Sphingosine kinase 1 mediates head & neck squamous cell carcinoma invasion through sphingosine 1-phosphate receptor 1

Cancer Cell International, Aug 2014

Background Head and neck squamous cell carcinoma (HNSCC) is characterized by aggressive loco-regional invasion. Sphingosine kinase1 (SphK1), an enzyme in sphingolipid metabolism, is emerging as a key player in HNSCC pathology. The observation that SphK1 is overexpressed in all HNSCC stages and is associated with depth of tumor invasion, metastasis and clinical failure underscores the importance of SphK1 in HNSCC pathology. Still, the mechanisms underlying SphK1 regulation of invasion have not been delineated. Therefore, we sought to mechanistically describe how SphK1 regulates invasion in HNSCC. Methods Invasion assays were used to measure invasive ability of SphK1 overexpressing human tongue squamous cell carcinoma (SCC-25 cells). Western blotting, quantitative qPCR, ELISA and zymography were used to measure the effect of SphK1 and sphingosine 1-phoshate receptor 1 (S1P1) on invasion measures, MMP-2/9, E-cadherin, EGFR, IL-6/STAT3, in SCC-25 cells. Results SphK1 expression is elevated in cells with an invasive phenotype as compared to non-invasive phenotype. We show SphK1 overexpression increased EGF-induced EGFR/ERK and AKT activity, increased matrix metalloproteinase (MMP)-2/9 mRNA and reduced E-cadherin. SphK1 overexpression also increased IL-6 concentration and EGF-induced STAT3 phosphorylation, exemplifying that SphK1 modulates IL-6/STAT3 signaling. Notably, we show that S1P1 knockdown reduced IL-6/STAT3 signaling, representing another pathway by which SphK1/S1P regulates invasion. Conclusions Taken together, our data suggest that SphK1 sits at the hub of multiple key signaling cascades, all which have been implicated in the regulation of invasiveness, making SphK1 an attractive target for the development of HNSCC therapies.

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Sphingosine kinase 1 mediates head & neck squamous cell carcinoma invasion through sphingosine 1-phosphate receptor 1

Cancer Cell International Sphingosine kinase 1 mediates head & neck squamous cell carcinoma invasion through sphingosine 1-phosphate receptor 1 Paulette M Tamashiro 1 Hideki Furuya 1 Yoshiko Shimizu 0 1 Toshihiko Kawamori 0 1 0 Department of Molecular Biosciences and Bioengineering, University of Hawaii at Manoa , Honolulu, HI 96818 , USA 1 Cancer Biology Program, University of Hawaii Cancer Center , 701 Ilalo Street, Honolulu, HI 96813 , USA Background: Head and neck squamous cell carcinoma (HNSCC) is characterized by aggressive loco-regional invasion. Sphingosine kinase1 (SphK1), an enzyme in sphingolipid metabolism, is emerging as a key player in HNSCC pathology. The observation that SphK1 is overexpressed in all HNSCC stages and is associated with depth of tumor invasion, metastasis and clinical failure underscores the importance of SphK1 in HNSCC pathology. Still, the mechanisms underlying SphK1 regulation of invasion have not been delineated. Therefore, we sought to mechanistically describe how SphK1 regulates invasion in HNSCC. Methods: Invasion assays were used to measure invasive ability of SphK1 overexpressing human tongue squamous cell carcinoma (SCC-25 cells). Western blotting, quantitative qPCR, ELISA and zymography were used to measure the effect of SphK1 and sphingosine 1-phoshate receptor 1 (S1P1) on invasion measures, MMP-2/9, E-cadherin, EGFR, IL-6/STAT3, in SCC-25 cells. Results: SphK1 expression is elevated in cells with an invasive phenotype as compared to non-invasive phenotype. We show SphK1 overexpression increased EGF-induced EGFR/ERK and AKT activity, increased matrix metalloproteinase (MMP)-2/9 mRNA and reduced E-cadherin. SphK1 overexpression also increased IL-6 concentration and EGF-induced STAT3 phosphorylation, exemplifying that SphK1 modulates IL-6/STAT3 signaling. Notably, we show that S1P1 knockdown reduced IL-6/STAT3 signaling, representing another pathway by which SphK1/S1P regulates invasion. Conclusions: Taken together, our data suggest that SphK1 sits at the hub of multiple key signaling cascades, all which have been implicated in the regulation of invasiveness, making SphK1 an attractive target for the development of HNSCC therapies. Sphingosine kinase1; Invasion; HNSCC; EGFR; STAT3; Sphingosine 1-phosphate; Sphingosine 1-phosphate receptor 1; IL-6; SCC-25 cells - Background Recurrence rates for advanced-stage head and neck squamous cell carcinoma (HNSCC) is greater than 50% [1] and the 5-year survival rate for HNSCC has not drastically improved over the last 30 years [2]. The bleak survival rate is due to late presentation, the subsequent delay of detection of lesions, invasion into loco-regional lymph nodes, a high rate of metastasis [2] and the limited availability of effective therapies. Sphingolipids play a crucial role in cancer pathogenesis by modulating cell signal transduction pathways to influence biological outcomes such as cell senescence, differentiation, apoptosis, migration, and proliferation [3-5]. Sphingosine kinase-1 (SphK1) is an important enzyme in sphingolipid metabolism which regulates tumor growth in HNSCC. For example, SphK1 knockdown results in lower cell proliferation and smaller HNSCC tumors [6], and SphK1 inhibition increases radiation sensitivity [7]. In addition to its well-documented role in cell proliferation, SphK1 also regulates invasion. SphK1 overexpression is positively associated with invasion, invasive morphology and cell diameter in esophageal squamous carcinoma cells (ESCC) [8]. In addition, immunodeficient mice subcutaneously injected with ESCC cells overexpressing SphK1 exhibited 6-fold greater lung metastasis compared to parent cells [8]. In clinical HNSCC samples, human SphK1 expression was significantly higher compared to normal mucosa, and this was positively associated with depth of tumor invasion, metastasis, and clinical failure [9]. Furthermore, SphK1 negative staining was associated with a 6.5-year survival post-surgery, while SphK1 positive staining was associated with only a 2-year survival period post-surgery [9]. It is not known whether SphK1 is directly involved in activation of epidermal growth factor receptor (EGFR) in HNSCC. However, it is known that SphK1 expression correlates with genes downstream of the EGFR pathway (i.e., amphiregulin, integrin5, epiregulin) in ESCC as demonstrated with microarray analyses [8]. Also, ESCC cells overexpressing SphK1 had greater phosphorylation of EGF, while cells transfected with siRNA against SphK1 showed reduced EGFR phosphorylation [8]. Another study showed treatment with EGFR alone increased invasion (~4) in carcinoma of the epiglottis (PCI-37A) and inhibition of EGFR inhibited invasion by 47-fold [10]. In addition, EGFR was also shown to mediate invasion in conjunction with signal transducer and activator of transcription 3 (STAT3) in HNSCC [11]. Together these studies suggest that a relationship may exist between SphK1, EGFR and STAT3 to affect invasive abi (...truncated)


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Paulette M Tamashiro, Hideki Furuya, Yoshiko Shimizu, Toshihiko Kawamori. Sphingosine kinase 1 mediates head & neck squamous cell carcinoma invasion through sphingosine 1-phosphate receptor 1, Cancer Cell International, 2014, pp. 76, 14, DOI: 10.1186/s12935-014-0076-x