肺癌患者全线粒体DNA体细胞性突变检测

Somatic mutation detection in complete mitochondrial DNA of lung cancer patients

目的 检测肺癌患者线粒体DNA体细胞性突变并探讨它在肿瘤发生中的作用。方法 利用时相温度梯度凝胶电泳对 16例肺癌患者的肿瘤及相应癌旁组织全线粒体DNA进行突变筛查 ,然后直接测序明确突变并进行分析。结果 在 10例 ( 62 .5 % )患者中发现 2 9个体细胞性突变 ,其中 2个位于rRNA区( 6.90 % ) ,10个位于mRNA区 ( 3 4.48% ) ,17个位于高变D环区 ( 5 8.62 % )。检测到 2例肿瘤和 3例癌旁组织具有常见的 4977bp缺失突变。除np3 0 3～ 3 0 9位点具有长度不稳定外 ,未发现其它微卫星区域不稳定。结论 肺癌患者的线粒体DNA存在高频率的体细胞性突变。

Objective To detect the somatic mutations in complete mitochondrial genome and to investigate the role of mtDNA mutation in the tumorigenesis of lung cancer. Methods DNA were extracted from sixteen lung cancer and corresponding normal tissues. The entire mitochondrial genome was amplified with 32 pairs of overlapping primers. mtDNA mutations were screened by temporal temperature gradient gel electrophoresis. mtDNA fragments showing different banding patterns between tumor and paracancerous tissues were sequenced to identify the exact mutations. The common 4 977 bp deletion was also analyzed in all sixteen tumor tissues as well as the matched paracancerous samples by PCR. Results Ten out of sixteen (62.5%) tumor tissues showed a total of 29 mutations. Half (5/10) of tumors with somatic mutation presented one mutation and the rests had multiple ones. Out of 29 mutations, 17 (58.62%) were in D-loop region, 2 (6.90%) in rRNA and 10 (34.48%) in mRNA. Among 10 mRNA mutations, 7 were silence and 3 were missense mutations. Five out of twenty-nine alterations were heteroplasmic to heteroplasmic change, one was homoplasmic to heteroplasmic and the remains (23/29, 79.3%) were homoplasmic to homoplasmic change. There were five common deletions found, two in tumor tissues and three in paracancerous tissues. There was no mitochondrial microsatellite instability, except for the short deletion or insertion in np303--309. Conclusion The high incidence of mtDNA mutations found in patients with lung cancer suggests that mtDNA alterations might play an important role in tumorigenesis of lung cancer. Further studies should be needed to determine the pathological effects of somatic mtDNA mutations in lung cancer.