sphingosine-1-phosphate and Head-and-Neck-Neoplasms

It has been documented previously that defects in the generation of C(18)-ceramide, a product of ceramide synthase 1 (CerS1), also known as longevity assurance gene 1 (hLASS1), play important roles in the pathogenesis and/or progression of HNSCC. However, whether altered levels of ceramide generation in HNSCC tumors have any clinical relevance remains unknown. In this study, the levels of endogenous ceramides were measured in tumor tissues of 45 HNSCC patients as compared to their normal tissues using high-pressure liquid chromatography/mass spectrometry (LC/MS), and then possible link between ceramide levels and the clinical parameters of HNSCC were examined. The data showed that the levels of C(16)-, C(24)-, C(24:1)-ceramides were significantly elevated in the majority of tumor tissues compared to their normal tissues, while the levels of only C(18)-ceramide were significantly decreased in HNSCC tumors, especially in tumor tissues of male patients. Importantly, it was also shown here that decreased C(18)-ceramide levels in HNSCC tumor tissues were significantly associated with the higher incidences of lymphovascular invasion, and pathologic nodal metastasis. Importantly, attenuation of C(18)-ceramide was also positively linked to the higher overall stages of the primary HNSCC tumors. Therefore, these data suggest, for the first time, that the defects in the generation/accumulation of C(18)-ceramide might have important clinical roles in HNSCC, especially in lymphovascular invasion and nodal disease.

Topics: Biomarkers, Tumor; Carcinoma, Squamous Cell; Case-Control Studies; Ceramides; Chromatography, Liquid; Female; Head and Neck Neoplasms; Humans; Larynx; Lymphatic Metastasis; Lysophospholipids; Male; Mass Spectrometry; Neoplasm Invasiveness; Neoplasm Recurrence, Local; Pharynx; Sphingosine

The purpose of this study was to determine the profile of bioactive sphingolipids in xenograft mouse tissues of head and neck squamous cell carcinoma. We utilized UHPLC-MS/MS to determine the profile of full set of ceramides, sphingosine, and sphingosine 1-phosphate in this xenograft mouse model. The tissues isolated and investigated were from brain, lung, heart, liver, spleen, kidney, bladder, tumors and blood. With the exception of equal volume of blood plasma (100ul), all tissues were studied with the same amount of protein (800ug). Results demonstrated that brain contained the highest level of ceramide and kidney had the highest level of sphingosine, whereas sphingosine 1-phosphate and dihydrosphingosine 1-phosphate were heavily presented in the blood. Brain also comprised the highest level of phospholipids. As for the species, several ceramides, usually present in very low amounts in cultured tumor cells, showed relatively high levels in certain tissues. This study highlights levels of bioactive sphingolipids profiles in xenograft mouse model of head and neck squamous cell carcinoma, and provides resources to investigate potential therapeutic targets and biomarkers that target bioactive sphingolipids metabolism pathways.

Topics: Animals; Carcinoma, Squamous Cell; Cell Line, Tumor; Ceramides; Chromatography, High Pressure Liquid; Disease Models, Animal; Head and Neck Neoplasms; Humans; Lysophospholipids; Mice, Nude; Sphingolipids; Sphingosine; Tandem Mass Spectrometry; Transplantation, Heterologous

Molecules that appear on the surface of tumor cells after their therapy treatment may have important roles either as damage-associated molecular patterns (DAMPs) or signals for phagocytes influencing the disposal of these cells. Treatment of SCCVII and CAL27 cells, models of mouse and human squamous cell carcinoma respectively, by photodynamic therapy (PDT) resulted in the presentation of ceramide and sphingosine-1-phosphate (S1P) on the cell surface. This was documented by anti-ceramide and anti-S1P antibody staining followed by flow cytometry. The exposure of these key sphingolipid molecules on PDT-treated tumor cells was PDT dose-dependent and it varied in intensity with different photosensitizers used for PDT. The above results, together with the finding that both ceramide and S1P can activate NFκB signaling in macrophages co-incubated with PDT-treated tumor cells, establish that these two sphingolipids can act as DAMPs stimulating inflammatory/immune reactions critical for tumor therapy response.

