Unique induction of p21(WAF1/CIP1)expression by vinorelbine in androgen-independent prostate cancer cells

To study the mechanisms of the development of hormone refractory prostate cancer, we established an androgen-independent (AI) prostate cancer cell line derived from hormone-dependent (AD) LNCaP cells. Our previous studies have demonstrated that AI cells are deficient in expression of p21(WAFl/CIP1) (p21) due to overexpressed AR and are resistant to apoptosis. In this study, the induction of p53 and p21 expression by vinorelbine (Navelbine) was compared between AD and AI cells in an attempt to understand the difference(s) in apoptotic signalling pathways in these cells. Using a series of deletion of p21 reporter constructs, we found that vinorelbine mediated p21 induction in a p53-dependent manner in AD cells. In contrast, p21 expression restored by vinorelbine in AI cells was found to be through both p53-dependent and-independent pathways. In the absence of two p53 binding sites, Spl-3 and Spl-4 sites, in the promoter of human p21 gene, were found to be required for vinorelbine-mediated p21 activation. No p21 induction was observed by paclitaxel in AI cells. Exposure of AI cells to paciltaxel followed by vinorelbine produced synergism. Our data, thus, provide a basis for the synergistic combination of vinorelbine and paclitaxel for the treatment of advanced prostate cancer.     

Differential targeting of the cyclin-dependent kinase inhibitor,p21CIP1/WAF1, by chelators with anti-proliferative activity in a range of tumor cell-types

Chelators such as 2-hydroxy-1-napthylaldehyde isonicotinoyl hydrazone (311) and di-2-pyridylketone-4,4-dimethyl-3-thiosemicarbazone (Dp44mT) target tumor cell iron pools and inhibit proliferation. These agents also modulate multiple targets, one of which is the cyclin-dependent kinase inhibitor, p21. Hence, this investigation examined the mechanism of action of these compounds in targeting p21. All the chelators up-regulated p21 mRNA in the five tumor cell-types assessed. In contrast, examining their effect on total p21 protein levels, these agents induced either: (1) down-regulation in MCF-7 cells; (2) up-regulation in SK-MEL-28 and CFPAC-1 cells; or (3) had no effect in LNCaP and SK-N-MC cells. The nuclear localization of p21 was also differentially affected by the ligands depending upon the cell-type, with it being decreased in MCF-7 cells, but increased in SK-MEL-28 and CFPAC-1 cells. Further studies assessing the mechanisms responsible for these effects demonstrated that p21 expression was not correlated with p53 status, suggesting a p53-independent mechanism. Considering this, we examined proteins that modulate p21 independently of p53, namely NDRG1, MDM2 and ΔNp63. These studies demonstrated that a dominant negative MDM2 isoform (p75^MDM2) closely resembled p21 expression in response to chelation in three cell lines. These data suggest MDM2 may be involved in the regulation of p21 by chelators.     

Reduced p21(WAF1/CIP1) protein expression is predominantly related to altered p53 in hepatocellular carcinomas

To investigate the relationship between the expression of p21(WAF1/CIP1) protein and p53 status and the possible role of the two proteins in hepatocellular carcinomas (HCCs), we examined the expression of p21(WAF1/CIP1) and p53 immunohistochemically in 81 tumours from 65 patients with hepatocellular carcinoma. p21(WAF1/CIP1) protein was absent from 59 of 81 tumours (72.8%), and altered p53 expression was found in 43 (53.1%). p21(WAF1/CIP1) expression was significantly associated with p53 status (P = 0.0008); 38 of 59 tumours lacking p21(WAF1/CIP1) protein were accompanied by altered p53 expression. Further analyses showed that p21(WAF1/CIP1) expression was inversely correlated with p53 expression in hepatitis C virus (HCV)-related HCCs, but not in HBV-related hepatocellular carcinomas and hepatocellular carcinomas without viral infection. All 11 tumours with intrahepatic metastasis showed altered p21(WAF1/CIP1) or p53 expression. In contrast, no intrahepatic metastasis was found in any of the 17 tumours without abnormal expression of either of the two proteins. These results suggest that: (1) different modes of p21(WAF1/CIP1) regulation are involved in HCCs differing in their hepatitis viral infection status, and p21(WAF1/CIP1) expression appears to be predominantly related to altered p53 in HCV-related HCCs; (2) disruption of the p53-p21(WAF1/CIP1) cell-cycle-regulating pathway may contribute to malignant progression of HCC.     

