The comparison of T98G cell response to trifluoperazine in 2D & 2.5D microenvironment

In cancer-drug interaction studies, 3D culture systems have been shown to mimic living organism conditions better than 2D culture environments. 2.5D cell culture environments have been developed to take advantage of 3D cell culture and mitigate its disadvantages. Trifluoperazine (TFP) is an FDA-approved antipsychotic and antiemetic drug used to treat schizophrenia. It has been shown to inhibit the proliferation and invasion of various cancer cells and to induce cell death in several cancer cell lines and animal models. T98G cells are a human glioblastoma cell line and are routinely used as an experimental model for the development of cancer therapies. There is a limited number of studies investigating the effect of TFP drug on T98G cell line in 2D culture. The aim of this thesis was to create an in vitro culture system for drug molecules that is cost-effective, efficient, and more similar to in vivo cell responses. The T98G glioblastoma cell line was evaluated for cell viability and morphology in the presence and absence of trifluoperazine (TFP) in 2.5D and 2D cell culture systems. For the 2.5D cell culture system, chitosan hydrogels cross-linked with genipin and glycerol phosphate were prepared and T98G cells were cultured directly on these wet hydrogels. The surface pore structure of the gels was examined by scanning electron microscopy. The response of the cells to Trifluoperazine (1 µM, 2 µM, 4 µM, 6 µM and 8 µM) drug molecule in 2D and 2.5D culture systems, cell viability was investigated by MTT assay and rezazurin methods. Changes in the cell nucleus were detected by DAPI staining. The IC50 concentrations of TFP on T98G cell line was calculated as 3.75 µM for 24 h and 2.57 µM for 48 h by MTT analysis for 2D culture on TCPS. IC50 concentrations of TFP on T98G cell line was calculated as 2.86 µM for 24 h and 2.64 µM for 48 h by resazurin analysis for 2D culture on TCPS. It was concluded that, IC50 concentrations calculated from MTT and resazurin analysis results were very close. DAPI staining also showed the condensed and small nuclei shape in T98G cells after 24 h application of TFP when compared to the control group. According to these results TFP doses were selected as 1-8 µM for 2.5D cell culture studies. 2.5D chitosan hydrogels were prepared using 0.5% acidic acid as a solvent, 100 µL glycerol phosphate (1 g/mL) and 1 mM genipin as physical and chemical crosslinkers. T98G cells were cultured on these hydrogels for 24 and 48 h. Viability of the cells were both determined by resazurin analysis. IC50 concentration of TFP for T98G cells cultured on 2.5D chitosan hydrogels were determined as 2.37 µM for 24 h culture and 1.94 µM for 48 h culture. In conclusion, it was observed that 2.5D chitosan hydrogels mimicked the 3D cell culture environment. The IC50 dose of cells cultured on 2.5D chitosan hydrogels was lower and closer to the IC50 value of cells cultured on 2D TCPS, indicating that T98G cells may be more sensitive to TFP drug on 2.5D hydrogels. The results showed that the hydrogels produced created in vitro and in vivo-like culture conditions for the cells and that the cell-drug response can be examined more closely than in vivo.