Inhibitory effect of c-Myc on p53-induced apoptosis in leukemia cells. Microarray analysis reveals defective induction of p53 target genes and upregulation of chaperone genes

Protocatechuic acid (PCA), one of the main metabolites of complex polyphenols, exerts numerous biological activities including antiapoptotic, anti-inflammatory, and antiatherosclerotic effects. Oxidised LDL have atherogenic properties by damaging arterial wall cells and inducing p53-dependent apoptosis in macrophages. This study was aimed at defining the molecular mechanism responsible for the protective effects of PCA against oxidative and proapoptotic damage exerted by oxLDL in J774 A.1 macrophages. We found that the presence of PCA in cells treated with oxLDL completely inhibited the p53-dependent apoptosis induced by oxLDL. PCA decreased oxLDL-induced ROS overproduction and in particular prevented the early increase of ROS. This decrease seemed to be the main signal responsible for maintaining the intracellular redox homeostasis hindering the activation of p53 induced by ROS, p38MAPK, and PKCδ. Consequently the overexpression of the proapoptotic p53-target genes such as p66Shc protein did not occur. Finally, we demonstrated that PCA induced the activation of JNK, which, in turn, determined the increase of nuclear Nrf2, leading to inhibition of the early ROS overproduction. We concluded that the antiapoptotic mechanism of PCA was most likely related to the activation of the JNK-mediated survival signals that strengthen the cellular antioxidant defences rather than to the PCA antioxidant power.

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Induction of p53-Dependent Apoptosis by Prostaglandin A2

Biomolecules ◽

10.3390/biom10030492 ◽

2020 ◽

Vol 10 (3) ◽

pp. 492 ◽

Cited By ~ 1

Author(s):

Su-Been Lee ◽

Sangsun Lee ◽

Ji-Young Park ◽

Sun-Young Lee ◽

Ho-Shik Kim

Keyword(s):

Target Genes ◽

P53 Gene ◽

Histone H2a ◽

Molecule Inhibitor ◽

Induction Of Apoptosis ◽

Prostaglandin A2 ◽

Induced Apoptosis ◽

Inhibitor Of Protein Synthesis ◽

P53 Target Genes ◽

Hct116 Cells

Prostaglandin (PG) A2, one of cyclopentenone PGs, is known to induce activation of apoptosis in various cancer cells. Although PGA2 has been reported to cause activation of apoptosis by altering the expression of apoptosis-related genes, the role of p53, one of the most critical pro-apoptotic genes, on PGA2-induced apoptosis has not been clarified yet. To address this issue, we compared the apoptosis in HCT116 p53 null cells (HCT116 p53-/-) to that in HCT116 cells containing the wild type p53 gene. Cell death induced by PGA2 was associated with phosphorylation of histone H2A variant H2AX (H2AX), activation of caspase-3 and cleavage of poly(ADP-ribose) polymerase 1 in HCT116 cells. Induction of apoptosis in PGA2-treated cells was almost completely prevented by pretreatment with a pan-caspase inhibitor, z-VAD-Fmk, or an inhibitor of protein synthesis, cycloheximide. While PGA2 induced apoptosis in HCT116 cells, phosphorylation of p53 and transcriptional induction of p53-target genes such as p21WAF1, PUMA, BAX, NOXA, and DR5 occurred. Besides, pretreatment of pifithrin-α (PFT-α), a chemical inhibitor of p53’s transcriptional activity, interfered with the induction of apoptosis in PGA2-treated HCT116 cells. Pretreatment of NU7441, a small molecule inhibitor of DNA-activated protein kinase (DNA-PK) suppressed PGA2-induced phosphorylation of p53 and apoptosis as well. Moreover, among target genes of p53, knockdown of DR5 expression by RNA interference, suppressed PGA2-induced apoptosis. In the meanwhile, in HCT116 p53-/- cells, PGA2 induced apoptosis in delayed time points and with less potency. Delayed apoptosis by PGA2 in HCT116 p53-/- cells was also associated with phosphorylation of H2AX but was not inhibited by either PFT-α or NU7441. Collectively, these results suggest the following. PGA2 may induce p53-dependent apoptosis in which DNA-PK activates p53, and DR5, a transcriptional target of p53, plays a pivotal role in HCT116 cells. In contrast to apoptosis in HCT116 cells, PGA2 may induce apoptosis in a fashion of less potency, which is independent of p53 and DNA-PK in HCT116 p53-/- cells

