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Hasan Mukhtar - One of the best experts on this subject based on the ideXlab platform.
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Fisetin and 5-fluorouracil: Effective combination for PIK3CA-mutant colorectal cancer.
International Journal of Cancer, 2019Co-Authors: Naghma Khan, Farah Jajeh, Emily L. Eberhardt, Devon Miller, Dawn M. Albrecht, Rachel Van Doorn, Melissa D. Hruby, Morgan E. Maresh, Linda Clipson, Hasan MukhtarAbstract:The normal colon epithelium is transformed into its neoplastic counterpart through a series of genetic alterations in driver genes including activating mutations in PIK3CA. Treatment often involves surgery followed by 5-fluorouracil (5-FU) based therapy, which has limited efficiency and serious side effects. We sought to determine whether Fisetin, a dietary flavonoid, alone or in combination with 5-FU affected tumorigenesis in the mammalian intestine. We first determined the effect of Fisetin, 5-FU or their combination on PIK3CA-mutant and PIK3CA wild-type colon cancer cells by assessing cell viability, colony formation, apoptosis and effects on PI3K/AKT/mTOR signaling. Treatment of PIK3CA-mutant cells with Fisetin and 5-FU reduced the expression of PI3K, phosphorylation of AKT, mTOR, its target proteins, constituents of mTOR signaling complex and this treatment increased the phosphorylation of AMPKα. We then determined whether Fisetin and 5-FU together or singly affected tumorigenesis in ApcMin/+ mice that also express constitutively active PI3K in the distal small intestine and colon. Tumor incidence was markedly lower in Fisetin-treated FC1 3K1 ApcMin/+ mice that also express constitutively active PI3K in distal small intestine and colon, as compared to control animals, indicating that Fisetin is a strong preventive agent. In addition, the combination of Fisetin and 5-FU also reduced the total number of intestinal tumors. Fisetin could be used as a preventive agent plus an adjuvant with 5-FU for the treatment of PIK3CA-mutant colorectal cancer.
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Fisetin targets yb 1 rsk axis independent of its effect on erk signaling insights from in vitro and in vivo melanoma models
Scientific Reports, 2018Co-Authors: Mario Sechi, Hasan Mukhtar, Rahul K Lall, Saheed O Afolabi, Anant Singh, Dinesh C Joshi, S Y Chiu, Deeba N SyedAbstract:The anti-proliferative activity of dietary flavonoid Fisetin has been validated in various cancer models. Establishing its precise mechanism of action has proved somewhat challenging given the multiplicity of its targets. We demonstrated that YB-1 promotes epithelial-to-mesenchymal transition and its inhibition suppressed tumor cell proliferation and invasion. The p90 ribosomal S6 kinase (RSK), an important ERK effector, activates YB-1 to drive melanoma growth. We found that Fisetin treatment of monolayer/3-D melanoma cultures resulted in YB-1 dephosphorylation and reduced transcript levels. In parallel, Fisetin suppressed mesenchymal markers and matrix-metalloproteinases in melanoma cells. Data from cell-free/cell-based systems indicated that Fisetin inhibited RSK activity through binding to the kinase. Affinity studies for RSK isoforms evaluated stronger interaction for RSK2 than RSK1. Competition assays performed to monitor binding responses revealed that YB-1 and RSK2 do not compete, rather binding of Fisetin to RSK2 promotes its binding to YB-1. Fisetin suppressed YB-1/RSK signaling independent of its effect on ERK, and reduced MDR1 levels. Comparable efficacy of Fisetin and vemurafenib for inhibiting melanoma growth was noted albeit through divergent modulation of ERK. Our studies provide insight into additional modes of regulation through which Fisetin interferes with melanoma growth underscoring its potential therapeutic efficacy in disease progression.
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dietary flavonoid Fisetin binds to β tubulin and disrupts microtubule dynamics in prostate cancer cells
Cancer Letters, 2015Co-Authors: Eiman Mukhtar, Vaqar M Adhami, Mario Sechi, Hasan MukhtarAbstract:Microtubule targeting based therapies have revolutionized cancer treatment; however, resistance and side effects remain a major limitation. Therefore, novel strategies that can overcome these limitations are urgently needed. We made a novel discovery that Fisetin, a hydroxyflavone, is a microtubule stabilizing agent. Fisetin binds to tubulin and stabilizes microtubules with binding characteristics far superior than paclitaxel. Surface plasmon resonance and computational docking studies suggested that Fisetin binds to β-tubulin with superior affinity compared to paclitaxel. Fisetin treatment of human prostate cancer cells resulted in robust up-regulation of microtubule associated proteins (MAP)-2 and -4. In addition, Fisetin treated cells were enriched in α-tubulin acetylation, an indication of stabilization of microtubules. Fisetin significantly inhibited PCa cell proliferation, migration, and invasion. Nudc, a protein associated with microtubule motor dynein/dynactin complex that regulates microtubule dynamics, was inhibited with Fisetin treatment. Further, Fisetin treatment of a P-glycoprotein overexpressing multidrug-resistant cancer cell line NCI/ADR-RES inhibited the viability and colony formation. Our results offer in vitro proof-of-concept for Fisetin as a microtubule targeting agent. We suggest that Fisetin could be developed as an adjuvant for treatment of prostate and other cancer types.
