Sublethal effects of niclosamide on the aquatic snail Pomacea canaliculata.

Pomacea canaliculata is a malignant invasive aquatic snail found worldwide, and niclosamide (NS) is one of the primary agents used for its control. NS applied to water will exist in non-lethal concentrations for some time due to degradation or water exchange, thus resulting in sublethal effects on environmental organisms. To identify sublethal effects of NS on Pomacea canaliculata, we studied the aspects of histopathology, oxygen-nitrogen ratio (RO∶N), enzyme activity determination, and gene expression. After LC30 NS treatment (0.310 g/L), many muscle fibers of the feet degenerated and some acinar vesicles of the hepatopancreas collapsed and dissolved. The oxygen-nitrogen ratio (RO∶N) decreased significantly from 15.0494 to 11.5183, indicating that NS had changed the metabolic mode of Pomacea canaliculata and shifted it primarily to protein catabolism. Transcriptome analysis identified the sublethal effects of LC30 NS on the snails at the transcriptional level. 386, 322, and 583 differentially expressed genes (DEGs) were identified in the hepatopancreas, gills, and feet, respectively. GO (Gene Ontology) functional analysis and KEGG (Kyoto Encyclopedia of Genes and Genomes) pathway annotations showed that DEGs in the hepatopancreas were mainly enriched for sugar metabolism, protein biosynthesis, immune response, and amino acid metabolism functional categories; DEGs in the gills were mainly enriched for ion transport and amino acid metabolism; DEGs in the feet were mainly enriched for transmembrane transport and inositol biosynthesis. In the future, we will perform functional validation of key genes to further explain the molecular mechanism of sublethal effects.

Disturbances to energy metabolism in juvenile lake sturgeon (Acipenser fulvescens) following exposure to niclosamide.

Since the 1960s, invasive sea lamprey (Petromyzon marinus) populations in the Laurentian Great Lakes have been controlled by applying two chemicals, 3-trifluoromethyl-4-nitrophenol (TFM) and 2',5-dichloro-4'-nitrosalicylanilide (niclosamide, aka. Bayluscide®), to streams infested with larval sea lamprey. These "lampricide" applications primarily rely on TFM, and are often combined with 1-2% niclosamide, which increases treatment effectiveness. Niclosamide is also used alone to treat lentic habitats and in rivers with high discharge. However, little is known about niclosamide's possible adverse physiological effects on non-target organisms. Of particular concern is the lake sturgeon (Acipenser fulvescens), which is threatened throughout the Great Lakes basin where its habitat often overlaps with larval lamprey. Because niclosamide is believed to impair ATP production by uncoupling oxidative phosphorylation, we determined how it altered metabolic processes and acid-base balance in young-of-the-year (YOY) lake sturgeon exposed to their 9-h LC50 of niclosamide (0.11 mg L-1) for 9 h. Exposure to niclosamide led to decreased brain ATP and glucose reserves, and increased lactate, with no effect on brain glycogen. In contrast, substantial (60%) reductions in glycogen were observed in liver, suggesting that hepatic glycogen reserves were mobilized to meet the brain's glucose requirements when ATP supply was impaired during niclosamide exposure. Disturbances in carcass included reduced phosphocreatine (65-70%), 2- and 4-fold increases in pyruvate and lactate, and a slight metabolic acidosis, characterized by a 0.1 unit decrease in intracellular pH (pHi). Each of these disturbances were corrected within 24 h following depuration in clean (niclosamide-free) water. We conclude that if lake sturgeon survive exposure to niclosamide, they are able to rapidly replenish their energy stores (glycogen, ATP, phosphocreatine) and correct any corresponding metabolic disturbances within 24 h.

Chemical mitochondrial uncouplers share common inhibitory effect on NLRP3 inflammasome activation through inhibiting NFκB nuclear translocation.

