Role Of P53 In Response Research Articles

Polyphenols as Antitumor Agents Targeting Key Players in Cancer-Driving Signaling Pathways.

Polyphenols constitute an important group of natural products that are traditionally associated with a wide range of bioactivities. These are usually found in low concentrations in natural products and are now available in nutraceuticals or dietary supplements. A group of polyphenols that include apigenin, quercetin, curcumin, resveratrol, EGCG, and kaempferol have been shown to regulate signaling pathways that are central for cancer development, progression, and metastasis. Here, we describe novel mechanistic insights on the effect of this group of polyphenols on key elements of the signaling pathways impacting cancer. We describe the protein modifications induced by these polyphenols and their effect on the central elements of several signaling pathways including PI3K, Akt, mTOR, RAS, and MAPK and particularly those affecting the tumor suppressor p53 protein. Modifications of p53 induced by these polyphenols regulate p53 gene expression and protein levels and posttranslational modifications such as phosphorylation, acetylation, and ubiquitination that influence stability, subcellular location, activation of new transcriptional targets, and the role of p53 in response to DNA damage, apoptosis control, cell- cycle regulation, senescence, and cell fate. Thus, deep understanding of the effects that polyphenols have on these key players in cancer-driving signaling pathways will certainly lead to better designed targeted therapies, with less toxicity for cancer treatment. The scope of this review centers on the regulation of key elements of cancer signaling pathways by the most studied polyphenols and highlights the importance of a profound understanding of these regulations in order to improve cancer treatment and control with natural products.

Contribution of genotoxic anticancer treatments to the development of multiple primary tumours in the context of germline TP53 mutations

IntroductionLi-Fraumeni syndrome (LFS), due to TP53 germline mutations, is characterised by a remarkably high incidence of multiple primary cancers (MPCs), and the key role of p53 in response to DNA damage questions the contribution of anticancer treatments to MPCs development. Materials and methodsWe first evaluated genotoxicity of X-rays and different classes of conventional chemotherapies, thanks to genotoxicity assays, based on the measurement of transcriptional response to DNA damage and performed in murine splenocytes, either exposed ex vivo or extracted from exposed mice. We then exposed a total of 208 Trp53Δ/Δ, wt/Δ or wt/wt mice to clinical doses of X-rays or genotoxic or non-genotoxic chemotherapies. Tumour development was monitored using whole-body magnetic resonance imaging and pathological examination at death. Results:X-rays and conventional chemotherapies, except mitotic spindle poisons, were found to be genotoxic in both p53 genotoxicity assays. Exposition to X-rays and the topoisomerase inhibitor etoposide, analysed as genotoxic anticancer treatment, drastically increase the tumour development risk in Trp53Δ/Δ and wt/Δ mice (hazard ration [HR] = 4.4, 95% confidence interval [CI] [2.2–8.8], p < 0.001*** and HR = 4.7, 95% CI [2.4–9.3], p < 0.001***, respectively). In contrast, exposure to the non-genotoxic mitotic spindle poison, docetaxel, had no impact on tumour development. ConclusionsThis study shows that radiotherapy and genotoxic chemotherapies significantly increase the risk of tumour development in a LFS mice model. These results strongly support the contribution of genotoxic anticancer treatments to MPC development in LFS patients. Therefore, to reduce the risk of MPCs in germline TP53 mutation carriers, radiotherapy should be avoided whenever possible, surgical treatment prioritised, and non-genotoxic treatments considered.

The role of p38 MAPK pathway in p53 compromised state and telomere mediated DNA damage response

There is an intricate balance of DNA damage response and repair which determines the homeostasis of human genome function. p53 protein is widely known for its role in cell cycle regulation and tumor suppressor activity. In case of several cancers where function of p53 gene gets compromised either by mutation or partial inactivation, the role of p53 in response to DNA damage needs to be supplemented by another molecule or pathway. Due to sedentary lifestyle and exposure to genotoxic agents, genome is predisposed to chronic stress, which ultimately leads to unrepaired or background DNA damage. p38 MAPK signaling pathway is strongly activated in response to various environmental and cellular stresses. DNA damage response and the repair options have crucial links with chromosomal integrity. Telomere that regulates integrity of genome is protected by a six member shielding unit called shelterin complex which communicates with other pathways for functionality of telomeres. There are evidences that p38 gets activated through ATM in response to DNA damage. Dysfunctional telomere leads to activation of ATM which subsequently activates p38 suggesting a crosstalk between p38, ATM and shelterin complex. This review focuses on activation of p38 in response to genotoxic stress induced DNA damage in p53 mutated or compromised state and its possible cross talk with telomere shelterin proteins. Thus p38 may act as an important target to treat various diseases and in majority of cancers in p53 mutated state.

