Modulation Of P53 Research Articles

Garlic (Allium sativum) as an antidote or a protective agent against natural or chemical toxicities: A comprehensive update review.

Garlic (Allium sativum, Liliaceae) is used widely as a spice and medicinal herb not only in its native region (Central Asia and northeastern Iran) but also all around the world. Garlic has abundance chemical compounds such as allicin, alliin, S-allyl cysteines, thiacremonone, diallyl-disulfide, diallylsulfide, and others. This medicinal plant and its constituents offer a lot of benefits including free-radical scavenging, anti-inflammatory, anticholesterolemic, anti-gastric ulcer, antimicrobial, anticancer, and antioxidant properties. Garlic also modulates the activity of several metabolizing enzymes. This review summarizes various in vitro and animal studies on the protective effects of garlic against natural and chemical toxicities. It has been shown that garlic and its major components can ameliorate the toxicity of different agents in brain, kidney, blood, liver, embryo, spleen, pancreas, heart, reproductive system in part through radical scavenging, antioxidant effect, reducing lipid peroxidation, anti-inflammatory, chelating agent, cytoprotective activities, increase protein synthesis in damaged tissues, suppressing apoptosis, modulation of p53, phosphoinositide 3-kinase, Akt, nuclear factor (erythroid-derived 2)-like 2, antioxidant responsive element, p38 MAPK, inducible nitric oxide synthase, cyclooxygenase-2, cytosolic phospholipases A2, cleaved-caspase-9, cleaved-caspase-3 Bcl-2, Bcl-2-associated X, peroxisome proliferator-activated receptor gamma, NF-jB, nuclear factor-kappaB signaling pathways and cytochrome P450 enzymes. With controlled clinical trials, garlic may be introduced as a universal antidote or protective plant against many toxic agents.

The Antiproliferative Activity of Oxypeucedanin via Induction of G2/M Phase Cell Cycle Arrest and p53-Dependent MDM2/p21 Expression in Human Hepatoma Cells.

Oxypeucedanin (OPD), a furocoumarin compound from Angelica dahurica (Umbelliferae), exhibits potential antiproliferative activities in human cancer cells. However, the underlying molecular mechanisms of OPD as an anticancer agent in human hepatocellular cancer cells have not been fully elucidated. Therefore, the present study investigated the antiproliferative effect of OPD in SK-Hep-1 human hepatoma cells. OPD effectively inhibited the growth of SK-Hep-1 cells. Flow cytometric analysis revealed that OPD was able to induce G2/M phase cell cycle arrest in cells. The G2/M phase cell cycle arrest by OPD was associated with the downregulation of the checkpoint proteins cyclin B1, cyclin E, cdc2, and cdc25c, and the up-regulation of p-chk1 (Ser345) expression. The growth-inhibitory activity of OPD against hepatoma cells was found to be p53-dependent. The p53-expressing cells (SK-Hep-1 and HepG2) were sensitive, but p53-null cells (Hep3B) were insensitive to the antiproliferative activity of OPD. OPD also activated the expression of p53, and thus leading to the induction of MDM2 and p21, which indicates that the antiproliferative activity of OPD is in part correlated with the modulation of p53 in cancer cells. In addition, the combination of OPD with gemcitabine showed synergistic growth-inhibitory activity in SK-Hep-1 cells. These findings suggest that the anti-proliferative activity of OPD may be highly associated with the induction of G2/M phase cell cycle arrest and upregulation of the p53/MDM2/p21 axis in SK-HEP-1 hepatoma cells.

