Lessons learnt from prenatal exome sequencing

Abstract

Background: Prenatal exome sequencing (ES) for monogenic disorders in fetuses with structural anomalies increases diagnostic yield. In England there is a national trio ES service delivered from two laboratories. To minimise incidental findings and reduce the number of variants investigated, analysis uses a panel of 1205 genes where pathogenic variants may cause abnormalities presenting prenatally. Here we review our laboratory's early experience developing and delivering ES to identify challenges in interpretation and reporting and inform service development.

Methods: A retrospective laboratory records review from 01.04.2020 to 31.05.2021.

Results: Twenty-four of 116 completed cases were identified as challenging including 13 resulting in difficulties in analysis and reporting, nine where trio inheritance filtering would have missed the diagnosis, and two with no prenatal diagnosis; one due to inadequate pipeline sensitivity, the other because the gene was not on the panel. Two cases with copy number variants identified were not detectable by microarray.

Conclusions: Variant interpretation requires close communication between referring clinicians, with occasional additional examination of the fetus or parents and communication of evolving phenotypes. Inheritance filtering misses ∼5% of diagnoses. Panel analysis reduces but does not exclude incidental findings. Regular review of published literature is required to identify new reports that may aid classification.

FIGURE 1

Variant prioritisation flowchart. AR, autosomal recessive; CNVs, copy number variants

FIGURE 2

KAT6A exon 13–17 deletion detected in C14. (A) Results from our CNV analysis tool indicating a deletion of exons 13–16 of the KAT6A gene. The blue dot represents normal copy number and red dot a deletion. The grey area shows the variance in read depth for the other samples on the run. (B) The sequencing reads at the identified breakpoints. The blue box shows the matching reference sequence from intron 13 that can be seen in the reads mapping to exon 17 with an insertion of GA in between. The red box indicates the matching reference sequence from exon 17 seen in intron 12, again with an insertion of GA. The CNV analysis results did not indicate a deletion of exon 17 however the results were out of the normal range. This is consistent with the breakpoint being within the exon and with it not being called due to the size of the exon. CNV, copy number variant; GA, gestational age

FIGURE 3

COL1A1 exon 40–43 deletion detected in C15. (A) Results from our CNV analysis tool indicating a deletion of exons 40–43 of the COL1A1 gene. The blue dot represents normal copy number and red dot a deletion. The grey area shows the variance in read depth for the other samples on the run. (B) The sequencing reads at the identified breakpoints. The blue box shows the matching reference sequence from intron 39 that can be seen in the reads mapping to exon 43 with insertion of GGGA from the exon 43 sequence in between. The red box indicates the matching reference sequence from exon 43 seen in intron 39, again with an insertion of GGGA. CNV, copy number variant