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Increased p53 Acetylation By SIRT1 Inhibition Is Required for Optimal Activation of p53 Activity and Significantly Enhances the Ability of HDM2 Inhibitors to Target CML LSC

Blood ◽

10.1182/blood.v124.21.4521.4521 ◽

2014 ◽

Vol 124 (21) ◽

pp. 4521-4521

Author(s):

Yann Pierre Kevin Duchartre ◽

Ling Li ◽

Tinisha McDonald ◽

YinWei Ho ◽

Yao-Te Hsieh ◽

...

Keyword(s):

Cell Cycle ◽

Stem Cells ◽

Transcriptional Activity ◽

Target Genes ◽

Myelogenous Leukemia ◽

Growth And Survival ◽

Cycle Arrest ◽

Cd34 Cells ◽

Induced Apoptosis ◽

P53 Target Genes

Abstract BCR-ABL tyrosine kinase inhibitors (TKI) are effective in inducing remissions and prolonging survival in chronic myelogenous leukemia (CML) patients, but do not eliminate leukemia stem cells (LSCs) that are responsible for establishment, maintenance and recurrence of the disease. We have shown that the NAD-dependent SIRT1 deacetylase is overexpressed in CML LSC (Li et al., Cancer Cell, 21:266, 2013). SIRT1 participates in the maintenance, growth and treatment resistance of CML LSC by deacetylation and inhibition of the p53 pathway that regulates cell cycle and apoptosis. Inhibition of SIRT1 using RNAi and the small molecule SIRT1 inhibitor Tenovin-6 (TV-6) inhibits growth and survival of CML LSC by itself and results in enhanced targeting of LSC in combination with TKI treatment. The effects of SIRT1 inhibition are related at least in part to enhanced acetylation of the p53 protein associated with enhanced p53 transcriptional activity. Here we examined the efficacy of an alternative strategy to activate p53, using inhibition of the p53 regulatory protein HDM2, in activating p53 transcriptional activity and inhibiting CML LSC growth and survival compared to SIRT1 inhibition. Nutlin-3 (Nut-3) is a small molecule HDM2 inhibitor that disrupts the p53-HDM2 interaction which has proceeded to clinical trials. Treatment of CML CP CD34+ cells with Nut-3 (1, 2, 5µM) increased expression of the p53 target genes p21 (associated with cell cycle arrest) and NOXA and PIG3 (associated with apoptosis), inhibited proliferation and induced apoptosis to a significantly lesser extent than TV-6 (1, 2 µM) (Nut 5 µM vs TV 2 µM ; p<0.0003) . However, Nutlin-3 enhanced p53 target gene expression and inhibited proliferation and survival of normal CD34+ cells to a similar extent as CML CD34+ cells. This was in contrast to TV-6 which although inhibiting proliferation of both CML and normal CD34+ cells, selectively induced apoptosis in CML compared to normal CD34+ cells. Treatment of CML CD34+ cells with the combination of Nut-3 (2, 5µM) and TV-6 (1µM) significantly increased the expression of p53 target genes (p21, PIG3, NOXA), and enhanced apoptosis of CML CD34+ cells compared to Nut-3 or TV-6 alone. 32D-BCR-ABL cells transduced with lentivirus vectors expressing p53 shRNA demonstrated significantly reduced apoptosis following treatment with the combination of Nut-3 and TV-6 compared to cells expressing control shRNA, indicating that the effects of this treatment are p53 dependent. Our results indicate that enhancement of p53 acetylation by SIRT1 inhibition is required for optimal activation of p53 transcriptional activity and induction of apoptosis in CML LSC. These results further support SIRT1 as a valid therapeutic target in CML, and suggest that addition of SIRT1 inhibitors may significantly enhance the ability of HDM2 inhibitors to eliminate CML LSC. Table 1 : Effects of Nutlin-3, Tenovin-6 or combination on the expression of apoptosis/cell cycle arrest related-genes, apoptosis and proliferation in cord blood and CML CD34+ cells. Figure 1 Figure 1. SEM values ; significance, compared to controls: ***p<0.001. ****p<0.0001. Key words : Chronic Myelogenous Leukemia (CML), hematopoietic stem cells, p53, SIRT1. Disclosures No relevant conflicts of interest to declare.

