Latest Publications

@article{boudny_novel_2019,

title = {Novel CHK1 inhibitor MU380 exhibits significant single-agent activity in TP53-mutated chronic lymphocytic leukemia cells.},

author = {Miroslav Boudny and Jana Zemanova and Prashant Khirsariya and Marek Borsky and Jan Verner and Jana Cerna and Alexandra Oltova and Vaclav Seda and Marek Mraz and Josef Jaros and Zuzana Jaskova and Michaela Spunarova and Yvona Brychtova and Karel Soucek and Stanislav Drapela and Marie Kasparkova and Jiri Mayer and Kamil Paruch and Martin Trbusek},

doi = {10.3324/haematol.2018.203430},

issn = {1592-8721 0390-6078},

year  = {2019},

date = {2019-12-01},

journal = {Haematologica},

volume = {104},

number = {12},

pages = {2443–2455},

abstract = {Introduction of small-molecule inhibitors of B-cell receptor signaling and BCL2 protein significantly improves therapeutic options in chronic lymphocytic leukemia. However, some patients suffer from adverse effects mandating treatment discontinuation, and cases with TP53 defects more frequently experience early progression of the disease. Development of alternative therapeutic approaches is, therefore, of critical importance. Here we report details of the anti-chronic lymphocytic leukemia single-agent activity of MU380, our recently identified potent, selective, and metabolically robust inhibitor of checkpoint kinase 1. We also describe a newly developed enantioselective synthesis of MU380, which allows preparation of gram quantities of the substance. Checkpoint kinase 1 is a master regulator of replication operating primarily in intra-S and G(2)/M cell cycle checkpoints. Initially tested in leukemia and lymphoma cell lines, MU380 significantly potentiated efficacy of gemcitabine, a clinically used inducer of replication stress. Moreover, MU380 manifested substantial single-agent activity in both TP53-wild type and TP53-mutated leukemia and lymphoma cell lines. In chronic lymphocytic leukemia-derived cell lines MEC-1, MEC-2 (both TP53-mut), and OSU-CLL (TP53-wt) the inhibitor impaired cell cycle progression and induced apoptosis. In primary clinical samples, MU380 used as a single-agent noticeably reduced the viability of unstimulated chronic lymphocytic leukemia cells as well as those induced to proliferate by anti-CD40/IL-4 stimuli. In both cases, effects were comparable in samples harboring p53 pathway dysfunction (TP53 mutations or ATM mutations) and TP53-wt/ATM-wt cells. Lastly, MU380 also exhibited significant in vivo activity in a xenotransplant mouse model (immunodeficient strain NOD-scid IL2Rγ(null) ) where it efficiently suppressed growth of subcutaneous tumors generated from MEC-1 cells.},

note = {Place: Italy},

keywords = {*Drug Synergism, *Mutation, Animals, Antimetabolites, Antineoplastic/pharmacology, Apoptosis, B-Cell/*drug therapy/genetics/pathology, Biomarkers, Cell Cycle, Cell Proliferation, Checkpoint Kinase 1/*antagonists & inhibitors, Chronic, Cultured, Deoxycytidine/analogs & derivatives/pharmacology, Drug resistance, Female, gemcitabine, Gene Expression Regulation, Humans, Inbred NOD, Leukemia, Lymphocytic, Mice, Neoplasm/drug effects, Neoplastic/*drug effects, Piperidines/*pharmacology, Protein Kinase Inhibitors/pharmacology, Pyrazoles/*pharmacology, Pyrimidines/*pharmacology, SCID, Tumor Cells, Tumor Suppressor Protein p53/*genetics, Tumor/genetics, Xenograft Model Antitumor Assays},

pubstate = {published},

tppubtype = {article}

}

@article{vargova_hypericin_2018,

title = {Hypericin affects cancer side populations via competitive inhibition of BCRP.},

author = {Jana Vargová and Jaromír Mikeš and Rastislav Jendželovský and Lucia Mikešová and Barbora Kuchárová and Ľubomír Čulka and Radek Fedr and Ján Remšík and Karel Souček and Alois Kozubík and Peter Fedoročko},

