Abstract
Valproic acid (VPA), a well-known histone deacetylase (HDAC) inhibitor, is used as an anti-cancer drug for various cancers, but the synergistic anti-cancer effect of VPA and doxorubicin (DOX) combination treatment and its potential underlying mechanism in hepatocellular carcinoma (HCC) remain to be elucidated. Here, we evaluate the mono- and combination-therapy effects of VPA and DOX in HCC and identify a specific and efficient, synergistic anti-proliferative effect of the VPA and DOX combination in HCC cells, especially HepG2 cells; this effect was not apparent in MIHA cells, a normal hepatocyte cell line. The calculation of the coefficient of drug interaction confirmed the significant synergistic effect of the combination treatment. Concurrently, the synergistic apoptotic cell death caused by the VPA and DOX combination treatment was confirmed by Hoechst nuclear staining and Western blot analysis of caspase-3 and poly (ADP-ribose) polymerase (PARP) activation. Co-treatment with VPA and DOX enhanced reactive oxygen species (ROS) generation and autophagy, which were clearly attenuated by ROS and autophagy inhibitors, respectively. Furthermore, as an indication of the mechanism underlying the synergistic effect, we observed that DOX internalization, which was induced in the VPA and DOX combination-treated group, occurred via by the caveolae-mediated endocytosis pathway. Taken together, our study uncovered the potential effect of the VPA and DOX combination treatment with regard to cell death, including induction of cellular ROS, autophagy, and the caveolae-mediated endocytosis pathway. Therefore, these results present novel implications in drug delivery research for the treatment of HCC.
Highlights
According to the World Health Organization (WHO), hepatocellular carcinoma (HCC) is the second most common cause of cancer-related deaths worldwide
The results showed that HCC cell viability was inhibited significantly in a dose-dependent manner, while little or no effect was observed in MIHA cells (Figure 1B,C)
We performed histone deacetylase (HDAC) activity assay and revealed that HDAC activity was expectedly attenuated by the Valproic acid (VPA) treatment, while the combination of VPA and DOX treatment did not show a significant (p = 0.679) reduction compared to only VPA treatment did (Figure 1F)
Summary
According to the World Health Organization (WHO), hepatocellular carcinoma (HCC) is the second most common cause of cancer-related deaths worldwide. Doxorubicin (DOX) is a well-known, potent drug for the treatment of breast cancer patients [3], even though it displays adverse side-effects in terms of cardiotoxicity, hepatotoxicity nephrotoxicity, typhlitis, and other toxicities [4,5]. For advanced HCC treatment, doxorubicin (DOX), an anthracycline anticancer agent, has been used frequently; it shows a maximum response rate of approximately 15–20% [6,7], which is not sufficient for HCC treatment. Previous studies have suggested a number of factors that might cause DOX resistance in HCC [8,9,10]. An in vitro study showed that doxorubicin resistance was more severe under hypoxic conditions compared to normoxic conditions in HCC cells [11]. For more efficient cancer treatment, a combination treatment of DOX with other anticancer drugs may be an effective approach to HCC therapy
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