Enhancement of in vitro and in vivo tumor cell radiosensitivity by the DNA methylation inhibitor zebularine.
Aberrant DNA hypermethylation is a frequent finding in tumor cells, which has suggested that inhibition of DNA methylation may be an effective cancer treatment strategy. Because DNA methylation affects gene expression and chromatin structure, parameters considered to influence radioresponse, we investigated the effects of the DNA methylation inhibitor zebularine on the radiosensitivity of human tumor cells. Three human tumor cell lines were used in this study (MiaPaCa, DU145, and U251) and the methylation status of three genes frequently hypermethylated in tumor cells (RASSF1A, HIC-1, and 14-3-3sigma) was determined as a function of zebularine exposure. Zebularine resulted in DNA demethylation in a time-dependent manner, with the maximum loss of methylation detected by 48 hours. Treatment of cells with zebularine for 48 hours also resulted in an increase in radiosensitivity with dose enhancement factors of >1.5. As a measure of radiation-induced DNA damage, gammaH2AX expression was determined. Whereas zebularine had no effect on radiation-induced gammaH2AX foci at 1 hour, the number of gammaH2AX foci per cell was significantly greater in the zebularine-treated cells at 24 hours after irradiation, suggesting the presence of unrepaired DNA damage. Zebularine administration to mice reactivated gene expression in U251 xenografts; irradiation of U251 tumors in mice treated with zebularine resulted in an increase in radiation-induced tumor growth delay. These results indicate that zebularine can enhance tumor cell radiosensitivity in vitro and in vivo and suggest that this effect may involve an inhibition of DNA repair.
Dote
Hideaki
H
Molecular Radiation Therapeutics Branch, National Cancer Institute, Bethesda, Maryland, USA.
Cerna
David
D
Burgan
William E
WE
Carter
Donna J
DJ
Cerra
Michael A
MA
Hollingshead
Melinda G
MG
Camphausen
Kevin
K
Tofilon
Philip J
PJ
D048948:14-3-3 Proteins; D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000528:radiation effects; D014408:Biomarkers, Tumor / Q000235:genetics; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects / Q000528:radiation effects; D003131:Combined Modality Therapy; D003562:Cytidine / Q000031:analogs & derivatives* / Q000494:pharmacology*; D019175:DNA Methylation / Q000187:drug effects*; D004268:DNA-Binding Proteins / Q000235:genetics; D004307:Dose-Response Relationship, Radiation; D005092:Exonucleases / Q000235:genetics; D005095:Exoribonucleases; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D051741:Kruppel-Like Transcription Factors; D008297:Male; D051379:Mice; D008819:Mice, Nude; D009363:Neoplasm Proteins / Q000235:genetics; D009374:Neoplasms, Experimental / Q000235:genetics / Q000473:pathology / Q000628:therapy*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013997:Time Factors; D014157:Transcription Factors / Q000235:genetics; D025521:Tumor Suppressor Proteins / Q000235:genetics; D023041:Xenograft Model Antitumor Assays / Q000379:methods
D048948:14-3-3 Proteins; D014408:Biomarkers, Tumor; D004268:DNA-Binding Proteins; C499808:HIC1 protein, human; D051741:Kruppel-Like Transcription Factors; D009363:Neoplasm Proteins; C411762:RASSF1 protein, human; D012333:RNA, Messenger; D014157:Transcription Factors; D025521:Tumor Suppressor Proteins; D003562:Cytidine; C009131:pyrimidin-2-one beta-ribofuranoside; D005092:Exonucleases; D005095:Exoribonucleases; C413442:SFN protein, human