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Biomedical subjects

Ygal Haupt

Publications and source records attributed to Ygal Haupt.

18 recordsLinked to original sources
PubMed2006

Celecoxib can induce cell death independently of cyclooxygenase-2, p53, Mdm2, c-Abl and reactive oxygen species.

Cell lines that do not overexpress functional cyclooxygenase-2 are resistant to the normal plasma levels of celecoxib achieved following oral ingestion. Cell growth inhibition was demonstrated after 24 h exposure to 80 micromol/l celecoxib while significant death was not detected at concentrations below 120 micromol/l following 24 h exposure. This growth inhibition and death induction was identified to be independent of p53 and Hdm2 in these cells, despite wild-type p53 stabilization and Hdm2 diminution in some lines. Cell death induced by celecoxib was preceded by the generation of reactive oxygen species within 4 h of drug exposure. The precise mechanism of elicitation of reactive oxygen species in these cells remains to be elucidated, although it was found to be independent of p53 and c-Abl, while in vitro, celecoxib enhanced superoxide radical production by xanthine oxidase. Importantly, the failure of anti-oxidants to protect from death indicates that celecoxib induces death independently of reactive oxygen species and that reactive oxygen species generation may be an insufficient trigger of death in p53-deficient cells.

Adenocarcinoma↗
PubMed2006

Importance of p53 for cancer onset and therapy.

Cancer predisposition, onset and therapeutic response can be critically determined by the integrity of the tumor suppressor p53. The majority of human cancers appear to exhibit either abnormal p53 or disrupted p53 activation pathways. Intervention to restore wild-type p53 activities is an attractive approach for cancer therapy. The manipulation of p53 and its targets is a challenging field that is still in its infancy, but witnessing some notable developments in the areas of p53 gene therapy, mutant reactivation and suppression of the negative p53 regulator Mdm2 using small molecules. In addition, wild-type p53 manipulation in healthy tissues of cancer patients in the context of chemotherapy and radiation therapies is offering the potential of enhanced patient recovery.

Animals↗
PubMed2005

C-Abl as a modulator of p53.

P53 is renowned as a cellular tumor suppressor poised to instigate remedial responses to various stress insults that threaten DNA integrity. P53 levels and activities are kept under tight regulation involving a complex network of activators and inhibitors, which determine the type and extent of p53 growth inhibitory signaling. Within this complexity, the p53-Mdm2 negative auto-regulatory loop serves as a major route through which intra- and extra-cellular stress signals are channeled to appropriate p53 responses. Mdm2 inhibits p53 transcriptional activities and through its E3 ligase activity promotes p53 proteasomal degradation either within the nucleus or following nuclear export. Upon exposure to stress signals these actions of Mdm2 have to be moderated, or even interrupted, in order to allow sufficient p53 to accumulate in an active form. Multiple mechanisms involving a variety of factors have been demonstrated to mediate this interruption. C-Abl is a critical factor that under physiological conditions is required for the maximal and efficient accumulation of active p53 in response to DNA damage. C-Abl protects p53 by antagonizing the inhibitory effect of Mdm2, an action that requires a direct interplay between c-Abl and Mdm2. In addition, c-Abl protects p53 from other inhibitors of p53, such as the HPV-E6/E6AP complex, that inhibits and degrades p53 in HPV-infected cells. Surprisingly, the oncogenic form of c-Abl, the Bcr-Abl fusion protein in CML cells, also promotes the accumulation of wt p53. However, in contrast to the activation of p53 by c-Abl, its oncogenic form, Bcr-Abl, counteracts the growth inhibitory activities of p53 by modulating the p53-Mdm2 loop. Thus, it appears that by modulating the p53-Mdm2 loop, c-Abl and its oncogenic forms critically determine the type and extent of the cellular response to DNA damage.

Animals↗
PubMed2005

Mdm2 in growth signaling and cancer.