Topics: Animals; Carcinoma, Squamous Cell; Cell Line, Tumor; Cell Membrane; Cell Separation; Ceramides; Disease Models, Animal; Flow Cytometry; Head and Neck Neoplasms; Lysophospholipids; Mice; Mice, Inbred C3H; Photochemotherapy; Receptors, Pattern Recognition; Sphingosine

Eradication of tumours by photodynamic therapy (PDT) is accompanied by marked changes in local sphingolipid (SL) engagement. Because of the heterogeneity of cellular composition, analysis of tumour tissue homogenates to quantify SL species is inadequate for evaluating their levels in parenchymal cancer cell population. By staining tumour-derived single cell suspensions with antibodies specific to ceramide and sphingosine 1-phosphate (S1P) followed by flow cytometry, we were able to document changes in the levels of these two key SLs in cancer cells and tumour-associated macrophages (TAMs) of mouse SCCVII tumours following PDT. The results confirm previously obtained indications that tumour treatment by PDT induces a marked rise in ceramide levels in cancer cells within these lesions. Cancer cells from PDT-treated SCCVII tumours undergoing apoptosis were found to have much higher ceramide levels and substantially lower S1P levels than their viable counterparts. Compared to cancer cells, considerably higher ceramide and S1P levels were consistently found in TAMs. Treatment of SCCVII tumour-bearing mice with ceramide analog LCL29 induced a rise in ceramide levels in TAMs but not in cancer cells. When combined with PDT, LCL29 treatment produced a further increase in ceramide levels in TAMs while having no evident impact on ceramide content in cancer cells within same tumours. The results highlight SLs as important participants in tumour response to PDT and potential adjuvant therapeutic targets to PDT.

Topics: Animals; Apoptosis; Carcinoma, Squamous Cell; Ceramides; Head and Neck Neoplasms; Indoles; Lysophospholipids; Macrophages; Mesoporphyrins; Mice; Mice, Inbred C3H; Organosilicon Compounds; Photochemotherapy; Photosensitizing Agents; Pyridinium Compounds; Sphingosine

Two anticancer agents, LCL85 and photodynamic therapy (PDT) were combined to test whether the combination PDT/LCL85 evokes changes in the sphingolipid (SL) profile and promotes cell death. Treatment of SCCVII mouse squamous carcinoma cells using the silicone phthalocyanine Pc 4 for PDT induced increases in the prodeath global ceramides/dihydroceramides (DHceramides), and no changes in the prosurvival sphingosine-1-phosphate (S1P). In contrast, after LCL85, the levels of most ceramides and DHceramides were reduced, whereas the levels of S1P were increased. After PDT/LCL85 the levels of global ceramides and DHceramides, and of S1P, were restored to resting levels. PDT/LCL85 also enhanced the levels of C18-, C20-, and C20:1-ceramide, and C18-DHceramide. Treatment with PDT, with or without LCL85, led to substantial reductions in sphingosine levels. PDT/LCL85 induced enhanced autophagy and caspase-3 activation. None of the treatments affected short-term viability of cells. In contrast, long-term clonogenic survival was reduced not only after PDT or LCL85, but even more after PDT/LCL85. Overall, our data show that short-term exposure to PDT/LCL85 led to distinct signature effects on the SL profile, enhanced autophagy, and caspase-3 activation without cell death. Long-term exposure to PDT/LCL85 enhanced overall cell killing, supporting translational potential of PDT/LCL85.

Topics: Animals; Antineoplastic Agents; Autophagy; Carcinoma, Squamous Cell; Caspase 3; Cell Line, Tumor; Ceramides; Head and Neck Neoplasms; Humans; Indoles; Lysophospholipids; Mice; Photochemotherapy; Photosensitizing Agents; Propanolamines; Pyridinium Compounds; Sphingolipids; Sphingosine