Abstract
Abstract
Liver cancer is listed among the most common causes o f cancer-related death. Nowadays the clinical diagnosis of early-stage hepatocellular carcinoma (HCC) has improved
significantly; nevertheless, HCC prognosis is still highly poor. Therefore, new effective and well-designed therapeutic strategies are urgently recommended. Ruthenium complexes represent a new promising family of metal-based anticancer complexes that exhibit reduced toxicity compared to platinum (II) complexes c urrently used clinically. These complexes show a novel mechanism of action with different spectrum of activity compared to platinum-based drugs. The present study was performed to investigate the effect of new formulated Rhuthenium complex {Bis (quinolin-8-olato) bis (triphenylphosphine) Ruthenium(II)} (Ru(quin)2) with or without sorafenib; the gold standard treatment for advanced HCC, as a combined treatment on different molecular signaling pathway such as ERK1/2, Stat3, AKT/m- TOR and Wnt/β-catenin that are responsible for HCC progression. In addition, the
cytotoxic effect of Ru(quin)2 was also examined on phase I and phase II drug metabolizing enzyme in vivo. HepG2 cells were treated with different concentration of Ru(quin)2 to evaluate its IC50 with or without 5µM sorafenib treatment to investigate morphological changes, apoptotic induction, histone release, caspase-3 activity and clonogenic formation. Furthermore, the protein expression levels of P-ERK1/2, Bax, β-catenin, cyclin D1, Stat3, m-TO R and S6- Kinase were also examined by western immunoblotting and gene expression of Aurora kinase B was assayed using qRT-PCR. Flow cytometric analysis was performed to analyze cell cycle machinery and immunocytochemical assay was used to visualize the nuclear and cytoplasmic localization of oncogenic β-catenin protein. In vitro data revealed that Ru(quin)2 exerts potent antitumor effect on HepG2 cells compared to sorafenib. There was a significant decrease in cell viability, proliferation and clonogenic inhibition of HepG2 cells after treatment for 24 h in a dose-dependent manner. In addition, a significant increase in histone release and caspase-3 activity was observed. Moreover, Ru(quin)2 significantly upregulated proapoptotic Bax protein and significantly downregulated the expression of Stat3, m- TOR, S6-Kinase, P-ERK1/2, oncogenic β-catenin and cyclin D1 which in turn was accompanied with G0/G1 cell cycle arrest and upregulation of Aurora kinase B. Immunocytochemical analysis of β-catenin showed a reduction in the nuclear localization and expression compared to control. Moreover, combined treatment of Ru(quin)2 and sorafenib showed more profound effect on proteins expression and apoptotic induction compared to each compound alone. In vivo results showed that Ru(quin)2 treatment was associated with stimulation of phase I and phase II drug metabolizingenzymes. The activities of phase 1 drug- metabolizing enzyme changed significantly in the low and high dose treated groups (2.5 mg/kg and 5 mg/kg every other day for 2 weeks) compared to control group. Meanwhile, Ru(quin)2 treatment of Albino mice upregulated CYP3A4 and NADPH-cytochrome-c-reductase expression compared to the control.
All in all, formulated Ru(quin)2 showed a promising inhibitor for HepG2 cell line proliferation, however further investigation for its complete mechanism is warranted.
Citation Format: Reem EL Sayed Bakir, Salah Ahmed Sheweita, Mohamed Ali Khalifa, Ahmed Samir Sultan. A novel Ruthenium complex induces apoptosis in HepG2 cell line [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 136.