The tumor microenvironment impairs CLL response to the exit signal from lymphoid tissues, sphingosine 1-phospate (S1P)

M. BORGE; F. REMES LENICOV; P.R. NANNINI; M.M. DE LOS RÍOS ALICANDU; H. FERNANDEZ GNECCO; R.F. BEZARES; P.E. MORANDE; P. OPPEZZO; M. GIORDANO; R. GAMBERALE

Colonia

Workshop; XV International Workshop on Chronic Lymphocytic Leukemia; 2013

European Haematology Association

The leukemic clone of CLL patients is characterized by a subtle balance between cells circulating in the blood and cells located in permissive niches in lymphoid organs, where CLL cells are activated and proliferate in close contact with stromal cells, nurse like cells (NLC) and activated T cells. While numerous reports have studied the mechanisms involved in CLL entry to lymphoid tissues, there is scarce information regarding the mechanisms involved in the leukemic clone exit from this permissive niche to circulation. Sphingosine-1 phosphate (S1P) is a bioactive sphingolipid which exerts most of its function as a specific ligand for a family of five G-protein-coupled receptors, being S1PR1 the main receptor involved in lymphocyte exit from lymphoid tissues. Our central hypothesis assumes that, by acting through S1PR1, S1P regulates the egress of the leukemic clone from lymphoid tissues to peripheral blood, a process that may be delayed by activatory signals delivered by the tumor microenvironment. To test our hypotheses we aimed to determine whether key signals from the microenvironment reduce the migratory response towards S1P. Thus, purified CLL cells where cultured with different stimulus, such as CXCL12, fibroblast CD40L+ and antibodies anti-IgM in order to induce BCR crosslinking. Then, the in vitro chemotaxis assay towards S1P was performed by employing the transwell system. We found that CXCL12 and the crosslinking of CD40 or surface IgM significantly reduce the migratory response towards S1P (Fig. 1 A) and diminish the expression of S1PR1 evaluated by qPCR (Fig. 1 B) and flow cytometry (Fig. 1 C). Similarly, co-cultures with autologous NLC, which represent an accurate model for the microenvironment in secondary lymphatic tissues, reduced the migratory response towards S1P (Fig. 1 D) and the expression of S1PR1 (Fig. 1 E and F). In conclusion, our data show that the supportive microenvironment reduce the expression of S1PR1 and the in vitro chemotaxis assay towards S1P. Altogether, these results suggest that the activation within lymphoid tissues may reduce the in vivo exit of CLL cells to circulation, extending the dwell time of the leukemic clone in the tissues that favor their accumulation. A better understanding of the role of S1P in CLL will contribute to the current knowledge of leukemic cell biology, will provide insights into the progression of the disease and might help to find potential targets for novel therapeutic treatments.