Activation of NLRP3 inflammasome is implicated in varieties of pathologies, the aim of the present study is to characterize the effect and mechanism of mitochondrial uncouplers on NLRP3 inflammasome activation by using three types of uncouplers, niclosamide, CCCP and BAM15. Niclosamide, CCCP and BAM15 inhibited LPS plus ATP-induced increases of NLRP3 protein and IL-1ß mRNA levels in RAW264.7 macrophages and THP-1 derived macrophages. Niclosamide, CCCP and BAM15 inhibited LPS plus ATP-induced increase of NFκB (P65) phosphorylation, and inhibited NFκB (P65) nuclear translocation in RAW264.7 macrophages. Niclosamide and BAM15 inhibited LPS-induced increase of IκBα phosphorylation in RAW264.7 macrophages, and the inhibitory effect was dependent on increased intracellular [Ca2+]i; however, CCCP showed no significant effect on IκBα phosphorylation in RAW264.7 macrophages stimulated with LPS. In conclusion, chemical mitochondrial uncouplers niclosamide, CCCP and BAM15 share common inhibitory effect on NLRP3 inflammasome activation through inhibiting NFκB nuclear translocation.

Maternal-to-zygotic transition as a potential target for niclosamide during early embryogenesis.

Niclosamide is an antihelminthic drug used worldwide for the treatment of tapeworm infections. Recent drug repurposing screens have highlighted the broad bioactivity of niclosamide across diverse mechanisms of action. As a result, niclosamide is being evaluated for a range of alternative drug-repurposing applications, including the treatment of cancer, bacterial infections, and Zika virus. As new applications of niclosamide will require non-oral delivery routes that may lead to exposure in utero, it is important to understand the mechanism of niclosamide toxicity during early stages of embryonic development. Previously, we showed that niclosamide induces a concentration-dependent delay in epiboly progression in the absence of effects on oxidative phosphorylation - a well-established target for niclosamide. Therefore, the overall objective of this study was to further examine the mechanism of niclosamide-induced epiboly delay during zebrafish embryogenesis. Based on this study, we found that (1) niclosamide exposure during early zebrafish embryogenesis resulted in a decrease in yolk sac integrity with a concomitant decrease in the presence of yolk sac actin networks and increase in cell size; (2) within whole embryos, niclosamide exposure did not alter non-polar metabolites and lipids, but significantly altered amino acids specific to aminoacyl-tRNA biosynthesis; (3) niclosamide significantly altered transcripts related to translation, transcription, and mRNA processing pathways; and (4) niclosamide did not significantly alter levels of rRNA and tRNA. Overall, our findings suggest that niclosamide may be causing a systemic delay in embryonic development by disrupting the translation of maternally-supplied mRNAs, an effect that may be mediated through disruption of aminoacyl-tRNA biosynthesis.

How do microplastics alter molluscicidal activity? Effects of weathered microplastics and niclosamide in developing freshwater snails.

Despite the wide distribution and persistence of microplastics (MPs), their interactive effects with molluscicides are unknown. Schistosomiasis, a neglected tropical disease, affects 236.6 million people worldwide. Niclosamide (NCL) is the only molluscicide recommended by the World Health Organization (WHO) and it is used to control the population of Schistosoma spp.'s intermediate host. Thus, this study aimed to evaluate of the interaction between polyethylene (PE) MPs and NCL, and their associated toxicity in the freshwater snail Biomphalaria glabrata (Say 1818). Weathered PE MPs were characterized and theoretical analysis of NCL-MP adsorption nature was made using quantum mechanical calculations. The toxicity of NCL isolated (0.0265 to 0.0809 mg L-1) and under interaction with PE MPs (3400 µg L-1) in B. glabrata embryos and newly hatched snails was analyzed. In silico analysis confirmed the adsorption mechanisms of NCL into PE MPs. PE MPs decreased the NCL toxicity to both B. glabrata developmental stages, increasing their survival and NCL lethal concentrations, indicating concerns regarding NCL use as molluscicide in aquatic environments polluted by MPs. In conclusion, MPs may change the efficiency of chemicals used in snail control programs.

Presumed veterinary niclosamide-induced retinal toxicity in a human: a case report.

