Role of Genetic Haploinsufficiency in the Biology and Targeted Therapy of Del(5q) Myelodysplastic Syndrome

The studies presented in this thesis focused on the identification of genes that play a central role in the pathogenesis of myelodysplastic syndromes with a deletion of the short arm (q) of chromosome 5, the del(5q) MDS. The overall goal of the projects was to gain a better understanding of how gene haploinsufficiency in (del)5q MDS leads to a clonal advantage, ineffective hematopoiesis and how haploinsufficiency can be targeted. Heterozygous deletion of RPS14 occurs in del(5q) myelodysplastic syndrome (MDS) and has been linked to impaired erythropoiesis, characteristic of this disease subtype. So far, it was not well understood how ribosomal haploinsufficiency affects protein synthesis and if differentially translated proteins have a impact on the erythroid differentiation defect (chapter 1). We generated a murine model with conditional inactivation of Rps14 and demonstrated a p53-dependent erythroid differentiation defect with apoptosis at the transition from polychromatic to orthochromatic erythroblasts resulting in age- and erythroid stress dependent progressive anemia, megakaryocyte dysplasia, and loss of hematopoietic stem cell (HSC) quiescence (chapter 2). Protein synthesis was significantly reduced in Rps14 haploinsufficient hematopoietic stem cells and in particular in erythroid progenitor cells relative to wild-type cells. As assessed by quantitative proteomics, Rps14 haploinsufficient erythroblasts expressed higher levels of proteins involved in innate immune signaling, notably the heterodimeric S100 calcium-binding proteins S100a8 and S100a9 (alarmins). S100a8 is functionally involved in the erythroid defect caused by the Rps14 deletion, as addition of recombinant S100a8 was sufficient to induce an erythroid differentiation defect in wild-type erythroid cells (phenocopy), and genetic inactivation of S100a8 expression rescued the erythroid differentiation defect of Rps14-haploIn the current study it was our goal to focus on important design criteria including (1) the opportunity for reproducible and user-defined properties such as the choice of substrate, extracellular matrix, cell types, and degradability; (2) the ability to induce vascularisation; (3) accessibility and suitability for complementary histological/cytological analysis; and, most importantly, (4) the functional ability to capture and retrieve endogenous hematopoietic cells efficiently

Puzzling pieces of chromosome 7 loss or deletion

In this issue of Blood, Inaba et al review the challenges and questions to be answered in the molecular and functional dissection of loss of chromosome 7 (monosomy 7 [-7]) and deletion of a segment of the long arm (del(7q)) found in patients with various syndromes involving the myeloid blood cell lineage

Understanding deregulated cellular and molecular dynamics in the haematopoietic stem cell niche to develop novel therapeutics for bone marrow fibrosis

Bone marrow fibrosis is the continuous replacement of blood-forming cells in the bone marrow with excessive scar tissue, leading to failure of the body to produce blood cells and ultimately to death. Myofibroblasts are fibrosis-driving cells and are well characterized in solid organ fibrosis, but their role and cellular origin in bone marrow fibrosis have remained obscure. Recent work has demonstrated that Gli1+ and leptin receptor+ mesenchymal stromal cells are progenitors of fibrosis-causing myofibroblasts in the bone marrow. Genetic ablation or pharmacological inhibition of Gli1+ mesenchymal stromal cells ameliorated fibrosis in mouse models of myelofibrosis. Conditional deletion of the platelet-derived growth factor (PDGF) receptor-α (PDGFRA) gene (Pdgfra) and inhibition of PDGFRA by imatinib in leptin receptor+ stromal cells suppressed their expansion and ameliorated bone marrow fibrosis. Understanding the cellular and molecular mechanisms in the haematopoietic stem cell niche that govern the mesenchymal stromal cell-to-myofibroblast transition and myofibroblast expansion will be critical to understand the pathogenesis of bone marrow fibrosis in both malignant and non-malignant conditions, and will guide the development of novel therapeutics. In this review, we summarize recent discoveries of mesenchymal stromal cells as part of the haematopoietic niche and as myofibroblast precursors, and discuss potential therapeutic strategies in the specific targeting of fibrotic transformation in bone marrow fibrosis

Malignant Transformation Involving CXXC4 Mutations Identified in a Leukemic Progression Model of Severe Congenital Neutropenia

Olofsen et al. show that acquisition of a mutation in Cxxc4 results in increased CXXC4 protein levels, reduced TET2 protein, increased inflammatory signaling, and leukemic progression of a CSF3R/RUNX1 mutant mouse model of severe congenital neutropenia (SCN).Severe congenital neutropenia (SCN) patients treated with CSF3/G-CSF to alleviate neutropenia frequently develop acute myeloid leukemia (AML). A common pattern of leukemic transformation involves the appearance of hematopoietic clones with CSF3 receptor (CSF3R) mutations in the neutropenic phase, followed by mutations in RUNX1 before AML becomes overt. To investigate how the combination of CSF3 therapy and CSF3R and RUNX1 mutations contributes to AML development, we make use of mouse models, SCN-derived induced pluripotent stem cells (iPSCs), and SCN and SCN-AML patient samples. CSF3 provokes a hyper-proliferative state in CSF3R/RUNX1 mutant hematopoietic progenitors but does not cause overt AML. Intriguingly, an additional acquired driver mutation in Cxxc4 causes elevated CXXC4 and reduced TET2 protein levels in murine AML samples. Expression of multiple pro-inflammatory pathways is elevated in mouse AML and human SCN-AML, suggesting that inflammation driven by downregulation of TET2 activity is a critical step in the malignant transformation of SCN

Heterogeneous bone-marrow stromal progenitors drive myelofibrosis via a druggable alarmin axis

Functional contributions of individual cellular components of the bone-marrow microenvironment to myelofibrosis (MF) in patients with myeloproliferative neoplasms (MPNs) are incompletely understood. We aimed to generate a comprehensive map of the stroma in MPNs/MFs on a single-cell level in murine models and patient samples. Our analysis revealed two distinct mesenchymal stromal cell (MSC) subsets as pro-fibrotic cells. MSCs were functionally reprogrammed in a stage-dependent manner with loss of their progenitor status and initiation of differentiation in the pre-fibrotic and acquisition of a pro-fibrotic and inflammatory phenotype in the fibrotic stage. The expression of the alarmin complex S100A8/S100A9 in MSC marked disease progression toward the fibrotic phase in murine models and in patient stroma and plasma. Tasquinimod, a small-molecule inhibiting S100A8/S100A9 signaling, significantly ameliorated the MPN phenotype and fibrosis in JAK2V617F-mutated murine models, highlighting that S100A8/S100A9 is an attractive therapeutic target in MPNs.Leimkühler and colleagues demonstrate that mesenchymal stromal progenitor cells are fibro