sphingosine-kinase and Carcinoma

Hypoxia facilitates an aggressive phenotype and immune evasion in solid tumors including bladder cancer (BC). Sphingosine kinase 1 (SphK1) is aberrantly expressed and correlated with poor prognosis in BC patients. However, its roles in hypoxia-evoked malignancies and immune evasion in BC remain elusive.. The expression of SphK1 in BC tissues was analysed using a bioinformatics database. BC cells were transfected with si-SphK1 or recombinant HIF-1α plasmids under hypoxic conditions. The mRNA level, activity and protein expression of SphK1 were determined. Transwell assay was performed to evaluate cell invasion. After co-culture with natural killer (NK) cells, NK cell cytotoxicity to BC cells was assessed. The involvement of sphingosine-1-phosphate (S1P)/HIF-1α signaling was analysed by ELISA, qRT-PCR and western blot.. UALCAN and GEPIA database confirmed high expression of SphK1 in BC tissues. Moreover, hypoxia increased the expression and activity of SphK1. Loss of SphK1 inhibited hypoxia-induced cell invasion. IL-2 induced NK cell activation by secreting TNF-α and IFN-γ. Hypoxia antagonized NK cell activation-evoked cytotoxicity to BC cells. Intriguingly, SphK1 knockdown reversed hypoxia-induced cell resistance to NK cell killing. Mechanically, SphK1 loss inhibited hypoxia-activated the S1P/HIF-1α signaling. However, S1P addition reversed the inhibitory effects of SphK1 down-regulation on hypoxia-activated S1P/HIF-1α signaling. Notably, reactivating HIF-1α overturned the suppressive roles of SphK1 loss in decreasing hypoxia-induced cell invasion and resistance to NK cell cytotoxicity.. Targeting SphK1 may inhibit hypoxia-evoked invasion and immune evasion via the S1P/HIF-1α signaling, indicating a promising therapeutic target for BC.

Topics: Carcinoma; Cell Death; Humans; Hypoxia; Interleukin-2; Killer Cells, Natural; Oncogenes; Phosphotransferases (Alcohol Group Acceptor); RNA, Messenger; Tumor Necrosis Factor-alpha; Urinary Bladder; Urinary Bladder Neoplasms

Sphingolipid metabolites are emerging as key signaling molecules in cancer. Sphingosine kinase 1 is up-regulated in many different types of human malignancies and plays a crucial role in cancer development and progression. The utility of sphingosine kinase 1 to act as a predictive biomarker in thyroid cancer remains unclear.. Sphingosine kinase 1 expression was evaluated using immunohistochemical staining in 110 formalin-fixed, paraffin-embedded papillary thyroid carcinoma tissue samples.. Sphingosine kinase 1 expression in papillary thyroid carcinoma tissue was significantly higher than in nodular goiter (p<0.001) or normal thyroid (p<0.001) tissue. Sphingosine kinase 1 was observed in the cytoplasm of tumor cells. Thirty-four (30.9%) of 110 papillary thyroid carcinomas exhibited high sphingosine kinase 1 expression, that was significantly associated with tumor multiplicity (p=0.004), extrathyroidal extension (p=0.013), presence of lymph node metastasis (p<0.001), and number of metastatic lymph nodes (p=0.042). In addition, high sphingosine kinase 1 expression was the only independent predictor of lymph node metastasis (p<0.001).. Sphingosine kinase 1 is involved in papillary thyroid carcinoma development and progression and can serve as a potential biomarker predictive of lymph node metastasis.