BACKGROUND: To report the first case of bull's eye maculopathy associated with veterinary niclosamide. CASE PRESENTATION: A 27-year-old Iranian female presented with a history of reduced vision and photopsia since 3 years, after accidental ingestion of four boluses of veterinary niclosamide. Fundus examination showed atrophy in parafoveal retinal pigmentary epithelium, appearing as bilateral bull's-eye maculopathy. Optical coherence tomography revealed disruption of the parafoveal ellipsoid zone and outer retinal thinning, appearing as a flying saucer sign. Electroretinography displayed decreased scotopic and photopic amplitudes with normal waveform in both eyes. The causality score was 4, showing "possible" retinopathy due to niclosamide according to Naranjo's causality assessment algorithm. Based on clinical and ancillary findings, a diagnosis of niclosamide-induced maculopathy was made. CONCLUSION: Veterinary niclosamide is an anthelmintic drug that in higher doses could be detrimental to the human retina. Awareness about its side effects and appropriate drug labeling could prevent accidental toxicity.

Exposition of Intermediate Hosts of Schistosomes to Niclosamide (Bayluscide WP 70) Revealed Significant Variations in Mortality Rates: Implications for Vector Control.

(1) Background: Schistosomiasis remains a public health issue in Cameroon. Snail control using Niclosamide can prevent schistosome transmission. It is safe to determine lethal concentrations for the population. This study aimed at assessing the toxicity of Niclosamide on different developmental stages of snail populations; (2) Methods: Snails were collected, identified, and reared in the laboratory. Egg masses and adult snails were exposed to Niclosamide, at increasing concentrations (0.06, 0.125, 0.25, 0.5, 1 mg/L for egg embryos and 0.06, 0.08, 0.1, 0.12, 0.14, 0.16, 0.18, 0.2 mg/L for adults). After 24 h exposure, egg masses and snails were removed from Niclosamide solutions, washed with source water and observed; (3) Results: Snail susceptibility was species and population dependent. For egg embryos, Biomphalaria pfeifferi was the most susceptible (LC50: 0.1; LC95: 6.3 mg/L) and Bulinus truncatus the least susceptible (LC50: 4.035; LC95: 228.118 mg/L). However, for adults, B. truncatus was the most susceptible (mortality rate: 100%). The LC50 and LC95 for Bi. camerunensis eggs were 0.171 mg/L and 1.102 mg/L, respectively, and were higher than those obtained for adults (0.0357 mg/L and 0.9634 mg/L); (4) Conclusion: These findings will guide the design of vector control strategies targeting these snail species in Cameroon.

[Activity of aromatic pyrrole-based compounds against of Schistosoma japonicum cercariae and acute toxicity to fish].

OBJECTIVE: To test the activity of aromatic pyrrole-based compounds against cercariae of Schistosoma japonicum and test their acute toxicity to fish. METHODS: A series of aromatic pyrrole-based compounds were synthesized using 4-benzyl-5-(trifluoromethyl)-1H-pyrrole-3-nitrile as the lead compound. The synthesized compounds were prepared into solutions at concentrations of 10.00, 1.00, 0.10, 0.01 mg/L, and the activity of these solutions against S. japonicum cercariae was tested in 30 min, while 0.10 mg/L and 0.01 mg/L niclosamide solutions served as a positive control and dechlorinated water with 1% dimethyl sulfoxide (DMSO) was used as a negative control, with 10 to 30 cercariae of S. japonicum in each group. In addition, the compounds were prepared into solutions at concentrations of 0.50, 0.25, 0.12, 0.06, 0.03 mg/L, and their toxicity to zebrafish was tested in 72 h, while 0.15 mg/L and 0.30 mg/L niclosamide solutions served as a positive control and dechlorinated water with 1% DMSO was used as a negative control, with 10 zebrafishes in each group. RESULTS: A total of 7 aromatic pyrrole-based compounds were successfully synthesized. Treatment with compounds 102, 104 and 106 at a concentration of 0.01 mg/L for 30 min killed all S. japonicum cercariae, and compounds 105 and 107 showed no activity against cercariae. No death of cercariae was found in the blank control group, while treatment with 0.10 mg/L niclosamide for 10 min caused a 100% mortality rate of S. japonicum cercariae and 0.01 mg/L niclosamide failed to kill S. japonicum cercariae. No zebrafish death was found 72 h post-treatment with compounds 101, 104 and 105 at a concentration of 0.03 mg/L, and exposure to compounds 102, 103 and 106 at a concentration of 0.03 mg/L for 12 h resulted in a 100% mortality rate of zebrafish. No zebrafish death occurred 72 h post-treatment with 0.50 mg/L Compound 104, and no zebrafish death was found in the blank control group, while treatment with 0.30 mg/L niclosamide for 24 h resulted in a 100% mortality rate of zebrafish. CONCLUSIONS: Compound 104 achieves a 100% mortality rate against S. japonicum cercariae at a concentration of 0.01 mg/L for 30 min, and causes no death of zebrafish at a concentration of 0.50 mg/L for 72 h, which may serve as a cercaricide candidate.