CBMT-48. TARGETING METABOLIC REPROGRAMMING WITH NANOCURCUMIN IN GLIOBLASTOMA MULTIFORME

Abstract Glioblastoma (GBM) is a highly glycolytic aggressive brain tumor characterized by increased proliferation and resistance to chemotherapy and radiotherapy. AMPK has been reported as tumor suppressor and reprograms the cellular metabolic pathways and produces a metabolic checkpoint on the cell cycle though mTORC1, p53 and other modulators involved in cell proliferation, growth, survival and autophagy. The AMPK activity is diminished in gastric, breast and ovarian tumor cells by activated PI3K-AKT pathways. Cancer cells are able to reprogram their energy metabolism to compensate their high bioenergetic demands needed for their aggressive growth and survival. Curcumin exhibits pleiotropic properties and activate MAPK and leads to suppress p53, Wnt/β-catenin, SHH and PI3K-AKT signaling pathways. Curcumin or diferuloylmethane is a yellow polyphenol extracted from the rhizome of turmeric (Curcuma longa). The absorption, biodistribution, metabolism, and elimination studies of curcumin have, unfortunately, shown only poor absorption, rapid metabolism, and elimination of curcumin as major reasons for poor bioavailability of this interesting polyphenolic compound. We have engineered a curcumin-based nanoparticle (Curc-NP) which demonstrates high water solubility. Curc-NP was effectively transported into the cells by nanoparticles through endocytosis and localized around the nuclei in the cytoplasms. In vitro studies proved that the cytotoxicity of Curc-NP is more effective against U-251 cell line in a dose-dependent manner. Systemic delivery of Curc-NP led to preferentially accumulation in an orthotopic preclinical glioma model minimizing systemic toxic effect. Multicolor microscopy images of the tumor tissue showed that Curc-NP particles were internalized inside tumor cells selectively and localized within nuclei. Curc-NP demonstrated to restore the dysregulated AMPK activity in glioma cells. Curc-NP-induced AMPK activation resulted in inhibition of oncogenic signalling pathways in glioma. Curc-NP-induced metabolic reprograming in glioma cells will be examined and the in vivo therapeutic efficacy of Curc-NP in an experimental rat model of GBM will also be evaluated.

Curcumin stabilizes p53 by interaction with NAD(P)H:quinone oxidoreductase 1 in tumor-derived cell lines

Curcumin is a natural phytochemical with potent anti-neoplastic properties including modulation of p53. Targeting p53 activity has been suggested as an important strategy in cancer therapy. The purpose of this study was to describe a mechanism by which curcumin restores p53 levels in human cancer cell lines.HeLa, SiHa, CaSki and MDA-MB-231 cells were exposed to curcumin and a pulse and chase and immunoprecipitation assays were performed. Here we showed that curcumin increases the half-life of p53 by a physical interaction between p53-NQO1 (p53 - NAD(P)H:quinone oxidoreductase 1) proteins after treatment with curcumin. Interestingly, the cell viability assay after treatment with curcumin showed that the cytotoxic activity was selectively higher in cervical cancer cells contained wild type p53 but not in breast cancer cells contained mutated p53. The cytotoxic effect of curcumin in cervical cancer cells was related to the complex p53-NQO1 that avoids the interaction between p53 and its negative regulator ubiquitin ligase E6-associated protein (E6AP). Finally, we demonstrated that in pancreatic epithelioid carcinoma cells (PANC1) that are knockout for NQO1, the reestablishment of NQO1 expression can stabilize p53 in presence of curcumin. Collectively, our findings showed that curcumin is necessary to promote the protein interaction of NQO1 with p53, therefore, it increases the half-life of p53, and permits the cytotoxic effect of curcumin in cancer cells containing wild type p53. Our findings suggest that the use of curcumin may reactivate the p53 pathway in cancer cells with p53 wild-type.

Kaempferia rotunda tuberous rhizome lectin induces apoptosis and growth inhibition of colon cancer cells in vitro

Kaempferia rotunda Linn. is a plant of ginger family and has many medicinal values in traditional applications, including as antimicrobial and anticancer agents. The present study aims to examine the anticancer activity of Kaempferia rotunda tuberous rhizome lectin (KRL, MW 29 ± 1.0 kDa) against colon cancer cells SW480 and SW48. KRL inhibited 67% and 59% of SW480 and SW48 cells growth respectively at the concentration of 1.0 mg/ml. The cells growth inhibition was a dose dependent manner. KRL treatments notably inhibited the colony formation capacity of the cancer cells. The surviving fractions of SW480 and SW48 cells treated with KRL significantly (p < 0.001) reduced compared to that of control cells. Significant increment of the apoptotic cells were noted following by G0/G1 or G2/M cell cycle arrest in KRL treated SW480 and SW48 cells, respectively. Modulation of PARP1, p53, p21, Bax and Bcl2 proteins expression was observed in treated cells in comparison to that of untreated cells. Furthermore, activation of caspase-3 and caspase-9 was noted in KRL treated cells and caspase-3 and caspase-9 inhibitors pre-treated cells were shown insensitive to KRL treatment. The results implied that KRL prevents SW480 and SW48 cells proliferation by the induction of apoptosis in the mitochondrial intrinsic pathway.

