Evaluation of p21WAF1/CIP1 and cyclin D1 expression in the progression of superficial bladder cancer

Immunoreactivity of p21^WAF1/CIP1 and cyclin D₁ proteins was assessed in a cohort of 207 patients with superficial (pTa-pT₁) bladder cancer followed up for a mean of 4.9 years. The results of the immunostainings were compared with T category, WHO grade, tumor cell proliferation rate (MIB-1 score), the expressions of p53 and bcl-2 as well as survival. Sixty-eight percent and 75% of the tumors were p21^WAF1/CIP1 positive (≥5% of cells positive) and cyclin D₁ positive (≥10% of cells positive), respectively. The p21^WAF1/CIP1 expression was related to cyclin D₁ immunolabelling (P < 0.001) but not to the other variables studied. The expression of cyclin D₁ was inversely associated with T category (P=0.001), WHO grade (P=0.006), MIB-1 score (P=0.014), p53 expression (P=0.001), and bcl-2 (P=0.011) immunoreactivity. In univariate analysis, T category (P=0.0001), WHO grade (P < 0.0001), MIB-1 score (P < 0.0001), bcl-2 (P=0.0092), p53 (P=0.0016) and p21^WAF1/CIP1 (P=0.009) expressions were significant prognostic factors with regard to tumor progression, whereas cyclin D₁ was without any prognostic significance (P=0.1). Out of 123 p21 positive tumors 21 progressed, whereas only 2 out of 58 p21 negative tumors progressed. In multivariate analysis, the MIB-1 score was the only independent predictor of cancer-specific survival (P=0.03), whereas tumor grade (P=0.002) and cyclin D₁ expression (P=0.04) were independent predictors of tumor recurrence. Only the WHO grade (P=0.04) retained its prognostic value indicating the risk of progression. We suggest that in superficial bladder cancer p21^WAF1/CIP1 and cyclin D₁ immunohistochemistry provide no additional prognostic information compared with already established prognostic factors for predicting the risk of progressive disease. Received: 13 September 1999 / 22 March 2000     

p53、p16、Cyclin D1表达与胃癌增殖细胞核抗原的关系

目的 探讨ｐ５３、ｐ１６、Ｃｙｃｌｉｎ Ｄ１表达与胃癌细胞增生状况的关系。方法 采用免疫组化ＡＢＣ法检测５８例胃癌中ｐ５３、ｐ１６、ＣｙｃｌｉｎＤ１和ＰＣＮＡ的表达状况,并对ＰＣＮＡ免疫阳性细胞作半定量分析。结果 ｐ５３、ｐ１６、Ｃｙｃｌｉｎ Ｄ１阳性表达率分别为５１．７％（３０／５８）、４８．３％（２８／５８）、５３．４％（３１／５８）;ｐ５３、Ｃｙｃｌｉｎ Ｄ１阳性组织中,单位面积ＰＣＮＡ阳性     

Expression of p16 and cyclin D1 in bladder cancer and correlation in cancer progression

INTRODUCTION: Gene p16 encodes a cyclin-dependent kinase inhibitor which functions to regulate cyclin D1, cell cycle progression and malignancies. The relationship between p16 and cyclin D1 is thought to alter bladder cancer formation and tumor progression. We aimed to investigate the expression of p16 and cyclin D1 genes in order to evaluate their clinical significance in bladder cancer. MATERIALS AND METHODS: Tissue samples from 67 patients with transitional cell carcinoma were examined with an immunohistochemical stain for the expression of p16 and cyclin D1 genes. The expression rate was compared to 12 normal urinary bladder mucosa samples obtained from transurethral surgery from noncancer patients. RESULTS: The results revealed significant differences between normal bladder mucosa (100%) and cancer tissue (40.3%) for the positive staining of p16 protein (p < 0.001), while positive staining for the cyclin D1 protein in the patient group (67.2%) was significantly higher (p = 0.003) than that in the control group (16.7%). With the progression of tumor grade and clinical staging the positive rate of p16 gene expression increased, whereas, that of cyclin D1 decreased. Expression of the p16 gene in the non-invasive group was greater than that in the invasive group and a lower expression rate of the cyclin D1 gene in the non-invasive group compared to the invasive group. CONCLUSIONS: The results revealed that expression of the p16 gene is inversely proportional to the expression of the cyclin D1 gene. Therefore, abnormal expression of the p16 and cyclin D1 genes play important roles in tumorigenesis and tumor progression.     