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abstract 2531 Fisetin enhances the efficacy of cabazitaxel an in vitro and in vivo study in prostate cancer
Cancer Research, 2015Co-Authors: Eiman Mukhtar, Vaqar M Adhami, Mario Sechi, Hasan MukhtarAbstract:Proceedings: AACR 106th Annual Meeting 2015; April 18-22, 2015; Philadelphia, PA Despite advances in screening and treatment, prostate cancer (PCa) remains a leading cause of death among American men. A wide range of novel therapies have been introduced for treatment of PCa which lead to dramatic regression of the primary tumor. However, responses are often short-lived due to the development of resistant cancer cells. Cabazitaxel, a microtubule targeting agent, is approved for treatment of metastatic hormone-refractory PCa in patients pretreated with docetaxel. However, toxicity and resistance are associated with cabazitaxel treatment. Therefore, an effective approach that can achieve long-term improvement is urgently needed for the management of PCa. One of the strategies proposed for overcoming resistance is combination therapy. Natural dietary substances have received considerable attention as chemoprevention and chemotherapeutic agents because of their unique property that allows them to interact with multiple targets in cancer cells. Fisetin, a small molecule, has emerged as one of the natural agent with enormous potential in cancer chemoprevention and chemotherapy. This study was designed to investigate the effect of a combination of Fisetin and cabazitaxel to achieve maximum therapeutic benefits, reduce dose and toxicity, and to minimize or delay the induction of drug resistance. We observed that treatment of PCa cells with Fisetin (20-80 μM) for 24-48 hours and then cabazitaxel (20-80 nM) for 24 hours significantly retarded the growth of PCa cells when compared to cabazitaxel or Fisetin alone. The combination resulted in 80% inhibition of cell growth when compared to cabazitaxel (30%) or Fisetin alone (18%). In addition, the combination (fisein, 20μM, cabazitaxel 5 nM) treatment significantly inhibited colony formation compared to each agent alone. We next implanted 22Rν1 cells for determining the effects of Fisetin, cabazitaxel and their combination. A total of 24 athymic nude male mice (6-8) weeks of age were injected subcutaneously with 1×106 22Rν1 cells. Two weeks later, tumor bearing mice were randomly divided into four groups (n = 6) and treated with Fisetin (20 mg/kg; 3 times/week), cabazitaxel (5 mg/kg; once/week), and combination of Fisetin (20 mg/kg; 3 times/week) and cabazitaxel (5 mg/kg; once/week), or vehicle (control) via intraperitoneal route. We observed that treatment with Fisetin alone resulted in 14% inhibition of tumor growth; cabazitaxel treatment alone resulted in 36% inhibition whereas combination treatment with Fisetin and cabazitaxel resulted in 53% inhibition of tumor growth compared with the control group. Tissue staining with proliferation marker Ki67 confirmed the in-vivo effect of Fisetin and cabazitzxel on inhibition of cell proliferation. This study potentiates the benefit of combination of Fisetin and cabazitaxel that could eliminate rapidly proliferating and resistant cells in PCa and possibly other cancer types. Citation Format: Eiman Mukhtar, Vaqar Mustafa Adhami, Mario Sechi, Hasan Mukhtar. Fisetin enhances the efficacy of cabazitaxel: an in vitro and in vivo study in prostate cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2531. doi:10.1158/1538-7445.AM2015-2531
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involvement of er stress and activation of apoptotic pathways in Fisetin induced cytotoxicity in human melanoma
Archives of Biochemistry and Biophysics, 2014Co-Authors: Deeba N Syed, Rahul K Lall, Jean Christopher Chamcheu, Omar Mohammad Haidar, Hasan MukhtarAbstract:Abstract The prognosis of malignant melanoma remains poor in spite of recent advances in therapeutic strategies for the deadly disease. Fisetin, a dietary flavonoid is currently being investigated for its growth inhibitory properties in various cancer models. We previously showed that Fisetin inhibited melanoma growth in vitro and in vivo. Here, we evaluated the molecular basis of Fisetin induced cytotoxicity in metastatic human melanoma cells. Fisetin treatment induced endoplasmic reticulum (ER) stress in highly aggressive A375 and 451Lu human melanoma cells, as revealed by up-regulation of ER stress markers including IRE1α, XBP1s, ATF4 and GRP78. Time course analysis indicated that the ER stress was associated with activation of the extrinsic and intrinsic apoptotic pathways. Fisetin treated 2-D melanoma cultures displayed autophagic response concomitant with induction of apoptosis. Prolonged treatment (16 days) with Fisetin in a 3-D reconstituted melanoma model resulted in inhibition of melanoma progression with significant apoptosis, as evidenced by increased staining of cleaved Caspase-3 in the treated constructs. However, no difference in the expression of autophagic marker LC-3 was noted between treated and control groups. Fisetin treatment to 2-D melanoma cultures resulted in phosphorylation and activation of the multifunctional AMP-activated protein kinase (AMPK) involved in the regulation of diverse cellular processes, including autophagy and apoptosis. Silencing of AMPK failed to prevent cell death indicating that Fisetin induced cytotoxicity is mediated through both AMPK-dependent and -independent mechanisms. Taken together, our studies confirm apoptosis as the primary mechanism through which Fisetin inhibits melanoma cell growth and that activation of both extrinsic and intrinsic pathways contributes to Fisetin induced cytotoxicity.