IASPP mediates p53 selectivity through a modular mechanism fine-tuning DNA recognition

The most frequently mutated protein in human cancer is p53, a transcription factor (TF) that regulates myriad genes instrumental in diverse cellular outcomes including growth arrest and cell death. Cell context-dependent p53 modulation is critical for this life-or-death balance, yet remains incompletely understood. Here we identify sequence signatures enriched in genomic p53-binding sites modulated by the transcription cofactor iASPP. Moreover, our p53-iASPP crystal structure reveals that iASPP displaces the p53 L1 loop-which mediates sequence-specific interactions with the signature-corresponding base-without perturbing other DNA-recognizing modules of the p53 DNA-binding domain. A TF commonly uses multiple structural modules to recognize its cognate DNA, and thus this mechanism of a cofactor fine-tuning TF-DNA interactions through targeting a particular module is likely widespread. Previously, all tumor suppressors and oncoproteins that associate with the p53 DNA-binding domain-except the oncogenic E6 from human papillomaviruses (HPVs)-structurally cluster at the DNA-binding site of p53, complicating drug design. By contrast, iASPP inhibits p53 through a distinct surface overlapping the E6 footprint, opening prospects for p53-targeting precision medicine to improve cancer therapy.

P53 supports endothelial barrier function via APE1/Ref1 suppression

The tumor suppressor protein P53 is strongly involved in orchestrating cellular defenses in the diverse variety of human tissues. Anomalies to lung endothelium permeability are streaming severe consequences towards human health, often associated with fatal outcomes. Ongoing investigations suggest that P53 exerts a prominent strategic role in crucial signaling cascades, in charge of both the maintenance and defense of pulmonary endothelium against toxic intruders. The current study employs human and bovine lung microvascular cells, as well as pharmacologic and genetic P53 modulators to demonstrate the negative regulation of APE1/Ref1 by P53. Moreover, it includes real time measurements of endothelial permeability, to reveal the disruptive role of APE1/Ref1 towards endothelial integrity. Those findings supports our efforts to elucidate the highly sophisticated regulatory network that enact endothelial adaptations under the plethora of challenging environmental factors.

SIRT1 suppresses p53-dependent apoptosis by modulation of p21 in osteoblast-like MC3T3-E1 cells exposed to fluoride

Fluoride is very crucial for development of teeth and bones. Excessive fluoride, however, causes damage to teeth and bones resulting in serious public health problem. SIRT1 regulates physiological and pathological processes such as apoptosis and cell cycle. Although SIRT1 inhibits p53-mediated transactivation, how SIRT1 regulates p53 in fluorosis remains unclear. This study aims to investigate the involvement of SIRT1 in fluoride-induced cell cycle arrest and apoptosis in MC3T3-E1 cells and the underlying mechanism. Cell apoptosis was determined using Annexin V-FITC/PI dual staining, cell cycle detected with PI staining, intracellular ROS levels measured with DCFH-DA probe, and apoptosis-related protein expressions determined using Western blotting. Results showed that there was a promotion in apoptosis rate, intracellular ROS levels, the ratio of Bax/Bcl-2, protein expression (Cyt c, Caspase-3, p53, Ac-p53 and p21) and blockage of S phase after cells were exposed to NaF. Afterwards, the influence of SIRT1 on apoptosis was explored after SRT1720 (SIRT1 activator) and Ex-527 (SIRT1 inhibitor) was introduced. Results indicated that SRT1720 in combination with fluoride significantly decreased the intracellular ROS levels, the protein expression of Caspase-3, Ac-p53 and p21 and alleviated apoptosis, while it was reversed by Ex-527. Collectively, SIRT1 plays an essential role in protection against fluoride-induced oxidative stress and mitochondria-dependent apoptosis in MC3T3-E1 cells. The SIRT1/p53/p21 pathway may be a potential therapeutic target for fluorosis.

