Using Proteomics to 1) Identify the Bone Marrow Homing Receptors Expressed on Human Hematopoietic Stem Cells and 2) Elucidate Critical Signaling Pathways Responsible for the Blockage of Hematopoietic Differentiation in Leukemia
AuthorsChin, Chee J.
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AbstractSuccessful hematopoiesis requires the trafficking of hematopoietic stem/progenitor cells (HSPCs) to their bone marrow (BM) niche, where they can differentiate to produce all blood lineages. Leukemia arises when there is a blockage of differentiation and uncontrolled proliferation in the hematopoietic cells during their development. To refine therapies for leukemia, this study sought to improve the homing of healthy donor HSPCs for better transplantation and to find new candidates for differentiating and blocking proliferation in leukemic cells. Characterizing the molecular effectors mediating cell migration forms the basis for improving clinical transplantation of HSPCs. E-selectin/ligand interactions play a critical role in the homing of HSPCs to the BM, however, the identity of E-selectin ligands remains elusive. We aimed to use mass spectrometry (MS) to fully analyze the E-selectin ligands expressed on HSPCs. Immunoprecipitation studies coupled with MS confirmed the expression of three known E-selectin ligands, the hematopoietic cell E-/L-selectin ligand (HCELL), P-selectin glycoprotein ligand-1 (PSGL-1) and CD43, and revealed the presence of many interesting candidates on HSPCs-like cell line and on primary human BM CD34+ cells. The MS dataset represents a rich resource for further characterization of E-selectin ligands, which will lead to improvement of HSPCs transplantation. 4 Understanding the critical pathways underlying the initiation and maintenance of leukemia plays a key role in treating acute myeloid leukemia (AML). Ligation of the glycoprotein, CD44, using monoclonal antibodies or its natural ligand, hyaluronic acid, drives the differentiation of immature leukemic cells towards mature terminally differentiated cells, inhibits their proliferation and in some case induces their apoptosis. The aim of this study is to characterize the phosphoproteome of AML cells in response to CD44-induced differentiation. This will afford novel insights into the signaling events critical for proliferation and differentiation in AML cells. Stable isotope labeling with amino acids in cell culture is used to quantify the changes in the global phosphorylation profile upon CD44 activation. Preliminary results showed that the heavy isotope was fully incorporated and that anti-CD44 induction of myeloid differentiation was not perturbed in our culture conditions. We anticipate that this study will bring forth new candidates for a targeted therapy of AML.