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Nobiletin Inhibits Non-Small-Cell Lung Cancer by Inactivating WNT/β-Catenin Signaling through Downregulating miR-15-5p

Evidence-based Complementary and Alternative Medicine ◽

10.1155/2021/7782963 ◽

2021 ◽

Vol 2021 ◽

pp. 1-9

Author(s):

Sang Hyup Han ◽

Jeong Hee Han ◽

Wan Joo Chun ◽

Sang Soo Lee ◽

Hae Sung Kim ◽

...

Keyword(s):

Lung Cancer ◽

Small Cell Lung Cancer ◽

Cell Lung Cancer ◽

Target Genes ◽

Inhibitory Effect ◽

Soft Agar ◽

Small Cell ◽

Small Cell Lung ◽

Protein Levels ◽

Induced Apoptosis

Background. Nobiletin is a natural compound with anticancer activity; however, the mechanism is not clear. Methods. The inhibitory effect of nobiletin on non-small-cell lung cancer (NSCLC) cells was examined using soft agar, Transwell, and apoptosis analyses. Cancer stemness was measured by sphere assay. Genes and miRNAs regulated by nobiletin were identified by whole-genome sequencing. Protein levels were detected by western blot and immunofluorescence assays. Results. Nobiletin significantly inhibited NSCLC cell colony formation and sphere formation and induced apoptosis. Nobiletin upregulated negative regulators of WNT/β-catenin signaling, including NKD1, AXIN2, and WIF1, while it inhibited the expression of β-catenin and its downstream genes, including c-Myc, c-Jun, and cyclin D1. Furthermore, we identified that GN inhibits miR-15-5p expression in NSCLC cells and that NKD1, AXIN2, and WIF1 are the target genes of miR-15-5p. Conclusions. Nobiletin has a strong inhibitory effect on NSCLC, and nobiletin plays an anticancer role by inhibiting miR-15-5p/β-catenin signaling in NSCLC.

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p53 Mediates Colistin-Induced Autophagy and Apoptosis in PC-12 Cells

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00641-16 ◽

2016 ◽

Vol 60 (9) ◽

pp. 5294-5301 ◽

Cited By ~ 11

Author(s):

Ling Zhang ◽

Daoyuan Xie ◽

Xueping Chen ◽

Maria L. R. Hughes ◽

Guozheng Jiang ◽

...

Keyword(s):

