The unsequenceable: novel, easy to adapt method for the major ADTKD gene MUC1

Introduction: Clinical symptoms of autosomal dominant tubulointerstitial kidney disease (ADTKD) are often unspecific and patients are not, late or incorrectly diagnosed. Progressive tubulointerstitial fibrosis may finally lead to end-stage renal failure. An early and correct diagnosis is of major importance, however genetic analysis is challenged by the complex architecture of MUC1 with a variable number of tandem repeats (VNTR). Due to this complexity, very little is known about MUC1 variants beyond the „typical“ 1 bp frameshift alteration insC. We report the use of a novel, easy to adapt methodology called MGM-MUC1-VNTR as a supplementary diagnostic tool for short-read-sequencing (SRS) data of MUC1 in a cohort of 434 patients. Methods: A large cohort of patients diagnosed with ADTKD or allied chronic kidney diseases initially underwent SRS based custom-panel or whole exome sequencing (WES) analysis. SRS data were screened in parallel with the established VNtyper tool and our MGM-MUC1-VNTR algorithm to detect MUC1 VNTR frameshift mutations beyond the well-established insC variant. Novel putative variants were further validated by enrichment-based long-read sequencing. Results: With the implementation of MGM-MUC1-VNTR we were able to detect various atypical frameshift variants in previously unsolved cases of existing SRS data. Convincing larger and smaller frameshift deletions were detected in nine samples out of six unrelated pedigrees. In three of these families the causative mutations were confirmed by sequence-capture based LRS enabling refinement and deeper analysis of existing SRS data as well as definition of the VNTR composition. Conclusions: Our novel approach allows for easy and efficient detection of a wide range of pathogenic MUC1 alterations complementing established approaches. This algorithm can be easily adapted and implemented in routine diagnostics and will empower labs to (re-) analyze their (already existing) data with minimal cost and effort. Due to its ease of use our approach will help to broaden the mutational spectrum and identify further patients which might be of increasing importance in view of potential emerging genotype-driven therapeut