Targeting ceramide synthase 6–dependent metastasis-prone phenotype in lung cancer cells
Motoshi Suzuki, … , Mamoru Kyogashima, Takashi Takahashi
Motoshi Suzuki, … , Mamoru Kyogashima, Takashi Takahashi
Published January 4, 2016; First published December 7, 2015
Citation Information: J Clin Invest. 2016;126(1):254-265. https://doi.org/10.1172/JCI79775.
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Categories: Research Article Oncology

Targeting ceramide synthase 6–dependent metastasis-prone phenotype in lung cancer cells

Abstract

Sphingolipids make up a family of molecules associated with an array of biological functions, including cell death and migration. Sphingolipids are often altered in cancer, though how these alterations lead to tumor formation and progression is largely unknown. Here, we analyzed non–small-cell lung cancer (NSCLC) specimens and cell lines and determined that ceramide synthase 6 (CERS6) is markedly overexpressed compared with controls. Elevated CERS6 expression was due in part to reduction of microRNA-101 (miR-101) and was associated with increased invasion and poor prognosis. CERS6 knockdown in NSCLC cells altered the ceramide profile, resulting in decreased cell migration and invasion in vitro, and decreased the frequency of RAC1-positive lamellipodia formation while CERS6 overexpression promoted it. In murine models, CERS6 knockdown in transplanted NSCLC cells attenuated lung metastasis. Furthermore, combined treatment with l-α-dimyristoylphosphatidylcholine liposome and the glucosylceramide synthase inhibitor D-PDMP induced cell death in association with ceramide accumulation and promoted cancer cell apoptosis and tumor regression in murine models. Together, these results indicate that CERS6-dependent ceramide synthesis and maintenance of ceramide in the cellular membrane are essential for lamellipodia formation and metastasis. Moreover, these results suggest that targeting this homeostasis has potential as a therapeutic strategy for CERS6-overexpressing NSCLC.

Authors

Motoshi Suzuki, Ke Cao, Seiichi Kato, Yuji Komizu, Naoki Mizutani, Kouji Tanaka, Chinatsu Arima, Mei Chee Tai, Kiyoshi Yanagisawa, Norie Togawa, Takahiro Shiraishi, Noriyasu Usami, Tetsuo Taniguchi, Takayuki Fukui, Kohei Yokoi, Keiko Wakahara, Yoshinori Hasegawa, Yukiko Mizutani, Yasuyuki Igarashi, Jin-ichi Inokuchi, Soichiro Iwaki, Satoshi Fujii, Akira Satou, Yoko Matsumoto, Ryuichi Ueoka, Keiko Tamiya-Koizumi, Takashi Murate, Mitsuhiro Nakamura, Mamoru Kyogashima, Takashi Takahashi

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Figure 6

DMPC liposome induces CERS6-dependent apoptosis.

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DMPC liposome induces CERS6-dependent apoptosis. (A) MTT assays were per...
(A) MTT assays were performed for determining the viability of a panel of cell lines after treatment with 100 μM DMPC liposome (mean ± SD; n = 4–8). In the presence of 100 μM DMPC liposome, cell viability was plotted against cell doubling time. (B) Effects of 100 nm myriocin, 50 μm FB1, or control vehicle on LNM35 cells treated with 100 μM DMPC liposome (mean ± SD). (C) Quantification of ceramides in LNM35 cells treated with 200 μM DMPC liposome for 8 hours (mean ± SD; n = 3). The experiments were replicated, with similar results obtained. (D) Effects of CERS6 silencing by siCERS6-1 in LNM35 cells and of CERS6 overexpression in BEAS-2B cells (mean ± SD; n = 3). Cells were treated with 100 μM DMPC liposome. The experiments were replicated, with similar results obtained. (E) Effects of siCERS6-1 and/or 200 μM DMPC liposome on ceramide composition in LNM35 cells (mean ± SD; n = 3). The experiments were replicated, with similar results obtained. (F) LNM35 cells were incubated with 200 μM NBDPC containing liposome for 60 minutes and 4 μM ER-Tracker Red Dye for 30 minutes. Scale bar: 10 μm. The experiments were replicated, with similar results obtained. (G) Illustration of hypothetical pathway between deuterated DMPC and C16:0 ceramide. The relative amounts of deuterated or nondeuterated acyl and sphingoid chains in C16:0 ceramide were quantitated in the presence or absence of deuterated DMPC using LC/MS/MS analysis (mean ± SD).