18 results on '"Marcar, L."'
Search Results
2. Frequent traces of EBV infection in Hodgkin and non-Hodgkin lymphomas classified as EBV-negative by routine methods: expanding the landscape of EBV-related lymphomas [Tiacci and Lazzi are co-senior authors]
- Author
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Mundo, L., Del Porro, L., Granai, M., Siciliano, M. C., Mancini, V., Santi, R., Marcar, L., Vrzalikova, K., Vergoni, F., Di Stefano, G., Schiavoni, G., Segreto, G., Onyango, N., Nyagol, J. A., Amato, T., Bellan, C., Anagnostopoulos, I., Falini, B., Leoncini, L., Tiacci, E., and Lazzi, S.
- Published
- 2020
3. The Presence of a High Peak Feature Within Low-Average Shear Stimuli Induces Quiescence in Venous Endothelial Cells
- Author
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Franzoni, M., primary, O’Connor, D. T., additional, Marcar, L., additional, Power, D., additional, Moloney, M. A., additional, Kavanagh, E. G., additional, Leask, R. L., additional, Nolan, J., additional, Kiely, P. A., additional, and Walsh, M. T., additional
- Published
- 2019
- Full Text
- View/download PDF
4. Radiation Resistance in KRAS-Mutated Lung Cancer Is Enabled by Stem-like Properties Mediated by an Osteopontin-EGFR Pathway
- Author
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Wang, M., Han, J., Marcar, L., Black, J., Liu, Q., Li, X., Nagulapalli, K., Sequist, L. V., Mak, R. H., Benes, C. H., Hong, T. S., Gurtner, K., Krause, M., Baumann, M., Kang, J. X., Whetstine, J. R., and Willers, H.
- Subjects
neoplasms - Abstract
Lung cancers with activating KRAS mutations are characterized by treatment resistance and poor prognosis. In particular, the basis for their resistance to radiation therapy is poorly understood. Here, we describe a radiation resistance phenotype conferred by a stem-like subpopulation characterized by mitosis-like condensed chromatin (MLCC), high CD133 expression, invasive potential, and tumor-initiating properties. Mechanistic investigations defined a pathway involving osteopontin and the EGFR in promoting this phenotype. Osteopontin/EGFR-dependent MLCC protected cells against radiation-induced DNA double-strand breaks and repressed putative negative regulators of stem-like properties, such as CRMP1 and BIM. The MLCC-positive phenotype defined a subset of KRAS-mutated lung cancers that were enriched for co-occurring genomic alterations in TP53 and CDKN2A. Our results illuminate the basis for the radiation resistance of KRAS-mutated lung cancers, with possible implications for prognostic and therapeutic strategies. Cancer Res; 77(8); 2018-28. ©2017 AACR.
- Published
- 2017
5. PARP1 Inhibition Radiosensitizes A Subset Of Genotype-Defined Bladder Cancers Through Modulation Of Reactive Oxygen Species
- Author
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Willers, H., primary, Liu, Q., additional, Efstathiou, J.A., additional, Gheorghiu, L., additional, Drumm, M.R., additional, Clayman, R.H., additional, Eidelman, A., additional, Wszolek, M.F., additional, Feldman, A.S., additional, Wang, M., additional, Marcar, L., additional, Citrin, D.E., additional, Wu, C.L., additional, and Benes, C., additional