硒对乳腺癌细胞株p21^WAF1/CIP1启动子的调控作用

目的:研究硒对p21的转录调控及其调控位点.方法:通过向转染了重组质粒pGL3- p21p的乳腺癌细胞株MCF7先后加入不同的p21因子启动子的负调节因子和乳酸硒,对比分析荧光素酶表达活性,以确定硒对p21的转录调控及调控位点,并验证硒对癌细胞的生长的负调控作用.结果:perifosine、depsipeptide、apicidin、butyrate与硒共同诱导荧光素酶,酶活性表达无显著差异;而C-Myc与醋酸硒先后诱导酶活性表达差异显著.结论:硒对癌细胞具有诱导凋亡的作用,转录调节位点在p21启动子的sp1结合位点.     

p21^WAF1／CIP1在原发性肺癌中的表达

目的：探讨p21^WAF1/CIP1在肺癌中的生物学功能,方法：用免疫组化法检测62例肺癌和14例正常肺组织石蜡切片p21^WAF1/CIP1的染色强度。结果：1．p21^WAF1/CIP1定位于细胞核或细胞浆。2．小细胞肺癌是与非小细胞肺中核p21^WAF1/CIP1表达存在显著性差异（P＜0．005）,浆p21^WAF1/CIP1则不存在显著性差异（P＜0．005）。核与浆p21^WAF1/CIP1表达的对比在高,低分化肺癌中具有显著性差异P＜0．05）。结论：肺癌细胞p21^WAF1/CIP1的生物学功能可能依其定位、量表达的高低及组织学类型的不同而存在差异。     

胃肠道类癌中生长抑素和p21^WAF1/CIP1蛋白表达的意义

目的探讨生长抑素和p21WAF1/CIP1蛋白阳性表达与胃肠道类癌的组织分化、浸润和转移的关系.方法采用免疫组化S-P法对36例胃肠道类癌组织生长抑素和p21WAF1/CIP1蛋白的表达进行检测.结果 36例类癌组织中,生长抑素和p21WAF1/CIP1蛋白较多表达于高分化类癌组(P＜0.05),随着肿瘤的浸润和淋巴结转移,生长抑素阳性表达率显著降低(P＜0.01),p21WAF1/CIP1阳性表达差异有显著性(P＜0.05).结论生长抑素和p21WAF1/CIP1低表达在类癌的组织分化和发展中起着重要作用,可用于临床对患者进行预后判断.     

人脑胶质瘤组织中p21WAF1/CIP1表达及临床意义

[目的]探讨人脑胶质瘤组织中p21WAF1/CIP1基因蛋白表达水平与人脑胶质瘤恶性程度的关系。[方法]随机选取的Ⅰ￣Ⅳ人脑胶质瘤标本48例,正常外伤脑组织10例为对照。应用免疫组化技术(SP法)检测p21WAF1/CIP1基因蛋白在人脑胶质瘤中的表达水平。[结果]p21WAF1/CIP1基因蛋白在正常脑组织中均为阴性表达,而在胶质细胞瘤组织中表达增高,阳性率为70%￣78%,在Ⅰ～Ⅳ级胶质瘤组织中表达数值分别为2.11±0.10,1.44±0.56,1.0±0.12及0.89±0.32,表达水平随胶质瘤恶性程度的升高呈下降趋势,在高分化与低分化肿瘤之间存在显著性差异(P<0.05)。[结论]p21WAF1/CIP1可能参与胶质瘤的发生和发展,并可作为评估胶质瘤细胞瘤恶性程度以及预后的手段之一。     

MiR-224 promotes the chemoresistance of human lung adenocarcinoma cells to cisplatin via regulating G1/S transition and apoptosis by targeting p21WAF1/CIP1

Background:

Increasing evidence has shown that microRNAs (miRNAs) can serve as oncogenes and tumour suppressors to participate in tumour development. However, the roles of miRNAs in chemoresistance of human lung adenocarcinoma (LA) remain largely undefined.