Reversible disruptions to energy supply and acid-base balance in larval sea lamprey exposed to the pesticide: Niclosamide (2',5-dichloro-4'-nitrosalicylanilide).

Since the 1960s, chemical control of larval sea lamprey has been achieved using the pesticides 3-trifluoromethyl-4-nitrophenol (TFM) and niclosamide (Bayluscide®). Much more potent, niclosamide is often used as an adjuvant for TFM, and on its own to treat lentic habitats, rivers with high discharge and currents, and for population surveys. Yet, little is known about its mode of action or physiological effects on sea lamprey. Like TFM, niclosamide is thought to impair mitochondrial ATP production by uncoupling oxidative phosphorylation. We therefore tested the hypothesis that niclosamide would result in metabolic perturbations and disturbances to acid-base balance in larval lamprey due to their need to balance ATP supply with ATP demands. When larval sea lamprey were exposed to the nominal 9-h niclosamide LC50 (0.11 mg L-1) over 9 h, it resulted in significant decreases in brain, phosphocreatine (35 %) and glycogen (50 %), accompanied by a 5-fold increase in lactate. In carcass, there were 25-30 % decreases in glycogen, corresponding increases in pyruvate and lactate, and a pronounced 0.5 unit decrease in intracellular pH. Calculation of the NAD+/NADH ratio in the carcass indicated that neither oxygen delivery nor the flux of reducing equivalents through the mitochondrial electron transport chain were impaired by niclosamide, supporting the hypothesis that niclosamide interferes with mitochondrial ATP production by uncoupling oxidative phosphorylation. Thus, greater reliance on glycogen, characterized by higher rates of glycolysis, temporarily mitigates the corresponding shortfall in ATP supply caused by niclosamide. Notably, all lamprey that survived niclosamide exposure readily restored ATP, phosphocreatine, glycogen and acid-base balance after recovery in niclosamide-free water. This resilience suggests that sea lamprey that survive or escape niclosamide treatment could compromise sea lamprey control efforts by subsequently completing their larval stage and developing into parasitic juvenile sea lamprey that could ultimately threaten Great Lake's fisheries populations.

The potential endocrine disruption mechanism of anthelmintic drug niclosamide by activating estrogen receptors and estrogen-related receptors.

Niclosamide (NIC), a helminthic drug used widely for controlling schistosomiasis, can reportedly disrupt the endocrine system. However, its underlying mechanisms are still unclear. In this study, we revealed the potential endocrine disruption mechanism of NIC by activating estrogen receptors (ERs) and estrogen-related receptors (ERRs). The binding potency of NIC with ERα, ERß and ERRγ were determined by fluorescence competitive binding assays, which shows an IC50 (the concentration of NIC needed to displace 50 % of the probe from the receptor) of 90 ± 4.1, 10 ± 1.7 nM and 0.59 ± 0.07 nM respectively. The IC50 for ERRγ is the lowest one among the three detected receptors, which is three orders of magnitude lower than the known agonist GSK4716.The transcriptional activities of NIC on ERs and ERRs were detected by MVLN cells (stably transfected with ERs reporter gene) and HeLa cells (transiently transfected with ERRs reporter gene)-based luciferase reporter gene assay. The lowest observable effective concentration (LOEC) ranked as follows: ERRγ (0.5 nM) < ERRα (10 nM) < ERs (100 nM). The maximum observed induction rate for ERRγ (294 %) was higher than that for ERRα (191 %). The maximum observed induction rate of NIC for ERs was 30 % relative to 17ß-estradiol. In addition, we simulated the interactions of NIC with ERs and ERRs by molecular docking. NIC could dock into the ligand binding pockets of ERs and ERRs and form hydrogen bonds with different amino acids. The binding energy ranked as follows: ERRγ (-8.90 kcal/mol) < ERß (-7.57 kcal/mol) < ERRα (-7.15 kcal/mol) < ERα (-6.53 kcal/mol), which implied that NIC bound to ERRγ with higher binding affinity than the other receptors. Overall, we clarify that ERRγ might be the dominant target for NIC in cells rather than ERRα and ERs. We reveal potential novel mechanisms for the endocrine disruption effects of NIC by activating both ERRs and ERs at environmentally-related nanomolar levels.

