Predictive Models for Mechanism of Action Classification from Phenotypic Assay Data – Application to Phenotypic Drug Discovery
•Download as PPTX, PDF•
2 likes•842 views
Predictive Models for Mechanism of Action Classification from Phenotypic Assay Data – Application to Phenotypic Drug Discovery Presentation at SLAS 2014 conference in San Diego, 21 January 2014
Recommended
Recommended
More Related Content
What's hot
What's hot (20)
Viewers also liked
Viewers also liked (20)
Similar to Predictive Models for Mechanism of Action Classification from Phenotypic Assay Data – Application to Phenotypic Drug Discovery
Similar to Predictive Models for Mechanism of Action Classification from Phenotypic Assay Data – Application to Phenotypic Drug Discovery (20)
Recently uploaded
Recently uploaded (20)
Predictive Models for Mechanism of Action Classification from Phenotypic Assay Data – Application to Phenotypic Drug Discovery
- 1. Predictive Models for Mechanism of Action Classification from Phenotypic Assay Data – Application to Phenotypic Drug Discovery Ellen L. Berg, PhD 21 January 2014 SLAS 2014, San Diego, CA
- 2. • Problem: - Drug discovery productivity is at an all time low - We are swimming in oceans of data • High throughput technologies • New assay models and platforms • Needed: - New tools or new approaches - Framework for integrating information Extracting Meaning from Complex Data
- 3. The Challenge of Drug Discovery Scale (meters) molecules pathways cells tissues humans 10-9 M 10-8 M 10-7 M 10-6 M 10-5 M 10-4 M 10-3 M 10-2 M 10-1 M 1 M Human exposureMolecular targets 3 • Human biology is complex • Multiple modular, highly interconnected networks
- 4. Context is Key • Target validation - Biology has a modular architecture - Function depends on “context” • Target selectivity (poly-pharmacy) - Most drugs interact with more than one target 4
- 5. BioMAP® Technology Platform BioMAP® Assay Systems Reference Profile Database Predictive Informatics Tools Standardized human primary cell disease models Database of reference profiles Analysis and data mining tools A Primary Human Cell and Co-Culture-Based Assay Platform for PDD 5
- 6. Human primary cell-based assays Tissue & disease models BioMAP® Systems – Key Features 6 • Primary human cell types • Physiologically relevant “context” - Complex activation settings - Co-cultures • Translational biomarker endpoints
- 7. Feature Mice Man Lifespan 2 Years 70 Years Size 60 g 60 kg Environment Animal facility, cage- mates Outside world, people, animals, etc. Why Human? • Key differences between mouse and man: - DNA repair mechanisms - Control of blood flow, hemostasis - Immune system status 7
- 8. • Two approaches: - (1) Measure everything • Whole genome mRNA, proteomics, metabolomics, etc. - (2) Measure what is “decision-making” • Translational biomarkers, known disease biomarkers, are downstream of multiple pathways and integrate information Why Translational Biomarkers? mRNA, epigenome Phospho-sites, intracellular proteins, metabolome Cell surface, secreted molecules 8
- 9. BioMAP Profiling: Example Profile Reference p38 MAPK Inhibitor Logexpressionratio (Drug/DMSOcontrol) Control (no drug) 99% significance envelope BioMAP Systems Readout Parameters (Biomarkers) Dose Response Cytotoxicity Readouts 9 BioMAP profiles retain shape over multiple concentrations