Genetic and biochemical evidence have demonstrated a direct link between Mdm2 and cancer development. Elevated expression of Mdm2 is observed in a significant proportion of different types of cancer. The major contribution of Mdm2 to the development of cancer is through a tight inhibition of the activities and stability of the tumor suppressor p53. However, extensive studies over the past few years have identified p53-independent functions of Mdm2, in the regulation of several important cellular processes and multiple signaling pathways. The promotion of cell cycle progression by Mdm2 is mediated via p53 inhibition, and by regulating the pRb/E2F complex. Mdm2 is an important mediator of growth and survival signaling in the PI3K/Akt pathway, an activator of certain steroid hormone receptors, and an inhibitor of the TGF-beta growth restrictive pathway. Thus, the impact on these pathways by deregulated Mdm2, as often observed in cancer, can be oncogenic in a permissible environment. This renders Mdm2 as an important target for the development of anti-cancer drugs.

Animals↗
PubMed2005

Mutations in proline 82 of p53 impair its activation by Pin1 and Chk2 in response to DNA damage.

Tumor suppression by the p53 protein largely depends on the elimination of damaged cells by apoptosis. Mutations in the polyproline region (PPR) of p53 impair its apoptotic function. Deletion of the PPR renders p53 more sensitive to inhibition by Mdm2 via an unknown mechanism. We have explored the mechanism by which the PPR modulates the p53/Mdm2 loop. Proline 82 of p53 was identified to be essential for its interaction with the checkpoint kinase 2 (Chk2) and consequent phosphorylation of p53 on serine 20, following DNA damage. These physical and functional interactions are regulated by Pin1 through cis-trans isomerization of proline 82. Our study unravels the pathway by which Pin1 activates p53 in response to DNA damage and explains how Pin1 protects p53 from Mdm2. Further, we propose a role for Pin1-dependent induction of p53 conformational change as a mechanism responsible for the enhanced interaction between p53 and Chk2 following DNA damage. Importantly, our findings elucidate the selection for mutations in the Pin1 target Thr81/Pro82 motif within the PPR of p53 in human cancer.

Animals↗
PubMed2004

Treatment of chronic myeloid leukemia cells with imatinib (STI571) impairs p53 accumulation in response to DNA damage.

Chronic myelogenous leukaemia (CML) is induced by the Bcr-Abl fusion protein. Inhibition of Bcr-Abl by STI571 is widely used to treat CML patients. Unlike in most cancer types, the frequency of p53 mutations in CML is low. Here, we investigated the effect of STI571 treatment of CML cells on p53 regulation. Exposure of CML cells, including established cell lines and freshly isolated cells from patients, to STI571 reduced p53 protein levels, and severely impaired its accumulation in response to DNA damage. This may be explained by the status of p53 serine 20 phosphorylation. In non-stressed CML cells, serine 20 of p53 is constitutively phosphorylated by Chk1, and is inhibited by STI571. In response to DNA damage, however, this phosphorylation is mediated by Chk1 and Chk2, and is only partially inhibited by STI571. CML cells expressing wild-type p53 are more resistant to treatment with STI571, but moderately more sensitive to DNA damage, than CML cells lacking p53. An enhanced induction of apoptosis by STI571 and DNA damage is observed in CML cells bearing wild-type p53, but not in cells lacking functional p53. This implies that the status of p53 may affect the response of CML cells to this combined treatment.

Antineoplastic Agents↗
PubMed2004

p53 Regulation: a family affair.

The p53 protein averts tumor formation by preventing the proliferation of damaged cells. The presence of functional p53 is critical for efficient and proper cellular responses to a variety of stress conditions. Interestingly, p63 and p73, which are the homologous ancestors of p53, retain a broader set of activities than their progeny, particularly during early embryonic development. The link of these homologues to cancer and their effect on p53 tumor suppression is only beginning to be unravelled. The tight regulation of p53 is governed by the Mdm2 E3 ligase, but also by at least two other E3 ligases. Recent findings suggest fine-tuning of p53 regulation through changes in the ratio of p53 and Mdm2. This regulation of p53 is modulated by the Mdm2 homologue, Mdmx. Genetic studies reveal the critical role Mdmx plays in p53 regulation, although the mode of action is yet to be fully explored. The relief of p53 from this tight regulation is imperative in order for it to respond to stress signals. An intriguing player in this process is the prolyl isomerase Pin1, which induces a conformational change in p53, and more recently identified, also in p73, in response to DNA damage. This complex network of regulation emerges as a family affair. This wealth of knowledge has been translated into the development of novel anti-cancer strategies based on the p53 status in the cancer cell.