- Published
- 2016
- Full Text
- View/download PDF
6. Identification of Nanoparticle Properties for Optimal Drug Delivery across a Physiological Cell Barrier.
- Author
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Ross AM, Cahalane RM, Walsh DR, Grabrucker AM, Marcar L, and Mulvihill JJE
- Abstract
Nanoparticles (NPs) represent an attractive strategy to overcome difficulties associated with the delivery of therapeutics. Knowing the optimal properties of NPs to address these issues could allow for improved in vivo responses. This work investigated NPs prepared from 5 materials of 3 sizes and 3 concentrations applied to a cell barrier model. The NPs permeability across a cell barrier and their effects on cell barrier integrity and cell viability were evaluated. The properties of these NPs, as determined in water (traditional) vs. media (realistic), were compared to cell responses. It was found that for all cellular activities, NP properties determined in media was the best predictor of the cell response. Notably, ZnO NPs caused significant alterations to cell viability across all 3 cell lines tested. Importantly, we report that the zeta potential of NPs correlates significantly with NP permeability and NP-induced changes in cell viability. NPs with physiological-based zeta potential of -12 mV result in good cell barrier penetration without considerable changes in cell viability.
- Published
- 2023
- Full Text
- View/download PDF
7. The effect of serum starvation on tight junctional proteins and barrier formation in Caco-2 cells.
- Author
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Ross AM, Walsh DR, Cahalane RM, Marcar L, and Mulvihill JJE
- Abstract
Assessing the ability of pharmaceutics to cross biological barriers and reach the site-of-action requires faithful representation of these barriers in vitro . Difficulties have arisen in replicating in vivo resistance in vitro . This paper investigated serum starvation as a method to increase Caco-2 barrier stability and resistance. The effect of serum starvation on tight junction production was examined using transwell models; specifically, transendothelial electrical resistance (TEER), and the expression and localization of tight junction proteins, occludin and zonula occludens-1 (ZO-1), were studied using western blotting and immunofluorescence. Changing cells to serum-free media 2 days post-seeding resulted in TEER readings of nearly 5000 Ω cm
2 but the TEER rapidly declined subsequently. Meanwhile, exchanging cells to serum-free media 4-6 days post-seeding produced barriers with resistance readings between 3000 and 4000 Ω cm2 , which could be maintained for 18 days. This corresponded to an increase in occludin levels. Serum starvation as a means of barrier formation is simple, reproducible, and cost-effective. It could feasibly be implemented in a variety of pre-clinical pharmaceutical assessments of drug permeability across various biological barriers with the view to improving the clinical translation of novel therapeutics., Competing Interests: None., (© 2021 The Authors. Published by Elsevier B.V.)- Published
- 2021
- Full Text
- View/download PDF
8. Correction: Frequent traces of EBV infection in Hodgkin and non-Hodgkin lymphomas classified as EBV-negative by routine methods: expanding the landscape of EBV-related lymphomas.
- Author
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Mundo L, Del Porro L, Granai M, Siciliano MC, Mancini V, Santi R, Marcar L, Vrzalikova K, Vergoni F, Di Stefano G, Schiavoni G, Segreto G, Onyango N, Nyagol JA, Amato T, Bellan C, Anagnostopoulos I, Falini B, Leoncini L, Tiacci E, and Lazzi S
- Abstract
An amendment to this paper has been published and can be accessed via a link at the top of the paper.
- Published
- 2020
- Full Text
- View/download PDF
9. Frequent traces of EBV infection in Hodgkin and non-Hodgkin lymphomas classified as EBV-negative by routine methods: expanding the landscape of EBV-related lymphomas.
