Acid ceramidase and its inhibitors: a de novo drug target and a new class of drugs for killing glioblastoma cancer stem cells with high efficiency

Abstract

Glioblastoma remains the most common, malignant primary cancer of the central nervous system with a low life expectancy and an overall survival of less than 1.5 years. The treatment options are limited and there is no cure. Moreover, almost all patients develop recurrent tumors, which typically are more aggressive. Therapeutically resistant glioblastoma or glioblastoma stem-like cells (GSCs) are hypothesized to cause this inevitable recurrence. Identifying prognostic biomarkers of glioblastoma will potentially advance knowledge about glioblastoma tumorigenesis and enable discovery of more effective therapies. Proteomic analysis of more than 600 glioblastoma-specific proteins revealed, for the first time, that expression of acid ceramidase (ASAH1) is associated with poor glioblastoma survival. CD133+ GSCs express significantly higher ASAH1 compared to CD133- GSCs and serum-cultured glioblastoma cell lines, such as U87MG. These findings implicate ASAH1 as a plausible independent prognostic marker, providing a target for a therapy tailored toward GSCs. We further demonstrate that ASAH1 inhibition increases cellular ceramide level and induces apoptosis. Strikingly, U87MG cells, and three different patient-derived glioblastoma stem-like cancer cell lines were efficiently killed, through apoptosis, by three different known ASAH1 inhibitors with IC50's ranging from 11–104 μM. In comparison, the standard glioblastoma chemotherapy agent, temozolomide, had minimal GSC-targeted effects at comparable or even higher concentrations (IC50 > 750 μM against GSCs). ASAH1 is identified as a de novo glioblastoma drug target, and ASAH1 inhibitors, such as carmofur, are shown to be highly effective and to specifically target glioblastoma GSCs. Carmofur is an ASAH1 inhibitor that crosses the blood-brain barrier, a major bottleneck in glioblastoma treatment. It has been approved in Japan since 1981 for colorectal cancer therapy. Therefore, it is poised for repurposing and translation to glioblastoma clinical trials.

Role of Ceramidases in Sphingolipid Metabolism and Human Diseases

“…Doan et al identified Carmofur as an ASAH1 inhibitor that can cross the blood-brain barrier. It is highly effective and targets glioblastoma cancer stem cells [116]. Carmofur is an approved drug against colorectal cancer in Japan [117].…”

Sphingosine-1-Phosphate in the Tumor Microenvironment: A Signaling Hub Regulating Cancer Hallmarks

“…In support, it was reported that administration of the functional S1P antagonist FTY720 to nude mice led to downregulation of S1P receptors, induced apoptosis in GSCs, and was synergistic with TMZ in promoting cytotoxicity [232]. Of relevance, the apoptosis induced by inhibiting SphKs was shown to be highly effective, and to specifically target GSCs [220], known as resistant cells to the standard GBM chemotherapy agent TMZ.…”

“…In particular, it was found that SphK1 inhibition: (1) leads to cell death by inducing apoptosis of human GBM cells and xenografts, and reduces survival in orthotopic GBM [72,177]; (2) sensitizes GBM cells, and different cancer cells, to several cytotoxic drugs [216]; (3) is effective in potentiating the cytotoxicity of both TMZ and radiation therapy in various human GBM cell lines [217,218]; and (4) induces apoptosis and inhibits colony formation in TMZ-resistant GBM cells [219]. Of relevance, recent studies on GBM irradiated cells revealed that S1P is upregulated, and SphK1 gene significantly induced following radiation, suggesting that S1P links to radio-resistance and increased aggressiveness of irradiated GBM cells [69,220].…”

Targeting the Sphingolipid System as a Therapeutic Direction for Glioblastoma

“…Subsequent inhibition of acid ceramidase using N-oleoylethanolamine, was found to suppress this increase in ASAH1 expression, resulting in increased ceramide accumulation [149]. Elevated ASAH1 levels have been reported in the tumors of patients with newly diagnosed GBM, as well as radiotherapy-treated GBM, correlating with poor survival [165,166]. This elevation of ASAH1 in response to radiation has also been reported in both adult (U87) and pediatric (SJGBM-2) GBM cell lines, as well as patient tissue samples [165,166].…”

“…Elevated ASAH1 levels have been reported in the tumors of patients with newly diagnosed GBM, as well as radiotherapy-treated GBM, correlating with poor survival [165,166]. This elevation of ASAH1 in response to radiation has also been reported in both adult (U87) and pediatric (SJGBM-2) GBM cell lines, as well as patient tissue samples [165,166]. The mechanism by which this resistance occurs is thought to be through a reduction in ceramide accumulation, and an upregulation of S1P, resulting in increased cell viability [166].…”