BACKGROUND The American College of Medical Genetics and Genomics (ACMG)/Association for Molecular Pathology (AMP) clinical variant interpretation guidelines established criteria for different types of evidence. This includes the strong evidence codes PS3 and BS3 for "well-established" functional assays demonstrating a variant has abnormal or normal gene/protein function, respectively. However, they did not provide detailed guidance on how functional evidence should be evaluated, and differences in the application of the PS3/BS3 codes are a contributor to variant interpretation discordance between laboratories. This recommendation seeks to provide a more structured approach to the assessment of functional assays for variant interpretation and guidance on the use of various levels of strength based on assay validation. METHODS The Clinical Genome Resource (ClinGen) Sequence Variant Interpretation (SVI) Working Group used curated functional evidence from ClinGen Variant Curation Expert Panel-developed rule specifications and expert opinions to refine the PS3/BS3 criteria over multiple in-person and virtual meetings. We estimated the odds of pathogenicity for assays using various numbers of variant controls to determine the minimum controls required to reach moderate level evidence. Feedback from the ClinGen Steering Committee and outside experts were incorporated into the recommendations at multiple stages of development. RESULTS The SVI Working Group developed recommendations for evaluators regarding the assessment of the clinical validity of functional data and a four-step provisional framework to determine the appropriate strength of evidence that can be applied in clinical variant interpretation. These steps are as follows: (1) define the disease mechanism, (2) evaluate the applicability of general classes of assays used in the field, (3) evaluate the validity of specific instances of assays, and (4) apply evidence to individual variant interpretation. We found that a minimum of 11 total pathogenic and benign variant controls are required to reach moderate-level evidence in the absence of rigorous statistical analysis. CONCLUSIONS The recommendations and approach to functional evidence evaluation described here should help clarify the clinical variant interpretation process for functional assays. Further, we hope that these recommendations will help develop productive partnerships with basic scientists who have developed functional assays that are useful for interrogating the function of a variety of genes.

中文翻译：

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使用 ACMG/AMP 序列变异解释框架应用功能证据 PS3/BS3 标准的建议。

背景 美国医学遗传学和基因组学学会 (ACMG)/分子病理学协会 (AMP) 临床变异解释指南为不同类型的证据建立了标准。这包括强有力的证据代码 PS3 和 BS3，用于“完善的”功能分析，分别证明变体具有异常或正常的基因/蛋白质功能。然而，他们没有提供关于如何评估功能证据的详细指导，PS3/BS3 代码应用的差异是导致实验室之间变异解释不一致的一个原因。该建议旨在提供一种更加结构化的方法来评估功能检测，以进行变异解释，并指导基于检测验证的各种强度水平的使用。方法 临床基因组资源 (ClinGen) 序列变异解释 (SVI) 工作组使用来自 ClinGen 变异管理专家小组制定的规则规范和专家意见的精选功能证据，通过多次面对面和虚拟会议完善 PS3/BS3 标准。我们使用各种数量的变异对照来估计致病性的几率，以确定达到中等水平证据所需的最低对照。来自 ClinGen 指导委员会和外部专家的反馈在多个发展阶段被纳入建议。结果 SVI 工作组为评估人员制定了关于评估功能数据临床有效性的建议，并制定了一个四步临时框架以确定可应用于临床变异解释的适当证据强度。这些步骤如下：(1) 定义疾病机制，(2) 评估现场使用的一般化验类别的适用性，(3) 评估特定化验实例的有效性，以及 (4) 将证据应用于个体变异解释。我们发现，在没有严格统计分析的情况下，至少需要 11 个总致病性和良性变异对照才能达到中等水平的证据。结论 此处描述的功能证据评估的建议和方法应有助于阐明功能测定的临床变异解释过程。此外，我们希望这些建议将有助于与基础科学家建立富有成效的合作伙伴关系，这些基础科学家已经开发出可用于研究各种基因功能的功能测定。