A sphingosine kinase inhibitor combined with temozolomide induces glioblastoma cell death through accumulation of dihydrosphingosine and dihydroceramide, endoplasmic reticulum stress and autophagy

Abstract

Glioblastomas (GBMs) are very aggressive tumors with low chemosensitivity. The DNA-alkylating agent temozolomide (TMZ) is currently the most efficient chemotoxic drug for GBM therapy; however, many patients develop resistance to TMZ. Combining TMZ with another agent could present an improved treatment option if it could overcome TMZ resistance and avoid side effects. Sphingosine kinase inhibitors (SKIs) have emerged as anticancer agents. Sphingosine kinases are often overexpressed in tumors where their activity of phosphorylating sphingosine (Sph) contributes to tumor growth and migration. They control the levels of the pro-apoptotic ceramide (Cer) and Sph and of the pro-survival sphingosine-1 phosphate. In the present work, TMZ was combined with a specific SKI, and the cytotoxic effect of each drug alone or in combination was tested on GBM cell lines. The combination of sublethal doses of both agents resulted in the cell death potentiation of GBM cell lines without affecting astrocyte viability. It triggered a caspase-3-dependent cell death that was preceded by accumulation of dihydrosphingosine (dhSph) and dihydroceramide (dhCer), oxidative stress, endoplasmic reticulum stress, and autophagy. Autophagy was identified as the crucial switch that facilitated induction of this cell death potentiation. The sublethal dose of the inhibitor induced these stress events, whereas that of TMZ induced the destructive autophagy switch. Remarkably, neither Cer nor Sph, but rather the Cer intermediates, dhSph and dhCer, was involved in the cytotoxicity from the combination. Cell lines sensitive to the combination expressed low levels of the antioxidant enzyme glutathione peroxidase-1, indicating this enzyme as a potential marker of sensitivity to such treatment. This work shows for the first time a strong interaction between a SKI and TMZ, leading to a tumor cell-specific death induction. It further demonstrates the biological relevance of dihydrosphingolipids in cell death mechanisms and emphasizes the potential of drugs that affect sphingolipid metabolism for cancer therapy.

SKI-178: A multitargeted inhibitor of sphingosine kinase and microtubule dynamics demonstrating therapeutic efficacy in acute myeloid leukemia models

“…Indeed, SKIs, other inhibitors of Cer metabolism and Cer itself all synergistically enhance the apoptotic efficacy of MTAs. 39–49 More recently, it has been suggested that Cer may actually be required for paclitaxel induced mitochondrial outer membrane permeabilization (MOMP) and apoptosis. 50 Mechanistically, sphingolipids have also been shown to activate the pro-apoptotic Bcl-2 family proteins Bak and Bax.…”

“…Indeed, there are numerous examples in the literature of SKIs interacting synergistically with both targeted and cytotoxic chemotherapies. 39,68–70 A highly selective non-cytotoxic SphK inhibitor, such as PF-543, may be an ideal adjuvant to current chemotherapeutic regimens. In such an instance, inhibition of S1P production and/or accumulation of Cer could enhance induction of apoptosis of the standard-of-care regimens.…”

Evaluation of Consistency in Spheroid Invasion Assays

“…We compared the effects of applying a sphingosine kinase inhibitor (SKI), known to impair survival and invasion of the glioma cells18, on spheroids generated after 72 h (87 spheroids of 90) and 144 h (78 spheroids of 90) (Fig. 6).…”

Recent advances in the development of sphingosine kinase inhibitors

“…Nevertheless, SKI-II has been shown to decrease cellular S1P, increase cellular ceramides and sphingosine, and induce apoptosis [66,68]. Furthermore, it has been demonstrated to enhance the effects of other anti-cancer agents on cell lines, including sensitising a panel of glioblastoma lines to temozolomide [69], chemo-resistant MCF7-TN-R breast cancer cells to doxorubicin [70], and head and neck squamous cell carcinoma cell lines to the EGF receptor inhibitor, Cetuximab [71].…”