Inhibition of STAT3 signaling contributes to the anti-melanoma effects of chrysoeriol

Abstract

Background: Melanoma is an aggressive malignancy with a high mortality rate. Signal transducer and activator of transcription 3 (STAT3), an oncoprotein, is considered as an effective target for treating melanoma. Chrysoeriol is a flavonoid compound, and possesses anti-tumor activity in lung cancer, breast cancer and multiple myeloma; while whether it has anti-melanoma effects is still not known. Chrysoeriol has been shown to restrain STAT3 signaling in an inflammation mouse model.

Purpose: In this study, the anti-melanoma effects of chrysoeriol and the involvement of STAT3 signaling in these effects were investigated.

Study design and methods: CCK8 assays, 5-ethynyl-2'-deoxyuridine (EdU) staining, Annexin V-FITC/PI staining, Western blot analyses of cleaved caspase-9 and wound healing assays were used to study the anti-melanoma effects of chrysoeriol in cell models. A B16F10 melanoma bearing mouse model was used to evaluate the in vivo anti-melanoma effects of chrysoeriol. Indicators of cell proliferation, cell apoptosis and angiogeneis in melanoma tissues were detected by immunohistochemistry (IHC) staining and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) staining. Immune cells in melanoma tissues were analyzed by flow cytometry. STAT3-overactivated cell models were used to investigate the involvement of STAT3 signaling in the anti-melanoma effects of chrysoeriol. Molecular dynamics (MD) simulations and surface plasmon resonance (SPR) assays were conducted to determine whether chrysoeriol binds to Src, an upstream kinase of STAT3.

Results: The results of cell experiments showed that chrysoeriol dose-dependently inhibited viability, proliferation and migration of, and induced apoptosis in, A375 and B16F10 melanoma cells. Chrysoeriol inhibited the phosphorylation of STAT3, and downregulated the expression of STAT3-target genes involved in melanoma growth and metastasis. Mouse studies showed that chrysoeriol restrained melanoma growth and tumor-related angiogenesis, and altered compositions of immune cells in melanoma microenvironment. Chrysoeriol also inhibited STAT3 signaling in B16F10 allografts. Chrysoeriol's viability-inhibiting effects were attenuated by over-activating STAT3 in A375 cells. Furthermore, chrysoeriol bound to the protein kinase domain of Src, and suppressed Src phosphorylation in melanoma cells and tissues.

Conclusion: This study, for the first time, demonstrates that chrysoeriol has anti-melanoma effects, and these effects are partially due to inhibiting STAT3 signaling. Our findings indicate that chrysoeriol has the potential to be developed into an anti-melanoma agent.

Keywords: Chrysoeriol; Melanoma; STAT3; Src; Tumor microenvironment.