doi = {10.1016/j.biopha.2018.01.074},

issn = {1950-6007 0753-3322},

year  = {2018},

date = {2018-03-01},

journal = {Biomedicine & pharmacotherapy = Biomedecine & pharmacotherapie},

volume = {99},

pages = {511–522},

abstract = {OBJECTIVE: Cancer stem-like cells (CSLCs) are considered a root of tumorigenicity and resistance. However, their identification remains challenging. The use of the side population (SP) assay as a credible marker of CSLCs remains controversial. The SP assay relies on the elevated activity of ABC transporters that, in turn, can be modulated by hypericin (HYP), a photosensitizer and bioactive compound of St. John's Wort (Hypericum perforatum), a popular over-the-counter antidepressant. Here we aimed to comprehensively characterize the SP phenotype of cancer cells and to determine the impact of HYP on these cells. METHODS: Flow cytometry and sorting-based assays were employed, including CD24-, CD44-, CD133-, and ALDH-positivity, clonogenicity, 3D-forming ability, ABC transporter expression and activity, and intracellular accumulation of HYP/Hoechst 33342. The tumorigenic ability of SP, nonSP, and HYP-treated cells was verified by xenotransplantation into immunodeficient mice. RESULTS: The SP phenotype was associated with elevated expression of several investigated transporters and more intensive growth in non-adherent conditions but not with higher clonogenicity, tumorigenicity or ALDH-positivity. Despite stimulated BCRP level and MRP1 activity, HYP reversibly decreased the SP proportion, presumably via competitive inhibition of BCRP. HYP-selected SP cells acquired additional traits of resistance and extensively eliminated HYP. CONCLUSIONS: Our results suggest that SP is not an unequivocal CSLC-marker. However, SP could play an important role in modulating HYP-treatment and serve as a negative predictive tool for HYP-based therapies. Moreover, the use of supplements containing HYP by cancer patients should be carefully considered, due to its proposed effect on drug efflux and complex impact on tumor cells, which have not yet been sufficiently characterized.},

note = {Place: France},

keywords = {ABC transporters, Aldehyde Dehydrogenase/metabolism, Animals, Anthracenes, ATP Binding Cassette Transporter, Biomarkers, Cancer stem-like cells, Carcinogenesis/drug effects/metabolism/pathology, Cell Line, Cellular/drug effects/metabolism/pathology, Clone Cells, Drug resistance, Humans, Hypericin, Member 1/metabolism, Member 2/*metabolism, Mice, Neoplasm Proteins/*metabolism, Neoplasms/*metabolism/*pathology, Neoplastic Stem Cells/drug effects/metabolism/pathology, Perylene/*analogs & derivatives/pharmacology, Phenotype, SCID, Side population, Side-Population Cells/drug effects/*pathology, Spheroids, St. John’s wort, Subfamily B, Subfamily G, Substrate Specificity/drug effects, Survival Analysis, Tumor, Tumor/metabolism},

pubstate = {published},

tppubtype = {article}

}

@article{remsik_plasticity_2018,

title = {Plasticity and intratumoural heterogeneity of cell surface antigen expression in breast cancer.},

author = {Ján Remšík and Radek Fedr and Jiří Navrátil and Lucia Binó and Eva Slabáková and Pavel Fabian and Marek Svoboda and Karel Souček},

doi = {10.1038/bjc.2017.497},

issn = {1532-1827 0007-0920},

year  = {2018},

date = {2018-03-01},

journal = {British journal of cancer},

volume = {118},

number = {6},

pages = {813–819},

abstract = {Background:The intratumoural heterogeneity, often driven by epithelial-to-mesenchymal transition (EMT), significantly contributes to chemoresistance and disease progression in adenocarcinomas. Methods:We introduced a high-throughput screening platform to identify surface antigens that associate with epithelial–mesenchymal plasticity in well-defined pairs of epithelial cell lines and their mesenchymal counterparts. Using multicolour flow cytometry, we then analysed the expression of 10 most robustly changed antigens and identified a 10-molecule surface signature, in pan-cytokeratin-positive/EpCAM-positive and -negative fractions of dissociated breast tumours. Results:We found that surface CD9, CD29, CD49c, and integrin ß5 are lost in breast cancer cells that underwent EMT in vivo. The tetraspanin family member CD9 was concordantly downregulated both in vitro and in vivo and associated with epithelial phenotype and favourable prognosis. Conclusions:We propose that overall landscape of 10-molecule surface signature expression reflects the epithelial–mesenchymal plasticity in breast cancer.},

note = {Place: England},

keywords = {Antigens, Biomarkers, Breast Neoplasms/*genetics/*immunology/pathology, Cell Line, Cell Plasticity/immunology, Cellular Reprogramming/physiology, Epithelial-Mesenchymal Transition/immunology, Female, Flow Cytometry, Genetic, High-Throughput Screening Assays, Humans, Neoplasm Metastasis, Neoplasm/*biosynthesis/immunology, Surface/*biosynthesis/immunology, Tetraspanin 29/biosynthesis/immunology, Transcription, Tumor},

pubstate = {published},

tppubtype = {article}

}

@article{samadder_synthesis_2017,

title = {Synthesis and Profiling of a Novel Potent Selective Inhibitor of CHK1 Kinase Possessing Unusual N-trifluoromethylpyrazole Pharmacophore Resistant to Metabolic N-dealkylation.},

author = {Pounami Samadder and Tereza Suchánková and Ondřej Hylse and Prashant Khirsariya and Fedor Nikulenkov and Stanislav Drápela and Nicol Straková and Petr Vaňhara and Kateřina Vašíčková and Hana Kolářová and Lucia Binó and Miroslava Bittová and Petra Ovesná and Peter Kollár and Radek Fedr and Milan Ešner and Josef Jaroš and Aleš Hampl and Lumír Krejčí and Kamil Paruch and Karel Souček},

