[论文解读] Beyond the exome: what's next in diagnostic testing for Mendelian conditions

Monica H. Wojcik, Chloe M. Reuter+22 more|arXiv (Cornell University)|Jan 18, 2023

Genomics and Rare Diseases被引用 5

一句话总结

本文回顾了外显子组测序在诊断单基因遗传病方面的局限性，并提出了一套结构化的下一步诊断检测框架，包括全基因组测序、RNA测序以及长读长测序和光学基因组分析等新兴技术。文章强调基于表型和既往检测结果的临床决策，倡导在未确诊病例中转诊至联盟机构以提高外显子组测序之外的诊断率。

ABSTRACT

Despite advances in clinical genetic testing, including the introduction of exome sequencing (ES), more than 50% of individuals with a suspected Mendelian condition lack a precise molecular diagnosis. Clinical evaluation is increasingly undertaken by specialists outside of clinical genetics, often occurring in a tiered fashion and typically ending after ES. The current diagnostic rate reflects multiple factors, including technical limitations, incomplete understanding of variant pathogenicity, missing genotype-phenotype associations, complex gene-environment interactions, and reporting differences between clinical labs. Maintaining a clear understanding of the rapidly evolving landscape of diagnostic tests beyond ES, and their limitations, presents a challenge for non-genetics professionals. Newer tests, such as short-read genome or RNA sequencing, can be challenging to order and emerging technologies, such as optical genome mapping and long-read DNA or RNA sequencing, are not available clinically. Furthermore, there is no clear guidance on the next best steps after inconclusive evaluation. Here, we review why a clinical genetic evaluation may be negative, discuss questions to be asked in this setting, and provide a framework for further investigation, including the advantages and disadvantages of new approaches that are nascent in the clinical sphere. We present a guide for the next best steps after inconclusive molecular testing based upon phenotype and prior evaluation, including when to consider referral to a consortium such as GREGoR, which is focused on elucidating the underlying cause of rare unsolved genetic disorders.

研究动机与目标

• 解决尽管广泛使用外显子组测序，单基因遗传病仍存在高比例未确诊病例的问题。
• 识别阻碍外显子组测序后诊断解决的关键技术、生物学和解释性局限。
• 提供基于表型和既往检测结果选择下一代遗传检测的临床决策框架。
• 评估新兴基因组技术（如短读长和长读长全基因组测序、RNA测序以及光学基因组分析）在临床诊断中的实用性和局限性。
• 推动在未确诊罕见病病例中及时转诊至专门的联盟机构（如GREGoR），以提高诊断效率。

提出的方法

• 系统性回顾外显子组测序后单基因遗传病诊断面临的当前挑战。
• 评估新兴基因组技术，包括短读长全基因组测序、RNA测序、长读长测序以及光学基因组分析。
• 开发基于表型的算法，以指导外显子组测序后的下一步检测选择。
• 分析变异解释的挑战，包括非编码区变异、复杂调控元件以及不完全的基因型-表型关联。
• 整合临床与研究联盟模式（如GREGoR）作为解决未确诊病例的关键资源。
• 综合临床决策路径，整合新技术与专家咨询。

实验结果

研究问题

• RQ1 外显子组测序后单基因遗传病诊断失败的主要原因是什么？
• RQ2 外显子组测序的技术和生物学局限如何导致未确诊病例？
• RQ3 下一代基因组检测方法（如全基因组测序和RNA测序）在临床和应用上的优缺点是什么？
• RQ4 临床医生在何种情况下应考虑将未确诊的单基因遗传病转诊至专门的研究联盟（如GREGoR）？
• RQ5 表型信息如何指导选择外显子组测序之外的最佳下一步基因组检测？

主要发现

• 超过50%的疑似单基因遗传病患者在外显子组测序后仍未能确诊，凸显了诊断中的关键空白。
• 短读长全基因组测序和RNA测序通过检测外显子组测序遗漏的非编码区变异、剪接位点破坏及基因表达异常，提高了诊断率。
• 长读长测序和光学基因组分析在检测结构变异和复杂基因组重排方面展现出潜力，但受限于临床可及性和标准化程度。
• 基于表型的检测选择显著提高了诊断效率并减少了不必要的检测。
• 向GREGoR等研究联盟转诊可提高未确诊病例的诊断解决率，通过支持高级数据分析和协作变异解读。
• 缺乏外显子组测序后检测的标准化临床指导，仍是广泛实施下一代基因组检测的主要障碍。