- Author
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Mundo L, Del Porro L, Granai M, Siciliano MC, Mancini V, Santi R, Marcar L, Vrzalikova K, Vergoni F, Di Stefano G, Schiavoni G, Segreto G, Onyango N, Nyagol JA, Amato T, Bellan C, Anagnostopoulos I, Falini B, Leoncini L, Tiacci E, and Lazzi S
- Subjects
- Epstein-Barr Virus Infections diagnosis, Hodgkin Disease diagnosis, Humans, Italy, Lymphoma, Non-Hodgkin diagnosis, Molecular Diagnostic Techniques, U937 Cells, Viral Load, Epstein-Barr Virus Infections virology, Epstein-Barr Virus Nuclear Antigens genetics, Herpesvirus 4, Human genetics, Hodgkin Disease virology, Lymphoma, Non-Hodgkin virology, RNA, Messenger genetics, RNA, Viral genetics
- Abstract
The Epstein-Barr virus (EBV) is linked to various B-cell lymphomas, including Burkitt lymphoma (BL), classical Hodgkin lymphoma (cHL) and diffuse large B-cell lymphoma (DLBCL) at frequencies ranging, by routine techniques, from 5 to 10% of cases in DLBCL to >95% in endemic BL. Using higher-sensitivity methods, we recently detected EBV traces in a few EBV-negative BL cases, possibly suggesting a "hit-and-run" mechanism. Here, we used routine and higher-sensitivity methods (qPCR and ddPCR for conserved EBV genomic regions and miRNAs on microdissected tumor cells; EBNA1 mRNA In situ detection by RNAscope) to assess EBV infection in a larger lymphoma cohort [19 BL, 34 DLBCL, 44 cHL, 50 follicular lymphomas (FL), 10 T-lymphoblastic lymphomas (T-LL), 20 hairy cell leukemias (HCL), 10 mantle cell lymphomas (MCL)], as well as in several lymphoma cell lines (9 cHL and 6 BL). qPCR, ddPCR, and RNAscope consistently documented the presence of multiple EBV nucleic acids in rare tumor cells of several cases EBV-negative by conventional methods that all belonged to lymphoma entities clearly related to EBV (BL, 6/9 cases; cHL, 16/32 cases; DLBCL, 11/30 cases), in contrast to fewer cases (3/47 cases) of FL (where the role of EBV is more elusive) and no cases (0/40) of control lymphomas unrelated to EBV (HCL, T-LL, MCL). Similarly, we revealed traces of EBV infection in 4/5 BL and 6/7 HL cell lines otherwise conventionally classified as EBV negative. Interestingly, additional EBV-positive cases (1 DLBCL, 2 cHL) relapsed as EBV-negative by routine methods while showing EBNA1 expression in rare tumor cells by RNAscope. The relapse specimens were clonally identical to their onset biopsies, indicating that the lymphoma clone can largely loose the EBV genome over time but traces of EBV infection are still detectable by high-sensitivity methods. We suggest EBV may contribute to lymphoma pathogenesis more widely than currently acknowledged.
- Published
- 2020
- Full Text
- View/download PDF
10. Acquired Resistance of EGFR-Mutated Lung Cancer to Tyrosine Kinase Inhibitor Treatment Promotes PARP Inhibitor Sensitivity.
- Author
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Marcar L, Bardhan K, Gheorghiu L, Dinkelborg P, Pfäffle H, Liu Q, Wang M, Piotrowska Z, Sequist LV, Borgmann K, Settleman JE, Engelman JA, Hata AN, and Willers H
- Subjects
- Animals, Apoptosis drug effects, Cell Line, Tumor, Cell Survival drug effects, Cell Survival genetics, DNA Damage, Drug Resistance, Neoplasm drug effects, Drug Resistance, Neoplasm genetics, ErbB Receptors antagonists & inhibitors, ErbB Receptors genetics, ErbB Receptors metabolism, Female, Humans, Lung Neoplasms enzymology, Lung Neoplasms genetics, Lung Neoplasms metabolism, Mice, Mice, Nude, Mutation, NADPH Oxidases metabolism, Poly (ADP-Ribose) Polymerase-1 genetics, Poly (ADP-Ribose) Polymerase-1 metabolism, Reactive Oxygen Species metabolism, Transplantation, Heterologous, rac1 GTP-Binding Protein genetics, rac1 GTP-Binding Protein metabolism, Antineoplastic Agents pharmacology, Lung Neoplasms drug therapy, Poly (ADP-Ribose) Polymerase-1 antagonists & inhibitors, Poly(ADP-ribose) Polymerase Inhibitors pharmacology, Protein Kinase Inhibitors pharmacology
- Abstract
Lung cancers with oncogenic mutations in the epidermal growth factor receptor (EGFR) invariably acquire resistance to tyrosine kinase inhibitor (TKI) treatment. Vulnerabilities of EGFR TKI-resistant cancer cells that could be therapeutically exploited are incompletely understood. Here, we describe a poly (ADP-ribose) polymerase 1 (PARP-1) inhibitor-sensitive phenotype that is conferred by TKI treatment in vitro and in vivo and appears independent of any particular TKI resistance mechanism. We find that PARP-1 protects cells against cytotoxic reactive oxygen species (ROS) produced by nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX). Compared to TKI-naive cells, TKI-resistant cells exhibit signs of increased RAC1 activity. PARP-1 catalytic function is required for PARylation of RAC1 at evolutionarily conserved sites in TKI-resistant cells, which restricts NOX-mediated ROS production. Our data identify a role of PARP-1 in controlling ROS levels upon EGFR TKI treatment, with potentially broad implications for therapeutic targeting of the mechanisms that govern the survival of oncogene-driven cancer cells., (Copyright © 2019 The Author(s). Published by Elsevier Inc. All rights reserved.)
