Beetroot (Beta vulgaris) potential in preventing colorectal cancer using in-silico analysis

Abstract

Colorectal cancer (CRC) is one of the leading causes of cancer-related mortality worldwide, necessitating the need for an effective therapeutic strategy. Beta vulgaris (beetroot) possesses active compounds that exert anti-cancer properties. The aim of this study was to evaluate the potential of beetroot as a preventative agent against the progression of CRC using differentially expressed gene (DEG) analysis and network pharmacology approaches. The protein-protein interaction network and molecular docking analyses were employed to assess the key interactions of beetroot active compounds with CRC-related target protein. Cytotoxicity of beetroot extract was experimentally evaluated using 3-(4,5-dimethylthiazol-2-yl)-2,5 diphenyl tetrazolium bromide (MTT) assay on the HT29 cell line. The result of this study showed that protein in the cell cycle was significantly enriched in CRC, with cyclin-dependent kinase 4 (CDK4) gene as one of the specific genes. Quercetin, galangin, hesperidin, farrerol, and betanin were the most typical compounds of beetroot based on the Comparative Toxicogenomics Database (CTD). Molecular docking studies revealed the strong binding affinity between quercetin (-7.04 kcal/mol) and bentanin (-8.11 kcal/mol) with CDK4. Beetroot demonstrated anticancer properties against the HT29 cell line with IC₅₀ value of 39.03±1.4 µg/mL. In conclusion, the beetroot extract has inhibitory activity against HT29 cell line proliferation, highlighting its potential in preventing the development of CRC through the substantial suppression of gene expression within the cell cycle pathway.

Figure 1.

Volcano plot exhibiting significant differentially expressed gene (DEG) analysis of colorectal primary tumor cells and normal cells. Red and blue dots represent significant DEGs with p<0.05 and 1≤log2FC≤-1. Black dots represent non-significant DEGs.

Figure 2.

Protein network related to ulcerative colitis (UC) pathways in cancer and cell cycle with significant gene expression regulation. The thickness of the line indicates the more substantial the data supporting the protein/gene network interaction. Cluster 1: green nodes; cluster 2: red node; cluster 3: blue node.

Figure 3.

Mechanism of Beta vulgaris on the cell cycle (KEGG modification). Genes marked in red are genes that can influence regulation due to the addition of the active compound B. vulgaris.

Figure 4.

Gene ontology analysis related to cell cycle based on biological process (A), molecular function (B), and cellular components (C). The graphs were illustrated with blue to red gradient corresponding to the highest and lowest value, respectively.

Figure 5.

Chromatogram of GC-MS analysis of beetroot extract.

Figure 6.

Two- and three-dimensional structures for the interactions of CDK4 with quercetin (A) and betanin (B).

Figure 7.

Linear regression curve of log concentration against % inhibitions of beetroot extract (A) and doxorubicin (B).