鼻咽癌中EB病毒LMP1基因N端Xho I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Xho I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Xho I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Xho I酶切位点的丢失。63例鼻咽癌组织中有50例(79．37％)出现Xho I酶切位点的丢失(Xho I—loss),还有4例(6．34％,)为Xho I酶切位点部分丢失,只有9例(14．29％)未见Xho I酶切位点的丢失(wt-Xho I)。除了Xho I酶切位点的丢失(nt：169423～169428;GAGCTC→GA□TCTC)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—Xho I,而在鼻咽癌组织中主要为Xho I-loss。因此,我们认为EB病毒LMP1基因N-末端区Xho I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。

Loss of an XhoI-site within N-terminal region of Epstein-Barr virus LMP1 gene in nasopharyngeal carcinoma

BACKGROUND & OBJECTIVE: It is well known that Epstein-Barr virus(EBV) LMP1 gene is involved in nasopharyngeal carcinogenesis. This research was designed to investigate the loss of an Xho I-site within the N-terminus of Epstein-Barr virus(EBV) latent membrane protein 1(LMP1) gene isolated from nasopharyngeal carcinoma (NPC) in Guangdong for further understanding the sequence variation of LMP1 gene involved in carcinogenesis. METHODS: Sixty-three fresh nasopharyngeal biopsies taken from the patients with nasopharyngeal carcinoma were collected in Cancer Center of Sun Yat-sen University. The peripheral blood mononuclear cells (PBMCs) obtained from 10 healthy EBV carriers were as control. The QIAamp DNA Mini Kits were used for extracting the DNA of biopsies and PBMCs. The N-terminus of EBV LMP1 gene was amplified using nested polymerase chain reaction (PCR) and then followed by Xho I enzyme digestion. Bidirectional solid-phase sequencing of the PCR products was performed using four-colored fluorescence terminator sequencing method. RESULTS: No loss of an Xho I-site within N-terminus of EBV LMP1 gene (wt-Xho I) was detected in PBMCs of all 10 carriers. The loss of an Xho I-site (Xho I-loss) was demonstrated in 50 cases (50/63, 79.36%) and the partial loss was demonstrated in 4 cases (4/63, 6.35%). The loss of an Xho I-site was not found in 9 cases (9/63, 14.29%). Besides loss of an Xho I-site (nt:169423-169428; GAGCTC --> GATCTC), 4 additional missense point mutations were found. CONCLUSION: According to the results obtained from this investigation, the PBMCs of 10 EBV carriers residing in Guangdong merely contain EBV variant with wt-Xho I. On the contrary, the EBV variant with XhoI-loss becomes the predominant variant detected in NPC tissues. So, the genomic variation within N-terminus (loss of an Xho I-site and other missense point mutations) of EBV LMP1 gene might be developed in the process of nasopharyngeal carcinogenesis.