- 10. BioMAP Profiling: Example Profile Reference p38 MAPK Inhibitor Logexpressionratio (Drug/DMSOcontrol) 10 Activities relevant to the role of p38 in monocyte / Th1-type inflammation p38 kinase is important for Th1-dependent inflammatory responses Takanami-Ohnishi Y, et al., Essential role of p38 mitogen-activated protein kinase in contact hypersensitivity. J Biol Chem. 2002, 277:37896-903. IL-8 HLA-DR Monocyte activation IL-6IL-1aCD38 HLA-DR TNF-a
- 11. BioMAP Profiling: Example Profile Reference p38 MAPK Inhibitor Logexpressionratio (Drug/DMSOcontrol) 11 Activities relevant to anti-thrombotic effects of p38 inhibitors Tissue factor is the primary cellular initiator of coagulation p38α deficiency impairs thrombus formation Sakurai K, et al. Role of p38 mitogen-activated protein kinase in thrombus formation. J Recept Signal Transduct Res. 2004;24(4):283-96. Tissue Factor
- 12. BioMAP Profiling: Example Profile Reference p38 MAPK Inhibitor Logexpressionratio (Drug/DMSOcontrol) 12 Activities relevant to side effects – clinical finding: skin rash Upregulation of VCAM and ITAC are characteristic of skin hyperreactivity Melikoglu M, et al., Characterization of the divergent wound-healing responses occurring in the pathergy reaction and normal healthy volunteers. J Immunol. 2006, 177:6415-21. ITAC VCAM MMP1 VCAM
- 13. Similarity Analysis of Profiles • Highly correlated Similar - Pearson’s correlation of r > 0.7 • Low correlation Not similar - Pearson’s correlation of r < 0.7 13
- 14. Microtubule Stabilizers Mitochondrial ET chain Retinoids Hsp90 CDK NFkB MEK DNA synthesis JNK Protein synthesis Microtubule Destabilizers Estrogen R PI-3K Ca++ Mobilization BioMAP Data Can Cluster Compounds According to Mechanisms of Action mTOR PKC Activation p38 MAPK HMG-CoA reductase Calcineurin Transcription 14 Each circle represents a compound tested at a single dose Lines are drawn between compounds whose profiles are similar (r > 0.7) Figure adopted from Berg, JPTox Meth. 2010
- 15. Microtubule Stabilizers Mitochondrial ET chain Retinoids Hsp90 CDK NFkB MEK DNA synthesis JNK Protein synthesis Microtubule Destabilizers Estrogen R PI-3K Ca++ Mobilization BioMAP Data Can Cluster Compounds According to Mechanisms of Action mTOR PKC Activation p38 MAPK HMG-CoA reductase Calcineurin Transcription p38 MAPK Calcineurin mTOR Mitochondrial ATPase 15 Each circle represents a compound tested at a single dose Lines are drawn between compounds whose profiles are similar (r > 0.7) Figure adopted from Berg, JPTox Meth. 2010
- 16. Consensus Profiles for Mechanism Classes p38 MAPK inhibitor 1 p38 MAPK inhibitor 2 p38 MAPK inhibitor 3 • Profiles for target-selective compounds can be used to define a mechanism class (Berg, Yang & Polokoff, 2013) • Consensus profile reflects target-specific biology 16 1 1 1 1 1 1 1 1 1
- 17. Mechanism Class Consensus Profiles AhR Agonist Calcineurin Inhibitor EGFR Inhibitor EP Agonist ER Agonist GR Agonist (Full) H1 Antagonist HDAC Inhibitor HMG-CoA Reductase Inhibitor Hsp90 Inhibitor IKK2 Inhibitor IL-17A Agonist JAK Inhibitor MEK Inhibitor Microtubule Disruptor Microtubule Stabilizer Mitochondrial Inhibitor mTOR Inhibitor p38 MAPK Inhibitor PDE IV Inhibitor PI3K Inhibitor PKC (c+n) Inhibitor Proteasome Inhibitor RAR/RXR Agonist SR Ca++ ATPase Inhibitor Src Family Inhibitor TNF-alpha Antagonist Vitamin D Receptor Agonist • Dense coverage of biology • Multiple pathway classes detected per assay MechanismClasses BioMAP Assay / Endpoints CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD54/ICAM−1 CD62E/E−Selectin CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR Proliferation SRB Visual CCL2/MCP−1 CCL26/Eotaxin−3 CD106/VCAM−1 CD62P/P−selectin CD87/uPAR SRB VEGFR2 CCL2/MCP−1 CD106/VCAM−1 CD142/TissueFactor CD40 CD62E/E−Selectin CXCL8/IL−8 IL−1alpha M−CSF sPGE2 SRB sTNF−alpha CCL2/MCP−1 CD38 CD40 CD62E/E−Selectin CD69 CXCL8/IL−8 CXCL9/MIG PBMCCytotoxicity Proliferation SRB CD87/uPAR CXCL10/IP−10 CXCL9/MIG HLA−DR IL−1alpha MMP−1 PAI−I SRB TGF−betaI tPA uPA CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR IL−6 LDLR M−CSF Proliferation SerumAmyloidA SRB CD106/VCAM−1 CollagenIII CXCL10/IP−10 CXCL8/IL−8 CXCL9/MIG EGFR M−CSF MMP−1 PAI−I Proliferation_72hr SRB TIMP−1 −1.5 −1.4 −1.3 −1.2 −1.1 −1.0 −0.9 −0.8 −0.7 −0.6 −0.5 −0.4 −0.3 −0.2 −0.1 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 LogRatio Profiles Cyclopamine 40 uM Cyclopamine 13.333 u... Cyclopamine 4.444 uM Cyclopamine 1.482 uM 3C 4H LPS SAg BE3C CASM3C HDF3CGF CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD54/ICAM−1 CD62E/E−Selectin CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR Proliferation SRB Visual CCL2/MCP−1 CCL26/Eotaxin−3 CD106/VCAM−1 CD62P/P−selectin CD87/uPAR SRB VEGFR2 CCL2/MCP−1 CD106/VCAM−1 CD142/TissueFactor CD40 CD62E/E−Selectin CXCL8/IL−8 IL−1alpha M−CSF sPGE2 SRB sTNF−alpha CCL2/MCP−1 CD38 CD40 CD62E/E−Selectin CD69 CXCL8/IL−8 CXCL9/MIG PBMCCytotoxicity Proliferation SRB CD87/uPAR CXCL10/IP−10 CXCL9/MIG HLA−DR IL−1alpha MMP−1 PAI−I SRB TGF−betaI tPA uPA CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR IL−6 LDLR M−CSF Proliferation SerumAmyloidA SRB CD106/VCAM−1 CollagenIII CXCL10/IP−10 CXCL8/IL−8 CXCL9/MIG EGFR M−CSF MMP−1 PAI−I Proliferation_72hr SRB TIMP−1 −1.5 −1.4 −1.3 −1.2 −1.1 −1.0 −0.9 −0.8 −0.7 −0.6 −0.5 −0.4 −0.3 −0.2 −0.1 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 LogRatio Profiles Cyclopamine 40 uM Cyclopamine 13.333 u... Cyclopamine 4.444 uM Cyclopamine 1.482 uM 3C 4H LPS SAg BE3C CASM3C HDF3CGF CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD54/ICAM−1 CD62E/E−Selectin CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR Proliferation SRB Visual CCL2/MCP−1 CCL26/Eotaxin−3 CD106/VCAM−1 CD62P/P−selectin CD87/uPAR SRB VEGFR2 CCL2/MCP−1 CD106/VCAM−1 CD142/TissueFactor CD40 CD62E/E−Selectin CXCL8/IL−8 IL−1alpha M−CSF sPGE2 SRB sTNF−alpha CCL2/MCP−1 CD38 CD40 CD62E/E−Selectin CD69 CXCL8/IL−8 CXCL9/MIG PBMCCytotoxicity Proliferation SRB CD87/uPAR CXCL10/IP−10 CXCL9/MIG HLA−DR IL−1alpha MMP−1 PAI−I SRB TGF−betaI tPA uPA CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR IL−6 LDLR M−CSF Proliferation SerumAmyloidA SRB CD106/VCAM−1 CollagenIII CXCL10/IP−10 CXCL8/IL−8 CXCL9/MIG EGFR M−CSF MMP−1 PAI−I Proliferation_72hr SRB TIMP−1 CCL2/MCP−1 CD54/ICAM−1 −1.5 −1.4 −1.3 −1.2 −1.1 −1.0 −0.9 −0.8 −0.7 −0.6 −0.5 −0.4 −0.3 −0.2 −0.1 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 LogRatio Profiles Cyclopamine 40 uM Cyclopamine 13.333 u... Cyclopamine 4.444 uM Cyclopamine 1.482 uM 3C 4H LPS SAg BE3C CASM3C HDF3CGF K CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD54/ICAM−1 CD62E/E−Selectin CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR Proliferation SRB Visual CCL2/MCP−1 CCL26/Eotaxin−3 CD106/VCAM−1 CD62P/P−selectin CD87/uPAR SRB VEGFR2 CCL2/MCP−1 CD106/VCAM−1 CD142/TissueFactor CD40 CD62E/E−Selectin CXCL8/IL−8 IL−1alpha M−CSF sPGE2 SRB sTNF−alpha