Stacey Andrew W, Nakamichi Kenji, Huey Jennifer, Stevens Jeffrey, Waligorski Natalie, Crotty Erin E, Van Gelder Russell N, Mustafi Debarshi
Department of Ophthalmology and Roger and Angie Karalis Johnson Retina Center, University of Washington, Seattle, Washington, USA.
Division of Ophthalmology, Seattle Children's Hospital, Seattle, Washington, USA.
JCI Insight. 2024 Dec 26;10(4):e188216. doi: 10.1172/jci.insight.188216.
BACKGROUNDCurrent clinical sequencing methods cannot effectively detect DNA methylation and allele-specific variation to provide parent-of-origin information from the proband alone. Parent-of-origin effects can lead to differential disease, and the inability to assign parent of origin in de novo cases limits prognostication in the majority of affected individuals with retinoblastoma, a hereditary cancer with suspected parent-of-origin effects.METHODSTo directly assign parent of origin in patients with retinoblastoma, we extracted genomic DNA from blood samples for sequencing using a programmable, targeted, single-molecule, long-read DNA genomic and epigenomic approach. This allowed germline variant calling and simultaneous haplotype-resolved CpG methylation in participants with familial (n = 7) and de novo (n = 9) retinoblastoma.RESULTSTargeted long-read sequencing allowed phasing genomic variation with a differentially methylated region in intron 2 of the retinoblastoma gene to confirm parent of origin in known familial samples. This approach allowed us to directly assign parent of origin in simple and complex de novo cases from the proband alone. The ability to assign parent of origin in all retinoblastoma cases showed that harboring disease-causing variants on the paternally inherited allele, whether arising familially or de novo, was associated with more advanced cancer staging at presentation and significantly greater risk of chemotherapy failure (P = 0.002).CONCLUSIONThis study demonstrates the diagnostic potential of multiomic long-read profiling to unveil the parent-of-origin effect in hereditary cancer. The approach in this work will be instrumental in assigning parent of origin to other genetic diseases using local and distant imprinting signals in the genome.FUNDINGNational Eye Institute, NIH; Gerber Foundation; Research to Prevent Blindness; Angie Karalis Johnson Fund; Dawn's Light Foundation; and Mark J. Daily, MD Research Fund.
背景
当前的临床测序方法无法有效地检测DNA甲基化和等位基因特异性变异，从而无法仅从先证者那里获取亲源信息。亲源效应可导致疾病差异，而在新发病例中无法确定亲源限制了大多数视网膜母细胞瘤（一种怀疑存在亲源效应的遗传性癌症）患者的预后。
方法
为了直接确定视网膜母细胞瘤患者的亲源，我们从血液样本中提取基因组DNA，使用一种可编程、靶向、单分子、长读长的DNA基因组和表观基因组方法进行测序。这使得我们能够在家族性视网膜母细胞瘤患者（n = 7）和新发视网膜母细胞瘤患者（n = 9）中进行种系变异检测并同时进行单倍型解析的CpG甲基化分析。
结果
靶向长读长测序能够将基因组变异与视网膜母细胞瘤基因内含子2中的一个差异甲基化区域进行定相，以确认已知家族性样本中的亲源。这种方法使我们能够仅从先证者直接确定简单和复杂新发病例的亲源。在所有视网膜母细胞瘤病例中确定亲源的能力表明，无论家族性还是新发，父系遗传等位基因上携带致病变异都与发病时更晚期的癌症分期以及化疗失败的显著更高风险相关（P = 0.002）。
结论
本研究证明了多组学长读长分析在揭示遗传性癌症亲源效应方面的诊断潜力。这项工作中的方法将有助于利用基因组中的局部和远距离印记信号为其他遗传疾病确定亲源。
资助
美国国立卫生研究院国家眼科研究所；格伯基金会；预防失明研究；安吉·卡拉利斯·约翰逊基金；道恩之光基金会；以及马克·J·戴利医学研究基金。
