P213: Interdisciplinary variant re-classification: FGFR3 as an example of genotypic investigation in suspected skeletal dysplasia population

Introduction:The fibroblast growth factor receptor 3 (FGFR3) gene is linked to skeletal dysplasias like achondroplasia (ACH) and hypochondroplasia (HCH), inherited in an autosomal dominant pattern.Most ACH cases (80%) and a substantial portion of HCH cases (50-70%) are de novo variants, highlighting the genetic complexity of these disorders.Despite advancements in understanding disease mechanisms and developing treatments, inconclusive genetic test results remain a significant challenge, as shown by a recent cohort study where 5,011 patients with skeletal disorders received gene panel testing, which resulted in 80 inconclusive FGFR3 molecular results (compared to 232 confirmed molecular diagnoses in FGFR3).Historically, interpreting uncertain results was limited in part by the lack of high-throughput functional assays, which are particularly useful in cases of limited evidence from clinical or other data per classification criteria.However, new assay methods are emerging.In parallel, real-world data from biobanks, registries, and clinical records provide valuable resources for evidence gathering to support variant reclassification.This effort requires coordinated involvement from healthcare professionals, researchers, statisticians, and laboratories.To ensure accurate reclassification, the process must follow predefined, evidence-based criteria and be conducted by impartial reviewers.Methods: To demonstrate the impact of a collaborative approach to reclassifying FGFR3 variants of uncertain significance (VUS), we initiated a project with the following steps:1. Data Collection: De-identified clinical data associated with FGFR3 VUS were collected via the Discover Dysplasias no-cost testing program from December 2019 to April 2024 (LabCorp Genetics, formerly Invitae).2. Assessment of Existing Databases: Data from external sources, including the UK Biobank (UKBB), gnomAD, and other relevant databases, were assessed to identify frequency, phenotypic correlations, and potential pathogenicity of the variants.3. Clinician Researcher Involvement: Clinician researchers with expertise in skeletal dysplasias and genetic testing contributed to the data interpretation and provided clinical context for the variants under review.4. Objective Review Process: An established process was used to review all available evidence objectively.This included using pre-defined criteria for variant classification to ensure consistency and impartiality in re-evaluation decisions.Results: A total of 168 inconclusive FGFR3 molecular results were reviewed, involving 119 unique nucleotide variants of uncertain significance (113 unique predicted protein coding variants; 137 unique ordering clinicians).As of November 15 th , 2024, 53 clinicians were reached out to and 20 clinician interviews have been conducted.Data from ClinVar, UK Biobank (UKBB), and gnomAD have been reviewed.Clinician interviews included patient chart review and family pedigree discussion, which provided clear phenotypic evidence to support potential variant reclassification.To illustrate the evidence used in the process and demonstrate potential impact to patient care, we highlight examples including p.Val555Leu (4 patients; 2 nucleotide variants), p.Ser249Phe (2 patients; 1 nucleotide variant), and p.Asp758Asn (8 patients; 1 nucleotide variant).Conclusion: In many cases, concerted collection and analysis of clinical and laboratory data are required to facilitate variant classification (in addition to publicly available sources).Further interdisciplinary work and collaboration across the healthcare system can improve variant classification and patient care.Where available, functional assays can provide the crucial missing evidence to support variant classification.