p53 Gene mutations in superficial bladder cancer

OBJECTIVES: To assess the presence of p53 gene mutations in superficial tumors of the urinary bladder (transitional cell carcinoma) and their relationship to classic prognostic factors for cancer recurrence and progression. To analyze the implication of these mutations on the P53 protein structure. MATERIALS AND METHODS: Observational, cross-sectional study of 90 consecutive patients, 60 with superficial transitional cell carcinoma (pTa and pT1) and 30 without neoplastic disease (control group). Samples of bladder tumor and control normal mucosa were analyzed by polymerase chain reaction-single-strand conformation polymorphism (PCR-SSCP) to detect p53 mutations in exons 5-9. Automatic sequencing was used to characterize the mutations and their effect on the P53 protein was analyzed. Bivariate analysis was used to assess the association with other prognostic factors. RESULTS: PCR-SSCP found no mutations in any control group patient, whereas 38.3% of patients with superficial transitional cell carcinoma had one or more mutations in the exons analyzed. Thirty mutations were sequenced; all were point mutations and 86.67% were considered relevant for the P53 structure. A total of 93.3% of the mutations were located in highly conserved regions and 73.3% in mutational hot spots. The highest cell differentiation grades and pT1 stage were associated with a higher incidence of p53 gene mutations. Previous recurrences and other tumor-related histological variables were not associated with a higher percentage of mutations. CONCLUSION: Mutations at p53 did not appear in healthy bladder mucosa and were significantly more frequent in pT1 and high-grade (G-II and G-III) tumors. All mutations detected were point mutations and most caused considerable P53 structural abnormalities, implying major repercussions on P53 function. These data suggest that certain p53 mutations may have prognostic value, even though they were not associated with other classic recurrence and tumor progression parameters. Future analyses of the progress of patients with superficial bladder transitional cell carcinoma and mutated p53 will help clarify this aspect.     

p16和cyclin D1在膀胱癌中的表达及其相关性研究

目的研究ｐ１６和ｃｙｃｌｉｎＤ１在膀胱癌组织中的表达,探讨其与膀胱癌生物学行为的关系。方法应用免疫组化法观察５０例膀胱癌ｐ１６和ｃｙｃｌｉｎＤ１的表达情况。结果（１）膀胱癌组织中ｐ１６阳性率为４４％,明显低于正常膀胱粘膜（Ｐ＜０．０１）;ｃｙｃｌｉｎＤ１阳性率为６２％,明显高于正常膀胱粘膜（Ｐ＜０．０１）。随着肿瘤恶性程度的增高和临床分期的进展,ｐ１６阳性率下降,ｃｙｃｌｉｎＤ１阳性率升高。ｐ１６在未复发组、存活组的表达率显著高于复发组、死亡组（Ｐ＜０．０１）。ｃｙｃｌｉｎＤ１在存活组的表达率显著低于死亡组（Ｐ＜０．０５）。（２）ｐ１６与ｃｙｃｌｉｎＤ１的表达呈负相关。（３）有１５％的病例存在ｐ１６和ｃｙｃｌｉｎＤ１同时表达或失表达。结论ｐ１６与ｃｙｃｌｉｎＤ１异常表达在膀胱癌的发生、发展过程中起重要作用。     

Immunohistochemical Expression of p16, p21, p27 and Cyclin D1 in Oral Nevi and Melanoma

The acquisition of abnormalities at G1/S is considered a crucial step in the genesis and progression of melanoma. The expression of cell cycle regulators has also been used in various neoplasms as an adjunct to diagnosis. The aim of this study was to compare the expression of p16, p21, p27 and cyclin D1 in oral nevi and melanomas. Expression of these cell cycle regulatory proteins was evaluated by immunohistochemistry in 51 oral melanocytic lesions, including 38 intramucosal nevi and 13 primary oral melanomas. p16 and p27 were highly expressed in intramucosal nevi, whereas p21 and cyclin D1 expression was higher in oral melanomas. The results indicate that p21 and cyclin D1 may be involved in the development of oral melanomas, and eventually they may be useful in the differential diagnoses of oral benign and malignant melanocytic lesions.     

Immunohistochemical study of p53 and Ki-67 antigen expression in bladder carcinoma]

We examined the relationship between the expression of mutant p53 and Ki-67 antigens in urinary bladder transitional cell carcinoma and the pathological and clinical findings. Tissues were obtained from 28 patients with bladder carcinoma who underwent total or partial cystectomy. An ABC immunostaining method and two primary antibodies (DO-7 and MIB-1 antibodies) were used. The percentages of p53 and Ki-67 antigen-positive cells to the total number of cells were regarded as the p53 and Ki-67 labeling indices (LI) respectively. There were no statistically significant correlations between p53 LI and the histological grade or stage, although p53 LI increased slightly in the high grade and high stage group. There was a statistically significant correlation between Ki-67 LI and the histological grade and stage (p < 0.05). The correlation between p53 LI and Ki-67 LI was linear. Some cases had a p53 LI below the mean even though the Ki-67 LI was higher. The clinical course was characteristic of superficial bladder carcinoma initially, but progressed to invasive bladder carcinoma over the next several years. These results suggest that even cases initially diagnosed as superficial bladder carcinoma with a low p53 LI may progress to invasive bladder carcinoma in subsequent years. Therefore, it is important that the patient be followed-up.     

Loss of P16 expression and chromosome 9p21 LOH in predicting outcome of patients affected by superficial bladder cancer

OBJECTIVES: To evaluate the prognostic role of p16 expression and loss of heterozygosity (LOH) on chromosome 9p21 in patients affected by low-grade (G1-G2) urothelial bladder cancer. METHODS: Fifty-six consecutive patients with diagnosis of urothelial bladder cancer were enrolled in this prospective study. LOH analysis was performed on a blood/tumor pair sample of each patient, by using polymerase chain reaction analysis. The D9S171 (9p21) locus on chromosome 9 was investigated. All tumors were stained immunohistochemically for p16. Data from p16 and LOH analyses were compared with follow-up data to evaluate the prognostic role of these molecular markers. RESULTS: Loss of p16 expression was found in 33 patients (58.9%) and was significantly associated with the reduced recurrence-free probability (P < 0.0001). No correlations were reported with stage (P = 0.162) or grade (P = 0.051). Forty-three patients (76.7%) showed LOH on chromosome 9p21 (D9S171). A significant association was observed between loss of p16 expression and LOH on chromosome 9p21 (P = 0.005). The Kaplan-Meier curves showed a significant correlation between recurrence-free status and p16 expression (P = 0.0001). By multivariate analysis, p16 expression (P = 0.002) and number of lesions (P = 0.002) were identified as independent tumor recurrence factors. CONCLUSIONS: Our study highlights the prognostic role of p16 in predicting the recurrence-free probability in patients affected by low-grade urothelial bladder and highlights the fact that this method could be used in everyday urologic clinical practice to better characterize the natural history of urothelial bladder carcinomas.     

早期阴茎癌中p53、p16、EGFR、cyclinD1的表达

目的研究免疫组化指标p53、p16、EGFR、cyclinDl在早期阴茎癌中的表达及其与肿瘤复发的关系。方法回顾性分析2006年3月至2010年5月实施局部广泛切除术的早期阴茎癌33例。采用EnV确on／HRP两步法检测p53、p16、EGFR和cyclingD1的表达。结果33例中可获得完整病例资料的18例,平均年龄47．7（32-70）岁,平均随访时间21．9（5--45）个月。其中L期1例、T1a期12例、T1b期3例、T2期2例。由于样本量问题四项指标均无统计学意义。结论p53蛋白高表达是早期阴茎癌发生术后复发的可能危险因素,尚需进一步大样本研究证实。     

尼古丁对膀胱癌p53基因表达的相关研究

目的：探讨尼古丁与癌基因表达和膀胱癌生物学行为的关系。方法：对40只大鼠以10％BBN为致癌剂膀胱灌注诱发膀胱癌,其中30只分25、15、5mg／kg剂量以咽管灌胃方式给予尼古丁,10只作为对照,应用免疫组化方法对膀胱癌模型中p53蛋白进行检测。结果：不同剂量的尼古丁干预下的p53蛋白表达的阳性率分别为60％、30％、20％。p53蛋白表达阳性率与给药剂量、给药时间呈正相关。结论：尼古丁对癌基因的异常表达及协同作用,在膀胱癌的发生发展中起一定作用。     

p53 and microvessel density in primary resection specimens of superficial bladder cancer

PURPOSE: p53 Regulates angiogenesis in fibrosarcoma and correlative studies suggest a similar role for muscle invasive bladder cancer. We evaluated the associations of p53 status and microvessel density with pathological features and clinical outcomes in a large population of patients with superficial bladder cancer. In addition, we assessed the correlation of p53 status with microvessel density, which would suggest the regulation of angiogenesis by p53. MATERIALS AND METHODS: We stained 84 primary bladder resection specimens, including 55 stage pTa, 29 stage pT1, 27 grade 1, 35 grade 2 and 22 grade 3 samples, for p53, CD31 and CD34. The relationships of p53 or microvessel density and tumor stage-grade or clinical recurrence-progression were analyzed by analysis of variance and pairwise comparison analysis for least significant difference, and Pearson correlation coefficients. Only patients with no previous biopsy were included in analysis to preclude interference by granulation tissue related neovascularization. The 4 samples with significant inflammation were also excluded from study. RESULTS: At a mean followup of 33 months (range 1 to 93) 34 of 84 patients (40.4%) experienced 1 or more tumor recurrences and 10 (11.9%) had stage and/or grade progression. Statistically significant associations were observed of p53 immunostaining and microvessel density with tumor stage and grade (p <0.05). However, the association of p53 status with microvessel density was weak and not statistically significant. Similar results were observed for the CD31 and CD34 based estimates of microvessel density. Neither p53 status nor microvessel density correlated with recurrence or progression. CONCLUSIONS: Our study confirms the strong association of p53 and microvessel density with the well established prognostic factors of grade and stage in superficial bladder cancer, supporting other evidence of an important role for p53 and angiogenesis in the tumor biology of this disease. However, our data argue against a primary role of p53 in the regulation of angiogenesis in superficial bladder cancer. This study, which to our knowledge is the first to focus on primary resection specimens, suggests that other genetic or environmental factors may contribute to the regulation of angiogenesis in superficial bladder cancer.     

性激素受体在瘢痕中的表达及与细胞周期调控蛋白相互关系的研究

目的　了解雄激素受体 (AR)、雌激素受体 (ER)在病理性瘢痕中的表达及其与细胞周期调节蛋白D1(cyclinD1)、p16之间的相互关系 ,以探讨他们在瘢痕形成过程中的作用及机制。方法　采用免疫组化方法 (SP法 )对 30例瘢痕标本进行研究 ,以正常皮肤组织为对照 ,观察上述指标的表达。结果　正常皮肤及普通瘢痕成纤维细胞中所有指标均为阴性 ;增生性瘢痕与瘢痕疙瘩成纤维细胞中cyclinD1、p16、AR与正常皮肤相比差异均有显著性意义 (P <0 0 5 ) ;瘢痕疙瘩成纤维细胞cyclinD1和AR的表达高于增生性瘢痕 ,且有显著性意义 (P <0 0 5 ) ;p16在瘢痕疙瘩成纤维细胞的表达比增生性瘢痕为高 ,但两者之间差异无显著性意义。在病理性瘢痕中cyclinD1和AR的表达具有明显的相关性。结论　AR在病理性瘢痕的发生及发展中起一定的作用 ,它可能是通过与其配体结合后促使与cyclinD1有关的基因表达而发挥作用的。在瘢痕疙瘩里可能存在cyclinD1的促细胞增生作用超过P16细胞抑制 ,所以细胞呈现持续增殖状态 ;而在增生性瘢痕里cyclinD1与p16可能处于相对的平衡状态 ,细胞生长具有一定的自限性。     

p53、p21、Ki-67和VEGF与膀胱癌分级、分期以及预后的关系

目的：探讨p53、p21、Ki-67和VEGF的表达与膀胱癌的病理分级、分期以及预后足否相关。方法：应用免疫组织化学染色的方法,对40例手术证实的膀胱移行细胞癌患者的病理切片进行p53、p2l、Ki-67和VEGF的化学染色。将免疫组化结果与病理分级、分期以及预后情况进行分析。结果：在40个肿榴标本中．p53、p21、Ki-67和VEGF的表达有改变的分别有31个（77.5％）．22个（55.0％）．16个（40.0％）．17个（42.5％）：其中至少1个标记物表达异常35例（87.5％）．而4个标记物均表达异常者有7例（17.5％）。患者平均随访51个月。除了Ki-67、VEGF与病理分级以及Ki-67与分期之间无统计学意义外．4个标记物多少都与膀胱癌的病理分级、分期相关。p53（＋）／p21（-）以及4个标记物同时异常是与疾病相关死亡率有关的独立因素（P＜0.05,P＜0.01）。而Ki-67、VEGF均不是膀胱癌相关死亡率的独立因素（P〉0．05）。越多标记物表达异常,则膀胱癌的死亡率增加。结论：p53、p21、Ki-67和VEGF多少都与膀胱癌的病理分级、分期相关。联合检测p53、p21、Ki-67和VEGF可以更加准确地预测膀胱癌的预后。     

Immunohistochemical analysis of p53 and ras p21 expression in colorectal adenomas and early carcinomas

To further investigate whether multiple genetic changes are involved in the development of colorectal cancer, we performed an immunohistochemical analysis of p53 and ras p21 protein expression in 139 specimens of colorectal adenoma with varying degrees of dysplasia, 57 specimens of early cancer with an adenomatous component, and 12 specimens of superficial early cancer without any adenomatous component. Positive p53 staining was found in 15% of the adenomas with moderate dysplasia and in 42% of the adenomas with severe dysplasia or intramucosal carcinoma (IMCA). Positive immunostaining of p53 was observed to be significantly correlated with the degree of dysplasia and the depth of invasion, as was the expression of ras p21. However, a closer correlation was observed with the increasing size of the adenomas. Furthermore, p53 staining was positive in 42% of the 12 superficial early cancer specimens, while ras staining was positive in only 1 specimen (8%). These results indicate that p53 gene overexpression may play some biological role in both the adenoma-to-carcinoma sequence and in de novo cancer development, whereas ras p21 expression may not be as involved in de novo cancer development as in the malignant conversion of colorectal adenomas.     

p53, p21/WAF1, pRb的表达与T1G3膀胱癌预后的关系

目的　探讨p5 3,p2 1/WAF1和pRb的异常表达在T1G3 膀胱癌预后判断中的价值。　方法　对 4 7例T1G3 膀胱癌患者进行术后随访 ,采用p5 3,p2 1/WAF1和pRb单克隆抗体对手术标本行免疫组化染色。将肿瘤进展情况与染色结果和与预后相关的临床指标进行分析。　结果　 39例手术保留膀胱的患者总进展率为 5 9% ,其中 9例发生远处转移。 8例行膀胱全切术的患者 1例于术后 2年发现肺转移。肿瘤细胞核p5 3,p2 1/WAF1和pRb蛋白的异常表达率分别为 6 6 .7%、5 1.4 %和 71.8%。多因素分析显示 ,p5 3、pRb同时异常表达 (P <0 .0 5 )和p5 3、p2 1/WAF1、pRb三者同时异常表达(P <0 .0 1)与肿瘤进展显著相关。　结论　p5 3,p2 1/WAF1和pRb的表达与T1G3 膀胱癌患者的预后密切相关     

组织芯片技术应用于检测乳腺癌组织中p16和cyclin D1的表达

目的:探讨乳腺癌组织中多肿瘤抑制蛋白（p16）和细胞周期蛋白（cyclin D1）的表达及其意义。方法:采用组织芯片技术制作80例乳腺癌组织芯片，同时用SP免疫组织化学方法检测乳腺癌组织芯片中p16和cyclin D1的表达。结果:80例乳腺癌中p16和 cyclin D1的阳性率分别为40.0%和53.8%。乳腺癌中p16低表达与cyclin D1的高表达呈负相关（r<-0.49）;p16低表达、cyclin D1高表达与乳腺癌的淋巴结转移密切相关。结论:p16和cyclin D1的表达水平可以作为评估乳腺癌预后的参考指标。应用组织芯片大规模高效检测临床组织样本是可行的，具有快速、方便、经济、准确的特点。     

Correlation and prognostic significance of p53, p21WAF1/CIP1 and Ki-67 expression in patients with superficial bladder tumors treated with bacillus Calmette-Guerin intravesical therapy

PURPOSE: We determine if, before intravesical bacillus-Calmette Guerin (BCG) therapy, p53, p21WAF-1-CIP1 (a critical downstream effector of p53 pathway of cell growth control, inhibiting cyclin dependent kinases) and the cell proliferation marker Ki-67 (MIB-1) could be used as prognostic markers of response to BCG in patients with superficial bladder tumors. MATERIALS AND METHODS: The study included 47 patients with superficial bladder tumors at high risk for recurrence or progression treated with 6 weekly intravesical BCG instillations. We analyzed p53, p21 and Ki-67 on paraffin embedded samples by immunohistochemistry and the percentage of positive cells was determined in a blinded fashion. Quantitative immunostaining was analyzed in relation to time to recurrence and progression using univariate or multivariate analysis and the Kaplan-Meier method. RESULTS: During a mean followup of 24.6 months 23 of the 47 patients (48.9%) presented with tumor recurrence and 10 (21.2%) had later progression to invasive disease. A p21 over expression (greater than 10%) was observed in 23 tumors (48.9%) and positively correlated with p53 (p = 0.0097) but not with Ki-67 (p = 0.327). Of the tumors 18 (38.2%) were p53 and p21 negative. Among p21 positive tumors 15 (65.2%) were p53 and p21 positive, suggesting that p21 may also be regulated by p53 independent pathways. However, p53 did not act as a predictor of recurrence or progression. In contrast, using Kaplan-Meier curves p21 over expression (greater than 10%) and Ki-67 at a 25% cutoff were associated with shorter recurrence-free survival (both p = 0.02 log rank test) but they did not predict additional information about risk of progression. However, multivariate analysis failed to demonstrate any independent prognostic value for p21 or Ki-67 in contrast to tumor stage. CONCLUSIONS: Our results indicate that p21WAF-1-CIP1 seems to be regulated by p53 independent pathways in superficial bladder cancer. The present study did not indicate an independent prognostic significance in patients treated with BCG for p53, p21WAF-1-CIP1 or Ki-67 markers. Larger prospective studies are needed to evaluate further the independent value of these biological markers in superficial bladder cancer management.     

p16,cyclin D1和PCNA表达在胃癌早期诊断中的意义

目的 探讨Ｐ１６,ｃｙｃｌｉｎ Ｄ１和增殖细胞核抗原（ＰＣＮＡ）表达在胃癌早期诊断中的意义。方法 采用免疫组织化学（Ｓ－Ｐ）方法和显微摄像计算机图像分析技术,研究１８５例各类胃粘膜病变和４２例早期胃癌中Ｐ１６,ｅｙｅｌｉｎ Ｄ１和ＰＣＮＡ的表达情况。结果 Ｐ１６蛋白阳性率在慢性浅表性胃炎组（Ａ组）、慢性萎缩性胃炎组（Ｂ组）和会肠化生萎缩悸胃炎组（Ｃ组）均显著高于早期胃癌组（Ｅ组）（Ｐ〈０．０５～０     

病理性瘢痕中p53和cyclinD1的mRNA表达的实验研究

目的探讨病理性瘢痕中p53、cyclinD1的表达及在瘢痕疙瘩与增生性瘢痕中表达的区别,以进一步了解瘢痕疙瘩与增生性瘢痕的形成及发展的机理.方法手术切取病理性瘢痕标本作为实验组,以普通瘢痕作为对照组,运用RT-PCR方法测定病理性瘢痕中p53和cyclinD1的mRNA表达情况.结果病理性瘢痕中p53和cyclinD1的mRNA表达与对照组比较差异有显著意义(P＜0.05);而瘢痕疙瘩中p53的mRNA表达明显高于增生性瘢痕,两者比较差异有显著意义(P＜0.05).结论病理性瘢痕中p53和cyclinD1的异常表达,在病理性瘢痕的形成及发展中起重要作用.