Farrukh Afaq - One of the best experts on this subject based on the ideXlab platform.
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Fisetin inhibits uvb induced cutaneous inflammation and activation of pi3k akt nfκb signaling pathways in skh 1 hairless mice
Photochemistry and Photobiology, 2015Co-Authors: Harish C Pal, Craig A Elmets, Mohammad Athar, Farrukh AfaqAbstract:Solar ultraviolet B (UVB) radiation has been shown to induce inflammation, DNA damage, p53 mutations and alterations in signaling pathways eventually leading to skin cancer. In this study, we investigated whether Fisetin reduces inflammatory responses and modulates PI3K/AKT/NFκB cell survival signaling pathways in UVB-exposed SKH-1 hairless mouse skin. Mice were exposed to 180 mJ cm−2 of UVB radiation on alternate days for a total of seven exposures, and Fisetin (250 and 500 nmol) was applied topically after 15 min of each UVB exposure. Fisetin treatment to UVB-exposed mice resulted in decreased hyperplasia and reduced infiltration of inflammatory cells. Fisetin treatment also reduced inflammatory mediators such as COX-2, PGE2 as well as its receptors (EP1–EP4) and MPO activity. Furthermore, Fisetin reduced the level of inflammatory cytokines TNFα, IL-1β and IL-6 in UVB-exposed skin. Fisetin treatment also reduced cell proliferation markers as well as DNA damage as evidenced by increased expression of p53 and p21 proteins. Further studies revealed that Fisetin inhibited UVB-induced expression of PI3K, phosphorylation of AKT and activation of the NFκB signaling pathway in mouse skin. Overall, these data suggest that Fisetin may be useful against UVB-induced cutaneous inflammation and DNA damage.
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Fisetin inhibits growth induces g2 m arrest and apoptosis of human epidermoid carcinoma a431 cells role of mitochondrial membrane potential disruption and consequent caspases activation
Experimental Dermatology, 2013Co-Authors: Harish C Pal, Samriti Sharma, Craig A Elmets, Mohammad Athar, Farrukh AfaqAbstract:Non-melanoma skin cancers (NMSCs), one of the most common neoplasms, cause serious morbidity and mortality. Therefore, identification of non-toxic phytochemicals for prevention/treatment of NMSCs is highly desirable. Fisetin (3,3',4',7-tetrahydroxyflavone), a dietary flavonoid, present in fruits and vegetables possesses anti-oxidant and antiproliferative properties. The aim of this study was to investigate the chemotherapeutic potential of Fisetin in cultured human epidermoid carcinoma A431 cells. Treatment of A431 cells with Fisetin (5-80 μm) resulted in a significant decrease in cell viability in a dose- and time-dependent manner. Employing clonogenic assay, we found that Fisetin treatment significantly reduced colony formation in A431 cells. Fisetin treatment of A431 cells resulted in G₂ /M arrest and induction of apoptosis. Furthermore, treatment of A431 cells with Fisetin resulted in (i) decreased expression of anti-apoptotic proteins (Bcl2; Bcl-xL and Mcl-1); (ii) increased expression of pro-apoptotic proteins (Bax, Bak and Bad); (iii) disruption of mitochondrial potential; (iv) release of cytochrome c and Smac/DIABLO from mitochondria; (v) activation of caspases; and (vi) cleavage of Poly(ADP-ribose) polymerase (PARP) protein. Pretreatment of A431 cells with the pan-caspase inhibitor (Z-VAD-FMK) blocked Fisetin-induced cleavage of caspases and PARP. Taken together, these data provide evidence that Fisetin possesses chemotherapeutic potential against human epidermoid carcinoma A431 cells. Overall, these results suggest that Fisetin could be developed as a novel therapeutic agent for the management of NMSCs.
