Dehydrocurvularin,A Microbial Secondary Metabolite,Inhibits Breast Cancer By Blocking STAT3 Activation

Background:Breast cancer is one of the most common malignancies and currently ranks first among women worldwide in the incidence of malignant tumors.Its prevalence and mortality are high and tend to be younger.Despite the development of multiple treatments in recent years,breast cancer remains a major public health problem that threatens women's health.The malignant proliferation and metastasis of cancer cells is an important cause of death in breast cancer patients.Therefore,studying metastasis and proliferation is an extremely urgent scientific problem in the field of breast cancer prevention and treatment.The activation of Signal Transducer and Activator of Transcription 3(STAT3)is closely related to the biological behavior of tumors,and is directly involved in abnormal cell proliferation and malignant transformation,thus becoming an important target for tumor therapy.By screening microbial secondary metabolite libraries,we found that a metabolite Dehydrocurvularin(DCV)has anti-inflammatory and anti-cancer effects,but the mechanism of action is unclear.Objective:To explore the inhibitory effect of DCV on breast cancer cells and its inhibitory mechanism on STAT3 activityMethods:MTT assay was used to determine the effect of DCV on the survival rate of breast cancer MDA-MB-231,MDA-MB-468 cells and normal breast MCF-10 A cells.Scratch test,Transwell test and colony formation assay were used to detect the effect of DCV on the migration,invasion and colony formation of MDA-MB-231 cells;Flow cytometry and Western blot were used to study the effect of DCV on apoptosis.Luciferase assay was used to detect the effect of DCV on constitutive and IL-6(Interleukin-6)-induced STAT3 transcriptional activity.Western blot was used to detect the effect of DCV on constitutive and IL-6-induced phosphorylation of STAT3,the effect of?-interferon(Interferon-?,IFN-?)on STAT1,STAT5 activity and STAT3 upstream signals(IL-6,JAK kinase,protein tyrosine phosphatase(PTPs));Molecular docking simulation was used to study the interaction between DCV and STAT3.Western blot was used to detect the effect of DCV on the activationof STAT3 after treatment with DL-Dithiothreitol(DTT)and Glutathione(GSH).Results:The MTT results showed that DCV could significantly inhibited the proliferation of breast cancer MDA-MB-231 and MDA-MB-468 cells in a time and dose-dependent manner,and was safe with a small toxic side effect on normal breast MCF-10 A cells.The results of scratch test,Transwell test and colony formation showed that DCV significantly inhibited the migration,invasion and colony formation ability of breast cancer cells.Western blot showed that apoptosis was not obvious at 24 hours,but DCV promoted apoptosis at high doses within 48 hours.Luciferase assay and Western blot showed that DCV significantly inhibited constitutive and activation of STAT3 induced by IL-6.Similar to inhibition of IL-6-induced STAT3 phosphorylation,DCV also inhibited IFN-?-induced STAT3 phosphorylation,but not IFN-?-induced STAT1 and STAT5 activity.Western blot showed that DCV did not affect the upstream signals of STAT3(IL-6,JAKs kinase,protein tyrosine phosphatase(PTPs)).The chemical structure of DCV and molecular docking simulation suggest that DCV may directly interact with STAT3,and DTT/GSH treatment reverses the inhibitory effect of DCV on STAT3 activation.Conclusions:(1)DCV inhibits breast cancer cell growth.(2)DCV inhibits the growth of breast cancer cells by inducing apoptosis.(3)DCV selectively inhibits STAT3 activity,and the mechanism may be covalent binding to STAT3 by forming a disulfide bond.