Chemosensitivity Is Differentially Regulated by SDF-1/CXCR4 and SDF-1/CXCR7 Axes in Acute Lymphoblastic Leukemia.

Abstract 2441

SDF-1 (CXCL12) is a chemokine vastly secreted from bone marrow-stromal cells (BM-SC), and plays a pivotal role for multiple biological events in a variety of cells. Hematopoietic stem cells express its receptor CXCR4 at high levels and the SDF-1/CXCR4 system is profoundly implicated in their homing to BM-SC resulting in their chemoresistance. Although important roles of the SDF-1/CXCR4 system for acute myeloid leukemia (AML) have been extensively characterized, its biological implication in acute lymphoblastic leukemia (ALL) remains not fully elucidated. Recently, an alternative SDF-1 receptor CXCR7 was identified, but its functional role in ALL is almost unclear. In the present study, we compared the SDF-1/CXCR4 and SDF-1/CXCR7 systems, particularly about effects on induction of anti-leukemic agent-induced apoptosis, by using established ALL cell lines with MLL gene rearrangement (n=9) and inhibitors against CXCR4 and CXCR7 respectively. Surface CXCR4 expression was very high in all of the cell lines, while surface CXCR7 expression was limited regardless of its abundant cytoplasmic expression. Neither addition of CXCR4 antagonist AMD3100 (1μM) nor CXCR7 antagonist CCX733 (250nM) in culture media showed a significant effect on proliferation of cell lines, suggesting that both systems have not an essential role in proliferation. The chemotactic assay using the Transwell showed a marked migration of cells from the upper to the lower chamber when SDF-1 (10–100nM) or confluent BM-SC line KM104 was present in the lower chamber, and its elicited migration was almost canceled by pretreatment with AMD3100, but only very modestly by pretreatment with CCX733. Next, changes in sensitivity to cytosine arabinoside (Ara-C) after pretreatment of cell lines with either of antagonists was examined on flow cytometry after 48-hour incubation of Ara-C (100nM). Of importance, the Annexin V-labeled apoptotic population with AMD3100 (1μM) pretreatment was significantly decreased as compared with that without pretreatment. It should be noted that when myeloid HL60 cells were used in this experiment, AMD3100 pretreatment rather enhanced Ara-C-induced apoptosis. To address molecular mechanism of AMD3100-induced chemoresistance, the chip analysis was performed and revealed that 24-hour AMD3100 plus Ara-C treatment induced a decrease in RNAs of several caspases and pro-apoptotic molecules as compared with Ara-C alone. In contrast, the Annexin V-labeled apoptotic population with CCX733 pretreatment was significantly increased in a dose-dependant manner of CCX733 (10–500nM) as compared with that without pretreatment. Taken together, SDF-1/CXCR4 and SDF-1/CXCR7 axes are differentially implicated in chemosensitivity of ALL cells that are not adherent to BM-SC; up-regulation of sensitivity via SDF-1/CXCR4 axis vs. down-regulation of sensitivity via SDF-1/CXCR7 axis. Of clinical importance, in contrast to AML, blockade of the SDF-1/CXCR4 axis in ALL facilitate chemoresistance, suggesting that a clinical application of CXCR4 antagonist currently undergoing in AML patients should be carefully considered in ALL patients.