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.
Highlights
Glioblastoma remains the most common, malignant primary cancer of the central nervous system despite many decades of research [1]
Our results showed that CD133+ glioblastoma stem-like cells (GSCs) express a very high level of ASAH1 compared to CD133- GSCs and non-stem cancer cells, such as U87MG cells
The correlation between protein levels and survival was evaluated by graphing mass spectral count, which correlates with protein level, against survival
Summary
Glioblastoma remains the most common, malignant primary cancer of the central nervous system despite many decades of research [1]. Recent findings have suggested that glioblastoma stem-like cells (GSCs) may play a significant role in the resistance of cancer to chemotherapy and radiotherapy [22, 23]. The lack of effective treatment for glioblastoma, together with the recent findings regarding the role of GSCs, has generated intense interest in developing new biomarkers and GSC-targeted therapies to reduce tumor recurrence and improve patient survival. Mass-spectrometry (MS)based proteomics analysis is emerging as a viable, high throughput method for discovering disease biomarkers by simultaneous, efficient quantitative analysis of many targets Recent optimization of this method by us for analyzing protein markers in glioblastoma has been developed using banked human glioblastoma specimens associated with clinical parameters and outcome data from our institutional Brain and Spinal Cord Tissue Bank [28]. Due to its high level of expression in GSCs, ASAH1 inhibition is proposed as a new antiglioblastoma therapy that targets GSCs
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