摘要
研究背景和目的
宫颈癌是世界上第二大常见的妇科肿瘤,尤其是在发展中国家。在发达国家,每10万妇女发现有10个宫颈癌患者,而在某些发展中国家每10万妇女发现有60个宫颈癌患者。绝大多数的宫颈癌由宫颈上皮内瘤样病变(cervical intra-epithelial neoplasia, CIN)逐渐发展而来。CIN是局限于宫颈上皮内,具有不同的组织学改变的宫颈癌前病变,从CIN发展到浸润癌大约需要10年的时间。如果在CIN或早期浸润癌阶段就能及早发现并给以极积治疗,治愈的机会非常高。因此,为了降低宫颈癌的发病率和死亡率,迫切地需要建立有效的宫颈癌筛查方案,尤其是在发展中国家。
细胞学检查和人类乳头状瘤病毒DNA检查是世界卫生组织强力推荐的宫颈癌筛查方法。据报道,在一些采取了有组织的筛查方法的国家,传统细胞学涂片检查已经降低了70%以上宫颈癌的发生率。尽管如此,细胞学涂片检查的灵敏度低、假阴性率高、检测结果不稳定。与传统的细胞学涂片相比,液基细胞学检查方法的应用增加了人群筛查的效能但并没有显著提高其灵敏度。由于细胞学检查的灵敏度低,一些癌前病变不能被早期发现,因而得不到有效的治疗而进展为浸润性宫颈癌。
HPV感染是宫颈癌发生的必需因素,99.7%的浸润性宫颈癌中可以发现HPV的感染。目前为止,100多种HPV被鉴定出来,大约13种HPV (16、18、31、33、35、39、45、51、52、56、58、59、68、73和82)被鉴定为高危型,它们可以转化生殖道上皮细胞并能引起浸润性宫颈癌。在这些高危型HPV中,最常见的是HPV16和18,与宫颈癌的发展密切相关。高危型HPV感染和宫颈癌发展的因果关系使得HPV检查具有了宫颈癌筛查的潜在作用。HCⅡ是美国食品和药物管理局推荐的HPV DNA的检测方法,能一次性检测出18种HPV,包括13种高危型别(16、18、31、33、35、39、45、51、52、56、58、59、68、73和82型)和5种低危型别(6,11,42,43 and 44型),但是不能鉴定是哪一型别的HPV存在,也不能区别是暂时感染还是持续感染。有趣地是,大多数HPV感染是暂时感染,机体可以将其自发清除而不能引起CINⅡ以上的病变,而只有高危型HPV的持续感染才能导致宫颈癌的发生。HPV DNA检查用于筛查CINⅡ以上宫颈病变具有更稳定更灵敏的优点,但特异性不如常规细胞学检查方法高。这是因为大多数HPV感染是暂时感染,机体可以自发清除而不引起CINⅡ以上宫颈病变。只有高危型HPV的持续感染(占HPV感染的10-20%)才能引起宫颈癌的发生。因此,需要找到更特异的生物学标记来增加CINⅡ以上病变筛查的特异性。这也许会使得暂时感染高危型HPV的妇女特别是细胞学检查结果正常的妇女避免进行不必要的随访。这也可能会减少对某些患者的过度诊断和治疗。
除了高危HPV感染,一些染色体异常和基因组不稳定性变化是癌前病变进展为浸润性宫颈癌的必要途径。在这些异常变化中,最常见的畸变是3q26的扩增,该区域包含人端粒酶RNA基因(TERC基因),70%左右的宫颈癌中发现有该染色体区域的异常扩增。端粒酶通过提供端粒稳定性和调节端粒长度来维持染色体的稳定。端粒酶主要由两个部分组成,一个是RNA亚单位为端粒重复序列提供模板,一个是蛋白亚单位催化端粒酶的合成。TERC基因编码端粒酶RNA单位来维持细胞分裂时端粒的长度。当TERC基因过表达时,细胞可以逃避凋亡而导致癌症的发生。最近的一些研究发现：采用荧光原位杂交技术检测TERC基因,结果发现该基因的扩增频繁地出现在宫颈鳞状细胞癌、高度宫颈上皮内瘤变及低度宫颈上皮内瘤变中且扩增频率随着宫颈病变严重程度的增加而增加,表明TERC基因可能是宫颈癌筛查的一个有效分子标志。Alameda F等研究发现,FISH检测3q26扩增可以作为低度宫颈上皮内瘤变病人持续或进展的预测指标。可见,对TERC基因扩增的检测有助于宫颈癌的筛查及早期诊断。目前,荧光原位杂交(fluorescence in hybridization, FISH)技术被认为是检测这一基因扩增的标准方法。FISH技术操作简单、重复性好、稳定性高,并具有较好的灵敏性及特异性,因此很适合于临床检测。目前已广泛应用于细胞遗传学、肿瘤生物学、基因定位、基因扩增和产前诊断等领域。
在本研究中,我们采用荧光原位杂交技术检测宫颈脱落上皮细胞hTERC基因扩增情况,并与细胞学、HPV结果、病理学结果比较,评估TERC基因检测对于宫颈癌筛查及CIN病情进展风险预测的临床意义,从而为中国妇女宫颈癌的筛查、早期诊断及预后评估提供新的途径。
方法/材料
收集南方医科大学附属南方医院2008年3月至12月的门诊接受筛查者或接受治疗前的住院患者120例,并取得患者的知情同意。所有的研究对象均排除急性感染、妊娠、手术切除及放化疗史。对每一位研究对象进行TERC基因的FISH检测,HPV DNA检测、细胞学检查和组织病理学检查。从宫颈鳞柱交接部采集脱落上皮细胞,然后将其贮存在液基细胞学专用储存瓶和HPV专用储存瓶中用于FISH检测和HPV检测。根据细胞学检查结果,120名研究对象分为正常21例、ASCUS 8例、ASCH 5例、LSIL 22例、HSIL 52例和SCC 12例。根据组织病理学结果,120名研究对象分为正常20例、CINⅠ14例、CINⅡ35例、CINⅢ36例和宫颈癌15例。
结果
TERC基因扩增的阳性率在正常、CINⅠ、CINⅡ、CINⅢ和宫颈癌组中分别是0.00%、7.14%、65.71%、97.22%和100.00%。不同宫颈病变组,TERC基因扩增阳性率有显著差异(P<0.01),随宫颈病变程度加重,TERC基因扩增阳性率显著增高。除正常组与CINⅠ、CINⅡ/Ⅲ与宫颈癌组之外,其余各组两两比较差异均有统计学意义(P<0.005)。TERC基因扩增阳性的异常细胞平均数在正常、CINⅠ、CINⅡ、CINⅢ和宫颈癌组中分别是0.70、3.57、10.97、17.86和35.87。不同宫颈病变组,TERC基因扩增的异常细胞平均数有显著性差异(P=0.000),随宫颈病变程度加重,TERC基因扩增阳性的异常细胞数增多。组间两两比较,各组异常细胞数差异均有统计学意义(P=0.000)。宫颈病变程度越高,TERC基因扩增阳性的异常细胞的核型表现越复杂。在正常对照组,异常信号的模式只有2:3(CSP3:TERC)一种。在CINⅠ组,异常信号的模式是2:3和2:4,分别占总的异常信号的82%和18%。在CINⅡ或更高病变组,出现了2:5,4:4,5:5,6:6和更复杂的信号类型,且复杂信号类型的比例增加。在SCC组中,出现更复杂的异常信号的模式,如2:5,5:5,6:6,其中,5:5信号类型占总的异常信号模式的22.3%。以FISH结果阳性为标准诊断CINⅡ及以上宫颈病变的敏感性和特异性分别是84.9%、97.1%。以HPV DNA检测结果阳性为标准诊断CINⅡ及以上宫颈病变的敏感性和特异性分别是96.5%、52.9%。与HPV-DNA检测比较,FISH检测诊断CINⅡ及以上宫颈病变的特异性显著提高而敏感性并没有明显降低。
结论
TERC基因的异常扩增在宫颈癌的发展过程中起重要作用。采用FISH技术检测TERC基因扩增,具有较高的灵敏度和特异度,在宫颈癌及其癌前期病变的筛查和早期诊断中有重要临床价值。在细胞学检查和HPV检测筛查宫颈癌的同时,采用FISH技术检测TERC基因扩增,可以显著提高筛查的特异度而不明显降低其灵敏度,有望成为辅助宫颈癌及癌前病变诊断及预测癌前期病变是否向宫颈癌进展的重要指标,为宫颈癌的临床诊断和治疗提供更多的依据。此外,FISH检测TERC基因可以有效地对不同级别的CIN进行区分,因此可以作为一个可靠的检测手段辅助CIN级别的明确诊断,指导CIN的合理治疗。
Background and objectives
Cervical cancer is the second common gynecological tumor worldwide and more prevalent in developing countries, with an incidence ranging from 10 per 100,000 women in industrialized countries to 60 per 100,000 women in some developing countries. The vast majority of cervical cancer has gradually evolved from cervical intraepithelial neoplasia (CIN). CIN is a series of cervical precancerous lesions which are limited to cervical intraepithelial and have different histological changes. It generally takes about 10 years time for the development from CIN to invasive carcinoma. Cervical lesions can be cure eassily if they can be found and given effective treatments in CIN or early invasive carcinoma stage. Therefore, it is imperative to establish effective cancer screening programs, especially in developing countries, in order to reduce the morbidity and mortality of cervical cancer.
Cytopathologic examination and human papillomavirus (HPV) DNA testing are strongly recommended by World Health Organization (WHO) for cervical cancer screening. The conventional Pap smear has been reported to reduce the incidence of cervical cancer by more than 70% in some countries with organized screening program. However, it suffers from relatively low sensitivity and high false negative rate and has variable test results. Compared with conventional Pap smears, the introduction of liquid-based cytology methods has increased the efficacy of population screening but not significantly improved the poor sensitivity. Due to poor sensitivity of cytology, some patients with precancerous lesions can not be detected at early stage and fail to receive effective treatment to progression to invasive cervical cancer.
Infection with HPV is a necessary factor of cervical cancer, which can be detected in 99.7% of invasive cervical carcinomas. So far, more than 100 different HPV types have been identified, and approximately 13 HPV (type 16,18,31,33,35, 39,45,51,52,56,58,59,68,73 and 82) are identified as high risk HPV (HR-HPV) that can transform cells in the genital tract and lead to invasive cancer. Among these HR-HPV types, HPV 16 and 18 are the most commonly identified types in cervical cancer worldwide and strongly related to the development of cervical cancer. The causative relationship between HR-HPV infection and the development of cervical cancer provides potential roles for HPV testing in cervical screening. Hybrid captureⅡ(HCⅡ), approved by the US Food and Drug Administration (FDA), can detected 18 types of HPV, containing 13 high risk types and 5 low-risk types (6,11,42,43 and 44), but can not identify which HPV types are present and discriminate between persistent and transient infections. HPV DNA testing is more reliable and more sensitive, but has less specificity and positive predictive value (PPV) than routinely performed cytology for detection of CINⅡor higher grade lesions (≥CINⅡ). This is because most HPV infections are transient infections that can be cleared spontaneously and do not cause≥CINⅡand only persistent infections with HR-HPV, which accounts for 10-20% of HPV infections, is associated with the development of cervical cancer. Interestingly, most HPV infections are transient infections that can be cleared spontaneously and do not cause≥CINⅡand only persistent infection with HR-HPV is associated with the development of cervical cancer. HPV DNA testing is more reliable and more sensitive but has less specificity and PPV than routinely performed cytology for detection of≥CINⅡ. Thus, more specific biomarkers need to be added to HPV DNA testing to increase the specificity for detecting≥CINⅡ. This may significantly reduce the number of women, particularly those with normal cytology, who are subjected to unnecessary follow-up based on a transient HP-HPV infection. This also may highly reduce the number of women who suffer from over-diagnosis and over-treatment.
In addition to HR-HPV infection, some chromosomal abnormalities and genomic instabilities are necessary for precancerous lesions to progress to invasive cervical cancer. Among these abnormalities, the most common aberration is gain of 3q26 containing the TERC gene which can be detected in approximately 70% of cervical carcinomas. Telomerase is involved in the maintenance of chromosomes by providing telomere stability and regulating telomere length. Telomerase is composed of two main components, an RNA subunit (hTERC) that serves as a template for telomere addition and a protein subunit (hTERT) that acts to catalyzes telomere synthesis. TERC gene encodes for an RNA unit of telomerase that maintains the length of telomeres through cellular divisions. When this gene is overexpressed, the cells avoid undergoing apoptosis, potentially leading to tumorigenesis. Several recent studies have demonstrated that gain of TERC gene as measured by fluorescence in situ hybridization (FISH) occurs frequently in cervical squamous cell carcinoma, high-grade and light-grade cervical intraepithelial neoplasia and that the frequency of TERC gain increase with the severity of cervical lesions, indicating that TERC might be a useful molecular marker for primary cervical screening. Alameda F found that FISH detection of 3q26 amplification can be used as a persistence-progression indicator in low-grade cervical intraepithelial neoplasia. So, TERC amplification detection of FISH may assist in cervical cancer screening and early diagnosis. At present, fluorescence in situ hybridization (FISH) technology is considered as the standard method of detection of TERC gene amplification. FISH technique is simple, reproducible and has high stability and better sensitivity and specificity, so it is suitable for clinical testing. FISH has been widely used in the areas of cellular genetics, tumor biology, gene mapping, gene amplification and prenatal diagnosis.
In the present study, we detected TERC gene amplification of cervical exfoliated epithelial cells by fluorescence in situ hybridization, and compared these results with cytology, HPV results and histopathology results. Our purpose was to assess the clinical significance of FISH detection of TERC amplification in cervical cancer screening and CIN progression prediction. It may provide a new approach for cervical cancer screening, early diagnosis and progression prediction in Chinese women.
Methods/Materials
A total of 120 patients with informed consent were recruited from Nafang Hospital affiliated to Southern Medical University. All of them were the inpatients or outpatients of the hospital from March 2008 to December 2008 and excluded acute pelvic infection, pregnancy, hysterectomy, pelvic radiotherapy and chemotherapy. Each of them was subject to FISH analysis with the TERC-specific probe, HPV DNA testing, liquid-based cytology and histopathological examination in a blind fashion. Cervical cells collected from squamocolumnar junction of the cervix were stored in liquid-based cytology storage solution used for liquid-based cytology and FISH detection and HPV-DNA test dedicated storage solution used for HPV DNA testing, respectively. The biopsy specimens were also taken from the squamocolumnar junction of the cervix under colposcopy-directed. Either cytological or histopathological examination was confirmed by the experienced pathologists. The cytological diagnoses were categorized as follows:normal (21 cases), atypical squamous cells of undetermined significance (ASCUS,8 cases), atypical squamous cells-cannot exclude a high-grade lesion (ASCH,5 cases), low-grade squamous intraepithelial lesion (LSIL,22 cases), high-grade squamous intraepithelial lesion (HSIL,52 cases), and squamous cervical cancer (SCC,12 cases). The histologic diagnoses were categorized as follows:20 cases of normal,14 cases of CINI,35 cases of CINⅡ,36 cases of CINIⅡ, and 15 cases of SCC.
Results
TERC amplification was observed in 0 of 20 (0.00%) of normal and inflammatory cases, in 1 of 14 (7.14%) of CINⅠcases, in 23 of 35 (65.71%) of CINⅡcases, in 35 of 36 (97.22%) of CINⅢcases and in 15 of 15 (100.00%) of SCC cases, respectively. TERC amplification was significantly associated with cervical lesions according to cytological and pathological evaluation (P<0.001), and the positive rate of TERC amplification increased with the increasing severity of cervical lesions. Except of normal and CINⅠ, CINⅡ/Ⅲand SCC, the positive rate of TERC amplification were significantly different between other groups (P<0.005). In normal and inflammatory, CINⅠ, CINⅡ, CINⅢand SCC, the mean of abnormal cells number with TERC amplification was 0.70,3.57,10.97,17.86 and 35.87, respectively. The mean of abnormal cells number with TERC amplification was significantly different among different cervical lesions group classified by histopathological evaluation (P=0.000). Comparison between two groups, the mean of abnormal cells number with TERC amplification was significantly different among any two different cervical lesions groups (P=0.000).000). The complexity of abnormal signal pattern of TERC gene amplification increased as the severity of cervical lesions. In normal control group, the abnormal signal pattern was only 2:3. In CINⅠgroup, the abnormal signal pattern were 2:3 and 2:4, respectively accounting for 82% and 18% of the total abnormal signal pattern. In CINⅡor higher lesions group,2:5,4:4,5:5,6:6 and more complex signal types were found and the proportion of complex signal types increased. In SCC group, the abnormal signal pattern appeared more complex types such as 2:5,5:5,6:6, in which the signal type of 5:5 accounted for 22.3% of the total abnormal signal pattern. With FISH positive results as the standard, the sensitivity and specificity of detection of≥CINⅡlesions were 84.9% and 97.1%, respectively. With HPV DNA positive results as the standard, the sensitivity and specificity of detection of≥CINⅡlesions were 96.5% and52.9%, respectively. The specificity of FISH for detecting CINⅡor more severe cervical lesions (≥CINⅡ) were obviously higher than those of HPV DNA testing.
Conclusions
TERC gene amplification play an important role in the development of cervical cancer. With high sensitivity and specificity, detection of TERC gene amplification using FISH has important clinical value in the screening and early diagnosis of cervical cancer and precancerous lesions. In addition to cytology and HPV testing, detection of TERC gene amplification using FISH method, significantly improve the specificity of screening for cervical cancer without significantly reducing its sensitivity. It may be serve as an important biological marker of screening for cervical cancer and precancerous lesions and a progression indicator of CIN and may provide more evidence for the clinical diagnosis and treatment of cervical cancer. In addition, as FISH detection of TERC gene can effectively distinguish between different levels of CIN, it can be used as a reliable measure to assist in diagnosis of CIN grade and guide the rational treatment of CIN.
引文
[1]Parkin D M, Bray F, Ferlay J, et al. Estimating the world cancer burden: Globocan 2000.[J]. Int J Cancer,2001,94(2):153-156.
[2]Parkin D M, Bray F, Ferlay J, et al. Global cancer statistics,2002.[J]. CA Cancer J Clin,2005,55(2):74-108.
[3]Peto J, Gilham C, Fletcher O, et al. The cervical cancer epidemic that screening has prevented in the UK.[J]. Lancet,2004,364(9430):249-256.
[4]Naryshkin S. The false-negative fraction for Papanicolaou smears:how often are "abnormal" smears not detected by a "standard" screening cytologist?[J]. Arch Pathol Lab Med,1997,121(3):270-272.
[5]Gill G W. Blinded review of Papanicolaou smears.[J]. Cancer,2005,105(2): 53-55.
[6]Jr Wright T C. Cervical cancer screening in the 21st century:is it time to retire the PAP smear?[J]. Clin Obstet Gynecol,2007,50(2):313-323.
[7]Coste J, Cochand-Priollet B, de Cremoux P, et al. Cross sectional study of conventional cervical smear, monolayer cytology, and human papillomavirus DNA testing for cervical cancer screening.[J]. BMJ,2003,326(7392):733.
[8]Zur H H. Papillomaviruses and cancer:from basic studies to clinical application.[J]. Nat Rev Cancer,2002,2(5):342-350.
[9]Walboomers J M, Jacobs M V, Manos M M, et al. Human papillomavirus is a necessary cause of invasive cervical cancer worldwide.[J]. J Pathol,1999, 189(1):12-19.
[10]Cogliano V, Baan R, Straif K, et al. Carcinogenicity of human papillomaviruses. [J]. Lancet Oncol,2005,6(4):204.
[11]Zur H H. Oncogenic DNA viruses.[J]. Oncogene,2001,20(54):7820-7823.
[12]Mayrand M H, Duarte-Franco E, Rodrigues I, et al. Human papillomavirus DNA versus Papanicolaou screening tests for cervical cancer.[J]. N Engl J Med,2007,357(16):1579-1588.
[13]Koshiol J, Lindsay L, Pimenta J M, et al. Persistent human papillomavirus infection and cervical neoplasia:a systematic review and meta-analysis.[J]. Am J Epidemiol,2008,168(2):123-137.
[14]Ostor A G, Mulvany N. The pathology of cervical neoplasia.[J]. Curr Opin Obstet Gynecol,1996,8(1):69-73.
[15]Snijders P J, Steenbergen R D, Heideman D A, et al. HPV-mediated cervical carcinogenesis:concepts and clinical implications.[J]. J Pathol,2006,208(2): 152-164.
[16]Munger K, Baldwin A, Edwards K M, et al. Mechanisms of human papillomavirus-induced oncogenesis.[J]. J Virol,2004,78(21):11451-11460.
[17]Wentzensen N, Vinokurova S, von Knebel D M. Systematic review of genomic integration sites of human papillomavirus genomes in epithelial dysplasia and invasive cancer of the female lower genital tract.[J]. Cancer Res, 2004,64(11):3878-3884.
[18]von Knebel D M. New markers for cervical dysplasia to visualise the genomic chaos created by aberrant oncogenic papillomavirus infections.[J]. Eur J Cancer,2002,38(17):2229-2242.
[19]Graham D A, Southern S A, Mcdicken I W, et al. Interphase cytogenetic evidence for distinct genetic pathways in the development of squamous neoplasia of the uterine cervix.[J]. Lab Invest,1998,78(3):289-296.
[20]Bulten J, Melchers W J, Kooy-Smits M M, et al. Numerical aberrations of chromosome 1 in cervical intraepithelial neoplasia are strongly associated with infection with high-risk human papillomavirus types.[J]. J Pathol,2002, 198(3):300-309.
[21]Hopman A H, Smedts F, Dignef W, et al. Transition of high-grade cervical intraepithelial neoplasia to micro-invasive carcinoma is characterized by integration of HPV 16/18 and numerical chromosome abnormalities.[J]. J Pathol,2004,202(1):23-33.
[22]Melsheimer P, Vinokurova S, Wentzensen N, et al. DNA aneuploidy and integration of human papillomavirus type 16 e6/e7 oncogenes in intraepithelial neoplasia and invasive squamous cell carcinoma of the cervix uteri.[J]. Clin Cancer Res,2004,10(9):3059-3063.
[23]Heselmeyer K, Schrock E, Du Manoir S, et al. Gain of chromosome 3q defines the transition from severe dysplasia to invasive carcinoma of the uterine cervix.[J]. Proc Natl Acad Sci U S A,1996,93(1):479-484.
[24]Heselmeyer-Haddad K, Janz V, Castle P E, et al. Detection of genomic amplification of the human telomerase gene (TERC) in cytologic specimens as a genetic test for the diagnosis of cervical dysplasia. [J]. Am J Pathol,2003, 163(4):1405-1416.
[25]Heselmeyer-Haddad K, Sommerfeld K, White N M, et al. Genomic amplification of the human telomerase gene (TERC) in pap smears predicts the development of cervical cancer.[J]. Am J Pathol,2005,166(4):1229-1238.
[26]Yang Y C, Shyong W Y, Chang M S, et al. Frequent gain of copy number on the long arm of chromosome 3 in human cervical adenocarcinoma.[J]. Cancer Genet Cytogenet,2001,131(1):48-53.
[27]Huang F Y, Kwok Y K, Lau E T, et al. Genetic abnormalities and HPV status in cervical and vulvar squamous cell carcinomas.[J]. Cancer Genet Cytogenet, 2005,157(1):42-48.
[28]Sugita M, Tanaka N, Davidson S, et al. Molecular definition of a small amplification domain within 3q26 in tumors of cervix, ovary, and lung.[J]. Cancer Genet Cytogenet,2000,117(1):9-18.
[29]Cohen S B, Graham M E, Lovrecz G O, et al. Protein composition of catalytically active human telomerase from immortal cells.[J]. Science,2007, 315(5820):1850-1853.
[30]Gilley D, Tanaka H, Herbert B S. Telomere dysfunction in aging and cancer.[J]. Int J Biochem Cell Biol,2005,37(5):1000-1013.
[31]Greider C W. Telomerase activation:one step on the road to cancer?[J].1999, 15(3):109-112.
[32]Wong J M, Collins K. Telomere maintenance and disease.[J]. Lancet,2003, 362(9388):983-988.
[33]Royle N J, Mendez-Bermudez A, Gravani A, et al. The role of recombination in telomere length maintenance.[J]. Biochem Soc Trans,2009,37(Pt 3): 589-595.
[34]Li Y, Li H, Yao G, et al. Inhibition of telomerase RNA (hTR) in cervical cancer by adenovirus-delivered siRNA.[J]. Cancer Gene Ther,2007,14(8): 748-755.
[35]Caraway N P, Khanna A, Dawlett M, et al. Gain of the 3q26 region in cervicovaginal liquid-based pap preparations is associated with squamous intraepithelial lesions and squamous cell carcinoma.[J]. Gynecol Oncol,2008, 110(1):37-42.
[36]Seppo A, Jalali G R, Babkowski R, et al. Gain of 3q26:A genetic marker in low-grade squamous intraepithelial lesions (LSIL) of the uterine cervix[J]. 2009,114(1):80-83.
[37]Hopman A H, Theelen W, Hommelberg P P, et al. Genomic integration of oncogenic HPV and gain of the human telomerase gene TERC at 3q26 are strongly associated events in the progression of uterine cervical dysplasia to invasive cancer.[J]. J Pathol,2006,210(4):412-419.
[38]Caraway N P, Khanna A, Dawlett M, et al. Gain of the 3q26 region in cervicovaginal liquid-based pap preparations is associated with squamous intraepithelial lesions and squamous cell carcinoma[J].2008,110(1):37-42.
[39]Alameda F, Espinet B, Corzo C, et al.3q26 (hTERC) gain studied by fluorescence in situ hybridization as a persistence-progression indicator in low-grade squamous intraepithelial lesion cases.[J]. Hum Pathol,2009.
[40]Bosch F X, Manos M M, Munoz N, et al. Prevalence of human papillomavirus in cervical cancer:a worldwide perspective. International biological study on cervical cancer (IBSCC) Study Group.[J]. J Natl Cancer Inst,1995,87(11): 796-802.
[41]Bosch F X, de Sanjose S. Chapter 1:Human papillomavirus and cervical cancer--burden and assessment of causality.[J]. J Natl Cancer Inst Monogr, 2003(31):3-13.
[42]Arbyn M, Sasieni P, Meijer C J L M, et al. Chapter 9:Clinical applications of HPV testing: A summary of meta-analyses[J].2006,24(Supplement 3): S78-S89.
[43]Ho G Y, Burk R D, Klein S, et al. Persistent genital human papillomavirus infection as a risk factor for persistent cervical dysplasia.[J]. J Natl Cancer Inst, 1995,87(18):1365-1371.
[44]Rosa M I, Medeiros L R, Bozzetti M C, et al. Accuracy of telomerase in cervical lesions:a systematic review.[J]. Int J Gynecol Cancer,2007,17(6): 1205-1214.
[45]Hopman A H, Theelen W, Hommelberg P P, et al. Genomic integration of oncogenic HPV and gain of the human telomerase gene TERC at 3q26 are strongly associated events in the progression of uterine cervical dysplasia to invasive cancer.[J]. J Pathol,2006,210(4):412-419.
[46]Takac I, Arko D, Kodric T, et al. Human telomerase gene amplification and high-risk human papillomavirus infection in women with cervical intra-epithelial neoplasia.[J]. J Int Med Res,2009,37(5):1588-1595.
[47]Caraway N P, Khanna A, Dawlett M, et al. Gain of the 3q26 region in cervicovaginal liquid-based pap preparations is associated with squamous intraepithelial lesions and squamous cell carcinoma.[J]. Gynecol Oncol,2008, 110(1):37-42.
[48]Mehes G, Speich N, Bollmann M, et al. Chromosomal aberrations accumulate in polyploid cells of high-grade squamous intraepithelial lesions (HSIL).[J]. Pathol Oncol Res,2004,10(3):142-148.
[49]Kokalj-Vokac N, Kodric T, Erjavec-Skerget A, et al. Screening of TERC gene amplification as an additional genetic diagnostic test in detection of cervical preneoplastic lesions.[J]. Cancer Genet Cytogenet,2009,195(1):19-22.
[50]Jr Wright T, Massad L S, Dunton C J, et al.2006 consensus guidelines for the management of women with abnormal cervical screening tests.[J]. J Low Genit Tract Dis,2007,11(4):201-222.
[51]Riethdorf S, Riethdorf L, Schulz G, et al. Relationship between telomerase activation and HPV 16/18 oncogene expression in squamous intraepithelial lesions and squamous cell carcinomas of the uterine cervix.[J]. Int J Gynecol Pathol,2001,20(2):177-185.
[52]Reesink-Peters N, Helder M N, Wisman G B, et al. Detection of telomerase, its components, and human papillomavirus in cervical scrapings as a tool for triage in women with cervical dysplasia.[J]. J Clin Pathol,2003,56(1):31-35.
[1]Parkin D M, Bray F. Chapter 2:The burden of HPV-related cancers[J]. Vaccine. 2006,24 Suppl 3:S3-S11.
[2]Cohen S B, Graham M E, Lovrecz G O, et al. Protein composition of catalytically active human telomerase from immortal cells[J]. Science.2007, 315(5820):1850-1853.
[3]Nakamura T M, Morin G B, Chapman K B, et al. Telomerase catalytic subunit homologs from fission yeast and human[J]. Science.1997,277(5328):955-959.
[4]Feng J, Funk W D, Wang S S, et al. The RNA component of human telomerase[J]. Science.1995,269(5228):1236-1241.
[5]Mitchell J R, Wood E, Collins K. A telomerase component is defective in the human disease dyskeratosis congenita[J]. Nature.1999,402(6761):551-555.
[6]Kirwan M, Dokal I. Dyskeratosis congenita:a genetic disorder of many faces[J]. Clin Genet.2008,73(2):103-112.
[7]Colgin L M, Reddel R R. Telomere maintenance mechanisms and cellular immortalization[J]. Curr Opin Genet Dev.1999,9(1):97-103.
[8]Royle N J, Mendez-Bermudez A, Gravani A, et al. The role of recombination in telomere length maintenance[J]. Biochem Soc Trans.2009,37(Pt 3):589-595.
[9]Rooney P H, Murray G I, Stevenson D A, et al. Comparative genomic hybridization and chromosomal instability in solid tumours[J]. Br J Cancer. 1999,80(5-6):862-873.
[10][Mosse Y P, Greshock J, Margolin A, et al. High-resolution detection and mapping of genomic DNA alterations in neuroblastoma[J]. Genes Chromosomes Cancer.2005,43(4):390-403.
[11]Coe B P, Henderson L J, Garnis C, et al. High-resolution chromosome arm 5p array CGH analysis of small cell lung carcinoma cell lines[J]. Genes Chromosomes Cancer.2005,42(3):308-313
[12]Sugita M, Tanaka N, Davidson S, et al. Molecular definition of a small amplification domain within 3q26 in tumors of cervix, ovary, and lung[J]. Cancer Genet Cytogenet.2000,117(1):9-18.
[13]Grm H S, Bergant M, Banks L. Human papillomavirus infection, cancer & therapy[J]. Indian J Med Res.2009,130(3):277-285.
[14]Bhatla N, Moda N. The clinical utility of HPV DNA testing in cervical cancer screening strategies[J]. Indian J Med Res.2009,130(3):261-265.
[15]Walboomers J M, Jacobs M V, Manos M M, et al. Human papillomavirus is a necessary cause of invasive cervical cancer worldwide[J]. J Pathol.1999, 189(1):12-19.
[16]Wentzensen N, Vinokurova S, von Knebel D M. Systematic review of genomic integration sites of human papillomavirus genomes in epithelial dysplasia and invasive cancer of the female lower genital tract[J]. Cancer Res.2004,64(11): 3878-3884.
[17]von Knebel D M. New markers for cervical dysplasia to visualise the genomic chaos created by aberrant oncogenic papillomavirus infections[J]. Eur J Cancer. 2002,38(17):2229-2242.
[18]Graham D A, Southern S A, Mcdicken I W, et al. Interphase cytogenetic evidence for distinct genetic pathways in the development of squamous neoplasia of the uterine cervix[J]. Lab Invest.1998,78(3):289-296.
[19]Bulten J, Melchers W J, Kooy-Smits M M, et al. Numerical aberrations of chromosome 1 in cervical intraepithelial neoplasia are strongly associated with infection with high-risk human papillomavirus types[J]. J Pathol.2002,198(3): 300-309.
[20]Hopman A H, Smedts F, Dignef W, et al. Transition of high-grade cervical intraepithelial neoplasia to micro-invasive carcinoma is characterized by integration of HPV 16/18 and numerical chromosome abnormalities[J]. J Pathol. 2004,202(1):23-33.
[21]Melsheimer P, Vinokurova S, Wentzensen N, et al. DNA aneuploidy and integration of human papillomavirus type 16 e6/e7 oncogenes in intraepithelial neoplasia and invasive squamous cell carcinoma of the cervix uteri[J]. Clin Cancer Res.2004,10(9):3059-3063.
[22]Heselmeyer K, Schrock E, Du Manoir S, et al. Gain of chromosome 3q defines the transition from severe dysplasia to invasive carcinoma of the uterine cervix[J]. Proc Natl Acad Sci U S A.1996,93(1):479-484.
[23]Heselmeyer-Haddad K, Janz V, Castle P E, et al. Detection of genomic amplification of the human telomerase gene (TERC) in cytologic specimens as a genetic test for the diagnosis of cervical dysplasia[J]. Am J Pathol.2003, 163(4):1405-1416.
[24]Heselmeyer-Haddad K, Sommerfeld K, White N M, et al. Genomic amplification of the human telomerase gene (TERC) in pap smears predicts the development of cervical cancer[J]. Am J Pathol.2005,166(4):1229-1238.
[25]Yang Y C, Shyong W Y, Chang M S, et al. Frequent gain of copy number on the long arm of chromosome 3 in human cervical adenocarcinoma[J]. Cancer Genet Cytogenet.2001,131(1):48-53.
[26]Huang F Y, Kwok Y K, Lau E T, et al. Genetic abnormalities and HPV status in cervical and vulvar squamous cell carcinomas[J]. Cancer Genet Cytogenet. 2005,157(1):42-48.
[27]Mehes G, Speich N, Bollmann M, et al. Chromosomal aberrations accumulate in polyploid cells of high-grade squamous intraepithelial lesions (HSIL)[J]. Pathol Oncol Res.2004,10(3):142-148.
[28]Hopman A H, Theelen W, Hommelberg P P, et al. Genomic integration of oncogenic HPV and gain of the human telomerase gene TERC at 3q26 are strongly associated events in the progression of uterine cervical dysplasia to invasive cancer[J]. J Pathol.2006,210(4):412-419.
[29]Naryshkin S. The false-negative fraction for Papanicolaou smears:how often are "abnormal" smears not detected by a "standard" screening cytologist?[J]. Arch Pathol Lab Med.1997,121(3):270-272.
[30]Gill G W. Blinded review of Papanicolaou smears.[J]. Cancer.2005,105(2): 53-55.
[31]Jr Wright T C. Cervical cancer screening in the 21st century:is it time to retire the PAP smear?[J]. Clin Obstet Gynecol.2007,50(2):313-323.
[32]Coste J, Cochand-Priollet B, de Cremoux P, et al. Cross sectional study of conventional cervical smear, monolayer cytology, and human papillomavirus DNA testing for cervical cancer screening[J]. BMJ.2003,326(7392):733.
[33]Mayrand M H, Duarte-Franco E, Rodrigues I, et al. Human papillomavirus DNA versus Papanicolaou screening tests for cervical cancer[J]. N Engl J Med. 2007,357(16):1579-1588.
[34]Soder A I, Hoare S F, Muir S, et al. Amplification, increased dosage and in situ expression of the telomerase RNA gene in human cancer[J]. Oncogene.1997, 14(9):1013-1021.
[35]Caraway N P, Khanna A, Dawlett M, et al. Gain of the 3q26 region in cervicovaginal liquid-based pap preparations is associated with squamous intraepithelial lesions and squamous cell carcinoma[J]. Gynecol Oncol.2008, 110(1):37-42.
[36]Seppo A, Jalali G R, Babkowski R, et al. Gain of 3q26:A genetic marker in low-grade squamous intraepithelial lesions (LSIL) of the uterine cervix[J].2009, 114(1):80-83.
[37]Hopman A H, Theelen W, Hommelberg P P, et al. Genomic integration of oncogenie HPV and gain of the human telomerase gene TERC at 3q26 are strongly associated events in the progression of uterine cervical dysplasia to invasive cancer[J]. J Pathol.2006,210(4):412-419.
[38]Caraway N P, Khanna A, Dawlett M, et al. Gain of the 3q26 region in cervicovaginal liquid-based pap preparations is associated with squamous intraepithelial lesions and squamous cell carcinoma[J].2008,110(1):37-42.
[39]Zhang Y, Wang X, Ma L, et al. Clinical significance of hTERC gene amplification detection by FISH in the screening of cervical lesions[J]. J Huazhong Univ Sci Technolog Med Sci.2009,29(3):368-371.
[40]Heselmeyer-Haddad K, Sommerfeld K, White N M, et al. Genomic amplification of the human telomerase gene (TERC) in pap smears predicts the development of cervical cancer[J]. Am J Pathol.2005,166(4):1229-1238.
[41]Alameda F, Espinet B, Corzo C, et al.3q26 (hTERC) gain studied by fluorescence in situ hybridization as a persistence-progression indicator in low-grade squamous intraepithelial lesion cases[J]. Hum Pathol.2009.
[42]Andersson S, Wallin K L, Hellstrom A C, et al. Frequent gain of the human telomerase gene TERC at 3q26 in cervical adenocarcinomas[J]. Br J Cancer. 2006,95(3):331-338.
[43]Yatabe N, Kyo S, Kondo S, et al.2-5A antisense therapy directed against human telomerase RNA inhibits telomerase activity and induces apoptosis without telomere impairment in cervical cancer cells[J]. Cancer Gene Ther. 2002,9(7):624-630.
[44]Li Y, Li H, Yao G, et al. Inhibition of telomerase RNA (hTR) in cervical cancer by adenovirus-delivered siRNA[J]. Cancer Gene Ther.2007,14(8):748-755.
[45]Liu B, Qu Y, Liu S, et al. Inhibition of telomerase in tumor cells by ribozyme targeting telomerase RNA component[J]. Sci China C Life Sci.2002,45(1): 87-95.
[46]赵家明,李明意,杨展,等.脱氧核酶对鼻咽癌细胞端粒酶活性的抑制作用[J].华中科技大学学报(医学版)[J].2005(3).
