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<script type="text/javascript" src="/corehtml/pmc/jatsreader/ptpmc_3.22/js/jr.boots.min.js"> </script><title>Modulators of STAT Transcription Factors for the Targeted Therapy of Cancer (STAT3 Activators) - Probe Reports from the NIH Molecular Libraries Program - NCBI Bookshelf</title>
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<meta name="citation_inbook_title" content="Probe Reports from the NIH Molecular Libraries Program [Internet]">
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<meta name="citation_title" content="Modulators of STAT Transcription Factors for the Targeted Therapy of Cancer (STAT3 Activators)">
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<meta name="citation_publisher" content="National Center for Biotechnology Information (US)">
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<meta name="citation_date" content="2010/12/16">
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<meta name="citation_author" content="F Madoux">
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<meta name="citation_author" content="M Koenig">
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<meta name="citation_author" content="E Nelson">
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<meta name="citation_author" content="S Chowdhury">
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<meta name="citation_author" content="M Cameron">
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<meta name="citation_author" content="B Mercer">
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<meta name="citation_author" content="W Roush">
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<meta name="citation_author" content="D Frank">
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<meta name="citation_author" content="P Hodder">
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<meta name="citation_pmid" content="21433395">
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<meta name="citation_fulltext_html_url" content="https://www.ncbi.nlm.nih.gov/books/NBK51964/">
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<meta name="DC.Title" content="Modulators of STAT Transcription Factors for the Targeted Therapy of Cancer (STAT3 Activators)">
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<meta name="DC.Type" content="Text">
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<meta name="DC.Publisher" content="National Center for Biotechnology Information (US)">
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<meta name="DC.Contributor" content="F Madoux">
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<meta name="DC.Contributor" content="M Koenig">
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<meta name="DC.Contributor" content="E Nelson">
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<meta name="DC.Contributor" content="S Chowdhury">
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<meta name="DC.Contributor" content="M Cameron">
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<meta name="DC.Contributor" content="B Mercer">
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<meta name="DC.Contributor" content="W Roush">
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<meta name="DC.Contributor" content="D Frank">
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<meta name="DC.Contributor" content="P Hodder">
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<meta name="DC.Date" content="2010/12/16">
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<meta name="description" content="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. STATs are activated in response to growth factors, cytokines, and G-CSF binding to cell surface receptor tyrosine kinases. Although structurally similar, the seven STAT family members possess diverse biological roles. For example, STAT1 activation is pro-inflammatory and anti-proliferative, while STAT3 activation is anti-inflammatory and pro-apoptotic. 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. Further, blocking STAT3 interaction with the epidermal growth factor receptor (EGFR) using peptide aptamers has been shown to reduce tumor growth. Due to the diverse roles and potent phenotypes associated with STAT signaling, the identification of selective potentiators of STAT3 activity may lead to pharmacological tools for cancer, wound healing, and inflammatory diseases. In response to the chief goal of this project, the identified small molecular probe ML115 (CID-619100 is a potent and selective activator of transcription 3 (STAT3), with a low nanomolar EC50 for STAT3, and is inactive against the related STAT1 and NFκB anti-targets. QPCR experiments further confirm that the probe upregulates transcription of the oncogene BCL3. In addition, the probe does not exhibit cytotoxicity in the parental cell lines used for the STAT3 assay (HT-1080) and STAT1 assay (NIH-3T3).">
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<meta name="og:title" content="Modulators of STAT Transcription Factors for the Targeted Therapy of Cancer (STAT3 Activators)">
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<meta name="og:description" content="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. STATs are activated in response to growth factors, cytokines, and G-CSF binding to cell surface receptor tyrosine kinases. Although structurally similar, the seven STAT family members possess diverse biological roles. For example, STAT1 activation is pro-inflammatory and anti-proliferative, while STAT3 activation is anti-inflammatory and pro-apoptotic. 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. Further, blocking STAT3 interaction with the epidermal growth factor receptor (EGFR) using peptide aptamers has been shown to reduce tumor growth. Due to the diverse roles and potent phenotypes associated with STAT signaling, the identification of selective potentiators of STAT3 activity may lead to pharmacological tools for cancer, wound healing, and inflammatory diseases. In response to the chief goal of this project, the identified small molecular probe ML115 (CID-619100 is a potent and selective activator of transcription 3 (STAT3), with a low nanomolar EC50 for STAT3, and is inactive against the related STAT1 and NFκB anti-targets. QPCR experiments further confirm that the probe upregulates transcription of the oncogene BCL3. In addition, the probe does not exhibit cytotoxicity in the parental cell lines used for the STAT3 assay (HT-1080) and STAT1 assay (NIH-3T3).">
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id="jr-fip-info-p"><a id="jr-fip-prev" class="wsprkl btn" title="Jump to previuos match">◀</a><button id="jr-fip-matches">no matches yet</button><a id="jr-fip-next" class="wsprkl btn" title="Jump to next match">▶</a></nav></nav></div><div id="jr-epub-interstitial" class="hidden"></div><div id="jr-content"><article data-type="main"><div class="main-content lit-style" itemscope="itemscope" itemtype="http://schema.org/CreativeWork"><div class="meta-content fm-sec"><div class="fm-sec"><h1 id="_NBK51964_"><span class="title" itemprop="name">Modulators of STAT Transcription Factors for the Targeted Therapy of Cancer (STAT3 Activators)</span></h1><p class="contribs">Madoux F, Koenig M, Nelson E, et al.</p><p class="fm-aai"><a href="#_NBK51964_pubdet_">Publication Details</a></p></div></div><div class="jig-ncbiinpagenav body-content whole_rhythm" data-jigconfig="allHeadingLevels: ['h2'],smoothScroll: false" itemprop="text"><div id="_abs_rndgid_" itemprop="description"><p>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. STATs are activated in response to growth factors, cytokines, and G-CSF binding to cell surface receptor tyrosine kinases. Although structurally similar, the seven STAT family members possess diverse biological roles. For example, STAT1 activation is pro-inflammatory and anti-proliferative, while STAT3 activation is anti-inflammatory and pro-apoptotic. 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. Further, blocking STAT3 interaction with the epidermal growth factor receptor (EGFR) using peptide aptamers has been shown to reduce tumor growth. Due to the diverse roles and potent phenotypes associated with STAT signaling, the identification of selective potentiators of STAT3 activity may lead to pharmacological tools for cancer, wound healing, and inflammatory diseases. In response to the chief goal of this project, the identified small molecular probe ML115 (CID-619100 is a potent and selective activator of transcription 3 (STAT3), with a low nanomolar EC50 for STAT3, and is inactive against the related STAT1 and NFκB anti-targets. QPCR experiments further confirm that the probe upregulates transcription of the oncogene BCL3. In addition, the probe does not exhibit cytotoxicity in the parental cell lines used for the STAT3 assay (HT-1080) and STAT1 assay (NIH-3T3).</p></div><div class="h2"></div><p><b>Assigned Assay Grant #:</b> 1 X01 MH079826-01</p><p><b>Screening Center Name & PI:</b> Scripps Research Institute Molecular Screening Center (SRIMSC); Hugh Rosen</p><p><b>Chemistry Center Name & PI:</b> SRIMSC; Hugh Rosen</p><p><b>Assay Submitter & Institution:</b> David Frank, Dana Farber Cancer Institute</p><p><b>Summary Bioassay Identifier (AID):</b>
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<a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/1805" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-1805</a></p><div id="a_probestruct"><h2 id="_a_probestruct_">Probe Structure & Characteristics: <a href="/pcsubstance/?term=ML115[synonym]" ref="pagearea=body&targetsite=entrez&targetcat=term&targettype=pubchem">ML115</a></h2><div id="ml115.fu1" class="figure"><div class="graphic"><img src="/books/NBK51964/bin/ml115fu1.jpg" alt="Image ml115fu1" /></div></div><div class="iconblock whole_rhythm clearfix ten_col table-wrap" id="figml115tu1"><a href="/books/NBK51964/table/ml115.tu1/?report=objectonly" target="object" title="Table" class="img_link icnblk_img figpopup" rid-figpopup="figml115tu1" rid-ob="figobml115tu1"><img class="small-thumb" src="/books/NBK51964/table/ml115.tu1/?report=thumb" src-large="/books/NBK51964/table/ml115.tu1/?report=previmg" alt="STAT3 Activator Probe Information (Isoxazole Carboxamide Scaffold; MLS 000522376)." /></a><div class="icnblk_cntnt"><h4 id="ml115.tu1"><a href="/books/NBK51964/table/ml115.tu1/?report=objectonly" target="object" rid-ob="figobml115tu1">Table</a></h4><p class="float-caption no_bottom_margin">STAT3 Activator Probe Information (Isoxazole Carboxamide Scaffold; MLS 000522376). </p></div></div></div><div id="ml115.s1"><h2 id="_ml115_s1_">Recommendations for scientific use of the probe (<a href="/pcsubstance/?term=ML115[synonym]" ref="pagearea=body&targetsite=entrez&targetcat=term&targettype=pubchem">ML115</a>)</h2><div id="ml115.introa"><h3>Current state of the art limitations addressed by the probe</h3><p>As prescribed in the CPDP, the chief goal for this probe development project was to find a selective activator the signal transducer and activator of transcription 3 (STAT3). In response to this goal, the probe compound <a href="/pcsubstance/?term=ML115[synonym]" ref="pagearea=body&targetsite=entrez&targetcat=term&targettype=pubchem">ML115</a> (CID6619100), belonging to the isoxazole carboxamide scaffold, is claimed as a potent and selective activator of STAT3. This probe has a low nanomolar EC<sub>50</sub> for STAT3 and is inactive against the related STAT1 and NFκB anti-targets. As expected for a useful STAT3 activator, QPCR experiments (run by the assay provider) confirm that the probe upregulates transcription of the oncogene BCL3. In addition, the probe does not exhibit cytotoxicity in the parental cell lines used for the STAT3 assay (HT-1080) and STAT1 assay (NIH-3T3). In contrast, prior art Compound 1 is a non-selective xanthocillin reported to induce phosphorylation of STAT3, Jak2, and STAT5 in a human leukemia cell line [<a class="bibr" href="#ml115.r1" rid="ml115.r1">1</a>]. As a potent EGFR tyrosine kinase inhibitor, prior art Compound 56 (CID2857) demonstrated potency in the STAT3 activator assay; however it was equally potent in the STAT1 activator assay.</p><p>Probe <a href="/pcsubstance/?term=ML115[synonym]" ref="pagearea=body&targetsite=entrez&targetcat=term&targettype=pubchem">ML115</a> was identified from the STAT3 uHTS campaign. It was selected as a starting point for medicinal chemistry because it increased expression of BCL3, a known STAT3-dependent oncogene. It was selected as a probe since analogs that were generated during the probe development effort were either less potent against STAT3 (analogs 1, 3 and 4), or did not significantly increase BCL3 transcription (e.g., analog 2). This compound does not share structural similarities with the known STAT3 activators Compound 1 and 56. The HTS campaign directly yielded several very potent compounds belonging to the isoxazole carboxamide scaffold: <a href="https://pubchem.ncbi.nlm.nih.gov/substance/14735210" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">SID-14735210</a>, 7965603, and 14733223. All three compounds were re-synthesized to confirm the activity. <a href="https://pubchem.ncbi.nlm.nih.gov/substance/14735210" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">SID-14735210</a> was found to be highly selective and it was active in the Assay Provider’s qPCR assay. Analogs 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. In addition, several analogs were purchased or synthesized to establish a more detailed SAR. Following testing of these analogs and initial lead compounds in several cell-based studies by the assay provider, <a href="https://pubchem.ncbi.nlm.nih.gov/substance/14735210" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">SID-14735210</a> (CID6619100) exhibited the characteristics of the desired probe and was assigned a probe number (<a href="/pcsubstance/?term=ML115[synonym]" ref="pagearea=body&targetsite=entrez&targetcat=term&targettype=pubchem">ML115</a>). This novel probe compound reported here acts to enhance STAT3 potentiation/activation mediated by exposure to the known inflammatory cytokine, interleukin-6 (IL-6). This probe acts in part by enhancing the transcriptional activity of the STAT3 promoter. This activity is restricted to the STAT3 signaling pathway, as no activation of the STAT1 or NFκ B pathways was detected.</p></div><div id="ml115.introb"><h3>What the probe will be used for</h3><p>Taken together, the results of the screening campaign, assay provider studies, and various probe development assays described herein validate the selection of CID6619100 as a potent and selective STAT3 activator probe with cell-based activity. In the light that the prior art STAT3 activators have off-target activity, the probe identified here is useful for oncologic research aiming to elucidate the STAT3 signaling pathway, with minimal effect on related STAT1 and NFκB pathways.</p><p>The maintenance of pluripotency in embryonic stem cells has been shown to be dependent on the continued activation of STAT3, at least in murine systems. This likely reflects the fact that among STAT3 target genes are genes that promote self-renewal such as <i>KLF4</i> and <i>BCL6</i>. Thus, a small molecule activator of STAT3 may be useful in maintaining embryonic stem cells in an undifferentiated state without the need for feeder cells or other exogenous biomolecules.</p><p>The ability of a small molecule to specifically enhance STAT3-dependent gene expression raises a number of mechanistic questions that such a probe could help address. For example, this finding suggests that there are unique mediators of STAT3-dependent gene expression (but not STAT1- or NF-kB-dependent gene expression) that can be modulated pharmacologically. Identifying the cellular target(s) of this probe thus provides a unique opportunity to delineate these specific transcriptional mediators that have heretofore been undiscovered. These transcriptional paradigms offer intriguing and complex biological questions that the current probe can be used to investigate.</p></div><div id="ml115.introc"><h3>Expected end-users of the probe in the research community</h3><p>Probe <a href="/pcsubstance/?term=ML115[synonym]" ref="pagearea=body&targetsite=entrez&targetcat=term&targettype=pubchem">ML115</a> can be used by academic researchers studying cell biology, molecular biology, and tumor biology, breast cancer biology in particular. In addition, the identified probe will be useful for any researcher working in the field of drug discovery.</p></div><div id="ml115.introd"><h3>Relevant biology of the probe</h3><p>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 [<a class="bibr" href="#ml115.r2" rid="ml115.r2">2</a>, <a class="bibr" href="#ml115.r3" rid="ml115.r3">3</a>]. STATs are activated in response to growth factors, cytokines, and G-CSF binding to cell surface receptor tyrosine kinases [<a class="bibr" href="#ml115.r2" rid="ml115.r2">2</a>–<a class="bibr" href="#ml115.r4" rid="ml115.r4">4</a>]. 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 [<a class="bibr" href="#ml115.r3" rid="ml115.r3">3</a>]. Although structurally similar, the seven STAT family member (STATs 1, 2, 3, 4, 5a, 5b, and 6) possess diverse biological roles [<a class="bibr" href="#ml115.r3" rid="ml115.r3">3</a>]. For example, STAT1 activation is pro-inflammatory and anti-proliferative, while STAT3 activation is anti-inflammatory and pro-apoptotic [<a class="bibr" href="#ml115.r3" rid="ml115.r3">3</a>]. STAT1 is largely responsible for mediating the effects of IFN-gamma, while STAT3 is predominantly involved in IL-6 signaling [<a class="bibr" href="#ml115.r5" rid="ml115.r5">5</a>]. STAT1 induces expression of genes that inhibit the cell cycle, and thus STAT1 is considered to have tumor suppressor properties [<a class="bibr" href="#ml115.r6" rid="ml115.r6">6</a>]. 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 [<a class="bibr" href="#ml115.r2" rid="ml115.r2">2</a>, <a class="bibr" href="#ml115.r5" rid="ml115.r5">5</a>, <a class="bibr" href="#ml115.r7" rid="ml115.r7">7</a>]. Blocking STAT3 interaction with the epidermal growth factor receptor (EGFR) using peptide aptamers has been shown to reduce tumor growth [<a class="bibr" href="#ml115.r8" rid="ml115.r8">8</a>]. Due to the diverse roles and potent phenotypes associated with STAT signaling, the identification of selective potentiators of STAT3 activity may lead to pharmacological tools for cancer, wound healing, and inflammatory diseases.</p><p>Finally, targeted activation of STAT3 may be important in preventing unwanted cell death, as in a myocardial infarction or stroke, and thus STAT3 activators may have independent medicinal uses. In fact, a recent (OCT2010) report shows that leptin signalling is able to reduce cardiac injury in post infracted heart tissue by acting directly on cardiomyocytes to increase STAT3 (and AMPK) signalling, decreasing apoptosis, cardiac hypertrophy, and inflammation [<a class="bibr" href="#ml115.r9" rid="ml115.r9">9</a>]. As a result, the identification of selective STAT3 modulators may provide useful tools for exploring STAT3 biology in cardiovascular disease and inflammatory disease.</p></div></div><div id="ml115.s2"><h2 id="_ml115_s2_">1. Introduction</h2><p>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 [<a class="bibr" href="#ml115.r2" rid="ml115.r2">2</a>, <a class="bibr" href="#ml115.r3" rid="ml115.r3">3</a>]. STATs are activated in response to growth factors, cytokines, and G-CSF binding to cell surface receptor tyrosine kinases [<a class="bibr" href="#ml115.r2" rid="ml115.r2">2</a>–<a class="bibr" href="#ml115.r4" rid="ml115.r4">4</a>]. 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 [<a class="bibr" href="#ml115.r3" rid="ml115.r3">3</a>]. Although structurally similar, the seven STAT family member (STATs 1, 2, 3, 4, 5a, 5b, and 6) possess diverse biological roles [<a class="bibr" href="#ml115.r3" rid="ml115.r3">3</a>]. For example, STAT1 activation is pro-inflammatory and anti-proliferative, while STAT3 activation is anti-inflammatory and pro-apoptotic [<a class="bibr" href="#ml115.r3" rid="ml115.r3">3</a>]. STAT1 is largely responsible for mediating the effects of IFN-γ while STAT3 is predominantly involved in IL-6 signaling [<a class="bibr" href="#ml115.r5" rid="ml115.r5">5</a>]. STAT1 induces expression of genes that inhibit the cell cycle, and thus STAT1 is considered to have tumor suppressor properties [<a class="bibr" href="#ml115.r6" rid="ml115.r6">6</a>]. 