CCL2/MCP−1 CD38 CD40 CD62E/E−Selectin CD69 CXCL8/IL−8 CXCL9/MIG PBMCCytotoxicity Proliferation SRB CD87/uPAR CXCL10/IP−10 CXCL9/MIG HLA−DR IL−1alpha MMP−1 PAI−I SRB TGF−betaI tPA uPA CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR IL−6 LDLR M−CSF Proliferation SerumAmyloidA SRB CD106/VCAM−1 CollagenIII CXCL10/IP−10 CXCL8/IL−8 CXCL9/MIG EGFR M−CSF MMP−1 PAI−I Proliferation_72hr SRB TIMP−1 CCL2/MCP−1 CD54/ICAM−1 −1.5 −1.4 −1.3 −1.2 −1.1 −1.0 −0.9 −0.8 −0.7 −0.6 −0.5 −0.4 −0.3 −0.2 −0.1 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 LogRatio Profiles Cyclopamine 40 uM Cyclopamine 13.333 u... Cyclopamine 4.444 uM Cyclopamine 1.482 uM 3C 4H LPS SAg BE3C CASM3C HDF3CGF K CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD54/ICAM−1 CD62E/E−Selectin CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR Proliferation SRB Visual CCL2/MCP−1 CCL26/Eotaxin−3 CD106/VCAM−1 CD62P/P−selectin CD87/uPAR SRB VEGFR2 CCL2/MCP−1 CD106/VCAM−1 CD142/TissueFactor CD40 CD62E/E−Selectin CXCL8/IL−8 IL−1alpha M−CSF sPGE2 SRB sTNF−alpha CCL2/MCP−1 CD38 CD40 CD62E/E−Selectin CD69 CXCL8/IL−8 CXCL9/MIG PBMCCytotoxicity Proliferation SRB CD87/uPAR CXCL10/IP−10 CXCL9/MIG HLA−DR IL−1alpha MMP−1 PAI−I SRB TGF−betaI tPA uPA CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR IL−6 LDLR M−CSF Proliferation SerumAmyloidA SRB CD106/VCAM−1 CollagenIII CXCL10/IP−10 CXCL8/IL−8 CXCL9/MIG EGFR M−CSF MMP−1 PAI−I Proliferation_72hr SRB TIMP−1 CCL2/MCP−1 CD54/ICAM−1 CXCL10/IP−10 IL−1alpha MMP−9 −1.5 −1.4 −1.3 −1.2 −1.1 −1.0 −0.9 −0.8 −0.7 −0.6 −0.5 −0.4 −0.3 −0.2 −0.1 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 LogRatio Profiles Cyclopamine 40 uM Cyclopamine 13.333 u... Cyclopamine 4.444 uM Cyclopamine 1.482 uM 3C 4H LPS SAg BE3C CASM3C HDF3CGF KF3C CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD54/ICAM−1 CD62E/E−Selectin CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR Proliferation SRB Visual CCL2/MCP−1 CCL26/Eotaxin−3 CD106/VCAM−1 CD62P/P−selectin CD87/uPAR SRB VEGFR2 CCL2/MCP−1 CD106/VCAM−1 CD142/TissueFactor CD40 CD62E/E−Selectin CXCL8/IL−8 IL−1alpha M−CSF sPGE2 SRB sTNF−alpha CCL2/MCP−1 CD38 CD40 CD62E/E−Selectin CD69 CXCL8/IL−8 CXCL9/MIG PBMCCytotoxicity Proliferation SRB CD87/uPAR CXCL10/IP−10 CXCL9/MIG HLA−DR IL−1alpha MMP−1 PAI−I SRB TGF−betaI tPA uPA CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR IL−6 LDLR M−CSF Proliferation SerumAmyloidA SRB CD106/VCAM−1 CollagenIII CXCL10/IP−10 CXCL8/IL−8 CXCL9/MIG EGFR M−CSF MMP−1 PAI−I Proliferation_72hr SRB TIMP−1 CCL2/MCP−1 CD54/ICAM−1 CXCL10/IP−10 IL−1alpha MMP−9 −1.5 −1.4 −1.3 −1.2 −1.1 −1.0 −0.9 −0.8 −0.7 −0.6 −0.5 −0.4 −0.3 −0.2 −0.1 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 LogRatio Profiles Cyclopamine 40 uM Cyclopamine 13.333 u... Cyclopamine 4.444 uM Cyclopamine 1.482 uM 3C 4H LPS SAg BE3C CASM3C HDF3CGF KF3C CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD54/ICAM−1 CD62E/E−Selectin CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR Proliferation SRB Visual CCL2/MCP−1 CCL26/Eotaxin−3 CD106/VCAM−1 CD62P/P−selectin CD87/uPAR SRB VEGFR2 CCL2/MCP−1 CD106/VCAM−1 CD142/TissueFactor CD40 CD62E/E−Selectin CXCL8/IL−8 IL−1alpha M−CSF sPGE2 SRB sTNF−alpha CCL2/MCP−1 CD38 CD40 CD62E/E−Selectin CD69 CXCL8/IL−8 CXCL9/MIG PBMCCytotoxicity Proliferation SRB CD87/uPAR CXCL10/IP−10 CXCL9/MIG HLA−DR IL−1alpha MMP−1 PAI−I SRB TGF−betaI tPA uPA CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR IL−6 LDLR M−CSF Proliferation SerumAmyloidA SRB CD106/VCAM−1 CollagenIII CXCL10/IP−10 CXCL8/IL−8 CXCL9/MIG EGFR M−CSF MMP−1 PAI−I Proliferation_72hr SRB TIMP−1 CCL2/MCP−1 CD54/ICAM−1 CXCL10/IP−10 IL−1alpha MMP−9 SRB TGF−betaI −1.5 −1.4 −1.3 −1.2 −1.1 −1.0 −0.9 −0.8 −0.7 −0.6 −0.5 −0.4 −0.3 −0.2 −0.1 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 LogRatio Profiles Cyclopamine 40 uM Cyclopamine 13.333 u... Cyclopamine 4.444 uM Cyclopamine 1.482 uM 3C 4H LPS SAg BE3C CASM3C HDF3CGF KF3CT CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD54/ICAM−1 CD62E/E−Selectin CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR Proliferation SRB Visual CCL2/MCP−1 CCL26/Eotaxin−3 CD106/VCAM−1 CD62P/P−selectin CD87/uPAR SRB VEGFR2 CCL2/MCP−1 CD106/VCAM−1 CD142/TissueFactor CD40 CD62E/E−Selectin CXCL8/IL−8 IL−1alpha M−CSF sPGE2 SRB sTNF−alpha CCL2/MCP−1 CD38 CD40 CD62E/E−Selectin CD69 CXCL8/IL−8 CXCL9/MIG PBMCCytotoxicity Proliferation SRB CD87/uPAR CXCL10/IP−10 CXCL9/MIG HLA−DR IL−1alpha MMP−1 PAI−I SRB TGF−betaI tPA uPA CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR IL−6 LDLR M−CSF Proliferation SerumAmyloidA SRB CD106/VCAM−1 CollagenIII CXCL10/IP−10 CXCL8/IL−8 CXCL9/MIG EGFR M−CSF MMP−1 PAI−I Proliferation_72hr SRB TIMP−1 CCL2/MCP−1 CD54/ICAM−1 CXCL10/IP−10 IL−1alpha MMP−9 SRB TGF−betaI −1.5 −1.4 −1.3 −1.2 −1.1 −1.0 −0.9 −0.8 −0.7 −0.6 −0.5 −0.4 −0.3 −0.2 −0.1 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 LogRatio Profiles Cyclopamine 40 uM Cyclopamine 13.333 u... Cyclopamine 4.444 uM Cyclopamine 1.482 uM 3C 4H LPS SAg BE3C CASM3C HDF3CGF KF3CT CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD54/ICAM−1 CD62E/E−Selectin CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR Proliferation SRB Visual CCL2/MCP−1 CCL26/Eotaxin−3 CD106/VCAM−1 CD62P/P−selectin CD87/uPAR SRB VEGFR2 CCL2/MCP−1 CD106/VCAM−1 CD142/TissueFactor CD40 CD62E/E−Selectin CXCL8/IL−8 IL−1alpha M−CSF sPGE2 SRB sTNF−alpha CCL2/MCP−1 CD38 CD40 CD62E/E−Selectin CD69 CXCL8/IL−8 CXCL9/MIG PBMCCytotoxicity Proliferation SRB CD87/uPAR CXCL10/IP−10 CXCL9/MIG HLA−DR IL−1alpha MMP−1 PAI−I SRB TGF−betaI tPA uPA CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR IL−6 LDLR M−CSF Proliferation SerumAmyloidA SRB CD106/VCAM−1 CollagenIII CXCL10/IP−10 CXCL8/IL−8 CXCL9/MIG EGFR M−CSF MMP−1 PAI−I Proliferation_72hr SRB TIMP−1 CCL2/MCP−1 CD54/ICAM−1 CXCL10/IP−10 IL−1alpha MMP−9 SRB TGF−betaI TIMP−2 −1.5 −1.4 −1.3 −1.2 −1.1 −1.0 −0.9 −0.8 −0.7 −0.6 −0.5 −0.4 −0.3 −0.2 −0.1 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 LogRatio Profiles Cyclopamine 40 uM Cyclopamine 13.333 u... Cyclopamine 4.444 uM Cyclopamine 1.482 uM 3C 4H LPS SAg BE3C CASM3C HDF3CGF KF3CT CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD54/ICAM−1 CD62E/E−Selectin CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR Proliferation SRB Visual CCL2/MCP−1 CCL26/Eotaxin−3 CD106/VCAM−1 CD62P/P−selectin CD87/uPAR SRB VEGFR2 CCL2/MCP−1 CD106/VCAM−1 CD142/TissueFactor CD40 CD62E/E−Selectin CXCL8/IL−8 IL−1alpha M−CSF sPGE2 SRB sTNF−alpha CCL2/MCP−1 CD38 CD40 CD62E/E−Selectin CD69 CXCL8/IL−8 CXCL9/MIG PBMCCytotoxicity Proliferation SRB CD87/uPAR CXCL10/IP−10 CXCL9/MIG HLA−DR IL−1alpha MMP−1 PAI−I SRB TGF−betaI tPA uPA CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR IL−6 LDLR M−CSF Proliferation SerumAmyloidA SRB CD106/VCAM−1 CollagenIII CXCL10/IP−10 CXCL8/IL−8 CXCL9/MIG EGFR M−CSF MMP−1 PAI−I Proliferation_72hr