Project Details

Detection of Hidden Variants in Cystic Kidney Disease Using Long-Read Sequencing in Serum and Urine

Alexandra Keefe, MD, PhD

Summary

BACKGROUND: Autosomal dominant polycystic kidney disease (ADPKD) is the most common heritable kidney disease in children, however standard DNA sequencing misses many disease-causing variants. Long-read sequencing (LRS) is an emerging technology which reads much longer DNA molecules, and can identify variants missed by standard testing, such as structural variants (i.e. deletions, insertions, inversions). Having a precise molecular diagnosis in AKPKD is critical because genetic etiology influences phenotypic severity and long-term treatment options. GAP: Detecting missing variants is best performed on affected tissue (i.e. kidney cells), but biopsies are rarely performed in AKPKD since the disease is diagnosed via non-invasive ultrasound. Urine, which contains kidney cell-derived DNA, provides a unique specimen for DNA sequencing, as it simultaneously allows for tissue-specific genetic testing but through a non-invasive modality – a type of “liquid biopsy”. Performing LRS of urine-derived DNA for the detection of genetic causes of ADPKD is entirely novel, and potentially can provide a cost-effective, fast, non-invasive first line clinical approach for determining the molecular cause of ADPKD, which may be superior to current clinical standards. HYPOTHESIS: I hypothesize that by combining emerging new technology (LRS) and tissue-specific sequencing (urine) in unsolved ADPKD cases, I will identify previously “hidden” disease-causing variants. METHODS: To test this hypothesis I will 1) Validate that targeted LRS can detect the same pathogenic variant in both serum and urine samples from patients with confirmed pathogenic variants in known ADPKD genes and 2) Perform targeted LRS of serum and urine-derived DNA in individuals with AKPKD with no known pathogenic variant after standard commercial DNA sequencing. RESULTS: Pending. IMPACT: Aim 1 will provide a proof of concept that urine-derived DNA is non-inferior to blood-based sequencing in ADPKD, and Aim 2 will show that candidate disease-causing variants are missed by standard clinical testing, but can be identified by LRS in urine. Together, these aims will demonstrate that LRS of urine-derived DNA can shorten time to diagnosis, increase the diagnostic rate, and reduce barriers to comprehensive testing, which are all crucial for clinical decision making as early intervention impacts long-term outcomes for children with ADPKD.