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 [<a class="bibr" href="#ml115.r2" rid="ml115.r2">2</a>, <a class="bibr" href="#ml115.r5" rid="ml115.r5">5</a>, <a class="bibr" href="#ml115.r7" rid="ml115.r7">7</a>]. Blocking STAT3 interaction with the epidermal growth factor receptor (EGFR) using peptide aptamers has been shown to reduce tumor growth [<a class="bibr" href="#ml115.r8" rid="ml115.r8">8</a>]. 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. A common goal of the HTS screens is to identify compounds which act selectively on either STAT1 or STAT3. Compounds will be considered selective based upon their ability to modulate only one member of the STAT family. Compounds of interest should exhibit high potency and selectivity. Test compounds that do not affect STAT1::luciferase or NFkB::luciferase activities will be considered selective activators of STAT3. The novel probe identified here met these criteria.</p></div><div id="ml115.s3"><h2 id="_ml115_s3_">2. Materials and Methods</h2><div id="ml115.s4"><h3>2.1. Assay Details</h3><p>Following primary HTS in singlicate to identify STAT3 activators (<a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/871" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-871</a>), confirmation of hit activity in triplicate (<a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/1267" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-1267</a>), counterscreening in triplicate against NFκ B (<a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/1309" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-1309</a>) and STAT3 (<a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/1318" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-1318</a>) to determine selectivity, followed by titration assays to determine compound potency (<a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/1398" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-1398</a>) and selectivity (<a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/1406" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-1406</a>), certain compounds were identified as possible candidates for probe development. Probes were identified (<a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/2049" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-2049</a>). Compared to previously described compounds, the probe described herein offers greatly improved STAT3 selectivity and potency.</p><div class="iconblock whole_rhythm clearfix ten_col table-wrap" id="figml115tu2"><a href="/books/NBK51964/table/ml115.tu2/?report=objectonly" target="object" title="Table" class="img_link icnblk_img figpopup" rid-figpopup="figml115tu2" rid-ob="figobml115tu2"><img class="small-thumb" src="/books/NBK51964/table/ml115.tu2/?report=thumb" src-large="/books/NBK51964/table/ml115.tu2/?report=previmg" alt="PubChem BioAssay Table." /></a><div class="icnblk_cntnt"><h4 id="ml115.tu2"><a href="/books/NBK51964/table/ml115.tu2/?report=objectonly" target="object" rid-ob="figobml115tu2">Table</a></h4><p class="float-caption no_bottom_margin">PubChem BioAssay Table. </p></div></div><div class="iconblock whole_rhythm clearfix ten_col table-wrap" id="figml115tu3"><a href="/books/NBK51964/table/ml115.tu3/?report=objectonly" target="object" title="Table" class="img_link icnblk_img figpopup" rid-figpopup="figml115tu3" rid-ob="figobml115tu3"><img class="small-thumb" src="/books/NBK51964/table/ml115.tu3/?report=thumb" src-large="/books/NBK51964/table/ml115.tu3/?report=previmg" alt="Table of Assay Rationale and Screening Statistics." /></a><div class="icnblk_cntnt"><h4 id="ml115.tu3"><a href="/books/NBK51964/table/ml115.tu3/?report=objectonly" target="object" rid-ob="figobml115tu3">Table</a></h4><p class="float-caption no_bottom_margin">Table of Assay Rationale and Screening Statistics. </p></div></div><p>Descriptions of each assay are below.</p><div id="ml115.s5"><h4>STAT3 Activation Assays (<a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/871" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-871</a>, <a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/1267" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-1267</a>, <a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/1398" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-1398</a>, and <a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/2049" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-2049</a>)</h4><p>This STAT3 potentiator assay was run simultaneously in the same plates as the STAT3 antagonist campaign (PubChem <a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/862" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-862</a>). The cells were grown in T-175 flasks in Dulbecco's Modified Eagle's Media (DMEM) supplemented with 10% v/v fetal bovine serum and antibiotics (50 ug/mL each of penicillin and streptomycin, 100 ug/mL neomycin) at 37 degrees C in an atmosphere of 5% CO2 and 95% relative humidity (RH). Cells were harvested by trypsinization when they passed 70% confluency.</p><p>Cells were resuspended in growth medium without phenol red, at a density of 1.88 million cells/mL and filtered through a 0.7 micron filter. Next, 4 ul of well-mixed cell suspension (7,520 cells per well) were dispensed into each well of 1536-well plates. The assay was started by immediately dispensing 28 nL of test compound (5.5 micromolar final nominal concentration in DMSO) or DMSO (less than 0.6% final concentration) to sample or control wells, respectively. The plates were then incubated for 1 hour at 37 degrees C (5% CO2, 95% RH). Then, 28nL of nifuroxazide (100 µM final concentration in DMSO) were added to a subset of control wells, to monitor that the assay was functioning properly. Next, 1 ul of human recombinant IL-6 (final nominal concentration 33 ng/mL) was dispensed into sample wells. This IL-6 concentration resulted in approximately 80% STAT3::luciferase reporter activity. Next, the plates were incubated for 6 hours at 37 degrees C (5% CO2, 95% RH). The assay was stopped by dispensing 5 ul of SteadyLite HTS luciferase substrate at room temperature to each well, followed by incubation at room temperature for 15 minutes. Luminescence was measured on the ViewLux plate reader.</p><p>Prior to % potentiation calculations, luminescence values were corrected by subtracting background luminescence, measured in wells containing media and substrate only.</p><p>The percent potentiation, referred as percent activation in the formula and data results was defined using the following mathematical formula: %Activity = 100*(Test_Compound/Median_High_Control)</p><p>Where:</p><p>Test_Compound is defined as the luminescence value of a well containing test compound.</p><p>Median_High_Control is defined as the median luminescence value of sample wells treated with 33 ng/mL IL-6 (IL-6 EC80, set as 100% activation).</p><p>A mathematical algorithm was used to determine nominally activating compounds in the primary screen. Two values were calculated: (1) the average percent potentiation of all compounds tested, and (2) three times their standard deviation. The sum of these two values was used as a cutoff parameter, i.e. any compound that exhibited greater % potentiation than the cutoff parameter was declared active.</p><p>The reported Pubchem_Activity_Score has been normalized. % Activation values of greater than or equal to 200 are reported as activity score 100. Negative % activation values are reported as activity score zero.</p><div id="ml115.s6"><h5>Reagents</h5><p>Dulbecco's Modified Eagle's Media I (Invitrogen, part# 11965-092)</p><p>Dulbecco's Modified Eagle's Media, no Phenol Red (Invitrogen, part# 21063-029)</p><p>Fetal Bovine Serum (Hyclone, part# SH30088-03)</p><p>100X Penicillin-Streptomycin-Neomycin mix (Invitrogen, part# 15640-055).</p><p>Neomycin B (Sigma-Aldrich, St. Louis, MO, part 33492)</p><p>Recombinant human IL-6 (Peprotech, Rocky Hill, NJ, part# 200-06)</p><p>Nifuroxazide (Sigma-Aldrich, part# N 2641)</p><p>SteadyLite HTS Assay Kit (PerkinElmer, part# 6016989)</p></div></div><div id="ml115.s7"><h4>STAT1 Activation Counterscreens (<a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/932" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-932</a>, <a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/1318" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-1318</a>, <a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/1406" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-1406</a>, and <a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/2049" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-2049</a>)</h4><div id="ml115.s8"><h5>Assay Overview</h5><p>Potentiation of STAT1 activity was measured using a murine NIH 3T3 fibroblast cell line that stably expresses a STAT1-luciferase construct. In this primary assay approximately 195,000 test compounds from the MLSCN library were screened for their ability to increase IFN-gamma-mediated STAT1::luciferase reporter activity. Cells were exposed to test compounds from the MLSCN library, followed by treatment with IFN-gamma. Changes in STAT1::luciferase activity were monitored by measuring luminescence. As designed, a STAT1 potentiator will increase IFN-gamma-mediated STAT1 transcription, thus increasing the activation of the luciferase reporter gene, and increasing luminescence.</p></div><div id="ml115.s9"><h5>Protocol Summary</h5><p>This STAT1 potentiator assay was run simultaneously in the same plates as the STAT1 antagonist campaign. The cells were grown in T-175 flasks in Dulbecco's Modified Eagle's Media (DMEM) supplemented with 10% v/v fetal bovine serum and antibiotics (50 ug/mL each of penicillin and streptomycin) at 37 degrees C in an atmosphere of 5% CO2 and 95% relative humidity (RH). Cells were harvested by trypsinization when they passed 70% confluency.</p><p>Cells were resuspended in growth medium without phenol red, at a density of 1.88 million cells/mL and filtered through a 0.7 micron filter. Next, 4 ul of well-mixed cell suspension (7,520 cells per well) were dispensed into each well of 1536-well plates. The assay was started by immediately dispensing 28 nL of test compound (5.5 µM final nominal concentration) in DMSO to sample wells, or DMSO alone (0.6% final concentration) to control wells. The plates were then incubated for 1 hour at 37 degrees C (5% CO2, 95% RH). Next, 1 ul of human recombinant IFN-gamma (final nominal EC80 concentration of 3.0 ng/mL, set as 100% activation) was dispensed to sample and control wells. The plates were then incubated for 6 hours at 37 degrees C (5% CO2, 95% RH). The assay was stopped by dispensing 5 ul of SteadyLite HTS luciferase substrate at room temperature to each well, followed by incubation at room temperature for 15 minutes. Luminescence was measured on the ViewLux plate reader. The percent activity was defined using the following mathematical formula: %Activation = 100*(Test_Compound/Median_High_Control)</p><p>Where:</p><p>Test_Compound is defined as the luminescence value of IFN-gamma-treated wells containing test compound.</p><p>Median_High_Control is defined as the median luminescence value of control wells containing IFN-gamma.</p><p>A mathematical algorithm was used to determine nominally activating compounds in the primary screen. Two values were calculated: (1) the average percent activation of all compounds tested, and (2) three times their standard deviation. The sum of these two values was used as a cutoff parameter (127.73%), i.e. any compound that exhibited greater % activation than the cutoff parameter was declared active.</p></div><div id="ml115.s10"><h5>Reagents</h5><p>Dulbecco's Modified Eagle's Media I (Invitrogen, part# 11965-092)</p><p>Dulbecco's Modified Eagle's Media, no Phenol Red (Invitrogen, part# 21063-029)</p><p>Fetal Bovine Serum (Hyclone, part# SH30088-03)</p><p>100X Penicillin-Streptomycin-Neomycin mix (Invitrogen, part# 15640-055).</p><p>Recombinant human IFN-gamma (R&D Systems, part# 485-MI)</p><p>SteadyLite HTS Assay Kit (PerkinElmer, part# 6016989)</p></div></div><div id="ml115.s11"><h4>NFkB Activation Counterscreens (<a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/1309" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-1309</a> and <a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/2049" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-2049</a>)</h4><div id="ml115.s12"><h5>Assay Overview</h5><p>The purpose of this assay is to determine whether a subset of compounds identified as active in a previous set of experiments entitled, “Primary cell-based high throughput screening assay to measure STAT3 activation” (PubChem <a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/871" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-871</a>) were nonselective due to activation of NF-kappaB. The compounds selected for testing in this AID met at least the two following criteria: 1) they were declared active in <a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/871" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-871</a>; and 2) they were declared inactive in a previous set of experiments entitled, “Primary cell-based high throughput screening assay to measure STAT1 activation” (PubChem <a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/932" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-932</a>).</p><p>In this assay activation of NF-kB transcription was measured using a human HEK 293T cell line that stably expresses a human NF-kB::luciferase construct. Test compounds were screened for their ability to increase TNF-alpha-mediated NF-kB::luciferase reporter activity. Cells were exposed to test compounds, followed by treatment with TNF-alpha to activate NF-kB transcription. Changes in NF-kB::luciferase activity were monitored by measuring luminescence. An activator will increase TNF-alpha-mediated NF-kB transcription, thus activating the luciferase reporter gene, and increasing well luminescence. As designed, test compounds that activate NF-kB activity are considered non-selective activators.</p></div><div id="ml115.s13"><h5>Protocol Summary</h5><p>The activator and inhibitor counterscreen assays using NF-kB::luciferase cells were run simultaneously. HEK 293T cells were grown in T-175 flasks in Dulbecco's Modified Eagle's Media (DMEM) supplemented with 10% v/v fetal bovine serum and antibiotics (50 micrograms/ml each of penicillin and streptomycin, 100 micrograms/mL neomycin and 1 microgram/mL of puromycin) at 37 degrees C in an atmosphere of 5% CO2 and 95% relative humidity (RH).</p><p>Prior to the start of the assay the cells were resuspended at a density of 0.94 million cells/mL in phenol red-free growth medium, and filtered through a 0.7 micron filter. Next, 4 ul of cell suspension (3,760 cells per well) were dispensed into each well of 1536-well plates. Cells were allowed to adhere to plates by incubation at 37 degrees C, 5% CO2, and 95% RH for 18 hours. The assay was started by dispensing 28 nL of test compound (5.7 µM final nominal concentration) in DMSO to sample wells, while High Control wells received DMSO (0.6% final concentration; set as 100% activation). A subset of wells received MG-132 (100 µM final nominal concentration in DMSO) to monitor that the assay was functioning properly. The plates were then incubated for 1 hour at 37 degrees C (5% CO2, 95% RH). Next, 1 ul of human recombinant TNF-alpha (5.6 ng/ml final concentration) was dispensed into sample and all control wells. This TNF-alpha concentration resulted in approximately 80% NF-kappaB::luciferase reporter activity. Next, plates were incubated for 6 hours at 37 degrees C (5% CO2, 95% RH). The assay was stopped by dispensing 5 ul of SteadyLite HTS luciferase substrate at room temperature to each well, followed by incubation at room temperature for 15 minutes. Luminescence was measured on the ViewLux plate reader.</p><p>The percent activation was defined using the following mathematical formula: %Activation = 100*(Test_Compound/Median_High_Control)</p><p>Where:</p><p>Test_Compound is defined as the luminescence value of a well containing TNF-alpha and test compound.</p><p>Median_High_Control is defined as the median luminescence of wells containing TNF-alpha and DMSO.</p><p>Test compounds were assayed in triplicate at a single concentration of 5.7 micromolar. A mathematical algorithm was used to determine nominally activating compounds. Two values were calculated: (1) the average percent activation of all 1280 wells defined as “Median_Low_Control” (i.e. a “DMSO plate”), and (2) three times their standard deviation. The sum of these two values was used as a cutoff parameter: any compound that exhibited greater % activation than the cutoff parameter was declared active.</p><p>The reported Pubchem_Activity_Score has been normalized to 100% of the highest observed activation. Negative % activation values were assigned an activity score of zero.</p></div><div id="ml115.s14"><h5>Reagents</h5><p>Dulbecco's Modified Eagle's Media I (Invitrogen, part 11965-092)</p><p>Dulbecco's Modified Eagle's Media, no Phenol Red (Invitrogen, part 21063-029)</p><p>Fetal Bovine Serum (Hyclone, part SH30088-03)</p><p>100X Penicillin-Streptomycin-Neomycin mix (Invitrogen, part 15640-055)</p><p>Neomycin B (Sigma-Aldrich, part 33492)</p><p>Puromycin (Sigma-Aldrich, part P8833)</p><p>Recombinant human TNF-alpha (Invitrogen, part PHC3016)</p><p>MG-132 (Tocris Cookson Ltd, part 1748-5)</p><p>SteadyLite HTS Assay Kit (PerkinElmer, part 6016989)</p><p>T175 Flasks (Corning, part 431080)</p><p>1536-well plates (Greiner, part 789173)</p></div></div><div id="ml115.s15"><h4>BCL3 QPCR Activation Counterscreens (<a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/2049" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-2049</a>)</h4><p>The purpose of this assay is to determine whether compounds of interest can activate expression of the STAT3 target gene, BCL3. The assay protocol has been described [<a class="bibr" href="#ml115.r10" rid="ml115.r10">10</a>]. Briefly, cells were incubated with either compound or vehicle, and RNA was isolated using the RNeasy kit (QIAGEN, Valencia, CA). cDNA was generated using the Taqman reverse transcription kit (Applied Biosystems, Foster City, CA). Quantitative real-time polymerase chain reaction (PCR) was performed in triplicate using SYBR green master mix (Applied Biosystems) on a model 7500 real time PCR system (Applied Biosystems). Data are expressed as the mean fold change plus or minus SE of 3 replicates. Each assay was repeated at least 3 times.</p></div><div id="ml115.s16"><h4>Cell Viability Counterscreen (<a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/2049" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-2049</a>)</h4><p>The purpose of this assay is to determine whether compounds identified as active in the STAT3 activation assay were false positives resulting from an increase in well luminescence due to increased cell proliferation. This assay employs the CellTiter-Glo luminescent reagent, which contains luciferase to catalyze the oxidation of beetle luciferin to oxyluciferin and light in the presence of cellular ATP. As designed, compounds that promote cell proliferation will increase cell numbers and ATP levels, resulting in increased well luminescence. Compounds were assayed in a 10-point 1:3 dilution series starting at a nominal concentration of 56 micromolar under conditions strictly identical to the STAT3 activation assay (6 hours incubation time, HT-1080 cells)</p></div></div><div id="ml115.s17"><h3>2.2. Probe Chemical Characterization</h3><div class="iconblock whole_rhythm clearfix ten_col table-wrap" id="figml115tu4"><a href="/books/NBK51964/table/ml115.tu4/?report=objectonly" target="object" title="Table" class="img_link icnblk_img figpopup" rid-figpopup="figml115tu4" rid-ob="figobml115tu4"><img class="small-thumb" src="/books/NBK51964/table/ml115.tu4/?report=thumb" src-large="/books/NBK51964/table/ml115.tu4/?report=previmg" alt="Image " /></a><div class="icnblk_cntnt"><h4 id="ml115.tu4"><a href="/books/NBK51964/table/ml115.tu4/?report=objectonly" target="object" rid-ob="figobml115tu4">Table</a></h4></div></div><div class="iconblock whole_rhythm clearfix ten_col table-wrap" id="figml115tu5"><a href="/books/NBK51964/table/ml115.tu5/?report=objectonly" target="object" title="Table" class="img_link icnblk_img figpopup" rid-figpopup="figml115tu5" rid-ob="figobml115tu5"><img class="small-thumb" src="/books/NBK51964/table/ml115.tu5/?report=thumb" src-large="/books/NBK51964/table/ml115.tu5/?report=previmg" alt="Table of Probe Properties (ML115)." /></a><div class="icnblk_cntnt"><h4 id="ml115.tu5"><a href="/books/NBK51964/table/ml115.tu5/?report=objectonly" target="object" rid-ob="figobml115tu5">Table</a></h4><p class="float-caption no_bottom_margin">Table of Probe Properties (ML115). </p></div></div><p>Graphical representation of the stability of the new probe compound is shown below. The new probe has been tested in numerous cell-based assays, demonstrating its cellular permeability, solubility, and stability.