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Fisetin induces autophagic cell death through suppression of mtor signaling pathway in prostate cancer cells
Carcinogenesis, 2010Co-Authors: Yewseok Suh, Farrukh Afaq, Naghma Khan, Jeremy J Johnson, Fatima H Khusro, Hasan MukhtarAbstract:The mammalian target of rapamycin (mTOR) kinase is an important component of PTEN/PI3K/Akt signaling pathway, which is frequently deregulated in prostate cancer (CaP). Recent studies suggest that targeting PTEN/PI3K/Akt and mTOR signaling pathway could be an effective strategy for the treatment of hormone refractory CaP. Here, we show that the treatment of androgen-independent and PTEN-negative human CaP PC3 cells with Fisetin, a dietary flavonoid, resulted in inhibition of mTOR kinase signaling pathway. Treatment of cells with Fisetin inhibited mTOR activity and downregulated Raptor, Rictor, PRAS40 and GβL that resulted in loss of mTOR complexes (mTORC)1/2 formation. Fisetin also activated the mTOR repressor TSC2 through inhibition of Akt and activation of AMPK. Fisetin-mediated inhibition of mTOR resulted in hypophosphorylation of 4EBP1 and suppression of Cap-dependent translation. We also found that Fisetin treatment leads to induction of autophagic-programmed cell death rather than cytoprotective autophagy as shown by small interfering RNA Beclin1-knockdown and autophagy inhibitor. Taken together, we provide evidence that Fisetin functions as a dual inhibitor of mTORC1/2 signaling leading to inhibition of Cap-dependent translation and induction of autophagic cell death in PC3 cells. These results suggest that Fisetin could be a useful chemotherapeutic agent in treatment of hormone refractory CaP.
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Fisetin a novel dietary flavonoid causes apoptosis and cell cycle arrest in human prostate cancer lncap cells
Carcinogenesis, 2008Co-Authors: Naghma Khan, Farrukh Afaq, Deeba N Syed, Hasan MukhtarAbstract:467 Prostate cancer (PCa) has become one of the most frequently diagnosed malignancies in the United States with similar trend in most Western countries. The substantial mortality and morbidity associated with PCa and its poor treatment options have led a surge to develop novel means for its prevention. The development of chemoprevention strategies against PCa would have huge impact, both medically and economically. Because of its ubiquity and disease burden, PCa is an attractive target for chemoprevention. Fisetin (3,39,49,7-tetrahydroxyflavone), is found in fruits andvegetables, such as strawberry, apple, persimmon, grape, onionand cucumber. The aim of this study was to determine the effect of Fisetin on cell-growth and apoptosis in human PCa cells. Treatment of Fisetin (10-60 μM; 48 h) was found to result in a decrease in the viability of LNCaP, CWR22Rυ1 and PC-3 cells but had only minimal effect on normal prostate epithelial PrEC cells as assessed by MTT assay. To decipher the mechanism of growth inhibition of Fisetin, we selected LNCaP cells for further studies. Treatment of LNCaP cells with Fisetin (10-60 μM; 48 h) also resulted in G1-phase arrest (as assessed by DNA cell cycle analysis). Fisetin also caused a marked decrease in the protein expression of cyclins D1, D2 and E and their activating partners cdks 2, 4 and 6 with concomitant induction of WAF1/p21 and KIP1/p27. Treatment of LNCaP cells with Fisetin also resulted in significant apoptosis in a dose-dependent fashion as determined by ELISA. A significant increase in the proteinexpression of Bax, Bak, Bad and Bid (proapoptotic) and decrease in Bcl-2 and BCl-xL (antiapoptotic) was observed on treatment with Fisetin in a dose-dependent fashion thus further confirming the induction of apoptotic process. Additionally, PARP cleavage analysis showed that the full size PARP (116 KD) protein was cleaved to yield an 85 KD fragment after treatment of cells with Fisetin. Fisetin treatment of cells also resulted in induction of mitochondrial release of cytochrome c into cytosol and downregulation of XIAP and upregulation of Smac/DIABLO. Treatment of cells with Fisetin also resulted in significant activation of caspase-3, -8 and -9. Pretreatment of cells with caspase inhibitor (Z-VAD-FMK) blocked Fisetin-induced activation of caspases as detected by immunoblot analysis and immunofluorescence staining. These data provide the first evidence that Fisetin could be developed as an agent against prostate cancer.
Yenchou Chen - One of the best experts on this subject based on the ideXlab platform.
