sphingosine-1-phosphate and Leukemia--Lymphocytic--Chronic--B-Cell

BCR signaling pathway inhibitors such as ibrutinib, idelalisib, and fostamatinib (respective inhibitors of Bruton's tyrosine kinase, PI3Kδ, and spleen tyrosine kinase) represent a significant therapeutic advance in B cell malignancies, including chronic lymphocytic leukemia (CLL). These drugs are distinctive in increasing blood lymphocytes while simultaneously shrinking enlarged lymph nodes, suggesting anatomical redistribution of CLL cells from lymph nodes into the blood. However, the mechanisms underlying this phenomenon are incompletely understood. In this study, we showed that the egress receptor, sphingosine-1-phosphate (S1P) receptor 1 (S1PR1), was expressed at low levels in normal germinal centers and CLL lymph nodes in vivo but became upregulated on normal B cells and, to a variable and lesser extent, CLL cells following in vitro incubation in S1P-free medium. Spontaneous recovery of S1PR1 expression on normal B and CLL cells was prevented by BCR cross-linking, whereas treatment of CLL cells with idelalisib increased S1PR1 expression and migration toward S1P, the greatest increase occurring in cases with unmutated IgH V region genes. Intriguingly, ibrutinib and fostamatinib had no effect on S1PR1 expression or function. Conversely, chemokine-induced migration, which requires integrin activation and is essential for the entry of lymphocytes into lymph nodes as well as their retention, was blocked by ibrutinib and fostamatinib, but not idelalisib. In summary, our results suggest that different BCR signaling inhibitors redistribute CLL cells from lymph nodes into the blood through distinct mechanisms: idelalisib actively promotes egress by upregulating S1PR1, whereas fostamatinib and ibrutinib may reduce CLL cell entry and retention by suppressing chemokine-induced integrin activation.

Topics: Adenine; Agammaglobulinaemia Tyrosine Kinase; Aminopyridines; Antineoplastic Agents; B-Lymphocytes; Case-Control Studies; Cell Movement; Class I Phosphatidylinositol 3-Kinases; Gene Expression Regulation, Leukemic; Germinal Center; Human Umbilical Vein Endothelial Cells; Humans; Integrins; Intracellular Signaling Peptides and Proteins; Leukemia, Lymphocytic, Chronic, B-Cell; Lymph Nodes; Lysophospholipids; Morpholines; Oxazines; Piperidines; Primary Cell Culture; Protein Kinase Inhibitors; Protein-Tyrosine Kinases; Purines; Pyrazoles; Pyridines; Pyrimidines; Quinazolinones; Receptors, Antigen, B-Cell; Receptors, Lysosphingolipid; Signal Transduction; Sphingosine; Sphingosine-1-Phosphate Receptors; Syk Kinase

Chronic lymphocytic leukemia (CLL) is characterized by the progressive accumulation of clonal B lymphocytes. Proliferation occurs in lymphoid tissues upon interaction of leukemic cells with a supportive microenvironment. Therefore, the mobilization of tissue-resident CLL cells into the circulation is a useful therapeutic strategy to minimize the reservoir of tumor cells within survival niches. Because the exit of normal lymphocytes from lymphoid tissues depends on the presence of sphingosine-1 phosphate (S1P) and the regulated expression of S1P receptor-1 (S1PR1), we investigated whether the expression and function of S1PR1 can be modulated by key microenvironment signals. We found that activation of CLL cells with CXCL12, fibroblast CD40L(+), BCR cross-linking, or autologous nurse-like cells reduces their S1PR1 expression and the migratory response toward S1P. Moreover, we found that S1PR1 expression was reduced in the proliferative/activated subset of leukemic cells compared with the quiescent subset from the same patient. Similarly, bone marrow-resident CLL cells expressing high levels of the activation marker CD38 showed a lower expression of S1PR1 compared with CD38(low) counterparts. Finally, given that treatment with BCR-associated kinase inhibitors induces a transient redistribution of leukemic cells from lymphoid tissues to circulation, we studied the effect of the Syk inhibitors piceatannol and R406 on S1PR1 expression and function. We found that they enhance S1PR1 expression in CLL cells and their migratory response toward S1P. Based on our results, we suggest that the regulated expression of S1PR1 might modulate the egress of the leukemic clone from lymphoid tissues.

Topics: ADP-ribosyl Cyclase 1; Adult; Aged; Aged, 80 and over; Animals; B-Lymphocytes; CD40 Ligand; Cell Movement; Chemokine CXCL12; Female; Gene Expression; Gene Expression Regulation, Neoplastic; Humans; Intracellular Signaling Peptides and Proteins; Leukemia, Lymphocytic, Chronic, B-Cell; Lysophospholipids; Male; Membrane Glycoproteins; Mice; Middle Aged; Oxazines; Protein-Tyrosine Kinases; Proto-Oncogene Proteins c-bcr; Pyridines; Receptors, CXCR4; Receptors, Lysosphingolipid; Sphingosine; Sphingosine-1-Phosphate Receptors; Stilbenes; Syk Kinase; Tumor Cells, Cultured; Tumor Microenvironment

FTY720 is an immunosuppressant developed to prevent organ transplant rejection. Recent studies indicate an additional role for FTY720 in inducing cell apoptosis. We demonstrate here that FTY720 mediates toxic effects in cell lines representing different B-cell malignancies and primary B cells from patients with chronic lymphocytic leukemia (CLL). In contrast to previous reports in T-cell lines, FTY720-induced toxicity in the Raji cell line and primary CLL B cells is independent of activation of caspases or poly(ADP-ribose) polymerase processing. Further, pancaspase inhibitor Z-VAD-fmk failed to rescue these cells from apoptosis mediated by FTY720. FTY720 induced down-regulation of Mcl-1 but not Bcl-2 in CLL B cells. Overexpression of Bcl-2 failed to protect transformed B cells from FTY720-induced apoptosis, suggesting a Bcl-2-independent mechanism. Interestingly, FTY720 induced protein phosphatase 2a (PP2a) activation and downstream dephosphorylation of ERK1/2, whereas okadaic acid at concentrations that inhibited the FTY720-induced PP2a activation also resulted in inhibition of FTY720-mediated apoptosis and restoration of baseline ERK1/2 phosphorylation in primary CLL cells, indicating a role for PP2a activation in FTY720-induced cytotoxicity. Further, FTY720 treatment resulted in significant prolonged survival in a xenograft severe combined immunodeficiency (SCID) mouse model of disseminated B-cell lymphoma/leukemia. These results provide the first evidence for the potential use of FTY720 as a therapeutic agent in a variety of B-cell malignancies, including CLL.

Topics: Animals; Apoptosis; B-Lymphocytes; Caspases; Cell Line; Cysteine Endopeptidases; Down-Regulation; Fingolimod Hydrochloride; Humans; Immunosuppressive Agents; Leukemia, Lymphocytic, Chronic, B-Cell; Lysophospholipids; Mice; Mice, SCID; Myeloid Cell Leukemia Sequence 1 Protein; Neoplasm Proteins; Precursor Cell Lymphoblastic Leukemia-Lymphoma; Propylene Glycols; Protein Phosphatase 2; Proto-Oncogene Proteins c-bcl-2; Signal Transduction; Sphingosine; Xenograft Model Antitumor Assays