Investigation of the protein alkylation sites of the STAT3:STAT3 inhibitor Stattic by mass spectrometry

Heidelberger, Sibylle; Zinzalla, Giovanna; Antonow, Dyeison; Essex, Samantha; Basu, Bristi; Palmer, Jonathan M.; Husby, Jarmila; Jackson, Paul J.; Rahman, Khondaker Miraz; Wilderspin, Andrew F.; Zloh, Mire; Thurston, David E.

doi:10.1016/j.bmcl.2013.05.066

Cited by 46 publications

(51 citation statements)

References 26 publications

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“…Stattic inhibits STAT3 activity via cysteine alkylation [21] and despite a cysteine residue residing in the SH2 domain (Cys687), we did not detect any disruption of STAT3-peptide binding with stattic up to 600μM (Figure 3A). S3I-1757 has been reported as an SH2 domain inhibitor of STAT3 and our results support that S3I-1757 can bind the SH2 domain and prevent protein-protein interactions at this interface, with an IC 50 of 7.39 ± 0.95μM in the FP assay (Figure 3B).…”

Section: Resultsmentioning

confidence: 82%

“…Due to the difficulty in developing small molecules capable of disrupting protein-protein interactions over a large surface area, while maintaining drug-like properties, there are a limited number of SH2 domain inhibitors that have reached pre-clinical and clinical trials [10, 15, 17, 18]. Stattic was one of the first small molecule inhibitors of STAT3 to be identified [10, 19], and it was initially believed to be a SH2 domain inhibitor; however, it was later determined that its inhibitory activity was due to modification of the protein through covalent interactions with cysteine residues, many of which reside in the DNA-binding domain (DBD) of STAT3 (Figures 1B and 1C) [20, 21]. Therefore, there has been interest in developing small molecules that disrupt STAT3 transcriptional activity by targeting the DBD of STAT3 (Figure 1) [22–24].…”

Section: Introductionmentioning

confidence: 99%

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Evaluation of quantitative assays for the identification of direct signal transducer and activator of transcription 3 (STAT3) inhibitors

et al. 2016

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Section: Resultsmentioning

confidence: 82%

Section: Introductionmentioning

confidence: 99%

Evaluation of quantitative assays for the identification of direct signal transducer and activator of transcription 3 (STAT3) inhibitors

et al. 2016

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“…Buettner et al (23) demonstrated that the small molecule STAT3 inhibitor C48 alkylates Cys-468 in the STAT3 DNA binding domain, although at a high compound concentration. Stattic, a STAT3 inhibitor that has been shown to prevent STAT3 dimerization and phosphorylation (29), was shown to alkylate four cysteine residues in unphosphorylated STAT3 (Cys-251, Cys-259, Cys-367, and Cys-426) (24). Furthermore, it has been shown that the transcriptional activity of STAT3 is sensitive to changes in cellular redox conditions as treatment of STAT3 with hydrogen peroxide leads to covalent modifications (oxidation) of specific cysteine residues (25).…”

Section: Discussionmentioning

confidence: 99%

“…With the reactive potential of galiellalactone toward biological nucleophiles in consideration, we were interested in investigating whether galiellalactone can alkylate STAT3 and thereby inhibit the DNA binding as there is precedence that direct covalent modification of STAT3 with small molecules (23,24) or through cysteine oxidation (25) can block the transcriptional activity of STAT3. The aim of the present study is to elucidate in more detail the mechanism of action of galiellalactone using human prostate cancer cells as a model.…”

mentioning

confidence: 99%

Galiellalactone Is a Direct Inhibitor of the Transcription Factor STAT3 in Prostate Cancer Cells

Don‐Doncow

Escobar

Johansson

et al. 2014

Journal of Biological Chemistry

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Background: STAT3 is constitutively active in castration-resistant prostate cancer and the fungal metabolite galiellalactone inhibits STAT3 signaling. Results: Galiellalactone binds covalently to one or more cysteines in STAT3 and prevents STAT3 binding to DNA. Conclusion: Galiellalactone inhibits STAT3 signaling by binding directly to STAT3. Significance: Galiellalactone is a promising direct STAT3 inhibitor for treatment of castration-resistant prostate cancer.

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“…Therefore, STAT3 may be involved in the increase in EPOR expression. The specificity of Stattic has been challenged [44]. However, Stattic has been used as a specific STAT3 inhibitor in numerous studies [45,46].…”

Section: Discussionmentioning

confidence: 99%

Investigation of the pronounced erythropoietin-induced reduction in hyperglycemia in type 1-like diabetic rats

Kuo

Cheng

et al. 2018

Endocr J

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Abstract. Erythropoietin (EPO) is known to stimulate erythropoiesis after binding with its specific receptor. In clinics, EPO is widely used in hemodialyzed patients with diabetes. However, changes in the expression of the erythropoietin receptor (EPOR) under diabetic conditions are still unclear. Therefore, we investigated EPOR expression both in vivo and in vitro. Streptozotocin-induced type 1-like diabetic rats (STZ rats) were used to evaluate the blood glucose-lowering effects of EPO. The expression and activity of the transducer and activator of transcription 3 (STAT3), the potential signaling molecule, was investigated in cultured rat skeletal myoblast (L6) cells incubated in high-glucose (HG) medium to mimic the in vivo changes. The EPO-induced reduction in hyperglycemia was more pronounced in diabetic rats. The increased EPOR expression in the soleus muscle of diabetic rats was reversed by the reduction in hyperglycemia. Glucose uptake was also increased in highglucose (HG)-treated L6 cells. Western blotting results indicated that the EPO-induced hyperglycemic activity was enhanced mainly through an increase in EPOR expression. Increased EPOR expression was associated with the enhanced nuclear expression of STAT3 in HG-exposed L6 cells. In addition, treatment with siRNA specific to STAT3 reversed the increased expression of EPOR observed in these cells. Treatment with Stattic at a dose sufficient to inhibit STAT3 reduced the expression level of EPOR in STZ rats. In conclusion, the increased expression of EPOR by hyperglycemia is mainly associated with an augmented expression of nuclear STAT3, which was identified both in vivo and in vitro in the present study. STZ rats, Streptozotocin-induced type 1-like diabetic rats; STAT3, transducer and activator of transcription 3; HG, high glucose; 2-NBDG, 2-(N-(7-Nitrobenz-2-oxa-1,3-diazol-4-yl)Amino)-2-Deoxyglucose; PBS, phosphate buffer solution; RIPA buffer, radioimmuno-precipitation assay buffer; HMBS, hydroxymethylbilane synthase

show abstract

Investigation of the protein alkylation sites of the STAT3:STAT3 inhibitor Stattic by mass spectrometry

Cited by 46 publications

References 26 publications

Evaluation of quantitative assays for the identification of direct signal transducer and activator of transcription 3 (STAT3) inhibitors

Evaluation of quantitative assays for the identification of direct signal transducer and activator of transcription 3 (STAT3) inhibitors

Galiellalactone Is a Direct Inhibitor of the Transcription Factor STAT3 in Prostate Cancer Cells

Investigation of the pronounced erythropoietin-induced reduction in hyperglycemia in type 1-like diabetic rats

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