A SINGLE CELL STUDY OF LYMPHOMA EVOLUTION

Abstract

Loss-of-function mutations of CREBBP are highly prevalent in lymphoid malignancies. We have demonstrated, that loss of Crebbp in murine haematopoietic stem and progenitor cells (HSPC) leads to an increased development of B-cell lymphomas that are preceded by a distinct and functionally validated pre-malignant phase. We now use this mouse model to characterise the molecular mechanisms of lymphoma evolution at single cell resolution and predict that transcriptional heterogeneity related to loss of Crebbp function facilitates transformation. We have probed lymphoma development using (sc)RNA-Seq (10x Genomics) analysis of B220+ splenocytes from four longitudinal stages of disease evolution (age-matched wild type, healthy Crebbp deficient mice, Crebbp deficient pre-malignant and Crebbp deficient malignant mice). This allowed us to generate an atlas of WT B220+ splenocytes, and provided a blueprint to map the subsequent course of malignant progression in Crebbp-/- mice. Cellularly, we identified B1 cells and distinct clusters of aberrant germinal centre B cells to be significantly expanded across disease evolution. Molecularly, differential gene expression profiling identified a number of direct Crebbp targets to be down-regulated during lymphoma evolution, including the Myc-interacting tumour suppressor Bin1. Intriguingly, the fatty acid transporter CD36 (a cancer stem cell marker) is specifically upregulated in the aberrant germinal centre clusters. We are now validating these markers in human datasets and undertaking functional experiments. These data have allowed us to computationally map the trajectory of malignant B-cell transformation, identify critical transcriptional targets differentially expressed during lymphoma evolution and have highlighted potential new targets for therapeutic intervention.