sphingosine-1-phosphate and Leukemia

Since the discovery and initial characterizations of sphingolipids (SLs) in 1884, extensive research has established that these molecules not only are structural components of eukaryotic membranes but they are also critical bioactive lipids involved in fundamental cellular processes such as proliferation, differentiation, apoptosis, inflammation, migration, and autophagy. Altered SL metabolism has been observed in many pathological conditions including hematological malignancies. Thus, targeting the SL pathway to induce lipid changes to counteract specific pathologies is currently being pursued as a promising, novel therapeutic intervention. In this review, we discuss the general characteristics of the SL pathway, illustrating those features relevant to the understanding of the role of SLs in leukemia, and we address novel SL-targeting therapeutic approaches.

Topics: Animals; Antineoplastic Agents; Ceramides; Humans; Leukemia; Lysophospholipids; Signal Transduction; Sphingolipids; Sphingosine

Bioactive phospholipids, including sphingosine-1-phosphate (S1P), ceramide-1-phosphate (C1P), lysophosphatidylcholine (LPC), and its derivative lysophosphatidic acid (LPA), have emerged as important mediators regulating the trafficking of normal and cancer cells. While the role of S1P in regulating migration of hematopoietic cells is well established, in this work we compared its biological effects to the effects of C1P, LPC, and LPA. We employed 10 human myeloid and lymphoid cell lines as well as blasts from AML patients. We observed that human leukemic cells express functional receptors for phospholipids and respond to stimulation by phosphorylation of p42/44 MAPK and AKT. We also found that bioactive phospholipids enhanced cell migration and adhesion of leukemic cells by downregulating expression of HO-1 and iNOS in a p38 MAPK-dependent manner but did not affect cell proliferation. By contrast, downregulation of p38 MAPK by SB203580 enhanced expression of HO-1 and iNOS and decreased migration of leukemic cells in vitro and their seeding efficiency to vital organs in vivo after injection into immunodeficient mice. Based on these findings, we demonstrate that, besides S1P, human leukemic cells also respond to C1P, LPC, and LPA. Since the prometastatic effects of bioactive phospholipids in vivo were mediated, at least in part, by downregulating HO-1 and iNOS expression in a p38 MAPK-dependent manner, we propose that inhibitors of p38 MAPK or stimulators of HO-1 activity will find application in inhibiting the spread of leukemic cells in response to bioactive phospholipids.

Topics: Animals; Blast Crisis; Cell Adhesion; Cell Line, Tumor; Cell Movement; Cell Proliferation; Ceramides; Fibronectins; Hematopoietic Stem Cells; Heme Oxygenase-1; Humans; Leukemia; Lysophospholipids; Mice, SCID; Nitric Oxide Synthase Type II; p38 Mitogen-Activated Protein Kinases; Phospholipids; Receptors, Cell Surface; Sphingosine

Sphingosine 1-phosphate (S1P) is a bioactive lipid that has been identified as an accelerant of cancer progression. The sphingosine kinases (SphKs) are the sole producers of S1P, and thus, SphK inhibitors may prove effective in cancer mitigation and chemosensitization. Of the two SphKs, SphK1 overexpression has been observed in a myriad of cancer cell lines and tissues and has been recognized as the presumptive target over that of the poorly characterized SphK2. Herein, we present the design and synthesis of amidine-based nanomolar SphK1 subtype-selective inhibitors. A homology model of SphK1, trained with this library of amidine inhibitors, was then used to predict the activity of additional, more potent, inhibitors. Lastly, select amidine inhibitors were validated in human leukemia U937 cells, where they significantly reduced endogenous S1P levels at nanomolar concentrations.

Topics: Amidines; Antineoplastic Agents; Cell Line, Tumor; Chemistry, Pharmaceutical; Drug Design; Enzyme Inhibitors; Gene Expression Regulation, Leukemic; Humans; Kinetics; Leukemia; Lysophospholipids; Models, Chemical; Models, Molecular; Phosphotransferases (Alcohol Group Acceptor); Sphingosine; U937 Cells

We recently reported increased sphingosine kinase 1 (SPHK1) and decreased neutral sphingomyelinase 2 (NSMase2) gene expression in myelodysplastic syndromes and acute leukemia. This alteration is supposed to change the cellular sphingolipid metabolites; however, positive correlations were observed between daunorubicin (DA)-IC50 and the SPHK1 message but not between DA-IC50 and NSMase2 messages, when 16 different leukemia cell lines were used to analyze the relationship between gene expressions and chemosensitivity against DA. Using two cell lines with either the highest or lowest SPHK1 expression, cellular ceramides and sphingosine 1-phosphate (S1P) were quantified by liquid chromatography/mass spectrometry. Increased ceramide was observed in DA-sensitive, but not in DA-resistant cell lines treated with low doses of DA. Upon DA treatment, S1P decreased more in the sensitive cell lines than in resistant cell lines. A SPHK inhibitor recovered the DA sensitivity of DA-resistant cells. The modulation of SPHK1 gene expression by either overexpression or using siRNA affected the DA sensitivity of representative cell lines. Results clearly show that SPHK1 is both a good marker to predict the DA sensitivity of leukemia cells and a potential therapeutic target for leukemia with high SPHK1 expression, and suggest that the sphingolipid rheostat plays a significant role in DA-induced cytotoxicity.