Animals↗
PubMed2004

Improving cancer therapy through p53 management.

The tumor suppressor p53 normally acts to appropriately coordinate cellular responses to stress stimuli. When p53 activity is disabled, the onset of malignancy is a potential consequence. Engendering wild type p53 activities in cells that lack these functions is an approach that is currently being explored for cancer therapy. Eliciting elevated levels of active p53, imparting p53 activities through gene therapy, compelling mutant p53 to perform normal functions, manipulating p53 regulators, and activating p53 effectors are all approaches that are currently being developed. In this review we will provide a synopsis of the most promising 'p53-based' strategies for fighting cancer, both those under clinical trial and recent innovative concepts.

Animals↗
PubMed2004

Manipulation of the tumor suppressor p53 for potentiating cancer therapy.

The tumor suppressor p53 is a linchpin in the regulation of appropriate cellular responses to various stress conditions. Inactivation of the functions of this critical participant can have diabolical consequences, in particular the development of malignant diseases. Elicitation of appropriate p53 functions is an attractive strategy for combating cancer. Triggering p53 responses, reconstituting p53 activities through gene therapy, coercing mutant p53 to perform normal functions, manipulating p53 regulators, and activating p53 effectors are all approaches that are currently being developed. Here, we will overview 'p53-based' strategies for fighting cancer, both those under clinical trial and recent innovative concepts.

Animals↗
PubMed2003

Apoptosis - the p53 network.

Exposure to cellular stress can trigger the p53 tumor suppressor, a sequence-specific transcription factor, to induce cell growth arrest or apoptosis. The choice between these cellular responses is influenced by many factors, including the type of cell and stress, and the action of p53 co-activators. p53 stimulates a wide network of signals that act through two major apoptotic pathways. The extrinsic, death receptor pathway triggers the activation of a caspase cascade, and the intrinsic, mitochondrial pathway shifts the balance in the Bcl-2 family towards the pro-apoptotic members, promoting the formation of the apoptosome, and consequently caspase-mediated apoptosis. The impact of these two apoptotic pathways may be enhanced when they converge through Bid, which is a p53 target. The majority of these apoptotic effects are mediated through the induction of specific apoptotic target genes. However, p53 can also promote apoptosis by a transcription-independent mechanism under certain conditions. Thus, a multitude of mechanisms are employed by p53 to ensure efficient induction of apoptosis in a stage-, tissue- and stress-signal-specific manner. Manipulation of the apoptotic functions of p53 constitutes an attractive target for cancer therapy.

Animals↗
PubMed2003

The promyelocytic leukemia protein protects p53 from Mdm2-mediated inhibition and degradation.

The p53 protein is kept labile under normal conditions. This regulation is governed largely by its major negative regulator, Mdm2. In response to stress however, p53 accumulates and becomes activated. For this to occur, the inhibitory effects of Mdm2 have to be neutralized. Here we investigated the role of the promyelocytic leukemia protein (PML) in the activation of p53 in response to stress. We found that PML is critical for the accumulation of p53 in response to DNA damage under physiological conditions. PML protects p53 from Mdm2-mediated ubiquitination and degradation, and from inhibition of apoptosis. PML neutralizes the inhibitory effects of Mdm2 by prolonging the stress-induced phosphorylation of p53 on serine 20, a site of the checkpoint kinase 2 (Chk2). PML recruits Chk2 and p53 into the PML nuclear bodies and enhances p53/Chk2 interaction. Our results provide a novel mechanistic explanation for the cooperation between PML and p53 in response to DNA damage.

Active Transport, Cell Nucleus↗
PubMed2003

P53 licensed to kill? Operating the assassin.

