Mitotic catastrophe induced by mictotubule-targeting drugs such as benzimidazole carbamates has been demonstrated to be an efficient mechanism for suppression of tumor cells growth and proliferation, with variable resulting endpoints. The present study was designed to explore some of these endpoints; i.e. the apoptosis as well as autophagy and their related signaling in several stabilized cell lines as well as human explant melanoma cells treated with flubendazole (FLU). FLU-induced mitotic catastrophe resulted in mitochondrial and caspase-dependent apoptosis, which occurred at various rates in all treated cells during 96 h of treatment. The process was characterized by enhanced transcriptional activity of TP53 and NF-κB as well as upregulated Noxa expression. Also, inactivation of Bcl-2, BclXL and Mcl-1 proteins by JNK mediated phosphorylation was observed. Although increased autophagic activity took place in treated cells too, no discernible functional linkage with ongoing cell death process was evidenced. Together these results advance our evidence over the effectiveness of FLU cytotoxicity-related killing of melanoma cells while calling for more extensive testing of melanoma samples as a prerequisite of further preclinical evaluation of FLU antineoplastic potential.
- MeSH
- antitumorózní látky farmakologie MeSH
- apoptóza účinky léků MeSH
- autofagie účinky léků MeSH
- buněčný cyklus účinky léků MeSH
- cytochromy c metabolismus MeSH
- JNK mitogenem aktivované proteinkinasy metabolismus MeSH
- lidé středního věku MeSH
- lidé MeSH
- mebendazol analogy a deriváty farmakologie MeSH
- melanom farmakoterapie metabolismus MeSH
- membránový potenciál mitochondrií účinky léků MeSH
- mitóza MeSH
- nádorové buněčné linie MeSH
- nádorový supresorový protein p53 metabolismus MeSH
- protein MCL-1 metabolismus MeSH
- protoonkogenní proteiny c-bcl-2 metabolismus MeSH
- senioři MeSH
- Check Tag
- lidé středního věku MeSH
- lidé MeSH
- mužské pohlaví MeSH
- senioři MeSH
- Publikační typ
- časopisecké články MeSH
Flubendazole (FLU) is a widely used anthelmintic drug belonging to benzimidazole group. Recently, several studies have been published demonstrating its potential to inhibit growth of various tumor cells including those derived from colorectal cancer, breast cancer or leukemia via several mechanisms. In the present study we have investigated cytotoxic effects of FLU on malignant melanoma using A-375, BOWES and RPMI-7951 cell lines representing diverse melanoma molecular types. In all three cell lines, FLU inhibited cell growth and proliferation and disrupted microtubule structure and function which was accompanied by dramatic changes in cellular morphology. In addition, FLU-treated cells accumulated at the G2/M phase of cell cycle and displayed the features of mitotic catastrophe characterized by formation of giant cells with multiple nuclei, abnormal spindles and subsequent apoptotic demise. Although this endpoint was observed in all treated melanoma lines, our analyses showed different activated biochemical signaling in particular cells, thus suggesting a promising treatment potential of FLU in malignant melanoma warranting its further testing.
BACKGROUND: Colon cancer is the most common type of gastrointestinal cancer. Despite advances during the last two decades, the efficacy of colorectal cancer treatment is still insufficient and new anticancer agents are necessary. METHODS: In our study, colon cancer cells derived from a primary tumor (SW480) and lymph node metastasis (SW620) from the same patient were used and compared. The effect of flubendazole (FLU) on cell adhesion and migration was monitored using the x-CELLigence Real-Time Cell Analysis system. Expressions of molecules involved in adhesion and migration were analyzed using RT-PCR and western blot. Furthermore, RNA silencing of nuclear factor-κB in SW620 cells was used to determine the involvement of the NF-κB p65 regulation pathway in FLU action. RESULTS: FLU significantly suppressed the adhesion of SW480 cells and reduced the expression of adhesion markers (ICAM-1, αE-catenin; β-catenin; integrin α5 and β1). Moreover, a significant anti-migratory potential of FLU was manifested in the SW620 cells. In addition, FLU suppressed the phosphorylation of NF-κB p65 and potentiated the suppression of several metastatic markers (ICAM-1, EpCAM, integrin α5, β1, α-tubulin) caused by NF-κB p65 silencing. CONCLUSION: FLU has a significant anti-migratory effect in intestinal cancer cell SW480 and its lymph node metastatic cells SW620. FLU decreases the expression of some proteins involved in metastatic processes and inhibits activation of NF-κB p65.
- MeSH
- antitumorózní látky chemie farmakologie MeSH
- buněčná adheze účinky léků MeSH
- léky antitumorózní - screeningové testy MeSH
- lidé MeSH
- mebendazol analogy a deriváty chemie farmakologie MeSH
- molekulární struktura MeSH
- nádorové buňky kultivované MeSH
- nádory tračníku farmakoterapie patologie MeSH
- pohyb buněk účinky léků MeSH
- proliferace buněk účinky léků MeSH
- vztah mezi dávkou a účinkem léčiva MeSH
- vztahy mezi strukturou a aktivitou MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Although veterinary anthelmintics represent an important source of environmental pollution, the fate of anthelmintics and their effects in plants has not yet been studied sufficiently. The aim of our work was to identify metabolic pathways of the two benzimidazole anthelmintics fenbendazole (FBZ) and flubendazole (FLU) in the ribwort plantain (Plantago lanceolata L.). Plants cultivated as in vitro regenerants were used for this purpose. The effects of anthelmintics and their biotransformation products on plant oxidative stress parameters were also studied. The obtained results showed that the enzymatic system of the ribwort plantain was able to uptake FLU and FBZ, translocate them in leaves and transform them into several metabolites, particularly glycosides. Overall, 12 FLU and 22 FBZ metabolites were identified in the root, leaf base and leaf top of the plant. Concerning the effects of FLU and FBZ, both anthelmintics in the ribwort plantain cells caused significant increase of proline concentration (up to twice), a well-known stress marker, and significant decrease of superoxide dismutase activity (by 50%). In addition, the activities of four other antioxidant enzymes were significantly changed after either FLU or FBZ exposition. This could indicate a certain risk of oxidative damage in plants influenced by anthelmintics, particularly when they are under other stress conditions.
- MeSH
- anthelmintika metabolismus toxicita MeSH
- biotransformace MeSH
- fenbendazol metabolismus toxicita MeSH
- mebendazol analogy a deriváty metabolismus toxicita MeSH
- metabolické sítě a dráhy účinky léků MeSH
- oxidační stres účinky léků MeSH
- Plantago účinky léků enzymologie růst a vývoj MeSH
- veterinární léky metabolismus toxicita MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
Benzimidazole anthelmintics flubendazole and mebendazole are microtubule-targeting drugs that showed considerable anti-cancer activity in different preclinical models. In this study, the effects of flubendazole and mebendazole on proliferation, migration and cadherin switching were studied in a panel of oral cell lines in vitro. Both compounds reduced the viability of the PE/CA-PJ15 and H376 oral squamous carcinoma cells and of the premalignant oral keratinocytes DOK with the IC50 values in the range of 0.19-0.26 μM. Normal oral keratinocytes and normal gingival fibroblasts were less sensitive to the treatment. Flubendazole and mebendazole also reduced the migration of the PE/CA-PJ15 cell in concentrations that had no anti-migratory effects on the normal gingival fibroblasts. Levels of the focal adhesion kinase FAK, Rho-A and Rac1 GTPases and the Rho guanine nucleotide exchange factor GEF-H1 were decreased in both PE/CA-PJ15 cells and gingival fibroblasts following treatment. Both drugs also interfered with cadherin switching in the model of TGF-β-induced epithelial to mesenchymal transition (EMT) in the DOK cell line. Levels of N-cadherin were reduced in the TGF-β induced cells co-treated with flubendazol and mebendazole in very low concentration (50 nM). These results suggest direct effects of both benzimidazoles on selected processes of EMT in oral cell lines such as cadherin switching as well as cellular migration.
- MeSH
- buněčné linie MeSH
- cdc42 protein vázající GTP metabolismus MeSH
- epitelo-mezenchymální tranzice účinky léků MeSH
- fokální adhezní kinasa 1 metabolismus MeSH
- kadheriny metabolismus MeSH
- lidé MeSH
- mebendazol analogy a deriváty farmakologie MeSH
- nádory úst metabolismus patologie MeSH
- pohyb buněk účinky léků MeSH
- proliferace buněk účinky léků MeSH
- rhoA protein vázající GTP metabolismus MeSH
- spinocelulární karcinom metabolismus patologie MeSH
- transformující růstový faktor beta farmakologie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Haemonchus contortus (family Trichostrongylidae, Nematoda), a hematophagous gastrointestinal parasite found in small ruminants, has a great ability to develop resistance to anthelmintic drugs. We studied the biotransformation of the three benzimidazole anthelmintics: albendazole (ABZ), ricobendazole (albendazole S-oxide; RCB) and flubendazole (FLU) in females and males of H. contortus in both a susceptible ISE strain and resistant IRE strain. The ex vivo cultivation of living nematodes in culture medium with or without the anthelmintics was used. Ultrasensitive UHPLC/MS/MS analysis revealed 9, 7 and 12 metabolites of ABZ, RCB and FLU, respectively, with most of these metabolites now described in the present study for the first time in H. contortus. The structure of certain metabolites shows the presence of biotransformation reactions not previously reported in nematodes. There were significant qualitative and semi-quantitative differences in the metabolites formed by male and female worms. In most cases, females metabolized drugs more extensively than males. Adults of the IRE strain were able to form many more metabolites of all the drugs than adults of the ISE strain. Some metabolites were even found only in adults of the IRE strain. These findings suggest that increased drug metabolism may play a role in resistance to benzimidazole drugs in H. contortus.
- MeSH
- albendazol analogy a deriváty metabolismus farmakologie MeSH
- anthelmintika metabolismus farmakologie MeSH
- biochemické jevy MeSH
- biotransformace MeSH
- Haemonchus metabolismus MeSH
- hemonchóza farmakoterapie parazitologie veterinární MeSH
- léková rezistence * MeSH
- mebendazol analogy a deriváty metabolismus farmakologie MeSH
- nemoci ovcí farmakoterapie parazitologie MeSH
- ovce parazitologie MeSH
- sexuální faktory MeSH
- tandemová hmotnostní spektrometrie MeSH
- zvířata MeSH
- Check Tag
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Flubendazole is a widely used anthelmintic drug belonging to benzimidazole group. The molecular mechanism of action of flubendazole is based on its specific binding to tubulin, which results in disruption of microtubule structure and function, and in the interference with the microtubule-mediated transport of secretory vesicles in absorptive tissues of helminths. The microtubule-disrupting properties of benzimidazole derivatives raised recently interest in these compounds as possible anti-cancer agents. In this minireview flubendazole effects towards selected human malignant cells including myeloma, leukemia, neuroblastoma, breast cancer, colorectal cancer and melanoma are discussed along with basic data on its pharmacokinetics, metabolism and toxicity.
- MeSH
- leukemie farmakoterapie MeSH
- lidé MeSH
- mebendazol analogy a deriváty terapeutické užití MeSH
- mikrotubuly metabolismus MeSH
- mnohočetný myelom farmakoterapie MeSH
- stárnutí buněk účinky léků MeSH
- tubulin metabolismus MeSH
- viabilita buněk MeSH
- vztah mezi dávkou a účinkem léčiva MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- přehledy MeSH
Benzimidazoles anthelmintics, which enter into environment primarily through excretion in the feces or urine of treated animals, can affect various organisms and disrupt ecosystem balance. The present study was designed to test the phytotoxicity and biotransformation of the three benzimidazole anthelmintics albendazole (ABZ), fenbendazole (FBZ) and flubendazole (FLU) in the harebell (Campanula rotundifolia). This meadow plant commonly grows in pastures and comes into contact with anthelmintics through the excrements of treated animals. Suspensions of harebell cells in culture medium were used as an in vitro model system. ABZ, FLU and FBZ were not found to be toxic for harebell cells, which were able to metabolize ABZ, FLU and FBZ via the formation of a wide scale of metabolites. Ultrahigh-performance liquid chromatography coupled with high mass accuracy tandem mass spectrometry (UHPLC-MS/MS) led to the identification of 24, 18 and 29 metabolites of ABZ, FLU and FBZ, respectively. Several novel metabolites were identified for the first time. Based on the obtained results, the schemes of the metabolic pathways of these anthelmintics were proposed. Most of these metabolites can be considered deactivation products, but a substantial portion of them may readily be decomposed to biologically active substances which could negatively affect ecosystems.