</p><div id="ml115.fu2" class="figure"><div class="graphic"><img src="/books/NBK51964/bin/ml115fu2.jpg" alt="Image ml115fu2" /></div></div><p><b>Development of LC-MS/MS analytical methods</b>–All analytical methods will be in MRM mode where the parent ion is selected in Q1 of the mass spectrometer. The parent ion is fragmented and a characteristic fragment ion is monitored in Q3. MRM mass spectroscopy methods are particularly sensitive because additional time is spent monitoring the desired ions and not sweeping a large mass range. Methods will be rapidly set up using Automaton<sup>®</sup> (Applied Biosystems), where the compounds are listed with their name and mass in an Excel datasheet. Compounds are submitted in a 96 well plate to the HPLC autosampler and are slowly injected without a column present. A narrow range centered on the indicated mass is scanned to detect the parent ion. The software then evaluates a few pre-selected parameters to determine conditions that maximize the signal for the parent ion. The molecule is then fragmented in the collision cell of the mass spectrometer and fragments with m/z larger than 70 but smaller than the parent mass are determined. Three separate collision energies are evaluated to fragment the parent ion and the largest three ions are selected. Each of these three fragment ions are further optimized and the best fragment is chosen. The software then inserts the optimized masses and parameters into a template method and saves it with a unique name that indicates the individual compound being optimized. Spectra for the parent ion and the fragmentation pattern are saved and can be reviewed later.</p><p><b>Solubility in PBS.</b> The solubility of compounds was tested in phosphate buffered saline, pH 7.4. Compounds were inverted for 24 hours in test tubes containing 1–2 mg of compound with 1 mL of PBS. The samples were centrifuged and analyzed by HPLC (Agilent 1100 with diode-array detector). Peak area was compared to a standard of known concentration. In cases when the concentration was too low for UV analysis or when the compound did not possess a good chromophore, LC-MS-MS analysis was used.</p><p><b>Stability in PBS.</b> Demonstration of stability in PBS was conducted under conditions likely to be experienced in a laboratory setting. The compound was dissolved in 1 ml of PBS at a concentration of 10 μM unless its maximum solubility was insufficient to achieve this concentration. Low solubility compounds were tested between ten and fifty percent of their solubility limit. The solution was immediately aliquoted into seven standard polypropylene microcentrifuge tubes which were stored at ambient temperature in a block microcentrifuge tube holder. Individual tubes were frozen at −80°C at 0, 1, 2, 4, 8, 24, and 48 hours. The frozen samples were thawed in a room temperature and an equal volume of acetonitrile was added prior to determination of concentration by LC-MS/MS.</p><p><b>Determination of Glutathione reactivity.</b> One microliter of a 10 mM compound stock solution was added to 1 ml of a freshly prepared solution of 100 μM reduced glutathione. Final compound concentration is 10 μM unless solubility limited. The solution was allowed to incubate at 37°C for two hours prior to being directly analyzed by for glutathione adduct formation. LC-MS/MS analyses of GSH adducts was performed on an API 4000 Q-TrapTM mass spectrometer equipped with a Turboionspray source (Applied Biosystems, Foster City, CA). Two methodologies were utilized, a negative precursor ion (PI) scan of m/z 272, corresponding to GSH fragmenting at the thioether bond, and a neutral loss scan of −129 AMU was used to detect GSH adducts. This triggered positive ion enhanced resolution and enhanced product ion scans [<a class="bibr" href="#ml115.r11" rid="ml115.r11">11</a>, <a class="bibr" href="#ml115.r12" rid="ml115.r12">12</a>]</p></div><div id="ml115.s18"><h3>2.3. Probe Preparation: Synthesis of STAT3 Activator Probe and Analogs</h3><p>The STAT3 activator probe and analogs were synthesized by coupling the appropriate substituted aniline with 5-substituted-isoxazole-3-carboxylic acids, using standard peptide coupling reagents. <a class="figpopup" href="/books/NBK51964/table/ml115.t1/?report=objectonly" target="object" rid-figpopup="figml115t1" rid-ob="figobml115t1">Table 1</a> lists structures of the probe compound (<b>1a</b>) and selected analogs that were synthesized according to Scheme 1. These data show that the activity of these compounds as STAT3 activators is sensitive to the substituent at the 5-position of the isoxazole ring (compare data for <b>1a</b>–<b>1d</b>, which shows that a cyclopropyl (<b>1a</b>) or isopropyl (<b>1c</b>) group at this position leads to superior STAT3 activation than compounds with propyl (<b>1b)</b> or methyl (<b>1d</b>) groups at this position. The activity of these compounds as STAT3 activators is also dependent on the substituents and substitution pattern of the aniline ring (compare <b>1a</b>, <b>1e</b>, <b>1f</b>, and <b>1g</b>). Ultimately, compound <b>1a</b> was selected as the probe owing to its potency (EC<sub>50</sub> = 2 nM) in the primary assay, together with its superior performance in the secondary assays performed in the assay provider’s laboratory.</p><div class="iconblock whole_rhythm clearfix ten_col table-wrap" id="figml115t1"><a href="/books/NBK51964/table/ml115.t1/?report=objectonly" target="object" title="Table 1" class="img_link icnblk_img figpopup" rid-figpopup="figml115t1" rid-ob="figobml115t1"><img class="small-thumb" src="/books/NBK51964/table/ml115.t1/?report=thumb" src-large="/books/NBK51964/table/ml115.t1/?report=previmg" alt="Table 1. Substituted isoxazole carboxamides as potent STAT3 activators." /></a><div class="icnblk_cntnt"><h4 id="ml115.t1"><a href="/books/NBK51964/table/ml115.t1/?report=objectonly" target="object" rid-ob="figobml115t1">Table 1</a></h4><p class="float-caption no_bottom_margin">Substituted isoxazole carboxamides as potent STAT3 activators. </p></div></div><p>The probe was selected from a collection of 14 analogs, 11 of which were obtained by chemical synthesis. A more extensive chemistry SAR analysis was not performed, owing to the very high potency of the probe.</p><div id="ml115.fu3" class="figure"><div class="graphic"><img src="/books/NBK51964/bin/ml115fu15.jpg" alt="Image ml115fu15" /></div></div><p><b>Synthesis and analytical data for</b>
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<b><i>N</i></b><b>-(4-chloro-2,5-dimethoxyphenyl)-5-cyclopropyl-1,2-oxazole-3-carboxamide (1a).</b> A mixture of 5-cyclopropylisoxazole-3-carboxylic acid (185 mg, 1.21 mmol), 4-Chloro-2,5-dimethoxyaniline (342 mg, 1.82 mmol), EDC (353 mg, 1.84 mmol), HOBt (258 mg, 1.91 mmol), DIEA (0.42 mL) in DMF was stirred at rt for three days. Saturated aqueous sodium bicarbonate was added; it was extracted with ethyl acetate and dried over sodium sulfate. The solvent was removed and the residue was purified by silica gel chromatography. The title compound was obtained as beige solid (purity: 98% by analytical HPLC).</p><p>Spectroscopic data for the probe. <sup>1</sup>H-NMR (400 MHz, d<sub>6</sub>-DMSO) δ 9.40 (s, 1H), 7.89 (s, 1H), 7.23 (s, 1H), 6.64 (s, 1H), 3.84 (s, 3H), 3.80 (s, 3H), 2.26-2.22 (m, 1H), 1.16-1.11 (m, 2H), 0.99-0.95 (m, 2H); <sup>13</sup>C-NMR (100 MHz, d<sub>6</sub>-DMSO) δ 177.28, 158.61, 156.67, 148.18, 143.97, 125.45, 116.56, 113.38, 106.90, 98.26, 56.76, 56.43, 8.73, 7.70; MS (m/z): 323 [M+1]<sup>+</sup>.</p><p>The spectroscopic data and chemical purity of the probe analogs were in complete agreement with the assigned structures.</p></div></div><div id="ml115.s19"><h2 id="_ml115_s19_">3. Results</h2><div id="ml115.s20"><h3>3.1. Summary of Screening Results</h3><p>Following primary HTS in singlicate to identify STAT3 activators (<a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/871" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-871</a>), confirmation of hit activity in triplicate (<a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/1267" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-1267</a>), counterscreening in triplicate against NFκB (<a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/1309" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-1309</a>) and STAT3 (<a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/1318" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-1318</a>) to determine selectivity, followed by titration assays to determine compound potency (<a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/1398" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-1398</a>) and selectivity (<a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/1406" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-1406</a>), certain compounds were identified as possible candidates for probe development. A novel probe was identified (<a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/2049" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-2049</a>). Compared to previously described compounds, the probe described herein offers greatly improved STAT3 selectivity and potency.</p><div class="iconblock whole_rhythm clearfix ten_col table-wrap" id="figml115tu6"><a href="/books/NBK51964/table/ml115.tu6/?report=objectonly" target="object" title="Table" class="img_link icnblk_img figpopup" rid-figpopup="figml115tu6" rid-ob="figobml115tu6"><img class="small-thumb" src="/books/NBK51964/table/ml115.tu6/?report=thumb" src-large="/books/NBK51964/table/ml115.tu6/?report=previmg" alt="Summary Table of Screening Results." /></a><div class="icnblk_cntnt"><h4 id="ml115.tu6"><a href="/books/NBK51964/table/ml115.tu6/?report=objectonly" target="object" rid-ob="figobml115tu6">Table</a></h4><p class="float-caption no_bottom_margin">Summary Table of Screening Results. </p></div></div></div><div id="ml115.s21"><h3>3.2. Dose Response Curves for Probe</h3><div id="ml115.fu4" class="figure"><div class="graphic"><img src="/books/NBK51964/bin/ml115fu16.jpg" alt="Image ml115fu16" /></div></div><p><b><a href="/pcsubstance/?term=ML115[synonym]" ref="pagearea=body&targetsite=entrez&targetcat=term&targettype=pubchem">ML115</a> is a potent, dose-dependent STAT3 activator.</b>
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<a href="/pcsubstance/?term=ML115[synonym]" ref="pagearea=body&targetsite=entrez&targetcat=term&targettype=pubchem">ML115</a> was tested in the cell-based STAT3 and STAT1 assays. Due to the very low EC50 values of this compound, activities were assessed using 2 overlapping 10-point, 1:3 serial dilutions starting at 56 μM and 5.6 μM. The probe displayed a dose-dependent activation with a calculated EC50 of 2nM in the STAT3 assay and no apparent activity in the STAT1 assay.</p></div><div id="ml115.s22"><h3>3.3. Scaffold/Moiety Chemical Liabilities</h3><p>Analog 4 from a commercial source was initially found to be very potent (5 nM). The structure of this compound as well as all the other potent analogs was confirmed by re-synthesis in our lab. However, in a subsequent round Analog 4 from both the commercial source and on-house TSRI synthesis was far less potent (2 μm and 7 μm respectively). Repeated testing of this analog was not done as we identified several analogs with confirmed potency and structure in the low nanomolar range.</p><div class="iconblock whole_rhythm clearfix ten_col table-wrap" id="figml115tu7"><a href="/books/NBK51964/table/ml115.tu7/?report=objectonly" target="object" title="Table" class="img_link icnblk_img figpopup" rid-figpopup="figml115tu7" rid-ob="figobml115tu7"><img class="small-thumb" src="/books/NBK51964/table/ml115.tu7/?report=thumb" src-large="/books/NBK51964/table/ml115.tu7/?report=previmg" alt="STAT3 Activators SAR Table (Isoxazole Carboxamide Scaffold)." /></a><div class="icnblk_cntnt"><h4 id="ml115.tu7"><a href="/books/NBK51964/table/ml115.tu7/?report=objectonly" target="object" rid-ob="figobml115tu7">Table</a></h4><p class="float-caption no_bottom_margin">STAT3 Activators SAR Table (Isoxazole Carboxamide Scaffold). </p></div></div></div><div id="ml115.s23"><h3>3.4. Cellular Activity</h3><p>The probe (<a href="/pcsubstance/?term=ML115[synonym]" ref="pagearea=body&targetsite=entrez&targetcat=term&targettype=pubchem">ML115</a>/CID 6619100/<a href="https://pubchem.ncbi.nlm.nih.gov/substance/14735210" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">SID-14735210</a>/SR01-000122599) is active in a variety of cell-based assays performed by the SRIMSC and assay provider. These assays were performed to determine the probe’s selectivity for STAT3 in comparison to the related STAT1, STAT5, and NFkB signaling pathways. The results of these studies demonstrated that the probe does not increase STAT1, STAT5 or NFkB signaling pathways. In addition the cell proliferation assays reveal that the selective STAT3 activation is not due to an increase in cell growth. As a result, <a href="/pcsubstance/?term=ML115[synonym]" ref="pagearea=body&targetsite=entrez&targetcat=term&targettype=pubchem">ML115</a> is a selective and potent STAT3 activator probe.</p></div><div id="ml115.s24"><h3>3.5. Profiling Assays</h3><p>In addition to the above cellular assays, we have also obtained profiling results for the STAT3 activator probe at 10 µM using the NCI-60 DTP Human Tumor Cell Line Screen (Background and Methods can be found at <a href="http://dtp.nci.nih.gov/branches/btb/ivclsp.html" ref="pagearea=body&targetsite=external&targetcat=link&targettype=uri">http://dtp.nci.nih.gov/branches/btb/ivclsp.html</a>). This profiling screen examined the effect of the probe at a single high concentration (10 μM) on cellular growth of 60 well characterized cell lines during 48 hours of exposure (longer than the 6–8 hour exposure used in our cell-based reporter assays). Despite the use of this concentration which is several orders of magnitude above the EC50 of this probe, the mean cell growth among all 60 cell lines was 95% of that seen in untreated cells <b>(refer to figure below of the NCI60 screen)</b>. Thus, it is clear that this probe is non-toxic across the dose range utilized in these experiments.</p><div id="ml115.fu5" class="figure"><div class="graphic"><img src="/books/NBK51964/bin/ml115fu32.jpg" alt="Image ml115fu32" /></div></div></div></div><div id="ml115.s25"><h2 id="_ml115_s25_">4. Discussion</h2><div id="ml115.s26"><h3>4.1. Comparison to existing art and how the new probe is an improvement</h3><p>Probe <a href="/pcsubstance/?term=ML115[synonym]" ref="pagearea=body&targetsite=entrez&targetcat=term&targettype=pubchem">ML115</a> was identified from the STAT3 uHTS campaign. It was selected as a starting point for medicinal chemistry because it increased expression of BCL3, a known STAT3-dependent oncogene. It was selected as a probe since analogs that were generated during the probe development effort were either less potent against STAT3 (analogs 1, 3 and 4), or did not significantly increase BCL3 transcription (e.g., analog 2). This compound does not share structural similarities with the known STAT3 activators Compound 1 and 56.</p><p>The current prior art for STAT3 activators include the 2 compounds below:</p><div id="ml115.fu6" class="figure"><div class="graphic"><img src="/books/NBK51964/bin/ml115fu33.jpg" alt="Image ml115fu33" /></div></div><p>The first prior art compound (above) is known as <b>Compound 56</b> ([4-[(3-Bromophenyl)amino]-6,7-diethoxyquinazoline], is soluble in DMSO (200mg/ml) and commercially available Calbiochem (<a href="https://pubchem.ncbi.nlm.nih.gov/substance/26758491" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">SID-26758491</a>; CID 2857; part number 234505). It is a suspected carcinogen and teratogen. Compound 56 is a cell-permeable, reversible, potent and specific inhibitor of the tyrosine kinase activity of the epidermal growth factor receptor (EGFR) (IC50 = 6 pM). The compound binds competitively to the ATP site [<a class="bibr" href="#ml115.r13" rid="ml115.r13">13</a>]. Most importantly, although compound 56 exhibited potency in the STAT3 activator assay, it was equally potent in the STAT1 activator assay. Thus the new probe <a href="/pcsubstance/?term=ML115[synonym]" ref="pagearea=body&targetsite=entrez&targetcat=term&targettype=pubchem">ML115</a> is an improvement over compound 56 since it is selective.</p><div id="ml115.fu7" class="figure"><div class="graphic"><img src="/books/NBK51964/bin/ml115fu34.jpg" alt="Image ml115fu34" /></div></div><p>The second prior art compound is <b>Compound 1</b> (above). Compound 1 is a xanthocillin isolated from a marine fungus and has been reported to induce the phosphorylation of STAT3, as well as Jak2 and STAT5, in human leukemia cell line [<a class="bibr" href="#ml115.r14" rid="ml115.r14">14</a>]. Thus, it is non-selective. Compound 1 is a thrombopoietin mimetic small molecule. No information is available on its mechanism of action, nor of its ability to potentiate IL-6-induced STAT3 specifically. Compound 1 was reported as cytotoxic to HeLa cancer cells [<a class="bibr" href="#ml115.r1" rid="ml115.r1">1</a>], and exhibited anti-viral activity as shown by inhibition of plaque formation of Newcastle disease virus (NDV) without cytotoxicity. Finally, Compound <b>1</b> was reported to inhibit the biosynthesis of prostaglandin E2 and F2α at IC<sub>50</sub> values of 0.2 μM, but the mechanism of this activity is not known [<a class="bibr" href="#ml115.r15" rid="ml115.r15">15</a>]. As expected for a useful STAT3 activator, QPCR experiments (run by the assay provider) confirm that probe <a href="/pcsubstance/?term=ML115[synonym]" ref="pagearea=body&targetsite=entrez&targetcat=term&targettype=pubchem">ML115</a> upregulates transcription of the oncogene BCL3. In addition, the probe does not exhibit cytotoxicity in the parental cell lines used for the STAT3 assay (HT-1080) and STAT1 assay (NIH-3T3). In contrast, prior art Compound 1 is a non-selective xanthocillin reported to induce phosphorylation of STAT3, Jak2, and STAT5 in a human leukemia cell line [<a class="bibr" href="#ml115.r15" rid="ml115.r15">15</a>].</p><p>As shown in the table below, the prior art compounds are not selective for STAT3. The probe is also significantly more potent than prior art compound 56. As a result, the new probe (<a href="/pcsubstance/?term=ML115[synonym]" ref="pagearea=body&targetsite=entrez&targetcat=term&targettype=pubchem">ML115</a>) with its nanomolar potency is a considerable improvement over the prior art, and will serve as a valuable tool to elucidate STAT3 biology and signaling in a variety of cell types and disease models.