Lethal variants in humans: lessons learned from a large molecular autopsy cohort

Abstract

Background: Molecular autopsy refers to DNA-based identification of the cause of death. Despite recent attempts to broaden its scope, the term remains typically reserved to sudden unexplained death in young adults. In this study, we aim to showcase the utility of molecular autopsy in defining lethal variants in humans.

Methods: We describe our experience with a cohort of 481 cases in whom the cause of premature death was investigated using DNA from the index or relatives (molecular autopsy by proxy). Molecular autopsy tool was typically exome sequencing although some were investigated using targeted approaches in the earlier stages of the study; these include positional mapping, targeted gene sequencing, chromosomal microarray, and gene panels.

Results: The study includes 449 cases from consanguineous families and 141 lacked family history (simplex). The age range was embryos to 18 years. A likely causal variant (pathogenic/likely pathogenic) was identified in 63.8% (307/481), a much higher yield compared to the general diagnostic yield (43%) from the same population. The predominance of recessive lethal alleles allowed us to implement molecular autopsy by proxy in 55 couples, and the yield was similarly high (63.6%). We also note the occurrence of biallelic lethal forms of typically non-lethal dominant disorders, sometimes representing a novel bona fide biallelic recessive disease trait. Forty-six disease genes with no OMIM phenotype were identified in the course of this study. The presented data support the candidacy of two other previously reported novel disease genes (FAAH2 and MSN). The focus on lethal phenotypes revealed many examples of interesting phenotypic expansion as well as remarkable variability in clinical presentation. Furthermore, important insights into population genetics and variant interpretation are highlighted based on the results.

Conclusions: Molecular autopsy, broadly defined, proved to be a helpful clinical approach that provides unique insights into lethal variants and the clinical annotation of the human genome.

Keywords: BMPR1A; EHBP1L1; Embryonic lethal; FAAH2; MSN; founder; loss of function; multi-locus.

Fig. 1

Pie chart summary of the study. A Age at death of the study cases. B Molecular tools used for the analysis in the study. C Molecular outcome of the study. D Inheritance pattern of the variants (pathogenic and likely pathogenic) identified in the study

Fig. 2

Identification of EHBP1L1-related lethal phenotype. A, B Pedigree of 14DG1037 and sequence chromatogram of EHBP1L1:NM_001099409.3:c.3333_3346delinsAGAGTCAGTAGCA variant. C, D Pedigree of 18DG0247 and RT-PCR of the variant EHBP1L1:NM_001099409:c.4004-1G>A) confirming aberrant splicing with skipping of exon-14. E Sketch for EHBP1L1 transcript and protein, arrows denote mutated bases and residues

Fig. 3

Identification of FAAH2-related lethal phenotype. A Pedigree of 18DG0989 (died during the neonatal period). B Images of the patient with facial dysmorphism, significant hypotonia, and mechanical ventilator dependence. C Sanger sequence of FAAH2:NM_174912:c.1175G>A:p.(Trp392*) and sketch of FAAH2 protein domains

Fig. 4

Identification of MSN-related lethal phenotype. A Pedigree of 16DG0125 (two siblings died during childhood). B, C CT scan showing widespread ground glass change with no significant interstitial thickening. D, E Sequence chromatogram of MSN:NM_002444.3:c.1056delG, MTTP:NM_000253: c.2620A>G variants, and sketches of MSN and MTTP protein domains; arrows denote mutated residues

Fig. 5

RNA-seq identifies EPG5-related lethal phenotype. A, Pedigree of 16DG1465 with deletion of EPG5 exon-1 (two siblings died during infancy). B Image of the patient with facial dysmorphism. C MRI showing absent corpus callosum. D A sashimi plot showing the base-level density of reads mapped to a genomic region surrounding the ~70K deletion site. The x-axis represents the genomic coordinate in hg38. The y-axis represents per-base read counts. On the bottom, the boxes are annotated exons, the horizontal lines are introns, and the left-facing arrowheads indicate the negative strand. Arcs connecting exons represent splice junction reads. The plot shows that no reads are mapped to exon 1 of EPG5 and that 22 split-reads are mapped to an aberrant splicing junction between exon 2 of PSTPIP2 and exon 2 of EPG5 that are ~84Kbp apart