Gene Expression Profiling of CD34+ Cells in Patients with Myelodysplastic Syndromes
Authors: Pellagatti Andrea , Cazzola Mario , Giagounidis Aristoteles , Perry Janet , Malcovati Luca , Porta Matteo Della , Jädersten Martin , Killick Sally , Hellström-Lindberg Eva , Wainscoat James S. , Boultwood Jacqueline 
Published: October 2019

Abstract
The myelodysplastic syndromes (MDS) are a heterogeneous group of hematopoietic malignancies, characterized by blood cytopenias, ineffective hematopoiesis and a hypercellular bone marrow. We have investigated the gene expression profiles of a large group of patients with MDS in order to better understand the molecular pathogenesis of this disorder. The CD34+ cells obtained from 154 MDS patients and 17 healthy individuals were analyzed using Affymetrix U133 Plus2.0 arrays. 38 genes were up-regulated by >2-fold in at least 77 MDS patients, and pathway analysis using these genes showed that the interferon signalling pathway was significantly deregulated (p=0.0006). Indeed IFIT1, the most up-regulated gene (up-regulated in 110 of 154 MDS patients), is an interferon-stimulated gene (ISG). Other ISGs, which mediate growth inhibitory effects of interferon, such as IFITM1, IFI44L and IFIT3, were markedly up-regulated in the majority of MDS patients. Up-regulation of ISGs is a major feature of MDS and may be responsible for some of the hematological characteristics of this disorder, such as peripheral blood cytopenias. We investigated differences in gene expression that could distinguish MDS patients according to their FAB subtype classification (48 patients with RA, 44 patients with RARS and 62 patients with RAEB). Hierarchical clustering performed using the 773 significantly differentially expressed probe sets identified showed that MDS patients with RARS constitute the most homogeneous group, while MDS patients with RA and RAEB show more overlap. RARS gene expression profile was characterized by up-regulation of mitochondrial-related genes and by down-regulation of ABCB7, a gene mutated in the rare inherited X-linked sideroblastic anemia with ataxia (XLSA/A). Moreover, a good separation between the 20 patients with RARS and the 20 patients with RCMD-RS was obtained by hierarchical clustering using the 86 significantly differentially expressed genes between these two WHO subgroups. One of the most significant genes was MFN1, which is essential for mitochondrial fusion and maintenance of mitochondrial morphology. The association of distinct gene expression profiles with specific cytogenetic groups was also determined, and we were able to separate by hierarchical clustering MDS patients with del(5q), patients with −7/del(7q) and patients with trisomy 8. The expression profile of patients with the del(5q) was characterized by down-regulation of genes mapping to chromosome 5q. Genes differentially expressed in patients with −7/del(7q) include LOX and UBE2H, while genes differentially expressed in patients with trisomy 8 include HRSP12 and TPM4. These findings suggest distinct molecular pathogenetic pathways for MDS patients with del(5q), −7/del(7q) and trisomy 8. In order to identify differences in gene expression associated with MDS disease progression, we compared the 48 patients with early MDS (RA) and the 35 patients with advanced MDS (RAEB2). Hierarchical clustering performed using 1081 significantly differentially expressed probe sets resulted in a good separation between MDS patients with RA and patients with RAEB2. LEF1, a regulator of neutrophilic granulopoiesis, was the most significant differentially expressed gene with higher expression levels in patients with RA and decreasing in patients with RAEB2. Other genes showing higher expression levels in patients with RA, decreasing in patients with RAEB2, include CASC5, a cancer susceptibility candidate gene, and RBBP8, a gene that plays a role in DNA-damage-induced cell cycle checkpoint control. Several genes mapping to the cell cycle pathway were significantly deregulated between early and advanced MDS. This study provides new important insights into the pathophysiology of MDS and represents a first step towards determining pathway signatures in MDS as a guide to targeted therapies.