Study of the Roles of SDF1 and CXCR4 in Hematopoiesis

Division Of Basic Sciences - Nci

Investigators

Linked publications, trials & patents

Abstract

Previous studies from many groups, including our group, have identified a critical role of the chemokine CXCL12/SDF1 and its receptor CXCR4 in the retention and mobilization of myeloid cells from the bone marrow. CXCL12 is mostly produced by the non-hematopoietic cells of bone marrow, particular in the CAR (CXCL12 abundant reticular) and in other mesenchymal cells in the bone marrow. The bone marrow hematopoietic cells generally express CXCR4, and the signaling interaction between CXCL12/CXCR4 serves to retain hematopoietic cells in the bone marrow. G-CSF is a critical physiological regulator of granulopoiesis: mice carrying homozygous deletions of colony-stimulating factor (G-CSF) or its receptor are severely neutropenic, and dominant-negative mutations of G-CSFR have been linked to severe defects of granulopoiesis. Administration of G-CSF induces an expansion of myeloid lineage cells in the bone marrow, and promotes the release of mature myeloid cells and hematopoietic progenitor cells from the bone marrow to the peripheral blood. Thus, G-CSF at pharmacologic concentrations is widely used to induce granulopoiesis and to mobilize hematopoietic progenitors to the peripheral blood. Functionally, G-CSF reduces CXCR4 and CXCL12 levels in the bone marrow through a variety of mechanisms. As a consequence, the glue that retains hematopoietic cells in the bone marrow is loosened and massive exit of mature and immature hematopoietic cells is achieved. A CXCR4 competitive inhibitor, AMD3100/Plerixafluor, has been approved by FDA as a mobilizing agent for hematopoietic precursors in conjunction with G-CSF. Recently, our studies on HSPC mobilization have detected an important role of the receptor/ligand pair EphrinB2/EphB4. We unveiled the mutually exclusive bone marrow distribution of EphB4 receptors in the sinusoids and EphrinB2 ligands in hematopoietic cells, and discovered an EphB4/EphrinB2-dependent pathway that controls HSPCs mobilization. Blockade of EphB4/EphrinB2 reduced HSPCs and other myeloid cells mobilization to the circulation. In murine cancer models, in which hematopoietic cells derived from the bone marrow promote tumor growth, EphB4/EphrinB2 blockade reduced tumor infiltration with HSPCs and tumor progression. These results identify EphB4/EphrinB2 signaling as critical to hematopoietic cells mobilization from bone marrow and provide a new strategy for reducing cancer progression by targeting the bone marrow. Other ongoing studies stem from previous observations in the neuronal system, particularly in the cerebellum, showing a biochemical link between EphrinB2 and CXCR4 that regulates cerebellar cell motility. More specifically, these studies showed that PDZ-dependent signaling by EphrinB2 recruits the PDZ-containing RGS3 protein, which in turn can bind and shut down CXCR4 signaling. In so doing, active EphrinB2 regulates CXCL12-induced motility of CXCR4-positive neuronal cells. We hypothesized that this interaction may also occur in the context of the bone marrow where CXCL12 (in non-hematopoietic cells), CXCR4 and EphrinB2 (in hematopoietic cells) are abundant. In proof-of-principle experiments we have documented the proximal co-localization of CXCR4 and EphrinB2 in mouse hematopoietic cells. Initial mapping of sites where these molecules associate in the bone marrow showed a broad pattern, with some degree of signal concentration at more proximal to the bone. This mapping will continue by imaging in WT and genetically-modified mouse lines. Other ongoing studies aim at defining the epigenome of hematopoietic cells at bulk and single-cell resolution. Current technological limitations have prevented effective multiplex analysis of the epigenome at a single-cell level. To overcome this limitation, we have developed a new technology that relies on the establishment of re-usable single-cells and proximity ligation-based mapping of epigenetic marks to the genome. This technology has successfully been applied to single-cell analysis of a model hematopoietic cell line (K562 cells). After further development, we intend to utilize this technique to evaluate the epigenome of hematopoietic cells during cell division.