The Bcl-2 protein family comprises both pro- and antiapoptotic members that control the permeabilization of the mitochondrial outer membrane, a crucial step in the modulation of apoptosis. Recent research has demonstrated that the carboxyl-terminal transmembrane domain (TMD) of some Bcl-2 protein family members can modulate apoptosis; however, the transmembrane interactome of the antiapoptotic protein Mcl-1 remains largely unexplored. Here, we demonstrate that the Mcl-1 TMD forms homooligomers in the mitochondrial membrane, competes with full-length Mcl-1 protein with regards to its antiapoptotic function, and induces cell death in a Bok-dependent manner. While the Bok TMD oligomers locate preferentially to the endoplasmic reticulum (ER), heterooligomerization between the TMDs of Mcl-1 and Bok predominantly takes place at the mitochondrial membrane. Strikingly, the coexpression of Mcl-1 and Bok TMDs produces an increase in ER mitochondrial-associated membranes, suggesting an active role of Mcl-1 in the induced mitochondrial targeting of Bok. Finally, the introduction of Mcl-1 TMD somatic mutations detected in cancer patients alters the TMD interaction pattern to provide the Mcl-1 protein with enhanced antiapoptotic activity, thereby highlighting the clinical relevance of Mcl-1 TMD interactions.

• MeSH
• apoptóza fyziologie   MeSH
• buněčná smrt fyziologie   MeSH
• endoplazmatické retikulum metabolismus   MeSH
• HeLa buňky MeSH
• lidé MeSH
• mitochondriální membrány metabolismus   MeSH
• mitochondrie metabolismus   MeSH
• protein MCL-1 metabolismus   MeSH
• proteinové domény MeSH
• protoonkogenní proteiny c-bcl-2 metabolismus   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Etoposide is commonly used as a monotherapy or in combination with other drugs for cancer treatments. In order to increase the drug efficacy, ceaseless search for novel combinations of drugs and supporting molecules is under way. MiRNAs are natural candidates for facilitating drug effect in various cell types. We used several systems to evaluate the effect of miR-29 family on etoposide toxicity in HeLa cells. We show that miR-29b significantly increases etoposide toxicity in HeLa cells. Because Mcl-1 protein has been recognized as a miR-29 family target, we evaluated downregulation of Mcl-1 protein splicing variant expression induced by miR-29 precursors and confirmed a key role of Mcl-1 protein in enhancing etoposide toxicity. Despite downregulation of Mcl-1 by all three miR-29 family members, only miR-29b significantly enhanced etoposide toxicity. We hypothesized that this difference may be linked to the change in Mcl-1L/Mcl-1S ratio induced by miR-29b. We hypothesized that the change could be due to miR-29b nuclear shuttling. Using specifically modified miR-29b sequences with enhanced cytosolic and nuclear localization we show that there is a difference, albeit statistically non-significant. In conclusion, we show that miR-29b has the synergistic effect with etoposide treatment in the HeLa cells and that this effect is linked to Mcl-1 protein expression and nuclear shuttling of miR-29b.

• MeSH
• buněčný cyklus účinky léků   MeSH
• down regulace MeSH
• etoposid toxicita   MeSH
• fytogenní protinádorové látky toxicita   MeSH
• HeLa buňky MeSH
• lidé MeSH
• mikro RNA metabolismus   MeSH
• protein MCL-1 genetika   metabolismus   MeSH
• viabilita buněk účinky léků   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

