Mutations in the mitochondrial genome confer resistance of cancer cells to anticancer drugs

已经确定，大多数癌细胞在线粒体基因组（mtDNA）中存在同质体细胞突变。我们发现mtDNA的体细胞突变与抗癌药物耐受有关。我们使用跨线粒体杂交细胞（cybrids）通过排除核背景的任何影响来揭示mtDNA突变在胰腺癌中的作用。通过用含有mtDNA突变的去核胰腺癌细胞（CFPAC-1,CAPAN-2）衍生的mtDNA重新填充缺乏mtDNA的HeLa构建Cybrids。我们构建了几个从癌细胞中获得突变的杂合体，以及那些从健康个体中获得野生mtDNA的杂合体。我们比较了突变体和野生杂交体对staurosporine（STS）、5FU、CDDP的体外抗性。实验表明，除了高温外，突变细胞系对药物的抗性比野生细胞系更强。接下来，对亚g1群体进行检测，以估计细胞凋亡过程中的核DNA断裂。STS处理在野生细胞系中比在突变细胞系中更显著地增加了亚g1群体。值得注意的是，细胞色素c氧化酶抑制剂阻止了STS野生杂交体亚g1群体的增加，这表明呼吸链活性参与了STS诱导的细胞凋亡。此外，我们还研究了cyt。线粒体免疫染色释放C。野生杂交细胞大部分失活，抗cyt染色。C抗体，表示cyt。C释放到细胞质中。相反，突变体细胞维持线粒体膜电位和细胞共定位。c.这些结果表明胰腺癌的mtDNA突变抑制了cyt。C依赖性细胞凋亡。我们的研究结果表明，mtDNA突变可以赋予癌细胞化疗耐药性。

It has been established that the majority of cancer cells harbor homoplasmic somatic mutations in the mitochondrial genome(mtDNA).We show that somatic mutations in mtDNA are involved in anticancer drug-tolerance.We used trans-mitochondrial hybrid cells (cybrids) to reveal the role of mutations in mtDNA in the pancreatic cancer by excluding any effects of the nuclear background.Cybrids were constructed by repopulating HeLa devoid of mtDNA with mtDNA derived from enucleated the pancreatic cancer cells(CFPAC-1,CAPAN-2)harboring mtDNA mutations. We constructed several cybrids with mutations derived from the cancer cells as well as those with wild mtDNA derived healthy individuals. We compared the mutant and wild cybrids in resistance against staurosporine(STS), 5FU, CDDP in vitro.The experiment revealed mutant cybrids were more resistant against the drugs than wild cybrids except hyperthermia. Next, Sub-G1 populations were examined to estimate nuclear DNA fragmentation during apoptosis. STS treatment increased Sub-G1 populations more greatly in wild cybrids than in mutant cybrids. Notably, an inhibitor of cytchrome c oxidase prevented the increase in Sub-G1 population of wild cybrids with STS, suggesting respiratory chain activity is involved in STS-induced apoptosis. Furthermore, we investigated cyt. c release from mitochondriaby immunostaining. Most cells of wild cybrid lost m and were stained with anti-cyt. c antibody,indicating cyt. c release to the cytosol. In contrast, mutant cybrids maintained mitochondrial membrane potential and co-localization of cyt. c. These results indicate that mtDNA mutations of the pancreatic cancer inhibit cyt. c -dependent apoptosis.Our results demonstrate that mtDNA mutationscan confer chemoresistance on cancer cells.