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.

• Klíčová slova
• Apoptosis, Autophagy, Flubendazole, Melanoma, Mitotic catastrophe, Necrosis,
• 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
• protinádorové látky farmakologie   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
• Názvy látek
• BCL2 protein, human MeSH Prohlížeč
• cytochromy c MeSH
• flubendazole MeSH Prohlížeč
• JNK mitogenem aktivované proteinkinasy MeSH
• MCL1 protein, human MeSH Prohlížeč
• mebendazol MeSH
• nádorový supresorový protein p53 MeSH
• protein MCL-1 MeSH
• protinádorové látky MeSH
• protoonkogenní proteiny c-bcl-2 MeSH

The specific effects of sodium selenite (selenite) on a chemoresistant human bladder cancer cell line RT-112/D21 were investigated during 72 h. Selenite at low concentration of 2.5 μmol (otherwise tolerated in normal urothelial cells UROtsa) suppressed growth and proliferation of the tested cancer cells via induced oxidative stress. Selenite further altered mitochondrial functions (i.e. decreased mitochondrial membrane potential, increased production of superoxide and reduced ATP synthesis), disrupted lysosomal membranes and activated autophagy. These changes in selenite-exposed cells ultimately resulted in their demise via necrosis and other cell death modality displaying heterotypic apoptotic and autophagic features.

• Klíčová slova
• Autophagic cell death, Bladder cancer, Lysosomes, Mitochondria, Necrosis, Selenite,
• MeSH
• apoptóza účinky léků   MeSH
• autofagie účinky léků   MeSH
• buněčná smrt účinky léků   MeSH
• chemorezistence účinky léků   MeSH
• lidé MeSH
• lyzozomy účinky léků   metabolismus   MeSH
• membránový potenciál mitochondrií účinky léků   MeSH
• mitochondrie účinky léků   metabolismus   MeSH
• nádorové buněčné linie MeSH
• nádory močového měchýře farmakoterapie   metabolismus   patologie   MeSH
• proliferace buněk účinky léků   MeSH
• seleničitan sodný farmakologie   MeSH
• vztah mezi dávkou a účinkem léčiva MeSH
• vztahy mezi strukturou a aktivitou MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• seleničitan sodný 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.

• Klíčová slova
• Flubendazole, Melanoma, Microtubules, Mitotic catastrophe, apoptosis,
• MeSH
• antinematodní látky toxicita   MeSH
• apoptóza účinky léků   MeSH
• lidé MeSH
• mebendazol analogy a deriváty   toxicita   MeSH
• melanom * MeSH
• mitóza účinky léků   MeSH
• nádorové buněčné linie MeSH
• proliferace buněk účinky léků   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• antinematodní látky MeSH
• flubendazole MeSH Prohlížeč
• mebendazol MeSH

We have investigated the cytotoxicity and specific effects of selenite in human bladder cancer cell line RT-112 and its clonogenic variant RT-112 HB. Selenite inhibited cell growth and proliferation in both cell lines. Treated cells developed extensive vacuolization which was dose independent but occurring in differing time frames. Ultrastructure analysis revealed that the observed vacuoles are damaged mitochondria and potentially other subcellular compartments. Selenite-specific effects on mitochondria were further confirmed by mitochondrial membrane potential analysis, changes in ATP production and generation of superoxide. Simultaneously, selenite induced DNA damage in treated cells with activation of p53, PARP-1 and JNK and suppressed autophagy. Cells ultimately died via a combination of apoptosis, necrosis and a distinct type of cell death featuring "vacuolar shrinkage", loss of adherence and absence of secondary necrosis as well as other classical markers of either apoptosis or autophagy. The significant presence of so called necroptosis was also not confirmed as the specific inhibitor necrostatin-1 could not prevent cell death. These results thus confirm the toxicity of selenite in bladder cancer cells while pointing at potentially new mechanism of action of this compound in this model.

• Klíčová slova
• Bladder cancer, DNA damage, Mitochondria, Necroptosis, Necrosis, Selenite,
• MeSH
• apoptóza účinky léků   MeSH
• kyselina seleničitá toxicita   MeSH
• lidé MeSH
• membránový potenciál mitochondrií účinky léků   MeSH
• mitochondrie účinky léků   fyziologie   MeSH
• nádorové buněčné linie MeSH
• nádorový supresorový protein p53 metabolismus   MeSH
• nádory močového měchýře metabolismus   patologie   MeSH
• nekróza chemicky indukované   MeSH
• poškození DNA * MeSH
• superoxidy metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• kyselina seleničitá MeSH
• nádorový supresorový protein p53 MeSH
• superoxidy MeSH