The human immune response to RSV includes a loop between the tumor suppressor p53 and expression of APOBEC3 genes

BackgroundRespiratory syncytial virus (RSV) infects nearly all children by theirsecond year of life with re‐infections occurring throughout adolescence andadulthood. RSV infection typicallyresults in mild host symptoms. However, susceptible individuals can develop severe lower respiratory tract infectionsand bronchiolitis, sometimes leading to hospitalization or death. Recently we established that in addition toits protective role against cancer development, the tumor suppressor p53 alsocontributes to host immune responses regulating the expression of several genesincluding the APOBEC3 family of DNA cytidine deaminases (A3), consideredsentinels in innate immune response against viral infections.ObjectiveUsinghuman lung cancer cells, we tested the role of p53 in response to RSV infectionand its replication through the regulation of the A3 gene family as well asother components of the immune response.MethodsA549 and H1299 cells were infected with RSV (Strain A2). Cells were thenharvested 72–120 hours post‐infection and qPCR and Nanostring arrays? were usedto evaluate mRNA expression of RSV‐g, P53, A3s and other immune targets. Expression of RSV was assessed? by ConfocalMicroscopy and cytofluorometry while Annexin V/PI assay was used to evaluateapoptosis.ResultsIn response to acute and chronic RSV infection in human lung cancer cells, p53 was activated and had an impact on RSV replication, which involves theexpression in a p53 dependent manner of several members of the A3 gene familyas well as other innate immune genes. APOBEC3G (A3G) was highly induced in a dose‐ and time‐dependent mannerafter RSV only in lung cancer cells harboring WT p53. Accordingly, overexpression of A3G in lungcancer cells reduced RSV replication. Furthermore, a Nanotstring transcriptome analysis (immune code set) after RSV infectionrevealed a strong p53 dependent regulation of immune response genes, includingseveral components of the innate immune antiviral response. In addition, RSV infection induced p53phosphorylation at serine 15 and triggered p53‐dependent apoptosis in p53 functional cells while p53 deficiency correlated with increased viralreplication and increased syncytia phenotype.ConclusionsOverall, our findings establish that the “guardian of the genome” role ascribed to p53 also extends to a unique component of the immune system – the A3 genes – revealing novel components in the antiviralimmune response against RSV infection.Support or Funding InformationSupported by NIEHS Division of Intramural Research Program.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

The Combination of Metformin and Valproic Acid Induces Synergistic Apoptosis in the Presence of p53 and Androgen Signaling in Prostate Cancer.

We investigated the potential of combining the hypoglycemic drug metformin (MET) and the antiepileptic drug valproic acid (VPA), which act via different biochemical pathways, to provide enhanced antitumor responses in prostate cancer. Prostate cancer cell lines (LNCaP and PC-3), normal prostate epithelial cells (PrEC), and patient-derived prostate tumor explants were treated with MET and/or VPA. Proliferation and apoptosis were assessed. The role of p53 in response to MET + VPA was assessed in cell lines using RNAi in LNCaP (p53+) and ectopic expression of p53 in PC-3 (p53-). The role of the androgen receptor (AR) was investigated using the AR antagonist enzalutamide. The combination of MET and VPA synergistically inhibited proliferation in LNCaP and PC-3, with no significant effect in PrEC. LNCaP, but not PC-3, demonstrated synergistic intrinsic apoptosis in response to MET + VPA. Knockdown of p53 in LNCaP (p53+, AR+) reduced the synergistic apoptotic response as did inhibition of AR. Ectopic expression of p53 in PC-3 (p53-, AR-) increased apoptosis in response to MET + VPA. In patient-derived prostate tumor explants, MET + VPA also induced a significant decrease in proliferation and an increase in apoptosis in tumor cells. In conclusion, we demonstrate that MET + VPA can synergistically kill more prostate cancer cells than either drug alone. The response is dependent on the presence of p53 and AR signaling, which have critical roles in prostate carcinogenesis. Further in vivo/ex vivo preclinical studies are required to determine the relative efficacy of MET + VPA as a potential treatment for prostate cancer. Mol Cancer Ther; 16(12); 2689-700. ©2017 AACR.

