Expression of p21(WAF1) and p53 and polymorphism of p21(WAF1) gene in gastric carcinoma

AIM: To investigate the relationship between expression of p21(WAF1) and p53 gene, and to evaluate the deletion and polymorphism of p21(WAF1) gene in gastric carcinoma (GC). METHODS: Expression of p21 and p53 proteins, and deletion and polymorphism of p21 gene in GC were examined by streptavidin-peroxidase conjugated method (SP) and polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) respectively. RESULTS: The expression of p21 and p53 was found in 100% (20/20) and 0% (0/20) of normal gastric mucosae(NGM), 92.5% (37/40) and 15.0% (6/40) of dysplasia (DP) and 39.8% (43/108) and 56.5% (61/108) of GC, respectively. The positive rate of p21 in GC was lower than that in NGM and DP (P<0.05), while the positive rate of p53 in GC was higher than that in NGM and DP (P<0.05). p21 and p53 were significantly expressed in 63.3% (19/30) and 36.7% (11/30), 35.0% (14/40) and 77.5% (31/40), 26.7% (4/15) and 80.0% (12/15), 30.8% (4/13) and 30.8% (4/13), and 20.0% (2/10) and 30.0% (3/10) of well-differentiated, poorly-differentiated, undifferentiated carcinomas, mucoid carcinomas and signet ring cell carcinomas. The expression of p21 in well-differentiated carcinomas was significantly higher than that in poorly-differentiated, un-differentiated, mucoid carcinomas and signet ring cell carcinomas (P<0.05). Contrarily, The expression of p53 was increased from well-differentiated to poorly-differentiated and un-differentiated carcinomas (P<0.05). The expression of p21 and p53 in paired primary and metastatic GC (35.3% and 70.6%) was different from non-metastatic GC (62.5% and 42.5%) markedly (P<0.05). The expression of p21 in invasive superficial muscle (60.0%) was higher than that in invasive deep muscle or total layer (35.2%) (P<0.05) and was higher in TNM stages I (60.0%) and II (56.2%) than in stages III (27.9%) and IV (22.2%) (P<0.05), whereas the expression of p53 did not correlate to invasion depth or TNM staging (P>0.05). The exoression patterns of p53+/p21-, and of p53-/p21+ were found in 5.0% and 82.5% of DP. There was a significant correlation between expression of p21 and p53 (P<0.05). But there was no significant correlation between expression of both in GC (P>0.05). There was no deletion in exon 2 of p21 gene in 30 cases of GC and 45 cases of non-GC, but polymorphism of p21 gene at exon 2 was found in 26.7% (8/30) of GC and 8.9% (4/45) of non-GC, a significant difference was found between GC and non-GC (P<0.05). There was no significant relation between p21 expression of polymorphism (37.5%, 3/8) and non-polymorphism (45.5%, 10/22) in GC (P>0.05). CONCLUSION: The loss of p21 protein and abnormal expression of p53 are related to carcinogenesis, differentiation and metastasis of GC. The expression of p21 is related to invasion and clinical staging in GC intimately. The expression of p21 protein depends on p53 protein largely in NGM and DP, but not in GC. No deletion of p21 gene in exon 2 can be found in GC. The polymorphism of p21 gene might be involved in gastric carcinogenesis.There is no significant association between polymorphism of p21 gene and expression of p21 protein.     

ATP-dependent activation of p21WAF1/CIP1-associated Cdk2 by Cdc6

When cells progressing in mid-S phase are damaged with a base-modifying chemical, they arrest in S phase long after the CHK1 checkpoint signal fades out, partly because of p53-mediated long-lasting induction of the cyclin-dependent kinase inhibitor p21(WAF1/CIP1). We have recently found that enforced expression of Cdc6, the assembler of prereplicative complexes, markedly advances recovery from the prolonged S-phase arrest and reactivation of Cdk2 despite the presence of a high level of induced p21. Here, we report that Cdc6 protein can activate p21-associated Cdk2 in an ATP-dependent manner in vitro. Consistently, Cdc6 mutated for ATPase or a putative cyclin binding motif is no longer able to activate the Cdk2 in vitro or promote reinitiation of S-phase progression and reactivation of Cdk2 in vivo. These results reveal the never anticipated function of Cdc6 and redefine its role in the control of S-phase progression in mammalian cells.     