High-throughput transcriptome sequencing reveals the developmental toxicity mechanisms of niclosamide in zebrafish embryo.

Niclosamide (NIC) is the most widely used molluscicides for preventing the occurrence of schistosomiasis disease, and its residues can be found in various environmental samples. However, the toxicity mechanism of NIC during early developmental stage remains largely unknown. In the present study, zebrafish embryos were acutely exposed to NIC at an environmentally realistic concentration (0 and 40 µg/L) until 120 h post-fertilization. Transcriptomic sequencing was performed to provide mechanistic insight into developmental impairment. Pathway enrichment analyses found that biological processes related to lipid metabolism were significantly affected in exposed zebrafish larvae. Consistently, biochemical measurements showed that NIC developmental exposure depleted lipid storage, elevated lipid utilization, but inhibited lipid synthesis. Furthermore, as characterized by pathway enrichment and hormonal levels, steroid hormone biosynthesis was also significantly disrupted by NIC exposure in zebrafish larvae, indicating the endocrine disrupting potential of NIC. Detoxifying phase I and II processes (e.g., metabolism, conjugation and elimination) were significantly activated by NIC exposure. Overall, our findings suggest that NIC developmental exposure at an environmentally realistic concentration disturbs the lipid metabolism, induces endocrine disruption and initiates detoxifying capacity in zebrafish larvae, which will provide preliminary clues for developmental toxicity mechanisms of NIC.

[Molluscicidal effect and toxicity of META-Li on Oncomelania snails in the mountain area].

The molluscicidal effect and toxicity of META-Li were evaluated and compared with niclosamide in Yushan county of Jiangxi Province in August 2008. There are four groups named as META-Li ridge group (1 g/m2), META-Li field group (1 g/m2), niclosamide group (2 g/m2) and control group. At 3 d, 7 d, 15 d and 2 months after drugs sprayed, the corrected mortalities of snails in the 3 groups were considerably higher than those of control (P<0.05). The corrected mortalities in META-Li field group were significantly lower than those of META-Li ridge group and niclosamide group (P<0.05). No adverse effect was observed to non-target organisms in META-Li groups, while fishes, snails and frogs all died at the first day after treated with niclosamide.

Toxicity of a molluscicide candidate PPU07 against Oncomelania hupensis (Gredler, 1881) and local fish in field evaluation.

Schistosomiasis japonica caused by Schistosoma japonicum infection is recognized as a considerable economic and public health concern in Asia. Oncomelania hupensis is the sole intermediate host of S. japonicum. The only molluscicide recommended by World Health Organization (WHO) since 1960s is relative toxic to other aquatic species. In this article, we evaluated the novel molluscicide PPU07 in field trials on their efficiency against O. hupensis and toxicity for local fish. 25% PPU07 sulfate WP exhibited similar molluscicidal effect at 2.0 g/m2 and 2.0 g/m3 in the spraying and immersion trials with the WHO recommended molluscicide niclosamide (1 g/m2 and 1 g/m3). The mortality rates reached 95% and 96%, respectively. Moreover, little toxicity was observed for local fish and other aquatic organisms at the effective molluscicidal concentrations. In all, 25% PPU07 sulfate WP is a promising molluscicide for snail control, particularly in semi-commercial or commercial aquaculture ponds.

Control of invasive sea lampreys using the piscicides TFM and niclosamide: Toxicology, successes & future prospects.

