Evidence for 28 genetic disorders discovered by combining healthcare and research data.

De novo mutations in protein-coding genes are a well-established cause of developmental disorders1. However, genes known to be associated with developmental disorders account for only a minority of the observed excess of such de novo mutations1,2. Here, to identify previously undescribed genes associated with developmental disorders, we integrate healthcare and research exome-sequence data from 31,058 parent-offspring trios of individuals with developmental disorders, and develop a simulation-based statistical test to identify gene-specific enrichment of de novo mutations. We identified 285 genes that were significantly associated with developmental disorders, including 28 that had not previously been robustly associated with developmental disorders. Although we detected more genes associated with developmental disorders, much of the excess of de novo mutations in protein-coding genes remains unaccounted for. Modelling suggests that more than 1,000 genes associated with developmental disorders have not yet been described, many of which are likely to be less penetrant than the currently known genes. Research access to clinical diagnostic datasets will be critical for completing the map of genes associated with developmental disorders.

Autosomal dominant ApoA4 mutations present as tubulointerstitial kidney disease with medullary amyloidosis.

Sporadic cases of apolipoprotein A-IV medullary amyloidosis have been reported. Here we describe five families found to have autosomal dominant medullary amyloidosis due to two different pathogenic APOA4 variants. A large family with autosomal dominant chronic kidney disease (CKD) and bland urinary sediment underwent whole genome sequencing with identification of a chr11:116692578 G>C (hg19) variant encoding the missense mutation p.L66V of the ApoA4 protein. We identified two other distantly related families from our registry with the same variant and two other distantly related families with a chr11:116693454 C>T (hg19) variant encoding the missense mutation p.D33N. Both mutations are unique to affected families, evolutionarily conserved and predicted to expand the amyloidogenic hotspot in the ApoA4 structure. Clinically affected individuals suffered from CKD with a bland urinary sediment and a mean age for kidney failure of 64.5 years. Genotyping identified 48 genetically affected individuals; 44 individuals had an estimated glomerular filtration rate (eGFR) under 60 ml/min/1.73 m2, including all 25 individuals with kidney failure. Significantly, 11 of 14 genetically unaffected individuals had an eGFR over 60 ml/min/1.73 m2. Fifteen genetically affected individuals presented with higher plasma ApoA4 concentrations. Kidney pathologic specimens from four individuals revealed amyloid deposits limited to the medulla, with the mutated ApoA4 identified by mass-spectrometry as the predominant amyloid constituent in all three available biopsies. Thus, ApoA4 mutations can cause autosomal dominant medullary amyloidosis, with marked amyloid deposition limited to the kidney medulla and presenting with autosomal dominant CKD with a bland urinary sediment. Diagnosis relies on a careful family history, APOA4 sequencing and pathologic studies.

De novo mutations in regulatory elements in neurodevelopmental disorders.

We previously estimated that 42% of patients with severe developmental disorders carry pathogenic de novo mutations in coding sequences. The role of de novo mutations in regulatory elements affecting genes associated with developmental disorders, or other genes, has been essentially unexplored. We identified de novo mutations in three classes of putative regulatory elements in almost 8,000 patients with developmental disorders. Here we show that de novo mutations in highly evolutionarily conserved fetal brain-active elements are significantly and specifically enriched in neurodevelopmental disorders. We identified a significant twofold enrichment of recurrently mutated elements. We estimate that, genome-wide, 1-3% of patients without a diagnostic coding variant carry pathogenic de novo mutations in fetal brain-active regulatory elements and that only 0.15% of all possible mutations within highly conserved fetal brain-active elements cause neurodevelopmental disorders with a dominant mechanism. Our findings represent a robust estimate of the contribution of de novo mutations in regulatory elements to this genetically heterogeneous set of disorders, and emphasize the importance of combining functional and evolutionary evidence to identify regulatory causes of genetic disorders.

Pathogenicity and selective constraint on variation near splice sites.

Mutations that perturb normal pre-mRNA splicing are significant contributors to human disease. We used exome sequencing data from 7833 probands with developmental disorders (DDs) and their unaffected parents, as well as more than 60,000 aggregated exomes from the Exome Aggregation Consortium, to investigate selection around the splice sites and quantify the contribution of splicing mutations to DDs. Patterns of purifying selection, a deficit of variants in highly constrained genes in healthy subjects, and excess de novo mutations in patients highlighted particular positions within and around the consensus splice site of greater functional relevance. By using mutational burden analyses in this large cohort of proband-parent trios, we could estimate in an unbiased manner the relative contributions of mutations at canonical dinucleotides (73%) and flanking noncanonical positions (27%), and calculate the positive predictive value of pathogenicity for different classes of mutations. We identified 18 patients with likely diagnostic de novo mutations in dominant DD-associated genes at noncanonical positions in splice sites. We estimate 35%-40% of pathogenic variants in noncanonical splice site positions are missing from public databases.

Contribution of retrotransposition to developmental disorders.

Mobile genetic Elements (MEs) are segments of DNA which can copy themselves and other transcribed sequences through the process of retrotransposition (RT). In humans several disorders have been attributed to RT, but the role of RT in severe developmental disorders (DD) has not yet been explored. Here we identify RT-derived events in 9738 exome sequenced trios with DD-affected probands. We ascertain 9 de novo MEs, 4 of which are likely causative of the patient's symptoms (0.04%), as well as 2 de novo gene retroduplications. Beyond identifying likely diagnostic RT events, we estimate genome-wide germline ME mutation rate and selective constraint and demonstrate that coding RT events have signatures of purifying selection equivalent to those of truncating mutations. Overall, our analysis represents a comprehensive interrogation of the impact of retrotransposition on protein coding genes and a framework for future evolutionary and disease studies.

Towards programming immune tolerance through geometric manipulation of phosphatidylserine.

The possibility of engineering the immune system in a targeted fashion using biomaterials such as nanoparticles has made considerable headway in recent years. However, little is known as to how modulating the spatial presentation of a ligand augments downstream immune responses. In this report we show that geometric manipulation of phosphatidylserine (PS) through fabrication on rod-shaped PLGA nanoparticles robustly dampens inflammatory responses from innate immune cells while promoting T regulatory cell abundance by impeding effector T cell expansion. This response depends on the geometry of PS presentation as both PS liposomes and 1 micron cylindrical PS-PLGA particles are less potent signal inducers than 80 × 320 nm rod-shaped PS-PLGA particles for an equivalent dose of PS. We show that this immune tolerizing effect can be co-opted for therapeutic benefit in a mouse model of multiple sclerosis and an assay of organ rejection using a mixed lymphocyte reaction with primary human immune cells. These data provide evidence that geometric manipulation of a ligand via biomaterials may enable more efficient and tunable programming of cellular signaling networks for therapeutic benefit in a variety of disease states, including autoimmunity and organ rejection, and thus should be an active area of further research.