Myosin light chains-1 (MLC-1) have been recently associated with the markers of heart function (NYHA, LVEF, NT-proBNP). Verification of the relationship between markers of heart function (New York Heart Association classification (NYHA), left ventricle ejection fraction determination (LVEF), N terminal prohormone of natriuretic peptide B type BNP (NT-proBNP) and concentrations of myosin light chains-1 (MLC-1) was assessed. Patients examined for dyspnea without signs of acute coronary syndrome. All patients underwent echocardiography (calculation of left ventricle ejection fraction--LVEF) and in the serum of all subjects NT-proBNP (ELEIA) and MLC-1 (ELISA) were determined. In the 38 patients (21 men, 17 women), mean age of 58 years (+/-12 years as 1 SD), a significant negative correlation was found between NT-proBNP and LVEF (r = - 0.47; p = 0.02, Spearman). The median levels of NT pro-BNP were closely associated with NYHA classification (type II--584 ng/l, type III--2792 ng/l, type IV--6400 ng/l; p < 0.05). Individuals with clinical NYHA IV differed significantly in median MLC-1 concentrations from persons with clinical NYHA classification II and III (type II--5.7 ng/l, type III--8.9 ng/l, type IV--17 ng/l; p < 0.05). A significant negative correlation between MLC-1 and LVEF (-0.35; p < 0.03) and significant positive correlations between MLC-1 and NT-proBNP (0.42; p < 0.012) were found. In conclusion MLC-1 cannot be used as a diagnostic marker in differential diagnosis of dyspnea.

• MeSH
• biologické markery krev   MeSH
• diferenciální diagnóza MeSH
• dyspnoe etiologie   MeSH
• lehké řetězce myosinu krev   MeSH
• lidé středního věku MeSH
• lidé MeSH
• natriuretický peptid typu B krev   MeSH
• peptidové fragmenty krev   MeSH
• ROC křivka MeSH
• senzitivita a specificita MeSH
• srdeční selhání komplikace   diagnóza   patofyziologie   MeSH
• tepový objem MeSH
• Check Tag
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH

Inhibitors of cyclin-dependent kinases 9 have been developed as potential anticancer drugs for the treatment of multiple myeloma. We have previously prepared a library of arylazo-3,5-diaminopyrazole inhibitors of CDKs. Here, we describe a novel member, AAP1742 (CDK9 inhibition with IC(50) = 0.28 μm), that reduces the viability of multiple myeloma cell lines in low micromolar concentrations. Consistent with inhibition of CDK9, AAP1742 decreases the phosphorylation of RNA polymerase II and inhibits mRNA synthesis of anti-apoptotic proteins Mcl-1, Bcl-2, and XIAP, followed by apoptosis in the RPMI-8226 cell line in a dose- and a time-dependent manner. These results are consistent with the biochemical profile of AAP1742 and further suggest cellular inhibition of CDK9 as a possible target for anticancer drugs.

• MeSH
• aktivace enzymů účinky léků   MeSH
• apoptóza účinky léků   MeSH
• azosloučeniny chemie   farmakologie   MeSH
• cyklin-dependentní kinasa 9 antagonisté a inhibitory   metabolismus   MeSH
• down regulace účinky léků   MeSH
• fosforylace účinky léků   MeSH
• lidé MeSH
• messenger RNA metabolismus   MeSH
• mitochondrie metabolismus   MeSH
• mnohočetný myelom metabolismus   patologie   MeSH
• nádorové buněčné linie MeSH
• protein MCL-1 genetika   metabolismus   MeSH
• protinádorové látky chemie   farmakologie   MeSH
• protoonkogenní proteiny c-bcl-2 genetika   metabolismus   MeSH
• pyrazoly chemie   farmakologie   MeSH
• RNA-polymerasa II metabolismus   MeSH
• X-vázaný inhibitor apoptózy genetika   metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

• Publikační typ
• abstrakt z konference MeSH

Accumulation of senescent cells in tissues with advancing age participates in the pathogenesis of several human age-associated diseases. Specific senescent secretome, the resistance of senescent cells to apoptotic stimuli, and lack of immune system response contribute to the accumulation of senescent cells and their adverse effects in tissues. Inhibition of antiapoptotic machinery, augmented in senescent cells, by BCL-2 protein family inhibitors represents a promising approach to eliminate senescent cells from tissues. This study aimed to explore synergistic and selective senolytic effects of anti-apoptotic BCL-2 family targeting compounds, particularly BH3 mimetics. Using human non-transformed cells RPE-1, BJ, and MRC-5 brought to ionizing radiation-, oncogene-, drug-induced and replicative senescence, we found synergy in combining MCL-1 selective inhibitors with other BH3 mimetics. In an attempt to uncover the mechanism of such synergy, we revealed that the surviving subpopulation of cells resistant to individually applied ABT-737/ABT-263, MIK665, ABT-199, and S63845 BCL-2 family inhibitors showed elevated MCL-1 compared to untreated control cells indicating the presence of a subset of cells expressing high MCL-1 levels and, therefore, resistant to BCL-2 inhibitors within the original population of senescent cells. Overall, we found that combining BCL-2 inhibitors can be beneficial for eliminating senescent cells, thereby enabling use of lower, potentially less toxic, doses of drugs compared to monotherapy, thereby overcoming the resistance of the subpopulation of senescent cells to monotherapy.