Topics: Antibiotics, Antineoplastic; Biomarkers; Cell Line, Tumor; Daunorubicin; Drug Resistance, Neoplasm; Gene Expression Profiling; Humans; Leukemia; Lysophospholipids; Phosphotransferases (Alcohol Group Acceptor); Sphingosine

Sphingosine 1-phosphate (S-1P) has been implicated as a second messenger preventing apoptosis by counteracting activation of executioner caspases. Here it is reported that S-1P prevents apoptosis and executioner caspase-3 activation by inhibiting the translocation of cytochrome c and Smac/DIABLO from mitochondria to the cytosol induced by anti-Fas, tumor necrosis factor-alpha (TNF-alpha), serum deprivation, and cell-permeable ceramides in the human acute leukemia Jurkat, U937, and HL-60 cell lines. Furthermore, the tumor promoter 12-O-tetradecanoyl-phorbol-13-acetate, which stimulates sphingosine kinase, the enzyme responsible for S-1P production, also inhibits cytochrome c and Smac/DIABLO release. In contrast, dimethylsphingosine (DMS), a specific inhibitor of sphingosine kinase, sensitizes cells to cytochrome c and Smac/DIABLO release triggered by anti-Fas, TNF-alpha, serum deprivation, or ceramide. DMS-induced mitochondrial apoptogenic factor leakage can likewise be overcome by S-1P cotreatment. Hence, S-1P, likely generated through a protein kinase C- mediated activation of sphingosine kinase, inhibits the apoptotic cascade upstream of the release of the mitochondrial apoptogenic factors, cytochrome c, and Smac/DIABLO in human acute leukemia cells.

Topics: Apoptosis; Apoptosis Regulatory Proteins; Carrier Proteins; Ceramides; Culture Media, Serum-Free; Cytochrome c Group; HL-60 Cells; Humans; Intracellular Signaling Peptides and Proteins; Jurkat Cells; Leukemia; Lysophospholipids; Mitochondria; Mitochondrial Proteins; Protein Transport; Receptors, Tumor Necrosis Factor; Sphingosine; Tetradecanoylphorbol Acetate; U937 Cells

Phorbol esters exert a dual function in human leukemia cells, induction of differentiation and activation of integrin-mediated functions. Here we have shown that the plastic adherence of phorbol ester-treated U937 cells is mediated by expression of integrin Mac-1 (CD11b/CD18) on the cell surface and that these adherent cells exhibit anoikis (apoptosis when adherent cells are detached or adherence is inhibited). We used U937-derived clones overexpressing either antisense RNAs antisense to CD11b and CD18 mRNAs or mRNA from a truncated mutant CD11b gene. We have also shown that apoptosis in non-adherent cells or anoikis was mediated by sphingosine and that survival of adherent cells was achieved by a shift of the dynamic balance between sphingosine and sphingosine 1-phosphate toward the latter by adherence-activated sphingosine 1-kinase.

Topics: Apoptosis; CD18 Antigens; Cell Adhesion; Cell Survival; Humans; Integrins; Leukemia; Lysophospholipids; Macrophage-1 Antigen; Monocytes; Phorbol 12,13-Dibutyrate; Sphingosine; Tumor Cells, Cultured

Exogenous sphingosine 1-phosphate (S1P) induced Ca2+ mobilization, in association with an increase in inositol polyphosphate production reflecting activation of phospholipase C in HL60 leukemia cells. The increase in intracellular Ca2+ concentration ([Ca2+]i) induced by S1P was inhibited by an appropriate treatment of the cells with pertussis toxin (PTX), U73122 (a phospholipase C inhibitor) or phorbol 12-myristate 13-acetate (PMA). In parallel with the Ca2+ response, these agents also inhibited inositol polyphosphate production. The S1P-induced Ca2+ response was also attenuated in the dibutyryl cAMP-induced differentiated cells, where GTP-binding protein-induced Ca2+ response suggested to be enhanced. Lysophosphatidic acid (LPA) also increased [Ca2+]i in the cels, but the maximal response was about half of that of S1P, and furthermore PTX and dibutyryl cAMP treatment hardly affected the LPA-induced Ca2+ mobilization. We conclude that exogenous S1P mobilizes Ca2+ through phospholipase C activation. The S1P-induced enzyme activation is at least partly mediated by PTX-sensitive GTP-binding protein-coupled receptors which may be different from LPA receptors.

Topics: Calcium; Enzyme Activation; GTP-Binding Proteins; HL-60 Cells; Humans; Inositol Phosphates; Leukemia; Lysophospholipids; Pertussis Toxin; Sphingosine; Type C Phospholipases; Virulence Factors, Bordetella