Cordyceps sinensis relieves non-small cell lung cancer by inhibiting the MAPK pathway

Abstract

Objective: To determine the pharmacodynamic mechanism underlying Cordyceps sinensis relief in a murine model of non-small cell lung cancer (NSCLC).

Methods: We created a murine model of NSCLC and studied the potential molecular mechanism by which C. sinensis relieved NSCLC using a combination of transcriptomics, proteomics, and experimental validation.

Results: C. sinensis markedly suppressed the fluorescence values in mice with NSCLC, improved the pathologic morphology of lung tissue, ameliorated inflammatory cytokines (tumor necrosis factor-alpha, interleukin-6, interleukin-10, and the oxidative stress indicators superoxide dismutase, malondialdehyde, and glutathione peroxidase). Transcriptomics results showed that the therapeutic effect of C. sinensis was primarily involved in the differentiation and activation of T cells. Based on the proteomic results, C. sinensis likely exerted a protective effect by recruiting immune cells and suppressing tumor cell proliferation via the MAPK pathway. Finally, the experimental validation results indicated that C. sinensis significantly decreased the VEGF and Ki67 expression, downregulated RhoA, Raf-1, and c-fos expression, which are related to cell migration and invasion, increased the serum concentration of hematopoietic factors (EPO and GM-CSF), and improved the percentage of immune cells (natural killer cells, dendritic cells, and CD4⁺ and CD8⁺ lymphocytes), which enhanced immune function.

Conclusions: Based on our preclinical study, C. sinensis was shown to exert a protective effect on NSCLC, primarily by inhibiting the MAPK pathway.

Keywords: Cordyceps Sinensis; MAPK pathway; NSCLC; Proteomics; Transcriptomics.

Fig. 1

Cordyceps sinensis inhibited lung cancer in mice with NSCLC. A Fluorescence photographs of mice were obtained 2 weeks after administration (n = 3; **P < 0.01 and ***P < 0.001 versus the model; ^&P < 0.05, vs. PD1). B Body weight for mice in the different treatment groups (n = 6). C Microscopic morphology of the lung tumor tissues in each group (HE, 20×). a Control group (Ctrl); b Model group (Model); c C. sinensis group (5 g/kg, BH); d 2 mg/kg PD1 inhibitor group (PD1); e 2 mg/kg PD1 inhibitor + 1 g/kg C. sinensis group (PD1 + BL); f 2 mg/kg PD1 inhibitor + 5 g/kg C. sinensis group (PD1 + BH).

Fig. 2

Influence of Cordyceps sinensis on inflammatory cytokines and indicators of oxidative stress in serum from mice with NSCLC. A–C Serum TNF-α, IL-6, and IL-10 concentrations in different groups. D–F Serum oxidative stress indicator (MDA, GSH-Px, and SOD) production in mice. Results are expressed as mean ± SD for n = 6 mice in each group. ^###P < 0.001 vs. control; *P < 0.05, **P < 0.01, ***P < 0.001 vs. model; ^&P < 0.05, ^&&P < 0.01, ^&&&P < 0.001, vs. PD1

Fig. 3

DEG analysis of C. sinensis against NSCLC. Volcano maps showing DEGs for (A) mice with NSCLC versus healthy mice and (B) mice with NSCLC versus BH mice. C, D GO (BP) enrichment results of DEGs in the model/control and BH/model groups. E–H GSEA enrichment results of DEGs in the model/control and BH/model groups

Fig. 4

DEP analysis of Cordyceps sinensis against NSCLC. A Flow chart of proteomics analysis. Volcano maps showing DEPs for B the model versus control groups and C the BH versus model groups. D, E GO (BP) enrichment result of DEPs in the model/control and BH/model groups

Fig. 5

Influence of Cordyceps sinensis on the levels of MAPK protein expression signaling pathway protein expression in mice with NSCLC. A, B Immunohistochemical staining (20×) was performed using different types of antibodies. Tumor expression of Ki67 and VEGF was analyzed via immunohistochemistry. Ki-67 is a protein marker of proliferation encoded by the MKI67 gene. VEGF, vascular endothelial growth factor. C, D Protein level analysis by Western blot in tumor tissues of key targets, including RhoA, Raf-1, c-fos, ERK1/2, MEK1/2 and their phosphorylated forms in the MAPK signaling pathway, respectively. E The serum hematopoietic growth factor (EPO and GM-CSF) concentration in mice from all groups. F Proportions of CD4⁺T cells, CD8⁺T cells, DCs, and NK cells in different groups. All values are expressed as the mean ± SD with n = 6 in each group. ^###P < 0.001, vs. Control; *P < 0.05, **P < 0.01, ***P < 0.001, vs. Model; ^&P < 0.05, ^&&P < 0.01, ^&&&P < 0.001, vs. PD1