Roles of computational modelling in understanding p53 structure, biology, and its therapeutic targeting.

The transcription factor p53 plays pivotal roles in numerous biological processes, including the suppression of tumours. The rich availability of biophysical data aimed at understanding its structure–function relationships since the 1990s has enabled the application of a variety of computational modelling techniques towards the establishment of mechanistic models. Together they have provided deep insights into the structure, mechanics, energetics, and dynamics of p53. In parallel, the observation that mutations in p53 or changes in its associated pathways characterize several human cancers has resulted in a race to develop therapeutic modulators of p53, some of which have entered clinical trials. This review describes how computational modelling has played key roles in understanding structural-dynamic aspects of p53, formulating hypotheses about domains that are beyond current experimental investigations, and the development of therapeutic molecules that target the p53 pathway.

Thymoquinone, an Active Constituent of Black Seed Attenuates CCl4 Induced Liver Injury in Mice via Modulation of Antioxidant Enzymes, PTEN, P53 and VEGF Protein.

AIM:The present study was undertaken to evaluate the possible protective role of thymoquinone on CCl4-induced hepatotoxicity.METHODS:The activities of liver function enzymes and antioxidant enzymes were measured. Haematoxylin-Eosin staining was performed to analyze the live tissue alterations. Additionally, expression pattern of different proteins was evaluated through immunohistochemistry staining.RESULTS:The antioxidants enzymes activities were decreased significantly in the CCl4 induced group whereas recovery/increase of antioxidant enzymes was observed when thymoquinone was given to the mice. Moreover, thymoquinone administration significantly decrease the serum levels of alanine aminotransferase (ALT), alkaline phosphatase (ALP), and serum aspartate aminotransferase (AST). Liver tissue alterations were noted in CCl4 treated group whereas treatment with thymoquinone significantly prevented the CCl4-induced histological alteration. The expression of PTEN protein was high in CCl4 plus thymoquinone treated group while the loss of PTEN protein expression was observed in CCl4 treated group. Moreover, high expression of P53 protein was noticed in CCl4 treated the group as compared to CCl4 plus thymoquinone group. Difference in expression pattern of PTEN and p53 protein in CCl4 group and thymoquinone plus CCl4 treated group was statically significant (p < 0.05). Besides, expression of VEGF was high in CCl4 treated group as well as thymoquinone plus CCl4 treated group and difference in expression pattern was statically insignificant (p > 0.05).CONCLUSION:Our results suggest that thymoquinone can protect CCl4 induced liver damage and could be a preventive drug in the development of novel therapeutic agents for liver diseases.

A Novel Small Molecule p53 Stabilizer for Brain Cell Differentiation.

Brain tumor, as any type of cancer, is assumed to be sustained by a small subpopulation of stem-like cells with distinctive properties that allow them to survive conventional therapies and drive tumor recurrence. Thus, the identification of new molecules capable of controlling stemness properties may be key in developing effective therapeutic strategies for cancer by inducing stem-like cells differentiation. Spiropyrazoline oxindoles have previously been shown to induce apoptosis and cell cycle arrest, as well as upregulate p53 steady-state levels, while decreasing its main inhibitor MDM2 in the HCT116 human colorectal carcinoma cell line. In this study, we made modifications in this scaffold by including combinations of different substituents in the pyrazoline ring in order to obtain novel small molecules that could modulate p53 activity and act as differentiation inducer agents. The antiproliferative activity of the synthesized compounds was assessed using the isogenic pair of HCT116 cell lines differing in the presence or absence of the p53 gene. Among the tested spirooxindoles, spiropyrazoline oxindole 1a was selective against the cancer cell line expressing wild-type p53 and presented low cytotoxicity. This small molecule induced neural stem cell (NSC) differentiation through reduced SOX2 (marker of multipotency) and increased βIII-tubulin (marker of neural differentiation) which suggests a great potential as a non-toxic inducer of cell differentiation. More importantly, in glioma cancer cells (GL-261), compound 1a reduced stemness, by decreasing SOX2 protein levels, while also promoting chemotherapy sensitization. These results highlight the potential of p53 modulators for brain cell differentiation, with spirooxindole 1a representing a promising lead molecule for the development of new brain antitumor drugs.

