Anticancer Effect of Cycas media: Molecular Basis Through Modulation of PI3K/AKT/mTOR Signaling Pathway

Abstract

Many researchers are focusing on screening the biological activities of plants owing to their safety and possible pharmacological actions. Consequently, we aimed to explore the antiproliferative and cytotoxic properties of Cycas media methanolic extract on HepG2 cell lines. Moreover, we also explore the antitumor action against the experimentally induced solid Ehrlich carcinoma (SEC) model and investigate the possible involved molecular mechanisms. Also, the antibacterial action of the extract was elucidated. Different concentrations of the extract were incubated with HepG2 to determine cytotoxicity, followed by cell cycle analysis. The in vivo experiment was accomplished by grouping the animals into four different groups (n = 10); normal control, SEC, C. media 100, and C. media 200. The extract was administered at 100 and 200 mg/kg. Tumor volume, tumor inhibition rate, toxicity profile, and antioxidant biomarkers were determined. Moreover, the PI3K/AKT/mTOR signaling pathway was investigated as a possible underlying antitumor mechanism. The tumor control group showed a remarkable upregulation for PI3K, p-AKT, and p-mTOR, along with downregulation for the antioxidant SOD and GPX4, as well as decreased levels of GSH and MDA. C. media extract reversed these parameters to a significant level and the higher dose showed a superior antitumor effect. C. media extract showed antiproliferative effects against HepG2 cells, along with a suppressive action on the PI3K/AKT/mTOR pathway and an antioxidant effect. Additionally, C. media had antibacterial consequences against S. aureus isolates with minimum inhibitory concentrations from 32 to 128 µg/mL. It also caused a noteworthy growth delay as well as a notable reduction in the membrane integrity of S. aureus isolates. These beneficial outcomes suggest C. media to have potential antitumor and antibacterial activities.

Keywords: Cycas media; EAC; HepG2; PI3K/AKT/mTOR; Staphylococcus aureus; antitumor.

Figure 1

Mass/mass spectra showed the fragmentation pattern of major identified compounds.

Figure 2

Cellular viability of HepG2 cells exposed to 2, 1, 0.5, 0.25, 0.125, and 0.0625 mg/mL of C. media extract. The IC₅₀ value was found to be 0.5183 mg/mL calculated from 3 repeats.

Figure 3

Effect of 500 µg/mL of C. media extract on cell cycle distribution of treated and control HepG2 cells. The cell cycle phases were analyzed after propidium iodide labeling. Data are presented as mean ± SD of three different experiments, (p < 0.01), a: significant versus control and b: significant versus DMSO.

Figure 4

Effect of C. media extract on (a) tumor volume and (b) tumor inhibition rate in SEC-bearing mice. Data are presented as mean ± SD for 10 mice and were analyzed using one-way ANOVA followed by Tukey post hoc test. p < 0.05, a: significant versus tumor control and b: significant versus C. media extract 100. SEC: solid Ehrlich carcinoma.

Figure 5

Toxicity profile of C. media extract on hepatic, cardiac, and renal biomarkers in SEC-bearing mice. (a) ALT, (b) AST, (c) cTnT, (d) CK-MB, (e) urea, and (f) creatinine. Data are presented as mean ± SD for 10 mice and were analyzed using one-way ANOVA followed by Tukey post hoc test. p < 0.05. a: significant versus normal control, b: significant versus disease control, c: significant versus C. media extract 100. SEC: solid Ehrlich carcinoma, ALT: alanine aminotransferase, AST: aspartate aminotransferase, cTnT: cardiac troponin, CK-MB: creatin kinase MB.

Figure 6

Effect of C. media extract on oxidative stress biomarkers and antioxidant enzymes in SEC-bearing mice. (a) GSH, (b) MDA, (c) SOD, and (d) GPX4. Data are presented as mean ± SD for 10 mice and were analyzed using one-way ANOVA followed by Tukey post hoc test. p < 0.05, a: significant versus normal control, b: significant versus disease control, and c: significant versus C. media extract 100. SEC: solid Ehrlich carcinoma, GSH: glutathione, MDA: malondialdehyde, SOD: superoxide dismutase, GPX4: glutathione peroxidase 4.

Figure 7

Modulation of PI3K/AKT/mTOR protein expression in the tumor tissue by C. media extract. Data are presented as mean ± SD for 3 readings per sample for 3 mice and were analyzed using one-way ANOVA followed by Tukey post hoc test. p < 0.05. a: significant versus normal control, b: significant versus disease control, c: significant versus C. media extract 100. SEC: solid Ehrlich carcinoma, PI3K: phosphatidylinositol 3-kinase, p-AKT: phosphorylated protein kinase B, p-mTOR: phosphorylated mammalian target of rapamycin.

Figure 8

An illustrative graph for the influence of C. media on the growth kinetics of S. aureus isolates.

Figure 9

A descriptive diagram displaying the influence of C. media on the membrane integrity of S. aureus isolate.