</p><div class="iconblock whole_rhythm clearfix ten_col table-wrap" id="figml115tu8"><a href="/books/NBK51964/table/ml115.tu8/?report=objectonly" target="object" title="Table" class="img_link icnblk_img figpopup" rid-figpopup="figml115tu8" rid-ob="figobml115tu8"><img class="small-thumb" src="/books/NBK51964/table/ml115.tu8/?report=thumb" src-large="/books/NBK51964/table/ml115.tu8/?report=previmg" alt="Comparative Activity Table: Probe and Prior Art Compounds." /></a><div class="icnblk_cntnt"><h4 id="ml115.tu8"><a href="/books/NBK51964/table/ml115.tu8/?report=objectonly" target="object" rid-ob="figobml115tu8">Table</a></h4><p class="float-caption no_bottom_margin">Comparative Activity Table: Probe and Prior Art Compounds. </p></div></div></div><div id="ml115.s27"><h3>4.2. Mechanism of Action Studies</h3><p>In addition to the cellular and profiling assays above, it was important to determine whether the STAT3 activator probe (<a href="/pcsubstance/?term=ML115[synonym]" ref="pagearea=body&targetsite=entrez&targetcat=term&targettype=pubchem">ML115</a>) could lead to induction of relevant STAT3 target genes. Thus, the assay provider performed quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR) to examine the mRNA expression of the STAT3 target gene, BCL3. These assays are described [<a class="bibr" href="#ml115.r10" rid="ml115.r10">10</a>]. Briefly, cells were incubated with either compound or vehicle, and RNA was isolated using the RNeasy kit (QIAGEN, Valencia, CA). cDNA was generated using the Taqman reverse transcription kit (Applied Biosystems, Foster City, CA). Quantitative real-time polymerase chain reaction (PCR) was performed in triplicate using SYBR green master mix (Applied Biosystems) on a model 7500 real time PCR system (Applied Biosystems). Data are expressed as the mean fold change plus or minus SE of 3 replicates, normalized to cells treated with IL-6 alone. Each assay was repeated at least 3 times. Graphical results of BCL3 mRNA expression are below. The results show that the probe (<a href="https://pubchem.ncbi.nlm.nih.gov/substance/14735210" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">SID-14735210</a>) exhibits robust induction of BCL3, consistent with its ability to activate STAT3 signaling.</p><div id="ml115.fu8" class="figure"><div class="graphic"><img src="/books/NBK51964/bin/ml115fu35.jpg" alt="Image ml115fu35" /></div></div><p>Going forward, we would very much like to have biological data reflecting the increased STAT3-dependent gene transcription induced by these compounds. In simple cell culture experiments looking at phenotypes like proliferation or survival, we have not been able to discern an effect from increased STAT3-dependent gene expression. There are many more stringent phenotypes which might be useful for this purpose, such as colony formation, migration, invasion, differentiation, and apoptotic potential which may reflect this effect, though we have not examined those as of yet. Similarly, physiological readouts like cytoprotection in models of cardiac or cerebral ischemia may also reflect the effect of enhanced STAT3 function. Our hope of course, is to examine these effects through collaborations with labs that routinely study these processes.</p><p>Finally, although we do not know the specific upstream mechanism by which this novel STAT3 activator probe (<a href="/pcsubstance/?term=ML115[synonym]" ref="pagearea=body&targetsite=entrez&targetcat=term&targettype=pubchem">ML115</a>) enhances STAT3-dependent gene expression, we do know that it very potent (IC50 2nM) and importantly, it is inactive against the related transcriptional modifiers STAT1 and NFkB, making <a href="/pcsubstance/?term=ML115[synonym]" ref="pagearea=body&targetsite=entrez&targetcat=term&targettype=pubchem">ML115</a> a significant improvement in cell-based systems over the current art.</p></div><div id="ml115.s28"><h3>4.3. Planned Future Studies</h3><p>As indicated by the assay provider, future studies with the probe will fall into three spheres. First, through an ongoing synthetic chemistry effort we will make structural analogues of the identified probes to be tested for enhanced STAT3-dependent gene transcription in the luciferase system and subsequently with endogenous STAT3 target genes. This will provide key insights into structure-function relationships of these probes.</p><p>Second, through a series of experiments analyzing both the kinetics and magnitude of STAT3 phosphorylation, nuclear localization, DNA binding, and co-activator recruitment, we will identify the direct molecular mechanism(s) by which each probe is able to enhance STAT3-dependent gene expression. We will also make use of our ability to perform large scale RNA interference-based screens to identify direct molecular targets by which these probes exert their effects.</p><p>Finally, we will examine the biological effects of the probes in both cell culture and animal models. Among the cell-based phenotypes to be explored will be colony formation, migration, invasion, and apoptotic potential. The underlying hypothesis for potential medical applications is that a STAT3 activator would accelerate wound repair. As noted, STAT3 plays an important role in maintaining the pluripotent state of embryonic stem cells, and thus the effects of these probes on self-renewal and differentiation in stem cell systems will also be explored. Lastly, STAT3 activation may be important in mitigating cell death in cardiac or cerebral ischemia, the first and third leading causes of death in the United States. Thus, we will explore the effects of the STAT3 activators in physiological models studying these two diseases.</p></div></div><div id="ml115.s29"><h2 id="_ml115_s29_">5. References</h2><dl class="temp-labeled-list"><dl class="bkr_refwrap"><dt>1.</dt><dd><div class="bk_ref" id="ml115.r1">Takahashi C, Sekita S, Yoshihira K, Natori S. The structures of toxic metabolites of Aspergillus candidus. II. The compound B (xanthoascin), a hepato- and cardio-toxic xanthocillin analog. <span><span class="ref-journal">Chem Pharm Bull (Tokyo). </span>1976;<span class="ref-vol">24</span>(10):2317–21.</span> PMID 1017077. [<a href="https://pubmed.ncbi.nlm.nih.gov/1017077" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pubmed">PubMed<span class="bk_prnt">: 1017077</span></a>]</div></dd></dl><dl class="bkr_refwrap"><dt>2.</dt><dd><div class="bk_ref" id="ml115.r2">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. <span><span class="ref-journal">Cancer Res. </span>2005;<span class="ref-vol">65</span>(12):5054–62.</span> PMID 15958548. [<a href="https://pubmed.ncbi.nlm.nih.gov/15958548" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pubmed">PubMed<span class="bk_prnt">: 15958548</span></a>]</div></dd></dl><dl class="bkr_refwrap"><dt>3.</dt><dd><div class="bk_ref" id="ml115.r3">Schindler C, Levy DE, Decker T. JAK-STAT signaling: from interferons to cytokines. <span><span class="ref-journal">J Biol Chem. </span>2007;<span class="ref-vol">282</span>(28):20059–63.</span> PMID 17502367. [<a href="https://pubmed.ncbi.nlm.nih.gov/17502367" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pubmed">PubMed<span class="bk_prnt">: 17502367</span></a>]</div></dd></dl><dl class="bkr_refwrap"><dt>4.</dt><dd><div class="bk_ref" id="ml115.r4">Germain D, Frank DA. Targeting the cytoplasmic and nuclear functions of signal transducers and activators of transcription 3 for cancer therapy. <span><span class="ref-journal">Clin Cancer Res. </span>2007;<span class="ref-vol">13</span>(19):5665–9.</span> PMID 17908954. [<a href="https://pubmed.ncbi.nlm.nih.gov/17908954" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pubmed">PubMed<span class="bk_prnt">: 17908954</span></a>]</div></dd></dl><dl class="bkr_refwrap"><dt>5.</dt><dd><div class="bk_ref" id="ml115.r5">Levy DE, Darnell JE Jr. Stats: transcriptional control and biological impact. <span><span class="ref-journal">Nat Rev Mol Cell Biol. </span>2002;<span class="ref-vol">3</span>(9):651–62.</span> PMID 12209125. [<a href="https://pubmed.ncbi.nlm.nih.gov/12209125" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pubmed">PubMed<span class="bk_prnt">: 12209125</span></a>]</div></dd></dl><dl class="bkr_refwrap"><dt>6.</dt><dd><div class="bk_ref" id="ml115.r6">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. <span><span class="ref-journal">Clin Cancer Res. </span>2003;<span class="ref-vol">9</span>(6):2166–72.</span> PMID 12796382. [<a href="https://pubmed.ncbi.nlm.nih.gov/12796382" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pubmed">PubMed<span class="bk_prnt">: 12796382</span></a>]</div></dd></dl><dl class="bkr_refwrap"><dt>7.</dt><dd><div class="bk_ref" id="ml115.r7">Takeda K, Kaisho T, Yoshida N, Takeda J, Kishimoto T, Akira S. Stat3 activation is responsible for IL-6-dependent T cell proliferation through preventing apoptosis: generation and characterization of T cell-specific Stat3-deficient mice. <span><span class="ref-journal">J Immunol. </span>1998;<span class="ref-vol">161</span>(9):4652–60.</span> PMID 9794394. [<a href="https://pubmed.ncbi.nlm.nih.gov/9794394" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pubmed">PubMed<span class="bk_prnt">: 9794394</span></a>]</div></dd></dl><dl class="bkr_refwrap"><dt>8.</dt><dd><div class="bk_ref" id="ml115.r8">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. <span><span class="ref-journal">J Biol Chem. </span>2003;<span class="ref-vol">278</span>(39):37610–21.</span> PMID 12842895. [<a href="https://pubmed.ncbi.nlm.nih.gov/12842895" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pubmed">PubMed<span class="bk_prnt">: 12842895</span></a>]</div></dd></dl><dl class="bkr_refwrap"><dt>9.</dt><dd><div class="bk_ref" id="ml115.r9">McGaffin KR, Witham WG, Yester KA, Romano LC, O'Doherty RM, McTiernan CF, O'Donnell CP. Cardiac-specific leptin receptor deletion exacerbates ischaemic heart failure. <span><span class="ref-journal">Cardiovasc Res. </span>2010.</span> PMID 20833647. [<a href="/pmc/articles/PMC3002875/" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pmc">PMC free article<span class="bk_prnt">: PMC3002875</span></a>] [<a href="https://pubmed.ncbi.nlm.nih.gov/20833647" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pubmed">PubMed<span class="bk_prnt">: 20833647</span></a>]</div></dd></dl><dl class="bkr_refwrap"><dt>10.</dt><dd><div class="bk_ref" id="ml115.r10">Nelson EA, Walker SR, Kepich A, Gashin LB, Hideshima T, Ikeda H, Chauhan D, Anderson KC, Frank DA. Nifuroxazide inhibits survival of multiple myeloma cells by directly inhibiting STAT3. <span><span class="ref-journal">Blood. </span>2008;<span class="ref-vol">112</span>(13):5095–102.</span> PMID 18824601. [<a href="/pmc/articles/PMC2597607/" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pmc">PMC free article<span class="bk_prnt">: PMC2597607</span></a>] [<a href="https://pubmed.ncbi.nlm.nih.gov/18824601" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pubmed">PubMed<span class="bk_prnt">: 18824601</span></a>]</div></dd></dl><dl class="bkr_refwrap"><dt>11.</dt><dd><div class="bk_ref" id="ml115.r11">Li X, He Y, Ruiz CH, Koenig M, Cameron MD, Vojkovsky T. Characterization of dasatinib and its structural analogs as CYP3A4 mechanism-based inactivators and the proposed bioactivatio pathways. <span><span class="ref-journal">Drug Metab Dispos. </span>2009;<span class="ref-vol">37</span>(6):1242–50.</span> PMID 19282395. [<a href="/pmc/articles/PMC3202349/" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pmc">PMC free article<span class="bk_prnt">: PMC3202349</span></a>] [<a href="https://pubmed.ncbi.nlm.nih.gov/19282395" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pubmed">PubMed<span class="bk_prnt">: 19282395</span></a>]</div></dd></dl><dl class="bkr_refwrap"><dt>12.</dt><dd><div class="bk_ref" id="ml115.r12">Li X, Kamenecka TM, Cameron MD. Bioactivation of the epidermal growth factor receptor inhibitor gefitinib: implications for pulmonary and hepatic toxicities. <span><span class="ref-journal">Chem Res Toxicol. </span>2009;<span class="ref-vol">22</span>(10):1736–42.</span> PMID 19803472. [<a href="https://pubmed.ncbi.nlm.nih.gov/19803472" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pubmed">PubMed<span class="bk_prnt">: 19803472</span></a>]</div></dd></dl><dl class="bkr_refwrap"><dt>13.</dt><dd><div class="bk_ref" id="ml115.r13">Bridges AJ, Zhou H, Cody DR, Rewcastle GW, McMichael A, Showalter HD, Fry DW, Kraker AJ, Denny WA. Tyrosine kinase inhibitors. 8. An unusually steep structure-activity relationship for analogues of 4-(3-bromoanilino)-6,7-dimethoxyquinazoline (PD 153035), a potent inhibitor of the epidermal growth factor receptor. <span><span class="ref-journal">J Med Chem. </span>1996;<span class="ref-vol">39</span>(1):267–76.</span> PMID 8568816. [<a href="https://pubmed.ncbi.nlm.nih.gov/8568816" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pubmed">PubMed<span class="bk_prnt">: 8568816</span></a>]</div></dd></dl><dl class="bkr_refwrap"><dt>14.</dt><dd><div class="bk_ref" id="ml115.r14">Sakai R, Nakamura T, Nishino T, Yamamoto M, Miyamura A, Miyamoto H, Ishiwata N, Komatsu N, Kamiya H, Tsuruzoe N. Xanthocillins as thrombopoietin mimetic small molecules. <span><span class="ref-journal">Bioorg Med Chem. </span>2005;<span class="ref-vol">13</span>(23):6388–93.</span> PMID 16115772. [<a href="https://pubmed.ncbi.nlm.nih.gov/16115772" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pubmed">PubMed<span class="bk_prnt">: 16115772</span></a>]</div></dd></dl><dl class="bkr_refwrap"><dt>15.</dt><dd><div class="bk_ref" id="ml115.r15">Kitahara N, Endo A. Xanthocillin X mono methyl ether, a potent inhibitor of prostaglandin biosynthesis. <span><span class="ref-journal">J Antibiot (Tokyo). </span>1981;<span class="ref-vol">34</span>(12):1556–61.</span> PMID 7333967. [<a href="https://pubmed.ncbi.nlm.nih.gov/7333967" ref="pagearea=cite-ref&targetsite=entrez&targetcat=link&targettype=pubmed">PubMed<span class="bk_prnt">: 7333967</span></a>]</div></dd></dl></dl></div><div id="bk_toc_contnr"></div></div></div><div class="fm-sec"><h2 id="_NBK51964_pubdet_">Publication Details</h2><h3>Author Information and Affiliations</h3><p class="contrib-group"><h4>Authors</h4><span itemprop="author">F Madoux</span>,<sup>1</sup> <span itemprop="author">M Koenig</span>,<sup>2</sup> <span itemprop="author">E Nelson</span>,<sup>3</sup> <span itemprop="author">S Chowdhury</span>,<sup>2</sup> <span itemprop="author">M Cameron</span>,<sup>4</sup> <span itemprop="author">B Mercer</span>,<sup>1</sup> <span itemprop="author">W Roush</span>,<sup>2</sup> <span itemprop="author">D Frank</span>,<sup>3</sup> and <span itemprop="author">P Hodder</span><sup>1</sup>.<sup><img src="/corehtml/pmc/pmcgifs/corrauth.gif" alt="corresponding author" /></sup></p><h4>Affiliations</h4><div class="affiliation"><sup>1</sup>
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Lead Identification Division, Translational Research Institute, Scripps Florida, C130 Scripps Way, Jupiter, Fl, 33458</div><div class="affiliation"><sup>2</sup>
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Department of Chemistry, Scripps Florida, C130 Scripps Way, Jupiter, Fl, 33458</div><div class="affiliation"><sup>3</sup>
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Department of Medical Oncology, Dana Farber Cancer Institute, 44 Binney Street, Boston, MA 02115</div><div class="affiliation"><sup>4</sup>
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Department of Molecular Therapeutics, Scripps Florida, C130 Scripps Way, Jupiter, Fl, 33458</div><div class="affiliation"><sup><img src="/corehtml/pmc/pmcgifs/corrauth.gif" alt="corresponding author" /></sup>
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To whom correspondence should be addressed,
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<span class="before-email-separator"></span><span class="email-label">Email: </span><a href="mailto:dev@null" data-email="ude.sppircs@preddoh" class="oemail">ude.sppircs@preddoh</a></div><h3>Publication History</h3><p class="small">Received: <span itemprop="datePublished">August 27, 2009</span>; Last Update: <span itemprop="dateModified">December 16, 2010</span>.</p><h3>Copyright</h3><div><div class="half_rhythm"><a href="/books/about/copyright/">Copyright Notice</a></div></div><h3>Publisher</h3><p>National Center for Biotechnology Information (US), Bethesda (MD)</p><h3>NLM Citation</h3><p>Madoux F, Koenig M, Nelson E, et al. Modulators of STAT Transcription Factors for the Targeted Therapy of Cancer (STAT3 Activators) 2009 Aug 27 [Updated 2010 Dec 16]. In: Probe Reports from the NIH Molecular Libraries Program [Internet]. Bethesda (MD): National Center for Biotechnology Information (US); 2010-. <span class="bk_cite_avail"></span></p></div><div class="small-screen-prev"><a href="/books/n/mlprobe/ml116/?report=reader"><svg xmlns="http://www.w3.org/2000/svg" viewBox="0 0 100 100" preserveAspectRatio="none"><path d="M75,30 c-80,60 -80,0 0,60 c-30,-60 -30,0 0,-60"></path><text x="20" y="28" textLength="60" style="font-size:25px">Prev</text></svg></a></div><div class="small-screen-next"><a href="/books/n/mlprobe/ml114/?report=reader"><svg xmlns="http://www.w3.org/2000/svg" viewBox="0 0 100 100" preserveAspectRatio="none"><path d="M25,30c80,60 80,0 0,60 c30,-60 30,0 0,-60"></path><text x="20" y="28" textLength="60" style="font-size:25px">Next</text></svg></a></div></article><article data-type="fig" id="figobml115fu1"><div id="ml115.fu1" class="figure"><div class="graphic"><img data-src="/books/NBK51964/bin/ml115fu1.jpg" alt="Image ml115fu1" /></div></div></article><article data-type="table-wrap" id="figobml115tu1"><div id="ml115.tu1" class="table"><h3><span class="title">STAT3 Activator Probe Information (Isoxazole Carboxamide Scaffold; MLS 000522376)</span></h3><p class="large-table-link" style="display:none"><span class="right"><a href="/books/NBK51964/table/ml115.tu1/?report=objectonly" target="object">View in own window</a></span></p><div class="large_tbl" id="__ml115.tu1_lrgtbl__"><table class="no_top_margin"><thead><tr><th id="hd_h_ml115.tu1_1_1_1_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">CID/ML#</th><th id="hd_h_ml115.tu1_1_1_1_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Target Name</th><th id="hd_h_ml115.tu1_1_1_1_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">EC50 (nM) [SID, AID]</th><th id="hd_h_ml115.tu1_1_1_1_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Anti-target</th><th id="hd_h_ml115.tu1_1_1_1_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">EC50 (μM) [SID, AID]</th><th id="hd_h_ml115.tu1_1_1_1_6" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Fold Selective</th><th id="hd_h_ml115.tu1_1_1_1_7" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Secondary Assays: EC50 (nM) [SID, AID]</th></tr></thead><tbody><tr><td headers="hd_h_ml115.tu1_1_1_1_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">CID 6619100/<a href="/pcsubstance/?term=ML115[synonym]" ref="pagearea=body&targetsite=entrez&targetcat=term&targettype=pubchem">ML115</a></td><td headers="hd_h_ml115.tu1_1_1_1_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">STAT3</td><td headers="hd_h_ml115.tu1_1_1_1_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">2.0 nM [<a href="https://pubchem.ncbi.nlm.nih.gov/substance/14735210" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">SID-14735210</a>, <a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/2049" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-2049</a>] Active</td><td headers="hd_h_ml115.tu1_1_1_1_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">STAT1</td><td headers="hd_h_ml115.tu1_1_1_1_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">> 56 µM [<a href="https://pubchem.ncbi.nlm.nih.gov/substance/14735210" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">SID-14735210</a>, <a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/2049" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-2049</a>] Inactive</td><td headers="hd_h_ml115.tu1_1_1_1_6" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;"><b>>28,000 fold selective</b></td><td headers="hd_h_ml115.tu1_1_1_1_7" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;"><b>NFkB Counterscreen (3X%ACT)</b>: 103.19%
