Summary of probe development efforts to identify inhibitors of signal transducer and activator of transcription 3 (STAT3)
Name: Summary of probe development efforts to identify inhibitors of signal transducer and activator of transcription 3 (STAT3) ..more
Source (MLSCN Center Name): The Scripps Research Institute Molecular Screening Center
Center Affiliation: The Scripps Research Institute (TSRI)
Assay Provider: David Frank, Dana Farber Cancer Institute
Network: Molecular Library Screening Center Network (MLSCN)
Grant Proposal Number: 1 X01 MH079826-01
Grant Proposal PI: David Frank, Dana Farber Cancer Institute
External Assay ID: STAT3_INH_LEADS_SUMMARY
Name: Summary of probe development efforts to identify inhibitors of signal transducer and activator of transcription 3 (STAT3)
The signal transducer and activator of transcription (STAT) family of transcription factors transduce signals from a variety of extracellular stimuli and are important mediators of inflammation, cell survival, differentiation, and proliferation (1, 2). STATs are activated in response to growth factors, cytokines, and G-CSF binding to cell surface receptor tyrosine kinases (1-3). In resting cells STATs are inactive in the cytoplasm. In response to stimuli, STATs are phosphorylated by the Janus-activated kinases (Jaks), which induces STAT dimerization and nuclear translocation, where STATs bind to specific enhancer elements in target genes (2). Although structurally similar, the seven STAT family member (STATs 1, 2, 3, 4, 5a, 5b, and 6) possess diverse biological roles (2). For example, STAT1 activation is pro-inflammatory and anti-proliferative, while STAT3 activation is anti-inflammatory and pro-apoptotic (2). STAT1 is largely responsible for mediating the effects of IFN- γ, while STAT3 is predominantly involved in IL-6 signaling (4). STAT1 induces expression of genes that inhibit the cell cycle, and thus STAT1 is considered to have tumor suppressor properties (5). Studies show that STAT3 is activated in a majority of breast and prostate cancers, and that STAT3 inhibition using RNA interference or a dominant negative leads to reduced cell proliferation, survival, and wound healing (1, 4, 6). Blocking STAT3 interaction with the epidermal growth factor receptor (EGFR) using peptide aptamers has been shown to reduce tumor growth (7). Due to the diverse roles and potent phenotypes associated with STAT signaling, the identification of selective modulators of STAT3 activity may lead to pharmacological tools for cancer, wound healing, and inflammatory diseases.
Summary of Probe Development Effort:
Following primary HTS in singlicate to identify STAT3 inhibitors (AID 862), confirmation of hit activity in triplicate (AID 1265), counterscreening in triplicate against NFkB (AID 1308) and STAT3 (AID 1317) to determine selectivity, followed by titration assays to determine compound potency (AID 1399) and selectivity (AID 1411), a compound (SID 24825594 (liquid)) belonging to the thienopyrimidine scaffold was identified as a candidate for probe development (AID 2078). This compound does not share structural similarities with the known STAT3 inhibitor and state of the art, nifuroxazide (SID 11532872). Analogs belonging to the thienopyrimidine scaffold were purchased in powder form or re-ordered from the MLSMR in liquid form and tested in dose response assays against both STAT3 and STAT1, as well as additional counterscreening assays to determine cytotoxicity. SID 87349854/SID 87326012 (powder) confirmed activity and is claimed as a potent, selective, and non-toxic STAT3 inhibitor probe.
The above probe development effort resulted in the identification of one probe. A probe report has been published (http://mlpcn.florida.scripps.edu/index.php/probes/probe-reports.html). A probe report for SID 87349854/SID 87326012 can be found in the Molecular Libraries Bookshelf (PubMed Books) (http://www.ncbi.nlm.nih.gov/books) under ML116.
1. Alvarez JV, Febbo PG, Ramaswamy S, Loda M, Richardson A, Frank DA. Identification of a genetic signature of activated signal transducer and activator of transcription 3 in human tumors. Cancer Res. 2005 Jun 15;65(12):5054-62.
2. Schindler C, Levy DE, Decker T. JAK-STAT signaling: from interferons to cytokines. J Biol Chem. 2007 Jul 13;282(28):20059-63.
3. Germain D, Frank DA. Targeting the cytoplasmic and nuclear functions of signal transducers and activators of transcription 3 for cancer therapy. Clin Cancer Res. 2007 Oct 1;13(19):5665-9.
4. Levy DE, Darnell JE Jr. Stats: transcriptional control and biological impact. Nat Rev Mol Cell Biol. 2002 Sep;3(9):651-62.
5. Battle TE, Wierda WG, Rassenti LZ, Zahrieh D, Neuberg D, Kipps TJ, Frank DA. In vivo activation of signal transducer and activator of transcription 1 after CD154 gene therapy for chronic lymphocytic leukemia is associated with clinical and immunologic response. Clin Cancer Res. 2003 Jun;9(6):2166-72.
6. Takeda, K. Takeda K, Kaisho T, Yoshida N, Takeda J, Kishimoto T, Akira S.1998. Stat3 activation is responsible for IL-6-dependent T cell proliferation through preventing apoptosis: generation and characterization of T cell- specific Stat3-deficient mice. J. Immunol. 161:4652-4660.
7. Buerger C, Nagel-Wolfrum K, Kunz C, Wittig I, Butz K, Hoppe-Seyler F, Groner B. Sequence-specific peptide aptamers, interacting with the intracellular domain of the epidermal growth factor receptor, interfere with Stat3 activation and inhibit the growth of tumor cells. J Biol Chem. 2003 Sep 26;278(39):37610-21.
Summary AID, STAT3, signal transducer and activator of transcription 3, acute-phase response factor, APRF, inhibitor, inhibition, U3A, transcription factor, luciferase, luminescence, reporter, dose response, counterscreen, 1536, HTS, assay, Scripps, Scripps Florida, Scripps Research Institute Molecular Screening Center, SRIMSC, Molecular Libraries Probe Production Centers Network, MLPCN.
Please see Related Bioassays for protocols performed in this probe development effort.
Categorized Comment - additional comments and annotations
From MLP Probe Report:
Probe count: 1
MLP Probe ML# for probe 1: ML116
PubChem Substance ID (SID) for probe 1: 24825594,87326012
PubChem Compound ID (CID) for probe 1: 2100018
Probe type for probe 1: Modulator
IC50/EC50 (nM) for probe 1: 4200
Target for probe 1: STAT3 (gi: 13272532)
Disease relevance for probe 1: Cancer
Anti-target for probe 1: STAT1
Fold selectivity for probe 1: STAT3 selective
NCBI Book chapter link for probe 1: http://www.ncbi.nlm.nih.gov/books/NBK56232/ (ID: 2509544)
Grant number for probe 1: MH079826-01
NCBI Book chapter title for probe 1: Modulators of STAT Transcription Factors for the Targeted Therapy of Cancer (STAT3 Inhibitors)
Data Table (Concise)