Prenatal Exome Sequencing Analysis in Fetuses With Structural Anomalies: A Multicenter Prospective Cohort Study With Practical Implications

ABSTRACT Objective To evaluate the diagnostic value of prenatal exome sequencing (ES) integrated with copy number variant (CNV) and single nucleotide variant (SNV) analysis (ES‐CNV/SNV) in fetuses with structural anomalies following negative chromosomal microarray analysis (CMA) and karyotyping, and to delineate the practical challenges encountered during its clinical implementation in prenatal settings. Methods In this multicenter prospective cohort study (2018–2021), 275 fetuses with structural anomalies were categorized into three groups: sporadic single‐system ( n = 128), sporadic multisystem ( n = 88), and recurrent anomalies ( n = 59). Trio‐based ES‐CNV/SNV analysis was used to identify SNVs, CNVs, or compound heterozygous CNV/SNV combinations— the latter being defined as two distinct pathogenic variants (a CNV and an SNV) coexisting within the same gene, resulting in biallelic gene dysfunction, which is pivotal for autosomal recessive disorders. Incremental diagnostic yields across groups, the incidence of CNV/SNV combinations, and expanded prenatal phenotypic‐genotypic correlations were analyzed. Results ES‐CNV/SNV analysis improved the diagnostic yield to 29.45% (81/275) compared with conventional chromosome analysis combined with CMA in fetuses with malformations. The highest diagnostic yield was observed in the recurrent anomaly group (40.68%, 24/59), followed by the multisystem (28.41%, 25/88) and single‐system anomaly groups (25.00%, 32/128). Notably, 1.45% of cases harbored compound CNV/SNV combinations, underscoring the diagnostic potential of ES‐CNV/SNV in autosomal recessive disorders. In addition, this study expanded prenatal phenotypic‐genotypic correlations for 36 variants and five genes. Conclusion ES‐CNV/SNV analysis enhances prenatal diagnostic precision by detecting compound CNV/SNV combinations that are undetectable by standard testing. Recurrent anomalies demonstrated a greater diagnostic benefit than sporadic single‐system or multisystem anomalies, underscoring the utility of this method in high‐risk cases. This study broadens the prenatal phenotypic spectrum and deepens our understanding of developmental mechanisms.