Methods:

On the basis of miRNA microarray data, miR-224 was identified as the most upregulated miRNA in cisplatin (DDP; cis-diamminedichloroplatinum II)-resistant A549 cells compared with parental A549 cells. The aim of our study was to investigate the roles of miR-224 in the formation of DDP-resistant phenotype of LA cells and its possible molecular mechanisms.

Results:

Here we showed that miR-224 could promote the in vitro and in vivo DDP resistance of LA cells via regulating G₁/S cell cycle transition and apoptosis. p21^WAF1/CIP1, a potent cyclin-dependent kinase inhibitor, was identified as the direct and functional target gene of miR-224. Overexpression of p21^WAF1/CIP1 could phenocopy the effect of miR-224 downregulation and silencing of p21^WAF1/CIP1 could partially reverse the effect of miR-224 downregulation on DDP resistance of DDP-resistant LA cells. In addition, miR-224 could affect the G₁/S transition of cell cycle and apoptosis in LA cells through the p21^WAF1/CIP1-pRb pathway and the intrinsic mitochondrial death pathway. Furthermore, miR-224 was found to be downregulated in DDP-responding LA tissues, and its expression was inversely correlated with p21^WAF1/CIP1. Multivariate analyses indicated that the status of miR-224 might be an independent prognostic factor for predicting the survival of LA patients.

Conclusions:

Our findings shed novel light on the roles of miR-224/p21^WAF1/CIP1 signalling in the DDP resistance of LA cells, and targeting it will be a potential strategic approach for reversing the DDP resistance in human LAs.     

Prognostic Significance of p53 and p21WAF1/CIP1 Immunoreactivity and Tumor Micronecrosis for Recurrence of Meningiomas

Recurrence is an important factor for prognosis of meningioma patients, this also occurring with some lesions diagnosed histopathologically as benign. To analyze their relationships with clinicopathological factors, p53 and p21^WAF1/CIP1 immunoreactivity, 80 meningiomas were classified into four groups with regard to the World Health Organization (WHO) histological classification and recurrence: 40 cases of Group I (typical type)-NR (no recurrence); five cases of Group I-R (recurrence); 20 cases of Group II (atypical or anaplastic type)-NR and 15 cases of Group II-R.Micronecrosis was detected in 25% of Group II-NR and 73.3% of Group II-R (P=0.007, odds ratio (OR) =8.25, 95% confidence interval (CI) =1.79–38.01). Patients receiving radiation therapy had a lower risk of recurrence (P=0.041, OR =0.20, 95% CI =0.05–0.85). Immunoreactivity for p53 protein was positive in 22% of Group I and 54% or Group II (P=0.005), and in 80% of Group I-R and 15% of Group I-NR (P=0.006, OR = 22.7, 95% CI = 2.15–239.4). p21^WAF1/CIP1 protein was detected in 22% of Group I and 48% of Group II (P=0.017), but with no link to recurrence. Multivariate analysis also showed p53 immunoreactivity in Group I (benign lesions) and micronecrosis in Group II (atypical/anaplastic meningiomas) to be strong prognostic factors for recurrence (P<0.05). These results indicate that p53 immunoreactivity and micronecrosis can help predicting recurrence of meningiomas.     

