FANCONI ANEMIA:GENOTYPE-PHENOTYPE CORRELATIONS

范可尼贫血：基因型-表型相关性

基本信息

批准号：
9571134
负责人：
settara chandrasekharappa
金额：
$ 78.54万
依托单位：
NATIONAL HUMAN GENOME RESEARCH INSTITUTE
依托单位国家：
美国
项目类别：
财政年份：
资助国家：
美国
起止时间：
至
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/9571134
关键词：
Adopted Affect Age Alleles BRCA2 gene Back Blood Cells Cell Fraction Clinical Management Clinical Treatment Collaborations Complement Congenital Abnormality Copy Number Polymorphism DNA DNA Repair Pathway Diagnosis Disease Enrollment Evaluation Exons FANCB gene FANCG gene Family Family-Based Registry Fanconi Anemia Complementation Group A Protein Fanconi&apos s Anemia Fibroblasts Frequencies Gene Mutation General Population Genes Genome Genomic DNA Genomics Genotype Goals Head and Neck Squamous Cell Carcinoma Hematopoietic Heterozygote Hybridization Array Inherited International Life Link Malignant neoplasm of pancreas Manuscripts Massive Parallel Sequencing Methodology Molecular Diagnosis Mosaicism Mutation Myeloid Leukemia Pancytopenia Patients Phenotype Predisposition Preparation Promoter Regions Proteins Protocols documentation Publications Publishing RNA Splicing RNA analysis Rare Diseases Registries Reporting Risk Role SNP array Siblings Techniques Technology Testing Time Tissues United States National Institutes of Health Universities Variant Zebrafish blood fractionation cohort comparative genomic hybridization disease-causing mutation genetic variant insertion/deletion mutation malignant breast neoplasm null mutation repaired

项目摘要

Once diagnosed with Fanconi anemia (FA), identification of the causative gene and the mutations is an arduous task. FA genes are large, with multiple exons, and harbor a wide spectrum of compound heterozygous mutations spread throughout the gene including large genomic deletions. Thus, molecular diagnosis of a large number of families enrolled in the International Fanconi Anemia Registry (IFAR) remained unknown. Within the last two years, there were reports of five new FA genes being identified, thus bringing the number of known FA genes to 21. Though FA patients can carry mutations in any of the 21 known genes, about two-thirds are affected by mutations in FANCA. Our current efforts are focused on employing massively parallel sequencing technologies to sequence large (2-3 Mb) regions of the genome, targeting all FA and FA-related DNA-repair pathway genes. We also adopt Comparative Genome Hybridization arrays (aCGH), and SNP arrays to explore large-size copy number variants in a similar set of genes. So far, through our collaboration with the Rockefeller University, we have identified bi-allelic mutations in 500 IFAR families, and these include 313 FANCA, 16 FANCB, 54 FANCC, 15 FANCD1, 21 FANCD2, seven FANCE, 15 FANCF, 23 FANCG, 11 FANCI, 15 FANCJ, five FANCL, four FANCN, three FANCP and one FANCT. A few highlights from this effort include: 1) identification of 16 patients with an X-linked FANCB mutation, a very rare FA group, and in four patients, the mutation was de novo; 2) Identification of rare disease-associated deletions in 148 families including 131 FANCA, 8 FANCC, 5 FANCD2, one FANCJ, one FANCI, and 2 FANCB using aCGH; 3) determination of the precise breakpoints for 90 deletions, and discovered the mechanisms leading to deletions; 4) found that fifty-five FANCA deletions overlapped exon 1 and extended beyond the 5 gene terminus, eliminating a putative promoter region and, likely expression. This was confirmed by RNA analysis. A manuscript describing characterization of both mutations in 160 FANCA patients in under revision. For the past two years, as a part of our detailed molecular diagnosis, we have been exploring causes and consequences of mosaicism in FA families. It is estimated that 20% of FA patients may display somatic mosaicism, a scenario where a fraction of cells from hematopoietic lineages may have lost, or repaired, one of the inherited mutations. This phenomenon results in a functional allele in the fraction of blood cells with reverse mosaicism (RM), and may often provide protection from hematopoietic diseases. Somatic mosaicism in a patient is evident when his/her blood cells were subjected to DNA breakage test at the time of diagnosis. We completed our study of three siblings in a family with mutations in FANCG displaying RM, each displaying a different mechanism that resulted in RM, and a manuscript describing these findings is under preparation. We also completed another study where a FANCB patient displayed RM. The patient harbored a 10kb intragenic duplication in FANCB. This duplication was unstable and reverted back to wild type in patient cells from peripheral blood and also, to some extent, from fibroblasts. A manuscript describing the mosaicism displayed by the FANCB patient is under revision. In an effort to evaluate the frequency of FA gene variants in patients diagnosed with HNSCC under the age of 50, we sequenced genomic DNA of 492 patients for variations in all FA genes. The findings from this study were published this year. In addition, we have generated zebrafish carrying mutations in each of the 19 FA genes. The initial characterization of FA null mutations in zebrafish is now complete and a manuscript describing these efforts is being prepared for publication.