Topics: Adult; Aged; Biomarkers, Tumor; Carcinoma; Carcinoma, Papillary; Disease Progression; Female; Humans; Immunohistochemistry; Lymphatic Metastasis; Male; Middle Aged; Phosphotransferases (Alcohol Group Acceptor); Predictive Value of Tests; Prognosis; Thyroid Cancer, Papillary; Thyroid Neoplasms; Tissue Array Analysis; Up-Regulation; Young Adult

The oncogenic roles of sphingosine kinase 1 (SphK1) in various cancers, including thyroid cancer, have been well demonstrated. However, the microRNAs (miRNAs) associated with the oncogenic roles of SphK1 remain largely unknown.. Global gene and miRNA expression in TPC1-Vector and TPC1-SphK1 cells was analyzed using digital gene expression (DGE) analysis and small RNA-seq, respectively. miRNA-mRNA interactions were explored by microT-CDS, and the predicted networks were visualized using CytoScape. In this study, we found that overexpression of SphK1 differentially regulates the expression of 46 miRNAs and 506 mRNAs in papillary thyroid cancer (PTC) TPC1 cells. Combining bioinformatics predictions of mRNA targets with DGE data on mRNA expression allowed us to identify the mRNA targets of deregulated miRNAs. The direct interaction between miR-144-3p and FN1, which mediates the pro-invasive role of SphK1 in PTC cells, was experimentally validated.. Our results demonstrated that SphK1 overexpression drives a regulatory network governing miRNA and mRNA expression in PTC cells. We also demonstrated the roles played by miR-144-3p and FN1 in mediating the oncogenic function of SphK1, which enhanced the understanding of the etiology of PTC.

Topics: Carcinoma; Carcinoma, Papillary; Gene Expression Profiling; Humans; MicroRNAs; Oncogenes; Phosphotransferases (Alcohol Group Acceptor); Thyroid Cancer, Papillary; Thyroid Neoplasms

Sphingosine kinase 1 (SPHK1) has been shown to be involved in the progression of various types of human cancers. We previously demonstrated that SPHK1 is overexpressed and associated with clinical stage, locoregional recurrence, distant metastasis and poor prognosis in nasopharyngeal carcinoma (NPC). However, the biological roles involving SPHK1 and its potential usefulness as a therapeutic target in NPC remain unknown. In this study, Blocking SPHK1 using siRNA or FTY720 (a SPHK1 inhibitor) significantly reduced proliferation and migration and increased cell cycle arrest and apoptosis in NPC cells. FTY720 also decreased SPHK1 activity, and overexpressing SPHK1 abrogated the FTY720-induced effects on cell viability. In addition, FTY720 sensitized NPC cells to radiotherapy by inhibiting SPHK1 activity in vitro and in vivo. Furthermore, high SPHK1 expression was associated with increased Ki-67 and p-Akt and decreased caspase-3 expression in human NPC specimens. These data suggest that SPHK1 might be a potential therapeutic target for NPC.

Topics: Animals; Antineoplastic Agents; Apoptosis; Carcinoma; Caspase 3; Cell Cycle Checkpoints; Cell Line, Tumor; Cell Movement; Cell Proliferation; Chemoradiotherapy; Dose-Response Relationship, Drug; Dose-Response Relationship, Radiation; Female; Fingolimod Hydrochloride; Humans; Ki-67 Antigen; Mice, Inbred BALB C; Mice, Nude; Molecular Targeted Therapy; Nasopharyngeal Carcinoma; Nasopharyngeal Neoplasms; Phosphorylation; Phosphotransferases (Alcohol Group Acceptor); Protein Kinase Inhibitors; Proto-Oncogene Proteins c-akt; RNA Interference; Signal Transduction; Time Factors; Transfection; Xenograft Model Antitumor Assays

We previously reported that sphingosine kinase 1 (SphK1), an enzyme that catalyzes the production of sphingosine-1-phosphate (SIP), is upregulated in human gastric cancer and predicts poor clinical outcome. In the present study, we used known differential effects of UV irradiation on human MGC-803 gastric cancer cells to determine their effect on SphK1 activity. Ectopic expression of SphK1 in MGC-803 gastric cancer cells markedly enhanced their resistance to UV irradiation, whereas silencing endogenous SphK1 with shRNAs weakened this ability. Furthermore, these anti-apoptotic effects were significantly associated with decrease of Bim, an apoptosis-related protein. We further demonstrated that SphK1 could downregulate the transcriptional activity of forkhead box O3a (FoxO3a) by inducing its phosphorylation, which was found to be associated with the PI3K/Akt signaling. Taken together, our study supports the theory that SphK1 confers resistance to apoptosis in gastric cancer cells via the Akt/FoxO3a/Bim pathway.

