title: Zebularine partially reverses GST methylation in prostate cancer cells and restores sensitivity to the DNA minor groove binder brostallicin.
abstract: Brostallicin is a DNA minor groove binder that shows enhanced antitumor activity in cells with high glutathione S-transferase (GST)/glutathione content. Prostate cancer cells present, almost invariably, methylation of the GSTP1 gene promoter and, as a consequence, low levels of GST-pi expression and activity. In these cells, brostallicin shows very little activity. We tested whether pretreatment of heavily GST-methylated prostate cancer cells with demethylating agents could enhance the activity of brostallicin. Human prostate cancer cells LNCaP and DU145 were used for these studies both in vitro and in vivo. The demethylating agent zebularine was used in combination with brostallicin. Methylation specific PCR and pyrosequencing were used to determine the level of GST methylation. Pretreatment with demethylating agents enhanced the in vitro activity of brostallicin in LNCaP cells. Zebularine, in particular, induced an enhancement of activity in vivo comparable to that obtained by transfecting the human GSTP1 gene in LNCaP cells in vitro. Molecular analysis performed on tumor xenografts in mice pretreated with zebularine failed to detect re-expression of GST-pi and demethylation of GSTP1. However, we found demethylation in the GSTM1 gene, with consequent re-expression of GST-mu at the mRNA level. These results indicate that zebularine, both in vitro and in vivo, enhances the activity of brostallicin and that this enhancement correlates with re-expression of GST-pi and GST-mu. These findings highlight the potential therapeutic value of combining demethylating agents and brostallicin in tumors with GST methylation that poorly respond to brostallicin.
authors.affiliation: Laboratory of Molecular Pharmacology; Department of Oncology; IRCCS-Istituto di Ricerche Farmacologiche Mario Negri; Milan, Italy.
authors.forename: Maria Antonietta
authors.initials: MA
authors.lastname: Sabatino
authors.forename: Cristina
authors.initials: C
authors.lastname: Geroni
authors.forename: Monica
authors.initials: M
authors.lastname: Ganzinelli
authors.forename: Roberta
authors.initials: R
authors.lastname: Ceruti
authors.forename: Massimo
authors.initials: M
authors.lastname: Broggini
mesh_terms: D000818:Animals; D000971:Antineoplastic Combined Chemotherapy Protocols / Q000627:therapeutic use*; D045744:Cell Line, Tumor; D003562:Cytidine / Q000031:analogs & derivatives* / Q000627:therapeutic use; D019175:DNA Methylation; D015972:Gene Expression Regulation, Neoplastic; D005982:Glutathione Transferase / Q000235:genetics* / Q000378:metabolism; D006146:Guanidines / Q000627:therapeutic use*; D006801:Humans; D008297:Male; D051379:Mice; D011471:Prostatic Neoplasms / Q000188:drug therapy* / Q000235:genetics* / Q000473:pathology; D011758:Pyrroles / Q000627:therapeutic use*; D011897:Random Allocation; D023041:Xenograft Model Antitumor Assays
chemical_list: D006146:Guanidines; D011758:Pyrroles; D003562:Cytidine; C009131:pyrimidin-2-one beta-ribofuranoside; D005982:Glutathione Transferase; C455279:brostallicin
keywords: DNA methylation; DNA reactive agents; combination chemotherapy; prostate cancer; reversal of drug resistance