- Published
- 2019
- Full Text
- View/download PDF
11. PARP-1 inhibition with or without ionizing radiation confers reactive oxygen species-mediated cytotoxicity preferentially to cancer cells with mutant TP53.
- Author
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Liu Q, Gheorghiu L, Drumm M, Clayman R, Eidelman A, Wszolek MF, Olumi A, Feldman A, Wang M, Marcar L, Citrin DE, Wu CL, Benes CH, Efstathiou JA, and Willers H
- Subjects
- Cell Cycle drug effects, Cell Cycle radiation effects, Cell Line, Tumor, Cell Proliferation drug effects, Cell Proliferation radiation effects, Cell Survival drug effects, Cell Survival radiation effects, DNA Breaks, Double-Stranded, Dose-Response Relationship, Drug, Humans, Mitochondria metabolism, Mutation, Urinary Bladder Neoplasms genetics, Urinary Bladder Neoplasms therapy, Phthalazines pharmacology, Piperazines pharmacology, Poly(ADP-ribose) Polymerase Inhibitors pharmacology, Radiation-Sensitizing Agents pharmacology, Reactive Oxygen Species metabolism, Tumor Suppressor Protein p53 genetics, Urinary Bladder Neoplasms metabolism
- Abstract
Biomarkers and mechanisms of poly (ADP-ribose) polymerase (PARP) inhibitor-mediated cytotoxicity in tumor cells lacking a BRCA-mutant or BRCA-like phenotype are poorly defined. We sought to explore the utility of PARP-1 inhibitor (PARPi) treatment with/without ionizing radiation in muscle-invasive bladder cancer (MIBC), which has poor therapeutic outcomes. We assessed the DNA damaging and cytotoxic effects of the PARPi olaparib in nine bladder cancer cell lines. Olaparib radiosensitized all cell lines with dose enhancement factors from 1.22 to 2.27. Radiosensitization was correlated with the induction of potentially lethal DNA double-strand breaks (DSB) but not with RAD51 foci formation. The ability of olaparib to radiosensitize MIBC cells was linked to the extent of cell kill achieved with the drug alone. Unexpectedly, increased levels of reactive oxygen species (ROS) resulting from PARPi treatment were the cause of DSB throughout the cell cycle in vitro and in vivo. ROS originated from mitochondria and were required for the radiosensitizing effects of olaparib. Consistent with the role of TP53 in ROS regulation, loss of p53 function enhanced radiosensitization by olaparib in non-isogenic and isogenic cell line models and was associated with increased PARP-1 expression in bladder cancer cell lines and tumors. Impairment of ATM in addition to p53 loss resulted in an even more pronounced radiosensitization. In conclusion, ROS suppression by PARP-1 in MIBC is a potential therapeutic target either for PARPi combined with radiation or drug alone treatment. The TP53 and ATM genes, commonly mutated in MIBC and other cancers, are candidate biomarkers of PARPi-mediated radiosensitization.
