CD62L Expression Is Associated With Chronic Lymphocytic Leukemia (CLL) Cell Survival In Vitro and Represents a Novel Therapeutic Target In CLL

Abstract

BackgroundRecent advances in the treatment of chronic lymphocytic leukemia (CLL) have improved overall patient survival, however, the disease remains incurable. There is accumulating evidence that CLL cell resistance to apoptosis is attributable to microenvironmental factors mediated by cell-cell interactions and dysregulation of cytokine signals. Despite this resistance to apoptosis in vivo, CLL cells undergo rapid spontaneous apoptosis when removed from the body. Recently, we and others have demonstrated that this in vitro apoptosis could be reduced by culture of CLL cells at high density and/or in the presence of accessory and stromal cells. A growing body of evidence indicates that alterations in the adhesion properties of neoplastic cells play a pivotal role in the development and progression of CLL. Methods and ResultsTo dissect the complex microenvironmental interactions present in vitro,we profiled the immunophoenotypic changes that occur in long-term CLL PBMC cultures using flow cytometry. Significant changes were observed for 25 antigens, with increases observed in the expression of CD26, CD40, CD58, CD62L and CD103 and decreased expression observed in CD11c, CD32, CD49f, CD62P, CD80, CD106, CD140a, CD141, CD184, CD206 and CD273. The most highly upregulated marker was CD62L (L-selectin, a homing receptor thought to play a role in CLL cell trafficking) and this expression was confirmed in a further subset of patient samples. Using confocal microscopy CD62L expression was present in proliferation and survival niches involved in CLL in the bone marrow and lymph nodes.The pro-survival role of CD62L was examined using a functional blocking antibody which resulted in the significant loss of CLL cell survival. PBMCs from normal healthy controls were unaffected by CD62L antibody treatment, which reinforces that the response is restricted to CLL cells. Furthermore, CD62L antibody treatment caused a specific reduction of CD5+/CD19+ cells with a 49% reduction when compared to untreated PBMCs. This cytotoxic mediated response of CD62L blockade was not abrogated by the presence of stromal cell line HS-5, or endothelial cell line HUVECs suggesting that anti-CD62L therapy may be effective in vivo where pro-survival microenvironmental interactions are intact. We also demonstated a significant increase in cytotoxic responses when anti-CD62L treatment was combined with both fludarabine and mafosfamide compared to either each agent alone, or any two agents combined. ConclusionImmunophenotypic analysis of CLL cultures demonstrated that the expression of several cell surface markers change throughout in vitro culture. These markers are suggestive of cell-cell interactions that clearly provide survival signals. Blocking the activation and homing marker, CD62L, regulates CLL cell survival in vitro and induces cell death equivalent to current CLL chemotherapeutics. Overall, CD62L is a novel prosurvival effector that may represent an attractive therapeutic target in CLL. Disclosures:No relevant conflicts of interest to declare.