Olsalazine pretreatment augments chemosensitivity of gemcitabine in hepatocellular carcinoma

Abstract

Recently, olsalazine a DNA hypomethylating agent was found to inhibit the growth of breast cancer cells. The present study was carried out to evaluate the effects of olsalazine pretreatment in the potentiation of chemosensitivity of gemcitabine for the treatment of hepatocellular carcinoma (HCC). In silico molecular docking was performed to analyze the interaction of olsalazine and gemcitabine with DNMT1 and DNA, respectively, using the AutoDock tools 1.5.6. Cytotoxicity of olsalazine, gemcitabine, and combination were measured on human HePG2 cells using MTT assay. Antiproliferative effects were assessed using animal model of N-nitrosodiethylamine and carbon tetrachloride-induced HCC. Treatment was initiated from 8th week of induction to 11th week and change in body weight, liver weight, and survival rate were measured. Following treatment, blood samples were collected for estimation serum biochemistry. Blood serum was used for the estimation of inflammatory cytokines tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6), C-reactive protein [CRP], lactate dehydrogenase (LDH), and P53 levels. Oxidative stress markers were measured in liver tissue homogenates. Histopathology and immunohistochemistry (IHC) were performed on liver sections to detect the morphological changes and P53 expression. Docking analysis revealed the interactions between olsalazine and DNMT1 with a binding energy score of -5.34 and gemcitabine and DNA with a binding energy score of -5.93. Olsalazine pretreatment potentiated the antiproliferative effect of gemcitabine in cell line study. In the group receiving olsalazine pretreatment showed significant reductions in relative liver weight and improved survival rate of gemcitabine treatment group. Serum biochemical markers: serum glutamate pyruvate transaminase, serum glutamic oxaloacetic transaminase, alkaline phosphatase, and bilirubin revealed improved liver functions. Olsalazine pretreatment also reduced the levels of inflammatory markers like CRP, LDH, TNF-α, and IL-6 and oxidative stress markers dose dependently. Histopathology and IHC showed improved liver morphology with potentiated the induction of P53 upon olsalazine pretreatment in combination with gemcitabine. In conclusion, sequential combination of olsalazine and gemcitabine improved the treatment outcomes during the progression of HCC.

Keywords: DNMT1; gemcitabine; hepatocellular carcinoma; olsalazine.