Identification of rare-disease genes using blood transcriptome sequencing and large control cohorts

It is estimated that 350 million individuals worldwide suffer from rare diseases, which are predominantly caused by mutation in a single gene¹. The current molecular diagnostic rate is estimated at 50%, with whole-exome sequencing (WES) among the most successful approaches^2–5. For patients in whom WES is uninformative, RNA sequencing (RNA-seq) has shown diagnostic utility in specific tissues and diseases^6–8. This includes muscle biopsies from patients with undiagnosed rare muscle disorders^6,9, and cultured fibroblasts from patients with mitochondrial disorders⁷. However, for many individuals, biopsies are not performed for clinical care, and tissues are difficult to access. We sought to assess the utility of RNA-seq from blood as a diagnostic tool for rare diseases of different pathophysiologies. We generated whole-blood RNA-seq from 94 individuals with undiagnosed rare diseases spanning 16 diverse disease categories. We developed a robust approach to compare data from these individuals with large sets of RNA-seq data for controls (n = 1,594 unrelated controls and n = 49 family members) and demonstrated the impacts of expression, splicing, gene and variant filtering strategies on disease gene identification. Across our cohort, we observed that RNA-seq yields a 7.5% diagnostic rate, and an additional 16.7% with improved candidate gene resolution.