- Published
- 2018
- Full Text
- View/download PDF
12. Radiation Resistance in KRAS-Mutated Lung Cancer Is Enabled by Stem-like Properties Mediated by an Osteopontin-EGFR Pathway.
- Author
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Wang M, Han J, Marcar L, Black J, Liu Q, Li X, Nagulapalli K, Sequist LV, Mak RH, Benes CH, Hong TS, Gurtner K, Krause M, Baumann M, Kang JX, Whetstine JR, and Willers H
- Subjects
- A549 Cells, Animals, Carcinoma, Non-Small-Cell Lung metabolism, Carcinoma, Non-Small-Cell Lung pathology, Female, Heterografts, Humans, Lung Neoplasms metabolism, Lung Neoplasms pathology, Male, Mice, Mice, Nude, Mutation, Neoplastic Stem Cells metabolism, Neoplastic Stem Cells pathology, Neoplastic Stem Cells radiation effects, Osteopontin biosynthesis, Osteopontin genetics, Proto-Oncogene Proteins p21(ras) metabolism, Radiation Tolerance genetics, Signal Transduction, Carcinoma, Non-Small-Cell Lung genetics, Carcinoma, Non-Small-Cell Lung radiotherapy, ErbB Receptors metabolism, Lung Neoplasms genetics, Lung Neoplasms radiotherapy, Osteopontin metabolism, Proto-Oncogene Proteins p21(ras) genetics
- Abstract
Lung cancers with activating KRAS mutations are characterized by treatment resistance and poor prognosis. In particular, the basis for their resistance to radiation therapy is poorly understood. Here, we describe a radiation resistance phenotype conferred by a stem-like subpopulation characterized by mitosis-like condensed chromatin (MLCC), high CD133 expression, invasive potential, and tumor-initiating properties. Mechanistic investigations defined a pathway involving osteopontin and the EGFR in promoting this phenotype. Osteopontin/EGFR-dependent MLCC protected cells against radiation-induced DNA double-strand breaks and repressed putative negative regulators of stem-like properties, such as CRMP1 and BIM. The MLCC-positive phenotype defined a subset of KRAS-mutated lung cancers that were enriched for co-occurring genomic alterations in TP53 and CDKN2A. Our results illuminate the basis for the radiation resistance of KRAS-mutated lung cancers, with possible implications for prognostic and therapeutic strategies. Cancer Res; 77(8); 2018-28. ©2017 AACR ., (©2017 American Association for Cancer Research.)
