Abstract:
Cancer is one of the leading causes of death worldwide. In order to improve current cancer treatment modalities, the development of new therapy options for cancer is an important and necessary task of current medical research. Oncolytic viruses represent a promising strategy for cancer treatment (so-called virotherapy). Oncolytic viruses are capable of selectively infecting, replicating within and killing tumor cells with moderate toxicity for healthy non-malignant tissues. However, primary resistances of tumor cells against all the available virotherapeutics, which are mainly used in monotherapy, limit the efficiency of virus-induced oncolysis. Thus, there is a clinical need to accurately characterize these viral resistance phenomena and to find ways to efficiently overcome them. Because of the impact on inducing cancer cell death and inhibition of tumor proliferation, histone deacetylase inhibitors (HDACi) are interesting combination partners for new therapy options and might be a key to overcome current high-grade tumor cell resistance to virotherapy. Previous studies have shown promising results concerning virotherapeutics as well as epigenetic compounds, both independently and in combinatorial treatment.
The aim of this thesis was to establish a combined application scheme of vaccinia virus GLV-1h68 and distinct HDACi compounds, such as resveratrol, kaempferol (both natural compounds) or SAHA (suberylanilide hydroxamic acid, an FDA approved non-natural/chemical HDACi agent) on a panel of four different human cell lines, i.e., three colorectal cancer cell lines and one renal adenocarcinoma cell line. A further aim was to determine whether the above mentioned HDACi compounds can enhance the potency of VACV GLV-1h68 in infection-resistant cancer cell lines.
After having determined suitable concentrations of each agent in a pre-testing phase, next cell viability, tumor cell proliferation and oncolytic effects were analyzed in co-treatment by viability assays (sulforhodamin B, CellTiter-Blue), quantification of lactate dehydrogenase (LDH) release, as well as by real time monitoring of tumor cell growth. To further evaluate possible effects on viral replication, virus growth curves were performed. Finally, also cell cycle analysis, area calculation of the cell coverage of the surface of the culture wells and a comprehensive fluorescence microscopy analysis were determined.
Our results indicate that co-treatment of HDACi compounds with vaccinia virus GLV-1h68 causes a moderate increase of cytotoxic effects compared to the respective single-agent treatments in all cell lines. In addition, co-treatment resulted in an increased virus-associated expression of GFP in ACHN, HCT-15 and HCT-116 cells when compared to the treatment with GLV-1h68 alone. Nevertheless, cytotoxic effects varied in all examined cell lines depending on the performed method.
The murine cell line Colon-26 behaved adverse regarding the virus-associated expression/cytotoxic effects as presented in different experiments, e.g., area calculation and fluorescence monitoring. Here, further investigations need to be performed to identify reasons for the above-mentioned effects. Furthermore, SAHA seemed to be the most potent epigenetic compound when compared to the other epigenetic compounds used in this study.
In conclusion, our data show that epigenetic compounds, such as resveratrol, kaempferol and SAHA are able to further enhance the potency of VACV GLV1-h68 in partially or highly resistant tumor cell lines when compared to either of the mono¬therapeutic approaches. Induction of immunostimulatory effects being triggered by the epigenetic compound could lead to further improved cytotoxic effects in cancer patients. These findings may have an enormous trans¬lational potential for treatment of human colorectal and/or renal carcinomas as well as other solid tumors and have to be tested in forthcoming virotherapeutic clinical trials [142].
