BACKGROUND: Cell cycle progression and leukemia development are tightly regulated processes in which even a small imbalance in the expression of cell cycle regulatory molecules and microRNAs (miRNAs) can lead to an increased risk of cancer/leukemia development. Here, we focus on the study of a ubiquitous, multifunctional, and oncogenic miRNA-hsa-miR-155-5p (miR-155, MIR155HG), which is overexpressed in malignancies including chronic lymphocytic leukemia (CLL). Nonetheless, the precise mechanism of how miR-155 regulates the cell cycle in leukemic cells remains the subject of extensive research. METHODS: We edited the CLL cell line MEC-1 by CRISPR/Cas9 to introduce a short deletion within the MIR155HG gene. To describe changes at the transcriptome and miRNome level in miR-155-deficient cells, we performed mRNA-seq/miRNA-seq and validated changes by qRT-PCR. Flow cytometry was used to measure cell cycle kinetics. A WST-1 assay, hemocytometer, and Annexin V/PI staining assessed cell viability and proliferation. RESULTS: The limited but phenotypically robust miR-155 modification impaired cell proliferation, cell cycle, and cell ploidy. This was accompanied by overexpression of the negative cell cycle regulator p21/CDKN1A and Cyclin D1 (CCND1). We confirmed the overexpression of canonical miR-155 targets such as PU.1, FOS, SHIP-1, TP53INP1 and revealed new potential targets (FCRL5, ISG15, and MX1). CONCLUSIONS: We demonstrate that miR-155 deficiency impairs cell proliferation, cell cycle, transcriptome, and miRNome via deregulation of the MIR155HG/TP53INP1/CDKN1A/CCND1 axis. Our CLL model is valuable for further studies to manipulate miRNA levels to revert highly aggressive leukemic cells to nearly benign or non-leukemic types.

• MeSH
• Leukemia, Lymphocytic, Chronic, B-Cell * genetics   pathology   MeSH
• Cyclin D1 genetics   metabolism   MeSH
• Cyclin-Dependent Kinase Inhibitor p21 * genetics   metabolism   MeSH
• Cell Cycle Checkpoints * genetics   MeSH
• Humans MeSH
• MicroRNAs * genetics   metabolism   MeSH
• Cell Line, Tumor MeSH
• Cell Proliferation genetics   MeSH
• Heat-Shock Proteins MeSH
• Gene Expression Regulation, Leukemic MeSH
• Carrier Proteins genetics   metabolism   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH

Impairment of the p53 pathway is a critical event in cancer. Therefore, reestablishing p53 activity has become one of the most appealing anticancer therapeutic strategies. Here, we disclose the p53-activating anticancer drug (3S)-6,7-bis(hydroxymethyl)-5-methyl-3-phenyl-1H,3H-pyrrolo[1,2-c]thiazole (MANIO). MANIO demonstrates a notable selectivity to the p53 pathway, activating wild-type (WT)p53 and restoring WT-like function to mutant (mut)p53 in human cancer cells. MANIO directly binds to the WT/mutp53 DNA-binding domain, enhancing the protein thermal stability, DNA-binding ability, and transcriptional activity. The high efficacy of MANIO as an anticancer agent toward cancers harboring WT/mutp53 is further demonstrated in patient-derived cells and xenograft mouse models of colorectal cancer (CRC), with no signs of undesirable side effects. MANIO synergizes with conventional chemotherapeutic drugs, and in vitro and in vivo studies predict its adequate drug-likeness and pharmacokinetic properties for a clinical candidate. As a single agent or in combination, MANIO will advance anticancer-targeted therapy, particularly benefiting CRC patients harboring distinct p53 status.