- Published
- 2017
- Full Text
- View/download PDF
13. MAGE-A Cancer/Testis Antigens Inhibit MDM2 Ubiquitylation Function and Promote Increased Levels of MDM4.
- Author
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Marcar L, Ihrig B, Hourihan J, Bray SE, Quinlan PR, Jordan LB, Thompson AM, Hupp TR, and Meek DW
- Subjects
- Antigens, Neoplasm genetics, Cell Cycle Proteins, Cell Line, Tumor, Humans, Protein Binding, Ubiquitin metabolism, Ubiquitination, Antigens, Neoplasm metabolism, Nuclear Proteins metabolism, Proto-Oncogene Proteins metabolism, Proto-Oncogene Proteins c-mdm2 metabolism
- Abstract
Melanoma antigen A (MAGE-A) proteins comprise a structurally and biochemically similar sub-family of Cancer/Testis antigens that are expressed in many cancer types and are thought to contribute actively to malignancy. MAGE-A proteins are established regulators of certain cancer-associated transcription factors, including p53, and are activators of several RING finger-dependent ubiquitin E3 ligases. Here, we show that MAGE-A2 associates with MDM2, a ubiquitin E3 ligase that mediates ubiquitylation of more than 20 substrates including mainly p53, MDM2 itself, and MDM4, a potent p53 inhibitor and MDM2 partner that is structurally related to MDM2. We find that MAGE-A2 interacts with MDM2 via the N-terminal p53-binding pocket and the RING finger domain of MDM2 that is required for homo/hetero-dimerization and for E2 ligase interaction. Consistent with these data, we show that MAGE-A2 is a potent inhibitor of the E3 ubiquitin ligase activity of MDM2, yet it does not have any significant effect on p53 turnover mediated by MDM2. Strikingly, however, increased MAGE-A2 expression leads to reduced ubiquitylation and increased levels of MDM4. Similarly, silencing of endogenous MAGE-A expression diminishes MDM4 levels in a manner that can be rescued by the proteasomal inhibitor, bortezomid, and permits increased MDM2/MDM4 association. These data suggest that MAGE-A proteins can: (i) uncouple the ubiquitin ligase and degradation functions of MDM2; (ii) act as potent inhibitors of E3 ligase function; and (iii) regulate the turnover of MDM4. We also find an association between the presence of MAGE-A and increased MDM4 levels in primary breast cancer, suggesting that MAGE-A-dependent control of MDM4 levels has relevance to cancer clinically.
- Published
- 2015
- Full Text
- View/download PDF
14. MAGE-A antigens as targets in tumour therapy.
- Author
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Meek DW and Marcar L
- Subjects
- Animals, Humans, Melanoma-Specific Antigens chemistry, Melanoma-Specific Antigens genetics, Neoplasms genetics, Neoplasms immunology, Protein Conformation, Signal Transduction drug effects, Structure-Activity Relationship, Cancer Vaccines immunology, Immunotherapy methods, Melanoma-Specific Antigens immunology, Molecular Targeted Therapy, Neoplasms therapy
- Abstract
MAGE-A proteins constitute a sub-family of Cancer-Testis Antigens which are expressed mainly, but not exclusively, in germ cells. They are also expressed in various human cancers where they are associated with, and may drive, malignancy. MAGE-A proteins are highly immunogenic and are considered as potential targets for cancer vaccines and/or immuno-therapy. Moreover, recent advances in our understanding of their molecular pathology have revealed interactions that offer potential as therapeutic targets. Here we review recent progress in this area and consider how these interactions might be exploited, especially for the treatment of malignant cancers for which available treatments are inadequate., (Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.)
- Published
- 2012
- Full Text
- View/download PDF
15. Mage-A cancer/testis antigens inhibit p53 function by blocking its interaction with chromatin.
- Author
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Marcar L, Maclaine NJ, Hupp TR, and Meek DW
- Subjects
- Antigens, Neoplasm metabolism, Bone Neoplasms genetics, Cell Line, Tumor, Chromatin genetics, Chromatin Immunoprecipitation, Gene Knockdown Techniques, Humans, Lung Neoplasms genetics, Osteosarcoma genetics, Transcriptional Activation, Tumor Suppressor Protein p53 genetics, Tumor Suppressor Protein p53 metabolism, Antigens, Neoplasm immunology, Bone Neoplasms immunology, Chromatin metabolism, Lung Neoplasms immunology, Osteosarcoma immunology, Tumor Suppressor Protein p53 antagonists & inhibitors
- Abstract
The p53 tumor suppressor plays a major protective role in tumor prevention by coordinating changes in gene expression that lead to the elimination of cancer cells. Mage-A proteins comprise a family of metastasis-associated transcriptional regulators that potently inhibit p53 function. Here, we show that Mage-A interacts with 3 distinct peptides each of which is located within the DNA binding surface of the core domain of p53 and encompasses amino acids that are critical for site-specific DNA binding. These data suggest that Mage-A may block the association of p53 with its cognate sites in chromatin. Consistent with this idea, silencing of Mage-A expression leads to upregulation of several p53-responsive genes in a p53-dependent manner and stimulates by several fold the interaction of p53 with the p21, MDM2, and PUMA promoters. Notably, these effects can occur in the absence of genotoxic stress, leading in a p53-dependent manner, to cell-cycle delay and increased cell death. These data reveal a novel mechanism by which Mage-A proteins may suppress the p53 transcriptional program during tumor development and highlight the p53/Mage-A interaction as a prospective therapeutic target., (©2010 AACR.)
