Exploring the pharmacological mechanisms of icaritin against nasopharyngeal carcinoma via network pharmacology and experimental validation

Abstract

Background: Icaritin is a natural product with a wide range of anti-tumor effects. However, its anti-tumor mechanism has not been thoroughly studied. This study examined the inhibitory effect of icaritin on nasopharyngeal cancer and its underlying mechanism using network pharmacology along with in vivo and in vitro experiments.Methods: MTT and clone formation assays were used to detect the effects of icaritin on the viability and proliferation of nasopharyngeal carcinoma cells, followed by the construction of a HONE1 xenograft tumor model to evaluate the anti-tumor efficacy of icaritin in vivo. A public database was used to predict prospective targets, built a protein-protein interaction (PPI) network, and analyze gene enrichment and biological processes. Based on network pharmacological data, cell cycle-related proteins were identified using western blotting. Besides, cell cycle distribution, apoptosis, and intracellular reactive oxygen species (ROS) generation were identified using flow cytometry. In addition, SA-β-Gal staining was performed to detect cellular senescence, and western blotting was performed to detect the expression of P53, P21, and other proteins to verify key signaling pathways.Results: Icaritin effectively inhibited the viability and proliferation of nasopharyngeal carcinoma cell lines and showed good anti-tumor activity against HONE1 nasopharyngeal carcinoma cells in vivo. Key protein targets, including AKT1, HSP90AA1, CDK4, CCND1, and EGFR, were screened using PPI network topology analysis. GO and KEGG analysis revealed that the cell cycle, p53 signaling, and cell senescence pathways may be the main regulatory pathways. Flow cytometry and western blot experiments showed that icaritin caused S-phase arrest and promoted an increase in ROS. SA-β-Gal staining showed that icaritin significantly induced cellular senescence, and western blotting showed that the expression of senescence-related proteins p53 and P21 increased significantly. Moreover, inhibition of ROS levels by N-Acetylcysteine (NAC) enhanced cell viability, reversed cellular senescence and reduced cellular senescence-associated protein expression.Conclusion: The results of network pharmacological analysis and in vivo and in vitro experiments showed that icaritin effectively inhibited the growth of nasopharyngeal carcinoma cells, promoted ROS production, induced cellular senescence, and inhibited tumor cells, which are related to the regulation of P53/P21 signal pathway.

New insights into the anticancer therapeutic potential of icaritin and its synthetic derivatives

“…The anticancer mechanisms of action of icaritin include inhibition of proliferation (Zhao et al, 2015) and ROS induction, (Liu et al, 2022) among others. Nevertheless, there is scarce information regarding clinical research on cancer therapy using icaritin (Table 4).…”

“…In nasopharyngeal carcinoma cell lines, icaritin showed good antitumor activity (inhibiting viability and proliferation and inducing S‐phase arrest). Moreover, the apoptotic activity induction was not significant, suggesting that this antitumor property is not a viable mechanism of icaritin for this cancer type (Liu et al, 2022).…”

Effect of Icaritin on non-small cell lung cancer NCI-H1299 cells

“…A CCK8 assay was used to detect cell proliferation. Cells were seeded in 96-well plates at a density of 5 × 10 3 cells per well, and after 24 h, different concentrations of icaritin (5,10,20,40,60,80, and 100 μM) were added to the cells, which were cultured for 24, 48, or 72 h. Then, 10 μL of CCK8 solution was added to the cultures, which were further incubated at 37°C for 2 h. The absorbance was measured at 450 nm using a standard instrument. Each group was tested in triplicate.…”

“…Icaritin is the hydrolytic form of icariin, which may be isolated from plants of the genus Epimedium; this compound exerts an anti-tumor effect in a variety of tumor types including lung cancer [3], breast cancer [4], prostate cancer [5], acute myeloid leukemia, and liver cancer [6,7]. The anti-tumor mechanism of icaritin primarily involves promoting tumor cell apoptosis and necrosis [8,9], inducing cell cycle arrest [10], inhibiting tumor angiogenesis [11,12], and interfering with tumorrelated signaling pathways, such as the MAPK/ERK/JNK and PTEN/Akt/HIF-1α pathways [13,14].…”

The efficacy of natural products for the treatment of nasopharyngeal carcinoma

“…Icaritin has good anti‐tumor effects both in vivo and in vitro. Studies have shown that icaritin can inhibit the proliferation of NPC cells by promoting ROS production, damaging DNA, and inducing S‐phase arrest, and its mechanism is related to the regulation of p53/p21 signaling pathway (Liu, Hu, et al., 2022).…”