Inhibition of histone deacetylases sensitizes glioblastoma cells to lomustine

TY - JOUR

T1 - Inhibition of histone deacetylases sensitizes glioblastoma cells to lomustine

AU - Staberg, Mikkel

AU - Michaelsen, Signe Regner

AU - Rasmussen, Rikke Darling

AU - Villingshøj, Mette

AU - Poulsen, Hans Skovgaard

AU - Hamerlik, Petra

PY - 2017/2/1

Y1 - 2017/2/1

N2 - PURPOSE: Glioblastoma (GBM) ranks among the deadliest solid cancers worldwide and its prognosis has remained dismal, despite the use of aggressive chemo-irradiation treatment regimens. Limited drug delivery into the brain parenchyma and frequent resistance to currently available therapies are problems that call for a prompt development of novel therapeutic strategies. While only displaying modest efficacies as mono-therapy in pre-clinical settings, histone deacetylase inhibitors (HDACi) have shown promising sensitizing effects to a number of cytotoxic agents. Here, we sought to investigate the sensitizing effect of the HDACi trichostatin A (TSA) to the alkylating agent lomustine (CCNU), which is used in the clinic for the treatment of GBM.METHODS: Twelve primary GBM cell cultures grown as neurospheres were used in this study, as well as one established GBM-derived cell line (U87 MG). Histone deacetylase (HDAC) expression levels were determined using quantitative real-time PCR and Western blotting. The efficacy of either CCNU alone or its combination with TSA was assessed using various assays, i.e., cell viability assays (MTT), cell cycle assays (flow cytometry, FACS), double-strand DNA break (DSB) quantification assays (microscopy/immunofluorescence) and expression profiling assays of proteins involved in apoptosis and cell stress (Western blotting and protein array).RESULTS: We found that the HDAC1, 3 and 6 expression levels were significantly increased in GBM samples compared to non-neoplastic brain control samples. Additionally, we found that pre-treatment of GBM cells with TSA resulted in an enhancement of their sensitivity to CCNU, possibly via the accumulation of DSBs, decreased cell proliferation and viability rates, and an increased apoptotic rate.CONCLUSION: From our data we conclude that the combined administration of TSA and CCNU eradicates GBM cells with a higher efficacy than either drug alone, thereby opening a novel avenue for the treatment of GBM.

AB - PURPOSE: Glioblastoma (GBM) ranks among the deadliest solid cancers worldwide and its prognosis has remained dismal, despite the use of aggressive chemo-irradiation treatment regimens. Limited drug delivery into the brain parenchyma and frequent resistance to currently available therapies are problems that call for a prompt development of novel therapeutic strategies. While only displaying modest efficacies as mono-therapy in pre-clinical settings, histone deacetylase inhibitors (HDACi) have shown promising sensitizing effects to a number of cytotoxic agents. Here, we sought to investigate the sensitizing effect of the HDACi trichostatin A (TSA) to the alkylating agent lomustine (CCNU), which is used in the clinic for the treatment of GBM.METHODS: Twelve primary GBM cell cultures grown as neurospheres were used in this study, as well as one established GBM-derived cell line (U87 MG). Histone deacetylase (HDAC) expression levels were determined using quantitative real-time PCR and Western blotting. The efficacy of either CCNU alone or its combination with TSA was assessed using various assays, i.e., cell viability assays (MTT), cell cycle assays (flow cytometry, FACS), double-strand DNA break (DSB) quantification assays (microscopy/immunofluorescence) and expression profiling assays of proteins involved in apoptosis and cell stress (Western blotting and protein array).RESULTS: We found that the HDAC1, 3 and 6 expression levels were significantly increased in GBM samples compared to non-neoplastic brain control samples. Additionally, we found that pre-treatment of GBM cells with TSA resulted in an enhancement of their sensitivity to CCNU, possibly via the accumulation of DSBs, decreased cell proliferation and viability rates, and an increased apoptotic rate.CONCLUSION: From our data we conclude that the combined administration of TSA and CCNU eradicates GBM cells with a higher efficacy than either drug alone, thereby opening a novel avenue for the treatment of GBM.

KW - Antineoplastic Agents, Alkylating/administration & dosage

KW - Antineoplastic Combined Chemotherapy Protocols/pharmacology

KW - Blotting, Western

KW - Brain Neoplasms/enzymology

KW - Cell Cycle/drug effects

KW - Cell Line, Tumor

KW - Cell Survival/drug effects

KW - Flow Cytometry

KW - Fluorescent Antibody Technique

KW - Glioblastoma/enzymology

KW - Histone Deacetylase Inhibitors/administration & dosage

KW - Humans

KW - Hydroxamic Acids/administration & dosage

KW - Lomustine/administration & dosage

KW - Real-Time Polymerase Chain Reaction

U2 - 10.1007/s13402-016-0301-9

DO - 10.1007/s13402-016-0301-9

M3 - Journal article

C2 - 27766591

SN - 2211-3428

VL - 40

SP - 21

EP - 32

JO - Cellular oncology (Dordrecht)

JF - Cellular oncology (Dordrecht)

IS - 1

ER -