Purpose: To investigate the number of rare missense variants observed in human genome sequences by ACMG/AMP PP3/BP4 evidence strength, following the calibrated PP3/BP4 computational recommendations. Methods: Missense variants from the genome sequences of 300 probands from the Rare Genomes Project with suspected rare disease were analyzed using computational prediction tools able to reach PP3_Strong and BP4_Moderate evidence strengths (BayesDel, MutPred2, REVEL, and VEST4). The numbers of variants at each evidence strength were analyzed across disease-associated genes and genome-wide. Results: From a median of 75.5 rare (≤1% allele frequency) missense variants in disease-associated genes per proband, a median of one reached PP3_Strong, 3-5 PP3_Moderate, and 3-5 PP3_Supporting. Most were allocated BP4 evidence (median 41-49 per proband) or were indeterminate (median 17.5-19 per proband). Extending the analysis to all protein-coding genes genome-wide, the number of PP3_Strong variants increased approximately 2.6-fold compared to disease-associated genes, with a median per proband of 1-3 PP3_Strong, 8-16 PP3_Moderate, and 10-17 PP3_Supporting. Conclusion: A small number of variants per proband reached PP3_Strong and PP3_Moderate in 3,424 disease-associated genes, and though not the intended use of the recommendations, also genome-wide. Use of PP3/BP4 evidence as recommended from calibrated computational prediction tools in the clinical diagnostic laboratory is unlikely to inappropriately contribute to the classification of an excessive number of variants as Pathogenic or Likely Pathogenic by ACMG/AMP rules.

中文翻译：

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评估校准 PP3/BP4 计算建议的证据产量

目的：按照校准的 PP3/BP4 计算建议，通过 ACMG/AMP PP3/BP4 证据强度调查在人类基因组序列中观察到的罕见错义变异的数量。方法：使用能够达到 PP3_Strong 和 BP4_Moderate 证据强度（BayesDel、MutPred2、REVEL 和 VEST4）的计算预测工具，对来自稀有基因组计划的 300 名疑似罕见疾病先证者的基因组序列的错义变异进行了分析。对疾病相关基因和全基因组范围内每个证据强度的变异数量进行了分析。结果：每个先证者疾病相关基因中平均有 75.5 个罕见（≤1% 等位基因频率）错义变异，其中 1 个达到 PP3_Strong、3-5 个 PP3_Moderate 和 3-5 个 PP3_Supporting。大多数分配了 BP4 证据（每个先证者中位数 41-49）或不确定（每个先证者中位数 17.5-19）。将分析扩展到全基因组范围内的所有蛋白质编码基因，与疾病相关基因相比，PP3_Strong 变异的数量增加了约 2.6 倍，每个先证者的中位数为 1-3 个 PP3_Strong、8-16 个 PP3_Moderate 和 10-17 个PP3_支持。结论：每个先证者的 3,424 个疾病相关基因中的少数变异达到 PP3_Strong 和 PP3_Moderate，虽然不是建议的预期用途，但也在全基因组范围内。按照临床诊断实验室校准计算预测工具的建议使用 PP3/BP4 证据不太可能不恰当地导致 ACMG/AMP 规则将过多的变异分类为致病性或可能致病性。