• MeSH
• Apoptosis drug effects   genetics   MeSH
• Cisplatin pharmacology   MeSH
• Doxorubicin pharmacology   MeSH
• Fluorouracil pharmacology   MeSH
• HCT116 Cells MeSH
• Colorectal Neoplasms drug therapy   genetics   metabolism   pathology   MeSH
• Cell Cycle Checkpoints drug effects   genetics   MeSH
• Humans MeSH
• Mice, Nude MeSH
• Mice MeSH
• Cell Line, Tumor MeSH
• Tumor Suppressor Protein p53 agonists   genetics   metabolism   MeSH
• Drug Discovery MeSH
• Cell Proliferation drug effects   MeSH
• Antineoplastic Agents chemical synthesis   pharmacology   MeSH
• Antineoplastic Combined Chemotherapy Protocols pharmacology   MeSH
• Pyrroles chemical synthesis   pharmacology   MeSH
• Gene Expression Regulation, Neoplastic MeSH
• Drug Synergism MeSH
• Thiazoles chemical synthesis   pharmacology   MeSH
• Protein Binding MeSH
• Xenograft Model Antitumor Assays MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Mice MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Colorectal cancer (CRC) continues to be one of the leading malignancies and causes of tumour-related deaths worldwide. Both impaired DNA repair mechanisms and disrupted telomere length homeostasis represent key culprits in CRC initiation, progression and prognosis. Mechanistically, altered DNA repair results in the accumulation of mutations in the genome and, ultimately, in genomic instability. DNA repair also determines the response to chemotherapeutics in CRC treatment, suggesting its utilisation in the prediction of therapy response and individual approach to patients. Telomere attrition resulting in replicative senescence, simultaneously by-passing cell cycle checkpoints, is a hallmark of malignant transformation of the cell. Telomerase is almost ubiquitous in advanced solid cancers, including CRC, and its expression is fundamental to cell immortalisation. Therefore, there is a persistent effort to develop therapeutics, which are telomerase-specific and gentle to non-malignant tissues. However, in practice, we are still at the level of clinical trials. The current state of knowledge and the route, which the research takes, gives us a positive perspective that the problem of molecular models of telomerase activation and telomere length stabilisation will finally be solved. We summarise the current literature herein, by pointing out the crosstalk between proteins involved in DNA repair and telomere length homeostasis in relation to CRC.

• MeSH
• Chromosomal Instability MeSH
• Telomere Homeostasis genetics   MeSH
• Colorectal Neoplasms drug therapy   genetics   metabolism   pathology   MeSH
• Cell Cycle Checkpoints genetics   MeSH
• Humans MeSH
• Cell Transformation, Neoplastic genetics   metabolism   MeSH
• DNA Repair genetics   MeSH
• Cellular Senescence genetics   MeSH
• Telomerase genetics   metabolism   MeSH
• Telomere metabolism   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Review MeSH

Long transgenes are often used in mammalian genetics, e.g., to rescue mutations in large genes. In the course of experiments addressing the genetic basis of hybrid sterility caused by meiotic defects in mice bearing different alleles of Prdm9, we discovered that introduction of copy-number variation (CNV) via two independent insertions of long transgenes containing incomplete Prdm9 decreased testicular weight and epididymal sperm count. Transgenic animals displayed increased occurrence of seminiferous tubules with apoptotic cells at 18 days postpartum (dpp) corresponding to late meiotic prophase I, but not at 21 dpp. We hypothesized that long transgene insertions could cause asynapsis, but the immunocytochemical data revealed that the adult transgenic testes carried a similar percentage of asynaptic pachytene spermatocytes as the controls. These transgenic spermatocytes displayed less crossovers but similar numbers of unrepaired meiotic breaks. Despite slightly increased frequency of metaphase I spermatocytes with univalent chromosome(s) and reduced numbers of metaphase II spermatocytes, cytological studies did not reveal increased apoptosis in tubules containing the metaphase spermatocytes, but found an increased percentage of tubules carrying apoptotic spermatids. Sperm counts of subfertile animals inversely correlated with the transcription levels of the Psmb1 gene encoded within these two transgenes. The effect of the transgenes was dependent on sex and genetic background. Our results imply that the fertility of transgenic hybrid animals is not compromised by the impaired meiotic synapsis of homologous chromosomes, but can be negatively influenced by the increased expression of the introduced genes.