SRB TIMP−1 CCL2/MCP−1 CD54/ICAM−1 CXCL10/IP−10 IL−1alpha MMP−9 SRB TGF−betaI TIMP−2 −1.5 −1.4 −1.3 −1.2 −1.1 −1.0 −0.9 −0.8 −0.7 −0.6 −0.5 −0.4 −0.3 −0.2 −0.1 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 LogRatio Profiles Cyclopamine 40 uM Cyclopamine 13.333 u... Cyclopamine 4.444 uM Cyclopamine 1.482 uM 3C 4H LPS SAg BE3C CASM3C HDF3CGF KF3CT CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD54/ICAM−1 CD62E/E−Selectin CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR Proliferation SRB Visual CCL2/MCP−1 CCL26/Eotaxin−3 CD106/VCAM−1 CD62P/P−selectin CD87/uPAR SRB VEGFR2 CCL2/MCP−1 CD106/VCAM−1 CD142/TissueFactor CD40 CD62E/E−Selectin CXCL8/IL−8 IL−1alpha M−CSF sPGE2 SRB sTNF−alpha CCL2/MCP−1 CD38 CD40 CD62E/E−Selectin CD69 CXCL8/IL−8 CXCL9/MIG PBMCCytotoxicity Proliferation SRB CD87/uPAR CXCL10/IP−10 CXCL9/MIG HLA−DR IL−1alpha MMP−1 PAI−I SRB TGF−betaI tPA uPA CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR IL−6 LDLR M−CSF Proliferation SerumAmyloidA SRB CD106/VCAM−1 CollagenIII CXCL10/IP−10 CXCL8/IL−8 CXCL9/MIG EGFR M−CSF MMP−1 PAI−I Proliferation_72hr SRB TIMP−1 CCL2/MCP−1 CD54/ICAM−1 CXCL10/IP−10 IL−1alpha MMP−9 SRB TGF−betaI TIMP−2 uPA −1.5 −1.4 −1.3 −1.2 −1.1 −1.0 −0.9 −0.8 −0.7 −0.6 −0.5 −0.4 −0.3 −0.2 −0.1 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 LogRatio Profiles Cyclopamine 40 uM Cyclopamine 13.333 u... Cyclopamine 4.444 uM Cyclopamine 1.482 uM 3C 4H LPS SAg BE3C CASM3C HDF3CGF KF3CT CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD54/ICAM−1 CD62E/E−Selectin CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR Proliferation SRB Visual CCL2/MCP−1 CCL26/Eotaxin−3 CD106/VCAM−1 CD62P/P−selectin CD87/uPAR SRB VEGFR2 CCL2/MCP−1 CD106/VCAM−1 CD142/TissueFactor CD40 CD62E/E−Selectin CXCL8/IL−8 IL−1alpha M−CSF sPGE2 SRB sTNF−alpha CCL2/MCP−1 CD38 CD40 CD62E/E−Selectin CD69 CXCL8/IL−8 CXCL9/MIG PBMCCytotoxicity Proliferation SRB CD87/uPAR CXCL10/IP−10 CXCL9/MIG HLA−DR IL−1alpha MMP−1 PAI−I SRB TGF−betaI tPA uPA CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR IL−6 LDLR M−CSF Proliferation SerumAmyloidA SRB CD106/VCAM−1 CollagenIII CXCL10/IP−10 CXCL8/IL−8 CXCL9/MIG EGFR M−CSF MMP−1 PAI−I Proliferation_72hr SRB TIMP−1 CCL2/MCP−1 CD54/ICAM−1 CXCL10/IP−10 IL−1alpha MMP−9 SRB TGF−betaI TIMP−2 uPA −1.5 −1.4 −1.3 −1.2 −1.1 −1.0 −0.9 −0.8 −0.7 −0.6 −0.5 −0.4 −0.3 −0.2 −0.1 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 LogRatio Profiles Cyclopamine 40 uM Cyclopamine 13.333 u... Cyclopamine 4.444 uM Cyclopamine 1.482 uM 3C 4H LPS SAg BE3C CASM3C HDF3CGF KF3CT CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD54/ICAM−1 CD62E/E−Selectin CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR Proliferation SRB Visual CCL2/MCP−1 CCL26/Eotaxin−3 CD106/VCAM−1 CD62P/P−selectin CD87/uPAR SRB VEGFR2 CCL2/MCP−1 CD106/VCAM−1 CD142/TissueFactor CD40 CD62E/E−Selectin CXCL8/IL−8 IL−1alpha M−CSF sPGE2 SRB sTNF−alpha CCL2/MCP−1 CD38 CD40 CD62E/E−Selectin CD69 CXCL8/IL−8 CXCL9/MIG PBMCCytotoxicity Proliferation SRB CD87/uPAR CXCL10/IP−10 CXCL9/MIG HLA−DR IL−1alpha MMP−1 PAI−I SRB TGF−betaI tPA uPA CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR IL−6 LDLR M−CSF Proliferation SerumAmyloidA SRB CD106/VCAM−1 CollagenIII CXCL10/IP−10 CXCL8/IL−8 CXCL9/MIG EGFR M−CSF MMP−1 PAI−I Proliferation_72hr SRB TIMP−1 CCL2/MCP−1 CD54/ICAM−1 CXCL10/IP−10 IL−1alpha MMP−9 SRB TGF−betaI TIMP−2 uPA −1.5 −1.4 −1.3 −1.2 −1.1 −1.0 −0.9 −0.8 −0.7 −0.6 −0.5 −0.4 −0.3 −0.2 −0.1 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 LogRatio Profiles Cyclopamine 40 uM Cyclopamine 13.333 u... Cyclopamine 4.444 uM Cyclopamine 1.482 uM 3C 4H LPS SAg BE3C CASM3C HDF3CGF KF3CT CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD54/ICAM−1 CD62E/E−Selectin CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR Proliferation SRB Visual CCL2/MCP−1 CCL26/Eotaxin−3 CD106/VCAM−1 CD62P/P−selectin CD87/uPAR SRB VEGFR2 CCL2/MCP−1 CD106/VCAM−1 CD142/TissueFactor CD40 CD62E/E−Selectin CXCL8/IL−8 IL−1alpha M−CSF sPGE2 SRB sTNF−alpha CCL2/MCP−1 CD38 CD40 CD62E/E−Selectin CD69 CXCL8/IL−8 CXCL9/MIG PBMCCytotoxicity Proliferation SRB CD87/uPAR CXCL10/IP−10 CXCL9/MIG HLA−DR IL−1alpha MMP−1 PAI−I SRB TGF−betaI tPA uPA CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR IL−6 LDLR M−CSF Proliferation SerumAmyloidA SRB CD106/VCAM−1 CollagenIII CXCL10/IP−10 CXCL8/IL−8 CXCL9/MIG EGFR M−CSF MMP−1 PAI−I Proliferation_72hr SRB TIMP−1 CCL2/MCP−1 CD54/ICAM−1 CXCL10/IP−10 IL−1alpha MMP−9 SRB TGF−betaI TIMP−2 uPA −1.5 −1.4 −1.3 −1.2 −1.1 −1.0 −0.9 −0.8 −0.7 −0.6 −0.5 −0.4 −0.3 −0.2 −0.1 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 LogRatio Profiles Cyclopamine 40 uM Cyclopamine 13.333 u... Cyclopamine 4.444 uM Cyclopamine 1.482 uM 3C 4H LPS SAg BE3C CASM3C HDF3CGF KF3CT CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD54/ICAM−1 CD62E/E−Selectin CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR Proliferation SRB Visual CCL2/MCP−1 CCL26/Eotaxin−3 CD106/VCAM−1 CD62P/P−selectin CD87/uPAR SRB VEGFR2 CCL2/MCP−1 CD106/VCAM−1 CD142/TissueFactor CD40 CD62E/E−Selectin CXCL8/IL−8 IL−1alpha M−CSF sPGE2 SRB sTNF−alpha CCL2/MCP−1 CD38 CD40 CD62E/E−Selectin CD69 CXCL8/IL−8 CXCL9/MIG PBMCCytotoxicity Proliferation SRB CD87/uPAR CXCL10/IP−10 CXCL9/MIG HLA−DR IL−1alpha MMP−1 PAI−I SRB TGF−betaI tPA uPA CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR IL−6 LDLR M−CSF Proliferation SerumAmyloidA SRB CD106/VCAM−1 CollagenIII CXCL10/IP−10 CXCL8/IL−8 CXCL9/MIG EGFR M−CSF MMP−1 PAI−I Proliferation_72hr SRB TIMP−1 CCL2/MCP−1 CD54/ICAM−1 CXCL10/IP−10 IL−1alpha MMP−9 SRB TGF−betaI TIMP−2 uPA −1.5 −1.4 −1.3 −1.2 −1.1 −1.0 −0.9 −0.8 −0.7 −0.6 −0.5 −0.4 −0.3 −0.2 −0.1 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 LogRatio Profiles Cyclopamine 40 uM Cyclopamine 13.333 u... Cyclopamine 4.444 uM Cyclopamine 1.482 uM 3C 4H LPS SAg BE3C CASM3C HDF3CGF KF3CT CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD54/ICAM−1 CD62E/E−Selectin CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR Proliferation SRB Visual CCL2/MCP−1 CCL26/Eotaxin−3 CD106/VCAM−1 CD62P/P−selectin CD87/uPAR SRB VEGFR2 CCL2/MCP−1 CD106/VCAM−1 CD142/TissueFactor CD40 CD62E/E−Selectin CXCL8/IL−8 IL−1alpha M−CSF sPGE2 SRB sTNF−alpha CCL2/MCP−1 CD38 CD40 CD62E/E−Selectin CD69 CXCL8/IL−8 CXCL9/MIG PBMCCytotoxicity Proliferation SRB CD87/uPAR CXCL10/IP−10 CXCL9/MIG HLA−DR IL−1alpha MMP−1 PAI−I SRB TGF−betaI tPA uPA CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR IL−6 LDLR M−CSF Proliferation SerumAmyloidA SRB CD106/VCAM−1 CollagenIII CXCL10/IP−10 CXCL8/IL−8 CXCL9/MIG EGFR M−CSF MMP−1 PAI−I Proliferation_72hr SRB TIMP−1 CCL2/MCP−1 CD54/ICAM−1 CXCL10/IP−10 IL−1alpha MMP−9 SRB TGF−betaI TIMP−2 uPA −1.5 −1.4 −1.3 −1.2 −1.1 −1.0 −0.9 −0.8 −0.7 −0.6 −0.5 −0.4 −0.3 −0.2 −0.1 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 LogRatio Profiles Cyclopamine 40 uM Cyclopamine 13.333 u... Cyclopamine 4.444 uM Cyclopamine 1.482 uM 3C 4H LPS SAg BE3C CASM3C HDF3CGF KF3CT 17
