The Comprehensive Effects of Carassius auratus Complex Formula against Lipid Accumulation, Hepatocarcinogenesis, and COVID-19 Pathogenesis via Stabilized G-Quadruplex and Reduced Cell Senescence
He-Yun Hsiao, Po-Jui Hsu, Bonifasius Putera Sampurna, Yu-Ju Lin, Kuan-Hao Lin, Yu-Ning Zhao, Horng-Dar Wang, Chiou-Hwa Yuh
First published: 22 March 2023 https://doi.org/10.1002/adbi.202200310
Carassius auratus complex formula (CACF) is a traditional Chinese medicine known for its antidiabetic effects. Hepatocellular carcinoma (HCC) is a major cause of cancer-related deaths worldwide, and there are currently no effective therapies for advanced HCC. This study explores the comprehensive effects and possible mechanisms of CACF on HCC. The results show that CACF reduces the viability of hepatoma cells in vitro, while benefiting normal hepatocytes. In addition, CACF inhibits hepatoma cell growth in a zebrafish xenotransplantation model and decreases lipid accumulation, represses inflammation and cell proliferation markers in fatty acid translocase (CD36) transgenic zebrafish, and inhibits the expression of cell proliferation and β-catenin downstream targets in telomerase (tert) transgenic zebrafish models. Ingenuity Pathway Analysis reveals that CACF exerts multiple functions, including reduction of inflammation and inhibition of lipid transporter and PPAR signaling pathway. Surprisingly, CACF also regulates the expression of genes and reduces coronavirus infection and pathogenesis in a zebrafish model. CACF treatment is validated to regulate the expression of genes for anti-coronavirus activity. Mechanistically, CACF stabilizes G-quadruplex and reduces cell senescence associated β-galactosidase activity. In summary, CACF may be a promising therapeutic agent with multiple functions including anticancer, anti-inflammation, and anti-microorganisms in a zebrafish model.
1.1 Therapeutics for Liver Cancer Is an Unmet Medical Need
Liver cancer is the fourth-most common malignant tumor disease worldwide. The most common type of primary liver cancer is hepatocellular carcinoma (HCC), which accounts for 90% of cases. The risk factors for hepatocellular carcinoma include infection by hepatitis B virus (HBV) and hepatitis C virus (HCV), nonalcoholic fatty liver (NAFLD), nonalcoholic steatohepatitis (NASH), alcohol consumption, and aflatoxin intake. NAFLD, which is most prevalent in industrialized countries, develops in four main stages: fatty liver (steatosis), steatohepatitis (NASH), fibrosis, and cirrhosis. With the advancement of anti-HBV vaccines and anti-HCV drugs, the incidence of virus-induced liver cancer gradually decreases, instead NASH-driven HCC increasingly becomes a global concern. Obesity and type 2 diabetes mellitus (T2DM) are two major factors linked to HCC, but the treatment for advanced HCC has limited selections, including sorafenib and other VEGF receptor inhibitors. The first-line treatment can only extend survival to 11–13 months and the second-line treatment to 8–10 months for advanced HCC patients. Moreover, cancer heterogeneity hampers the effectiveness of the therapeutics. Although atezolizumab (anti-PDL1) and bevacizumab (anti-VEGF) have increased the overall and progression-free survival in a phase III trial for advanced HCC and become the first line treatment, patients with NASH-driven HCC respond poorly and have compromised innate immunity. Therefore, effective treatment for advanced HCC is an urgent unmet medical need.
1.2 The Effects of Carassius auratus Complex Formula (CACF) and Traditional Chinese Medicine
CACF is composed of extracts from Carassius auratus, Rhizoma dioscoreae, Lycium chinense, and Rehmannia glutinosa Libosch. Oligosaccharides extracted from Rehmannia glutinosa Libosch can prevent ROS-related liver damage. A natural polysaccharide (RGP70-2) was isolated and purified from Rehmannia glutinosa Libosch, and may delay inflammation by regulating the ROS-NF-kappaB pathway. The compound extracted from Lycium chinense has antiapoptotic and anti-inflammatory effects, hepatic protection effects in mice, and lowers liver enzyme levels in human patients with mild hepatic dysfunction. Rhizoma dioscoreae extract was reported to regulate IL6/STAT3 signaling pathway to prevent the bone loss in rat model, and inhibit proinflammatory cytokines such as interleukin-1beta (IL-1beta) and tumor necrosis factor-alpha (TNF-alpha). Carassius auratus complex formula contains fish (Carassius auratus) and multiple herbs extract. These ingredients are used to treat diabetes since ancient China. CACF elicits antidiabetic effects in streptozotocin-induced diabetic mouse model under high-fat diet, and reduces triglyceride levels, maintains blood sugar control, and enhances insulin sensitivity. However, this is the only study on CACF. Therefore, the effects of CACF on anticancer, anti-inflammation, and the underlying mechanism remain unknown. In this study, we explore the comprehensive effects of CACF using in vitro and in vivo zebrafish models, also using whole transcriptomic analysis for identifying the genes/pathways regulated by CACF.
1.3 Overexpression of Cluster of Differentiation 36 (CD36) and Telomerase Reverse Transcriptase (TERT) in Hepatocellular Carcinoma (HCC)