diethoxyphosphoryloxymethyl-butanoate and Breast-Neoplasms

The histone deacetylase inhibitor (HDACI) butyroyloxymethyl diethylphosphate (AN-7) has been shown to synergize doxorubicin (Dox) anticancer activity while attenuating its cardiotoxicity. In this study we further explored the selectivity of AN-7's action in several cancer and normal cells treated with anticancer agents. The cells studied were murine mammary 4T1, human breast T47D and glioblastoma U251 cancer cell lines, neonatal rat cardiomyocytes, cardiofibroblasts and astrocytes, and immortalized cardiomyocyte H9C2 cells. Cell death, ROS production and changes in protein expression were measured and in vivo effects were evaluated in Balb-c mice. AN-7 synergized Dox and anti-HER2 cytotoxicity against mammary carcinoma cells with combination indices of 0.74 and 0.79, respectively, while it protected cardiomyocytes against their toxicity. Additionally AN-7 protected astrocytes from Dox-cytoxicity. Cell-type specific changes in the expression of proteins controlling survival, angiogenesis and inflammation by AN-7 or AN-7+Dox were observed. In mice, the protective effect of AN-7 against Dox cardiotoxicity was associated with a reduction in inflammatory factors. In summary, AN-7 augmented the anticancer activity of Dox and anti-HER2 and attenuated their toxicity against normal cells. AN-7 modulation of c-Myc, thrombospondin-1, lo-FGF-2 and other proteins were cell type specific. The effects of AN-7, Dox and their combination were preserved in vivo indicating the potential benefit of combining AN-7 and Dox for clinical use.

Topics: Angiogenic Proteins; Animals; Antibodies; Antineoplastic Combined Chemotherapy Protocols; Astrocytes; Brain Neoplasms; Breast Neoplasms; Butyrates; Cell Line, Tumor; Cell Survival; Cytoprotection; Dose-Response Relationship, Drug; Doxorubicin; Drug Synergism; Female; Fibroblasts; Glioblastoma; Histone Deacetylase Inhibitors; Humans; Inflammation Mediators; Inhibitory Concentration 50; Mice; Mice, Inbred BALB C; Myocytes, Cardiac; Organophosphorus Compounds; Rats; Reactive Oxygen Species; Receptor, ErbB-2; Time Factors

To investigate the anticancer activity and mode of action of butyroyloxymethyl-diethyl phosphate (AN-7), a prodrug of butyric acid and formaldehyde, as a single agent and in combination with doxorubicin in human carcinoma MCF-7 and the multidrug resistant MCF-7 Dx cell lines.. The anti-cancer activity of AN-7 as a single agent or in combination with doxorubicin was measured by the Hoechst cell viability and colony forming assays as well as by FACS analyses of cells stained with propidium iodide and annexin V-FITC. Modulations of protein expression and acetylation were measured by Western blot analyses. The number of doxorubicin-DNA adducts formed was evaluated using (14)C-labeled doxorubicin.. The AN-7 and homologous prodrugs exhibited similar growth inhibition effects against drug resistant and sensitive cells, and elicited their anticancer effect partially by inhibition of HDAC. The AN-7 transiently augmented histone acetylation and increase of p21 expression. Synergy between AN-7 and doxorubicin was demonstrated in the sensitive and the resistant cell lines by viability and colony formation assays and was further confirmed by FACS analysis showing an increase in cell mortality. The number of doxorubicin-DNA adducts in total genomic DNA isolated from cells treated with (14)C-labeled doxorubicin and AN-7 increased substantially compared to treatment with doxorubicin only. Treatment with AN-7 or doxorubicin increased p53 acetylation that was further potentiated by their combination.. The AN-7 combined with doxorubicin overcame drug resistance; at least in part by the intracellularly releasable formaldehyde that augmented formation of doxorubicin-DNA adducts and butyric acid that induced histone and p53 acetylation. Since the use of doxorubicin is limited by toxicity, the combination could offer an effective treatment modality with lower toxicity for breast cancer.

Topics: Acetylation; Antineoplastic Combined Chemotherapy Protocols; Breast Neoplasms; Butyrates; Cell Line, Tumor; Cell Survival; Cyclin-Dependent Kinase Inhibitor p21; Doxorubicin; Drug Resistance, Neoplasm; Drug Synergism; Female; Humans; Organophosphorus Compounds; Prodrugs; Tumor Suppressor Protein p53