P53 activation contributes to patulin-induced nephrotoxicity via modulation of reactive oxygen species generation.

Patulin is a major mycotoxin found in fungal contaminated fruits and their derivative products. Previous studies showed that patulin was able to induce increase of reactive oxygen species (ROS) generation and oxidative stress was suggested to play a pivotal role in patulin-induced multiple toxic signaling. The objective of the present study was to investigate the functional role of p53 in patulin-induced oxidative stress. Our study demonstrated that higher levels of ROS generation and DNA damage were induced in wild-type p53 cell lines than that found in either knockdown or knockout p53 cell lines in response to patulin exposure, suggesting p53 activation contributed to patulin-induced ROS generation. Mechanistically, we revealed that the pro-oxidant role of p53 in response to patulin was attributed to its ability to suppress catalase activity through up-regulation of PIG3. Moreover, these in vitro findings were further validated in the p53 wild-type/knockout mouse model. To the best of our knowledge, this is the first report addressing the functional role of p53 in patulin-induced oxidative stress. The findings of the present study provided novel insights into understanding mechanisms behind oxidative stress in response to patulin exposure.

P53 family members - important messengers in cell death signaling in photodynamic therapy of cancer?

TP53 is one of the genes most frequently inactivated in cancers. Mutations in TP53 gene are linked to worse prognosis and shorter overall survival of cancer patients. TP53 encodes a critical tumor suppressor, which dictates cell fate decisions upon stress stimuli. As a sensor of cellular stress, p53 is a relevant messenger of cell death signaling in ROS-driven photodynamic therapy (PDT) of cancer. The significant role of p53 in response to PDT has been reported for several clinically approved photosensitizers. Multiple reports described that wild-type p53 contributes to cell killing upon photodynamic therapy with clinically approved photosensitizers but the mechanism is still not fully understood. This work outlines the diverse functions of p53 family members in cancer cells' susceptibility and resistance to PDT. In summary p53 and p53 family members are emerging as important mediators of cell death signaling in photodynamic therapy of cancer, however the mechanism of cell death provoked during PDT might differ depending on the tissue type and the photosensitizer applied.

Abstract 3507: APR-246 reactivates mutant p53 in non-small cell lung cancer cell lines and sensitizes cells for CDDP treatment under normoxic and hypoxic conditions

Abstract Introduction: Tumors are often characterized by inactivating mutations in the TP53 gene. Mutations in this gene occur in about 50% of non-small cell lung cancers (NSCLC), and influence the response to common chemotherapeutics like CDDP (cisplatin). In addition, as hypoxic stress, which is often found in tumors, is able to induce p53 accumulation and p53 dependent apoptosis, the reactivation of mutant p53 might induce a strong cytotoxic effect in hypoxic tumor regions. Therefore, we treated NSCLC tumor cells with APR-246, a known activator of mutant p53, under normoxic and hypoxic conditions. In addition, we combined APR-246 with CDDP, to determine whether reactivation of mutant p53 could enhance the cytotoxic response of CDDP. Materials &amp; Methods: The NSCLC cell line CRL-5908, harboring a R273H p53 mutation, was used. To determine the role of mutant p53 in response to APR-246, CRL-5908 was transduced with a p53 shRNA lentiviral vector to obtain a p53 deficient sub cell line. The SRB-assay was used to determine the cytotoxic effect of both mono-and combination therapies after which possible synergism was calculated by the Chou-Talalay method, both under normoxic and hypoxic conditions (0% O2, 5% CO2, 95% N2). In addition, we determined the p53 protein level as well as the mRNA and protein levels of its main transcription targets MDM2, p21, PUMA and BAX. The induction of apoptosis and cell cycle arrest were determined by flowcytometric analysis. Results: Our results show a clear response of CRL-5908 to APR-246, which was stronger under hypoxic conditions compared to normoxic conditions (IC50: 9.5 ± 2.5 vs. 16.3 ± 3.4). We observed that CRL-5908 was less sensitive to CDDP monotherapy than the p53 wild type cell line A549 (IC50: 15.2 ± 3.3 vs. 6.2 ± 1.6). We were able to overcome this resistance to CDDP by combining the drug with APR-246 (IC20, IC40, IC60), which resulted in CDDP IC50 values of respectively 11.5 ± 4.0; 5.0 ± 2.2 and 6.5 ± 2.0 under normoxic conditions. Discussion: Our initial results show that APR-246 monotherapy is promising for the treatment of p53 mutant NSCLC, particularly under hypoxic conditions. In addition, APR-246 seems to sensitize cells for CDDP treatment when combined simultaneously. We will further study the underlying mechanisms, like the activation of p53 and the transcription of its main transcription target MDM2, p21, PUMA and BAX as well as apoptosis and cell cycle arrest. Moreover, we will compare simultaneous treatment of CDDP and APR-246 with sequential treatment in which CDDP is followed by APR-246 treatment, and further sort out the response of this combination therapy under hypoxic conditions, as well as the role of mutant p53. Citation Format: Christophe Deben, Vanessa Deschoolmeester, An Wouters, Jolien Van den Bossche, Julie Jacobs, Filip Lardon, Christian Rolfo, Patrick Pauwels. APR-246 reactivates mutant p53 in non-small cell lung cancer cell lines and sensitizes cells for CDDP treatment under normoxic and hypoxic conditions. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3507. doi:10.1158/1538-7445.AM2015-3507

PRAP1 is a novel executor of p53-dependent mechanisms in cell survival after DNA damage

p53 has a crucial role in governing cellular mechanisms in response to a broad range of genotoxic stresses. During DNA damage, p53 can either promote cell survival by activating senescence or cell-cycle arrest and DNA repair to maintain genomic integrity for cell survival or direct cells to undergo apoptosis to eliminate extensively damaged cells. The ability of p53 to execute these two opposing cell fates depends on distinct signaling pathways downstream of p53. In this study, we showed that under DNA damage conditions induced by chemotherapeutic drugs, gamma irradiation and hydrogen peroxide, p53 upregulates a novel protein, proline-rich acidic protein 1 (PRAP1). We identified functional p53-response elements within intron 1 of PRAP1 gene and showed that these regions interact directly with p53 using ChIP assays, indicating that PRAP1 is a novel p53 target gene. The induction of PRAP1 expression by p53 may promote resistance of cancer cells to chemotherapeutic drugs such as 5-fluorouracil (5-FU), as knockdown of PRAP1 increases apoptosis in cancer cells after 5-FU treatment. PRAP1 appears to protect cells from apoptosis by inducing cell-cycle arrest, suggesting that the induction of PRAP1 expression by p53 in response to DNA-damaging agents contributes to cancer cell survival. Our findings provide a greater insight into the mechanisms underlying the pro-survival role of p53 in response to cytotoxic treatments.

P53 Stabilization Induces Cell Growth Inhibition and Affects IGF2 Pathway in Response to Radiotherapy in Adrenocortical Cancer Cells

Adrenocortical carcinoma (ACC) is a very rare endocrine tumour, with variable prognosis, depending on tumour stage and time of diagnosis. However, it is generally fatal, with an overall survival of 5 years from detection. Radiotherapy usefulness for ACC treatment has been widely debated and seems to be dependent on molecular alterations, which in turn lead to increased radio-resistance. Many studies have shown that p53 loss is an important risk factor for malignant adrenocortical tumour onset and it has been reported that somatic mutations in TP53 gene occur in 27 to 70% of adult sporadic ACCs. In this study, we investigated the role of somatic mutations of the TP53 gene in response to ionizing radiation (IR). We studied the status of p53 in two adrenocortical cell lines, H295R and SW-13, harbouring non-functioning forms of this protein, owing to the lack of exons 8 and 9 and a point mutation in exon 6, respectively. Moreover, these cell lines show high levels of p-Akt and IGF2, especially H295R. We noticed that restoration of p53 activity led to inhibition of growth after transient transfection of cells with wild type p53. Evaluation of their response to IR in terms of cell proliferation and viability was determined by means of cell count and TUNEL assay.wtp53 over-expression also increased cell death by apoptosis following radiation in both cell lines. Moreover, RT-PCR and Western blotting analysis of some p53 target genes, such as BCL2, IGF2 and Akt demonstrated that p53 activation following IR led to a decrease in IGF2 expression. This was associated with a reduction in the active form of Akt. Taken together, these results highlight the role of p53 in response to radiation of ACC cell lines, suggesting its importance as a predictive factor for radiotherapy in malignant adrenocortical tumours cases.