ATM Dependent DUSP6 Modulation of p53 Involved in Synergistic Targeting of MAPK and p53 Pathways with Trametinib and MDM2 Inhibitors in Cutaneous Melanoma.

MAPK and p14ARF–MDM2–p53 pathways are critical in cutaneous melanomas. Here, synergistic combination of the MEK inhibitor, trametinib, with MDM2 inhibitors, nutlin-3/RG7388/HDM201, and the mechanistic basis of responses, for BRAFV600E and p53WT melanoma cells, are reported. The combination treatments induced higher levels of p53 target gene transcripts and protein products, resulting in increased cell cycle arrest and apoptosis compared with MDM2 inhibitors alone, suggesting trametinib synergized with MDM2 inhibitors via upregulation of p53-dependent pathways. In addition, DUSP6 phosphatase involvement was indicated by downregulation of its mRNA and protein following pERK reduction by trametinib. Furthermore, suppression of DUSP6 by siRNA, or inhibition with the small molecule inhibitor, BCI, at a dose without cytotoxicity, potentiated the effect of MDM2 inhibitors through increased ATM-dependent p53 phosphorylation, as demonstrated by complete reversal with the ATM inhibitor, KU55933. Trametinib synergizes with MDM2 inhibitors through a novel DUSP6 mechanism in BRAFV600E and p53WT melanoma cells, in which DUSP6 regulation of p53 phosphorylation is mediated by ATM. This provides a new therapeutic rationale for combination treatments involving activation of the ATM/p53 pathway and MAPK pathway inhibition.

Amentoflavone Induces Autophagy and Modulates p53.

Objective Amentoflavone is the main component of Selaginella tamariscina widely known as an oriental traditional medicinal stuff that has been known to have a variety of medicinal effects such as the induction of apoptosis, anti- metastasis, and anti-inflammation. However, the effect of amentoflavone on autophagy has not been reported until now. The aim of this study was to investigate whether amentoflavone has a positive effect on the induction of autophagy related to cell aging. Materials and Methods In this experimental study, the aging of young cells was induced by the treatment with insulin- like growth factor-1 (IGF-1) at 50 ng/mL three times every two days. The effect of amentoflavone on the cell viability was evaluated in A549 and WI-38 cells using 3-(4,5-dimethyl-2-yl)-2,5- diphenyl tetrazolium bromide (MTT) assay. The induction of autophagy was detected using autophagy detection kit. The expression of proteins related to autophagy and IGF-1 signaling pathway was examined by western blot analysis and immunofluorescence assay. Results First of all, it was found that amentoflavone induces the formation of autophagosome. In addition, it enhanced the expression level of Atg7 and increased the expression levels of Beclin1, Atg3, and LC3 associated with the induction of autophagy in immunofluorescence staining and western blot analyses. Moreover, amentoflavone inhibited the cell aging induced by IGF-1 and hydrogen peroxide. In particular, the levels of p53 and p-p21 proteins were increased in the presence of amentoflavone. Furthermore, amentoflavone increased the level of SIRT1 deacetylating p53. ConclusionOur results suggest that amentoflavone could play a positive role in the inhibition of various diseases associated with autophagy and the modulation of p53.