Topics: Apoptosis; Apoptosis Regulatory Proteins; Bcl-2-Like Protein 11; Carcinoma; Cell Line, Tumor; Down-Regulation; Forkhead Box Protein O3; Forkhead Transcription Factors; Gene Expression Regulation, Neoplastic; Gene Silencing; Humans; Lysophospholipids; Membrane Proteins; Phosphatidylinositol 3-Kinases; Phosphorylation; Phosphotransferases (Alcohol Group Acceptor); Proto-Oncogene Proteins; Proto-Oncogene Proteins c-akt; Reverse Transcriptase Polymerase Chain Reaction; RNA, Small Interfering; Signal Transduction; Sphingosine; Stomach Neoplasms; Ultraviolet Rays; Up-Regulation

To investigate the expression of sphingosine kinase 1 (SphK1) and NF-κB in colon carcinoma tissues and their correlation with clinicopathologic features.. Sixty-six paraffin-embedded colon carcinoma samples and 66 fresh colon carcinoma samples were tested using immunohistochemistry, RT-PCR and Western blot, respectively.. In 66 fresh colon carcinoma samples, the positive rate of SphK1 and NF-κB mRNA expression were 84.85%(56/66) and 74.24% (49/66), while the positive rate of SphK1 and NF-κB protein detected by Western blot were 78.79% (52/66) and 69.70% (46/66). The positive rates were higher than those in the adjacent tissues [mRNA: 63.64% (42/66), 48.49% (32/66); protein: 57.58% (38/66), 45.45% (30/66)] and the normal mucosa [mRNA: 42.42% (28/66), 25.76% (17/66); protein: 36.36% (24/66), 24.24% (16/66)], with statistical significances (all P values < 0.05). The mean expressive levels of SphK1 and NF-κB mRNA and protein in colon carcinoma were both significantly higher than those in the adjacent tissues and the normal mucosa (mRNA: 0.55 ± 0.06 vs 0.35 ± 0.05 vs 0.25 ± 0.05, 0.75 ± 0.06 vs 0.43 ± 0.05 vs 0.30 ± 0.04; protein: 0.77 ± 0.05 vs 0.38 ± 0.06 vs 0.12 ± 0.03, 0.45 ± 0.08 vs 0.23 ± 0.05 vs 0.13 ± 0.03; all P values < 0.05). There was a close correlation between SphK1 and NF-κB expression levels (r = 0.459, P = 0.036). The results of immunohistochemistry were similar to those of RT-PCR and Western blot. Overexpression of SphK1 and NF-κB in colon carcinoma was related with depth of invasion, distant and lymph node metastasis and Dukes' stages (all P values < 0.05). The expression of SphK1 was also related with differentiation (P < 0.05).. Overexpression of SphK1 and NF-κB may be involved in the pathogenesis and progression of colon carcinoma. Moreover, SphK1 and NF-κB may be correlated with the invasion and metastasis of colon carcinoma.

Topics: Adult; Aged; Blotting, Western; Carcinoma; Colonic Neoplasms; Female; Humans; Immunohistochemistry; Lymphatic Metastasis; Male; Middle Aged; Phosphotransferases (Alcohol Group Acceptor); Reverse Transcriptase Polymerase Chain Reaction; Transcription Factor RelA