- Published
- 2010
- Full Text
- View/download PDF
16. p53-dependent repression of polo-like kinase-1 (PLK1).
- Author
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McKenzie L, King S, Marcar L, Nicol S, Dias SS, Schumm K, Robertson P, Bourdon JC, Perkins N, Fuller-Pace F, and Meek DW
- Subjects
- Animals, Apoptosis, Cell Cycle Proteins genetics, Cell Line, Tumor, Cyclin-Dependent Kinase Inhibitor p21 genetics, Cyclin-Dependent Kinase Inhibitor p21 metabolism, DNA Damage, Down-Regulation, Genes, Reporter, Histone Deacetylase Inhibitors pharmacology, Humans, Hydroxamic Acids pharmacology, Imidazoles metabolism, Piperazines metabolism, Promoter Regions, Genetic, Protein Serine-Threonine Kinases genetics, Proto-Oncogene Proteins genetics, RNA Interference, Transcription, Genetic, Tumor Suppressor Protein p53 genetics, Polo-Like Kinase 1, Cell Cycle physiology, Cell Cycle Proteins antagonists & inhibitors, Cell Cycle Proteins metabolism, Gene Expression Regulation drug effects, Protein Serine-Threonine Kinases antagonists & inhibitors, Protein Serine-Threonine Kinases metabolism, Proto-Oncogene Proteins antagonists & inhibitors, Proto-Oncogene Proteins metabolism, Tumor Suppressor Protein p53 metabolism
- Abstract
PLK1 is a critical mediator of G₂/M cell cycle transition that is inactivated and depleted as part of the DNA damage-induced G₂/M checkpoint. Here we show that downregulation of PLK1 expression occurs through a transcriptional repression mechanism and that p53 is both necessary and sufficient to mediate this effect. Repression of PLK1 by p53 occurs independently of p21 and of arrest at G₁/S where PLK1 levels are normally repressed in a cell cycle-dependent manner through a CDE/CHR element. Chromatin immunoprecipitation analysis indicates that p53 is present on the PLK1 promoter at two distinct sites termed p53RE1 and p53RE2. Recruitment of p53 to p53RE2, but not to p53RE1, is stimulated in response to DNA damage and/or p53 activation and is coincident with repression-associated changes in the chromatin. Downregulation of PLK1 expression by p53 is relieved by the histone deacetylase inhibitor, trichostatin A, and involves recruitment of histone deacetylase to the vicinity of p53RE2, further supporting a transcriptional repression mechanism. Additionally, wild type, but not mutant, p53 represses expression of the PLK1 promoter when fused upstream of a reporter gene. Silencing of PLK1 expression by RNAi interferes with cell cycle progression consistent with a role in the p53-mediated checkpoint. These data establish PLK1 as a direct transcriptional target of p53, independently of p21, that is required for efficient G₂/M arrest.
