Methylation and mutations in RB1 and variants of synthetic folic acid metabolism

 描述(申请人提供)：50%的美国成年人食用合成叶酸补充剂。叶酸摄入量过高会导致CpG岛甲基化。最近的研究表明，高摄入量可能导致癌症的发生，特别是在那些不能有效代谢叶酸的人中，例如二氢叶酸还原酶(DHFR)基因19bp缺失多态的纯合子(Rs70991108)。纯合子(19%的美国成年人)将合成叶酸转化为生物叶酸的效率较低。DHFR19bpdel缺失(DHFR19bpdel)纯合子的女性生下视网膜母细胞瘤的风险更高，视网膜母细胞瘤是一种神经外胚层衍生的儿童早期癌症，已成为了解癌症发生的模型。在DHFR19bpdel纯合子妇女中，这种风险似乎增加了，她们在怀孕的前三个月服用了含有叶酸的产前补充剂。在视网膜母细胞瘤中，富含CpG的基因RB1中37%的致病突变是由于RB1中甲基化的胞嘧啶脱氨基造成的，而另外12%的突变是由RB1启动子的甲基化引起的。我们推测，这些与甲基化相关的RB1失活变化可能是由叶酸代谢功能较差的遗传变异和视网膜发育早期过度暴露引起的。小鼠模型表明，在神经发育的早期，对叶酸暴露特别敏感。在人类中，妊娠早期饮食不足的时期与亚稳等位基因的超甲基化有关。来自我们正在进行的墨西哥中部新诊断的视网膜母细胞瘤(平均年龄23个月)病例系列研究EpiRbMx的突变分析和甲基化阵列的试点数据表明，出生前具有效率较低的DHFR 19bpdel和高叶酸的妇女生下的孩子具有与甲基化相关的RB1失活突变或直接导致肿瘤形成的差异超甲基化的RB1启动子的更高风险。母体DHFR19bpdel纯合子与产前高叶酸摄入量可能导致发育易感细胞中RB1超甲基化，从而促进癌症的发生。基于我们试点数据的结果，我们建议在我们正在进行的EpiRbMx研究(成立于2003年)中使用现有的样本和来自337例患者的数据，包括母子血浆营养物质、饮食摄入量、基因和肿瘤的治疗前措施，以检验我们的假设，即母亲DHFR 19bpdel与产前叶酸暴露(使用血浆和摄入量措施)之间的关联：1)与甲基化相关的RB1突变，特别是在体细胞(非生殖系)RB1突变，以及2)RB1启动子的甲基化。我们将进一步研究DHFR和产前叶酸暴露对亚稳等位基因差异甲基化的影响，以证明它们的 对发育中的表观基因组的影响。鉴于RB1突变在预后较差的成人肿瘤中的高频率，如小细胞肺癌(54%)或胶质母细胞瘤(11%)，以及叶酸代谢效率较低的成年人中的高比例(19%)，我们的研究潜在的翻译影响很高，因为我们的结果可以为公共卫生政策和对合成叶酸暴露的评估提供参考。

 DESCRIPTION (provided by applicant): Synthetic folic acid supplements are consumed by 50% of US adults. High folic acid intake can result in CpG island hypermethylation. Recent studies suggest high intake may contribute to carcinogenesis particularly in those persons unable to metabolize folic acid efficiently such as persons homozygous for the 19bp deletion polymorphism in the dihydrofolate reductase(DHFR) gene (rs70991108). Homozygotes (19% of US adults) have less efficient conversion of synthetic folic acid into biological folate. Women homozygous for the DHFR19bp deletion (DHFR19bpdel) are at increased risk for having a child with retinoblastoma, a neuro-ectodermally derived early childhood cancer that has served as a model for understanding carcinogenesis. This risk appears elevated among women homozygous for DHFR19bpdel who consumed folic acid containing prenatal supplements in the first trimester. In retinoblastoma, 37% of disease-causing mutations in the CpG-rich gene, RB1, are due to deamination of methylated cytosines in RB1, while an additional 12% are caused by methylation of the RB1 promoter. We hypothesize that these methylation related RB1-inactivating changes may be induced by less functional genetic variations in folate metabolism and excess exposure in early retinal development. Murine models suggest particular sensitivity to folic acid exposure in early neural development. In humans, periods of dietary inadequacy in early gestation are associated with hyper-methylation at meta- stable epialleles. Pilot data from mutation assays and methylation arrays from our ongoing case-series study of newly diagnosed retinoblastoma (mean age 23 months) in Central Mexico, EpiRbMx, suggest that women who have the less efficient DHFR 19bpdel and high folic acid prenatally have higher risk of a child with a methylation related RB1-inactivating mutation or differentially hyper-methylated RB1 promoter directly resulting in tumor formation. Maternal DHFR19bpdel homozygosity combined with high prenatal folic acid intake may lead to RB1 hyper-methylation in developmentally susceptible cells, thereby contributing to carcinogenesis. Based on the results of our pilot data, we propose to use existing samples and data from 337 cases in our ongoing EpiRbMx study (founded in 2003) including pre-treatment measures of mother and child plasma nutrients, diet intake, genotype, and tumors, to test our hypothesis of an association between maternal DHFR 19bpdel and prenatal folic acid exposure (using plasma and intake measures) and 1) methylation related RB1 mutations, specifically in somatic (non-germline) RB1 mutations, and 2) methylation of the RB1 promoter. We will further examine the impact of DHFR and prenatal folic acid exposure on differential methylation in metastable epialleles to document their effect on the developing epigenome. Given the high frequency of RB1 mutations among poor prognosis adult tumors such as small cell lung cancer(54%) or glioblastoma(11%) and the high proportion (19%) of adults with less efficient folic acid metabolism, the potential translational impact of our study is high, as our results can inform public health policy and evaluation of synthetic folic acid exposure.