• MeSH
• Apoptosis genetics   MeSH
• DNA Breaks, Double-Stranded MeSH
• Fertility genetics   MeSH
• Genetic Background MeSH
• Cell Cycle Checkpoints genetics   MeSH
• Mice MeSH
• Pachytene Stage genetics   MeSH
• Sperm Count MeSH
• Spermatocytes metabolism   MeSH
• Testis anatomy & histology   metabolism   MeSH
• Transgenes * MeSH
• DNA Copy Number Variations * MeSH
• Organ Size MeSH
• Animals MeSH
• Check Tag
• Male MeSH
• Mice MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Protein phosphatase magnesium-dependent 1 delta (PPM1D) terminates cell response to genotoxic stress by negatively regulating the tumor suppressor p53 and other targets at chromatin. Mutations in the exon 6 of the PPM1D result in production of a highly stable, C-terminally truncated PPM1D. These gain-of-function PPM1D mutations are present in various human cancers but their role in tumorigenesis remains unresolved. Here we show that truncated PPM1D impairs activation of the cell cycle checkpoints in human non-transformed RPE cells and allows proliferation in the presence of DNA damage. Next, we developed a mouse model by introducing a truncating mutation in the PPM1D locus and tested contribution of the oncogenic PPM1DT allele to colon tumorigenesis. We found that p53 pathway was suppressed in colon stem cells harboring PPM1DT resulting in proliferation advantage under genotoxic stress condition. In addition, truncated PPM1D promoted tumor growth in the colon in Apcmin mice and diminished survival. Moreover, tumor organoids derived from colon of the ApcminPpm1dT/+ mice were less sensitive to 5-fluorouracil when compared to ApcminPpm1d+/+and the sensitivity to 5-fluorouracil was restored by inhibition of PPM1D. Finally, we screened colorectal cancer patients and identified recurrent somatic PPM1D mutations in a fraction of colon adenocarcinomas that are p53 proficient and show defects in mismatch DNA repair. In summary, we provide the first in vivo evidence that truncated PPM1D can promote tumor growth and modulate sensitivity to chemotherapy.

• MeSH
• Chromatin drug effects   MeSH
• Exons genetics   MeSH
• Fluorouracil pharmacology   MeSH
• Carcinogenesis drug effects   MeSH
• Cell Cycle Checkpoints genetics   MeSH
• Humans MeSH
• Mutation genetics   MeSH
• Mice MeSH
• Tumor Suppressor Protein p53 genetics   MeSH
• Colonic Neoplasms drug therapy   genetics   pathology   MeSH
• DNA Repair drug effects   MeSH
• DNA Damage drug effects   MeSH
• Cell Proliferation drug effects   MeSH
• Adenomatous Polyposis Coli Protein genetics   MeSH
• Protein Phosphatase 2C genetics   MeSH
• Gene Expression Regulation, Neoplastic drug effects   MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Mice MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Nádorová onemocnění jsou jednou z nejčastějších příčin úmrtí ve světě a postihují všechny věkové kategorie. Aktuální pokrok v terapii malignit je jak na poli chirurgické léčby, která stále zůstává stěžejní kurativní metodou časných stadií nemoci, tak na poli radioterapie, chemoterapie a v neposlední řadě i cílené léčby, která nám pomáhá překonávat limity standardní terapie nádorových onemocnění. I přes tento vývoj zůstávají některé malignity stále rezistentní vůči chemoterapii, radioterapii a často i primárně účinná cílená terapie po určité době naráží na vývoj sekundární rezistence. Role imunitního systému, respektive únik nádorových buněk z jeho dohledu, je ve vývoji nádorové choroby nezpochybnitelná. Právě ovlivnění jednotlivých kroků imunitní reakce se zdálo být velmi nadějným příslibem, jak ovlivnit průběh nádorového onemocnění a v posledních několika letech se tento předpoklad potvrzuje u stále se rozšiřujícího počtu diagnóz, kde moderní imunoterapie u pacientů prodlužuje přežívání. Klíčovou diagnózou se stalo onemocnění, které díky své chemorezistenci a radiorezistenci desítky let vyzývalo k objevení dalších možností léčby. Maligní melanom tak stál na začátku objevení účinnosti mnoha nových léků ze skupiny imunoterapeutik.

Tumor diseases are one of the most common causes of death worldwide and affect all ages. Current advances in malignant therapy are both in the field of surgical treatment, which remains a key curative method of early stages of the disease, as well as in the field of radiotherapy, chemotherapy and, last but not least, targeted treatments that help us to overcome the limits of standard cancer therapies. Even primarily effective targeted therapy often encounters the development of secondary resistance after a certain period of time. The role of the immune system, or the tumor cells escaping from its surveillance, is unquestionable in the development of cancer. Influencing the individual steps of the immune response seemed to be a very promising method how to influence the course of cancer, and in the last few years, this assumption has been confirmed by an ever-expanding number of diagnoses where modern immunotherapy in patients prolongs survival. The key diagnosis has become a disease that, thanks to its chemoresistance and radioresistance, for decades prompted the discovery of other treatment options.