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[<a href="https://pubchem.ncbi.nlm.nih.gov/substance/14735210" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">SID-14735210</a>, <a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/1309" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-1309</a>] Inactive<br /><br /><b>Cell Viability Counterscreen</b>: > 56 µM [<a href="https://pubchem.ncbi.nlm.nih.gov/substance/14735210" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">SID-14735210</a>, <a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/2049" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-2049</a>] (HT-1080 & NIH-3T3 cells) Inactive<br /><br /><b>BCL3 QPCR Assay</b>: [<a href="https://pubchem.ncbi.nlm.nih.gov/substance/14735210" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">SID-14735210</a>, <a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/2049" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-2049</a>] Active</td></tr></tbody></table></div></div></article><article data-type="table-wrap" id="figobml115tu2"><div id="ml115.tu2" class="table"><h3><span class="title">PubChem BioAssay Table</span></h3><p class="large-table-link" style="display:none"><span class="right"><a href="/books/NBK51964/table/ml115.tu2/?report=objectonly" target="object">View in own window</a></span></p><div class="large_tbl" id="__ml115.tu2_lrgtbl__"><table class="no_top_margin"><thead><tr><th id="hd_h_ml115.tu2_1_1_1_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">AID</th><th id="hd_h_ml115.tu2_1_1_1_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Assay Name</th><th id="hd_h_ml115.tu2_1_1_1_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Assay Type</th><th id="hd_h_ml115.tu2_1_1_1_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Target</th><th id="hd_h_ml115.tu2_1_1_1_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Powder Sample</th><th id="hd_h_ml115.tu2_1_1_1_6" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Compound Concentration</th></tr></thead><tbody><tr><td headers="hd_h_ml115.tu2_1_1_1_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;"><a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/871" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">871</a></td><td headers="hd_h_ml115.tu2_1_1_1_2" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;">Primary cell-based high throughput screening assay to measure STAT3 activation</td><td headers="hd_h_ml115.tu2_1_1_1_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Primary Assay (1X%ACT)</td><td headers="hd_h_ml115.tu2_1_1_1_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">STAT3</td><td headers="hd_h_ml115.tu2_1_1_1_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">No</td><td headers="hd_h_ml115.tu2_1_1_1_6" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">5.5 μM</td></tr><tr><td headers="hd_h_ml115.tu2_1_1_1_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;"><a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/932" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">932</a></td><td headers="hd_h_ml115.tu2_1_1_1_2" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;">Primary cell-based high throughput screening assay to measure STAT1 activation</td><td headers="hd_h_ml115.tu2_1_1_1_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Primary Assay (1X%ACT)</td><td headers="hd_h_ml115.tu2_1_1_1_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">STAT1</td><td headers="hd_h_ml115.tu2_1_1_1_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">No</td><td headers="hd_h_ml115.tu2_1_1_1_6" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">5.5 μM</td></tr><tr><td headers="hd_h_ml115.tu2_1_1_1_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;"><a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/1267" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">1267</a></td><td headers="hd_h_ml115.tu2_1_1_1_2" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;">Confirmation cell-based high throughput screening assay to measure STAT3 activation</td><td headers="hd_h_ml115.tu2_1_1_1_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Confirmation Assay (3X%ACT)</td><td headers="hd_h_ml115.tu2_1_1_1_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">STAT3</td><td headers="hd_h_ml115.tu2_1_1_1_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">No</td><td headers="hd_h_ml115.tu2_1_1_1_6" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">5.7 μM</td></tr><tr><td headers="hd_h_ml115.tu2_1_1_1_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;"><a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/1309" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">1309</a></td><td headers="hd_h_ml115.tu2_1_1_1_2" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;">Counterscreen assay for STAT3 activators: Cell-based high throughput assay to measure NF-kappaB activation</td><td headers="hd_h_ml115.tu2_1_1_1_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Counterscreen Assay (3X%ACT)</td><td headers="hd_h_ml115.tu2_1_1_1_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">NFκB</td><td headers="hd_h_ml115.tu2_1_1_1_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">No</td><td headers="hd_h_ml115.tu2_1_1_1_6" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">5.7 μM</td></tr><tr><td headers="hd_h_ml115.tu2_1_1_1_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;"><a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/1318" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">1318</a></td><td headers="hd_h_ml115.tu2_1_1_1_2" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;">Counterscreen assay for STAT3 activators: Cell-based high throughput assay to measure STAT1 activation</td><td headers="hd_h_ml115.tu2_1_1_1_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Counterscreen Assay (3X %ACT)</td><td headers="hd_h_ml115.tu2_1_1_1_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">STAT1</td><td headers="hd_h_ml115.tu2_1_1_1_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">No</td><td headers="hd_h_ml115.tu2_1_1_1_6" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">5.7 μM</td></tr><tr><td headers="hd_h_ml115.tu2_1_1_1_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;"><a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/1398" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">1398</a></td><td headers="hd_h_ml115.tu2_1_1_1_2" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;">Dose response cell-based assay to measure STAT3 activation</td><td headers="hd_h_ml115.tu2_1_1_1_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Dose Response (3X EC<sub>50</sub>)</td><td headers="hd_h_ml115.tu2_1_1_1_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">STAT3</td><td headers="hd_h_ml115.tu2_1_1_1_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">No</td><td headers="hd_h_ml115.tu2_1_1_1_6" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">10-point, 1:3 dilution starting at 55.7 μM</td></tr><tr><td headers="hd_h_ml115.tu2_1_1_1_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;"><a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/1406" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">1406</a></td><td headers="hd_h_ml115.tu2_1_1_1_2" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;">Dose response counterscreen assay for STAT3 activators: cell-based high throughput assay to measure STAT1 activation</td><td headers="hd_h_ml115.tu2_1_1_1_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Dose Response Counterscreen (3X EC<sub>50</sub>)</td><td headers="hd_h_ml115.tu2_1_1_1_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">STAT1</td><td headers="hd_h_ml115.tu2_1_1_1_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">No</td><td headers="hd_h_ml115.tu2_1_1_1_6" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">10-point, 1:3 dilution starting at 55.7 μM</td></tr><tr><td headers="hd_h_ml115.tu2_1_1_1_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;"><a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/1805" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">1805</a></td><td headers="hd_h_ml115.tu2_1_1_1_2" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;">Summary of probe development efforts to identify activators of signal transducer and activator of transcription 3 (STAT3)</td><td headers="hd_h_ml115.tu2_1_1_1_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Summary</td><td headers="hd_h_ml115.tu2_1_1_1_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">STAT3</td><td headers="hd_h_ml115.tu2_1_1_1_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">No</td><td headers="hd_h_ml115.tu2_1_1_1_6" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">N/A</td></tr><tr><td headers="hd_h_ml115.tu2_1_1_1_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;"><a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/2049" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">2049</a></td><td headers="hd_h_ml115.tu2_1_1_1_2" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;">Late stage results from the probe development effort to identify activators of signal transducer and activator of transcription 3 (STAT3).</td><td headers="hd_h_ml115.tu2_1_1_1_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Late Stage</td><td headers="hd_h_ml115.tu2_1_1_1_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">STAT3</td><td headers="hd_h_ml115.tu2_1_1_1_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Yes</td><td headers="hd_h_ml115.tu2_1_1_1_6" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Various</td></tr></tbody></table></div></div></article><article data-type="table-wrap" id="figobml115tu3"><div id="ml115.tu3" class="table"><h3><span class="title">Table of Assay Rationale and Screening Statistics</span></h3><p class="large-table-link" style="display:none"><span class="right"><a href="/books/NBK51964/table/ml115.tu3/?report=objectonly" target="object">View in own window</a></span></p><div class="large_tbl" id="__ml115.tu3_lrgtbl__"><table class="no_top_margin"><thead><tr><th id="hd_h_ml115.tu3_1_1_1_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:bottom;">AID</th><th id="hd_h_ml115.tu3_1_1_1_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:bottom;">Assay Rationale</th><th id="hd_h_ml115.tu3_1_1_1_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:bottom;">Assay Description</th><th id="hd_h_ml115.tu3_1_1_1_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:bottom;">Z′</th><th id="hd_h_ml115.tu3_1_1_1_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:bottom;">S:B</th></tr></thead><tbody><tr><td headers="hd_h_ml115.tu3_1_1_1_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;"><a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/871" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">871</a></td><td headers="hd_h_ml115.tu3_1_1_1_2" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;">Identify compounds that activate STAT3.</td><td headers="hd_h_ml115.tu3_1_1_1_3" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;">A human U3A fibrosarcoma cell line (deficient in STAT1) stably expressing a human STAT3-luciferase construct is incubated with test compounds in the presence of IL-6. Well luminescence is read after 6 hours. Compounds are tested in singlicate.</td><td headers="hd_h_ml115.tu3_1_1_1_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">0.84</td><td headers="hd_h_ml115.tu3_1_1_1_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">6.02</td></tr><tr><td headers="hd_h_ml115.tu3_1_1_1_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;"><a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/932" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">932</a></td><td headers="hd_h_ml115.tu3_1_1_1_2" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;">Identify compounds that activate STAT1.</td><td headers="hd_h_ml115.tu3_1_1_1_3" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;">A murine NIH 3T3 fibroblast cell line stably expressing a STAT1-luciferase construct is incubated with test compounds in the presence of IFNγ. Well luminescence is read after 6 hours. Compounds are tested in singlicate.</td><td headers="hd_h_ml115.tu3_1_1_1_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">0.81</td><td headers="hd_h_ml115.tu3_1_1_1_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">12.28</td></tr><tr><td headers="hd_h_ml115.tu3_1_1_1_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;"><a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/1267" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">1267</a></td><td headers="hd_h_ml115.tu3_1_1_1_2" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;">Confirm activity of compounds active in <a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/871" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-871</a>.</td><td headers="hd_h_ml115.tu3_1_1_1_3" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;">Same as <a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/871" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-871</a> except that compounds are tested in triplicate.</td><td headers="hd_h_ml115.tu3_1_1_1_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">0.87</td><td headers="hd_h_ml115.tu3_1_1_1_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">9.50</td></tr><tr><td headers="hd_h_ml115.tu3_1_1_1_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;"><a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/1309" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">1309</a></td><td headers="hd_h_ml115.tu3_1_1_1_2" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;">Determine whether compounds active in <a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/871" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-871</a> were nonselective due to activation of NF κB.</td><td headers="hd_h_ml115.tu3_1_1_1_3" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;">A human HEK 293T cell line stably expressing a human NFκ B::luciferase construct is incubated with test compounds in the presence of TNFα. Well luminescence is read after 6 hours. Compounds are tested in triplicate.</td><td headers="hd_h_ml115.tu3_1_1_1_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">0.85</td><td headers="hd_h_ml115.tu3_1_1_1_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">17.22</td></tr><tr><td headers="hd_h_ml115.tu3_1_1_1_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;"><a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/1318" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">1318</a></td><td headers="hd_h_ml115.tu3_1_1_1_2" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;">Determine whether compounds active in <a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/871" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-871</a> were nonselective due to activation of STAT1.</td><td headers="hd_h_ml115.tu3_1_1_1_3" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;">Same as <a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/932" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-932</a> except that compounds are tested in triplicate.</td><td headers="hd_h_ml115.tu3_1_1_1_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">0.83</td><td headers="hd_h_ml115.tu3_1_1_1_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">12.68</td></tr><tr><td headers="hd_h_ml115.tu3_1_1_1_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;"><a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/1398" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">1398</a></td><td headers="hd_h_ml115.tu3_1_1_1_2" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;">Determine STAT3 activation dose response curves for compounds active in <a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/871" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-871</a>.</td><td headers="hd_h_ml115.tu3_1_1_1_3" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;">Same as <a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/871" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-871</a> except that compounds are tested in triplicate using a 10-point dilution series starting at 55.7 μM.</td><td headers="hd_h_ml115.tu3_1_1_1_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">0.85</td><td headers="hd_h_ml115.tu3_1_1_1_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">9.15</td></tr><tr><td headers="hd_h_ml115.tu3_1_1_1_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;"><a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/1406" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">1406</a></td><td headers="hd_h_ml115.tu3_1_1_1_2" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;">Determine STAT1 activation dose response curves for compounds active in <a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/871" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-871</a>.</td><td headers="hd_h_ml115.tu3_1_1_1_3" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;">Same as <a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/932" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-932</a> except that compounds are tested in triplicate using a 10-point dilution series starting at 55.7 μM.</td><td headers="hd_h_ml115.tu3_1_1_1_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">0.84</td><td headers="hd_h_ml115.tu3_1_1_1_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">6.75</td></tr><tr><td headers="hd_h_ml115.tu3_1_1_1_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;"><a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/1805" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">1805</a></td><td headers="hd_h_ml115.tu3_1_1_1_2" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;">Summarize probe development efforts.</td><td headers="hd_h_ml115.tu3_1_1_1_3" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;">N/A</td><td headers="hd_h_ml115.tu3_1_1_1_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">N/A</td><td headers="hd_h_ml115.tu3_1_1_1_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">N/A</td></tr><tr><td headers="hd_h_ml115.tu3_1_1_1_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;"><a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/2049" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">2049</a></td><td headers="hd_h_ml115.tu3_1_1_1_2" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;">Describe the assays performed during the probe development effort.</td><td headers="hd_h_ml115.tu3_1_1_1_3" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;">STAT3 Activation Assays, STAT1 Activation Counterscreen Assays, NFkB Activation Counterscreen Assay as above, plus additional assays to determine compound cytotoxicity.