacophore.

A search query consisting of two aromatic centers and two cationic centers was defined based on previously identified small molecule inhibitors of the botulinum neurotoxin serotype A light chain (BoNT/A LC) and used to mine the National Cancer Institute Open Repository. Ten small molecule hits were identified, and upon testing, three demonstrated inhibitory activity. Of these, one was structurally unique, possessing a rigid diazachrysene scaffold. The steric limitations of the diazachrysene imposed a separation between the overlaps of previously identified inhibitors, revealing an extended binding mode. As a result, the pharmacophore for BoNT/A LC inhibition has been modified to encompass three zones. To demonstrate the utility of this model, a novel three-zone inhibitor was mined and its activity was confirmed.