• MeSH
• B7-H1 Antigen antagonists & inhibitors   therapeutic use   MeSH
• BCG Vaccine history   therapeutic use   MeSH
• History of Medicine MeSH
• Immunotherapy * methods   adverse effects   trends   MeSH
• Interferon-alpha history   therapeutic use   MeSH
• Interleukin-2 history   therapeutic use   MeSH
• Combined Modality Therapy * methods   trends   MeSH
• Cell Cycle Checkpoints * genetics   immunology   MeSH
• Medical Oncology methods   trends   MeSH
• Humans MeSH
• Antibodies, Monoclonal MeSH
• Biomarkers, Tumor MeSH
• Check Tag
• Humans MeSH
• Publication type
• Review MeSH

Ačkoli v obraně proti nádorům je imunitní systém většinou mnohem méně úspěšný než v obraně proti infekcím, v poslední době byly objasněny příčiny této skutečnosti a byly nalezeny způsoby, jak protinádorovou účinnost imunitního systému výrazně zlepšit. Byla identifikována řada více či méně specifických nádorových antigenů, byly objasněny mnohé buněčné a molekulární mechanismy, kterými se nádory brání proti imunitním zbraním. Velký význam mělo formulování a potvrzení platnosti hypotézy imunitní editace nádorů vysvětlující komplikované vztahy mezi nádory a imunitním systémem. Pro teoretické poznání tohoto vztahu bylo důležité objasnění několika úrovní tlumivých mechanismů, jimiž imunitní systém může paradoxně nádory chránit. Poznatky základního výzkumu vedly konečně k vývoji účinných imunoterapeutických postupů, které jsou nadále zdokonalovány a mohou se stát v blízké budoucnosti hlavními metodami protinádorové terapie.

Although the immune system is usually much less efficient in defense against tumors as compared to defense against infections, recent discoveries elucidated the causes of this phenomenon and made it possible to improve the efficiency of anti-tumor immune mechanisms. A number of more or less specific tumor antigens have been identified and several mechanisms involved in resistance of tumors to the immune weapons have been discovered. The formulation of tumor immune editing hypothesis and its validation was essential for current understanding of the complex relationship between the tumor and the immune system. The identification of several levels of immunosuppressive mechanisms has been crucial for understanding of the paradoxical phenomenon of immune protection of tumors. Decades of basic research culminated in development of efficient immunotherapeutic approaches which, after further improvements, have the potential to became major tools of anti-tumor therapies.

• MeSH
• Antigens, Neoplasm immunology   classification   therapeutic use   MeSH
• Immunologic Surveillance * physiology   immunology   MeSH
• Immune System immunology   drug effects   MeSH
• Immunotherapy * history   methods   trends   MeSH
• Combined Modality Therapy * methods   trends   MeSH
• Cell Cycle Checkpoints genetics   immunology   MeSH
• Medical Oncology methods   trends   MeSH
• Humans MeSH
• Antibodies, Monoclonal immunology   therapeutic use   MeSH
• Immune System Diseases complications   metabolism   MeSH
• Oncolytic Virotherapy methods   trends   MeSH
• Cancer Vaccines immunology   therapeutic use   MeSH
• Retroviridae genetics   immunology   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Review MeSH

Pokrok v protinádorové imunoterapii je v posledních deseti letech pozoruhodný. Koncept blokády kontrolních bodů imunitní odpovědi (tzv. checkpointů) pomocí monoklonálních protilátek (checkpoint inhibitorů) otevřel novou cestu v léčbě nádorů. Checkpoint inhibitory obnovují a zesilují protinádorovou aktivitu cytotoxických T lymfocytů. Jejich účinnost a přijatelná toxicita byla prokázána v řadě klinických studií. Checkpoint inhibitory jsou dnes schváleny k léčbě řady malignit a u ostatních běží intenzivní výzkum. Stále však neznáme odpovědi na řadu klíčových otázek. Více než cokoliv jiného naléhavě potřebujeme spolehlivé prediktivní biomarkery účinnosti a toxicity. Poté budeme schopni lépe vybírat vhodné pacienty k léčbě. Imunoterapie je právem považována za pátý pilíř protinádorové léčby, vedle chirurgie, radioterapie, chemoterapie a cílené léčby.