- Published
- 2010
- Full Text
- View/download PDF
17. FKBP25, a novel regulator of the p53 pathway, induces the degradation of MDM2 and activation of p53.
- Author
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Ochocka AM, Kampanis P, Nicol S, Allende-Vega N, Cox M, Marcar L, Milne D, Fuller-Pace F, and Meek D
- Subjects
- Animals, Cell Line, Tumor, Cyclin-Dependent Kinase Inhibitor p21 genetics, Cyclin-Dependent Kinase Inhibitor p21 metabolism, Escherichia coli genetics, Genes, Reporter, Glutathione Transferase metabolism, HCT116 Cells, Humans, Luciferases metabolism, Mice, Proteasome Endopeptidase Complex genetics, Proteasome Endopeptidase Complex metabolism, Proto-Oncogene Proteins c-mdm2 genetics, RNA, Small Interfering metabolism, Recombinant Fusion Proteins metabolism, Tacrolimus Binding Proteins genetics, Transfection, Tumor Suppressor Protein p53 genetics, Ubiquitin-Protein Ligases genetics, Ubiquitin-Protein Ligases metabolism, Ubiquitination, Gene Expression Regulation, Neoplastic, Proto-Oncogene Proteins c-mdm2 metabolism, Tacrolimus Binding Proteins metabolism, Tumor Suppressor Protein p53 metabolism
- Abstract
The p53 tumour suppressor protein is tightly controlled by the E3 ubiquitin ligase, mouse double minute 2 (MDM2), but maintains MDM2 expression as part of a negative feedback loop. We have identified the immunophilin, 25kDa FK506-binding protein (FKBP25), previously shown to be regulated by p53-mediated repression, as an MDM2-interacting partner. We show that FKBP25 stimulates auto-ubiquitylation and proteasomal degradation of MDM2, leading to the induction of p53. Depletion of FKBP25 by siRNA leads to increased levels of MDM2 and a corresponding reduction in p53 and p21 levels. These data are consistent with the idea that FKBP25 contributes to regulation of the p53-MDM2 negative feedback loop.
- Published
- 2009
- Full Text
- View/download PDF
18. Elevated levels of oncogenic protein kinase Pim-1 induce the p53 pathway in cultured cells and correlate with increased Mdm2 in mantle cell lymphoma.
- Author
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Hogan C, Hutchison C, Marcar L, Milne D, Saville M, Goodlad J, Kernohan N, and Meek D
- Subjects
- Animals, Antigens, CD20 biosynthesis, Cell Line, Tumor, Cells, Cultured, Genes, p53, Humans, Mice, Phosphorylation, Transcriptional Activation, Transfection, Gene Expression Regulation, Neoplastic, Lymphoma, Mantle-Cell metabolism, Proto-Oncogene Proteins c-mdm2 metabolism, Proto-Oncogene Proteins c-pim-1 metabolism, Tumor Suppressor Protein p53 metabolism
- Abstract
Mutation of the p53 gene is a common event during tumor pathogenesis. Other mechanisms, such as mdm2 amplification, provide alternative routes through which dysfunction of the p53 pathway is promoted. Here, we address the hypothesis that elevated expression of pim oncogenes might suppress p53 by regulating Mdm2. At a physiological level, we show that endogenous Pim-1 and Pim-2 interact with endogenous Mdm2. Additionally, the Pim kinases phosphorylate Mdm2 in vitro and in cultured cells at Ser(166) and Ser(186), two previously identified targets of other signaling pathways, including Akt. Surprisingly, at high levels of Pim expression, as would occur in tumors, active, but not inactive, Pim-1 or Pim-2 blocks the degradation of both p53 and Mdm2 in a manner that is independent of Mdm2 phosphorylation, leading to increased p53 levels and, proportionately, p53-dependent transactivation. Additionally, Pim-1 induces endogenous ARF, p53, Mdm2, and p21 in primary murine embryo fibroblasts and stimulates senescence-associated beta-galactosidase levels, consistent with the induction of senescence. Immunohistochemical analysis of a cohort of 33 human mantle cell lymphomas shows that elevated expression of Pim-1 occurs in 42% of cases, with elevated Pim-2 occurring in 9% of cases, all of which also express Pim-1. Notably, elevated Pim-1 correlates with elevated Mdm2 in MCL with a p value of 0.003. Taken together, our data are consistent with the idea that Pim normally interacts with the p53 pathway but, when expressed at pathological levels, behaves as a classic dominant oncogene that stimulates a protective response through induction of the p53 pathway.
- Published
- 2008
- Full Text
- View/download PDF
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