The invasion of the Laurentian Great Lakes of North America by sea lampreys (Petromyzon marinus) in the early 20th century contributed to the depletion of commercial, recreational and culturally important fish populations, devastating the economies of communities that relied on the fishery. Sea lamprey populations were subsequently controlled using an aggressive integrated pest-management program which employed barriers and traps to prevent sea lamprey from migrating to their spawning grounds and the use of the piscicides (lampricides) 3-trifluoromethyl-4-nitrophenol (TFM) and niclosamide to eliminate larval sea lampreys from their nursery streams. Although sea lampreys have not been eradicated from the Great Lakes, populations have been suppressed to less than 10% of their peak numbers in the mid-1900s. The ongoing use of lampricides provides the foundation for sea lamprey control in the Great Lakes, one of the most successful invasive species control programs in the world. Yet, significant gaps remain in our understanding of how lampricides are taken-up and handled by sea lampreys, how lampricides exert their toxic effects, and how they adversely affect non-target invertebrate and vertebrates species. In this review we examine what has been learned about the uptake, handling and elimination, and the mode of TFM and niclosamide toxicity in lampreys and in non-target animals, particularly in the last 10 years. It is now clear that the mode of TFM toxicity is the same in non-target fishes and lampreys, in which TFM interferes with oxidative phosphorylation by the mitochondria leading to decreased ATP production. Vulnerability to TFM is related to abiotic factors such as water pH and alkalinity, which we propose changes the relative amounts of the bioavailable un-ionized form of TFM in the gill microenvironment. Niclosamide, which is also a molluscicide used to control snails in areas prone to schistosomiasis infections of humans, also likely works by uncoupling oxidative phosphorylation, but less is known about other aspects of its toxicology. The effects of TFM include reductions in energy stores, particularly glycogen and high energy phosphagens. However, non-target fishes readily recover from sub-lethal TFM exposure as demonstrated by the rapid restoration of energy stores and clearance of TFM. Although both TFM and niclosamide are non-persistent in the environment and critical for sea lamprey control, increasing public and institutional concerns about pesticides in the environment makes it imperative to explore other means of sea lamprey control. Accordingly, we also address possible "next-generation" strategies of sea lamprey control including genetic tools such as RNA interference and CRISPR-Cas9 to impair critical physiological processes (e.g. reproduction, digestion, metamorphosis) in lamprey, and the use of green chemistry to develop more environmentally benign chemical methods of sea lamprey control.

[Effect of niclosamide on thyroid endocrine system of larval zebrafish].

OBJECTIVE: To examine the effect of niclosamide on thyroid endocrine disruption in larval zebrafish. METHODS: Zebrafish embryos (2 hours post-fertilization) were exposed to niclosamide at concentrations of 0, 5, 10, 20, 40 µg/L and 80 µg/L until 120 hours post-fertilization, and the body weight, hatching rate, malformation rate and survival rate of zebrafish embryos/larvae were measured. In addition, the triiodothyronine (T3) and thyroxin (T4) activities were determined in zebrafish, and the expression of tshß and ttr genes that were associated with the regulation of thyroid hormones was quantified using a quantitative real-time PCR (qPCR) assay. RESULTS: Following exposure to niclosamide, there was no concentration-dependent hatching rate (F = 0.947, P = 0.924) or body weight of larval zebrafish (F = 1.042, P = 0.409); however, there were concentration-dependent survival rate (F = 9.309, P = 0.005) and malformation rate (F = 14.900, P = 0.001). As compared to controls, exposure to niclosamide at concentrations of 40 µg/L and 80 µg/L resulted in a significant reduction in the survival rate (both P values < 0.05), and a marked rise in the malformation rate of larval zebrafish (both P values < 0.05). In addition, the T4 activity increased (R2 = 0.927, F = 6.858, P = 0.003) and T3 activity decreased (R2 = 0.925, F = 8.212, P = 0.001) in larval zebrafish with the concentration of niclosamide. qPCR assay determined up-regulation of tshß gene expression (R2 = 0.840, F = 9.032, P = 0.002) and down-regulation of ttr gene expression (R2 = 0.952, F = 9.130, P = 0.002). CONCLUSIONS: Niclosamide exposure at environmental related concentrations may cause thyroid endocrine disruption of larval zebrafish.

Potential of some monoterpenoids and their new N-methyl carbamate derivatives against Schistosomiasis snail vector, Biomphalaria alexandrina.

