1020 – HEMATOPOIETIC STEM CELL CLONAL TRACKING BY WHOLE GENOME SEQUENCING IDENTIFIES MULTIPLE INDEPENDENT SOMATIC GENETIC RESCUE MECHANISMS IN BONE MARROW FAILURE DISORDERS.

The congenital bone marrow failure syndrome Shwachman-Diamond Syndrome (SDS) is characterized by defective ribosome biogenesis, with a significant risk of developing a severe myelodysplastic syndrome or leukemia. The most common causal germ line mutations are in the SBDS gene. It remains unclear how the germline ribosomal dysfunction/stress affects individual hematopoietic stem cells and contributes to the aberrant population dynamics that drive specific clonal expansions and eventual leukemic transformation. To address this, we adopted a clonal tracking approach using whole genome sequencing of single hematopoietic stem and progenitor cell (HSPC)-derived colonies followed by phylogenetic tree construction to assess clonal relatedness in 9 SDS patients.  The mean mutation burden in SDS HSPCs increased linearly with age and the majority of mutations were C >T variants, consistent with mutational signatures in ageing. Single nucleotide variants in annotated genes across patients were found in known drivers TP53, EIF6 and other ribosomal proteins: all of which were mutually exclusive in different HSPC clones.  Distinct mutations in the same genes were found within the same patients, demonstrating that individual clones must acquire independent genetic rescue mechanisms. Whole genome sequencing permitted us to uncover a high degree of intra-patient heterogeneity, charting multiple independent paths that individual cells can take to combat defective ribosome deficiency caused by SBDS loss. This study sets the stage for functional analysis of potential leukemia driver mutations and longitudinal tracking of clonal dynamics throughout disease progression and re-enforces the value of using whole genome sequencing to track individual stem cell clones in disease.