Anticancer research · 2025
MGMT Promoter Methylation in Glioblastoma Stem Cells: Stability During Differentiation and Comparison With Surgically-resected Tumors.
СтатьяСравнительное исследованиеНазвание и аннотация: Перевод пока не подготовлен
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Yoshioka Masaki, Noguchi Shunsei, Iwadate Yasuo, Kobayashi Masayoshi, Motohashi Shinichiro, Higuchi Yoshinori
Аннотация
BACKGROUND/AIM: The prognosis and resistance to temozolomide in glioblastoma have been evaluated based on O6-methylguanine-DNA methyltransferase (MGMT) promoter methylation using bulk tumor samples resected via surgery. However, it remains unclear whether MGMT promoter methylation differs between glioblastoma stem cells and the overall tumor within the patient's brain. Chemoresistance of glioblastoma stem cells is a critical factor in understanding the refractory nature of the disease. This study aimed to assess MGMT promoter methylation in glioblastoma stem cells and compare it with that in differentiated tumor cells.
MATERIALS AND METHODS: Glioblastoma stem-like cells (GSCs) and differentiated GSCs (dGSCs) were cultured from surgically resected tumors. Following DNA extraction and bisulfite conversion, real-time methylation-specific PCR was performed to quantify MGMT promoter methylation in three tumor cell types: GSCs, dGSCs, and surgically resected tumor samples. MGMT protein and CD133 expression levels were assessed via flow cytometry.
RESULTS: Based on CD133 expression and microscopic evaluation, dGSCs exhibited a more differentiated phenotype than GSCs. No significant differences were observed in ΔCt values or MGMT promoter methylation ratios between GSCs and dGSCs. Similarly, ΔCt values and methylation ratios showed negligible differences between GSCs and surgically resected tumors. MGMT was highly expressed in both GSCs and dGSCs, consistent with an unmethylated MGMT promoter.
CONCLUSION: MGMT promoter methylation status in glioblastoma stem cells remained unchanged during differentiation and was comparable to that in surgically resected tumors. As temozolomide is presumed to be ineffective against glioblastoma stem cells with unmethylated MGMT promoters, MGMT blockade may be a beneficial therapeutic strategy.
MATERIALS AND METHODS: Glioblastoma stem-like cells (GSCs) and differentiated GSCs (dGSCs) were cultured from surgically resected tumors. Following DNA extraction and bisulfite conversion, real-time methylation-specific PCR was performed to quantify MGMT promoter methylation in three tumor cell types: GSCs, dGSCs, and surgically resected tumor samples. MGMT protein and CD133 expression levels were assessed via flow cytometry.
RESULTS: Based on CD133 expression and microscopic evaluation, dGSCs exhibited a more differentiated phenotype than GSCs. No significant differences were observed in ΔCt values or MGMT promoter methylation ratios between GSCs and dGSCs. Similarly, ΔCt values and methylation ratios showed negligible differences between GSCs and surgically resected tumors. MGMT was highly expressed in both GSCs and dGSCs, consistent with an unmethylated MGMT promoter.
CONCLUSION: MGMT promoter methylation status in glioblastoma stem cells remained unchanged during differentiation and was comparable to that in surgically resected tumors. As temozolomide is presumed to be ineffective against glioblastoma stem cells with unmethylated MGMT promoters, MGMT blockade may be a beneficial therapeutic strategy.