Bioinformatics study on clinical prognosis of liver cancer and targeted therapy with traditional Chinese medicine based on screening of key pathogenic genes
Primary liver cancer is the third leading cause of cancer-related deaths worldwide, and hepatocellular carcinoma (HCC) is the most common type of primary liver cancer. Its risk factors mainly include hepatitis B virus (HBV)/hepatitis C virus (HCV) infection, long-term excessive alcohol consumption, consumption of food contaminated with Aspergillus flavus, liver cirrhosis caused by various reasons, and a family history of hepatocellular carcinoma. Early diagnosis of hepatocellular carcinoma often relies on biomarkers, which are widely used in clinical practice. Biomarkers mainly come from serum, plasma, microRNAs in tissues, mutated genes, proteins, etc. Many patients diagnosed with liver cancer are usually in the late unresectable stage. After palliative treatment, the median survival time is only 6-12 months, and the 5-year survival rate is only 10%. Therefore, screening key pathogenic genes of liver cancer has important clinical significance for early diagnosis and analysis of liver cancer prognosis in patients with liver cancer.

The rapid development of high-throughput sequencing technology has changed the research mode of biomedical science. Gene sequencing provides a new understanding of tumor biology issues such as the pathogenesis of tumors, and has important reference value for the diagnosis, prognosis, and treatment selection of tumors. Bioinformatics algorithms are crucial for processing high-throughput omics data. This study takes high-throughput gene chip data mining as the starting point, introduces bioinformatics algorithm knowledge, obtains liver cancer related datasets from GEO database, and analyzes differentially expressed genes between liver cancer tissue and normal liver tissue. Further screening key pathogenic genes of liver cancer from differentially expressed genes, exploring the biological functions of key genes and their involvement in signaling pathways, mutations, immune infiltration, and their impact on clinical prognosis, and further screening potential traditional Chinese medicines for the treatment of liver cancer, aiming to provide theoretical reference and scientific basis for the clinical diagnosis, treatment, and prognosis judgment of liver cancer.

 

Liver cancer is one of the most common cancers worldwide, with approximately 840000 people suffering from it each year and at least 780000 deaths due to it. China is a big country with hepatitis. The increase of hepatitis incidence rate has increased the number of liver cancer patients. About 50% of liver cancer patients in the world come from China. At present, the treatment of early liver cancer mainly includes surgical resection, radiofrequency ablation, and liver transplantation. In the middle stage, a combination of various methods such as surgery, radiofrequency ablation, targeted therapy, immunotherapy, and chemotherapy are usually used; In the late stage, supportive treatment is mostly adopted. Liver cancer is a malignant tumor with a very high recurrence rate. With radical treatment methods such as surgical resection, the recurrence rate within 5 years is still as high as 77%. Therefore, clinical treatment with a combination of traditional Chinese and Western medicine and traditional Chinese medicine can be considered. Traditional Chinese medicine does not have a specific term for liver cancer. Based on its symptoms, it is often classified as a disease category such as “syndrome differentiation”, “liver accumulation”, “jaundice”, and “swelling”. The treatment of liver cancer with traditional Chinese medicine includes the treatment of traditional Chinese medicine compound, the treatment of unit Chinese medicine/traditional Chinese patent medicines and simple preparations, as well as external treatment such as acupuncture and moxibustion and acupoint application; The combination of traditional Chinese medicine and Western medicine mainly includes traditional Chinese medicine combined with surgical treatment, traditional Chinese medicine combined with radiofrequency ablation, traditional Chinese medicine combined with targeted therapy, and traditional Chinese medicine combined with chemotherapy, in order to reduce complications, alleviate side effects, and delay tumor recurrence and metastasis. This article analyzes the chip dataset of liver cancer from the molecular mechanism level by using bioinformatics related knowledge, mining its pathogenic key genes, and analyzing the significance of key genes in liver cancer prognosis. Furthermore, potential traditional Chinese medicines for treating liver cancer are screened through key gene screening. This study aims to provide scientific basis and reference for the differential diagnosis, prognosis, and treatment of liver cancer.

GO functional and KEGG pathway enrichment analysis revealed that differentially expressed genes are mainly involved in biological processes such as immune response, inflammatory response, and cell adhesion, involving signaling pathways including p53, mTOR, ErbB, etc. Research has shown that the reaction between tumor testis antigen and Sal like protein 4 with specific T cells may play an important role in controlling early liver cancer; The deletion of nucleotide de binding oligomerization domain 2 in the liver can induce the occurrence of liver cancer through inflammation, DNA damage, and genomic instability; Interferon – γ derived from NK cells can promote the occurrence of liver cancer through the epithelial cell adhesion molecule epithelial mesenchymal transition process in HBV transgenic mice; In addition, studies have shown that miR-621 can enhance the radiosensitivity of liver cancer cells by activating the p53 signaling pathway; The highly expressed proteasome activity subunit 4 can promote the proliferation of liver cancer cells through the mTOR signaling pathway; Epithelial V-like antigen 1 can upregulate the ErbB3-PI3K signaling pathway, thereby promoting the progression and metastasis of liver cancer.