• MeSH
• apoptóza MeSH
• lidé MeSH
• protein MCL-1 metabolismus   MeSH
• protoonkogenní proteiny c-bcl-2 * antagonisté a inhibitory   MeSH
• stárnutí buněk * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

In addition to its ability to act as a promising inducer of tumor-specific cell death, TRAIL has also been shown to stimulate signaling pathways leading to cancer cell survival. We examined the changes of anti-apoptotic Mcl-1 protein level following TRAIL treatment of human cell lines representing different stages of colon carcinogenesis-adenocarcinoma (HT-29, HCT116) or secondary metastasis (SW620), together with cell line derived from human fetal colon (FHC). While TRAIL was capable of triggering an anti-apoptotic signaling leading to significant early ERK-mediated transcriptional up-regulation of Mcl-1 in selected colon adenocarcinoma cell lines, none or very limited effects were demonstrated in cell lines derived from colon lymph node metastasis or fetal colon, respectively. We demonstrated an immediate impact of Mcl-1 protein level manipulations on the course of early acute apoptotic response of colon adenocarcinoma cells to TRAIL. It is therefore essential to consider the dynamics of modulation of Mcl-1 level and the balance between TRAIL-induced pro- and anti-apoptotic pathways when predicting the response of cells in different stages of cancer development, and designing the anticancer therapy using TRAIL.

• MeSH
• epitelové buňky metabolismus   MeSH
• lidé MeSH
• metastázy nádorů patologie   MeSH
• nádorové buněčné linie MeSH
• nádory tračníku metabolismus   patologie   MeSH
• protein TRAIL fyziologie   MeSH
• protoonkogenní proteiny c-bcl-2 genetika   MeSH
• regulace genové exprese MeSH
• viabilita buněk MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• práce podpořená grantem MeSH

Chromosomal rearrangements and copy number variation are frequently observed in cancer cells, including multiple myeloma (MM). Karyotypic abnormalities seen in MM cells correlate with the disease stage and drug responses. Here, we investigate the nuclear arrangement of the 1q21 region; amplification of this region is an important diagnostic and prognostic marker of MM. We examined the lymphoblastoid cell line CD138- ARH-77, multiple myeloma CD138+ MOLP-8 cells, and the CD138+ bone marrow fraction of patients diagnosed with MM. In this experimental system, we observed that gamma-radiation and selected cytostatic drugs such as melphalan and dexamethasone did not significantly alter the nuclear radial arrangement of the 1q21 region and other relevant regions of chromosome 1. Similarly, conserved nuclear radial positioning after cytostatic treatment was observed for the c-myc, TP53, CCND1, and IgH loci. When analyzed Mcl-1, a protein encoded by a gene mapped to the 1q21 region, we found that the variant Mcl1S is highly expressed in multiple myeloma MOLP-8 cells, but not in peripheral blood lymphocytes of healthy donors or lymphoblastoid ARH-77 cells; this is in contrast to the expression pattern of the Mcl-1L variant. On the basis of these observations we suggest that the 1q21 region is an important diagnostic marker of MM, particularly the gene encoding the Mcl-1S variant, which can be easily detected by western analysis.