Andrographolide enhances redox status of liver cells by regulating microRNA expression

Andrographis paniculata Nees and its principal compound andrographolide are well known for exerting beneficial effects by modulating signaling pathways in different biological systems. Our earlier studies have demonstrated the ability of andrographolide as well as andrographolide enriched extracts to activate Nrf2/HO-1 pathway through adenosine A2a receptor. Present study investigated ability of andrographolide to regulate Nrf2 induced antioxidant defense systems by miRNAs using HepG2 cells. Andrographolide strongly induced Nrf2 which in turn modulated enzymes of glutathione and thioredoxin antioxidant systems. It also regulated crucial transcription factors viz. hepatocyte nuclear factor alpha (HNF4A) and tumor suppressor protein 53 (p53). Downregulation of HNF4A by andrographolide led to decrease in miRNAs regulating Heme oxygenase-1 (miR-377) and glutathione cysteine ligase (miR-433). Upregulation of p53 on the other hand led to increase in miRNAs regulating thioredoxin interacting protein (miR-17, miR-224) and glutathione peroxidase (miR-181a). Involvement of p53 and HNF4A in modulation of these miRNAs was confirmed by chromatin immunoprecipitation assay. Overall, the work reveals that andrographolide through modulation of p53 and HNF4A, regulates miRNAs leading to upregulation of HO-1, glutathione and thioredoxin systems. Andrographolide thus, can play a beneficial role in modulating antioxidant defense in oxidative stress induced diseases such as diabetes, ageing etc.

Upregulation of microRNA-17-5p contributes to hypoxia-induced proliferation in human pulmonary artery smooth muscle cells through modulation of p21 and PTEN

BackgroundPulmonary arterial smooth muscle cell (PASMC) proliferation in response to hypoxia plays an important role in the vascular remodelling that occurs in hypoxic pulmonary hypertension. MicroRNAs (miRs) are emerging as important regulators in the progression of pulmonary hypertension. In this study, we investigated whether the expression of miR-17-5p is modulated by hypoxia and is involved in the hypoxia-induced proliferation of PASMCs.MethodsHuman PASMCs were cultured under hypoxic conditions. miR-17-5p expression was determined by real-time RT-PCR. A BrdU incorporation assay and time-lapse recording were utilized to determine cell proliferation and migration.ResultsPASMC proliferation was increased by moderate hypoxia (3% oxygen) but was reduced by severe hypoxia (0.1% oxygen) after 48 h. Moderate hypoxia induced miR-17-5p expression. Overexpression of miR-17-5p by transfection with miR-17-5p enhanced cell proliferation and migration in normoxia, whereas knockdown of miR-17-5p with anti-miR-17-5p inhibitors significantly reduced cell proliferation and migration. The expression of miR-17-5p target genes, specifically phosphatase and tensin homologue (PTEN) and cyclin-dependent kinase inhibitor 1 (p21WAF1/Cip1, p21), was reduced under moderate hypoxia in PASMCs. Under normoxia, overexpression of miR-17-5p in PASMCs reduced the expression of PTEN and p21.ConclusionOur data indicate that miR-17-5p might play a significant role in hypoxia-induced pulmonary vascular smooth muscle cell proliferation by regulating multiple gene targets, including PTEN and p21, and that miR-17-5p could be a novel therapeutic target for the management of hypoxia-induced PH.

Potentiating apoptosis and modulation of p53, Bcl2, and Bax by a novel chrysin ruthenium complex for effective chemotherapeutic efficacy against breast cancer.