结直肠癌中CDX2、p21~(H-ras)、p21~(WAF1)表达的临床意义

背景:CDX2与肠源性肿瘤的发生密切相关,而p21表达减低或缺失可能是结直肠癌发生、发展中的一个普遍事件。目的:探讨结直肠癌中CDX2、p21~(H-ras)、p21~(WAF1)表达的相关性和临床病理意义。方法:收集45例临床病理资料完整的结直肠癌手术切除标本,以免疫组化方法检测癌组织和相应癌旁组织中的CDX2、p21~(H-ras)、p21~(WAF1)蛋白表达。结果:结直肠癌组织中CDX2、p21~(H-ras)、p21~(WAF1)阳性率分别为86.7%、68.9%和35.6%,相应癌旁组织中阳性率分别为100.0%、37.8%和51.1%,CDX2和p21~(H-ras)在癌组织和癌旁组织中的阳性率差异有统计学意义(P=0.026和P=0.006);半定量分析结果显示CDX2和p21~(WAF1)在癌组织和癌旁组织中的表达水平差异有统计学意义(P=0.007和P=0.005)。癌组织中CDX2与p21~(H-ras)的表达存在相关性(r_s=0.501,P=0.000)。Dukes A、B期患者癌组织p21~(WAF1)阳性率显著高于C、D期患者(P=0.016),有淋巴结转移者p21~(WAF1)阳性率显著低于无淋巴结转移者(P=0.048),CDX2、p21~(H-ras)的表达与结直肠癌各临床病理特征均无相关性。结论:CDX2、p21~(H-ras)、p21~(WAF1)表达改变可能与结直肠癌的发生、发展相关。     

KLF6、p21^WAF1/CIP1、CyclinD1在结直肠癌中的表达及意义

目的研究转录因子KLF6、p21WAF1/C IP1及Cyc linD1在结直肠癌中的表达及意义。方法应用免疫组化Envision法对58例结直肠癌组织、20例结直肠黏膜慢性炎症组织中的KLF6、p21WAF1/C IP1及Cyc linD1蛋白表达进行检测。结果结直肠癌组织KLF6、p21WAF1/C IP1及Cyc linD1蛋白表达率均与结直肠黏膜慢性炎症组织比较有统计学差异（P〈0.05）;KLF6、p21WAF1/C IP1及Cyc linD1蛋白在结直肠癌组织中的表达与其浸润深度及预后有关（P〈0.05）。结论 KLF6、p21WAF1/C IP1及Cyc linD1在结直肠癌的发生发展中可能起重要作用。     

Plk1与p21WAF1/CIP1在肝细胞癌中的表达

目的 细胞周期抑制因子p21<'WAF1/CIP1>(p21)可在转录水平抑制Polo-like kinase1(Plk1)基因的表达,本文旨在探讨Plk1和p21蛋白在肝细胞癌中的表达及相关性.方法 应用Westem Blot方法检测10对原发性肝细胞癌和癌旁组织中Plk1和p21蛋白的表达情况.通过将pFlex-21表达质粒瞬时转染人肝癌细胞系HepG2细胞,进一步检测p21过表达对Plk1mRNA和蛋白表达的影响.结果 Plk1在10例肝癌组织中的表达均高于配对癌旁组织,p21蛋白在7对肝癌组织中的表达高于配对癌旁组织.HepG2细胞瞬时表达p21质粒后,Plk1的蛋白表达和mRNA水平均无变化(P>0.05).结论 Plk1和p21在肝细胞癌组织中的表达具有一定的肿瘤特异性.肝癌组织中p21蛋白过表达可能不具有抑制Plk1表达的作用,具体机制还有待于进一步研究.     