• MeSH
• buněčné jádro metabolismus   MeSH
• cyklin D1 analýza   MeSH
• interfáze MeSH
• lidé MeSH
• lidské chromozomy, pár 1 MeSH
• mapování chromozomů MeSH
• mnohočetný myelom diagnóza   genetika   patofyziologie   MeSH
• nádorové biomarkery analýza   MeSH
• nádorové buněčné linie MeSH
• protoonkogenní proteiny c-bcl-2 analýza   MeSH
• těžké řetězce imunoglobulinů genetika   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• práce podpořená grantem MeSH

MicroRNA hsa-miR-29 was connected to a number of malignancies. Its target genes are many, among them Mcl-1 that is expressed in three possible isoforms, one of which is anti-apoptotic and another one pro-apoptotic. Ratio of these two isoforms appears to affect cell response to external stimuli. We have demonstrated that miR-29b enhanced etoposide toxicity in HeLa cell line by modulating this ratio of Mcl-1 isoforms. However, it is not known whether the described miR-29 effect is common to various cancer types or even have the opposite effect. This represents a significant problem for possible future applications. In this report, we demonstrate that miR-29b affects toxicity of 60 μM etoposide in cell lines derived from selected malignancies. The mechanism, however, differs among the cell lines tested. Hep G2 cells demonstrated similar effect of miR-29b on etoposide toxicity as was described in HeLa cells, i.e. modulation of Mcl-1 expression. Target protein down-regulated by miR-29b resulting in enhanced etoposide toxicity in Caco-2 cells was, however, Bcl-2 protein. Moreover, H9c2, Hek-293 and ARPE-19 cell lines selected as a representatives of non-malignant cells, showed no effect of miR-29b on etoposide toxicity. Our data suggest that miR-29b could be a common enhancer of etoposide toxicity in malignant cells due to its modulation of Bcl family proteins.

• MeSH
• apoptóza účinky léků   genetika   MeSH
• buňky Hep G2 MeSH
• Caco-2 buňky MeSH
• etoposid * toxicita   farmakologie   MeSH
• fytogenní protinádorové látky farmakologie   toxicita   MeSH
• HEK293 buňky MeSH
• HeLa buňky MeSH
• lidé MeSH
• mikro RNA * genetika   metabolismus   MeSH
• protein MCL-1 * genetika   metabolismus   MeSH
• protoonkogenní proteiny c-bcl-2 genetika   metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

PURPOSE: Mantle cell lymphoma (MCL) is an aggressive subtype of B-cell non-Hodgkin lymphomas characterized by (over)expression of BCL2. A BCL2-targeting drug, venetoclax, has promising anticancer activity in MCL. We analyzed molecular mechanisms of venetoclax resistance in MCL cells and tested strategies to overcome it. EXPERIMENTAL DESIGN: We confirmed key roles of proapoptotic proteins BIM and NOXA in mediating venetoclax-induced cell death in MCL. Both BIM and NOXA are, however, differentially expressed in cell lines compared with primary cells. First, NOXA protein is significantly overexpressed in most MCL cell lines. Second, deletions of BIM gene harbored by three commonly used MCL cell lines (JEKO-1, MINO, and Z138) were not found by array comparative genomic hybridization using a validation set of 24 primary MCL samples. RESULTS: We demonstrated that MCL1 and NOXA play important roles in mediating resistance to venetoclax. Consequently, we tested an experimental treatment strategy based on cotargeting BCL2 with venetoclax and MCL1 with a highly specific small-molecule MCL1 inhibitor S63845. The combination of venetoclax and S63845 demonstrated synthetic lethality in vivo on a panel of five patient-derived xenografts established from patients with relapsed MCL with adverse cytogenetics. CONCLUSIONS: Our data strongly support investigation of venetoclax in combination with S63845 as an innovative treatment strategy for chemoresistant MCL patients with adverse cytogenetics in the clinical grounds.

• MeSH
• bicyklické sloučeniny heterocyklické farmakologie   MeSH
• chemorezistence MeSH
• lidé MeSH
• lokální recidiva nádoru farmakoterapie   metabolismus   patologie   MeSH
• lymfom z plášťových buněk farmakoterapie   metabolismus   patologie   MeSH
• myši inbrední NOD MeSH
• myši MeSH
• nádorové buněčné linie MeSH
• protein MCL-1 antagonisté a inhibitory   metabolismus   MeSH
• protinádorové látky farmakologie   MeSH
• protoonkogenní proteiny c-bcl-2 antagonisté a inhibitory   metabolismus   MeSH
• pyrimidiny farmakologie   MeSH
• sulfonamidy farmakologie   MeSH
• synergismus léků * MeSH
• thiofeny farmakologie   MeSH
• xenogenní modely - testy protinádorové aktivity MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH