CD38 Induces Homing of Chronic Lymphocytic Leukemia Cells to the Lymphoid Organs through a Functional Interplay with CXCR4.

Abstract 2328

Poster Board II-305

Chronic lymphocytic leukemia (CLL) is characterized by the expansion of a monoclonal population of mature B lymphocytes expressing surface CD5, CD23 and low levels of immunoglobulin (Ig). Originating in the bone marrow (BM), neoplastic cells accumulate in blood (the circulating pool) and in lymphoid organs (the residential pool). Data obtained ex vivo and in patients suggest that the proliferative core of the disease dwells within the residential pool, while little or no proliferation is seen among circulating CLL cells. Threrefore, homing of chronic lymphocytic leukemia (CLL) cells to sites favoring growth is a critical step in disease progression.

Human CD38 is a pleiotropic glycoprotein belonging to a complex family of genes coding for enzymes of the cell surface and involved in the catabolism of extracellular nucleotides. Along with the absence of mutations in the immunoglobulin variable region heavy chain gene (IgVH) and expression of the cytoplasmic kinase ZAP-70, CD38 expression above a critical threshold represents a reliable negative prognostic marker for CLL patients. The working hypothesis of our group is that CD38 is not merely a marker in CLL, but a cell surface receptor and adhesion molecule directly involved in the delivery of growth signals. Prolonged interactions between CD38 and its non-substrate ligand CD31 under static conditions increase survival of the CLL clone and sustain its proliferation in vitro. Here we show that CD38, is directly involved in chemokine-mediated homing of CLL cells. CD38 signals facilitate short- (ERK1/2 phosphorylation) and long-term (chemotaxis) effects induced by the CXCL12 chemokine. This was first shown by testing functional responses to CXCL12 in a large cohort of clinically and molecularly characterized patients. The superior chemotactic performance of CD38⁺ CLL cells was confirmed by comparing the CD38⁺ and CD38^- leukemic components within the same patient presenting with a bimodal clone. Assays using i) a specific inhibitor of CD38 enzymatic activities, ii) a panel of agonistic and blocking monoclonal antibodies (mAbs) and iii) microarray technology indicate that increased homing depends on the receptor functions of CD38. Indeed, interactions between CD38 and its non-substrate ligand CD31 define a genetic signature characterized by modulation of pathways involved in proliferation and migration of CLL cells. Conversely, blocking CD38 with domain-specific mAbs significantly weakens CXCL12 responses. This is attributed to physical proximity on the cell membrane between CD38 and CXCR4 (the CXCL12 receptor). The existence of a CD38/CXCR4 complex was demonstrated by i) co-immunoprecipitation, ii) confocal microscopy and iii) direct measurement of CXCL12 binding to its receptor. The relevance of CD38 in homing is substantiated by results from an in vivo mouse model, where CLL cells are allowed to home to lymphoid organs for a short period of time. The compromised homing ability of CLL cells to spleen and BM in the presence of blocking anti-CD38 mAbs confirms in vivo the in vitro evidence. Furthermore, the lack of effects induced by an isotype-matched mAb with the same anti-CD38 specificity convincingly rules out the possibility that the effects observed are secondary to sequestration of mAb-coated cells. Future studies will address the issue of whether CD38 may become a therapeutic target for selected CLL patients.