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wogonin and Fisetin induction of apoptosis through activation of caspase 3 cascade and alternative expression of p21 protein in hepatocellular carcinoma cells sk hep 1
Archives of Toxicology, 2002Co-Authors: Yenchou Chen, Shingchuan Shen, Woanruoh Lee, Hui Yi Lin, Chunming Shih, Lingling YangAbstract:Wogonin and Fisetin are flavonoids, which are widely distributed in plants. Our recent study demonstrated that, among seven structurally related flavonoids, wogonin and Fisetin showed the most potent apoptosis-inducing activities in human promyeloleukemic cells HL-60. In the present investigation, we performed molecular studies to assess the apoptotic effects of wogonin and Fisetin on hepatocellular carcinoma cells SK-HEP-1. Both wogonin and Fisetin showed dose-dependent cytotoxic effects on SK-HEP-1 cells, accompanied by DNA fragmentation. Microscopic observation under Giemsa staining showed that wogonin and Fisetin, at the dose of 80 µM, induced cellular swelling and the appearance of apoptotic bodies, characteristics of apoptosis, in SK-HEP-1 cells. Furthermore, flow cytometry analysis showed an increase of hypodiploid cells in wogonin- and Fisetin-treated SK-HEP-1 cells. These data demonstrated that wogonin and Fisetin were effective inducers of apoptosis in SK-HEP-1 cells. Treatment with an apoptosis-inducing concentration of wogonin or Fisetin caused induction of caspase 3/CPP32 activity, but not of caspase 1 activity. In addition, a caspase 3 inhibitor, Ac-DEVD-CHO, but not the caspase 1 inhibitor Ac-YVAD-CHO, reversed the cytotoxic effects of wogonin and Fisetin on SK-HEP-1 cells. Further, cleavage of caspase 3 substrates including poly(ADP-ribose) polymerase (PARP) and D4-GDI protein, and decrease of pro-caspase 3 protein were detected in wogonin- and Fisetin-treated SK-HEP-1 cells. Increase of p53 protein was associated with wogonin- and Fisetin-induced apoptosis; however, a p53-controlled gene, p21Waf/Cip-1, was only induced in wogonin- (not Fisetin-) treated SK-HEP-1 cells. Serum starvation elevated p21Waf/Cip-1 protein expression, and enhanced the apoptotic induction activity of wogonin (not fiseitn) in SK-HEP-1 cells. Our study has provided molecular evidence to demonstrate that wogonin and Fisetin had effective cytotoxic effects through apoptosis induction in hepatocellular carcinoma cells SK-HEP-1; activation of caspase 3 cascade, induction of p53 protein and alternative expression of p21Waf/Cip-1 protein were involved.
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wogonin and Fisetin induce apoptosis in human promyeloleukemic cells accompanied by a decrease of reactive oxygen species and activation of caspase 3 and ca2 dependent endonuclease
Biochemical Pharmacology, 2002Co-Authors: Woan Rouh Lee, Shingchuan Shen, Hui Yi Lin, Lingling Yang, Wen Chi Hou, Yenchou ChenAbstract:Abstract Seven structurally related flavonoids including luteolin, nobiletin, wogonin, baicalein, apigenin, myricetin and Fisetin were used to study their biological activities on the human leukemia cell line, HL-60. On MTT assay, wogonin, baicalein, apigenin, myricetin and Fisetin showed obvious cytotoxic effects on HL-60 cells, with wogonin and Fisetin being the most-potent apoptotic inducers among them. The cytotoxic effects of wogonin and Fisetin were accompanied by the dose- and time-dependent appearance of characteristics of apoptosis including DNA fragmentation, apoptotic bodies and the sub-G1 ratio. Treatment with an apoptosis-inducing concentration of wogonin or Fisetin causes rapid and transient induction of caspase 3/CPP32 activity, but not caspase 1 activity. Further, cleavage of poly(ADP-ribose) polymerase (PARP) and decrease of pro-caspase 3 protein were detected in wogonin- and Fisetin-treated HL-60 cells. An increase in the pro-apoptotic protein, bax, and a decrease in the anti-apoptotic protein, Mcl-1, were detected in Fisetin- and wogonin-treated HL-60 cells. However, Bcl-2, Bcl-XL, and Bad all remained unchanged in wogonin- and Fisetin-treated HL-60 cells. In vitro chromatin digestion revealed that endonuclease activity was profoundly enhanced in wogonin- and Fisetin-treated HL-60 cells, and the addition of ethylenediaminetetraacetic acid (EDTA) or ethyleneglycoltetraacetic acid (EGTA) into the reaction blocked endonuclease activation and at an optimum pH of 7.5. The caspase 3 inhibitor, Ac-DEVD-CHO, but not the caspase 1 inhibitor, Ac-YVAD-CHO, attenuated wogonin- and Fisetin-induced DNA ladders, PARP cleavage, and endonuclease activation. Pretreatment of HL-60 cells with N -acetyl-cysteine or catalase efficiently inhibited H 2 O 2 (200 μM)-induced apoptosis, but showed no inhibitory effect on wogonin- and Fisetin-induced DNA ladders, caspase 3 activation, or bax protein induction. Decrease in endogenous ROS production was detected in wogonin- and Fisetin-treated HL-60 cells by DCHF-DA assay. In conclusion, our experiments indicate that a decrease in intracellular peroxide level was involved in wogonin- and Fisetin-induced apoptosis; activation of caspase 3 and endonuclease, induction of bax protein and suppression of Mcl-1 protein were detected in the process.