Breast cancer is the most frequent cause of cancer in women. In the current study, transition metal ruthenium was complexed with flavonoid chrysin to evaluate the chemotherapeutic potential of this compound in Michigan Cancer Foundation-7 (MCF-7) human mammary cancer cell line and 7,12-dimethylbenz(α)anthracene-induced mammary cancer in female Sprague-Dawley rats. The characterizations of the complex were accomplished through UV-visible, NMR, IR, Mass spectra, and XRD techniques and antioxidant activity was assessed by DPPH, FRAP, and ABTS methods. In vitro studies included cell viability, cell cycle analysis, DNA fragmentation, and marker analysis by western blot analysis and found that complex treatment suppressed cell growth-induced cell cycle arrest and enhanced the induction of apoptosis in cancer cells. Moreover, complex treatment modulated signaling pathways including mTOR, VEGF, and p53 in the MCF-7 cells. Acute and subacute toxicity was performed in rats to determine the therapeutic doses. Breast cancer in rats was initiated by the administration of 7,12-dimethylbenz(α)anthracene (0.5 mg/100 g body weight) via single tail vein injection. The histopathological analysis after 24 weeks of carcinogenesis study depicted substantial repair of hyperplastic lesions. Immunohistochemical analysis revealed upregulation of Bax and p53 and downregulation of Bcl2 proteins and TUNEL assay showed an increase in apoptotic index in ruthenium-chrysin-treated groups as compared to the carcinogen control. Our findings from the in vitro and in vivo study support the continued investigation of ruthenium-chrysin complex possesses a potential chemotherapeutic activity against breast cancer and was efficient in reducing hyperplastic lesions in the mammary tissues of rats by inducing apoptosis.

PO-463 Augmentation of NAD+BY NQO1 activation attenuates cisplatin-mediated hearing impairment

IntroductionCisplatin [cis-diaminedichloroplatinum-II] is an extensively used chemotherapeutic agent, and one of its most adverse effects is ototoxicity. A number of studies have demonstrated that these effects are related to oxidative stress and DNA damage. However, the precise mechanism underlying cisplatin-associated ototoxicity is still unclear. The cofactor nicotinamide adenine dinucleotide (NAD+) has emerged as a key regulator of cellular energy metabolism and homeostasis. Although a link between NAD+-dependent molecular events and cellular metabolism is evident, it remains unclear whether modulation of NAD+ levels has an impact on cisplatin-induced hearing impairment.Material and methodsTo investigate whether augmentation of NAD+ by NQO1 activation using b-Lapachone (b-Lap) attenuates cisplatin-mediated hearing impairment, male C57BL/6 mice and NQO1 knockout mice on a C57BL/6 background were used. For analysis of the auditory threshold, auditory brainstem response (ABR) was recorded. For biochemical analysis, we measured the enzymatic activity of SIRT1, PARP1, ROS production, NAD+/NADH ratio, mRNA levels of miR-34a and pro-inflammatory cytokines. Immunohistochemistry and western blot analysis were also performed.Results and discussionsWe have demonstrated for the first time that both the protein expression level and the activity of SIRT1 were suppressed by the reduction of intracellular NAD+ levels in cisplatin-treated cochlear tissue. We also found that the decrease in SIRT1 protein expression and its activity after cisplatin exposure were mediated by the increase in transcriptional activity of p53 for miR-34a expression and PARP-1 activation causing NAD+-depletion, respectively. However, the increase in cellular NAD+ levels by NQO1 activation using b-Lap prevented mice from cisplatin-induced cochlear damage and hearing impairment through the modulation of PARP-1, SIRT1, p53, and NF-kB.ConclusionConsidering that b-Lap itself did not attenuate the tumoricidal effect of cisplatin, these results suggest that the direct modulation of the cellular NAD+ level by pharmacological agents could be a promising therapeutic strategy for enhancing the efficacy of cisplatin chemotherapy without its adverse effects.