The p53 protein is a key player in the cellular response to stress. Proper regulation of p53 is imperative for the suppression of tumor development. This regulation is largely governed by its master inhibitor, Mdm2, which both blocks p53 activities and promotes its destabilization. This tight regulation of p53 by Mdm2 must be interrupted under stress conditions in order for p53 to be stabilized in an active form. A combined action of partner proteins and modifying enzymes is essential for the relief of p53 from Mdm2. The recent revelation of p53 association with the PML-nuclear bodies provides one explanation of how this regulatory network is coordinated within the nucleus in response to certain stress conditions. Thus, it is not only the nature of the p53 regulatory complex but also the spatial and temporal context of this association that governs the output inhibitory signals mediated by p53.

ADP-Ribosylation Factors↗
PubMed2003

Flow cytometric analysis of p53-induced apoptosis.

The p53 protein is a key player in the cellular response to stress conditions. Activation of p53 induces growth inhibition in the form of cell growth arrest or apoptosis. The latter plays an important role in the tumor suppression function of p53. It is therefore of great interest to understand in detail the mechanisms by which p53 induces apoptosis. In this chapter, we describe a flow cytometric assay for the measurement of p53 apoptotic activity. This assay is applicable for exogenously expressed p53 by transfection as well as for endogenous p53. The p53 protein is detected by intra-cellular fluorescent staining of p53 or by tagging p53 with GFP. The extent of apoptosis in cells expressing p53 is determined by cell cycle distribution using flow cytometry. The protocol described here can be employed to study the regulation of p53-mediated apoptosis and can be broadly applied to other apoptotic proteins.

Animals↗
PubMed2002

Deconstruction of p53 functions and regulation.

The 11th international p53 workshop was held in Barcelona, Spain, May 15-18, 2002. The p53 workshop was organized by Carlos Cordón-Cardo (Memorial Sloan Kettering, New York). The workshop included 46 oral presentations and 206 posters.

Acetylation↗
PubMed2002

Tyrosine phosphorylation of Mdm2 by c-Abl: implications for p53 regulation.

The p53 tumor suppressor is inhibited and destabilized by Mdm2. However, under stress conditions, this downregulation is relieved, allowing the accumulation of biologically active p53. Recently we showed that c-Abl is important for p53 activation under stress conditions. In response to DNA damage, c-Abl protects p53 by neutralizing the inhibitory effects of Mdm2. In this study we ask whether this neutralization involves a direct interplay between c-Abl and Mdm2, and what is the contribution of the c-Abl kinase activity? We demonstrate that the kinase activity of c-Abl is required for maintaining the basal levels of p53 expression and for achieving maximal accumulation of p53 in response to DNA damage. Importantly, c-Abl binds and phosphorylates Mdm2 in vivo and in vitro. We characterize Hdm2 (human Mdm2) phosphorylation at Tyr394. Substitution of Tyr394 by Phe394 enhances the ability of Mdm2 to promote p53 degradation and to inhibit its transcriptional and apoptotic activities. Our results suggest that phosphorylation of Mdm2 by c-Abl impairs the inhibition of p53 by Mdm2, hence defining a novel mechanism by which c-Abl activates p53.

Amino Acid Substitution↗
PubMed2002

Facilitation of adenoviral wild-type p53-induced apoptotic cell death by overexpression of p33(ING1) in T.Tn human esophageal carcinoma cells.

To investigate the effect of p33(ING1) on wild-type p53 gene therapy, T.Tn human esophageal carcinoma cells were stably transfected with p33(ING1) cDNA. Infection with Ad-p53 (recombinant adenovirus containing wild-type p53) into p33-transfected cells reduced cell viability, while infection with empty vector had little effect. This reduced viability was shown to be due to apoptotic cell death by the TUNEL (terminal deoxynucleotidyl transferase-mediated nick end-labeling) assay. Following infection with Ad-p53, levels of p53 were similar in p33-expressing cells and in the parental line. However, levels of p21 and Mdm2 were elevated in p33-transfected cells. Nonetheless, this enhanced expression of Mdm2 appeared to be ineffective in downregulating p53. Transient transfection with mutant Mdm2 prior to Ad-p53 infection provided a significant protection as compared with cells transfected with wild-type Mdm2. These results imply a synergistic effect between p33 and p53 in the induction of apoptosis of human esophageal carcinoma cells. A role for Mdm2 in this synergism is suggested.

Adenoviridae↗