Target Genes ◽

Mammalian Target Of Rapamycin ◽

Inhibitory Effect ◽

Pc 12 Cells ◽

Link Type ◽

Cellular Events ◽

Cycle Arrest ◽

Induced Apoptosis ◽

20 Nm ◽

Amp Activated Protein Kinase

ABSTRACTThe mechanism of colistin-induced neurotoxicity is still unknown. Our recent study (L. Zhang, Y. H. Zhao, W. J. Ding, G. Z. Jiang, Z. Y. Lu, L. Li, J. L. Wang, J. Li, and J. C. Li, Antimicrob Agents Chemother 59:2189–2197, 2015,http://dx.doi.org/10.1128/AAC.04092-14; H. Jiang, J. C. Li, T. Zhou, C. H. Wang, H. Zhang, and H. Wang, Int J Mol Med 33:1298–1304, 2014,http://dx.doi.org/10.3892/ijmm.2014.1684) indicates that colistin induces autophagy and apoptosis in rat adrenal medulla PC-12 cells, and there is interplay between both cellular events. As an important cellular stress sensor, phosphoprotein p53 can trigger cell cycle arrest and apoptosis and regulate autophagy. The aim of the present study was to investigate the involvement of the p53 pathway in colistin-induced neurotoxicity in PC-12 cells. Specifically, cells were treated with colistin (125 μg/ml) in the absence and presence of a p53 inhibitor, pifithrin-α (PFT-α; 20 nM), for 12 h and 24 h, and the typical hallmarks of autophagy and apoptosis were examined by fluorescence/immunofluorescence microscopy and electron microscopy, real-time PCR, and Western blotting. The results indicate that colistin had a stimulatory effect on the expression levels of the target genes and proteins involved in autophagy and apoptosis, including LC3-II/I, p53, DRAM (damage-regulated autophagy modulator), PUMA (p53 upregulated modulator of apoptosis), Bax, p-AMPK (activated form of AMP-activated protein kinase), and caspase-3. In contrast, colistin appeared to have an inhibitory effect on the expression of p-mTOR (activated form of mammalian target of rapamycin), which is another target protein in autophagy. Importantly, analysis of the levels of p53 in the cells treated with colistin revealed an increase in nuclear p53 at 12 h and cytoplasmic p53 at 24 h. Pretreatment of colistin-treated cells with PFT-α inhibited autophagy and promoted colistin-induced apoptosis. This is the first study to demonstrate that colistin-induced autophagy and apoptosis are associated with the p53-mediated pathway.

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Growth Factor Independent-1 (Gfi1) As a New Target for Human Leukemia Therapy

Blood ◽

10.1182/blood.v118.21.560.560 ◽

2011 ◽

Vol 118 (21) ◽

pp. 560-560

Author(s):

Cyrus Khandanpour ◽

James D. Phelan ◽

Riyan Chen ◽

Shane Horman ◽

Lothar Vassen ◽

...

Keyword(s):