The progress in anticancer immunotherapy has been remarkable in the last decade. The concept of immune checkpoint blocade with monoclonal antibodies (checkpoint inhibitors) has opened a new way in the treatment of cancers. Immune checkpoint inhibitors restore and augment antitumor functions of cytotoxic T cells. Efficacy and acceptable safety of immune checkpoint inhibitors have been demonstrated in many clinical trials. Checkpoint inhibitors are now approved for treatment of variety cancers and intensive research on others is under way. There are still many challenges. More than anything else we urgently need reliable biomarkers of efficacy and toxicity. Then we will be able to better select suitable patients for treatment. Immunotherapy is rightly considered to be the fifth pillar of anticancer treatment, next to surgery, radiotherapy, chemotherapy and targeted therapy.

• MeSH
• CTLA-4 Antigen antagonists & inhibitors   immunology   therapeutic use   MeSH
• B7-H1 Antigen antagonists & inhibitors   immunology   therapeutic use   MeSH
• History of Medicine MeSH
• Immunotherapy * history   methods   trends   MeSH
• Combined Modality Therapy * methods   trends   MeSH
• Cell Cycle Checkpoints * genetics   immunology   drug effects   MeSH
• Medical Oncology history   methods   trends   MeSH
• Humans MeSH
• Melanoma drug therapy   MeSH
• Antibodies, Monoclonal therapeutic use   MeSH
• Head and Neck Neoplasms drug therapy   MeSH
• Lung Neoplasms drug therapy   MeSH
• Urologic Neoplasms drug therapy   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Research Support, Non-U.S. Gov't MeSH
• Review MeSH

• MeSH
• Early Detection of Cancer * methods   trends   MeSH
• Diagnostic Techniques, Respiratory System trends   MeSH
• Drug Therapy methods   trends   MeSH
• Immunotherapy methods   trends   MeSH
• Combined Modality Therapy * methods   trends   MeSH
• Cell Cycle Checkpoints genetics   immunology   drug effects   MeSH
• Humans MeSH
• Neoplasm Recurrence, Local diagnosis   therapy   MeSH
• Interdisciplinary Communication MeSH
• Lung Neoplasms diagnosis   classification   therapy   MeSH
• Carcinoma, Non-Small-Cell Lung * diagnosis   drug therapy   therapy   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Introductory Journal Article MeSH

Altered cell metabolism is a hallmark of cancer, and targeting specific metabolic nodes is considered an attractive strategy for cancer therapy. In this study, we evaluate the effects of metabolic stressors on the deregulated ERK pathway in melanoma cells bearing activating mutations of the NRAS or BRAF oncogenes. We report that metabolic stressors promote the dimerization of KSR proteins with CRAF in NRAS-mutant cells, and with oncogenic BRAF in BRAFV600E-mutant cells, thereby enhancing ERK pathway activation. Despite this similarity, the two genomic subtypes react differently when a higher level of metabolic stress is induced. In NRAS-mutant cells, the ERK pathway is even more stimulated, while it is strongly downregulated in BRAFV600E-mutant cells. We demonstrate that this is caused by the dissociation of mutant BRAF from KSR and is mediated by activated AMPK. Both types of ERK regulation nevertheless lead to cell cycle arrest. Besides studying the effects of the metabolic stressors on ERK pathway activity, we also present data suggesting that for efficient therapies of both genomic melanoma subtypes, specific metabolic targeting is necessary.

• MeSH
• Enzyme Activation MeSH
• Extracellular Signal-Regulated MAP Kinases metabolism   MeSH
• Stress, Physiological * MeSH
• Glucose metabolism   MeSH
• Glycolysis MeSH
• GTP Phosphohydrolases genetics   metabolism   MeSH
• Cell Cycle Checkpoints genetics   MeSH
• Humans MeSH
• Melanoma genetics   metabolism   MeSH
• Membrane Proteins genetics   metabolism   MeSH
• Protein Multimerization * MeSH
• Mutation MeSH
• Cell Line, Tumor MeSH
• Protein Kinases chemistry   genetics   metabolism   MeSH
• 14-3-3 Proteins chemistry   metabolism   MeSH
• raf Kinases chemistry   genetics   metabolism   MeSH
• Recombinant Fusion Proteins MeSH
• Oxygen Consumption MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH