Genotypic Analysis of Epstein-Barr Virus Associated with Nasopharyngeal Carcinoma of Japanese Patients

Types and certain genetic markers were studied for Epstein-Barr virus present in 10 specimens of nasopharyngeal carcinoma (NPC) from Japanese patients. The type 1 virus was predominant in our NPC specimens, as in the general Japanese population, and the type 2 virus was found only in one NPC specimen. The type C variant, which lacks the Bam HI site between the Bam HI-W1* and -I1* regions, appeared to be common among Japanese strains as in those in Southern China. The type f variant which has an extra Bam HI site in the Bam HI-F region and has been shown to be strongly associated with NPC in Southern China was found in only one NPC specimen. This virus strain was also the only type 2 virus among our specimens. The present study, therefore, does not show any specific association of the type f variant with NPC in Japan.     

鼻咽癌中EB病毒LMP1基因N端XhoⅠ酶切位点的丢失

背景与目的:众所周知, EB病毒 LMP1基因在鼻咽癌变过程起着一定的作用.本研究通过检测广东地区鼻咽癌组织 EB病毒 LMP1基因 N-末端区 XhoⅠ酶切位点的丢失,探讨 LMP1基因变异在鼻咽癌发生发展中的作用.方法 :收集中山大学肿瘤防治中心鼻咽癌患者鼻咽新鲜活检标本 63例.收集 EB病毒健康携带者外周血单个核细胞 (PBMCs) 10例作为对照.采用 QIAamp DNA Mini Kit和 QIAamp DNA Blood Mini Kit分别抽取组织和外周血单个核细胞的 DNA,应用巢式 PCR扩增 EB病毒 LMP1基因的 N-末端区,并用 XhoⅠ对扩增产物进行酶切.采用四色荧光 末端终止法对扩增产物进行序列分析.结果: 10例健康携带者外周血单个核细胞的 EB病毒 LMP1基因 N-末端区均未见 XhoⅠ酶切位点的丢失.63例鼻咽癌组织中有 50例(79.37%)出现 XhoⅠ酶切位点的丢失(XhoⅠ-loss),还有 4例(6.34%)为 XhoⅠ酶切位点部分丢失,只有 9例(14.29%)未见 XhoⅠ酶切位点的丢失(wt-XhoⅠ ).除了 XhoⅠ酶切位点的丢失 (nt: 169423～169428; GAGCTC→ GATCTC)外,还发现四个错义点突变.结论:本研究所检测的广东地区 EB病毒健康携带者外周血单个核细胞所携带的 EB病毒 LMP1基因为 wt-XhoⅠ,而在鼻咽癌组织中主要为 XhoⅠ-loss.因此,我们认为 EB病毒 LMP1基因 N-末端区 XhoⅠ酶切位点的丢失和其他的错义点突变可能是在鼻咽癌的发生发展过程中产生的.     

Expression of Epstein Barr Virus Encoded EBNA1 and LMP1 Oncoproteins in Nasopharyngeal Carcinomas from Northeast India

Background: Nasopharyngeal carcinoma (NPC), a malignancy arising from the epithelial lining of the nasopharynx, is distinct from others cancers in terms of its epidemiologic features. It is rare in most parts of the world except for a few regions with populations of Mongoloid origin. Objectives: To study the expression pattern of Epstein Barr virus (EBV) encoded oncoproteins EBNA1 and LMP1 in different histological types of NPC and to correlate expression patterns with sex, age and histological types. Materials and Methods: A total of 40 formalinfixed, paraffinembedded NPC biopsy samples and tissues from 20 healthy controls were collected to study the expression level of EBNA1 and LMP1 using immunohistochemistry. Results: EBNA1 and LMP1 expression was found in 92.5% and 90% respectively, of the cases and none of the control specimens. The expression patterns of EBNA1 and LMP1 were determined to be statistically significant (p<0.05) when correlated with sex, age and histological distributions. Also immunohistochemistry was found to be a sensitive technique in the detection of EBV. Conclusions: The study reveals that the potent oncoproteins EBNA1 and LMP1 were over expressed in our population cohort. Our findings are to some extent inconsistent with earlier reports as our population showed a higher expression of both EBNA1 and LMP1 compared to other studies.     

Impaired Long-term T Cell Immunity to Epstein-Barr Virus in Patients with Nasopharyngeal Carcinoma

The long-term T cell immunity to Epstein-Barr virus (EBV) is considered to play an important role in suppressing proliferation of EBV-infected B cells and outgrowth of EBV-associated tumors. It can be manifested and quantified by the EBV-induced focus regression assay. In the present study, we examined the strength of T cell immunity to EBV in patients with nasopharyngeal carcinoma (NPC) and other cancers originating from the head and neck region. In contrast to patients with other types of cancers, including EBV-negative NPC, patients with EBV-positive NPC were found to have a profound impairment in the long-term T cell immunity to EBV.     

鼻咽癌患者EB病毒与乙肝病毒双重感染的观察

目的：探讨鼻咽癌患者ＥＢ病毒和乙肝病毒感染的关系。方法检测５０５１健康康者和１２５例鼻咽癌患者的血清ＥＢＶ－ＩｇＡ／ＶＣＡ和ＨＢｓＡｇ。结果健康人群的ＥＢＶ－ＩｇＡ／ＶＣＡ和ＨＢｓＡｇ的阳性率分别为５．４％和１１．３％;健康人群ＥＢ病毒感染者的ＨＢｓＡｇ阳性率为９．７％;鼻咽癌患者的ＥＢＶ－ＩｇＡ／ＶＣＡ和ＨＢｓＡｇ的阳性率分别为９７．６％、２５．６％。鼻咽癌ＥＢ病毒、乙肝病毒双重感染率明显高于正     

鼻咽癌患者血浆游离EBV/DNA的定量检测及其临床意义

目的:探讨血浆EBV/DNA定量分析,在鼻咽癌早期诊断、临床分期、预后判断和监测放疗后转移复发中的临床意义.方法:采用荧光定量PCR方法定量检测经病理确诊为鼻咽癌的120例初治、90例放疗后随诊患者,其中包括60例放疗后持续缓解,30例远处转移和局部复发患者的血浆EBV/DNA含量.结果:初治、远处转移和局部复发的鼻咽癌患者血浆中游离的EBV/DNA检出率分别为96.0%、95.0%和100%,显著高于治疗后持续缓解鼻咽癌患者、健康对照者和非鼻咽癌的肿瘤患者;初治鼻咽癌患者各TNM分期之间血浆EBV/DNA拷贝数有显著统计学差异,晚期患者(Ⅲ Ⅳ)期血浆EBV/DNA中位拷贝数显著高于早期患者(I Ⅱ)期;初治患者治疗后已出现局部和远处转移者.治疗前血浆EBV/DNA中位数显著高于尚未出现复发转移患者:初治患者治疗前血浆EBV/DNA≥40 000拷贝/ml与＜40 000拷贝/ml两个水平,患者22个月无复发生存率分别为46.1%和92.9%,有显著统计学差异;放疗后复发、转移鼻咽癌患者血浆EBV/DNA的中位拷贝数显著高于治疗后持续缓解患者.结论:采用荧光定量PCR方法检测鼻咽癌患者血浆中游离的EBV/DNA是一种敏感可靠的方法,对于鼻咽癌早期诊断、鉴别诊断、分期、判断预后、监测治疗后复发和远处转移具有重要的临床意义,有可能成为鼻咽癌的血清肿瘤标记物.     

Rearranged Epstein-Barr Virus Genomes and Clonal Origin in Nasopharyngeal Carcinoma

Epstein-Barr virus (EBV) is known to be associated with two malignant diseases, nasopharyngeal carcinoma (NPC) and endemic Burkitt's lymphoma. In this study, the genomes of EBV in biopsy specimens from 4 NFC patients in Japan were analyzed using Southern blot hybridization. The NPC tissues of all examined cases contained rearranged EBV genomes whose Bam HI H fragments were larger than those of prototype EBV genomes. One of them had a Bam HI fragment containing contiguous sequences of Bam HI Y and H. A single-sized EBV DNA terminus was observed in these NPC tissues, implying the evolution of the carcinoma from a single EBV-infected cell.     

鼻咽癌患者血浆EB病毒DNA水平与肿瘤复发的关系

目的 :探讨鼻咽癌患者放射治疗后血浆EB病毒 (EBV)DNA水平与肿瘤复发的关系。方法 :11例临床缓解期患者和 9例临床复发患者分别抽血 3ml。所有的标本均采用荧光定量PCR的方法 ,在PE770 0型检测仪上定量检测血浆标本中EB病毒DNA的含量。结果 :肿瘤复发组 89% ( 8/ 9)的患者血浆中可检测到高拷贝数的EB病毒DNA ,中位浓度为4 70 0 0 0 (copies/ml) ,在临床缓解组患者中 ,仅 9% ( 1/ 11)的患者血浆中可检测到较高拷贝数的EB病毒DNA ,中位浓度为 0(copies/ml)。两组阳性率及中位浓度的比较均有显著性差异 (P <0 0 0 1)。结论 :鼻咽癌患者血浆EBVDNA水平与肿瘤复发关系密切 ,值得进一步研究。     

辅助化疗对高危远处转移鼻咽癌患者生存率的影响

目的　研究放疗后辅助化疗对高危远处转移鼻咽癌 (NPC)患者 5年生存率的影响。方法　将 164例高危远处转移鼻咽癌患者随机分为单纯放疗组和放疗后辅助化疗组。单纯放疗组 82例 ,患者采用常规分割放疗 ,鼻咽及转移性淋巴结DT70Gy/7W ,邻近淋巴引流区预防量 5 0Gy ;放疗后辅助化疗组 82例 ,患者放疗后采用顺铂为主联合氟脲嘧啶 (PF方案 )或足叶乙甙(EP方案 )等方案化疗 3～ 6个周期。结果　N2～ 3 期NPC(鳞癌 )患者中 ,单纯放疗组、放疗后辅助化疗组 5年生存率分别为 45 .2 %、63 .8% ,两者有显著性差异 (P <0 .0 5 ) ;鼻咽未分化癌患者中单纯化疗组、放疗后辅助化疗组 5年生存率分别为 47.1%、5 3 .8% ,两者无显著性差异。结论　对N2～ 3 期NPC(鳞癌 )患者放疗后辅以化疗可提高患者 5年生存率     

鼻咽癌血浆EB病毒DNA水平与肿瘤负荷的关系

目的探讨鼻咽癌血浆EB病毒(EBV)DNA水平与肿瘤负荷的关系。方法收集经病理检查证实、无远处转移的初治鼻咽癌患者共98例,治疗前行鼻咽及全颈CT,应用PACS中的多边形测量工具,逐层勾画鼻咽原发灶肿瘤及颈部转移淋巴结,采用面积求和法计算鼻咽原发灶肿瘤体积(TV)及颈部转移淋巴结体积(NV),以TV和NV之和表示总肿瘤负荷(TNV)。应用荧光定量PCR技术检测其治疗前血浆EBV DNA水平。结果全组TV、NV及TNV中位数(四分位数间距)分别为15.9cm3(9.1~29.2cm3)、17.1cm3(0.7~64.1cm3)、44.1cm3(20.4~87.4cm3)。全组血浆EBV DNA检出率为73.5%(72/98),血浆EBVDNA水平中位数(四分位数间距)为3.6×104 copies/ml(2.0×103~3.6×105 copies/ml)。按TV分组:≤20cm3、>20cm3者各58、40例,2组血浆EBV DNA水平中位数(四分位数间距)分别为9.5×103 copies/ml(0~1.6×105 copies/ml)、1.6×105 copies/ml(2.2×104~4.6×105 copies/ml),有非常显著性差异(P=0.001);按NV分组:≤20cm3、>20cm3者各52、46例,2组血浆EBV DNA水平中位数(四分位数间距)分别为2.2×104 copies/ml(0~1.8×105 copies/ml)、1.0×105 copies/ml(6.2×103~5.4×105 copies/ml),有显著性差异(P=0.033);按TNV分组:≤40cm3、>40cm3者各43、55例,2组血浆EBVDNA检出率分别为60.5%(26/43)、83.6%(46/55),有显著性差异(P=0.010);2组血浆EBVDNA水平中位数(四分位数间距)分别为1.7×104 copies/ml(0~1.3×105 copies/ml)、1.1×105 copies/ml(6.6×103~4.7×105 copies/ml),有显著性差异(P=0.014);血浆EBV DNA水平与TV、NV、TNV呈正相关关系。结论鼻咽癌患者治疗前血浆EBV DNA水平可在一定程度上反映其体内肿瘤负荷。     

鼻咽癌患者白细胞介素—2活性变化的研究

选出鼻咽癌病例及正常人对照各32例。测定放疗前后外周血IL-2活性。结果表明:病例组放疗前IL-2活性显著低下,放疗后明显回升,但仍低于对照组(P<0.01)。4例肿瘤残存者IL-2活性回升不良。讨论认为:肿瘤负荷可能是抑制IL-2活性的主要因素,IL-2活性检测可能作为监测肿瘤负荷及估计肿瘤患者预后的指标。     

趋化因子受体CXCR4在鼻咽癌中的表达及意义

目的探讨趋化因子受体CXCR4在人鼻咽癌中的表达及临床意义。方法采用Real-timeRT-PCR法检测鼻咽癌细胞株中CXCR4mRNA的表达,应用免疫组织化学法并结合组织阵列检测正常鼻咽组织、鼻咽癌和鼻咽癌淋巴结转移石蜡组织标本中CXCR4蛋白的表达情况。结果在7种鼻咽癌细胞株中,CXCR4基因在高成瘤、高转移潜能的5-8F细胞中表达水平最高,在无转移能力的610B细胞中表达最低。CXCR4蛋白在正常鼻咽组织、鼻咽癌和鼻咽癌淋巴结转移组织中表达差异具有统计学意义（P=0．002）;鼻咽癌组织中CXCR4表达高于正常鼻咽组（P=0．029）;伴发转移的鼻咽癌组织比未发生转移的鼻咽癌组织CXCR4表达增强,差异具有统计学意义（P=0．008）;淋巴结转移癌组织CX-CR4表达比鼻咽癌组织高（P=0．013）。结论本研究提示CXCR4表达与鼻咽癌转移存在密切关系,CXCR4表达可作为鼻咽癌转移过程中一个有价值的指标。     

Real-Time PCR Assay for Rapid Detection of GSTM1 Polymorphism in Nasopharyngeal Carcinoma Patients

Nasopharyngeal carcinoma (NPC) is a common public health problem in Thailand. Glutathione S-transferaseM1 gene deletion (GSTM1 null genotype) carriers have been reported to be at increased risk and therefore thisparameter is a potential marker for screening of NPC high-risk individuals. However, the conventional polymerasechain reaction (C-PCR) assay commonly used for GSTM1 null genotype detection is not suitable for mass screeningsince it is inconvenient, time consuming and unsafe due to the use of a toxic chemical. Currently, real-time PCR(R-PCR) assay is recommended for quicker and safer detection of various genetic polymorphisms. The aim ofthis study was to develop a SYBR green I R-PCR assay combined with melting curve analysis for GSTM1polymorphism detection in Thai NPC patients. The results were compared to those from the C-PCR assay usingDNA samples from peripheral blood leukocytes of 120 Thai NPC patients. The frequencies of GSTM1polymorphism detected by the R-PCR and the C-PCR were the same. Forty-eight individuals that were GSTM1+in the R-PCR assay showed 2 peaks with melting points of 82.5˚C and 87.5˚C that correlated with the appearanceof 2 DNA bands in the C-PCR assay (i.e., one for GSTM1 at 215 base pairs (bp) and one for β-globin at 268 bp).By contrast, 72 individuals that were GSTM1- in the R-PCR assay showed 1 peak with a melting point of 87.5˚Cthat correlated with the appearance of 1 DNA band for β-globin at 268 bp in the C-PCR assay. The R-PCR assayusing SYBR Green I and melting curve analysis for GSTM1 polymorphism detection was as reliable as the CPCRassay but was quicker and safer and more amenable to large scale screening in Thai NPC cases.     

2002—2013年上海市浦东新区居民鼻咽癌发病情况及其趋势分析

目的 分析上海市浦东新区居民2002—2013年鼻咽癌发病情况并预测其发展趋势,为鼻咽癌防治策略提供参考。方法 以2002—2013年上海市浦东新区常住户籍居民为研究对象,按1985年世界标准人口计算鼻咽癌标化发病率（age-standardized rate, ASR）,应用Join-point regression program分析率值的年均变化百分比（annual percent change, APC）进行趋势分析。结果 2002—2013年浦东新区新发鼻咽癌1 254例,男女发病人数比为2.63:1,合计粗发病率为3.97/10万人年,世界标化发病率为2.29/10万人年,逐年下降的趋势明显（APC=-5.89%, Z=4.77, P<0.001）;男性粗发病率为5.76/10万人年,世界标化发病率为3.36/10万人年,逐年下降的趋势明显（APC=-6.19%, Z=4.73, P<0.001）;女性粗发病率为2.18/10万人年,世界标化发病率为1.25/10万人年,逐年下降的趋势明显（APC=-5.56%,Z=3.87, P=0.003）; 居民鼻咽癌发病率在65~69岁时达到最高峰（11.52/10万人年）;男性鼻咽癌发病率在70~74岁时达到最高峰（18.56/10万人年） ; 女性鼻咽癌发病率在60~64岁时达到最高峰（6.17/10万人年）。结论 浦东新区鼻咽癌发病率近年来呈明显下降趋势, 相关部门应针对重点人群积极开展鼻咽癌病因学研究并应用早期诊断技术以进一步降低其发病率。     

血浆 EB病毒游离 DNA检测对监测鼻咽癌患者预后的意义

背景与目的:有报道 , 测定血浆中的 EB病毒游离 DNA( EBV-DNA)的拷贝数可作为诊断及监测鼻咽癌患者病情变化的手段之一.本研究旨在评价血浆 EBV-DNA检测在鼻咽癌患者预后监测上的价值, 并进一步与 VCA/IgA、 EA/IgA进行比较.方法:比较鼻咽癌放疗后 30例远处转移患者、 22例局部复发患者、 24例无 瘤生存者血浆中 EBV-DNA、 VCA/IgA、 EA/IgA水平.分别应用荧光定量 PCR方法检测血浆 EBV-DNA水平,免疫酶法检测 VCA/IgA、 EA/IgA;前瞻性观察 20例初诊鼻咽癌患者放疗前、放疗剂量达 40 Gy时及放疗结束时上述指标的变化. 结果:放疗后各组不同预后患者的血浆 EBV-DNA含量的中位数有显著性差异, 远处转移组为 135 100 copies/ml(四分线区域 5 525～ 1 003 750 copies/ml) >局部复发组的 20 500(四分线区域 0～ 58 500 copies/ml) > 无瘤生存组的 0 copy/ml(四分线区域 0～ 0 copy/ml), P均 < 0.05. 远处转移组的血浆 EBV-DNA水平高者较多, 当阳性标准为 1 000 000 copies/ml时,诊断远处转移组的敏感性为 27.3%,而诊断局部复发组的敏感性为 0.0%,特异性均为 100.0%.在初诊患者放疗前、放疗剂量达 40 Gy时及放疗结束时, EBV-DNA水平逐渐降低,平均含量分别为 32 050 copies/ml(四分线区域 3 880～ 317 750 copies/ml)、 0 copy/ml(四分线区域 0～ 14 375 copies/ml)、 0 copy/ml(四分线区域 0～ 2 940 copies/ml), P均 < 0.05, 而 VCA/IgA、 EA/IgA的水平未见明显变化. 结论: 血浆 EBV-DNA检测可用于监测鼻咽癌患者预后,其价值明显优于 VCA/IgA、 EA/IgA.     

XRCC1单核苷酸多态性与鼻咽癌铂类化疗敏感度的相关性

目的 探讨鼻咽癌铂类化疗敏感度与X射线交错互补修复基因1 codon194和codon399单核苷酸多态性的相关性。方法 收集广西医科大学第四附属医院2012年9月1日至2013年12月31日鼻咽部肿物活检确诊为鼻咽癌患者资料,采用限制性片段长度多态性聚合酶链反应技术检测鼻咽癌患者外周血DNA XRCC1 codon194和codon399单核苷酸多态性。顺铂+氟尿嘧啶方案诱导化疗2周期后复查MRI,按照RECIST 1.1标准评价其化疗敏感度,分析单核苷酸多态性（Single nucleotide polymorphism,SNP）与化疗敏感度的关系。结果 XRCC1 codon399 Gln/Gln基因型携带者化疗敏感度为Arg/Arg基因型携带者的3.500倍（P＜0.05）。XRCC1 codon399不含Arg基因型（即Gln/Gln）携带者化疗敏感度为含Arg基因型（Arg/Arg 和 Arg/Gln）携带者的3.274倍,（P＜0.05）。携带XRCC1 codon194各基因型患者化疗敏感度之间差异无明显统计学意义（P＞0.05）。结论 XRCC1 codon399 单核苷酸多态性有可能成为鼻咽癌铂类化疗敏感度的预测因子。     

分割放疗对鼻咽癌组织细胞增殖凋亡的影响

目的：探讨常规分割放疗和超分割放疗对鼻咽癌组织细胞增殖凋亡的影响。方法：采用ＴｄＴ酶介导的生物素化ｄｕｔｐ缺口末端标记技术（ＴＵＮＥＬ）和免疫组织化学Ｓ－Ｐ法,分别检测６０例鼻咽癌患者在放疗第２周和第４周时细胞凋亡率（ａｐｏｐｔｏｓｉｓ ｒａｔｅ,ＡＲ）和增殖细胞核抗原（ｐｒｏｌｉｆｅｒａｔｉｏｎ ｃｅｌｌ ｎｈｕｃｌｅａｒ ａｎｔｉｇｅｎ,ＰＣＮＡ）的表达。结果：常规分割放疗组放疗第４周后的ＡＲ     

Determination and Role of Epstein-Barr Virus in Patients With Lymphoproliferative Disorders

IntroductionEpstein-Barr virus (EBV) is associated with different types of human malignancies, including Burkitt lymphoma, nasopharyngeal carcinoma, and lymphomas. We retrospectively investigated the presence of EBV-DNA by real-time PCR in clinical samples of patients diagnosed as having hematologic malignancies while investigating the cause of lymphoproliferative disorders, and investigated its relationship to clinical manifestations.Patients and MethodsFifty clinical samples sent to Gazi University’s hematology clinics between November 2013 and March 2018 were included. EBV-DNA was investigated by real-time PCR method, and EBV-IgM and EBV-IgG antibodies were investigated by ELISA.ResultsFifty serum samples were investigated, and 10% (5/50) EBV-DNA positivity was determined in patients. Of the 5 patients with EBV-DNA positivity, 2 had acute lymphoblastic leukemia, 1 lymphoma, 1 T-cell lymphoma, and 1 B-cell lymphoma. Concomitant EBV-DNA and viral capsid antigen (VCA)-IgM positivity was not detected. The VCA-lgM test results of the all EBV-DNA–positive patients were negative and VCA-IgG positive (except for 1 patient). Regarding virus load, of the 5 samples, 2, 1, 1, and 1 of the samples had a virus load of 10², 10³, 10⁴, and 10⁵ copies/mL, respectively.ConclusionEBV infection is threatening in patients with hematologic malignancies and are diagnosed by serologic and molecular methods. As a result of the study, we suggest that the detection of EBV-DNA by real-time PCR in patients being admitted with lymphoproliferative diseases and diagnosed as acute lymphoblastic leukemia and lymphomas may be useful in follow-up and treatment.     

广东籍鼻咽癌患者家族聚集性研究

目的研究高发区广东省鼻咽癌的家族聚集性规律,为反映高发区鼻咽癌的遗传流行病学特点及遗传咨询提供资料。方法收集1998年1月~2000年8月在中山大学肿瘤防治中心治疗的全部广东籍初诊鼻咽癌患者共1142例的病案资料。按照事先制定的调查表,收集患者的一般资料及家族中的肿瘤情况。结果在广东籍人群中,21.9%的鼻咽癌患者具有肿瘤家族史,其中12.3%的患者有鼻咽癌家族史。家族中肿瘤患者70%左右发生于一级亲属中,家族中鼻咽癌患者在父母和兄弟姐妹中发生的比率相当。广东省内高发区的患者亲属同患鼻咽癌的比例为19.8%,明显高于省内其他地区同患鼻咽癌的比例8.5%,χ2=0.236,P<0.01。结论在鼻咽癌高发区广东省,鼻咽癌具有显著的家族聚集性,而且越高发的地区其家族聚集性越强。