title: Flavopiridol disrupts STAT3/DNA interactions, attenuates STAT3-directed transcription, and combines with the Jak kinase inhibitor AG490 to achieve cytotoxic synergy.
abstract: Up-regulated signal transducers and activators of transcription (STAT)-mediated signaling is believed to contribute to the pathogenesis of a variety of solid and hematologic cancers. Consequently, inhibition of STAT-mediated signaling has recently been proposed as a potential new therapeutic approach to the treatment of cancers. Having shown previously that the pan-cyclin-dependent kinase inhibitor flavopiridol binds to DNA and seems to kill cancer cells via that process in some circumstances, we evaluated the hypothesis that flavopiridol might consequently disrupt STAT3/DNA interactions, attenuate STAT3-directed transcription, and down-regulate STAT3 downstream polypeptides, including the antiapoptotic polypeptide Mcl-1. SDS-PAGE/immunoblotting and reverse transcription-PCR were used to assess RNA and polypeptide levels, respectively. DNA cellulose affinity chromatography and a nuclear elution assay were used to evaluate the ability of flavopiridol to disrupt STAT3/DNA interactions. A STAT3 luciferase reporter assay was used to examine the ability of flavopiridol to attenuate STAT3-directed transcription. Colony-forming assays were used to assess cytotoxic synergy between flavopiridol and AG490. Flavopiridol was found to (a) disrupt STAT3/DNA interactions (DNA cellulose affinity chromatography and nuclear elution assay), (b) attenuate STAT3-directed transcription (STAT3 luciferase reporter assay), and (c) down-regulate the STAT3 downstream antiapoptotic polypeptide Mcl-1 at the transcriptional level (reverse transcription-PCR and SDS-PAGE/immunoblotting). Furthermore, flavopiridol, but not the microtubule inhibitor paclitaxel, could be combined with the STAT3 pathway inhibitor AG490 to achieve cytotoxic synergy in A549 human non-small cell lung cancer cells. Collectively, these data suggest that flavopiridol can attenuate STAT3-directed transcription in a targeted fashion and may therefore be exploitable clinically in the development of chemotherapy regimens combining flavopiridol and other inhibitors of STAT3 signaling pathways.
authors.affiliation: Division of Developmental Oncology Research, Mayo Clinic, 200 First Street Southwest, Rochester, MN 55905, USA.
authors.forename: Yean Kit
authors.initials: YK
authors.lastname: Lee
authors.forename: Crescent R
authors.initials: CR
authors.lastname: Isham
authors.forename: Scott H
authors.initials: SH
authors.lastname: Kaufman
authors.forename: Keith C
authors.initials: KC
authors.lastname: Bible
mesh_terms: D000971:Antineoplastic Combined Chemotherapy Protocols; D002289:Carcinoma, Non-Small-Cell Lung / Q000188:drug therapy / Q000473:pathology; D004247:DNA / Q000378:metabolism; D015536:Down-Regulation; D004354:Drug Screening Assays, Antitumor; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005419:Flavonoids / Q000008:administration & dosage / Q000494:pharmacology*; D006801:Humans; D053613:Janus Kinase 1; D008175:Lung Neoplasms / Q000188:drug therapy / Q000473:pathology; D064549:Myeloid Cell Leukemia Sequence 1 Protein; D009363:Neoplasm Proteins / Q000187:drug effects / Q000235:genetics; D010750:Phosphoproteins / Q000187:drug effects / Q000378:metabolism; D010880:Piperidines / Q000008:administration & dosage / Q000494:pharmacology*; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000187:drug effects / Q000235:genetics; D012319:RNA Polymerase II / Q000187:drug effects / Q000378:metabolism; D050796:STAT3 Transcription Factor / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured; D020032:Tyrphostins / Q000494:pharmacology*
chemical_list: D004791:Enzyme Inhibitors; D005419:Flavonoids; D064549:Myeloid Cell Leukemia Sequence 1 Protein; D009363:Neoplasm Proteins; D010750:Phosphoproteins; D010880:Piperidines; D019253:Proto-Oncogene Proteins c-bcl-2; D050796:STAT3 Transcription Factor; D020032:Tyrphostins; C095512:alpha-cyano-(3,4-dihydroxy)-N-benzylcinnamide; C077990:alvocidib; D004247:DNA; D011505:Protein-Tyrosine Kinases; C507904:JAK1 protein, human; D053613:Janus Kinase 1; D012319:RNA Polymerase II