Predictive significance of the alterations of p16INK4A, p14ARF, p53, and proliferating cell nuclear antigen expression in the progression of cervical cancer.

PURPOSE: The purpose of this research was to evaluate the clinical significance of p16INK4A, p14ARF, p53, and proliferating cell nuclear antigen (PCNA) expression in tumor progression of cervical cancer. DESIGN: Seventeen patients (40 samples) with consecutive cervical lesions from normal squamous epithelium, inflammation of the cervix to cervical intraepithelial neoplasm (CIN) and invasive cervical squamous cell cancer (SCC), or from CIN to SCC were collected for this study. Expression of p16INK4A, p14ARF, p53, and PCNA were detected by immunohistochemistry on paraffin-embedded sections. Human papillomavirus DNA was detected simultaneously with PCR and typed according to its DNA sequence. RESULTS: p16INK4A overexpression was significantly higher in CIN (75%) and in SCC (75%) than in normal or inflammation of the cervix (12.5%; P < 0.01, P < 0.05, respectively). The positive rate of p14ARF expression was higher in SCC (83%) than in normal/inflammation of the cervix (25%; P < 0.05). PCNA expression was negative in normal or inflammation of the cervix, but an increased in expression was seen in 63.2% in CIN and 100% in SCC (P < 0.01, P < 0.05). When the time interval for disease progression from initial biopsy to CIN 3 or invasive cancer was compared with states of p16INK4A expression, cases stained positive for p16INK4A progressed within 64.2 months as compared with 122.3 months among those stained negatively (P < 0.01). Cases with increased p14ARF expression also had a short time interval for disease progression of 78.8 months as compared with 108.3 months in cases that were p14ARF negative. Cases with stable or decreased p53 expression had the shortest time interval for progression of 32.3 months in contrast to cases with no p53 expression (113.9 months). However, cases with increasing p53 expression progressed within 60.8 months. CONCLUSIONS: Our results suggested that altered states of p16INK4A, p14ARF, p53, and PCNA may be valuable markers to predict the progression of cervical neoplasia.     

The prognostic significance of p16(INK4a)/p14(ARF) locus deletion and MDM-2 protein expression in adult acute myelogenous leukemia

BACKGROUND: The p16(INK4a) locus encodes two distinct proteins, p16(INK4a) and p14(ARF). Although p16(INK4a) and p15(INK4b) are involved in the phosphorylation of the retinoblastoma (Rb) protein, p14(ARF) interacts with the MDM-2 oncoprotein antagonizing its function as a suppressor of p53. The role of deletions of p16(INK4a)/p14(ARF) and p15(INK4b) and expressions of MDM-2 in myeloid leukemias and its influence on prognosis remain unclear. METHODS: The authors analyzed deletions of p16(INK4)/p14(ARF) and p15(INK4b) in 74 adults with acute myeloid leukemia (AML) by Southern blotting. Western blotting was used to determine Rb protein phosphorylation in patients with deletions of p16(INK4)/p14(ARF) and p15(INK4b). Then, they analyzed the levels of MDM-2 protein expression and correlated it with prognosis in an expanded population of 79 adults with AML by immunoblot analysis and solid-phase radioimmunoassay. RESULTS: Deletions of p16(INK4a)/p14(ARF) and/or p15(INK4b) occurred in 4 of 74 patients (5%) (hemizygous in 3, homozygous in 1 patient). Although the complete remission (CR) rate was similar (79% vs. 50%; P = 0.187), CR duration (10 vs. 46 weeks; P < 0.001), event free survival rate (EFS; 6 vs. 85 weeks; P < 0.004) and overall survival rate (11 vs. 86 weeks; P = 0.001) were significantly shorter in patients with deletions of p16(INK4a)/p14(ARF) and/or p15(INK4b). Thirty-seven (47%) of 79 patients studied for MDM-2 showed increased MDM-2 expression. These patients had a significantly shorter EFS rate (50 vs. 64 weeks; P = 0.023) and a trend for shorter CR duration (24 vs. 53 weeks; P = 0.07). Overall survival rate was not significantly different (50 vs. 84 weeks; P = 0.136). CONCLUSIONS: The authors concluded that 1) deletions of p16(INK4a)/p14(ARF) and/or p15(INK4b) occur with low incidence in patients with AML; 2) patients with deletions of p16(INK4a)/p14(ARF) and/or p15(INK4b) have a significantly shorter CR duration, EFS rate, and overall survival rate than do patients without deletions; (3) overexpression of MDM-2 is common in AML and is associated with shorter CR duration and EFS rate. Mechanisms other than p14(ARF) deletion are responsible for MDM-2 overexpression, and this overexpression may play a role in the biology of the disease.     

非小细胞肺癌INK4a和ARF基因甲基化与其蛋白共表达关系的探讨

目的:分析INK4a和ARF基因启动子甲基化与其蛋白共表达之间的关系.方法:选择前期实验中p14ARF和p16INK4a蛋白共表达阴性的非小细胞肺癌(NSCLC)患者35例,共表达阳性的NSCLC患者20例作为研究对象,分别称为(p14 p16)阴性组和(p14 p16)阳性组.运用甲基化特异性PCR(MSP)方法对两组患者癌组织INK4a和ARF基因启动子的甲基化状态进行检测.结果:(p14 p16)阴性组有18例发生INK4a基因甲基化,(p14 p16)阳性组有2例发生INK4a基因甲基化,两组差异有显著意义(P＜0.01).(p14 p16)阴性组有8例发生ARF基因启动子甲基化,(p14 p16)阳性组有2例发生ARF基因启动子甲基化,两组差异无显著意义(P＞0.05).INK4a和ARF基因异常申基化相互之间无显著相关性(P＞0.05).结论:NSCLC患者肺癌组织INK4a基因甲基化是导致p16INK4a蛋白表达阴性的重要机制;INK4a和ARF基因甲基化可能是相对独立的事件.     

Two arginine rich domains in the p14ARF tumour suppressor mediate nucleolar localization

The INK4a/ARF locus encodes two distinct tumour suppressors, p16INK4a and p14ARF, that regulate cell cycle progression via the pRB and p53 pathways, respectively. The ARF protein inhibits hdm2 activity, leading to the stabilization of the p53 tumour suppressor and cell cycle inhibition. The amino-terminal domain of human p14ARF and of the mouse homologue, p19ARF, is sufficient for these effects. This domain is also sufficient for the nucleolar localization of the mouse ARF protein. In contrast, we show that the human ARF protein requires two arginine rich domains, one in the amino- and the other in the carboxy-terminus, for nucleolar targeting. The amino-terminal nucleolar-targeting domain of p14ARF is also important for ARF-hdm2 binding and cell cycle inhibition. The carboxy-terminal p14ARF nucleolar localization domain lies within the shared INK4a/ARF exon 2, and is mutated in a small number of melanoma-prone kindreds. The INK4a/ARF exon2-mutations could affect the function of both the p16INK4a and p14ARF tumour suppressors. Oncogene (2000).     

p16^INK4a和p14^ARF蛋白在宫颈癌和肺癌中的差异表达

[目的]研究宫颈癌和肺癌中p16INK4a和p14ARF蛋白表达水平的差异及意义。[方法]应用免疫组化方法检测50例宫颈癌和127例肺癌组织中p16INK4a和p14ARF蛋白表达。[结果]50例宫颈癌中,p16INK4a和p14ARF蛋白全部阳性表达;127例肺癌中,p16INK4a和p14ARF蛋白阳性表达率分别为61.42%和30.79%;宫颈癌和肺癌中p16INK4a和p14ARF蛋白表达差异均有显著性（P〈0.01）。[结论]p16INK4a和p14ARF蛋白在宫颈癌中过表达,而在肺癌中缺失表达。提示p16INK4a和p14ARF蛋白在宫颈癌和肺癌的发生发展中所起的作用和作用机制可能不同。     

Ratio of expression of p16INK4a to p14ARF correlates with the progression of non-small cell lung cancer.

The CDKN2 gene is located on the short arm of chromosome 9p and encodes two unrelated proteins, p16(INK4a) and p14(ARF), through the use of independent first exons and shared exons 2 and 3. p16(INK4a) is a cyclin-dependent kinase inhibitor, whereas p14(ARF) regulates the cell cycle through a p53 and MDM2-dependent pathway. We have examined the expression of p16(INK4a) and p14(ARF) using competitive RT-PCR in 60 non-small cell lung cancers (NSCLCs) and matching normal lung tissues. The intensities of bands for p16(INK4a) and p14(ARF) were nearly equal or the intensity of the p16(INK4a) band slightly exceeded that of p14(ARF) in the normal lung tissues (n = 60). In 38 tumors the intensity of the p16(INK4a) band was similar to or slightly weaker than that of p14(ARF). In 6 tumors the intensity of the p16(INK4a) band was weaker than that of p14(ARF). In 15 tumors the intensity of the p14(ARF) band was very strong and the p16(INK4a) band was barely visible. In only one tumor was the intensity of the p16(INK4a) band very strong, while the band of p14(ARF) was barely visible. The ratio of the intensity of p16(INK4a) to p14(ARF) had an interesting correlation with the tumor's clinicopathological characteristics. The p stage II - IV tumors had significantly lower p16(INK4a) to p14(ARF) ratios than the p stage I tumors (P = 0.036). The T2 - 4 tumors had significantly lower p16(INK4a) to p14(ARF) ratios than the T1 tumors (P = 0.005). The N1 - 3 tumors had significantly lower p16(INK4a) to p14(ARF) ratios than the N0 tumors (P = 0.014). Our results suggest that the ratio of expression of p16(INK4a) to p14(ARF) tends to decrease during the progression of NSCLC.     

Association between DNA methylation and shortened survival in patients with advanced colorectal cancer treated with 5-fluorouracil based chemotherapy.

PURPOSE: There are no good genomic markers of survival in patients with advanced colorectal cancer. The CpG island methylator phenotype (CIMP) marks a distinctive pathway in colorectal cancer. We sought to determine the prognostic significance of CIMP in advanced colorectal cancer patients treated with 5-fluorouracil (5-FU) in an Eastern Cooperative Oncology Group clinical trial. EXPERIMENTAL DESIGN: We studied 188 patients enrolled on protocol E2290, a five-arm trial comparing 5-FU, 5-FU in combination with N-phosphonoacetyl-l-aspartic acid, oral leucovorin, i.v. leucovorin, or IFNalpha-2a in patients with advanced colorectal cancer. Methylation of MINT1, MINT31, hMLH1, p14ARF, and p16INK4a in DNA extracted from formalin-fixed paraffin-embedded specimens was evaluated by combined bisulfite restriction analysis, and methylation of MINT2 was studied by methylation-specific PCR. RESULTS: Methylation frequencies were 21% for MINT1, 23% for MINT2, 24% for MINT31, 4% for hMLH1, 11% for p14ARF, and 17% for p16INK4a. Methylation of MINT1, MINT31, p14ARF, and p16INK4a were correlated, as expected. There was no association between methylation and clinicopathologic factors or response to therapy. Methylation of MINT1, MINT31, p14ARF, or p16INK4a was associated individually with shortened overall survival. Hazard ratios were 1.51 (P = 0.05) for MINT1, 1.70 (P = 0.006) for MINT31, 2.22 (P = 0.001) for p14ARF, and 1.51 (P = 0.05) for p16INK4a. Concurrent methylation of two or more genes of the CIMP-associated subset (MINT1, MINT31, p14ARF and p16INK4a) defined a group of cases with markedly reduced overall survival and hazard ratio was 3.22 (P < 0.0001 in multivariate analyses). CONCLUSIONS: CIMP is associated with poor survival in advanced colorectal cancer patients.     

Alterations of p16(INK4a) and p14(ARF) in patients with severe oral epithelial dysplasia

A number of genetic aberrations have been reported in end-stage squamous cell carcinoma of the head and neck, including p16(INK4a) and p14(ARF) (INK4a/ARF) inactivation rates of 70-85%. Still, the cell cycle-regulatory genes p16(INK4a) and p14(ARF) remain poorly understood in oral cavity premalignant lesions. This study evaluated INK4a/ARF locus alterations in 26 patients (28 samples) deemed to be at increased risk for malignant transformation to squamous cell carcinoma due to the diagnosis of severe oral epithelial dysplasia. Microscopically confirmed dysplastic oral epithelium and matching normal tissue were laser capture-microdissected from paraffin sections, DNA was isolated, and molecular techniques were used to evaluate p16(INK4a) and p14(ARF) gene deletion, mutation, loss of heterozygosity (LOH), and hypermethylation events. Deletion of exon 1beta, 1alpha, or 2 was detected in 3.8%, 11.5%, and 7.7% of patients, respectively. INK4a and ARF mutations were detected in 15.4% and 11.5% of patients with severe dysplasia of the oral epithelium. All identified mutations occurred in the INK4a/ARF conserved exon 2. Allelic imbalance was assessed using three markers previously reported to show high LOH rates in head and neck tumors. LOH was found in 42.1%, 35.0%, and 82.4% of patients for the markers IFNalpha, D9S1748, and D9S171, respectively. Hypermethylation of p16(INK4a) and p14(ARF) was detected in 57.7% and 3.8% of patients, respectively, using nested, two-stage methylation-specific PCR. The highest rates of p16(INK4a) hypermethylation occurred in lesions of the tongue and floor of the mouth. In addition, p16(INK4a) hypermethylation was significantly linked to LOH in two or more markers. These data support that INK4a/ARF locus alterations are frequent events preceding the development of oral cancer and that p16(INK4a) inactivation occurs to a greater extent in oral dysplasia than does p14(ARF) inactivation.     

CDKN2A基因座在膀胱癌中的表达及临床意义

目的:探讨CDKN2A基因座两种编码产物p16INK4A和p14ARF在膀胱癌组织中的表达及其临床意义。方法:采用免疫组织化学技术SP法观察13例正常膀胱组织和52例膀胱尿路上皮癌组织中p16INK4A和p14ARF的表达情况。结果:p16INK4A在正常组织、膀胱癌中阳性表达率分别为84.62%(11/13)、40.38%(21/52),两组间差异显著(P<0.01)。p14ARF在正常膀胱组织、膀胱癌中阳性表达率分别为92.31%(12/13)、23.08%(12/52),两组间差异显著(P<0.01)。p16INK4A和p14ARF的阳性表达率与肿瘤病理分级、临床分期呈负相关(P<0.05);两者在膀胱癌中的表达没有相关性(P>0.05),但他们均与患者的预后正相关(P<0.05)。结论:p16INK4A和p14ARF在膀胱癌中的表达异常与膀胱癌的发生发展密切相关,但它们所起的生物学作用不尽相同,联合检测两者的表达可能为膀胱癌的早期诊断及预后判断提供帮助。     

Deregulation of the TP53/p14ARF tumor suppressor pathway in low-grade diffuse astrocytomas and its influence on clinical course.

PURPOSE: The chromosome 9p21 region harbors three tumor suppressor genes, p14(ARF), p15(INK4b), and p16(INK4a), all of which can be targets for hypermethylation-associated inactivation in low-grade gliomas. p16(INK4a) and p15(INK4b) are critically involved in the RB1 pathway, whereas p14(ARF) acts as an upstream regulator of the TP53 pathway. The role of each tumor suppressor pathway in low-grade diffuse astrocytomas and their relationships with clinical behavior remain to be elucidated. EXPERIMENTAL DESIGN: We assessed the alterations of the RB1/CDK4/p16(INK4a)/p15(INK4b) and the TP53/MDM2/p14(ARF) pathways in 46 WHO grade II astrocytomas and analyzed their impact on prognosis. RESULTS: The TP53/MDM2/p14(ARF) pathway was altered in 32 of 46 cases (70%) by either TP53 mutation (25 cases) or p14(ARF) methylation (9 cases). The RB1/CDK4/p16(INK4a)/p15(INK4b) pathway was disrupted in 6 of 46 cases (13%) by either RB1 methylation (1 case), p16(INK4a) methylation (3 cases), or p15(INK4b) methylation or homozygous deletion (3 cases). Generally speaking, individual tumors thus tended to display alteration of only one component from both pathways. Any independently analyzed genetic alteration failed to provide statistically prognostic information. The alternate or simultaneous presence of TP53 mutation and p14(ARF) methylation emerged as a univariate predictor of a shorter progression-free survival (P = 0.0456) but was not statistically significant when age and extent of surgery were included in the analysis. CONCLUSIONS: Alternative disruption of the TP53/p14(ARF) pathway represents a frequent event in low-grade diffuse astrocytomas and correlates with an unfavorable clinical course. However, its value is unlikely to include prognostic utility that is independent of other conventional prognostic factors.     

Mechanisms of inactivation of p14ARF, p15INK4b, and p16INK4a genes in human esophageal squamous cell carcinoma.

The 9p21 gene cluster, harboring growth suppressive genes p14ARF, p15INK4b, and p16INK4a, is one of the major aberration hotspots in human cancers. It was shown that p14ARF and p16INK4a play active roles in the p53 and Rb tumor suppressive pathways, respectively, and p15INK4b is a mediator of the extracellular growth inhibition signals. To elucidate specific targets and aberrations affecting this subchromosomal region, we constructed a detailed alteration map of the 9p21 gene cluster by analyzing homozygous deletion, hypermethylation, and mutation of the p14ARF, p15INK4b, and p16INK4a genes individually in 40 esophageal squamous cell carcinomas (ESCCs) and compared the genetic alterations with mRNA expression in 18 of these samples. We detected aberrant promoter methylation of the p16INK4a gene in 16 (40%), of p14ARF in 6 (15%), and of p15INK4b in 5 (12.5%) tumor samples. Most p16INK4a methylations were exclusive, whereas all but one of the p14ARF/p15INK4b methylations were accompanied by concomitant p16INK4a methylation. We detected homozygous deletion of p16INK4a in 7 (17.5%), of p14ARF-E1beta in 13 (33%), and of p15INK4b in 16 (40%) tumor samples. Most deletions occurred exclusively on the E1beta-p15INK4b loci. Two samples contained p14ARF deletion but with p16INK4a and p15INK4b intact. No mutation was detected in the p14ARF and p16INK4a genes. Comparative RT-PCR showed good concordance between suppressed mRNA expression and genetic alteration for p15INK4b and p16INK4a genes in the 18 frozen samples, whereas 5 of the 13 cases with suppressed p14ARF mRNA expression contained no detectable E1beta alteration but aberrations in the p16INK4a locus. Our results show that in human ESCCs, p14ARF is a primary target of homozygous deletion along with p15INK4b, whereas p16INK4a is the hotspot of hypermethylation of the 9p21 gene cluster. The frequent inactivation of the p14ARF and p16INK4a genes may be an important mechanism for the dysfunction of both the Rb and p53 growth regulation pathways during ESCC development.     

p16INK4A immunohistochemical staining may be helpful in distinguishing branchial cleft cysts from cystic squamous cell carcinomas originating in the oropharynx

BACKGROUND:

We investigated p16^INK4A expression in branchial cleft cysts and its utility in distinguishing branchial cleft cysts from metastatic head and neck squamous cell carcinomas (SCCs) in fine‐needle aspiration biopsies (FNABs).

METHODS:

A study set comprising 41 resections (15 SCC and 26 branchial cleft cysts) and a test set of 15 FNABs (11 SCC and 4 branchial cleft cysts) were analyzed with p16^INK4A immunohistochemistry and human papillomavirus (HPV) polymerase chain reaction (PCR)/pyrosequencing. Cases with discrepant p16^INK4A and PCR/pyrosequencing results were further evaluated with HPV in situ hybridization (ISH). SCCs were divided into keratinizing SCC and nonkeratinizing SCC groups and site of origin.

RESULTS:

Metastatic oropharyngeal nonkeratinizing SCC in the study set exhibited diffuse, strong p16^INK4A (7 of 7) and HPV16 DNA positivity (6 of 6), while keratinizing SCC from the larynx and oral cavity was negative for p16^INK4A. p16^INK4A reactivity in the branchial cleft cyst study set was characterized by focal, strong staining (6 of 21) involving the superficial squamous epithelium. HPV DNA was identified in 7 of 19 branchial cleft cyst study set cases by PCR/pyrosequencing, but these cases were negative by HPV ISH. In the test set, oropharyngeal nonkeratinizing SCC exhibited diffuse, strong p16^INK4A (3 of 3) and HPV16 DNA (2 of 2), while metastatic keratinizing SCC was negative for p16^INK4A and HPV DNA. All 4 FNABs of branchial cleft cysts were negative for p16^INK4A. Diffuse, strong p16^INK4A correlated with oropharyngeal origin (P = .001) and nonkeratinizing morphology (P = .0001).

CONCLUSIONS:

Branchial cleft cysts can exhibit focal strong reactivity limited to the superficial squamous epithelium and glandular epithelium. Although p16^INK4A immunohistochemistry may be helpful in distinguishing oropharyngeal nonkeratinizing SCC from branchial cleft cysts in FNAB specimens, it is not helpful in cases of keratinizing SCC because these cases are typically negative for p16^INK4A. Cancer (Cancer Cytopathol) 2009. © 2009 American Cancer Society.     