doi = {10.1158/1535-7163.MCT-17-0018},

issn = {1538-8514 1535-7163},

year  = {2017},

date = {2017-09-01},

journal = {Molecular cancer therapeutics},

volume = {16},

number = {9},

pages = {1831–1842},

abstract = {Checkpoint-mediated dependency of tumor cells can be deployed to selectively kill them without substantial toxicity to normal cells. Specifically, loss of CHK1, a serine threonine kinase involved in the surveillance of the G(2)-M checkpoint in the presence of replication stress inflicted by DNA-damaging drugs, has been reported to dramatically influence the viability of tumor cells. CHK1's pivotal role in maintaining genomic stability offers attractive opportunity for increasing the selectivity, effectivity, and reduced toxicity of chemotherapy. Some recently identified CHK1 inhibitors entered clinical trials in combination with DNA antimetabolites. Herein, we report synthesis and profiling of MU380, a nontrivial analogue of clinically profiled compound SCH900776 possessing the highly unusual N-trifluoromethylpyrazole motif, which was envisioned not to undergo metabolic oxidative dealkylation and thereby provide greater robustness to the compound. MU380 is a selective and potent inhibitor of CHK1 which sensitizes a variety of tumor cell lines to hydroxyurea or gemcitabine up to 10 times. MU380 shows extended inhibitory effects in cells, and unlike SCH900776, does not undergo in vivo N-dealkylation to the significantly less selective metabolite. Compared with SCH900776, MU380 in combination with GEM causes higher accumulation of DNA damage in tumor cells and subsequent enhanced cell death, and is more efficacious in the A2780 xenograft mouse model. Overall, MU380 represents a novel state-of-the-art CHK1 inhibitor with high potency, selectivity, and improved metabolic robustness to oxidative N-dealkylation. Mol Cancer Ther; 16(9); 1831-42. ©2017 AACR.},

note = {Place: United States},

keywords = {Animal, Animals, Antineoplastic Agents/*chemical synthesis/*pharmacology, Apoptosis/drug effects, Biomarkers, Cell Cycle Checkpoints/drug effects, Cell Cycle/drug effects, Cell Line, Checkpoint Kinase 1/*antagonists & inhibitors, Dealkylation/drug effects, Disease Models, Dose-Response Relationship, Drug, Drug resistance, Humans, Methylation, Mice, Molecular Structure, Neoplasm/*drug effects, Protein Kinase Inhibitors/*chemical synthesis/*pharmacology, Pyrazoles/pharmacology, Pyrimidines/pharmacology, Tumor, Xenograft Model Antitumor Assays},

pubstate = {published},

tppubtype = {article}

}

@article{fedr_automatic_2013,

title = {Automatic cell cloning assay for determining the clonogenic capacity of cancer and cancer stem-like cells.},

author = {Radek Fedr and Zuzana Pernicová and Eva Slabáková and Nicol Straková and Jan Bouchal and Michal Grepl and Alois Kozubík and Karel Souček},

doi = {10.1002/cyto.a.22273},

issn = {1552-4930 1552-4922},

year  = {2013},

date = {2013-05-01},

journal = {Cytometry. Part A : the journal of the International Society for Analytical Cytology},

volume = {83},

number = {5},

pages = {472–482},

abstract = {The clonogenic assay is a well-established in vitro method for testing the survival and proliferative capability of cells. It can be used to determine the cytotoxic effects of various treatments including chemotherapeutics and ionizing radiation. However, this approach can also characterize cells with different phenotypes and biological properties, such as stem cells or cancer stem cells. In this study, we implemented a faster and more precise method for assessing the cloning efficiency of cancer stem-like cells that were characterized and separated using a high-speed cell sorter. Cell plating onto a microplate using an automatic cell deposition unit was performed in a single-cell or dilution rank mode by the fluorescence-activated cell sorting method. We tested the new automatic cell-cloning assay (ACCA) on selected cancer cell lines and compared it with the manual approach. The obtained results were also compared with the results of the limiting dilution assay for different cell lines. We applied the ACCA to analyze the cloning capacity of different subpopulations of prostate and colon cancer cells based on the expression of the characteristic markers of stem (CD44 and CD133) and cancer stem cells (TROP-2, CD49f, and CD44). Our results revealed that the novel ACCA is a straightforward approach for determining the clonogenic capacity of cancer stem-like cells identified in both cell lines and patient samples.},

note = {Place: United States},

keywords = {*Cell Proliferation, AC133 Antigen, Antigens, Biomarkers, CD/metabolism, Cell Adhesion Molecules/metabolism, Cell Line, Cell Survival, Colonic Neoplasms/metabolism/*pathology, Flow Cytometry/*methods, Glycoproteins/metabolism, Humans, Hyaluronan Receptors/metabolism, In Vitro Techniques, Integrin alpha6/metabolism, Male, Neoplasm/metabolism, Neoplastic Stem Cells/metabolism/*pathology, Peptides/metabolism, Prostatic Neoplasms/metabolism/*pathology, Tumor, Tumor Stem Cell Assay/*methods, Tumor/metabolism},

pubstate = {published},

tppubtype = {article}

}