In malignant melanoma complex reprogramming of cell death and survival pathways leads to increased chemoresistance and poor longer-term survival. Sulforaphane (SF) is a promising isothiocyanate compound occurring in cruciferous plants with reported antiproliferative and proapoptotic activity in several tumor cell lines including melanoma. In this work we investigated the effects of SF in several melanoma cell lines and fresh melanoma cultivates. We found that SF is cytotoxic and induces mitochondrial, caspase-dependent apoptosis in our study model, however with lower efficiency in fresh melanoma cultivates. Moreover, our results indicate that in melanoma cell lines and fresh melanoma cultivates SF induces multiple signaling including oxidative stress-mediated activation of DNA-damage response pathway, changes in p38 kinase activity and enhanced expression of Bax and Puma proapoptotic proteins. In addition, in SF-exposed p53-mutant melanoma cells Puma expression seem to be under p38 control and acts as a compensatory proapoptotic mechanism. Conversely, decreased apoptosis in SF-exposed melanoma cultivates might be attributed to Akt-mediated suppression of p38 as well as p53 activity. Together, our results suggest that SF inhibits growth and proliferation and induces mitochondrial apoptosis both in melanoma cell lines as well as in fresh melanoma cultivates. This proapoptotic effect might be enhanced in combination with Akt inhibitors, in particular in melanoma samples. SF is thus commendable for further preclinical testing, both as a single agent as well as in combination regimens.

• MeSH
• antikarcinogenní látky farmakologie   MeSH
• apoptóza účinky léků   MeSH
• isothiokyanatany farmakologie   MeSH
• kaspasy metabolismus   MeSH
• lidé MeSH
• melanom patologie   MeSH
• mitochondrie metabolismus   MeSH
• mitogenem aktivované proteinkinasy p38 metabolismus   MeSH
• nádorové buněčné linie účinky léků   MeSH
• nádorový supresorový protein p53 metabolismus   MeSH
• nádory kůže patologie   MeSH
• reaktivní formy kyslíku metabolismus   MeSH
• signální transdukce MeSH
• sulfoxidy MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• antikarcinogenní látky MeSH
• isothiokyanatany MeSH
• kaspasy MeSH
• mitogenem aktivované proteinkinasy p38 MeSH
• nádorový supresorový protein p53 MeSH
• reaktivní formy kyslíku MeSH
• sulforaphane MeSH Prohlížeč
• sulfoxidy MeSH

The role of p38 in irinotecan (CPT-11)-induced damage and cell death in colon cancer cell line SW620 was investigated. We demonstrate that CPT-11 treatment activates p38 in exposed cells, however with concentration dependent dynamics and differing consequences. Higher CPT-11 concentrations induce a massive early but relatively short-lasting p38 activity leading to apoptosis mediated by mitochondria and caspases. Pharmacological or siRNA inhibition of p38 then significantly prevents CPT-11-dependent cell death. Conversely, lower CPT-11 concentrations activate p38 in a delayed, however sustained manner, with apoptosis occurring only in a fraction of cells and in the absence of significant autophagy. Blocking p38 in thus treated cells increases their sensitivity toward CPT-11 and increases cell death. In summary, our results confirm the involvement of p38 in colon cancer cells response to CPT-11 while indicating a varying role of p38 in the final biological response.

• MeSH
• apoptóza účinky léků   MeSH
• autofagie účinky léků   MeSH
• časosběrné zobrazování MeSH
• cytochromy c metabolismus   MeSH
• fytogenní protinádorové látky farmakologie   MeSH
• irinotekan MeSH
• kamptothecin analogy a deriváty   farmakologie   MeSH
• kaspasy metabolismus   MeSH
• kvantitativní polymerázová řetězová reakce MeSH
• lidé MeSH
• membránový potenciál mitochondrií účinky léků   MeSH
• messenger RNA genetika   MeSH
• mitochondrie účinky léků   metabolismus   MeSH
• mitogenem aktivované proteinkinasy p38 genetika   metabolismus   MeSH
• nádorové buňky kultivované MeSH
• nádory tračníku farmakoterapie   genetika   patologie   MeSH
• polymerázová řetězová reakce s reverzní transkripcí MeSH
• poškození DNA účinky léků   MeSH
• western blotting MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• cytochromy c MeSH
• fytogenní protinádorové látky MeSH
• irinotekan MeSH
• kamptothecin MeSH
• kaspasy MeSH
• messenger RNA MeSH
• mitogenem aktivované proteinkinasy p38 MeSH

Various DNA-targeting agents may initiate p53-dependent as well as p53-independent response and subsequent apoptosis via alternative cellular systems which include for instance p73, caspase-2 or Bcl-2 family proteins. The scope of involvement of individual molecules in this process and the mechanisms governing their potential interplay are still not entirely understood, in particular in highly aggressive cancers such as in malignant melanoma. In this work we investigated the role and involvement of both p53-dependent and -independent mechanisms in selected melanoma cell lines with differing status of p53 using a model DNA topoisomerase I inhibitor camptothecin (CPT). Here we report that CPT induced in Bowes melanoma cells apoptosis which is essentially p53 and mitochondria-dependent but with some involvement of caspase-2 and p73. Conversely, in mutant p53 melanoma cells overall levels of CPT-induced apoptosis are significantly lower, with p73 and caspase-2 signaling playing important roles. In addition, in these cells the expression of micro RNAs family 34 (miR-34) were low compared to wild-type p53 cells. The ectopic expression of wild type p53 than restored apoptotic response of cells to CPT despite the fact that the expression of miR-34 and miR-155 were not influenced. These results suggest that CPT induces multivariate cellular stress responses including activation of DNA-damage response-p53 pathway as well as p53-independent signaling and their mutual crosstalk play the decisive role in the efficient triggering of apoptosis in melanoma cells.

• MeSH
• apoptóza účinky léků   MeSH
• cysteinové endopeptidasy genetika   metabolismus   MeSH
• DNA vazebné proteiny genetika   metabolismus   MeSH
• fytogenní protinádorové látky farmakologie   MeSH
• inhibitory topoisomerasy I farmakologie   MeSH
• jaderné proteiny genetika   metabolismus   MeSH
• kamptothecin farmakologie   MeSH
• kaspasa 2 genetika   metabolismus   MeSH
• kvantitativní polymerázová řetězová reakce MeSH
• lidé MeSH
• melanom genetika   metabolismus   patologie   MeSH
• mikro RNA genetika   metabolismus   MeSH
• mitochondrie účinky léků   genetika   metabolismus   MeSH
• nádorové buněčné linie MeSH
• nádorové supresorové proteiny genetika   metabolismus   MeSH
• nádorový supresorový protein p53 genetika   metabolismus   MeSH
• poškození DNA účinky léků   MeSH
• protein p73 MeSH
• protoonkogenní proteiny c-bcl-2 genetika   metabolismus   MeSH
• signální transdukce * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• CASP2 protein, human MeSH Prohlížeč
• cysteinové endopeptidasy MeSH
• DNA vazebné proteiny MeSH
• fytogenní protinádorové látky MeSH
• inhibitory topoisomerasy I MeSH
• jaderné proteiny MeSH
• kamptothecin MeSH
• kaspasa 2 MeSH
• mikro RNA MeSH
• nádorové supresorové proteiny MeSH
• nádorový supresorový protein p53 MeSH
• protein p73 MeSH
• protoonkogenní proteiny c-bcl-2 MeSH
• TP73 protein, human MeSH Prohlížeč

The cytotoxicity of topoisomerase I inhibiting camptothecin, topoisomerase II inhibiting etoposide and their combination were investigated in wild type p53 Bowes and mutant p53 SK-MEL-28 melanoma cell lines during 24h. A combination of camptothecin and etoposide (1 microg/ml + 10 microg/ml) proved to be efficient in both types of cell lines, although mutant p53 cells exhibited a higher resistance. Further studies proved that in Bowes cells, a combination of drugs induced p53-dependent mitochondrial apoptosis characterized by activation of caspases-8 and -2, -9 and -3 with some concurrent involvement of oxidative stress. In SK-MEL-28 cells, apoptosis was found to be mediated via increased oxidative stress, activation of stress kinases such as p38 and SAPK/JNK and mitochondrial dysfunction without significant involvement of p53 and its transactivated target genes. These results demonstrate efficiency of dual inhibition of topoisomerases in melanoma cells with functional as well as mutant p53 and point out at possible further investigation of this modality in preclinical and clinical oncology.

• MeSH
• apoptóza * MeSH
• DNA-topoisomerasy I metabolismus   MeSH
• DNA-topoisomerasy typu II metabolismus   MeSH
• etoposid aplikace a dávkování   MeSH
• imunoblotting MeSH
• inhibitory topoisomerasy I * MeSH
• inhibitory topoisomerasy II * MeSH
• JNK mitogenem aktivované proteinkinasy metabolismus   MeSH
• kamptothecin aplikace a dávkování   MeSH
• kaspasy metabolismus   MeSH
• lidé MeSH
• MAP kinasový signální systém účinky léků   MeSH
• melanom enzymologie   patologie   MeSH
• membránový potenciál mitochondrií účinky léků   MeSH
• mutace genetika   MeSH
• nádorové buňky kultivované MeSH
• nádorový supresorový protein p53 genetika   metabolismus   MeSH
• oxidační stres účinky léků   MeSH
• proliferace buněk účinky léků   MeSH
• protokoly protinádorové kombinované chemoterapie farmakologie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• DNA-topoisomerasy I MeSH
• DNA-topoisomerasy typu II MeSH
• etoposid MeSH
• inhibitory topoisomerasy I * MeSH
• inhibitory topoisomerasy II * MeSH
• JNK mitogenem aktivované proteinkinasy MeSH
• kamptothecin MeSH
• kaspasy MeSH
• nádorový supresorový protein p53 MeSH
• TP53 protein, human MeSH Prohlížeč

We characterized the effect of phytic acid (inositol hexaphosphate, IP6) as a potential adjuvant in treatment of colorectal carcinoma and evaluated the optimal concentration and treatment time to produe the maximal therapeutic effect. There is some evidence that myoinositol (Ins) can potentiate anti-cancer effects of IP6. Therefore, we tested both IP6 and Ins individually and in combination on human cell lines HT-29, SW-480 and SW-620 derived from colorectal carcinoma in different stages of malignancy. The effect of tested chemicals on the cells was measured using metabolic activity assay (WST-1), DNA synthesis assay (BrdU), protein synthesis assay (Brilliant Blue) and apoptosis (caspase-3 activity). We tested IP6 and Ins at three concentrations: 0.2, 1 and 5 mM for 24, 48 and 72 h. The concentrations and incubation periods were chosen according to low toxicity of the tested substance that was observed in a long-term clinical study. We found that all employed concentrations of IP6 or IP6/Ins decreased proliferation of the cell lines, with the maximum decrease being observed in HT-29 cells. Metabolic activity of treated cells differed in response to IP6 and IP6/Ins treatment; in HT-29 and SW-620 significant decrease was observed only at the highest concentration, whereas in SW-480 cells metabolic activity was lower at each concentration except 0.2 and 1 mM IP6 or IP6/Ins in 24-h incubation. The results from protein content assay corresponded to the results obtained from WST assay. In addition, we found maximum increase in caspase-3 activity at concentration 5 mM IP6 or IP6/Ins in HT-29 cells and with IP6 at concentration of 0.2 mM or IP6/Ins in SW-480 cells with clear indication of Ins enhancing the proapoptotic effect of IP6 in all the cell lines studied.

• MeSH
• apoptóza účinky léků   MeSH
• inositol farmakologie   MeSH
• kaspasa 3 metabolismus   MeSH
• kolorektální nádory farmakoterapie   patologie   MeSH
• kyselina fytová farmakologie   MeSH
• lidé MeSH
• nádorové buněčné linie MeSH
• nádorové proteiny analýza   MeSH
• proliferace buněk účinky léků   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• inositol MeSH
• kaspasa 3 MeSH
• kyselina fytová MeSH
• nádorové proteiny MeSH

Despite the reported cytotoxicity and apoptosis-inducing properties of sulforaphane (SF) in colon cancer cells, the details concerning individual mechanisms and signaling cascades underlying SF-mediated apoptosis remain unclear. To understand different aspects of SF-induced proapoptic signaling in advanced colon carcinoma, we investigated its mechanisms in metastatic SW620 cell line. Our results indicate that in SW620 cells SF acts to induce multivariate cascades including DNA-damage response pathway whose proapoptotic signaling is nevertheless reduced owing to the mutant status of p53 and caspase-2-JNK pathway which seems to complement and enhance p53-dependent signaling, however only in wild-type p53. Furthermore, both pathways require the active role of mitochondria and do not depend on generation of ROS, making SF an attractive chemopreventive agent whose antitumor properties should be further investigated in colon cancer.

• MeSH
• antikarcinogenní látky farmakologie   MeSH
• apoptóza účinky léků   MeSH
• cytochromy c metabolismus   MeSH
• isothiokyanatany MeSH
• JNK mitogenem aktivované proteinkinasy metabolismus   MeSH
• kaspasa 2 metabolismus   MeSH
• lidé MeSH
• membránový potenciál mitochondrií účinky léků   MeSH
• nádorové buněčné linie MeSH
• nádorový supresorový protein p53 metabolismus   MeSH
• nádory tračníku farmakoterapie   metabolismus   patologie   MeSH
• oxidační stres účinky léků   MeSH
• poškození DNA MeSH
• screeningové testy protinádorových léčiv MeSH
• sulfoxidy MeSH
• thiokyanatany farmakologie   MeSH
• viabilita buněk účinky léků   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• antikarcinogenní látky MeSH
• cytochromy c MeSH
• isothiokyanatany MeSH
• JNK mitogenem aktivované proteinkinasy MeSH
• kaspasa 2 MeSH
• nádorový supresorový protein p53 MeSH
• sulforaphane MeSH Prohlížeč
• sulfoxidy MeSH
• thiokyanatany MeSH