Sphingosine kinase 1 (SphK1) is an important regulator of apoptosis, survival, and proliferation in cancer cells. SphK1 expression in head and neck squamous cell cancer (HNSCC) cell lines and tumor tissue was assessed, and the efficacy of SphK1 knockdown in increasing tumor radiosensitivity was evaluated in vitro and in vivo.. Expression of SphK1 was determined by immunohistochemistry, Western blot, and real-time polymerase chain reaction (RT-PCR) in 34 prospectively collected HNSCC tumor samples. HNSCC cell lines squamous cell carcinoma (SCC)-15 and SCC-25 were treated with SphK1 inhibitor SKI-II and siRNA targeting SphK1 with and without radiation, and the cell viability was assessed. SCC-15 cells with and without transfection of SphK1 siRNA were then injected into athymic nude mice to develop tumor xenografts, and these 2 groups were further divided into 1 group that received radiation and 1 group that did not. Tumor size was measured over 18 days, when the animals were killed and the tumors were evaluated by immunohistochemistry.. SphK1 is found in both HNSCC cell lines and human tumor samples, with higher expression correlated with advanced tumor stage, nodal involvement, and recurrence. In vitro, both SCC-15 and SCC-25 were found to be radioresistant; however, they were sensitized by administration of SKI-II and transfection with siRNA targeting SphK1. In vivo, SphK1-siRNA transfected xenografts were decreased in size compared with both nonradiated control and radiated control mice, whereas mice with both SphK1-siRNA and radiation treatment showed a synergistic reduction in tumor volume. Histopathologic analysis demonstrated a decreased proliferative state in SphK1-siRNA transfected tumors.. SphK1 is upregulated in HNSCC, and inhibition of SphK1 sensitizes HNSCC to radiation-induced cytotoxicity.

Topics: Animals; Apoptosis; Blotting, Western; Carcinoma; Carcinoma, Squamous Cell; Disease Models, Animal; Enzyme Inhibitors; Head and Neck Neoplasms; Humans; Immunohistochemistry; In Vitro Techniques; Male; Mice; Mice, Inbred BALB C; Mice, Nude; Neoplasms, Squamous Cell; Phosphotransferases (Alcohol Group Acceptor); Polymerase Chain Reaction; Radiation Tolerance; Squamous Cell Carcinoma of Head and Neck; Thiazoles; Treatment Outcome

Sphingosine 1-phosphate (S1P) induces migration of the human thyroid follicular carcinoma cell line ML-1 by activation of S1P(1) and S1P(3) receptors, G(i) proteins, and the phosphatidylinositol 3-kinase-Akt pathway. Because sphingosine kinase isoform 1 (SK) recently has been implicated as an oncogene in various cancer cell systems, we investigated the functions of SK in the migration, proliferation and adhesion of the ML-1 cell line. SK overexpressing ML-1 cells show an enhanced secretion of S1P, which can be attenuated, by inhibiting SK activity and a multidrug-resistant transport protein (ATP-binding cassette transporter). Furthermore, overexpression of SK enhances serum-induced migration of ML-1 cells, which can be attenuated by blocking ATP-binding cassette transporter and SK, suggesting that the migration is mediated by autocrine signaling through secretion of S1P. Inhibition of protein kinase C alpha, with both small interfering RNA (siRNA) and small molecular inhibitors attenuates migration in SK overexpressing cells. In addition, SK-overexpressing cells show an impaired adhesion, slower cell growth, and an up-regulation of ERK1/2 phosphorylation, as compared with cells expressing a dominant-negative SK. Taken together, we present evidence suggesting that SK enhances migration of ML-1 cells by an autocrine mechanism and that the S1P-evoked migration is dependent on protein kinase C alpha, ERK1/2, and SK.

Topics: Autocrine Communication; Carcinoma; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cells, Cultured; Humans; Lysophospholipids; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Neoplasm Invasiveness; Oncogenes; Phosphorylation; Phosphotransferases (Alcohol Group Acceptor); Protein Kinase C-alpha; RNA, Small Interfering; Sphingosine; Thyroid Neoplasms; Transfection

