Sequence variation in mtDNA and extreme survival in glioblastoma

摘要 胶质母细胞瘤（GBM）是人类最致命的恶性肿瘤之一。中位生存期为12-15个月， 尽管采用了手术、放疗和同时使用替莫唑胺积极治疗。新的途径是 迫切需要开发有效的治疗方法。癌症中一个新兴的感兴趣领域是 线粒体，细胞器，通过氧化提供身体的大部分能量需求， 磷酸化（OXPHOS）。线粒体具有独立的环状16.5KB基因组（mtDNA）， 这是人类基因组中一个重要的、但经常被忽视的细胞质元件。线粒体DNA编码蛋白质 对OXPHOS至关重要，并且在人群中高度多态，还受到体细胞变异的影响。一 越来越多的证据表明，古代和近代的生殖系mtDNA变异和 线粒体DNA的体细胞改变可能在癌症风险和进展中起重要作用。在神经胶质瘤中， 有证据表明OXPHOS酶活性和组织mtDNA含量在耐药性中起作用， 肿瘤分级在这个R21中，我们建议进行第一次全面的生殖系和体细胞调查， GBM生存中的mtDNA变异。该项目是基于生殖细胞和肿瘤DNA的独特资源， 大量GBM患者，其统一的肿瘤和治疗数据以及生命状态可用。 生殖系mtDNA序列将使用高通量下一代口腔DNA测序来确定 样品我们将研究生殖系变异是否能区分3种罕见的“极端幸存者”（ES）， 15个月或更短的平均生存期（AS）（150例ES与450例AS年龄和 治疗）。线粒体单倍型群、常见个体变异以及罕见和单基因变异 在基因和功能途径内的总体考虑，都将检查与患者的相关性 结果。然后，我们将在探索性分析中调查肿瘤mtDNA中的体细胞变异是否有助于 患者存活率（120例GBM患者，具有配对的生殖系和新鲜冷冻的原发性肿瘤）。体细胞变体 将通过对天然肿瘤mtDNA进行测序来确定，采用高覆盖深度来捕获低覆盖深度。 个体内的频率变化。结构和单核苷酸变异将按类别、总数 负荷和预测的致病性与患者结局的相关性。该建议是响应 来自NCI的“挑衅性问题5”，例如：“ 线粒体异质性如何影响肿瘤发生或 进展？” (http http：//grants.nih.gov/grants/guide/rfa-files/RFA-CA-15-009.html）。成果将贡献新的 深入了解线粒体基因组在驱动GBM中的作用，并可能指向新的 针对线粒体功能的治疗策略。

ABSTRACT Glioblastoma (GBM) is among the most lethal human malignancies. Median survival is a dismal 12-15 months in spite of aggressive treatment with surgery, radiation and concurrent temozolomide. New avenues are urgently needed for development of effective therapeutics. An emerging area of interest in cancer is the role of the mitochondria, cellular organelles which provide most of the energy needs of the body via oxidative phosphorylation (OXPHOS). Mitochondria possess an independent circular 16.5KB genome (mtDNA) that is an essential, and often overlooked, cytoplasmic element of the human genome. mtDNA encodes proteins critical to OXPHOS and is highly polymorphic in human populations and also subject to somatic variation. A growing body of evidence indicates that ancient and recent germline mtDNA variants and the burden of somatic alterations in mtDNA may play important roles in cancer risk and progression. In glioma, recent evidence points to OXPHOS enzyme activity and tissue mtDNA content playing a role in drug resistance and tumor grade. In this R21, we propose to carry out the first comprehensive investigation of germline and somatic mtDNA variation in GBM survival. The project is based on a unique resource of germline and tumor DNA from large numbers of GBM patients for whom uniform tumor and treatment data, and vital status are available. Germline mtDNA sequence will be determined using high-throughput next-generation sequencing of oral DNA samples. We will investigate whether germline variants distinguish infrequent `extreme survivors' (ES) of 3 years or longer from average survivors (AS) of 15 months or less (150 ES versus 450 AS matched for age and treatment). Mitochondrial haplogroups, common individual variants, as well as rare and singleton variants considered in aggregate within genes and functional pathways, will all be examined for association with patient outcome. We will then investigate, in exploratory analysis, whether somatic variants in tumor mtDNA contribute to patient survival (120 GBM patients with paired germline and fresh frozen primary tumor). Somatic variants will be determined by sequencing native tumor mtDNA employing high depth of coverage to capture low frequency variants within individuals. Structural and single nucleotide variants will be considered by class, total burden and predicted pathogenicity for association with patient outcome. The proposal is responsive to “Provocative Question 5” from the NCI eg: “ How does mitochondrial heterogeneity influence tumorigenesis or progression?” (http://grants.nih.gov/grants/guide/rfa-files/RFA-CA-15-009.html). Results will contribute new insight on questions surrounding the role of the mitochondrial genome in driving GBM and may point to novel therapeutic strategies that target mitochondrial function.