Cell cycle control and apoptosis of p53-isogenic cell lines treated with Hsp90 inhibitors

3202 Most patients enrolled in clinical trials have advanced disease associated with loss or mutation of p53. The goal of new molecularly-targeted therapies is to increase the therapeutic index of drug resistant tumors, many of which have non-functional p53. However, there is limited knowledge on the role of p53 in response to these agents. Upon DNA damage, cells that express wild-type p53 undergo G1 and/or G2 arrest whereas mutant or null p53 cells are presumed to arrest transiently in G2. If the G2 checkpoint is abrogated, cells will undergo apoptosis. We tested the Hsp90 inhibitors, geldanamycin and its analogues, 17-DMAG and 17-AAG, and a checkpoint inhibitor, Gö6976, as potential abrogators of the G2 checkpoint in cells treated with traditional chemotherapeutic agents, such as Doxorubicin. Isogenic pairs of lymphoblastoid, breast, lung, and colon cancer cell lines were generated through retroviral transduction of siRNA for p53 or scramble (SC) encoding vectors. Survival, p53-dependent responses, cell cycle and apoptosis were used to determine if combining DNA damage and molecularly-targeted therapy leads to mitotic catastrophe and an increase in cell death. Sequential addition of DNA damaging agents with molecular therapies was synergistic in inducing cell death in cells that arrested in G2. Such sequential use of drugs may provide a rational approach to combination therapy in cancers with functional inactivation of p53, as a means to circumvent drug-resistance to traditional chemotherapeutic agents. No significant financial relationships to disclose.

The presence of p53 mutations in human osteosarcomas correlates with high levels of genomic instability.

The p53 gene is a critical tumor suppressor that is inactivated in a majority of cancers. The central role of p53 in response to stresses such as DNA damage, hypoxia, and oncogene activation underlies this high frequency of negative selection during tumorigenic transformation. Mutations in p53 disrupt checkpoint responses to DNA damage and result in the potential for destabilization of the genome. Consistent with this, p53 mutant cells have been shown to accumulate genomic alterations in cell culture, mouse models, and some human tumors. The relationship between p53 mutation and genomic instability in human osteosarcoma is addressed in this report. Similar to some other primary human tumors, the mutation of p53 correlates significantly with the presence of high levels of genomic instability in osteosarcomas. Surprisingly, osteosarcomas harboring an amplification of the HDM2 oncogene, which inhibits the tumor-suppressive properties of p53, do not display high levels of genomic instability. These results demonstrate that the inactivation of p53 in osteosarcomas directly by mutation versus indirectly by HDM2 amplification may have different cellular consequences with respect to the stability of the genome.

Role of p53 in response to antineoplastic therapy

Role of p53 in response to antineoplastic therapy

3 O - Reentry into the cell cycle of live-irradiated quiescent normal human diploid fibroblasts: its relationships with p53 induction or inhibition

In order to assess the possible role of p53 in response to DNA damage occuring in normal quiescent cells, human diploid fibroblasts in confluence for more than two weeks were exposed to ‘UVc irradiation (20 J/m²), with and without p53 antisens oligonucleotides, and time-course cell cycle analysis was performed by flow cytometry for the next 72 hrs. The accumulation of p53 in these irradiated cells at 24^th and the inhibition of this induction by p53 antisens oligonuclcotides were, verified by ELISA. After reseeding of control unirradiated fibroblasts, reentry into the cell cycle occured at the 48^th hr. In UVc-irradiated cells, absolutly no reentry into the cell cycle was observed. In quiescent irradiated fibroblasts in which p53 accumulation was inhibited by p53 antisens, a reentry into the cell cycle appeared as soon as the 24^th, i.e even before unirradiated control cells. These results show that in confluenl normal human diploid fibroblasts, the induction of p53 by a DNA damaging agent creates a complete block and inhibits reentry into the cell cycle. The inhibition of p53 induction not only overcomes this block, regardless of DNA damages, but also permits an earlier reentry into the cell cycle In conclusion, we demonstrate that p53 protein inhibits the reentry into the cell cycle of quiescent normal human fibroblasts exposed to DNA damaging agents. This suggests that p53 could induce a G0/G1block, in addition of a G1/S block which is moreover controversed. To clearly evaluate if this block is a G0/G1 or a G1/S block some more experiments are going on, including p27^kipl, p21^wafl/cipl and cyclin D expression studies