1. Inhibition of STAT3 by Anticancer Drug Bendamustine
- Author
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Naohisa Ogo, Yukie Inoue, Akira Asai, Kazunori Iwamoto, Kyoko Taguchi, Yutaka Uehara, Kenji Matsuno, and Daisuke Muraoka
- Subjects
0301 basic medicine ,Cell signaling ,lcsh:Medicine ,Protein domains ,Gene Expression ,Peptide ,Plasma protein binding ,Signal transduction ,Biochemistry ,Fluorophotometry ,law.invention ,SH2 domain ,Binding Analysis ,0302 clinical medicine ,Spectrum Analysis Techniques ,law ,Fluorescence Resonance Energy Transfer ,Bendamustine Hydrochloride ,Amino Acids ,lcsh:Science ,chemistry.chemical_classification ,Multidisciplinary ,Alanine ,Organic Compounds ,Precipitation Techniques ,Chemistry ,STAT signaling ,Spectrophotometry ,030220 oncology & carcinogenesis ,Biotinylation ,Physical Sciences ,Recombinant DNA ,medicine.symptom ,Cell Binding Assay ,Research Article ,Protein Binding ,STAT3 Transcription Factor ,Cell biology ,DNA binding assay ,Biology ,Research and Analysis Methods ,src Homology Domains ,03 medical and health sciences ,Structure-Activity Relationship ,Cell Line, Tumor ,medicine ,Genetics ,Structure–activity relationship ,Sulfur Containing Amino Acids ,Immunoprecipitation ,Humans ,Point Mutation ,Cysteine ,Binding site ,Phosphotyrosine ,Antineoplastic Agents, Alkylating ,Chemical Characterization ,Binding Sites ,lcsh:R ,Organic Chemistry ,Chemical Compounds ,Biology and Life Sciences ,Proteins ,Molecular biology ,030104 developmental biology ,Mechanism of action ,chemistry ,Cell culture ,lcsh:Q ,Drug Screening Assays, Antitumor ,Peptides - Abstract
Bendamustine (BENDA), which bears the bis(2-chloroethyl)amino moiety, is an alkylating agent that stops the growth of cancer cells by binding to DNA and interfering with its replication. However, the mechanism of action underlying its excellent clinical efficacy remains unclear. In this work, we report that BENDA inhibits signal transducer and activator of transcription 3 (STAT3). In an AlphaScreen-based biochemical assay using recombinant human STAT3, binding of STAT3-Src homology 2 (SH2) to the phosphotyrosine (pTyr, pY) peptide was inhibited by BENDA but not by the inactive metabolite dihydroxy bendamustine (HP2). When a single point mutation of C550A or C712A was introduced into recombinant human STAT3, its sensitivity to BENDA was substantially reduced, suggesting that these cysteine residues are important for BENDA to inhibit STAT3. Furthermore, BENDA suppressed the function of cellular STAT3 as a transcriptional activator in a human breast cancer cell line, MDA-MB-468, with constitutively activated STAT3. A competitive pull-down assay using biotinylated BENDA (Bio-BENDA) revealed that BENDA bound tightly to cellular STAT3, presumably through covalent bonds. Therefore, our results suggest that the anticancer effects of BENDA may be associated, at least in part, with its inhibitory effect on the SH2 domain of STAT3.
- Published
- 2017