PO-094 Persistent Myc activity prevents resolution of liver injury

IntroductionFas/CD95 is a member of the death receptor superfamily that can induce apoptosis and it functions as the key component of the extrinsic death pathway. This study reports on a relevant function of Fas death receptor that can regulate transcription factors p53, MYC, NF-κ B and pro-apoptotic gene PUMA, that are important in intrinsic death pathway in hepatocellular carcinoma (HCC).Material and methodsFor the in vivo study, genes Fas, and FasL from 111 samples (39 HCC, 30 peri-HCC, 31 cirrhosis, and 11 normal) from patients undergoing liver resection without any prior treatments were analysed. For the in vitro study, hepatocytes cell line IHH and human HCC cell lines HepG2, HUH7, JHH6 were used. Apoptosis-induction was performed by using anti-Fas (DX2) at a concentration of 250 ng/ml and 500 ng/ml for 24 hours. Flow cytometry and quantitative real-time PCR were performed to analyse the data.Results and discussionThe expressions of Fas and FasL mRNA were significantly noticed in HCC as compared to normal tissues. FasL mRNA was high expressed in viral-related compared to metabolic-related HCC while Fas was high in HCC tissues without vascular invasion compared to tissues with vascular invasion. For in vitro study, after treatment of anti-CD95, TNF-α was significantly up regulated in HEPG2 compare to IHH cells. Transcription factors p53, MYC and NF-κ B were down regulated in HEPG2 cells while up regulated or no difference change in IHH cells. Pro-apoptotic gene Puma was only up regulated in IHH cell line. Telomerase was up regulated in IHH and significantly down regulated in HEPG2.ConclusionsWe observed a modulation of transcription factors p53, MYC, NF-κ B and pro-apoptotic genes Puma, TNF-α, and Telomerase after induction by CD95 antibody in acute phase treatment for 24 hours. The expression of PUMA and MYC were regulated by the transcription factors p53, thus suggesting the involvement of Fas in p53-dependent apoptosis pathway.

A G3BP1-Interacting lncRNA Promotes Ferroptosis and Apoptosis in Cancer via Nuclear Sequestration of p53.

Long noncoding RNAs (lncRNA) have been associated with various types of cancer; however, the precise role of many lncRNAs in tumorigenesis remains elusive. Here we demonstrate that the cytosolic lncRNA P53RRA is downregulated in cancers and functions as a tumor suppressor by inhibiting cancer progression. Chromatin remodeling proteins LSH and Cfp1 silenced or increased P53RRA expression, respectively. P53RRA bound Ras GTPase-activating protein-binding protein 1 (G3BP1) using nucleotides 1 and 871 of P53RRA and the RRM interaction domain of G3BP1 (aa 177-466). The cytosolic P53RRA-G3BP1 interaction displaced p53 from a G3BP1 complex, resulting in greater p53 retention in the nucleus, which led to cell-cycle arrest, apoptosis, and ferroptosis. P53RRA promoted ferroptosis and apoptosis by affecting transcription of several metabolic genes. Low P53RRA expression significantly correlated with poor survival in patients with breast and lung cancers harboring wild-type p53. These data show that lncRNAs can directly interact with the functional domain of signaling proteins in the cytoplasm, thus regulating p53 modulators to suppress cancer progression.Significance: A cytosolic lncRNA functions as a tumor suppressor by activating the p53 pathway. Cancer Res; 78(13); 3484-96. ©2018 AACR.

Protective Effect of Cinnamon Zeylanicum, Berberis Vulgaris and Ulva Lactuca Extracts on Hepatocellular Toxicity Induced by Aspergillus Flavus Intake in Rats

Background: Aflatoxin B1 produced by the fungus Asperagillus flavus causes great economic losses and poses health hazards to human and animals through its toxic biological effects on liver, kidney and lungs. The aim of this work was to study the potential protective effect of Cinnamomum zeylanicum, Berberis vulgaris and Ulva lactuca extractson hepatocyte toxicity induced by A. flavus intakein rats. We also investigated the effect of A. flavus and the studied extracts on liver and kidney structure and function and the potential modulation of p53 and ICAM-1 gene expression as well as the liver antioxidant status. Our results showed a damaging effect of A. flavus intake on both liver and kidney as reflected by liver histopathological examination and the impaired liver and kidney functions measured by ALT, AST, Albumin, Urea, creatinine and glucose. Cinnamon and Berberis and Ulva pre-treatment kept these parameters to almost its normal levels compared to the induced unprotected animals. All tested extracts reduced the oxidative stress status and increased the antioxidant status by lowering TBARS and increasing NO levels significantly but had no significant effect on SOD activity. A.flavus intake causeda significant decline in both P53 and ICAM-1 gene expression; however, administration of Cinnamon or Berberis caused a significant increase in expression with Cinnamon causing the highest increase in p53 and Berberiscausing the highest increase in ICAM-1. In conclusion, we recommended the use of cinnamon Zeylanicum or Berberis vulgaris as protective natural antioxidants against hepatocellular aflatoxin induced toxicity.