Sphingosine 1-phosphate (S1P) the product of sphingosine kinase (SK) action plays an important role in various pathological conditions like inflammation and cancer. In this study, we show that in the human breast cancer cell line MCF7, epidermal growth factor (EGF) stimulates SK-1 activity in a biphasic manner with a first peak after 15 min and a second delayed activation occurring after 1 h up to 18 h and thereafter declining again. This delayed activation is accompanied by increased mRNA and protein expression of SK-1, but not SK-2. Mechanistically, the transcriptional upregulation is dependent on the classical mitogen-activated protein kinase, protein kinase C (PKC) and the phosphoinositide 3-kinase, since specific inhibitors of these enzymes all abolish the EGF-induced mRNA upregulation and activity of SK-1. Moreover, dexamethasone also suppressed EGF-induced SK-1 mRNA expression and activity which is reversed by the glucocorticoid receptor antagonist RU486. To see whether EGF-induced upregulation of SK-1 is of relevance for tumor progression, we investigated two hallmarks of carcinogenesis, i.e., cell proliferation and migration. Stimulation of cells with EGF leads to enhanced [(3)H]thymidine incorporation into DNA and also to stimulated migration in a modified Boyden chamber assay. When cells are depleted of SK-1, but not SK-2, by siRNA transfection or by dexamethasone treatment, EGF-induced proliferation and migration are drastically reduced. In summary, these data show that EGF causes an acute stimulation of SK-1 activity and, moreover, triggers a delayed SK activation which is due to increased gene transcription and de novo synthesis of SK-1, which in turn directs cells towards growth and increased motility. Thus, the sphingosine kinase-1 may represent a novel attractive target for cancer therapy.

Topics: Breast Neoplasms; Carcinoma; Cell Line, Tumor; Cell Movement; Cell Proliferation; Enzyme Inhibitors; Epidermal Growth Factor; Female; Humans; Mitogen-Activated Protein Kinases; Phosphatidylinositol 3-Kinases; Phosphoinositide-3 Kinase Inhibitors; Phosphotransferases (Alcohol Group Acceptor); Protein Kinase C; ras Proteins

Vascular endothelial growth factor (VEGF) signaling is critical to the processes of angiogenesis and tumor growth. Here, evidence is presented for VEGF stimulation of sphingosine kinase (SPK) that affects not only endothelial cell signaling but also tumor cells expressing VEGF receptors. VEGF or phorbol 12-myristate 13-acetate treatment of the T24 bladder tumor cell line resulted in a time- and dose-dependent stimulation of SPK activity. In T24 cells, VEGF treatment reduced cellular sphingosine levels while raising that of sphingosine-1-phosphate. VEGF stimulation of T24 cells caused a slow and sustained accumulation of Ras-GTP and phosphorylated extracellular signal-regulated kinase (phospho-ERK) compared with that after EGF treatment. Small interfering RNA (siRNA) that targets SPK1, but not SPK2, blocks VEGF-induced accumulation of Ras-GTP and phospho-ERK in T24 cells. In contrast to EGF stimulation, VEGF stimulation of ERK1/2 phosphorylation was unaffected by dominant-negative Ras-N17. Raf kinase inhibition blocked both VEGF- and EGF-stimulated accumulation of phospho-ERK1/2. Inhibition of SPK by pharmacological inhibitors, a dominant-negative SPK mutant, or siRNA that targets SPK blocked VEGF, but not EGF, induction of phospho-ERK1/2. We conclude that VEGF induces DNA synthesis in a pathway which sequentially involves protein kinase C (PKC), SPK, Ras, Raf, and ERK1/2. These data highlight a novel mechanism by which SPK mediates signaling from PKC to Ras in a manner independent of Ras-guanine nucleotide exchange factor.

Topics: Carcinoma; Endothelial Growth Factors; Endothelium, Vascular; Enzyme Activation; Enzyme Inhibitors; Humans; Intercellular Signaling Peptides and Proteins; Lymphokines; MAP Kinase Signaling System; Mitogen-Activated Protein Kinases; Phosphotransferases (Alcohol Group Acceptor); Protein Kinase C; ras GTPase-Activating Proteins; ras Guanine Nucleotide Exchange Factors; Tetradecanoylphorbol Acetate; Tumor Cells, Cultured; Urinary Bladder Neoplasms; Vascular Endothelial Growth Factor A; Vascular Endothelial Growth Factors