Cell Cycle ◽

T Cell ◽

B Cell ◽

Target Genes ◽

Cell Lymphoma ◽

Leukemic Cells ◽

Leukemia Cells ◽

Human Leukemia ◽

Cell Leukemia ◽

P53 Target Genes

Abstract Abstract 560 More than 50% of patients diagnosed with B or T-cell leukemia and lymphoma will fail current treatment protocols. This highlights the urgent need for new and improved therapies. Since the transcription factor Growth factor independent-1 (Gfi1) plays an important role in lymphoid differentiation, we explored whether it might be a suitable target for therapy. Using mouse models in which T-cell leukemia can be induced by transgenic expression of mutated forms of Notch1, by injection of the carcinogen N-Ethyl–nitrosourea (ENU) or infection with a Murine Moloney Leukemia Virus, we found that Gfi1 knockout mice had a significantly lower incidence and a longer latency period of T-ALL. To verify whether targeting Gfi1 would be a novel approach to treat B- or T-cell lymphoma, we used Mx1Cre Gfi1fl/fl mice. In these mice, injection of polyinosinic-polycytidylic acid (pIpC) activates the Mx1 promoter driven Cre expression, which ultimately leads to the deletion of the floxed Gfi1 alleles. As controls, we used Gfi1fl/fl mice, which lack the Cre recombinase and thus still express Gfi1 after pIpC injection. To elicit a T- or B-cell lymphoma, we used ENU injection combined with expression of a mutated Notch1 transgene for T-ALL, or transgenic over-expression of c-Myc for B-cell leukemia (Eμ-Myc). Using in-vivo ultrasound supported imaging, we observed complete regression of tumour masses when Gfi1 was eliminated in Mx1Cre Gfi1fl/fl mice, curing the mice of the either the T or B-cell malignancies. Strikingly, this effect was observed in the absence of any other treatment regimen. To explore the mechanisms underlying this phenomenon, we explanted tumor samples from mice, in which Gfi1 expression was either maintained or deleted and performed gene expression arrays. A comparative analysis of the array data demonstrated that loss of Gfi1 affects pathways of key importance for leukemia such as metabolism, cell cycle progression, basal transcription and apoptosis but also the response to DNA damage. It has been shown previously for non hematological malignancies that oncogenic transformation in conjuction with dysregulated cell cycle induces DNA strand breaks (DSBs), which leads to an increased p53 dependent apoptotic response in tumours compared to non-transformed cells. This forces the tumor cells to counteract this effect – for instance by selection for the loss of p53. Consistent with this concept, we noted that leukemic cells from our tumor models displayed a greater amount of DSBs and also higher rates of spontaneous apoptosis than normal cells. Interestingly, the number of apoptotic cells was further increased in those tumors where Gfi1 had been deleted. We hypothesized that Gfi1 protects leukemia cells against DSB induced apoptosis. To test this hypothesis, we irradiated in Gfi1+/+ and Gfi1−/− thymocytes, which induces DSB in thymocytes. In line with our hypothesis we found that Gfi1−/− thymocytes showed increased rates of apoptosis compared to irradiated Gfi1+/+ thymocytes and that loss of Gfi1 led to an increased induction of pro-apoptotic genes such as Bax, Noxa and Puma after irradiation. Since Bax, Noxa and Puma are all p53 target genes, we investigated a possible link between Gfi1 and p53 and found that (I) Gfi1 binds to p53, (II) that Gfi1 inhibits the transcription of p53 target genes and (III) that Gfi1 occupies p53 target gene at the same sites as p53. In summary, Gfi1 antagonizes p53 function and loss of Gfi1 sensitizes cells to p53-mediated apoptosis. Next, we used different human T-ALL cell lines and treated these cells either with sh-RNA lentivirus or morpholinos to abrogate GFI1 expression. In all cases, down-regulation of GFI1 expression led to increased apoptosis and impeded growth of human leukemia cells. Finally, we transplanted leukemic cells of T-ALL patients into NOD-Scid, IL2Rnull (NSG) mice, waited for the leukemic cells to engraft, and then injected GFI1 specific- or control morpholinos. While mice treated with control morpholino died of leukemia, the animals treated with GFI1-specific morpholinos survived showing a significant reduction of human leukemic cells in the blood, bone marrow and spleen, even with samples from patients who did not respond to first line therapy. Since morpholinos have received approval for use in humans, our data suggest that targeting GFI1 in human T-ALL patients may be a promising therapeutic target and a feasible way to complement current T-ALL treatment regimens. Disclosures: No relevant conflicts of interest to declare.

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Tauroursodeoxycholate protects from glycochenodeoxycholate-induced gene expression changes in perfused rat liver

Biological Chemistry ◽

10.1515/hsz-2019-0204 ◽

2019 ◽

Vol 400 (12) ◽

pp. 1551-1565

Author(s):

Martha Paluschinski ◽

Mirco Castoldi ◽

David Schöler ◽

Nils Bardeck ◽

Jessica Oenarto ◽

...

Keyword(s):

Gene Expression ◽

Rat Liver ◽

Target Genes ◽

Quantitative Polymerase Chain Reaction ◽

Gene Array ◽

Surrogate Marker ◽

Rat Hepatocytes ◽

Protective Effects ◽

Induced Apoptosis ◽

P53 Target Genes

Abstract Tauroursodeoxycholate (TUDC) is well known to protect against glycochenodeoxycholate (GCDC)-induced apoptosis in rat hepatocytes. In the present study, we analyzed whether TUDC also exerts protective effects by modulating GCDC-induced gene expression changes. For this, gene array-based transcriptome analysis and quantitative polymerase chain reaction (qPCR) were performed on RNA isolated from rat livers perfused with GCDC, TUDC or a combination of both (each 20 μm for 2 h). GCDC led to a significant increase of lactate dehydrogenase (LDH) into the effluent perfusate, which was prevented by TUDC. GCDC, TUDC and co-perfusion induced distinct gene expression changes. While GCDC upregulated the expression of several pro-inflammatory genes, co-perfusion with TUDC increased the expression of pro-proliferative and anti-apoptotic p53 target genes. In line with this, levels of serine20-phosphorylated p53 and of its target gene p21 were elevated by GCDC in a TUDC-sensitive way. GCDC upregulated the oxidative stress surrogate marker 8OH(d)G and the pro-apoptotic microRNAs miR-15b/16 and these effects were prevented by TUDC. The upregulation of miR-15b and miR-16 in GCDC-perfused livers was accompanied by a downregulation of several potential miR-15b and miR-16 target genes. The present study identified changes in the transcriptome of the rat liver which suggest, that TUDC is hepatoprotective by counteracting GCDC-induced gene expression changes.