Cell cycle inhibition by the anti-angiogenic agent TNP-470 is mediated by p53 and p21WAF1/CIP1

Angiogenesis has been demonstrated to be essential for tumor growth and metastasis, and inhibition of angiogenesis is emerging as a promising strategy for treating cancer. Among the most potent inhibitors of angiogenesis is the fumagillin family of natural products. An analog of fumagillin, known as TNP-470 or AGM-1470, has been undergoing clinical trials for treating a variety of cancers. TNP-470 has been shown to block endothelial cell cycle progression in the late G(1) phase. Although the direct molecular target for TNP-470 has been identified as the type 2 methionine aminopeptidase (MetAP2), how inhibition of this enzyme leads to cell cycle arrest has remained unclear. We report that treatment of endothelial and other drug-sensitive cell types leads to the activation of the p53 pathway, causing an accumulation of the G(1) cyclin-dependent kinase inhibitor p21(WAF1/CIP1). The requirement of p53 and p21(WAF1/CIP1) for the cell cycle inhibition by TNP-470 is underscored by the observation that cells deficient in p53 and p21(WAF1/CIP1) are resistant to TNP-470. These results shed significant light on the mechanism of cell cycle inhibition by TNP-470 and suggest an alternative method of activating p53 in endothelial cells to halt angiogenesis and tumor progression.     

p21WAF1 protein expression determined by quantitative immunoassay in relation to non-small-cell lung cancer aggressiveness

Purpose: p21WAF1, a cyclin-dependent kinase inhibitor, is an important mediator of the cell-cycle arrest and tumor suppression induced by the protein p53. Although alterations of the p53 gene and its overexpression are frequent in most malignancies, including non-small-cell lung cancer (NSCLC), and may be associated with poor patient prognosis, the clinical utility of p21WAF1 expression in NSCLC has not been established. Methods: We have used a commercial enzyme-linked immunosorbent assay (ELISA) kit for p21WAF1 to test soluble extracts of 54 NSCLC specimens with known clinicopathological properties. Results: There was no correlation between p21WAF1 and p53 concentrations, the latter being determined by a time-resolved immunofluorometric assay developed in-house. Furthermore, p21WAF1 levels were not associated with patient age, tumor/node/metastasis (TNM) stage, lymph node metastasis, histological grade or type, or smoking history, in Mann-Whitney analysis. χ²-tests, based on cutoffs equal to the 25th, 50th, or 75th percentiles of the p21WAF1 distribution, similarly did not reveal any statistically significant associations between p21WAF1 and other clinicopathological variables. Because of the small number of patients and the median follow-up of only 18 months, a meaningful survival analysis could not be performed. Conclusion: In summary, this preliminary study suggests that ELISA-quantified p21WAF1 levels in NSCLC extracts are weaker than p53 in terms of prognostic value and do not contribute to the further subclassification of patients. Received: 22 April 1999 / Accepted: 13 July 1999     

Plkl与p21~（WAF1/CIP1）在肝细胞癌中的表达

目的细胞周期抑制因子p21WAF1/CIP1（p21）可在转录水平抑制Polo-like kinase1（Plk1）基因的表达,本文旨在探讨Plk1和p21蛋白在肝细胞癌中的表达及相关性。方法应用Western Blot方法检测10对原发性肝细胞癌和癌旁组织中Plk1和p21蛋白的表达情况。通过将pFlex-p21表达质粒瞬时转染人肝癌细胞系HepG2细胞,进一步检测p21过表达对Plk1mRNA和蛋白表达的影响。结果 Plk1在10例肝癌组织中的表达均高于配对癌旁组织,p21蛋白在7对肝癌组织中的表达高于配对癌旁组织。HepG2细胞瞬时表达p21质粒后,Plk1的蛋白表达和mRNA水平均无变化（P〉0.05）。结论 Plk1和p21在肝细胞癌组织中的表达具有一定的肿瘤特异性。肝癌组织中p21蛋白过表达可能不具有抑制Plk1表达的作用,具体机制还有待于进一步研究。     

Expression and prognostic relevance of p21WAF1 in stage III esophageal squamous cell carcinoma

