Genexpressionsanalyse in unterschiedlichen malignen Tumorzelllinien nach Apoptose-Induktion durch Taurolidin mittels cDNA-Microarray

Abstract

Background: Taurolidine (TRD) was initially used as an anti-infective agent. Recently, TRD has been shown to have apoptosis inducing properties, but the mechanism of action is largely unknown. The aim of this study was to examine the function of TRD in different malignant cell lines on gene expression level. Material and Methods: Five different malignant cell lines, HT29 (colon), Chang Liver (liver), HT1080 (fibrosarcoma), AsPC-1 and BxPC-3 (pancreas) were incubated with increasing concentrations of TRD (100, 250, und 1000 μM) for 6h and 24h. Proliferation rate as well as proportions of viable, apoptotic and necrotic cells were determined by BrdU-Assay and FACS analysis (AnnexinV/ Propidiumiodide staining), respectively (four consecutive passages). Gene expression analysis was carried out by Agilent cDNA-Microarray to indentify genes which displayed conjoint regulation in all cell lines. Furthermore, candidate genes were subjected to Ingenuity Pathways Analysis® to elucidate signal transduction pathways. Results: Incubation with TRD resulted in a significant dose dependent reduction of viable cells after 6 h and 24 h – leading to viable cells between 25.7 % (HT1080) and 66.2 % (HT29). Among all concentrations, TRD 250 μM in all cell lines showed the strongest effects in terms of reduction of viable cells, enhancement of apoptotic cells (FACS analysis) and inhibition of proliferation (BrdU assay). Among the cell death associated genes with the strongest regulation in gene expression (factor x45–x8), pro-apoptotic transcription factors (EGR1, FOS, FosB), pro-apoptotic proteins of the GADD family (GADD45B, GADD45A, GADD34), cell cycle regulators (E2F3, E2F6) as well as pro-apoptotic proteins of the Bcl-2 family (PMAIP1) were identified. Conclusion: This is the first conjoint analysis of potential target genes of TRD which was performed simultaneously in different malignant cell lines. The results indicate that TRD might be involved in different signal transduction pathways leading to apoptosis.