Lingling Yang - One of the best experts on this subject based on the ideXlab platform.
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wogonin and Fisetin induction of apoptosis through activation of caspase 3 cascade and alternative expression of p21 protein in hepatocellular carcinoma cells sk hep 1
Archives of Toxicology, 2002Co-Authors: Yenchou Chen, Shingchuan Shen, Woanruoh Lee, Hui Yi Lin, Chunming Shih, Lingling YangAbstract:Wogonin and Fisetin are flavonoids, which are widely distributed in plants. Our recent study demonstrated that, among seven structurally related flavonoids, wogonin and Fisetin showed the most potent apoptosis-inducing activities in human promyeloleukemic cells HL-60. In the present investigation, we performed molecular studies to assess the apoptotic effects of wogonin and Fisetin on hepatocellular carcinoma cells SK-HEP-1. Both wogonin and Fisetin showed dose-dependent cytotoxic effects on SK-HEP-1 cells, accompanied by DNA fragmentation. Microscopic observation under Giemsa staining showed that wogonin and Fisetin, at the dose of 80 µM, induced cellular swelling and the appearance of apoptotic bodies, characteristics of apoptosis, in SK-HEP-1 cells. Furthermore, flow cytometry analysis showed an increase of hypodiploid cells in wogonin- and Fisetin-treated SK-HEP-1 cells. These data demonstrated that wogonin and Fisetin were effective inducers of apoptosis in SK-HEP-1 cells. Treatment with an apoptosis-inducing concentration of wogonin or Fisetin caused induction of caspase 3/CPP32 activity, but not of caspase 1 activity. In addition, a caspase 3 inhibitor, Ac-DEVD-CHO, but not the caspase 1 inhibitor Ac-YVAD-CHO, reversed the cytotoxic effects of wogonin and Fisetin on SK-HEP-1 cells. Further, cleavage of caspase 3 substrates including poly(ADP-ribose) polymerase (PARP) and D4-GDI protein, and decrease of pro-caspase 3 protein were detected in wogonin- and Fisetin-treated SK-HEP-1 cells. Increase of p53 protein was associated with wogonin- and Fisetin-induced apoptosis; however, a p53-controlled gene, p21Waf/Cip-1, was only induced in wogonin- (not Fisetin-) treated SK-HEP-1 cells. Serum starvation elevated p21Waf/Cip-1 protein expression, and enhanced the apoptotic induction activity of wogonin (not fiseitn) in SK-HEP-1 cells. Our study has provided molecular evidence to demonstrate that wogonin and Fisetin had effective cytotoxic effects through apoptosis induction in hepatocellular carcinoma cells SK-HEP-1; activation of caspase 3 cascade, induction of p53 protein and alternative expression of p21Waf/Cip-1 protein were involved.
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wogonin and Fisetin induce apoptosis in human promyeloleukemic cells accompanied by a decrease of reactive oxygen species and activation of caspase 3 and ca2 dependent endonuclease
Biochemical Pharmacology, 2002Co-Authors: Woan Rouh Lee, Shingchuan Shen, Hui Yi Lin, Lingling Yang, Wen Chi Hou, Yenchou ChenAbstract:Abstract Seven structurally related flavonoids including luteolin, nobiletin, wogonin, baicalein, apigenin, myricetin and Fisetin were used to study their biological activities on the human leukemia cell line, HL-60. On MTT assay, wogonin, baicalein, apigenin, myricetin and Fisetin showed obvious cytotoxic effects on HL-60 cells, with wogonin and Fisetin being the most-potent apoptotic inducers among them. The cytotoxic effects of wogonin and Fisetin were accompanied by the dose- and time-dependent appearance of characteristics of apoptosis including DNA fragmentation, apoptotic bodies and the sub-G1 ratio. Treatment with an apoptosis-inducing concentration of wogonin or Fisetin causes rapid and transient induction of caspase 3/CPP32 activity, but not caspase 1 activity. Further, cleavage of poly(ADP-ribose) polymerase (PARP) and decrease of pro-caspase 3 protein were detected in wogonin- and Fisetin-treated HL-60 cells. An increase in the pro-apoptotic protein, bax, and a decrease in the anti-apoptotic protein, Mcl-1, were detected in Fisetin- and wogonin-treated HL-60 cells. However, Bcl-2, Bcl-XL, and Bad all remained unchanged in wogonin- and Fisetin-treated HL-60 cells. In vitro chromatin digestion revealed that endonuclease activity was profoundly enhanced in wogonin- and Fisetin-treated HL-60 cells, and the addition of ethylenediaminetetraacetic acid (EDTA) or ethyleneglycoltetraacetic acid (EGTA) into the reaction blocked endonuclease activation and at an optimum pH of 7.5. The caspase 3 inhibitor, Ac-DEVD-CHO, but not the caspase 1 inhibitor, Ac-YVAD-CHO, attenuated wogonin- and Fisetin-induced DNA ladders, PARP cleavage, and endonuclease activation. Pretreatment of HL-60 cells with N -acetyl-cysteine or catalase efficiently inhibited H 2 O 2 (200 μM)-induced apoptosis, but showed no inhibitory effect on wogonin- and Fisetin-induced DNA ladders, caspase 3 activation, or bax protein induction. Decrease in endogenous ROS production was detected in wogonin- and Fisetin-treated HL-60 cells by DCHF-DA assay. In conclusion, our experiments indicate that a decrease in intracellular peroxide level was involved in wogonin- and Fisetin-induced apoptosis; activation of caspase 3 and endonuclease, induction of bax protein and suppression of Mcl-1 protein were detected in the process.