Some monoterpenoids and their corresponding new N-methyl carbamate derivatives were used to study their molluscicidal effect on Biomphalaria alexandrina, the snail-vector of Schistosoma mansoni in Egypt. Improving the efficacy of the tested monoterpenoids and/or their corresponding carbamates by either piperonyl butoxide (PBO) or triton X-100 (TX) was also performed. Thymol, ß-citronellol, carvacrol, and geraniol exhibited high molluscicidal activity against the snails. Geraniol, ß-citronellol, and carvacrol were strongly synergized by PBO but, an opposite trend was found with TX. The molluscicidal activity of geraniol or ß-citronellol when mixed with PBO was as potent as copper sulfate. Another attempt to improve the bioactivity of monoterpenoids was through their structure modifications. Thus, conversion of the tested monoterpenoids into their corresponding carbamates led to enhancement in the activity of aliphatic monoterpenoids and reduction in the aromatics.

Niclosamide Induces Epiboly Delay During Early Zebrafish Embryogenesis.

Niclosamide is an antihelminthic drug used worldwide for the treatment of tapeworm infections. Recent drug repurposing screens have revealed that niclosamide exhibits diverse mechanisms of action and, as a result, demonstrates promise for a number of applications, including the treatment of cancer, bacterial infections, and Zika virus. As new applications of niclosamide will require non-oral delivery routes that may lead to exposure in utero, the objective of this study was to investigate the mechanism of niclosamide toxicity during early stages of embryonic development. Using zebrafish as a model, we found that niclosamide induced a concentration-dependent delay in epiboly progression during late-blastula and early-gastrula, an effect that was dependent on exposure during the maternal-to-zygotic transition-a period characterized by degradation of maternally derived transcripts, zygotic genome activation, and initiation of cell motility. Moreover, we found that niclosamide did not affect embryonic oxygen consumption, suggesting that oxidative phosphorylation-a well-established target for niclosamide within intestinal parasites-may not play a role in niclosamide-induced epiboly delay. However, mRNA-sequencing revealed that niclosamide exposure during blastula and early-gastrula significantly impacted the timing of zygotic genome activation as well as the abundance of cytoskeleton- and cell cycle regulation-specific transcripts. In addition, we found that niclosamide inhibited tubulin polymerization in vitro, suggesting that niclosamide-induced delays in epiboly progression may, in part, be driven by disruption of microtubule formation and cell motility within the developing embryo.

Isolation, characterization using LC-ESI-QTOF, NMR and in vitro cytotoxicity assay of niclosamide forced degradation products.

The present study describes the isolation, characterization and in vitro cytotoxic effect of all forced degradation products of niclosamide (NCM) an anthelmintic class of drug used specifically to treat tapeworms. NCM was subjected to forced degradation involving hydrolysis (acidic, alkaline and neutral), oxidative, photolysis and thermal stress, as per ICH (Q1A (R2)) suggested conditions. The drug under hydrolytic (acidic and basic) conditions showed extensive degradation, while it was stable under neutral hydrolytic, oxidative, photolytic and thermal stress conditions. A total of four degradation products (DPs) were observed and chromatographic separation of drug and its degradation products were achieved on a reverse phase Fortis diphenyl column (150×4.6mm, 5µm) using 0.1% formic acid and acetonitrile as mobile phase in gradient mode. All the four degradation products were isolated by semi preparative LC and its structures were characterized and confirmed by high resolution MS and 1H NMR spectroscopic techniques. In view of safety aspects, cytotoxicity assay were carried out for NCM and its four degradation products on human mononuclear cells and cell lines depicting the major organelle: neuronal (Neuro 2a), hepatic (HepG 2) and alveolar (A549). NCM was found to be non toxic on human mononuclear cells and cell lines at tested concentrations. However DP-1, DP-2, DP-3 and DP-4 showed significant increase in LDH release as compared to control at a concentration of 100µM. DP-1 and DP-3 exhibited toxicity on A549 cells with an IC50 of 92.18±4.93µM and 65.42±6.29µM respectively. DP-2, DP-3 and DP-4 were cytotoxic to Neuro 2a cells with an IC50 of 63.62±3.85µM, 86.09±6.19µM and 42.81±8.10µM respectively. The degradation products were found to be nontoxic on HepG 2 cells.