</td><td headers="hd_h_ml115.tu3_1_1_1_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">N/A</td><td headers="hd_h_ml115.tu3_1_1_1_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">N/A</td></tr></tbody></table></div></div></article><article data-type="table-wrap" id="figobml115tu4"><div id="ml115.tu4" class="table"><p class="large-table-link" style="display:none"><span class="right"><a href="/books/NBK51964/table/ml115.tu4/?report=objectonly" target="object">View in own window</a></span></p><div class="large_tbl" id="__ml115.tu4_lrgtbl__"><table><thead><tr><th id="hd_h_ml115.tu4_1_1_1_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Compound</th><th id="hd_h_ml115.tu4_1_1_1_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">SR Number</th><th id="hd_h_ml115.tu4_1_1_1_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">MLS</th><th id="hd_h_ml115.tu4_1_1_1_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">CID</th><th id="hd_h_ml115.tu4_1_1_1_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">SID</th><th id="hd_h_ml115.tu4_1_1_1_6" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Solubility in PBS (µM)</th><th id="hd_h_ml115.tu4_1_1_1_7" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Michael Acceptor 100 μM GSH trap (Yes/No)</th><th id="hd_h_ml115.tu4_1_1_1_8" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Stability in PBS (t1/2 (hr)</th></tr></thead><tbody><tr><td headers="hd_h_ml115.tu4_1_1_1_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">PROBE (<a href="/pcsubstance/?term=ML115[synonym]" ref="pagearea=body&targetsite=entrez&targetcat=term&targettype=pubchem">ML115</a>)</td><td headers="hd_h_ml115.tu4_1_1_1_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">SR-01000122599</td><td headers="hd_h_ml115.tu4_1_1_1_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">MLS 000522376</td><td headers="hd_h_ml115.tu4_1_1_1_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">6619100</td><td headers="hd_h_ml115.tu4_1_1_1_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;"><a href="https://pubchem.ncbi.nlm.nih.gov/substance/14735210" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">SID-14735210</a> (liquid)<br /><a href="https://pubchem.ncbi.nlm.nih.gov/substance/87326010" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">SID-87326010</a> (powder)</td><td headers="hd_h_ml115.tu4_1_1_1_6" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;"><i>0.33</i></td><td headers="hd_h_ml115.tu4_1_1_1_7" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">No</td><td headers="hd_h_ml115.tu4_1_1_1_8" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">> 48 hr</td></tr></tbody></table></div></div></article><article data-type="table-wrap" id="figobml115tu5"><div id="ml115.tu5" class="table"><h3><span class="title">Table of Probe Properties (<a href="/pcsubstance/?term=ML115[synonym]" ref="pagearea=body&targetsite=entrez&targetcat=term&targettype=pubchem">ML115</a>)</span></h3><p class="large-table-link" style="display:none"><span class="right"><a href="/books/NBK51964/table/ml115.tu5/?report=objectonly" target="object">View in own window</a></span></p><div class="large_tbl" id="__ml115.tu5_lrgtbl__"><table class="no_top_margin"><tbody><tr><td rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">IUPAC Name</td><td rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">N-(4-chloro-2,5-dimethoxy phenyl)-5-cyclopropyl-1,2-isoxazole-3-carboxamide</td></tr><tr><td rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">MW</td><td rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">322.7436 [g/mol]</td></tr><tr><td rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">MF</td><td rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">C15H15ClN2O4</td></tr><tr><td rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">XLogP3-AA</td><td rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">2.7</td></tr><tr><td rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">H Donor</td><td rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">1</td></tr><tr><td rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">H Acceptor</td><td rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">5</td></tr><tr><td rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">Rotatable Bond Count</td><td rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">5</td></tr><tr><td rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">Tautomer Count</td><td rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">2</td></tr><tr><td rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">Exact Mass</td><td rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">322.072</td></tr><tr><td rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">MonoIsotopic Mass</td><td rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">322.072</td></tr><tr><td rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">Topological Polar surface area</td><td rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">73.6</td></tr><tr><td rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">Heavy Atom Count</td><td rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">22</td></tr><tr><td rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">Formal Charge</td><td rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">0</td></tr><tr><td rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">Complexity</td><td rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">407</td></tr><tr><td rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">Isotope Atom Count</td><td rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">0</td></tr><tr><td rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">Defined Atom Stereo Center Count; Undefined Count</td><td rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">0; 0</td></tr><tr><td rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">Defined Bond Stereo Center Count; Undefined Count</td><td rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">0; 0</td></tr><tr><td rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">Covalently-Bonded Unit Count</td><td rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">1</td></tr><tr><td rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">Vendor</td><td rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Chem Div</td></tr><tr><td rowspan="1" colspan="1" style="text-align:left;vertical-align:top;">Vendor Catalog Number</td><td rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">C226-2700</td></tr></tbody></table></div></div></article><article data-type="fig" id="figobml115fu2"><div id="ml115.fu2" class="figure"><div class="graphic"><img data-src="/books/NBK51964/bin/ml115fu2.jpg" alt="Image ml115fu2" /></div></div></article><article data-type="table-wrap" id="figobml115t1"><div id="ml115.t1" class="table"><h3><span class="label">Table 1</span><span class="title">Substituted isoxazole carboxamides as potent STAT3 activators</span></h3><p class="large-table-link" style="display:none"><span class="right"><a href="/books/NBK51964/table/ml115.t1/?report=objectonly" target="object">View in own window</a></span></p><div class="large_tbl" id="__ml115.t1_lrgtbl__"><table class="no_top_margin"><thead><tr><th id="hd_h_ml115.t1_1_1_1_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;"></th><th id="hd_h_ml115.t1_1_1_1_2" colspan="3" rowspan="1" style="text-align:center;vertical-align:middle;"><div class="graphic"><img src="/books/NBK51964/bin/ml115fu3.jpg" alt="Image ml115fu3.jpg" /></div></th></tr><tr><th headers="hd_h_ml115.t1_1_1_1_1" id="hd_h_ml115.t1_1_1_2_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">compound</th><th headers="hd_h_ml115.t1_1_1_1_2" id="hd_h_ml115.t1_1_1_2_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">R</th><th headers="hd_h_ml115.t1_1_1_1_2" id="hd_h_ml115.t1_1_1_2_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">R′</th><th headers="hd_h_ml115.t1_1_1_1_2" id="hd_h_ml115.t1_1_1_2_4" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;">STAT3 EC<sub>50</sub> (μM)</th></tr></thead><tbody><tr><td headers="hd_h_ml115.t1_1_1_1_1 hd_h_ml115.t1_1_1_2_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:top;"><b>1a</b></td><td headers="hd_h_ml115.t1_1_1_1_2 hd_h_ml115.t1_1_1_2_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:top;"><div class="graphic"><img src="/books/NBK51964/bin/ml115fu4.jpg" alt="Image ml115fu4.jpg" /></div></td><td headers="hd_h_ml115.t1_1_1_1_2 hd_h_ml115.t1_1_1_2_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:top;"><div class="graphic"><img src="/books/NBK51964/bin/ml115fu5.jpg" alt="Image ml115fu5.jpg" /></div></td><td headers="hd_h_ml115.t1_1_1_1_2 hd_h_ml115.t1_1_1_2_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:top;">0.002</td></tr><tr><td headers="hd_h_ml115.t1_1_1_1_1 hd_h_ml115.t1_1_1_2_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:top;"><b>1b</b></td><td headers="hd_h_ml115.t1_1_1_1_2 hd_h_ml115.t1_1_1_2_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:top;"><div class="graphic"><img src="/books/NBK51964/bin/ml115fu6.jpg" alt="Image ml115fu6.jpg" /></div></td><td headers="hd_h_ml115.t1_1_1_1_2 hd_h_ml115.t1_1_1_2_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:top;">-(CH<sub>2</sub>)<sub>2</sub>-CH<sub>3</sub></td><td headers="hd_h_ml115.t1_1_1_1_2 hd_h_ml115.t1_1_1_2_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:top;">0.008</td></tr><tr><td headers="hd_h_ml115.t1_1_1_1_1 hd_h_ml115.t1_1_1_2_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:top;"><b>1c</b></td><td headers="hd_h_ml115.t1_1_1_1_2 hd_h_ml115.t1_1_1_2_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:top;"><div class="graphic"><img src="/books/NBK51964/bin/ml115fu7.jpg" alt="Image ml115fu7.jpg" /></div></td><td headers="hd_h_ml115.t1_1_1_1_2 hd_h_ml115.t1_1_1_2_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:top;">-CH(CH<sub>3</sub>)<sub>2</sub></td><td headers="hd_h_ml115.t1_1_1_1_2 hd_h_ml115.t1_1_1_2_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:top;">0.001</td></tr><tr><td headers="hd_h_ml115.t1_1_1_1_1 hd_h_ml115.t1_1_1_2_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:top;"><b>1d</b></td><td headers="hd_h_ml115.t1_1_1_1_2 hd_h_ml115.t1_1_1_2_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:top;"><div class="graphic"><img src="/books/NBK51964/bin/ml115fu8.jpg" alt="Image ml115fu8.jpg" /></div></td><td headers="hd_h_ml115.t1_1_1_1_2 hd_h_ml115.t1_1_1_2_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:top;">-CH<sub>3</sub></td><td headers="hd_h_ml115.t1_1_1_1_2 hd_h_ml115.t1_1_1_2_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:top;">6.9</td></tr><tr><td headers="hd_h_ml115.t1_1_1_1_1 hd_h_ml115.t1_1_1_2_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:top;"><b>1e</b></td><td headers="hd_h_ml115.t1_1_1_1_2 hd_h_ml115.t1_1_1_2_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:top;"><div class="graphic"><img src="/books/NBK51964/bin/ml115fu9.jpg" alt="Image ml115fu9.jpg" /></div></td><td headers="hd_h_ml115.t1_1_1_1_2 hd_h_ml115.t1_1_1_2_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:top;"><div class="graphic"><img src="/books/NBK51964/bin/ml115fu10.jpg" alt="Image ml115fu10.jpg" /></div></td><td headers="hd_h_ml115.t1_1_1_1_2 hd_h_ml115.t1_1_1_2_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:top;">0.005</td></tr><tr><td headers="hd_h_ml115.t1_1_1_1_1 hd_h_ml115.t1_1_1_2_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:top;"><b>1f</b></td><td headers="hd_h_ml115.t1_1_1_1_2 hd_h_ml115.t1_1_1_2_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:top;"><div class="graphic"><img src="/books/NBK51964/bin/ml115fu11.jpg" alt="Image ml115fu11.jpg" /></div></td><td headers="hd_h_ml115.t1_1_1_1_2 hd_h_ml115.t1_1_1_2_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:top;"><div class="graphic"><img src="/books/NBK51964/bin/ml115fu12.jpg" alt="Image ml115fu12.jpg" /></div></td><td headers="hd_h_ml115.t1_1_1_1_2 hd_h_ml115.t1_1_1_2_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:top;">6.08</td></tr><tr><td headers="hd_h_ml115.t1_1_1_1_1 hd_h_ml115.t1_1_1_2_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:top;"><b>1g</b></td><td headers="hd_h_ml115.t1_1_1_1_2 hd_h_ml115.t1_1_1_2_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:top;"><div class="graphic"><img src="/books/NBK51964/bin/ml115fu13.jpg" alt="Image ml115fu13.jpg" /></div></td><td headers="hd_h_ml115.t1_1_1_1_2 hd_h_ml115.t1_1_1_2_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:top;"><div class="graphic"><img src="/books/NBK51964/bin/ml115fu14.jpg" alt="Image ml115fu14.jpg" /></div></td><td headers="hd_h_ml115.t1_1_1_1_2 hd_h_ml115.t1_1_1_2_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:top;">>5.6</td></tr></tbody></table></div></div></article><article data-type="fig" id="figobml115fu3"><div id="ml115.fu3" class="figure"><div class="graphic"><img data-src="/books/NBK51964/bin/ml115fu15.jpg" alt="Image ml115fu15" /></div></div></article><article data-type="table-wrap" id="figobml115tu6"><div id="ml115.tu6" class="table"><h3><span class="title">Summary Table of Screening Results</span></h3><p class="large-table-link" style="display:none"><span class="right"><a href="/books/NBK51964/table/ml115.tu6/?report=objectonly" target="object">View in own window</a></span></p><div class="large_tbl" id="__ml115.tu6_lrgtbl__"><table class="no_margin"><thead><tr><th id="hd_h_ml115.tu6_1_1_1_1" rowspan="2" colspan="1" headers="hd_h_ml115.tu6_1_1_1_1" style="text-align:center;vertical-align:middle;">Compound</th><th id="hd_h_ml115.tu6_1_1_1_2" rowspan="2" colspan="1" headers="hd_h_ml115.tu6_1_1_1_2" style="text-align:center;vertical-align:middle;">CID</th><th id="hd_h_ml115.tu6_1_1_1_3" rowspan="2" colspan="1" headers="hd_h_ml115.tu6_1_1_1_3" style="text-align:center;vertical-align:middle;">SID</th><th id="hd_h_ml115.tu6_1_1_1_4" rowspan="2" colspan="1" headers="hd_h_ml115.tu6_1_1_1_4" style="text-align:center;vertical-align:middle;">Target</th><th id="hd_h_ml115.tu6_1_1_1_5" rowspan="2" colspan="1" headers="hd_h_ml115.tu6_1_1_1_5" style="text-align:center;vertical-align:middle;">STAT3 EC50 (<a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/2049" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-2049</a>)</th><th id="hd_h_ml115.tu6_1_1_1_6" rowspan="2" colspan="1" headers="hd_h_ml115.tu6_1_1_1_6" style="text-align:center;vertical-align:middle;">Anti-target</th><th id="hd_h_ml115.tu6_1_1_1_7" rowspan="2" colspan="1" headers="hd_h_ml115.tu6_1_1_1_7" style="text-align:center;vertical-align:middle;">STAT1 EC50 (<a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/2049" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-2049</a>)</th><th id="hd_h_ml115.tu6_1_1_1_8" rowspan="2" colspan="1" headers="hd_h_ml115.tu6_1_1_1_8" style="text-align:center;vertical-align:middle;">STAT3 Selectivity</th><th id="hd_h_ml115.tu6_1_1_1_9" rowspan="2" colspan="1" headers="hd_h_ml115.tu6_1_1_1_9" style="text-align:center;vertical-align:middle;">BCL3 qPCR Assay</th><th id="hd_h_ml115.tu6_1_1_1_10" rowspan="2" colspan="1" headers="hd_h_ml115.tu6_1_1_1_10" style="text-align:center;vertical-align:middle;">NFκB 3X% ACT (<a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/1309" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-1309</a>)</th><th id="hd_h_ml115.tu6_1_1_1_11" colspan="2" rowspan="1" style="text-align:center;vertical-align:middle;">Viability Assay (<a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/2049" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-2049</a>)</th></tr><tr><th headers="hd_h_ml115.tu6_1_1_1_11" id="hd_h_ml115.tu6_1_1_2_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">HT-1080</th><th headers="hd_h_ml115.tu6_1_1_1_11" id="hd_h_ml115.tu6_1_1_2_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">NIH-3T3</th></tr></thead><tbody><tr><td headers="hd_h_ml115.tu6_1_1_1_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;"><b>Probe <a href="/pcsubstance/?term=ML115[synonym]" ref="pagearea=body&targetsite=entrez&targetcat=term&targettype=pubchem">ML115</a></b></td><td headers="hd_h_ml115.tu6_1_1_1_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">6619100</td><td headers="hd_h_ml115.tu6_1_1_1_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;"><a href="https://pubchem.ncbi.nlm.nih.gov/substance/14735210" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">14735210</a></td><td headers="hd_h_ml115.tu6_1_1_1_4" rowspan="8" colspan="1" style="text-align:center;vertical-align:middle;">STAT3</td><td headers="hd_h_ml115.tu6_1_1_1_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Active (2 nM)</td><td headers="hd_h_ml115.tu6_1_1_1_6" rowspan="6" colspan="1" style="text-align:center;vertical-align:middle;">STAT1</td><td headers="hd_h_ml115.tu6_1_1_1_7" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Inactive (>56 μM)</td><td headers="hd_h_ml115.tu6_1_1_1_8" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">>28,000</td><td headers="hd_h_ml115.tu6_1_1_1_9" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Active</td><td headers="hd_h_ml115.tu6_1_1_1_10" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Inactive</td><td headers="hd_h_ml115.tu6_1_1_1_11 hd_h_ml115.tu6_1_1_2_1 hd_h_ml115.tu6_1_1_2_2" colspan="2" rowspan="1" style="text-align:center;vertical-align:middle;">Inactive (>56 μM)</td></tr><tr><td headers="hd_h_ml115.tu6_1_1_1_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Analog 1</td><td headers="hd_h_ml115.tu6_1_1_1_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">5307721</td><td headers="hd_h_ml115.tu6_1_1_1_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;"><a href="https://pubchem.ncbi.nlm.nih.gov/substance/7965603" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">7965603</a></td><td headers="hd_h_ml115.tu6_1_1_1_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Active (8 nM)</td><td headers="hd_h_ml115.tu6_1_1_1_7" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Inactive (>56 μM)</td><td headers="hd_h_ml115.tu6_1_1_1_8" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">>7, 000</td><td headers="hd_h_ml115.tu6_1_1_1_9" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Active</td><td headers="hd_h_ml115.tu6_1_1_1_10" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Inactive</td><td headers="hd_h_ml115.tu6_1_1_1_11 hd_h_ml115.tu6_1_1_2_1 hd_h_ml115.tu6_1_1_2_2" colspan="2" rowspan="1" style="text-align:center;vertical-align:middle;">See below<sup>#</sup></td></tr><tr><td headers="hd_h_ml115.tu6_1_1_1_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Analog 2</td><td headers="hd_h_ml115.tu6_1_1_1_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">9550420</td><td headers="hd_h_ml115.tu6_1_1_1_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;"><a href="https://pubchem.ncbi.nlm.nih.gov/substance/14733223" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">14733223</a></td><td headers="hd_h_ml115.tu6_1_1_1_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Active (1 nM)</td><td headers="hd_h_ml115.tu6_1_1_1_7" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Inactive (>56 μM)</td><td headers="hd_h_ml115.tu6_1_1_1_8" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">>56,000</td><td headers="hd_h_ml115.tu6_1_1_1_9" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Inactive</td><td headers="hd_h_ml115.tu6_1_1_1_10" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Inactive</td><td headers="hd_h_ml115.tu6_1_1_1_11 hd_h_ml115.tu6_1_1_2_1 hd_h_ml115.tu6_1_1_2_2" colspan="2" rowspan="1" style="text-align:center;vertical-align:middle;">See below<sup>ψ</sup></td></tr><tr><td headers="hd_h_ml115.tu6_1_1_1_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Analog 3</td><td headers="hd_h_ml115.tu6_1_1_1_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">17253222</td><td headers="hd_h_ml115.tu6_1_1_1_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;"><a href="https://pubchem.ncbi.nlm.nih.gov/substance/57287818" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">57287818</a></td><td headers="hd_h_ml115.tu6_1_1_1_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Active (5 nM)</td><td headers="hd_h_ml115.tu6_1_1_1_7" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Inactive (>56 μM)</td><td headers="hd_h_ml115.tu6_1_1_1_8" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">>11,200</td><td headers="hd_h_ml115.tu6_1_1_1_9 hd_h_ml115.tu6_1_1_1_10 hd_h_ml115.tu6_1_1_1_11 hd_h_ml115.tu6_1_1_2_1 hd_h_ml115.tu6_1_1_2_2" colspan="4" rowspan="1" style="text-align:center;vertical-align:middle;">See below<sup>#</sup></td></tr><tr><td headers="hd_h_ml115.tu6_1_1_1_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Analog 4</td><td headers="hd_h_ml115.tu6_1_1_1_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">17173778</td><td headers="hd_h_ml115.tu6_1_1_1_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;"><a href="https://pubchem.ncbi.nlm.nih.gov/substance/57288067" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">57288067</a></td><td headers="hd_h_ml115.tu6_1_1_1_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Active (6980 nM)</td><td headers="hd_h_ml115.tu6_1_1_1_7" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Inactive (>56 μM)</td><td headers="hd_h_ml115.tu6_1_1_1_8" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">>8.1</td><td headers="hd_h_ml115.tu6_1_1_1_9 hd_h_ml115.tu6_1_1_1_10 hd_h_ml115.tu6_1_1_1_11 hd_h_ml115.tu6_1_1_2_1 hd_h_ml115.tu6_1_1_2_2" colspan="4" rowspan="1" style="text-align:center;vertical-align:middle;">See below<sup>#</sup></td></tr><tr><td headers="hd_h_ml115.tu6_1_1_1_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Analog 5</td><td headers="hd_h_ml115.tu6_1_1_1_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">5308825</td><td headers="hd_h_ml115.tu6_1_1_1_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;"><a href="https://pubchem.ncbi.nlm.nih.gov/substance/7967436" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">7967436</a></td><td headers="hd_h_ml115.tu6_1_1_1_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Inactive (<a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/871" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-871</a>)</td><td headers="hd_h_ml115.tu6_1_1_1_7" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Inactive (<a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/932" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-932</a>)</td><td headers="hd_h_ml115.tu6_1_1_1_8" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Cannot be determined</td><td headers="hd_h_ml115.tu6_1_1_1_9 hd_h_ml115.tu6_1_1_1_10 hd_h_ml115.tu6_1_1_1_11 hd_h_ml115.tu6_1_1_2_1 hd_h_ml115.tu6_1_1_2_2" colspan="4" rowspan="1" style="text-align:center;vertical-align:middle;">See below<sup>*</sup></td></tr><tr><td headers="hd_h_ml115.tu6_1_1_1_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Prior Art: Compound 1</td><td headers="hd_h_ml115.tu6_1_1_1_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">None</td><td headers="hd_h_ml115.tu6_1_1_1_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">None</td><td headers="hd_h_ml115.tu6_1_1_1_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Active (PMID 16115772)</td><td headers="hd_h_ml115.tu6_1_1_1_6" rowspan="1" colspan="1" style="text-align:center;vertical-align:top;">JAK and STAT5</td><td headers="hd_h_ml115.tu6_1_1_1_7" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Active (PMID 6115772)</td><td headers="hd_h_ml115.tu6_1_1_1_8" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Non-selective</td><td headers="hd_h_ml115.tu6_1_1_1_9 hd_h_ml115.tu6_1_1_1_10 hd_h_ml115.tu6_1_1_1_11 hd_h_ml115.tu6_1_1_2_1 hd_h_ml115.tu6_1_1_2_2" colspan="4" rowspan="1" style="text-align:center;vertical-align:middle;">See below<sup>^</sup></td></tr><tr><td headers="hd_h_ml115.tu6_1_1_1_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Prior Art: Compound 56</td><td headers="hd_h_ml115.tu6_1_1_1_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">2857</td><td headers="hd_h_ml115.tu6_1_1_1_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;"><a href="https://pubchem.ncbi.nlm.nih.gov/substance/26758491" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">26758491</a></td><td headers="hd_h_ml115.tu6_1_1_1_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Active (3-6 µM)</td><td headers="hd_h_ml115.tu6_1_1_1_6" rowspan="1" colspan="1" style="text-align:center;vertical-align:top;">STAT1</td><td headers="hd_h_ml115.tu6_1_1_1_7" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Active (6–12 µM)</td><td headers="hd_h_ml115.tu6_1_1_1_8" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Non-selective</td><td headers="hd_h_ml115.tu6_1_1_1_9 hd_h_ml115.tu6_1_1_1_10 hd_h_ml115.tu6_1_1_1_11 hd_h_ml115.tu6_1_1_2_1 hd_h_ml115.tu6_1_1_2_2" colspan="4" rowspan="1" style="text-align:center;vertical-align:middle;">See below<sup>†</sup></td></tr></tbody></table></div><div class="tblwrap-foot"><div><dl class="temp-labeled-list small"><dl class="bkr_refwrap"><dt>#</dt><dd><div id="ml115.tfn1"><p class="no_margin">This compound was not tested due to lower target potency compared to probe.