Deeba N Syed - One of the best experts on this subject based on the ideXlab platform.
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Fisetin targets yb 1 rsk axis independent of its effect on erk signaling insights from in vitro and in vivo melanoma models
Scientific Reports, 2018Co-Authors: Mario Sechi, Hasan Mukhtar, Rahul K Lall, Saheed O Afolabi, Anant Singh, Dinesh C Joshi, S Y Chiu, Deeba N SyedAbstract:The anti-proliferative activity of dietary flavonoid Fisetin has been validated in various cancer models. Establishing its precise mechanism of action has proved somewhat challenging given the multiplicity of its targets. We demonstrated that YB-1 promotes epithelial-to-mesenchymal transition and its inhibition suppressed tumor cell proliferation and invasion. The p90 ribosomal S6 kinase (RSK), an important ERK effector, activates YB-1 to drive melanoma growth. We found that Fisetin treatment of monolayer/3-D melanoma cultures resulted in YB-1 dephosphorylation and reduced transcript levels. In parallel, Fisetin suppressed mesenchymal markers and matrix-metalloproteinases in melanoma cells. Data from cell-free/cell-based systems indicated that Fisetin inhibited RSK activity through binding to the kinase. Affinity studies for RSK isoforms evaluated stronger interaction for RSK2 than RSK1. Competition assays performed to monitor binding responses revealed that YB-1 and RSK2 do not compete, rather binding of Fisetin to RSK2 promotes its binding to YB-1. Fisetin suppressed YB-1/RSK signaling independent of its effect on ERK, and reduced MDR1 levels. Comparable efficacy of Fisetin and vemurafenib for inhibiting melanoma growth was noted albeit through divergent modulation of ERK. Our studies provide insight into additional modes of regulation through which Fisetin interferes with melanoma growth underscoring its potential therapeutic efficacy in disease progression.
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involvement of er stress and activation of apoptotic pathways in Fisetin induced cytotoxicity in human melanoma
Archives of Biochemistry and Biophysics, 2014Co-Authors: Deeba N Syed, Rahul K Lall, Jean Christopher Chamcheu, Omar Mohammad Haidar, Hasan MukhtarAbstract:Abstract The prognosis of malignant melanoma remains poor in spite of recent advances in therapeutic strategies for the deadly disease. Fisetin, a dietary flavonoid is currently being investigated for its growth inhibitory properties in various cancer models. We previously showed that Fisetin inhibited melanoma growth in vitro and in vivo. Here, we evaluated the molecular basis of Fisetin induced cytotoxicity in metastatic human melanoma cells. Fisetin treatment induced endoplasmic reticulum (ER) stress in highly aggressive A375 and 451Lu human melanoma cells, as revealed by up-regulation of ER stress markers including IRE1α, XBP1s, ATF4 and GRP78. Time course analysis indicated that the ER stress was associated with activation of the extrinsic and intrinsic apoptotic pathways. Fisetin treated 2-D melanoma cultures displayed autophagic response concomitant with induction of apoptosis. Prolonged treatment (16 days) with Fisetin in a 3-D reconstituted melanoma model resulted in inhibition of melanoma progression with significant apoptosis, as evidenced by increased staining of cleaved Caspase-3 in the treated constructs. However, no difference in the expression of autophagic marker LC-3 was noted between treated and control groups. Fisetin treatment to 2-D melanoma cultures resulted in phosphorylation and activation of the multifunctional AMP-activated protein kinase (AMPK) involved in the regulation of diverse cellular processes, including autophagy and apoptosis. Silencing of AMPK failed to prevent cell death indicating that Fisetin induced cytotoxicity is mediated through both AMPK-dependent and -independent mechanisms. Taken together, our studies confirm apoptosis as the primary mechanism through which Fisetin inhibits melanoma cell growth and that activation of both extrinsic and intrinsic pathways contributes to Fisetin induced cytotoxicity.
