Characterization of phenothiazine-induced apoptosis in neuroblastoma and glioma cell lines: clinical relevance and possible application for brain-derived tumors.
In this study we aimed to (1). screen phenothiazines for cytotoxic activity in glioma, neuroblastoma, and primary mouse brain tissue; and (2). determine the mechanism of the cytotoxic effect (apoptosis, necrosis) and the roles of calmodulin inhibition and sigma receptor modulation. Rat glioma (C6) and human neuroblastoma (SHSY-5Y) cell lines were treated with different phenothiazines. All agents induced a dose-dependent decrease in viability and proliferation, with the highest activity elicited by thioridazine. Sensitivity to thioridazine of glioma and neuroblastoma cells was significantly higher (p < 0.05) than that of primary mouse brain culture (IC50 11.2 and 15.1 microM vs 41.3 microM, respectively). The N-mustard fluphenazine induced significantly lower cytotoxicity in glioma cells, compared to fluphenazine. The sigma receptor selective ligand (+)-SK&F10047 increased viability slightly while combined with fluphenazine; SK&F10047 did not alter fluphenazine activity. Flow cytometry of propidium iodide (PI)-stained glioma cells treated with thioridazine, fluphenazine, or perphenazine (6-50 microM) resulted in a concentration-dependent increase of fragmented DNA up to 94% vs 3% in controls by all agents. Thioridazine (12.5 microM)-treated glioma cells costained with PI and Hoechst 33342 revealed a red fluorescence of fragmented nuclei in treated cells and a blue fluorescence of intact control nuclei. After 4-h exposure to thioridazine (25 and 50 microM), a 25- to 30-fold increase in caspase-3 activity in neuroblastoma cells was noted. Overall, the marked apoptotic effect of phenothiazines in brain-derived cancer cells, and the low sensitivity of primary brain tissue suggest the potential use of selected agents as therapeutic modalities in brain cancer.
Laboratory of Biological Psychiatry, Felsenstein Medical Research Center, Campus Rabin, Petah-Tiqva 49100, Israel, and Sackler School of Medicine, Tel-Aviv University, Ramat Aviv, Israel. iritgil@post.tau.ac.il
Irit
I
Gil-Ad
Biana
B
Shtaif
Yechiel
Y
Levkovitz
Michal
M
Dayag
Ella
E
Zeldich
Abraham
A
Weizman
D000818:Animals; D000970:Antineoplastic Agents / Q000627:therapeutic use / Q000633:toxicity*; D017209:Apoptosis / Q000187:drug effects* / Q000502:physiology; D001932:Brain Neoplasms / Q000188:drug therapy*; D002147:Calmodulin / Q000187:drug effects / Q000378:metabolism; D053148:Caspase 3; D020169:Caspases / Q000187:drug effects / Q000378:metabolism; D002455:Cell Division / Q000187:drug effects / Q000502:physiology; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects / Q000502:physiology; D053938:DNA Fragmentation / Q000187:drug effects / Q000502:physiology; D004305:Dose-Response Relationship, Drug; D005476:Fluphenazine / Q000633:toxicity; D005910:Glioma / Q000188:drug therapy*; D006801:Humans; D051379:Mice; D008813:Mice, Inbred ICR; D009447:Neuroblastoma / Q000188:drug therapy*; D010620:Phenazocine / Q000031:analogs & derivatives* / Q000494:pharmacology; D010640:Phenothiazines / Q000627:therapeutic use / Q000633:toxicity*; D051381:Rats; D017480:Receptors, sigma / Q000187:drug effects / Q000378:metabolism; D013881:Thioridazine / Q000633:toxicity; D015854:Up-Regulation / Q000187:drug effects / Q000502:physiology
D000970:Antineoplastic Agents; D002147:Calmodulin; D010640:Phenothiazines; D017480:Receptors, sigma; C012134:SK&F 10047; C505407:CASP3 protein, human; C505409:Casp3 protein, mouse; C505410:Casp3 protein, rat; D053148:Caspase 3; D020169:Caspases; C031637:phenothiazine; D010620:Phenazocine; D013881:Thioridazine; D005476:Fluphenazine