Genomic instability, mutations and expression analysis of the tumour suppressor genes p14(ARF), p15(INK4b), p16(INK4a) and p53 in actinic keratosis

Actinic keratosis (AK) is a well-established pre-cancerous skin lesion that has the potential to progress to squamous cell carcinoma (SCC). We investigated the involvement of the CDKN2A, CDKN2B and p53 genes in AK and in the progression of AK to SCC. Mutational analysis on exons 1a, 1b and 2 of the CDKN2A locus and exon 1 of the CDKN2B locus as well as allelic imbalance was performed in 26 AK specimens. Expression levels of the genes p14(ARF), p15(INK4b), p16(INK4a) and p53 were examined in 16 AKs and 12 SCCs by real-time RT-PCR. A previously described polymorphism of p16(INK4a) (Ala148Thr) was detected at an allelic frequency of 12%. Six samples carried novel mutations at codon 71 of the CDKN2A locus and one sample presented an additional mutation at codon 65. Two AK samples carried a not-previously described non-UV type missense mutation at codon 184 (Val184Glu) of exon 1b in the p14(ARF) gene. Regarding the CDKN2B locus a new mutation at codon 50 (Ala50Thr) and another at codon 24 (Arg24Arg), were detected. Microsatellite instability (MSI) was found in 15% of AKs in at least one marker, indicating that genetic instability has some implication in the development of AK. Down-regulation of p16(INK4a) and p53 mRNA levels was noted in SCC compared to AK. TSGs expression levels in sun-exposed morphologically normal-appearing skin, suggests that abnormal growth stimuli might exist in these tissues as well. Furthermore, we suggest a possible role of p15(INK4b), independently from the intracellular pathway mediated by p16(INK4a), and of p14(ARF) in AK development, as well as in the progression of AK to SCC. The deregulation of the expression profiles of the CDKN2A, CDKN2B and p53 genes may, independently of mutations and LOH at 9p21, play a significant role in AK and progression of AK to SCC.     

Aberrations of the p14(ARF) and p16(INK4a) genes in renal cell carcinomas.

The INK4a / ARF locus on chromosome 9p21, which encodes two distinct genes, p14(ARF) and p16(INK4a), is frequently altered in human neoplasms. To investigate the potential roles of p14(ARF) and p16(INK4a) genes in human renal cell carcinomas (RCCs), we analyzed 6 human RCC cell lines and 91 primary RCCs for homozygous deletion, promoter hypermethylation and expression of the p14(ARF) and p16(INK4a) gene products using differential PCR, methylation-specific PCR, and immunohistochemistry, respectively. Five cell lines showed homozygous co-deletion of both genes and one demonstrated promoter hypermethylation of the p16(INK4a) gene only. Eight of 91 RCCs showed aberrations of p14(ARF) or p16(INK4a) status and six of these featured gross extension into the renal vein. The results suggest that p14(ARF) and p16(INK4a) aberrations may play roles in the relatively late stage of renal tumorigenesis associated with tumor progression.     

Progression of actinic keratosis to squamous cell carcinoma of the skin correlates with deletion of the 9p21 region encoding the p16(INK4a) tumor suppressor.

Actinic keratoses (AKs) are pre-neoplastic lesions that can develop into squamous cell carcinomas (SCCs) of the skin. Often AK and SCC have commonly altered p53. A status of another tumor suppressor, the p16(INK4a), was reported for SCC but not for AK. A comparative study of SCC and AK human samples by loss of heterozygosity (LOH) analysis determined that the p16(INK4a/ARF) locus is less frequently altered in AKs than in SCCs. These LOH data highly correlated with immunohistochemical findings demonstrating the presence of p16(INK4a) in the AK skin samples but its absence in SCC lesions. Our results imply that progression of AK into SCC may involve inactivation of p16(INK4a).     

5' cytosine-phospho-guanine island methylation is responsible for p14ARF inactivation and inversely correlates with p53 overexpression in resected non-small cell lung cancer.

PURPOSE AND EXPERIMENTAL DESIGN: The molecular mechanisms by which the p14ARF gene is altered in non-small cell lung cancer (NSCLC) are complex and unclear. Using genetic and epigenetic analyses, we examined various molecular alterations including the loss of protein and mRNA expression, and 5'CpG hypermethylation, allelic imbalance, and mutation of the p14ARF gene in a series of 102 NSCLC samples, in parallel with clinicopathological and prognostic analyses. To clarify the biological significance of p14ARF alterations, its relationship with p16INK4a and p53 alterations was also examined. RESULTS: We found that 34% of NSCLC patients had aberrant P14ARF protein expression, which was more frequent in adenocarcinomas (AD; 44%) than in squamous cell carcinomas (22%; P = 0.024). A high concordance was observed between alterations in protein and mRNA expression and 5'CpG hypermethylation (P 相似文献