The increasing incidence of liver cancer remains a major global health threat, meanwhile liver cancer is commonly diagnosed at an advanced stage, when few therapeutic choices are available to most HCC patients(Sim and Knox 2018). There is an urgent need to find more effective treatments and drugs. Recently, adjuvant chemotherapy based on traditional Chinese herbs provided a novel therapeutic option for the treatment of liver cancer in China. In the past, corosolic acid has received a lot of attentions for its pharmacological activities such as anti-inflammatory and anti-diabetic, recently, corosolic acid has become a research hotspot owing to its antitumor effects. It has been reported that corosolic acid induces apoptosis in lung adenocarcinoma cells by regulating the expression of anti-apoptotic proteins, corosolic acid induces apoptosis in gastric cancer cells via AMPK-mTOR pathway, and in colon cancer cells by activating the expression of caspase proteins, etc(Lee et al. 2010; Nho et al. 2013; Sung et al. 2014). In the treatment of liver cancer, studies have shown that corosolic acid can inhibit the growth of liver cancer cells in vitro and in vivo. Simultaneously, there is a more obvious inhibitory effect when corosolic acid is combined with sorafenib(Ku et al. 2015). Although some progress has been made regarding the mechanism of action for corosolic acid in hepatocellular carcinoma, there is still a long way to go if we wish to fully understand the potential targets and pharmacological mechanisms of corosolic acid in regulating hepatocellular carcinoma tumorigenesis. The good news is that the development of bioinformatics has provided a new boost to the identification of natural product targets, which has enabled the discovery of new target molecules for the action of corosolic acid in hepatocellular carcinoma through network pharmacology.
The Bel-7404 cell line is multidrug resistant and highly malignant. In this study, we identified 44 differential proteins in Bel-7404 before and after treatment with corosolic acid. These proteins were found to be highly involved in specific cellular processes, such as brain development, antimicrobial humoral immune response mediated by antimicrobial peptide, etc. To find genes related to hepatocellular carcinoma in corosolic acid potential targets, we analyzed the TCGA hepatocellular carcinoma dataset and identified four candidate genes for potential therapeutic targets of corosolic acid among 4498 differentially expressed genes in liver cancer, which were P4HA2, CCDC137, NDRG1 and SDC4. One of the molecules, P4HA2, is involved in collagen formation and is a key enzyme in this process(Myllyharju 2003). Meanwhile, collagen is an important component of the extracellular matrix (ECM)(Gorres and Raines 2010), which can ultimately promote tumor development by promoting tumor cell proliferation, invasion, migration, epithelial-mesenchymal transition (EMT), and chemoresistance(Yang et al. 2014; Zheng et al. 2017; Yamazaki et al. 2018). Recently, abnormal expression of P4HA2 has been continuously reported as a marker of poor disease prognosis in tumors such as liver cancer, cervical cancer, and ovarian cancer(Cao et al. 2020; Zhang and Zhao 2021; Shang et al. 2022). Simultaneously, it has also been shown that aspirin reduces the expression level of P4HA2 through the NF-κB and LMCD1-AS1/let-7g pathways, thereby inhibiting collagen deposition and preventing the progression of hepatocellular carcinoma(Wang et al. 2019). In the present study, we found that P4HA2 protein was overexpressed in HCC tissues, furthermore, upregulation of P4HA2 is correlated with poor prognosis. Additionally, we demonstrated that corosolic acid can significantly inhibit HCC cell proliferation and clone formation, induce cell cycle arrest and decreas the protein level of P4HA2 in vitro. Consequently, these results suggest that corosolic acid can inhibit the growth of hepatocellular carcinoma cell, given the important role of P4HA2 in tumorigenesis. We suggest that P4HA2 protein is a key target for the treatment of HCC with corosolic acid.
In summary, this study demonstrated changes in HCC cell expression after treatment with corosolic acid, P4HA2 was identified as a potential target in corosolic acid antitumor process. These findings will facilitate the clinical application of corosolic acid and the identification of novel therapeutic targets.