Induction of p53 and p21/WAF1/CIP1 expression by nitric oxide and their association with apoptosis in human cancer cells

In this study, human and rat cancer cells were used to investigate the expression of p53 and p21/WAF1/CIP1 and their association with apoptosis after exposure to nitric oxide (NO). It was found that NO induced nuclear accumulation of p53 protein in a dose- and time-dependent manner. The level of p53 protein was elevated by about fivefold compared with that of mock-treated cells 48 h after exposure to 300 ppm NO. The induction of p53 by NO was found by pulse-chase analysis to be mainly regulated by post-translational modification. The correlation between p53 status and apoptosis induced by NO in human cancer cells was also investigated in this study. We found that apoptosis was easily induced in cells containing wild-type p53 (COLO 205 and Hep G2) after exposure to NO. The p21/WAF1/CIP1 protein was induced by NO in cells containing wild-type p53 (Hep G2) but not in cells without p53 (Hep 3B) or with mutated p53 (HT-29). Our results indicate that wild-type p53 and p21/WAF1/CIP1 expression was elevated in human cancer cells by exposure to NO and suggest that this may eventually promote apoptosis. © 1996 Wiley-Liss, Inc.     

p21~(WAF1/CIP1)在乳腺癌中的表达及意义

目的　探讨p2 1WAF1/CIP1蛋白在乳腺癌中表达的临床意义。方法　运用免疫组化SP法半定量检测p2 1蛋白在癌旁正常乳腺组织、乳腺癌组织中的表达。结果　p2 1蛋白表达位于细胞核 ,呈棕黄色。在 2 0例癌旁正常乳腺组织中 ,无p2 1蛋白表达。在 69例乳腺癌组织中有 3 0例p2 1蛋白阳性表达。在乳腺癌组织中 ,随组织学分级升高 ,p2 1阳性率下降 (P <0 0 5 ) ,随临床分期升高 ,p2 1阳性率下降 (P <0 0 5 )。有淋巴结转移组p2 1阳性率低于无淋巴结转移组 (P <0 0 5 )。p2 1蛋白阳性表达者术后 5年无瘤生存率高于p2 1蛋白阴性者术后 5年无瘤生存率 (P <0 0 5 )。结论　p2 1蛋白可用来评估乳腺癌细胞分化情况及转移潜能 ,可判断乳腺癌患者预后。     

p21WAF1/CIP1和p53蛋白表达在胃癌发生发展过程中的研究

目的研究p21WAF1/CIP1和p53蛋白在胃癌发生发展过程中的作用及表达的临床病理意义.方法采用免疫组化SP法对正常胃粘膜、萎缩性胃炎伴肠上皮化生、萎缩性胃炎伴不典型增生组织各20例和78例胃癌组织标本进行p21WAF1/CIP1和p53蛋白检测.结果胃癌组织中p53蛋白阳性表达率高于正常胃粘膜、萎缩性胃炎伴肠上皮化生和不典型增生组(P＜0.05),而p21WAF1/CIP1蛋白阳性表达低于正常胃粘膜、萎缩性胃炎伴肠上皮化生组(P＜0.01)p21WAF1/CIP1、p53蛋白表达与胃癌的分化程度相关(P＜0.05);有淋巴结转移组p21WAF1/CIP1蛋白表达率低于无淋巴结转移组(P＜0.05),而有淋巴结转移组p53蛋白表达率高于无淋巴结转移组(P＜0.05);p53蛋白表达与胃癌浸润深度有关.结论p53蛋白高表达与p21WAF1/CIP1蛋白失表达可能参与胃癌的发生发展过程;检测p53和p21WAF1/CIP1蛋白作为反映胃癌病理学特点的参考指标可能有一定意义;p21WAF1/CIP1蛋白表达在胃癌可能存在非p53诱导表达途径.     

子宫内膜癌p21WAF1/CIP1蛋白的表达及其意义

背景与目的:探讨p21WAF1/CIP1蛋白在子宫内膜癌中的表达情况及其意义.材料与方法:应用免疫组织化学SP法检测30例正常子宫内膜,20例单纯性增生性宫内膜,22例不典型增生性宫内膜,57例子宫内膜癌(高分化32例,中分化12例,低分化13例)中p21WAF1/CIP1蛋白的表达. 结果:p21WAF1/CIP1蛋白表达于细胞核,子宫内膜癌中p21WAF1/CIP1蛋白的表达明显低于正常子宫内膜和单纯性增生性子宫内膜(P＜0.01),且p21WAF1/CIP1蛋白的表达与子宫内膜癌组织有无肌层浸润及临床分期明显相关(P均＜0.05).结论:p21WAF1/CIP1蛋白表达降低可能在子宫内膜癌的发生发展及判断预后方面具有重要作用.     