- MeSH
- albendazol metabolismus MeSH
- anthelmintika chemie metabolismus MeSH
- benzimidazoly chemie metabolismus MeSH
- biotransformace MeSH
- Campanulaceae cytologie metabolismus MeSH
- ekosystém MeSH
- feces chemie MeSH
- fenbendazol metabolismus MeSH
- kultivované buňky MeSH
- mebendazol analogy a deriváty metabolismus MeSH
- metabolické sítě a dráhy * MeSH
- tandemová hmotnostní spektrometrie metody MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
The present in vitro study was designed to test and compare anthelmintic activity, hepatotoxicity, and biotransformation of four selected aminoacetonitrile derivatives (AADs): monepantel (MOP, anthelmintic approved for the treatment), AAD-970, AAD-1154, and AAD-1336. Micro-agar larval development test, MTT test of cytotoxicity, and biotransformation study coupled with Ultra high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) technique were used for this purpose. Larvae of two Haemonchus contortus strains (drug susceptible and multi-drug resistant) and primary cultures of rat and ovine hepatocytes served as model systems. All AADs (including MOP) exhibited significant larvicidal effect in H. contortus susceptible as well as multi-resistant strains, much higher than those of reference anthelmintics thiabendazole and flubendazole. AAD-1154 provides the best results for most tested parameters among all AADs in this study. The cytotoxicity test showed that all AADs can be considered as nontoxic for hepatocytes. In the biotransformation study, Phase I and Phase II metabolites of AADs were identified and schemes of possible metabolic pathways in ovine hepatocytes were proposed. Biotransformation of MOP was much more extensive than biotransformation of other AADs. Based on obtained results, AAD-1154 and AAD-1336 can be considered as promising candidates for further in vivo testing.
- MeSH
- aminoacetonitrily analogy a deriváty analýza farmakokinetika toxicita MeSH
- anthelmintika analýza farmakokinetika toxicita MeSH
- biotransformace MeSH
- Haemonchus účinky léků MeSH
- hepatocyty metabolismus MeSH
- krysa rodu rattus MeSH
- kultivované buňky MeSH
- larva MeSH
- mebendazol analogy a deriváty analýza farmakokinetika MeSH
- ovce MeSH
- potkani Wistar MeSH
- tandemová hmotnostní spektrometrie MeSH
- thiabendazol analýza farmakokinetika MeSH
- vysokoúčinná kapalinová chromatografie MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- srovnávací studie MeSH
OBJECTIVES: Flubendazole (FLU), a member of benzimidazole family of anthelmintic drugs, is able to inhibit proliferation of various cancer cells. The aim of present study was to elucidate the mechanisms of antiproliferative effect of FLU on colorectal cancer cells in vitro. METHODS: The effect of FLU on proliferation, microtubular network, DNA content, caspase activation and senescence induction was studied in SW480 and SW620 cell lines. KEY FINDINGS: Flubendazole significantly affected cell proliferation in a pattern typical for mitotic inhibitor. This was accompanied by decrease in cyclin D1 levels, increase in cyclin B1 levels, activation of caspase 2 and caspase 3/7 and PARP cleavage. Morphological observations revealed disruption of microtubular network, irregular mitotic spindles, formation of giant multinucleated cells and increase in nuclear area and DNA content. In SW620 cell line, 37.5% giant multinucleated cells induced by FLU treatment showed positivity for SA-β-galactosidase staining. Cell lines were able to recover from the treatment and this process was faster in SW480 cells. CONCLUSION: Flubendazole in low concentration temporarily inhibits cell proliferation and induces mitotic catastrophe and premature senescence in human colon cancer cells in vitro.
- MeSH
- aparát dělícího vřeténka účinky léků ultrastruktura MeSH
- lidé MeSH
- mebendazol analogy a deriváty farmakologie MeSH
- mikrotubuly účinky léků ultrastruktura MeSH
- mitóza účinky léků MeSH
- nádorové buněčné linie MeSH
- obrovské buňky účinky léků ultrastruktura MeSH
- proliferace buněk účinky léků MeSH
- stárnutí buněk účinky léků MeSH
- tubulin metabolismus MeSH
- velikost buněčného jádra účinky léků MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
