P747: LR-MDS IS CHARACTERIZED BY DIFFERENTIAL EXPRESSION OF INFLAMMASOME-RELATED GENES IN SPECIFIC CELL POPULATIONS

Abstract

Background: Myelodysplastic Syndromes (MDS) are a group of hematopoietic neoplasms characterized by clonal expansion of hematopoietic stem and progenitor cells (HSPC) and peripheral blood cytopenia. MDS can develop from clonal hematopoiesis of indeterminate potential (CHIP) and progress through low-risk (LR) to high-risk (HR)-MDS and further to acute myeloid leukemia (AML). Recent findings emphasize the role of sterile inflammation in the bone marrow (BM) as a driver of neoplastic progression. Our previous analysis of whole BM mononuclear cells (BM-MNC) has revealed distinct expression patterns of inflammasome-related genes, including IL1B and IL18, to be associated with LR-MDS subgroups and clinical features. This raises the prospect of interrupting disease evolution at an early stage by targeted, personalized anti-inflammatory therapy. The development and stratification of such therapies will ultimately require more detailed knowledge of the variety and consequences of inflammation networks in MDS bone marrow. Aims: Our objective was to assess inflammasome-related gene expression levels in individual cell types in the BM of CHIP, LR-MDS and HR-MDS individuals as well as non-CHIP controls in order to determine a) which cell types contribute to the expression of inflammasome-related genes and b) whether specific disease states are associated with consistent patterns of cell-specific gene expression. Methods: Cryopreserved BM-MNC from 3 non-CHIP controls, 3 CHIP individuals, 14 LR-MDS and 5 HR-MDS patients were obtained from the MDS registry and the BoHemE Study (NCT02867085) at the University Hospitals in Dresden and Leipzig. Hematopoietic stem and progenitor cells (HSPC), monocytes, monocytic myeloid-derived suppressor cells (M-MDSC), polymorphonuclear MDSC (PMN-MDSC), B lymphocytes, T lymphocytes and CD45- cells (control) were sorted from thawed BM-MNC on a BD FACS Jazz. RNA was isolated and the gene expression of IL1B, IL18, S100A9, IRAK4, NLRP3, PYCARD, CASP1 and NLRC4 was assessed by RT-qPCR. Results: The majority of the inflammasome-related genes, including IL1B, NLRP3, NLRC4 and S100A9 were expressed in all disease states predominantly by monocytes and M-MDSC (p<0.001). However, IL18 was expressed at the highest level in HSPC. Consistent to our previous analysis of whole BM-MNC, we found an association between cell-specific gene expression and LR-MDS genetics, with HSPC-derived IL18 mRNA being highest in SF3B1-mutated LR-MDS, while monocytic IL1B mRNA was highest in del(5q) LR-MDS. Furthermore, an apparent progression in expression from non-CHIP through to LR-MDS was detected, with mRNA levels of IL1B and NLRP3 in monocytic cells and PYCARD and CASP1 in HSPC increasing from non-CHIP to CHIP between 1.5- and 7.9-fold. From CHIP to LR-MDS, mRNA levels of CASP1 and NLRC4 in monocytic cells and PYCARD both in HPSC and in monocytic cells increased by 1.8- to 2.1-fold. Summary/Conclusion: We identify features of inflammasome-related gene expression in individual cell populations and disease states reflecting the progression from non-CHIP through CHIP to MDS. Our analysis suggests strongly that high resolution studies of gene expression in specific populations or single cells will resolve a spectrum of MDS-related inflammation states relevant to prognosis and personalized therapy. Therefore, we are currently performing RNA-Seq from sorted cell populations in order to extend our analysis beyond the core inflammasome reported here.