</p></div></dd></dl><dl class="bkr_refwrap"><dt>ψ</dt><dd><div id="ml115.tfn2"><p class="no_margin">This compound was not tested due to inactivity in the qPCR assay.</p></div></dd></dl><dl class="bkr_refwrap"><dt>*</dt><dd><div id="ml115.tfn3"><p class="no_margin">This compound was not tested due to inactivity in the Primary screen.</p></div></dd></dl><dl class="bkr_refwrap"><dt>^</dt><dd><div id="ml115.tfn4"><p class="no_margin">This compound was not tested due to non-selectivity reported in PMID 16115772.</p></div></dd></dl><dl class="bkr_refwrap"><dt>†</dt><dd><div id="ml115.tfn5"><p class="no_margin">This compound was not tested due to non-selectivity as determined by the SRIMSC.</p></div></dd></dl></dl></div></div></div></article><article data-type="fig" id="figobml115fu4"><div id="ml115.fu4" class="figure"><div class="graphic"><img data-src="/books/NBK51964/bin/ml115fu16.jpg" alt="Image ml115fu16" /></div></div></article><article data-type="table-wrap" id="figobml115tu7"><div id="ml115.tu7" class="table"><h3><span class="title">STAT3 Activators SAR Table (Isoxazole Carboxamide Scaffold)</span></h3><p class="large-table-link" style="display:none"><span class="right"><a href="/books/NBK51964/table/ml115.tu7/?report=objectonly" target="object">View in own window</a></span></p><div class="large_tbl" id="__ml115.tu7_lrgtbl__"><table class="no_margin"><thead><tr><th id="hd_h_ml115.tu7_1_1_1_1" colspan="11" rowspan="1" style="text-align:center;vertical-align:middle;">Compound Information</th><th id="hd_h_ml115.tu7_1_1_1_2" colspan="7" rowspan="1" style="text-align:center;vertical-align:middle;">Screening Assays</th><th id="hd_h_ml115.tu7_1_1_1_3" colspan="5" rowspan="1" style="text-align:center;vertical-align:middle;">Probe Development Assays (<a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/2049" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-2049</a>)</th></tr><tr><th headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_1" id="hd_h_ml115.tu7_1_1_2_1" rowspan="2" colspan="1" style="text-align:center;vertical-align:middle;">Compound</th><th headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_2" id="hd_h_ml115.tu7_1_1_2_2" rowspan="2" colspan="1" style="text-align:center;vertical-align:middle;">Scripps ID</th><th headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_3" id="hd_h_ml115.tu7_1_1_2_3" rowspan="2" colspan="1" style="text-align:center;vertical-align:middle;">Structure</th><th headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_4" id="hd_h_ml115.tu7_1_1_2_4" rowspan="2" colspan="1" style="text-align:center;vertical-align:middle;">CID</th><th headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_5" id="hd_h_ml115.tu7_1_1_2_5" rowspan="2" colspan="1" style="text-align:center;vertical-align:middle;">SID</th><th headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_6" id="hd_h_ml115.tu7_1_1_2_6" rowspan="2" colspan="1" style="text-align:center;vertical-align:middle;">MLS ID</th><th headers="hd_h_ml115.tu7_1_1_1_1" id="hd_h_ml115.tu7_1_1_2_7" colspan="3" rowspan="1" style="text-align:center;vertical-align:middle;">Compound Source:</th><th headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_8" id="hd_h_ml115.tu7_1_1_2_8" rowspan="2" colspan="1" style="text-align:center;vertical-align:middle;">Vendor</th><th headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_9" id="hd_h_ml115.tu7_1_1_2_9" rowspan="2" colspan="1" style="text-align:center;vertical-align:middle;">Vendor Catalog ID</th><th headers="hd_h_ml115.tu7_1_1_1_2 hd_h_ml115.tu7_1_1_2_10" id="hd_h_ml115.tu7_1_1_2_10" rowspan="2" colspan="1" style="text-align:center;vertical-align:middle;">STAT3 Primary (<a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/871" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-871</a>) (%ACT)</th><th headers="hd_h_ml115.tu7_1_1_1_2 hd_h_ml115.tu7_1_1_2_11" id="hd_h_ml115.tu7_1_1_2_11" rowspan="2" colspan="1" style="text-align:center;vertical-align:middle;">STAT1 Primary (<a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/932" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-932</a>) (%ACT)</th><th headers="hd_h_ml115.tu7_1_1_1_2 hd_h_ml115.tu7_1_1_2_12" id="hd_h_ml115.tu7_1_1_2_12" rowspan="2" colspan="1" style="text-align:center;vertical-align:middle;">STAT3 Confirmation (<a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/1267" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-1267</a>) (%ACT)</th><th headers="hd_h_ml115.tu7_1_1_1_2 hd_h_ml115.tu7_1_1_2_13" id="hd_h_ml115.tu7_1_1_2_13" rowspan="2" colspan="1" style="text-align:center;vertical-align:middle;">NFkB Counterscreen (<a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/1309" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-1309</a>) (%ACT)</th><th headers="hd_h_ml115.tu7_1_1_1_2 hd_h_ml115.tu7_1_1_2_14" id="hd_h_ml115.tu7_1_1_2_14" rowspan="2" colspan="1" style="text-align:center;vertical-align:middle;">STAT1 Counterscreen (<a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/1318" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-1318</a>) (%ACT)</th><th headers="hd_h_ml115.tu7_1_1_1_2 hd_h_ml115.tu7_1_1_2_15" id="hd_h_ml115.tu7_1_1_2_15" rowspan="2" colspan="1" style="text-align:center;vertical-align:middle;">STAT3 EC50 (<a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/1398" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-1398</a>)</th><th headers="hd_h_ml115.tu7_1_1_1_2 hd_h_ml115.tu7_1_1_2_16" id="hd_h_ml115.tu7_1_1_2_16" rowspan="2" colspan="1" style="text-align:center;vertical-align:middle;">STAT1 EC50 (<a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/1406" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-1406</a>) (EC50)</th><th headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_17" id="hd_h_ml115.tu7_1_1_2_17" rowspan="2" colspan="1" style="text-align:center;vertical-align:middle;">STAT3 EC<sub>50</sub> (%ACT)</th><th headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_18" id="hd_h_ml115.tu7_1_1_2_18" rowspan="2" colspan="1" style="text-align:center;vertical-align:middle;">STAT1 EC<sub>50</sub> (%ACT)</th><th headers="hd_h_ml115.tu7_1_1_1_3" id="hd_h_ml115.tu7_1_1_2_19" colspan="2" rowspan="1" style="text-align:center;vertical-align:middle;">Cytotoxicity CC50 (max % Cytox)</th><th headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_20" id="hd_h_ml115.tu7_1_1_2_20" rowspan="2" colspan="1" style="text-align:center;vertical-align:middle;">qPCR Assay</th></tr><tr><th headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7" id="hd_h_ml115.tu7_1_1_3_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">MLS</th><th headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7" id="hd_h_ml115.tu7_1_1_3_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Sourced outside ML</th><th headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7" id="hd_h_ml115.tu7_1_1_3_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Chemistry: New Synth</th><th headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_19" id="hd_h_ml115.tu7_1_1_3_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">HT-1080</th><th headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_19" id="hd_h_ml115.tu7_1_1_3_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">NIH-3T3</th></tr></thead><tbody><tr><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;"><b>PROBE</b></td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">SR-01000122599</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_3" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;"><div class="graphic"><img src="/books/NBK51964/bin/ml115fu17.jpg" alt="Image ml115fu17.jpg" /></div></td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">6619100</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">14735210</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_6" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">MLS 000522376</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Yes</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Yes</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Yes</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_8" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Chem Div</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_9" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">C226-2700</td><td headers="hd_h_ml115.tu7_1_1_1_2 hd_h_ml115.tu7_1_1_2_10" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Active (159.3%)</td><td headers="hd_h_ml115.tu7_1_1_1_2 hd_h_ml115.tu7_1_1_2_11" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Inactive (99.1%)</td><td headers="hd_h_ml115.tu7_1_1_1_2 hd_h_ml115.tu7_1_1_2_12" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Active (149.9%)</td><td headers="hd_h_ml115.tu7_1_1_1_2 hd_h_ml115.tu7_1_1_2_13" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Inactive (103.2%)</td><td headers="hd_h_ml115.tu7_1_1_1_2 hd_h_ml115.tu7_1_1_2_14" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Inactive (97.9%)</td><td headers="hd_h_ml115.tu7_1_1_1_2 hd_h_ml115.tu7_1_1_2_15" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Active (>0.08 μM)</td><td headers="hd_h_ml115.tu7_1_1_1_2 hd_h_ml115.tu7_1_1_2_16" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Inactive (>55.7 μM)</td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_17" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;"><b>Active</b> 0.002 μM (97%)</td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_18" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Inactive >56 μM (17%)</td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_19 hd_h_ml115.tu7_1_1_3_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Inactive >56 μM (6.4%)</td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_19 hd_h_ml115.tu7_1_1_3_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Inactive >56 μM (8.35%)</td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_20" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Active</td></tr><tr><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Analog 1</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">SR-01000122581</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_3" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;"><div class="graphic"><img src="/books/NBK51964/bin/ml115fu18.jpg" alt="Image ml115fu18.jpg" /></div></td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">5307721</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">7965603</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_6" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">MLS 000116379</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Yes</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Yes</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Yes</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_8" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Chem Div</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_9" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">C226-3042</td><td headers="hd_h_ml115.tu7_1_1_1_2 hd_h_ml115.tu7_1_1_2_10" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Active (166.3%)</td><td headers="hd_h_ml115.tu7_1_1_1_2 hd_h_ml115.tu7_1_1_2_11" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Inactive (98.1%)</td><td headers="hd_h_ml115.tu7_1_1_1_2 hd_h_ml115.tu7_1_1_2_12" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Active (162.5%)</td><td headers="hd_h_ml115.tu7_1_1_1_2 hd_h_ml115.tu7_1_1_2_13" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Inactive (102.9%)</td><td headers="hd_h_ml115.tu7_1_1_1_2 hd_h_ml115.tu7_1_1_2_14" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Inactive (102.9%)</td><td headers="hd_h_ml115.tu7_1_1_1_2 hd_h_ml115.tu7_1_1_2_15" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Active (>0.229 μM)</td><td headers="hd_h_ml115.tu7_1_1_1_2 hd_h_ml115.tu7_1_1_2_16" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Inactive (>55.7 μM)</td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_17" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Active 0.008 μM (97%)</td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_18" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Inactive >56 μM (16%)</td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_19 hd_h_ml115.tu7_1_1_3_4 hd_h_ml115.tu7_1_1_3_5" colspan="2" rowspan="1" style="text-align:center;vertical-align:middle;">See below <sup>#</sup></td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_20" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Active</td></tr><tr><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Analog 2</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">SR-01000122754</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_3" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;"><div class="graphic"><img src="/books/NBK51964/bin/ml115fu19.jpg" alt="Image ml115fu19.jpg" /></div></td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">9550420</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">14733223</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_6" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">MLS 000520019</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Yes</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Yes</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Yes</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_8" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Chem Div</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_9" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">C226-2928</td><td headers="hd_h_ml115.tu7_1_1_1_2 hd_h_ml115.tu7_1_1_2_10" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Active (167.9%)</td><td headers="hd_h_ml115.tu7_1_1_1_2 hd_h_ml115.tu7_1_1_2_11" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Inactive (99.6%)</td><td headers="hd_h_ml115.tu7_1_1_1_2 hd_h_ml115.tu7_1_1_2_12" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Active (151.9%)</td><td headers="hd_h_ml115.tu7_1_1_1_2 hd_h_ml115.tu7_1_1_2_13" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Inactive (97%)</td><td headers="hd_h_ml115.tu7_1_1_1_2 hd_h_ml115.tu7_1_1_2_14" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Inactive (104.11%)</td><td headers="hd_h_ml115.tu7_1_1_1_2 hd_h_ml115.tu7_1_1_2_15" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Active (>0.229 μM)</td><td headers="hd_h_ml115.tu7_1_1_1_2 hd_h_ml115.tu7_1_1_2_16" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Inactive (>55.7 μM)</td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_17" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Active 0.001 μM (109%)</td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_18" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Inactive >56 μM (29%)</td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_19 hd_h_ml115.tu7_1_1_3_4 hd_h_ml115.tu7_1_1_3_5" colspan="2" rowspan="1" style="text-align:center;vertical-align:middle;">See below<sup>ψ</sup></td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_20" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Inactive</td></tr><tr><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Analog 3</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">SR-01000122601</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_3" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;"><div class="graphic"><img src="/books/NBK51964/bin/ml115fu20.jpg" alt="Image ml115fu20.jpg" /></div></td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">17253222</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">57287818</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_6" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">MLS 002473450</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">No</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Yes</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Yes</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_8" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Enamine</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_9" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">T6162698</td><td headers="hd_h_ml115.tu7_1_1_1_2 hd_h_ml115.tu7_1_1_2_10 hd_h_ml115.tu7_1_1_2_11 hd_h_ml115.tu7_1_1_2_12 hd_h_ml115.tu7_1_1_2_13 hd_h_ml115.tu7_1_1_2_14 hd_h_ml115.tu7_1_1_2_15 hd_h_ml115.tu7_1_1_2_16" colspan="7" rowspan="2" style="text-align:center;vertical-align:middle;">These compounds were not in the MLSMR collection</td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_17" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Active 0.005 μM (102%)</td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_18" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Inactive >56 μM (13%)</td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_19 hd_h_ml115.tu7_1_1_3_4 hd_h_ml115.tu7_1_1_3_5 hd_h_ml115.tu7_1_1_2_20" colspan="3" rowspan="1" style="text-align:center;vertical-align:middle;">See below <sup>#</sup></td></tr><tr><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Analog 4</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">SR-03000000868</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_3" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;"><div class="graphic"><img src="/books/NBK51964/bin/ml115fu21.jpg" alt="Image ml115fu21.jpg" /></div></td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">17173778</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">57288067</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_6" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">MLS 002473451</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">No</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Yes</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Yes</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_8" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Enamine</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_9" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">T5799570</td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_17" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Active 6.9 μM (73%)</td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_18" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Inactive >56 μM (19%)</td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_19 hd_h_ml115.tu7_1_1_3_4 hd_h_ml115.tu7_1_1_3_5 hd_h_ml115.tu7_1_1_2_20" colspan="3" rowspan="1" style="text-align:center;vertical-align:middle;">See below <sup>#</sup></td></tr><tr><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Analog 5</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">None</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_3" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;"><div class="graphic"><img src="/books/NBK51964/bin/ml115fu22.jpg" alt="Image ml115fu22.jpg" /></div></td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">5308825</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">7967436</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_6" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">MLS 000116339</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Yes</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Yes</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">No</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_8" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Chem Div</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_9" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">C226-2683</td><td headers="hd_h_ml115.tu7_1_1_1_2 hd_h_ml115.tu7_1_1_2_10" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Inactive (107%)</td><td headers="hd_h_ml115.tu7_1_1_1_2 hd_h_ml115.tu7_1_1_2_11" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Inactive (121.6%)</td><td headers="hd_h_ml115.tu7_1_1_1_2 hd_h_ml115.tu7_1_1_2_12 hd_h_ml115.tu7_1_1_2_13 hd_h_ml115.tu7_1_1_2_14 hd_h_ml115.tu7_1_1_2_15 hd_h_ml115.tu7_1_1_2_16 hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_17 hd_h_ml115.tu7_1_1_2_18 hd_h_ml115.tu7_1_1_2_19 hd_h_ml115.tu7_1_1_3_4 hd_h_ml115.tu7_1_1_3_5 hd_h_ml115.tu7_1_1_2_20" colspan="10" rowspan="1" style="text-align:center;vertical-align:middle;">See below<sup>*</sup></td></tr><tr><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Analog 6</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">SR-03000000865</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_3" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;"><div class="graphic"><img src="/books/NBK51964/bin/ml115fu23.jpg" alt="Image ml115fu23.jpg" /></div></td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">16352995</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">57288064</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_6" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Non-MLS</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">No</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Yes</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">T5640119</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_8" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Enamine</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_9" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">T5640119</td><td headers="hd_h_ml115.tu7_1_1_1_2 hd_h_ml115.tu7_1_1_2_10 hd_h_ml115.tu7_1_1_2_11 hd_h_ml115.tu7_1_1_2_12 hd_h_ml115.tu7_1_1_2_13 hd_h_ml115.tu7_1_1_2_14 hd_h_ml115.tu7_1_1_2_15 hd_h_ml115.tu7_1_1_2_16" colspan="7" rowspan="9" style="text-align:center;vertical-align:middle;">These compounds were not in the MLSMR collection</td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_17" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">9.62 μM (73%)</td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_18" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">41.4 μM (62%)</td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_19 hd_h_ml115.tu7_1_1_3_4 hd_h_ml115.tu7_1_1_3_5 hd_h_ml115.tu7_1_1_2_20" colspan="3" rowspan="9" style="text-align:center;vertical-align:middle;">See below<sup>#</sup></td></tr><tr><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Analog 7</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">SR-03000000871</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_3" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;"><div class="graphic"><img src="/books/NBK51964/bin/ml115fu24.jpg" alt="Image ml115fu24.jpg" /></div></td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">16027280</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">57288070</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_6" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Non-MLS</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">No</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Yes</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">G330-0382</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_8" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">ChemDiv</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_9" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">G330-0382</td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_17" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">>56 μM (38%)</td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_18" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">>56 μM (13%)</td></tr><tr><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Analog 8</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">SR-03000000869</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_3" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;"><div class="graphic"><img src="/books/NBK51964/bin/ml115fu25.jpg" alt="Image ml115fu25.jpg" /></div></td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">22427518</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">57288068</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_6" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Non-MLS</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">No</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Yes</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">C226-2648</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_8" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">ChemDiv</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_9" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">C226-2648</td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_17" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">>56 μM (63%)</td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_18" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">>56 μM (34%)</td></tr><tr><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Analog 9</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">SR-03000000870</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_3" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;"><div class="graphic"><img src="/books/NBK51964/bin/ml115fu26.jpg" alt="Image ml115fu26.jpg" /></div></td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">20950963</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">57288069</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_6" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Non-MLS</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">No</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Yes</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">G330-0344</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_8" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">ChemDiv</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_9" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">G330-0344</td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_17" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">>56 μM (24%)</td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_18" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">>56 μM (16%)</td></tr><tr><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Analog 10</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">SR-03000000864</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_3" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;"><div class="graphic"><img src="/books/NBK51964/bin/ml115fu27.jpg" alt="Image ml115fu27.jpg" /></div></td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">5302341</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">57288063</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_6" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Non-MLS</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">No</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Yes</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">A3975/0169384</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_8" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Chemical Block</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_9" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">A3975/0169384</td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_17" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">>56 μM (34%)</td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_18" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">>56 μM (17%)</td></tr><tr><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Analog 11</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">SR-03000001002</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_3" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;"><div class="graphic"><img src="/books/NBK51964/bin/ml115fu28.jpg" alt="Image ml115fu28.jpg" /></div></td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">24663726</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">85261423</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_6" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Non-MLS</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">No</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">No</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Yes</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_8" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">None</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_9" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">None</td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_17" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">>5.6 μM (28%)</td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_18" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">30 μM (65%)</td></tr><tr><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Analog 12</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">SR-03000001010</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_3" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;"><div class="graphic"><img src="/books/NBK51964/bin/ml115fu29.jpg" alt="Image ml115fu29.jpg" /></div></td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">40111489</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">85261424</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_6" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Non-MLS</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">No</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">No</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Yes</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_8" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">None</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_9" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">None</td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_17" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">6.08 μM (82%)</td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_18" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">3.05 μM (160%)</td></tr><tr><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Analog 13</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">SR-03000001011</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_3" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;"><div class="graphic"><img src="/books/NBK51964/bin/ml115fu30.jpg" alt="Image ml115fu30.jpg" /></div></td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">44241413</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">85261425</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_6" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Non-MLS</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">No</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">No</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Yes</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_8" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">None</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_9" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">None</td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_17" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">>5.6 μM (51%)</td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_18" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">58 μM (65%)</td></tr><tr><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Analog 14</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">SR-03000001012</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_3" rowspan="1" colspan="1" style="text-align:left;vertical-align:middle;"><div class="graphic"><img src="/books/NBK51964/bin/ml115fu31.jpg" alt="Image ml115fu31.jpg" /></div></td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">31775947</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">85261426</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_6" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Non-MLS</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">No</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">No</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_7 hd_h_ml115.tu7_1_1_3_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Yes</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_8" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">None</td><td headers="hd_h_ml115.tu7_1_1_1_1 hd_h_ml115.tu7_1_1_2_9" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">None</td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_17" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">>5.6 μM (25%)</td><td headers="hd_h_ml115.tu7_1_1_1_3 hd_h_ml115.tu7_1_1_2_18" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">5.23 μM (83%)</td></tr></tbody></table></div><div class="tblwrap-foot"><div><dl class="temp-labeled-list small"><dl class="bkr_refwrap"><dt>#</dt><dd><div id="ml115.tfn6"><p class="no_margin">This compound was not tested due to lower target potency compared to probe.</p></div></dd></dl><dl class="bkr_refwrap"><dt>ψ</dt><dd><div id="ml115.tfn7"><p class="no_margin">This compound was not tested due to inactivity in the qPCR assay.</p></div></dd></dl><dl class="bkr_refwrap"><dt>*</dt><dd><div id="ml115.tfn8"><p class="no_margin">This compound was not tested due to inactivity in the Primary screen.</p></div></dd></dl><dl class="bkr_refwrap"><dt>#</dt><dd><div id="ml115.tfn9"><p class="no_margin">These compounds were not tested due to lower target potency compared to probe.</p></div></dd></dl></dl></div></div></div></article><article data-type="fig" id="figobml115fu5"><div id="ml115.fu5" class="figure"><div class="graphic"><img data-src="/books/NBK51964/bin/ml115fu32.jpg" alt="Image ml115fu32" /></div></div></article><article data-type="fig" id="figobml115fu6"><div id="ml115.fu6" class="figure"><div class="graphic"><img data-src="/books/NBK51964/bin/ml115fu33.jpg" alt="Image ml115fu33" /></div></div></article><article data-type="fig" id="figobml115fu7"><div id="ml115.fu7" class="figure"><div class="graphic"><img data-src="/books/NBK51964/bin/ml115fu34.jpg" alt="Image ml115fu34" /></div></div></article><article data-type="table-wrap" id="figobml115tu8"><div id="ml115.tu8" class="table"><h3><span class="title">Comparative Activity Table: Probe and Prior Art Compounds</span></h3><p class="large-table-link" style="display:none"><span class="right"><a href="/books/NBK51964/table/ml115.tu8/?report=objectonly" target="object">View in own window</a></span></p><div class="large_tbl" id="__ml115.tu8_lrgtbl__"><table class="no_top_margin"><thead><tr><th id="hd_h_ml115.tu8_1_1_1_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Compound</th><th id="hd_h_ml115.tu8_1_1_1_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">CID</th><th id="hd_h_ml115.tu8_1_1_1_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">SID</th><th id="hd_h_ml115.tu8_1_1_1_4" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Target</th><th id="hd_h_ml115.tu8_1_1_1_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">STAT3 EC50 (<a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/2049" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-2049</a>)</th><th id="hd_h_ml115.tu8_1_1_1_6" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Anti-target</th><th id="hd_h_ml115.tu8_1_1_1_7" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">STAT1 EC50 (<a href="https://pubchem.ncbi.nlm.nih.gov/bioassay/2049" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">AID-2049</a>)</th><th id="hd_h_ml115.tu8_1_1_1_8" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">STAT3 Selectivity</th></tr></thead><tbody><tr><td headers="hd_h_ml115.tu8_1_1_1_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Probe <a href="/pcsubstance/?term=ML115[synonym]" ref="pagearea=body&targetsite=entrez&targetcat=term&targettype=pubchem">ML115</a></td><td headers="hd_h_ml115.tu8_1_1_1_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">6619100</td><td headers="hd_h_ml115.tu8_1_1_1_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;"><a href="https://pubchem.ncbi.nlm.nih.gov/substance/14735210" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">14735210</a></td><td headers="hd_h_ml115.tu8_1_1_1_4" rowspan="3" colspan="1" style="text-align:center;vertical-align:middle;">STAT3</td><td headers="hd_h_ml115.tu8_1_1_1_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Active (<b>2 nM</b>)</td><td headers="hd_h_ml115.tu8_1_1_1_6" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">STAT1</td><td headers="hd_h_ml115.tu8_1_1_1_7" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Inactive (<b>>56 μM</b>)</td><td headers="hd_h_ml115.tu8_1_1_1_8" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">>28,000</td></tr><tr><td headers="hd_h_ml115.tu8_1_1_1_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Prior Art: Compound 1</td><td headers="hd_h_ml115.tu8_1_1_1_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">None</td><td headers="hd_h_ml115.tu8_1_1_1_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">None</td><td headers="hd_h_ml115.tu8_1_1_1_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Active (PMID 16115772)</td><td headers="hd_h_ml115.tu8_1_1_1_6" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">JAK and STAT5</td><td headers="hd_h_ml115.tu8_1_1_1_7" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Active (PMID 6115772)</td><td headers="hd_h_ml115.tu8_1_1_1_8" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Non-selective</td></tr><tr><td headers="hd_h_ml115.tu8_1_1_1_1" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Prior Art: Compound 56</td><td headers="hd_h_ml115.tu8_1_1_1_2" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">2857</td><td headers="hd_h_ml115.tu8_1_1_1_3" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;"><a href="https://pubchem.ncbi.nlm.nih.gov/substance/26758491" ref="pagearea=body&targetsite=entrez&targetcat=link&targettype=pubchem">26758491</a></td><td headers="hd_h_ml115.tu8_1_1_1_5" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Active (3–6 µM)</td><td headers="hd_h_ml115.tu8_1_1_1_6" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">STAT1</td><td headers="hd_h_ml115.tu8_1_1_1_7" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Active (6–12 µM)</td><td headers="hd_h_ml115.tu8_1_1_1_8" rowspan="1" colspan="1" style="text-align:center;vertical-align:middle;">Non-selective</td></tr></tbody></table></div></div></article><article data-type="fig" id="figobml115fu8"><div id="ml115.fu8" class="figure"><div class="graphic"><img data-src="/books/NBK51964/bin/ml115fu35.jpg" alt="Image ml115fu35" /></div></div></article></div><div id="jr-scripts"><script src="/corehtml/pmc/jatsreader/ptpmc_3.22/js/libs.min.js"> </script><script src="/corehtml/pmc/jatsreader/ptpmc_3.22/js/jr.min.js"> </script></div></div>
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