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Fisetin inhibits human melanoma cell growth through direct binding to p70s6k and mtor findings from 3 d melanoma skin equivalents and computational modeling
Biochemical Pharmacology, 2014Co-Authors: Deeba N Syed, Vaqar M Adhami, Mario Sechi, Rahul K Lall, Jean Christopher Chamcheu, Mohammad Imran Khan, Hasan MukhtarAbstract:The incidence of melanoma continues to rise. Inspite of treatment advances, the prognosis remains grim once the disease has metastasized, emphasizing the need to explore additional therapeutic strategies. One such approach is through the use of mechanism-based dietary intervention. We previously showed that the flavonoid Fisetin inhibits melanoma cell proliferation, in vitro and in vivo. Here, we studied Fisetin-mediated regulation of kinases involved in melanoma growth and progression. Time-course analysis in 3-D melanoma constructs that transitioned from radial to vertical growth showed that Fisetin treatment resulted in significant decrease in melanocytic lesions in contrast to untreated controls that showed large tumor nests and invading disseminated cells. Further studies in melanoma cultures and mouse xenografts showed that Fisetin-mediated growth inhibition was associated with dephosphorylation of AKT, mTOR and p70S6K proteins. In silico modeling indicated direct interaction of Fisetin with mTOR and p70S6K with favorable free energy values. These findings were validated by cell-free competition assays that established binding of Fisetin to p70S6K and mTOR while little affinity was detected with AKT. Kinase activity studies reflected similar trend with % inhibition observed for p70S6K and mTOR at lower doses than AKT. Our studies characterized, for the first time, the differential interactions of any botanical agent with kinases involved in melanoma growth and demonstrate that Fisetin inhibits mTOR and p70S6K through direct binding while the observed inhibitory effect of Fisetin on AKT is mediated indirectly, through targeting interrelated pathways.
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Fisetin a novel dietary flavonoid causes apoptosis and cell cycle arrest in human prostate cancer lncap cells
Carcinogenesis, 2008Co-Authors: Naghma Khan, Farrukh Afaq, Deeba N Syed, Hasan MukhtarAbstract:467 Prostate cancer (PCa) has become one of the most frequently diagnosed malignancies in the United States with similar trend in most Western countries. The substantial mortality and morbidity associated with PCa and its poor treatment options have led a surge to develop novel means for its prevention. The development of chemoprevention strategies against PCa would have huge impact, both medically and economically. Because of its ubiquity and disease burden, PCa is an attractive target for chemoprevention. Fisetin (3,39,49,7-tetrahydroxyflavone), is found in fruits andvegetables, such as strawberry, apple, persimmon, grape, onionand cucumber. The aim of this study was to determine the effect of Fisetin on cell-growth and apoptosis in human PCa cells. Treatment of Fisetin (10-60 μM; 48 h) was found to result in a decrease in the viability of LNCaP, CWR22Rυ1 and PC-3 cells but had only minimal effect on normal prostate epithelial PrEC cells as assessed by MTT assay. To decipher the mechanism of growth inhibition of Fisetin, we selected LNCaP cells for further studies. Treatment of LNCaP cells with Fisetin (10-60 μM; 48 h) also resulted in G1-phase arrest (as assessed by DNA cell cycle analysis). Fisetin also caused a marked decrease in the protein expression of cyclins D1, D2 and E and their activating partners cdks 2, 4 and 6 with concomitant induction of WAF1/p21 and KIP1/p27. Treatment of LNCaP cells with Fisetin also resulted in significant apoptosis in a dose-dependent fashion as determined by ELISA. A significant increase in the proteinexpression of Bax, Bak, Bad and Bid (proapoptotic) and decrease in Bcl-2 and BCl-xL (antiapoptotic) was observed on treatment with Fisetin in a dose-dependent fashion thus further confirming the induction of apoptotic process. Additionally, PARP cleavage analysis showed that the full size PARP (116 KD) protein was cleaved to yield an 85 KD fragment after treatment of cells with Fisetin. Fisetin treatment of cells also resulted in induction of mitochondrial release of cytochrome c into cytosol and downregulation of XIAP and upregulation of Smac/DIABLO. Treatment of cells with Fisetin also resulted in significant activation of caspase-3, -8 and -9. Pretreatment of cells with caspase inhibitor (Z-VAD-FMK) blocked Fisetin-induced activation of caspases as detected by immunoblot analysis and immunofluorescence staining. These data provide the first evidence that Fisetin could be developed as an agent against prostate cancer.