London Calling 2023: The potential clinical utility of amplicon and targeted nanopore sequencing for rare disease diagnosis

Next-generation sequencing has driven research into rare diseases and molecular diagnostics for over a decade. Up to 45% of inherited retinal disease patients remain unsolved after clinical testing, with variants of uncertain significance (VUS), non-coding variants, whole-genome sequencing (WGS) intractable genes, and structural rearrangements thought to contribute.

We investigated the potential clinical utility of nanopore sequencing in a case-driven study to improve the detection and characterisation of variants in WGS unsolved patients recruited from the Inherited Eye Disease clinics at Moorfields Eye Hospital. Amplicon sequencing of retinal gene transcripts from patient blood demonstrated the previously undetectable splicing effect of coding and non-coding VUS. Targeted nanopore sequencing enabled read-through of the WGS-intractable OPN1LW/OPN1MW gene array, distant variant phasing, and detection of structural rearrangement. Nanopore sequencing is effective where traditional methods have failed to conclude diagnostic odysseys and may be an effective tool in the battery of molecular diagnostic tests.