The prognostic effect of p21^WAF1 expression on esophageal squamous cell carcinoma patients is controversial. Further clarifying the effect of this protein is beneficial for optimizing the patient outcomes. In the current study, we investigated the expression of p21^WAF1 protein in 189 specimens of stage III ESCC by immunohistochemistry. As shown by the Kaplan–Meier curve, the overall survival rate of the positive‐expression group was significantly higher than that of the negative‐expression group (P < 0.05). No significant correlation was observed between p21^WAF1 expression and clinicopathological parameters in terms of gender, age, tumor location, tumor grade, pathological stage, and number of regional lymph node metastases (P > 0.05). We concluded that p21^WAF1 played an intricate role in the tumorigenesis and development of ESCC. p21^WAF1 could serve as a positive prognostic predictor for stage III ESCC patients.     

HCC和HepG2细胞表达HCV C蛋白、p14ARF、p21WAF1的意义探讨

目的检测HCVC蛋白、p14、p21在HCC和表达野生p53HepG2中的表达，初步探讨C蛋白在HCC和HepG2中对p14-p53-p21凋亡通路的作用。方法收集42例HCC石蜡组织，采用免疫组织化学EnVision法检测HCC组织中核心蛋白、p14和p21的表达，用统计学方法及临床联系分析它们之间的关系；用细胞化学EnVision法和免疫荧光法检测核心蛋白、p53、p14和p21在HepG2细胞中的表达。结果C蛋白、p14和p21的阳性表达主要定位于细胞核膜和细胞核中；HCC组织中C蛋白、p14和p21阳性率分别为40．5％、45．24％、19．05％；3组间的Kruskal-Wallis检验P=0．03，差异显著；C蛋白与p14、p21间及p14与D21间蛋白阳性强度相关性分析显示，P值分别为0．000、0．43、-0．34，相关系数rs分别为0．64、-0．29、-0．33。HepG2细胞有较高的C蛋白和p53表达及少量的p14、p21蛋白表达。结论在C蛋白阳性的HCC中p14的表达与C蛋白有关，HCC中D21表达缺陷是十分常见的；C蛋白在HCC中可能影响p53通路，下调p21的表达，阻止其凋亡作用；HepG2细胞永生化特性可能与HCV或HCVC蛋白有关。     

Role of p53 and p21/WAF1 detection in patient selection for preoperative radiotherapy in rectal cancer patients

BACKGROUND: Recent studies showed that p53 and p21 may play major roles in determining tumor radiosensitivity through the apoptosis pathway. The aim of this study was to investigate the predicting value of radiosensitivity in human rectal carcinoma. METHODS: p53 and p21/WAF1 expressions in formalin fixed, paraffin-embedded, preradiation biopsy samples from 49 patients with primary rectal carcinoma were analyzed immunohistochemically. p53 and p21 expressions and their relationships with histopathologic changes after radiation and other clinical features were evaluated. RESULTS: Expressions of p53 and p21/WAF1 were 49 and 28.6 percent, respectively. In 36.7 percent of total tumors, significant histopathologic effect can be observed. There was a significant inverse expression of p53 and p21. Most of the p53(+) or p21(–) tumors were radioresistant, and the majority of p53(–) or p21(+) tumors were radiosensitive. Tumors size in the radiosensitive, p53(–), or p21(+) group decreased more significantly than in radioresistant, p53(+), or p21(–) group (P<0.01), and patients with radioresistant, p53(+), or p21(–) tumors had more local recurrence, more distant metastasis, and a shorter five-year survival rate than those with radiosensitive, p53(–), or p21(+) tumors, but without statistic significance. No statistically significant correlation can be observed between other tumor clinical features and radiosensitivity, p53, or p21 expressions. CONCLUSION: Immunohistochemistry detection of p53 and p21 expressions may be useful parameters for more radiosensitive patients selected for preoperative radiotherapy.Supported by a Grant-in-aid for Scientific Research from the Ministry of Education, Science and Culture of Japan.Presented at the meeting of the Asian-Pacific Congress of Gastroenterology, Yokohama, Japan, September 19 to 23, 1996.