Fisetin, a natural flavonoid, targets chemoresistant human pancreatic cancer AsPC-1 cells through DR3-mediated inhibition of NF-kappaB.
Death receptors of the tumor necrosis factor (TNF) receptor super family have been implicated in constitutive activation of nuclear factor-kappa B (NF-kappaB) in pancreatic cancer (PaC) cells. In this study, we demonstrate that fisetin, a natural flavonoid, induces apoptosis and inhibits invasion of chemoresistant PaC AsPC-1 cells through suppression of DR3-mediated NF-kappaB activation. Fisetin treatment resulted in dose-dependent inhibition of PaC cell growth and cell proliferation with concomitant induction of apoptosis. A cDNA array analysis revealed that fisetin modulates expression of more than 20 genes at transcription level with maximum decrease observed in DR3 expression and a parallel increase observed in the expression levels of IkappaBalpha, an NF-kappaB inhibitor. Down-regulation of DR3 in PaC cells was found to down regulate activated pNF-kappaB/p65, pIkBalpha/beta kinases (pIKK's), MMP9 and XIAP that mostly impart chemoresistance in PaC. Immunoblotting and EMSA analysis showed a marked decrease in pNF-kappaB and NF-kappaB DNA binding activity, respectively, with modest decrease in NF-kappaB promoter activity and significant decrease in MMP9 promoter activity with fisetin treatment. Importantly, consistent with these findings, we further found that transient down-regulation of DR3 by RNA interference significantly augmented fisetin induced changes in cell proliferation, cell invasion and apoptosis paralleled with decrease in pNF-kappaB, pIKKalpha/beta, MMP9, XIAP and NF-kappaB DNA binding activity. Blocking of DR3 receptor with an extra cellular domain blocking antibody demonstrated similar effects. These data provide evidence that fisetin could provide a biological rationale for treatment of pancreatic cancer or as an adjuvant with conventional therapeutic regimens.
Murtaza
Imtiyaz
I
Department of Dermatology, University of Wisconsin, Madison, WI 53706, USA. imz007@rediffmail.com
Adhami
Vaqar Mustafa
VM
Hafeez
Bilal Bin
BB
Saleem
Mohammad
M
Mukhtar
Hasan
H
D017209:Apoptosis / Q000187:drug effects; D015153:Blotting, Western; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D019008:Drug Resistance, Neoplasm; D024202:Electrophoretic Mobility Shift Assay; D005419:Flavonoids / Q000494:pharmacology*; D044948:Flavonols; D020869:Gene Expression Profiling; D006801:Humans; D020672:I-kappa B Proteins / Q000378:metabolism; D020780:Matrix Metalloproteinase 9 / Q000378:metabolism; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D009361:Neoplasm Invasiveness; D020411:Oligonucleotide Array Sequence Analysis; D010190:Pancreatic Neoplasms / Q000188:drug therapy* / Q000378:metabolism* / Q000473:pathology; D011401:Promoter Regions, Genetic; D034741:RNA, Small Interfering / Q000494:pharmacology; D053219:Receptors, Tumor Necrosis Factor, Member 25 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D051636:X-Linked Inhibitor of Apoptosis Protein / Q000378:metabolism
D005419:Flavonoids; D044948:Flavonols; D020672:I-kappa B Proteins; D016328:NF-kappa B; C000606204:NFKBIA protein, human; D034741:RNA, Small Interfering; D053219:Receptors, Tumor Necrosis Factor, Member 25; C506033:TNFRSF25 protein, human; D051636:X-Linked Inhibitor of Apoptosis Protein; C496866:XIAP protein, human; D000072000:NF-KappaB Inhibitor alpha; D020780:Matrix Metalloproteinase 9; C017875:fisetin