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381: Promoter Hypermethylation Profile of Kidney Cancer with New Pro-Apoptotic P53 Target Genes and Possible Clinical Implications

The Journal of Urology ◽

10.1016/s0022-5347(18)32637-5 ◽

2006 ◽

Vol 175 (4S) ◽

pp. 125-125

Author(s):

Frank Christoph ◽

Steffen Weikert ◽

Carsten Kempkensteffen ◽

Martin Schostak ◽

Hans Krause ◽

...

Keyword(s):

Kidney Cancer ◽

Target Genes ◽

Promoter Hypermethylation ◽

Clinical Implications ◽

P53 Target Genes

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Differential Activation of p53 Target Genes in Breast Cancer

10.21236/ada405348 ◽

2002 ◽

Author(s):

Edward Thornborrow ◽

James Manfredi

Keyword(s):

Breast Cancer ◽

Target Genes ◽

Differential Activation ◽

P53 Target Genes

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Synergistic Antitumor Effect of 5-Fluorouracil Combined with Constituents from Pleurospermum lindleyanum in Hepatocellular carcinoma SMMC-7721 Cells

Anti-Cancer Agents in Medicinal Chemistry ◽

10.2174/1871520620666200824094624 ◽

2020 ◽

Vol 20 ◽

Author(s):

Xiao-Feng Zhu ◽

Xiao-Jin Li ◽

Zhong-Lian Cao ◽

Xiu-Jie Liu ◽

Ping Yang ◽

...

Keyword(s):

Hepatocellular Carcinoma ◽

Treatment Effectiveness ◽

Synergistic Effects ◽

Folk Medicine ◽

Inhibitory Effect ◽

Caspase 9 ◽

Whole Plant ◽

Anti Cancer ◽

Induced Apoptosis

Background: A Chinese folk medicine plant Pleurospermum lindleyanum possesses pharmacological activities of heat-clearing, detoxifying and preventing from hepatopathy, coronary heart disease, hypertension, and high altitude sickness. We isolated and characterized its constituents to investigate its synergistic effects against human hepatoma SMMC-7721 cells. Objective: The aim of this study was to explore the synergistic anti-cancer activities of isolates from P. lindleyanum with 5-FU on hepatoma SMMC-7721 cells in vitro and their primary mechanisms. Methods: Sequential chromatographic techniques were conducted for the isolation studies. The isolates structures were established by spectroscopic analysis as well as X-ray crystallographic diffraction. Growth inhibition was detected by MTT assay. The isobologram method was used to assess the effect of drug combinations. Flow cytometry and western blot were used to examine apoptosis and protein expression. Results: A new coumarin (16), along with sixteen known compounds, were isolated from the whole plant of P. lindleyanum and their structures were elucidated by spectroscopic methods. Four coumarins (2, 3, 5, and 16), two flavonoids (8 and 9) and three phytosterols and triterpenes (12-14) were found to synergistically enhance the inhibitory effect of 5-FU against SMMC-7721 cells. Among them, compounds 3 and 16 exhibited the best synergistic effects with IC50 of 5-FU reduced by 16-fold and 22-fold possessing the minimum Combination Index (CI) 0.34 and 0.27. The mechanism of action of combinations might be through synergistic arresting for the cell cycle at G1 phases and the induction of apoptosis. Moreover, western blotting and molecular docking revealed that compounds 3 or 5 might promote 5-FU-induced apoptosis by regulating the expression of Caspase 9 and PARP. Conclusion: Constituents from P. lindleyanum may improve the treatment effectiveness of 5-FU against hepatocellular carcinoma cells.

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