Protein interaction analysis identified 10 key genes highly expressed in liver cancer tissues, namely CDC20 CCNB2、CCNB1、BUB1、BIRC5、AURKA、TOP2A、NCAPH、CDK1。 Prognostic analysis of these 10 key genes revealed that the high expression group of key genes significantly reduced the overall survival of liver cancer patients, and these key genes were positively correlated with the pathological analysis results of liver cancer. Research has shown that the expression of CDC20 in liver cancer tissues is higher than that in normal tissues (P<0.05), and liver cancer patients with high expression of CDC20 have a lower overall survival rate; Research has shown that liver cancer patients with high levels of CCNB2 have shorter 5-year overall survival and disease-free survival compared to those with low levels of CCNB2; Other studies have shown that higher levels of BUB1B can be detected in liver cancer tissues, and overexpression of BUB1B is positively correlated with adverse clinical and pathological manifestations.

Further expression validation and mutation analysis of 10 key genes revealed that they were highly expressed in liver cancer tissues, with protein expression levels of CDC20, CCNB1, TOP2A, and NCAPH increasing to varying degrees in liver cancer tissues; In addition, there is a certain degree of mutation in key genes in liver cancer tissues, and liver cancer patients carrying this mutated key gene have a poorer prognosis. The immune cell infiltration analysis of key genes found that the expression of key genes is positively correlated with the degree of infiltration of B cells, CD8+T cells, CD4+T cells, macrophages, neutrophils, and dendritic cells. Yang Gang et al. used immunohistochemistry to detect the protein expression level of CDC20 in liver cancer tissues, and the results showed that high expression of CDC20 was observed in 59.2% of hepatocellular carcinoma samples. Rong Min Hua et al. found that mRNA and protein of CCNB1 are overexpressed in hepatocellular carcinoma tissues, and CCNB1 may participate in the cell cycle of HCC by regulating DNA replication. These results indicate that the predicted results in this article have good reliability and accuracy.

The screening results of potential therapeutic drugs found that CCNB1, BIRC5, CDK1, and CDC20 screened a large number of traditional Chinese medicines, and CDK1, CCNB1, BIRC5, and TOP2A bound a large number of potential natural therapeutic ingredients. CDK1, CCNB1, and BIRC5 may be universal targets for drug therapy. Artemisia annua, galangal, and winter grass can act on 2-3 key genes and are potential traditional Chinese medicines for treating liver cancer. Research has shown that artemisinin is the active ingredient in Artemisia annua, which can regulate the proliferation, apoptosis, and angiogenesis of liver cancer cells through various pathways such as EGFR; The effective ingredient of galangal is galangin, which can prevent hepatocellular carcinoma through endoplasmic reticulum stress and mitochondrial dependent apoptosis; Winter grass can enhance the sensitivity of liver cancer to sorafenib anticancer effects by targeting the AKT pathway. According to traditional Chinese medicine theory, the main pathogenesis of hepatocellular carcinoma is “blood stasis”, “toxicity”, and “deficiency”. Traditional Chinese medicine plays a role in supporting the body, eliminating evil, attacking toxins, and fighting cancer at the molecular level, achieving a state of balance between yin and yang in the body. Compared with traditional Western medicine treatment, traditional Chinese medicine has potential advantages in the treatment of hepatocellular carcinoma, which are reflected in the small side effects, low incidence of adverse reactions, slow tumor growth, reduced recurrence and metastasis, improved quality of life for patients, and prolonged survival cycle.

In summary, this study found that the occurrence and development of liver cancer have the characteristics of multiple genes, pathways, and functions. A total of 10 key genes for liver cancer pathogenesis were screened and obtained; The expression levels of these genes and their protein products are elevated in liver cancer patients, and are closely related to the poor prognosis of liver cancer and the degree of immune cell infiltration; Further exploration revealed 36 potential traditional Chinese medicines for targeted treatment of liver cancer, including artemisia annua, galangal, and winter grass, as well as 105 natural active ingredients. The research results of this article can provide scientific basis for the clinical diagnosis, prognosis judgment, and related treatment of liver cancer.