Due to application of high-throughput sequencing techniques in diagnostics and clinical research, we identify more and more patients with variants of unknown significance (VUS) in known cancer
associated genes. The impact of these variants on tumorigenesis can only be determined using
functional tests that are specific for the gene in question. Correct classification of germline VUS is of utmost importance for diagnostics of patients and their families with hereditary cancers, allowing for individualized surveillance strategies and treatment decisions, ultimately leading to prevention of cancer in these families. We recently demonstrated that metabolic screening using mass spectrometry based measurements of tumor metabolites facilitates functional characterization of variants in Krebs cycle genes and can be utilized to reclassify VUS in such genes to improve clinical decision-making. This collaborative project aims to establish and utilize in vitro systems to reliable determine the pathogenicity of VUS in known cancer genes involved in metabolism and pseudohypoxia by combining state-of-the-art genetic engineering and analytical approaches.
- Establish isogenic cell models using CRISPR/Cas9 technology to evaluate the functionality of variants of unknown significance in SDHB, FH, and VHL.
- Set up and validate our in vitro pipeline using metabolite screening and pseudohypoxia activity assays.
- Functional in vitro testing of selected VUS from our available patient cohorts with our pipeline.
- Identify targetable phenotypes in our isogenic knockout cell lines using omics.
- The long-term goal of the presented project is to improve genetic diagnostics of patient with hereditary tumor syndromes by classifying VUS as either benign or pathogenic.