一旦被诊断出患有法科尼贫血（FA），病因基因和突变是一项艰巨的任务。 FA基因很大，具有多个外显子，并且具有广泛的复合杂合突变，包括大型基因组缺失。因此，对参加国际芬科尼贫血注册中心（IFAR）的大量家庭的分子诊断仍然未知。在过去的两年中，有报道称五个新的FA基因被鉴定出来，从而使已知的FA基因数量达到21个。尽管FA患者可以在21个已知基因中的任何一个中携带突变，但大约三分之二的人会受到FANCA突变的影响。我们目前的努力集中在采用大量平行的测序技术来对基因组的大（2-3 MB）区域进行序列，以瞄准所有FA和FA相关的DNA-REPAIR途径。我们还采用了比较基因组杂交阵列（ACGH）和SNP阵列来探索类似基因集中的大尺寸拷贝数变体。 So far, through our collaboration with the Rockefeller University, we have identified bi-allelic mutations in 500 IFAR families, and these include 313 FANCA, 16 FANCB, 54 FANCC, 15 FANCD1, 21 FANCD2, seven FANCE, 15 FANCF, 23 FANCG, 11 FANCI, 15 FANCJ, five FANCL, four FANCN, three FANCP and one FANCT. 这项工作中的一些亮点包括：1）鉴定16例X连锁Fancb突变的患者，一个非常罕见的FA组，在四名患者中，该突变是从头开始的； 2）在148个家庭中鉴定了罕见的疾病相关缺失，其中包括131个Fanca，8 Fancc，5 Fancd2，一个Fancj，一个Fancj，One Fanci和2 Fancb使用ACGH； 3）确定90个缺失的精确断点，并发现导致缺失的机制； 4）发现55 fanca的缺失重叠，并延伸到5个基因末端，从而消除了推定的启动子区域以及可能的表达。 RNA分析证实了这一点。在修订中，描述了160名FANCA患者的两种突变表征的手稿。在过去的两年中，作为我们详细的分子诊断的一部分，我们一直在探索FA家族中镶嵌的原因和后果。据估计，20％的FA患者可能表现出体细胞镶嵌性，这种情况可能会丢失或修复造血谱系中的一部分细胞之一。这种现象会导致血液细胞的功能性等位基因反向镶嵌（RM），并且通常可以保护免受造血疾病的保护。当患者的血液细胞在诊断时接受DNA断裂测试时，患者的体细胞镶嵌性很明显。我们完成了一个家庭中的三个兄弟姐妹的研究，其中粉丝中的突变显示了RM，每种兄弟姐妹都显示了导致RM的不同机制，并且描述这些发现的手稿正在准备中。我们还完成了另一项研究，其中粉丝患者显示了RM。该患者在Fancb中具有10KB的内部重复。这种重复是不稳定的，并从外周血，在某种程度上从成纤维细胞中恢复为野生型。描述粉丝患者展示的镶嵌的手稿正在修订。为了评估50岁以下HNSCC诊断的患者FA基因变异的频率，我们对所有FA基因的变异进行了492例患者的基因组DNA。这项研究的发现已于今年发表。此外，我们在19个FA基因中的每一个中都产生了斑马鱼携带突变。斑马鱼中FA Null突变的最初表征现已完成，并且正在为这些努力的手稿准备出版。