Cyclin D1 and retinoblastoma susceptibility gene alterations in non-small cell lung cancer

Among the major regulators of the G₁ restriction point are cyclin D1 and the retinoblastoma gene product (RB). In non-small cell lung cancer (NSCLC), the cyclin D1 gene is amplified/over-expressed in almost 50% of cases, and RB is inactivated in 6–32% of cases. It is of interest to evaluate concurrently the alterations of both genes on the same series of NSCLCs, to investigate whether cyclin D1 and RB alterations are alternative pathways leading to inactivation of the G₁ restriction point or if they can occur in the same tumor, possibly exerting an additive effect on cancer progression. We investigated a series of 57 NSCLCs, analyzing cyclin D1 and RB at the gene and protein levels by Southern blot, Northern blot and immunohistochemistry. The cyclin D1 gene was amplified in 18 cases. cyclin D1 immunoreactivity was seen in 25 tumors. Amplification and expression were significantly associated. RB immunohistochemical expression was absent in 9 of 42 informative cases. RB mRNA expression was low to absent in 9 of 45 informative cases. cyclin D1 amplification was associated with normal RB mRNA, and cyclin D1 over-expression was associated with normal RB immunoreactivity, supporting the hypothesis that alterations of cyclin D1 and RB are alternative mechanisms by which tumor cells may escape the G₁ restriction point. A concurrent alteration of RB and cyclin D1 was seen in a small subset of NSCLCs. Abnormalities of cyclin D1 and/or RB at the gene and/or expression level were present in more than 90% of cases, stressing that cyclin D1 and/or RB alterations represent an important step in lung tumorigenesis. Int. J. Cancer 75:187–192, 1998. © 1998 Wiley-Liss, Inc.     

Nonsurgical therapy for stages I and II non-small cell lung cancer

For patients who have stages I and II non-small cell lung cancer (NSCLC) and who are unable or unwilling to undergo surgical resection, nonsurgical treatment modalities have been used with curative intent. Conventionally fractionated radiotherapy has been the mainstay of nonsurgical therapy; however, advances in technology and the clinical application of radiobiologic principles have allowed more accurately targeted treatment that delivers higher effective doses to the tumor, while respecting the tolerance of surrounding normal tissues. This article discusses nonsurgical approaches to the treatment of early-stage NSCLC, including several promising techniques, such as radiation dose escalation, altered radiation fractionation, stereotactic radiotherapy, and radiofrequency ablation.     

非小细胞肺癌p16基因表达与放疗敏感性的关系

目的：研究非小细胞肺癌（NSCLC）p16基因的表达与放疗敏感性的关系。方法：采用免疫组化检测68例晚期NSCLC的p16表达,该组病例均予以总量为55Gy的常规放疗,分析p16的表达与放疗疗效之间的关系。结果：有57．4％的晚期肺癌p16阴性表达,并且与癌细胞的分化程度密切相关;68例NSCLC采用放射治疗的疗效与癌细胞p16表达高度相关,p16阳性表达的患者其疗效明显好于p16阴性组的疗效（P〈0．05）。结论：NSCLC中存在高频率的p16基因失活,并且p16基因的表达与否可能影响肿瘤细胞对放射治疗的敏感性,因此检测肺癌p16基因的表达可能有助于判断肺癌对于放射治疗的反应,并藉此判断患者的预后,这对于指导临床肿瘤治疗具有重要的参考意义。     

p16、Rb蛋白在非小细胞肺癌中的表达及意义

 目的 探讨视网膜母细胞瘤蛋白质（Rb）和p16蛋白在非小细胞肺癌（NSCLC）中的表达及其与临床病理因素之间的关系。方法 采用免疫组化SP法检测80例NSCLC组织中Rb和p16蛋白的表达。结果 在80例NSCLC组织中， 分别有56例（70.00 %）和47例（58.75 %）显示Rb和p16蛋白表达缺失。两者之间有一定相关性（P＜0.05）。p16蛋白在鳞状细胞癌比腺癌中有较高的表达（P＜0.05），而Rb蛋白的表达异常与肿瘤分期有关（P＜0.05）。结论 Rb／p16路径的中断是肺癌的发生、发展中常见事件。     

非小细胞肺癌中CDK4I的表达及其与预后关系的探讨

目的探讨CDK4I基因产物在人非小细胞肺癌中的表达及其与预后之间的关系.方法应用免疫组化法对146例人非小细胞肺癌中的CDK4I基因产物表达进行了研究,并以20例正常肺组织作对照.结果肺癌CDK41表达水平(58.82%)明显低于正常肺组织(82.06%) (P<0.05). CDK4I表达水平降低的程度与肺癌的转移有密切关系(P<0.05),而与肺癌的原发肿瘤大小,组织学类型,肿瘤部位和患者年龄均无明显关系(P>0.05).CDK41高表达组术后3年及5年生存率明显高于低表达组(P<0.05).结论 CDK41基因可能参与人非小细胞癌的发生、发展和转移过程.     

Prognostic significance of p21waf1, cyclin D1 and retinoblastoma expression detected by immunohistochemistry in non-small cell lung cancer.

p21waf1 is a downstream effector of p53, and mediates growth arrest by inhibiting the action of G1 cyclin-dependent kinases. Cyclin D1 is a cell-cycle regulator essential for G1 phases progression and a candidate proto-oncogene implicated in the pathogenesis of several human tumor types. Cyclin D1 overexpression and the absence of retinoblastoma (Rb) protein have been frequently seen in various types of cancer, including lung cancer. The aim of this study was to clarify the relationship between the expressions of p21waf1, cyclin D1, and Rb protein, and to investigate the correlation between these protein expressions and the clinical features of the patients, and their prognoses. We immunohistochemically examined 92 samples of resected non-small cell lung cancer for p21waf1, cyclin D1, and Rb expression. Of the 92 specimens examined, 43 cases (46.7%) showed p21waf1 expression, 23 cases (25.0%) showed cyclin D1 overexpression, and 61 cases (66.3%) showed Rb expression. No correlation was observed between the expressions of p21waf1, cyclin D1, and Rb. There was no association of p21waf1 and cyclin D1 immunoreactivity with gender, disease stage, or histological types of the tumor. Regarding the prognosis in 79 cases with complete resection, no statistical differences were observed according to the degree of expression of these three factors. However, when unfavorable prognostic factors were considered to be the positive expression of p21waf1, positive of cyclin D1, and negative of Rb, the 5-year disease-free survival rate in the group with 2 or 3 unfavorable prognostic factors was 21.1%, which was statistically poorer than the 45.4% in the group with 0 or 1 unfavorable prognostic factor (p=0.0138). We conclude that examination of the expression of cell cycle regulators, such as p21waf1, cyclin D1, and Rb, is useful as a prognostic indicator, when these proteins' expression is analyzed in combination.     

Cyclin D1 overexpression is an indicator of poor prognosis in resectable non-small cell lung cancer.

Cyclin D1 is one of the G1 cyclins that control cell cycle progression by allowing G1 to S transition. Overexpression of cyclin D1 has been postulated to play an important role in the development of human cancers. We have investigated the correlation between cyclin D1 overexpression and known clinicopathological factors and also its prognostic implication on resected non-small-cell lung cancer (NSCLC) patients. Formalin-fixed and paraffin-embedded tumour tissues resected from 69 NSCLC patients between stages I and IIIa were immunohistochemically examined to detect altered cyclin D1 expression. Twenty-four cases (34.8%) revealed positive immunoreactivity for cyclin D1. Cyclin D1 overexpression is significantly higher in patients with lymph node metastasis (50.0% vs 14.4%, P = 0.002) and with advanced pathological stages (I, 10%; II, 53.8%; IIIa, 41.7%, P = 0.048; stage I vs II, IIIa, P = 0.006). Twenty-four patients with cyclin D1-positive immunoreactivity revealed a significantly shorter overall survival than the patients with negativity (24.0 +/- 3.9 months vs 50.1 +/- 6.4 months, P = 0.0299). Among 33 patients between stages I and II, nine patients with cyclin D1-positive immunoreactivity had a much shorter overall survival (29.7 +/- 6.1 months vs 74.6 +/- 8.6 months, P = 0.0066). These results suggest that cyclin D1 overexpression is involved in tumorigenesis of NSCLCs from early stage and could be a predictive molecular marker for poor prognosis in resectable NSCLC patients, which may help us to choose proper therapeutic modalities after resection of the tumor.     

p21 is associated with cyclin D1, p16INK4a and pRb expression in resectable non-small cell lung cancer

p21 (p21WAF1/CIP1) is involved in cell cycle regulation, as an inhibitor of cyclin dependent kinases (CDK2, CDK4 and CDK6). However, subsequent in vitro studies have suggested that p21 may influence this process by an additional mechanism, in particular through the regulation of cyclin D1 subcellular localisation. This study of primary resectable non-small cell lung cancer (NSCLC) was designed to examine p21 functions in association with the expression of cyclin D1 (including its subcellular localisation), p16INK4a and pRb. p21 expression was examined in 50 NSCLC (stage I-IIIA) and in several normal lung samples all of which had previously been studied for cyclin D1 (DNA, RT-PCR, immunostaining), p16INK4a (DNA, RT-PCR, immunostaining), and pRb (immunostaining). As assessed by immunoblotting and immunostaining, p21 was expressed at low levels in normal lung tissue with immunoreactivity seen in a small number of bronchial epithelial cells only. In NSCLC, p21 expression (> or =10% of positive cells) was observed in 42% (21/50) of cases. High p21 expression was associated with well differentiated tumours (p = 0.01) and cyclin D1 nuclear staining (p = 0.02). Furthermore, we found an inverse correlation with p16INK4a (p = 0.004) and a direct correlation with pRb expression (p = 0.02). Risk of relapse was associated with p16INK4a and p21 status with no relapse in patients with normal p16INK4a and p21. Our results confirm that a large number of NSCLC have a low level of p21 expression. The associations of p21 and nuclear cyclin D1, pRb, p16INK4a support the relevance of pathways linked to lung carcinogenesis that involve p21 but may act in addition to direct CDK inhibition.     

Postsurgical adjuvant therapy in stages I, II, and IIIA non-small cell lung cancer

The prognostic importance of accurate staging of non-small cell lung cancer was established in 1974 and reaffirmed and refined in 1986. The concept of adjuvant therapy after pulmonary resection for lung cancer is justified by the behavior of the disease. The best available data pertinent to adjuvant therapy of lung cancer have been collected by The Lung Cancer Study Group over the past 13 years. These data are based on a commitment to prospective and standardized surgical staging as a basis for large-scale prospective randomized control trials. A treatment effect of combination chemotherapy has been detected for stage II and IIIA nonsquamous cancer and is suggested for squamous cancer as well. This treatment effect is of marginal clinical significance. Adjuvant therapy for stage I disease has not shown a detectable benefit. Adjuvant radiation therapy for stage II and IIIA squamous cell carcinoma likewise has not resulted in survival benefit. Systemic metastasis continues to be the major clinical problem in lung cancer treatment, and better systemic therapy is necessary to improve the outcome in this disease. However, some patients do benefit from adjuvant chemotherapy, and efforts to identify such patients prospectively are also the subject of current clinical research.     

p53、Cyclin D1及PCNA在非小细胞肺癌的表达

目的 研究非小细胞肺癌（NSCLC）中p53、Cyclin D1蛋白表达及与细胞增殖的关系。方法 应用免疫组织化学技术（SP法）检测74例NSCLC中p53、Cyclin D1蛋白和增殖细胞抗原（PCNA）的表达情况。结果 p53蛋白、CyclinD1蛋白在NSCLC中阳性表达率分别为55．41％和37．84％,均明显高于正常肺组织（P1＝0．0031,P2＝0．0429）。 p53蛋白表达与淋巴结转移有密切关系（P＝0．0222）。p53蛋白、CyclinD1蛋白表达分别与PCNA指数有密切关系（P1＝0．001,P2＝0．0009）。p53蛋白与CyclinD1蛋白表达之间未见明显关系（P＞0．05）。结论 p53蛋白和CyclinD1蛋白近表达参与了NSCLC的发生发展,有促进细胞增殖的作用。p53的诊断和评价NSCLC预后的重要参数。     

Adenovirus-mediated p16INK4a gene expression radiosensitizes non-small cell lung cancer cells in a p53-dependent manner

We examined the influence of adenovirus-mediated wild-type p16INK4a (Ad/p16) expression on the radiation sensitivity of NSCLC cell lines, all of which lacked constitutive p16INK4a but each of which varied in p53 status: A549 (-p16INK4a/ +pRb/wt-p53), H322 (-p16INK4a/ +pRb/mt-p53), and H1299 (-p16INK4a/ +pRb/deleted-p53). The in vitro clonogenic survival results indicate that Ad/p16 enhanced the radiosensitivity of A549 but not H322 or H1299. Further analysis indicated that the apoptosis induced by combination therapy using Ad/p16 plus irradiation was dependent on the endogenous p53 status of the cancer cells. We performed Western blotting to analyse the p53 protein expression of A549 cells treated with either Ad/p16 or Ad/Luc. Endogenous p53 protein levels were higher in A549 cells transfected with Ad/p16 than in those transfected with Ad/Luc. Furthermore, when wt-p53 protein expression was restored in H1299 using Ad/ p53, Ad/p16 stabilized p53 protein expression and radiosensitized the cells. These results suggest that Ad/ p16-induced stabilization of p53 protein may play an important role in Ad/p16 mediated radiosensitization by enhancing or restoring apoptosis properties. Thus, Ad/ p16 plus radiation in combination may be a useful gene therapy strategy for tumors that have wt-p53 but nonfunctional p16INK4a.     

Cyclin E expression, a potential prognostic marker for non-small cell lung cancers.

Cyclin E is a G1 cyclin that has been shown to be one of the key regulators of the G1-S transition and could consequently be a deregulated molecule in tumors. In the present study, we have characterized cyclin E expression by immunohistochemistry in 217 resected non-small cell lung cancers (NSCLCs) and found large variations in cyclin E expression among tumors. High-level cyclin E expression (a cyclin E-labeling index > or =30%), observed in 115 (53%) of 217 NSCLCs, was more frequently found in tumors from smokers than from nonsmokers (P = 0.001), in squamous cell carcinomas than in nonsquamous cell carcinomas (P = 0.0002), and in pT2-4 tumors than in pT1 tumors (P = 0.04) by the chi2 test. Multivariate logistic regression analysis for the correlation between cyclin E expression and various characteristics showed a significant association of high-level cyclin E expression with squamous cell carcinomas (P = 0.005). Patients with tumors having high-level cyclin E expression survived a significantly shorter time than patients with tumors having low-level expression, both among the 151 patients with potentially curatively resected NSCLCs (5-year survival rates, 48 and 63%, respectively; P = 0.03) and the 103 patients with p stage I NSCLCs (5-year survival rates, 57 and 81%, respectively; P = 0.007). High-level cyclin E expression was also a significant and independent unfavorable prognostic factor in both patients with potentially curatively resected NSCLCs (P = 0.01) and in those with p stage I NSCLCs (P = 0.03) by Cox's proportional hazards model analysis. These findings indicate that cyclin E may play a pivotal role for the biological behavior of NSCLCs, and that a high level of cyclin E expression may be a new prognostic marker for NSCLCs.     

p16和Rb基因产物在非小细胞肺癌的表达

目的　研究非小细胞肺癌 (NSCLC)中p16、Rb蛋白表达及二者的关系。方法　应用免疫组织化学技术 (SP法 )检测 74例NSCLC和 10例正常肺组织中p16、Rb蛋白的表达情况。结果　p16蛋白在NSCLC的缺失率为 45 .95 % ,显著高于正常肺组织 (P <0 .0 1) ,并与淋巴结转移有密切关系 (P <0 .0 1)。Rb蛋白在NSCLC的缺失率为 2 8.3 8% ,与正常肺组织差异无显著性 (P >0 .0 5 )。p16蛋白与Rb蛋白表达呈负相关 (P <0 .0 5 )。结论　p16缺失与可能NSCLC的发生、转移有密切关系。p16与Rb可能通过负反馈机制相互调节。     

p27表达水平与非小细胞肺癌预后关系的Meta分析

目的：采用Meta分析的方法,探讨p27在非小细胞肺癌（NSCLC）预后中的作用。方法：检索PubMed、CNKI中研究p27表达与NSCLC预后关系的文献,收集每篇文献的相对危险比（HR）及95％可信区间（95％CI）,应用Meta分析Dersimonian—Laird模型对文献进行定量综合分析。结果：共入选7篇文献,累计NSCI。C病例888例,其中p27阳性表达443例,阳性率49．9％。对入选7篇文献进行一致性检验,文献具有异质性（Q=102．13,P=0．000）,合并相对危险比为1．21（95％可信区间：1．13～1．29,P=0．000）。结论：p27的阳性表达可能是NSCLC的良好预后因素。     

非小细胞肺癌p16基因甲基化及缺失的研究

目的 研究p16基因在非小细胞肺癌组织中的甲基化和缺失情况，探讨其在肺癌诊断中的价值。方法 应用甲基化相关的PCR及双重PCR，检测50例肺癌组织和54例正常肺组织中p16基因第1外显子5′端启动子区域CpG岛甲基化及第2外显子缺失情况。结果 p16基因在非小细胞肺癌组织中甲基化率为32.0%(16/50)。缺失率为28.0%(14/50);54例正常肺组织甲基化率为3.7%(2/54)，缺失率为0，二组比较，甲基化率（Fisher's exact=0.000)及缺失率（Fisher's exact=0.000)均有显著性差异。结论 甲基化和缺失是非小细胞肺癌p16基因失活的主要形式，检测p16基因甲基化和缺失状态可能有助于肺癌的诊断。     

p21WAF1蛋白的表达与非小细胞肺癌增殖和预后的关系

目的检测人非小细胞肺癌(non-small cell lung cancer,NSCLC)组织中的p21WAF1/CIP1表达水平,探讨NSCLC中p21WAF1/CIP1的表达与细胞增殖活性和预后的关系.方法采用流式细胞术检测60例NSCLC中p21WAF1/CIP1的表达量及DNA含量;分析p21WAF1/CIP1表达水平与S期细胞比例、预后的关系.结果正常肺组织中未见p21WAF1/CIP1的表达,60例NSCLC组织中p21WAF1/CIP1阳性率为60%(36/60),p21WAF1/CIP1阳性表达者的S期细胞比例(10.87±0.69)%低于阴性表达者(13.30±0.93)%,t=3.232,P= 0.001.p21WAF1/CIP1的表达与患者的术后累积生存月数和5年生存率有关,p21WAF1/CIP1 阳性表达者的术后平均生存月数(40个月)明显高于阴性表达者(25个月),t=2.885,P=0.003;5年生存率(29%)也高于阴性表达者(9%),t=2.712,P=0.004.结论 p21WAF1/CIP1表达是判断NSCLC预后的一个可参考指标.     

周围型肺癌CT征象及预后与Cyclin D1表达的相关性研究

目的：探讨周围型肺癌CT征象及预后与Cyclin D1表达的关系。方法：利用免疫组化SP法及螺旋CT扫描,回顾分析92例经病理确诊的周围型肺癌Cyclin D1基因表达与CT征象及其预后的关系。结果：深分叶、棘突征、短细毛刺征、纵隔淋巴结转移与Cyclin D1表达有关,X^2值分别为6．139、4．727、4．756和5．220,P值均〈0．05;肿瘤大小、空洞、胸膜凹陷、组织类型、肿瘤分化程度、肿瘤TNM分期、年龄、性别与Cyclin D1表达无关,X^2值分别为0．244、0．004、0．411、1．523、0．391、0．040、1．104、1．896,P值均〉0．05。Cyelin D1过度表达与预后有关。Cyclin D1为一负性预后因子,其过度表达提示预后不良。结论：Cyclin D1表达在肺癌的发生、发展及CT征象中可能起重要作用,检测该基因并结合CT征象可作为临床诊断及预后评估的指标。