Digenic inheritance of human primary microcephaly delineates centrosomal and non-centrosomal pathways.

Primary microcephaly (PM) is characterized by a small head since birth and is vastly heterogeneous both genetically and phenotypically. While most cases are monogenic, genetic interactions between Aspm and Wdr62 have recently been described in a mouse model of PM. Here, we used two complementary, holistic in vivo approaches: high throughput DNA sequencing of multiple PM genes in human patients with PM, and genome-edited zebrafish modeling for the digenic inheritance of PM. Exomes of patients with PM showed a significant burden of variants in 75 PM genes, that persisted after removing monogenic causes of PM (e.g., biallelic pathogenic variants in CEP152). This observation was replicated in an independent cohort of patients with PM, where a PM gene panel showed in addition that the burden was carried by six centrosomal genes. Allelic frequencies were consistent with digenic inheritance. In zebrafish, non-centrosomal gene casc5 -/- produced a severe PM phenotype, that was not modified by centrosomal genes aspm or wdr62 invalidation. A digenic, quadriallelic PM phenotype was produced by aspm and wdr62. Our observations provide strong evidence for digenic inheritance of human PM, involving centrosomal genes. Absence of genetic interaction between casc5 and aspm or wdr62 further delineates centrosomal and non-centrosomal pathways in PM

Abstracts from the 50th European Society of Human Genetics Conference: Posters

International audienc

Elucidation of the phenotypic spectrum and genetic landscape in primary and secondary microcephaly

Purpose: Microcephaly is a sign of many genetic conditions but has been rarely systematically evaluated. We therefore comprehensively studied the clinical and genetic landscape of an unselected cohort of patients with microcephaly. Methods: We performed clinical assessment, high-resolution chromosomal microarray analysis, exome sequencing, and functional studies in 62 patients (58% with primary microcephaly [PM], 27% with secondary microcephaly [SM], and 15% of unknown onset). Results: We found severity of developmental delay/intellectual disability correlating with severity of microcephaly in PM, but not SM. We detected causative variants in 48.4% of patients and found divergent inheritance and variant pattern for PM (mainly recessive and likely gene-disrupting [LGD]) versus SM (all dominant de novo and evenly LGD or missense). While centrosome-related pathways were solely identified in PM, transcriptional regulation was the most frequently affected pathway in both SM and PM. Unexpectedly, we found causative variants in different mitochondria-related genes accounting for ~5% of patients, which emphasizes their role even in syndromic PM. Additionally, we delineated novel candidate genes involved in centrosome-related pathway (SPAG5, TEDC1), Wnt signaling (VPS26A, ZNRF3), and RNA trafficking (DDX1). Conclusion: Our findings enable improved evaluation and genetic counseling of PM and SM patients and further elucidate microcephaly pathways

A Solve-RD ClinVar-based reanalysis of 1522 index cases from ERN-ITHACA reveals common pitfalls and misinterpretations in exome sequencing

Purpose Within the Solve-RD project (https://solve-rd.eu/), the European Reference Network for Intellectual disability, TeleHealth, Autism and Congenital Anomalies aimed to investigate whether a reanalysis of exomes from unsolved cases based on ClinVar annotations could establish additional diagnoses. We present the results of the “ClinVar low-hanging fruit” reanalysis, reasons for the failure of previous analyses, and lessons learned. Methods Data from the first 3576 exomes (1522 probands and 2054 relatives) collected from European Reference Network for Intellectual disability, TeleHealth, Autism and Congenital Anomalies was reanalyzed by the Solve-RD consortium by evaluating for the presence of single-nucleotide variant, and small insertions and deletions already reported as (likely) pathogenic in ClinVar. Variants were filtered according to frequency, genotype, and mode of inheritance and reinterpreted. Results We identified causal variants in 59 cases (3.9%), 50 of them also raised by other approaches and 9 leading to new diagnoses, highlighting interpretation challenges: variants in genes not known to be involved in human disease at the time of the first analysis, misleading genotypes, or variants undetected by local pipelines (variants in off-target regions, low quality filters, low allelic balance, or high frequency). Conclusion The “ClinVar low-hanging fruit” analysis represents an effective, fast, and easy approach to recover causal variants from exome sequencing data, herewith contributing to the reduction of the diagnostic deadlock