ABSTRACT
Glioblastoma remains the most malignant of all primary adult brain tumours with poor patient survival and limited treatment options. This study adopts a drug repurposing approach by investigating the anti-cancer activity of a derivative of the antipsychotic drug phenothiazine (DS00329) in malignant U251 and U87 glioblastoma cells. Results from MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) and clonogenic assays showed that DS00329 inhibited short-term glioblastoma cell viability and long-term survival while sparing non-cancerous cells. Western blot analysis with an antibody to γH2AX showed that DS00329 induced DNA damage and flow cytometry and western blotting confirmed that it triggered a G1 cell cycle arrest which correlated with decreased levels in Cyclin A, Cyclin B, Cyclin D1 and cyclin dependent kinase 2 and an increase in levels of the cyclin dependent kinase inhibitor p21. DS00329 treated glioblastoma cells exhibited morphological and molecular markers typical of apoptotic cells such as membrane blebbing and cell shrinkage and an increase in levels of cleaved PARP. Flow cytometry with annexin V-FITC/propidium iodide staining confirmed that DS00329 induced apoptotic cell death in glioblastoma cells. We also show that DS00329 treatment of glioblastoma cells led to an increase in the autophagosome marker LC3-II and autophagy inhibition studies using bafilomycin A1 and wortmannin, showed that DS00329-induced-autophagy was a pro-death mechanism. Furthermore, DS00329 treatment of glioblastoma cells inhibited the phosphatidylinositol 3'-kinase/Akt cell survival pathway. Our findings suggest that DS00329 may be an effective treatment for glioblastoma and provide a rationale for further exploration and validation of the use of phenothiazines and their derivatives in the treatment of glioblastoma.
Subject(s)
Antineoplastic Agents/pharmacology , Glioblastoma/pathology , Phenothiazines/pharmacology , Antineoplastic Agents/therapeutic use , Apoptosis/drug effects , Autophagy/drug effects , Cell Cycle/drug effects , Cell Line, Tumor , Cell Proliferation/drug effects , Cell Survival/drug effects , DNA Damage , Drug Repositioning , Glioblastoma/drug therapy , Humans , Phenothiazines/therapeutic use , Phosphatidylinositol 3-Kinases/metabolism , Proto-Oncogene Proteins c-akt/metabolism , Signal Transduction/drug effectsABSTRACT
Halofantrine (HF) is a poorly water-soluble antimalarial drug with low bioavailability. Complex formation of HF.HCl and 2-hydroxypropyl-beta-cyclodextrin (HP-beta-CD) in aqueous solution and in solid state as well as the possibility of improving the solubility and dissolution rate of the drug though complexation with the cyclodextrin were investigated. Phase-solubility profile indicated that the solubility of the drug was significantly increased in the presence of HP-beta-CD and was classified as AL-type, indicating 1:1 stoichiometric inclusion complexes and an apparent stability constant value of 2300 M(-1). Solid inclusion complexes of HF, HCl and the cyclodextrin at 1:1 molar ratios were prepared by physical mixture, kneading, co-evaporation and freeze-drying methods and characterized by X-ray diffraction and Infra-red spectroscopy. The solubility and dissolution rates of HF.HCl from the complexes were determined and found to be dependent on the preparation method of the complexes. Dissolution profile of the drug was markedly enhanced by complex formation with the cyclodextrin and the product prepared by the freeze-drying method exhibited the most superior dissolution properties compared to the other methods used in this study. The results suggest that the complexation of HF.HCl with HP-beta-CD could improve therapeutic efficacy of the drug though enhanced absorption expected from increased drug dissolution.