Abstract
Heat shock factor 1 (HSF1) is a transcription factor for heat shock proteins (HSPs) expression that enhances the survival of cancer cells exposed to various stresses. HSF1 knockout suppresses carcinogen-induced cancer induction in mice. Therefore, HSF1 is a promising therapeutic and chemopreventive target. We performed cell-based screening with a natural compound collection and identified fisetin, a dietary flavonoid, as a HSF1 inhibitor. Fisetin abolished heat shock-induced luciferase activity with an IC50 of 14 μM in HCT-116 cancer cells. The treatment of HCT-116 with fisetin inhibited proliferation with a GI50 of 23 μM. When the cells were exposed to heat shock in the presence of fisetin, the induction of HSF1 target proteins, such as HSP70, HSP27 and BAG3 (Bcl-2-associated athanogene domain 3), were inhibited. HSP70/BAG3 complexes protect cancer cells from apoptosis by stabilizing anti-apoptotic Bcl-2 family proteins. The downregulation of HSP70/BAG3 by fisetin significantly reduced the amounts of Bcl-2, Bcl-xL and Mcl-1 proteins, subsequently inducing apoptotic cell death. Chromatin immunoprecipitation assays showed that fisetin inhibited HSF1 activity by blocking the binding of HSF1 to the hsp70promoter. Intraperitoneal treatment of nude mice with fisetin at 30mg/kg resulted in a 35.7% (P < 0.001) inhibition of tumor growth.
Introduction
Cancer cells have multiple oncogenes and a high degree of signal redundancy, which causes a single anticancer drug to have relatively weak therapeutic effects. Therefore, combination therapy or multi-targeted therapy is a way to solve these obstacles. Most natural compounds have multiple target molecules and thus are useful for treating cancers. A nutraceutical is any substance considered to be a food or part of a food that provides medical benefits (1,2). Intake of dietary foods, such as fruits or spices, has been reported to reduce the risk of cancer genesis.
Fisetin (3,7,3′,4′-tetrahydroxyflavone) is a flavonoid found in many edible fruits and vegetables, such as apples, grapes, kiwis, persimmons, strawberries, cucumbers and onions that has multiple properties, such as antioxidant (3) and anti-inflammatory (4) activity. The highest levels of fisetin (160 μg/g wet food) are found in strawberries (5). Fisetin induced apoptosis in LNCaP human prostate cancer cells (6), and mice receiving fisetin at 45mg/kg inhibited tumor growth by 74.8% through androgen receptor inhibition (7). Similarly, fisetin also inhibited melanoma cell growth by disrupting Wnt/β-catenin signaling, and mice receiving fisetin at 45mg/kg suppressed tumor growth by 66.6% (8). Furthermore, fisetin has been reported as an inhibitor of Aurora B kinase (9), NF-κB (10), mammalian target of rapamycin (11) and PI3K/AKT and mammalian target of rapamycin signaling (12). Recently Kang and colleagues (13) reported that fisetin induces apoptosis in human non-small cell lung cancer by downregulating Bcl-2 expression.
The heat shock response protects cells from a wide range of stresses, including heat shock, oxidative stress, heavy metals, fever or protein misfolding (reviewed in (14and 15)). The heat shock response is mainly mediated by heat shock transcription factor 1 (HSF1) that controls the transcription of heat shock proteins (HSPs), such as HSP70, HSP47, HSP27 and BAG3 (Bcl-2-associated athanogene domain 3). Cells overexpressing HSP70 or HSP27 had an increased tumorigenicity when inoculated into mice (16,17). The expression of hsp70 is induced by several oncogenes, such as H-rasval12 (18), c-myc (19), c-myb, SV40 large T antigen and adenovirus E1a (20).
HSF1 knockout suppresses chemically induced skin cancer (21) and hepatocellular carcinoma induced by procarcinogen diethylnitrosamine (22) in mice. In addition, HSF1 knockdown has a minimal effect on normal primary human cells but significantly impairs the proliferation of several human malignant cell lines (21). Similarly, the downregulation of HSP70 or HSP27 was found to inhibit cell proliferation and induce apoptosis (23,24), suggesting that HSF1 is a highly potent target for cancer therapy.
BAG3 is a HSF1-inducible gene and protects cancer cells from apoptosis by stabilizing the Bcl-2 family proteins, such as Bcl-2, Bcl-xL and Mcl-1 (25). The BAG1 family was first reported as a Bcl-2 interacting protein (26). Six BAG family members were reported to regulate HSP70 function either positively or negatively. Whereas BAG-1 interacts with the proteasome and increases the degradation of HSP70 client proteins (27), BAG3 inhibits their proteasomal degradation (28).
In this study, we identified fisetin as an HSF1 inhibitor. Fisetin decreased the levels of not only HSP70/BAG-3 but also Bcl-2 family proteins; these changes induced apoptosis of cancer cells. Furthermore, we demonstrated that the inhibition of HSF1 activity occurred by blocking HSF1 binding to target gene promoters. This study is the first report that the anticancer activity of fisetin involves HSF1 inhibition.
R E S T A T S O U R C E : Oxford_CARCINOGENESIS
Edited by Engadin, 13 May 2019 - 03:12 PM.