Asiatic Acid Promotes p21WAF1/CIP1 Protein Stability through Attenuation of NDR1/2 Dependent Phosphorylation of p21WAF1/CIP1 in HepG2 Human Hepatoma Cells

Previous studies have suggested anti-tumor effects of asiatic acid in some human cancer cell lines. This agent isreported to increase the levels of p21WAF1/CIP1 in human breast cancer cell lines. However, the molecular mechanismshave not been established. Here we report that asiatic acid up-regulates p21WAF1/CIP1 protein expression but notthe level of p21WAF1/CIP1 mRNA in HepG2 human hepatoma cells. Furthermore, we found that the asiatic acidinduced increase of p21WAF1/CIP1 protein was associated with decreased phosphorylation (ser-146) of p21WAF1/CIP1.Knockdown of NDR1/2 kinase, which directly phosphorylates p21WAF1/CIP1 protein at ser-146 and enhances itsproteasomal degradation, increased the levels of p21WAF1/CIP1 protein and eliminated the regulation of p21WAF1/CIP1 stability by asiatic acid. At the same time, the expression of NDR1/2 kinase decreased during treatment withasiatic acid in HepG2 cells. Moreover, asiatic acid inhibited the proliferation of HepG2 cells, this being attenuatedby knockdown of p21WAF1/CIP1. In conclusion, we propose that asiatic acid inhibits the expression NDR1/2 kinaseand promotes the stability of p21WAF1/CIP1 protein through attenuating NDR1/2 dependent phosphorylation ofp21WAF1/CIP1 in HepG2 cells.     

p21WAF1/CIP1基因的转录调控机制及其与肿瘤的关系

 p21WAF1/ CIP1基因参与多条信号通路，在细胞周期、细胞分化及凋亡等重要的细胞活动中具有重要作用。文章对p21WAF1/CIP1基因转录调控机制及其与肿瘤的关系进行阐述，以进一步明确其在肿瘤诊断、治疗和预后评价中的应用价值。     

Leptin-induced growth of human ZR-75-1 breast cancer cells is associated with up-regulation of cyclin D1 and c-Myc and down-regulation of tumor suppressor p53 and p21WAF1/CIP1

Summary Obesity has been recognized as a risk factor for breast cancer. Adipocyte-derived leptin may play as a paracrine regulator on the growth of breast cancer cells. Expression of both leptin and its OB-Rb receptor was detected in human breast cancer ZR-75-1 cells and further induced by leptin, suggesting that both expression and message mediation of leptin were autoregulated by itself. With cell counting and MTT assay, we had observed leptin stimulated ZR-75-1 growth in dose- and time-dependent manners. To study what steps of cell cycle progression leptin may involve in, we analyzed cell-cycle profile with flow cytometric analysis, mRNA and protein expressions of four cell-cycle regulators with RT-PCR and Western blotting analysis. Under the treatment of leptin, the G1 arrest of cells was reduced accompanied with up-regulation of G1 phase-specific cyclin D1 and proto-oncogene c-Myc, but down-regulation of cyclin-dependent kinase inhibitor p21^WAF1/CIP1 and tumor suppressor p53. Furthermore, JAK2 inhibitor AG490, PI3K/Akt inhibitor Wortmannin, and MEK/ERK1/2 inhibitor PD98059 were efficiently prevented leptin-promoted cell growth. Effect of cooperation between leptin and estrogen on ZR-75-1 growth had been observed. Collectively, the results showed that the proliferative effect of leptin on ZR-75-1 was associated with the up-regulation of cyclin D1 and c-Myc and down-regulation of tumor suppressor p53 and p21^WAF1/CIP1 plausibly through a hypothesized JAK2-PI3K/Akt-MEK/ERK pathway. The leptin- and OB-Rb-expressing capability of ZR-75-1 created a possible autocrine control of leptin, in which signal could be effectively amplified by itself, on cell growth.