- 18. • Can we use these signatures to build classifiers? - Predictive models for specific mechanism classes would enable automated mechanism assignments • Certain mechanisms are known to be associated with certain human outcomes (safety and/or toxicity) - Automated mechanism assignment could be used as a tool to help in compound assessment and prioritization - Triage of phenotypic drug discovery hits Consensus Profiles – Phenotypic Signatures 18
- 19. • Generate reference dataset - Compounds from 28 mechanism classes • Well characterized, target selective - Test in 8 BioMAP systems • Multiple concentrations • Build a series of Two-class models using support vector machines - Use profiles in the “known” class versus a “null” set - Model output is “decision value” Automated Mechanism Class Assignment - Predictive Models 19
- 20. • Test reference data set in each model - PPV – positive predictive value, for a given decision value cut off, what fraction (percentage) of profiles are correctly classified? (=TP/(TP+FP)) - Sensitivity – for a given decision value, what percentage of profiles were assigned to the class? (=TP/(TP+FN)) Assessing Model Performance Mitochondrial Inhibitor p38 MAPK Inhibitor Berg, Yang and Polokoff, 201320
- 21. Prediction Results – p38 MAPK • Among Phase II chemicals (800) tested as part of the EPA’s ToxCast program: - The 2 named p38 MAPK inhibitors were both classified as p38 MAPK inhibitors (highest decision values were 1.01 and 0.91) - Manuscript submitted (Nicole Kleinstreuer, Keith Houck et al) • Final results will depend on disclosure of target mechanisms for compounds donated by ToxCast Pharma partners
- 22. • Screening and library characterization • Triage of hits/actives from discovery programs - Phenotypic drug discovery programs • Elucidation of Adverse (Efficacy) Outcome Pathways - Connecting initiating events (targets) with clinical outcomes Applications 22
- 23. 2323 Source Library Type Conc. % Active % Cytotoxic A Secreted proteins 10 mg/ml 18% 0% B Peptides 1 mM 7% 0.2% C Kinase 3 mM 25% 6% D Diversity 3.3 mM 31% 6% E Natural Product 5 mM 50% 11% F Kinase 1.6 mM 66% 1% Screening and Library Characterization • Phenotypic assays higher hit rates • Small molecule libraries and collections can be prioritized
- 24. Mechanism Classification for Triage of Phenotypic Actives Environmental BioactivesKinase Focused Collection Mitochondria Microtubule cAMP Elevator mTOR Proteasome AhR EGFR Other Unclassified Mit Mic cAM mT Pro AhR EGF Oth Unc • ~50% of phenotypic actives from two collections could be classified • Mitochondrial and microtubule inhibitors are common mechanisms in both sets of compounds24
- 25. Adverse Outcome Pathway Framework MIE Key Event Adverse Outcome Key Event Key Event Molecular Initiating Event Clinical Effect • Framework for integrating mode of action hypotheses to outcomes for chemical risk assessment (OECD) • Focused on the clinical outcome
- 26. Mechanism Class Consensus Profiles AhR Agonist Calcineurin Inhibitor EGFR Inhibitor EP Agonist ER Agonist GR Agonist (Full) H1 Antagonist HDAC Inhibitor HMG-CoA Reductase Inhibitor Hsp90 Inhibitor IKK2 Inhibitor IL-17A Agonist JAK Inhibitor MEK Inhibitor Microtubule Disruptor Microtubule Stabilizer Mitochondrial Inhibitor mTOR Inhibitor p38 MAPK Inhibitor PDE IV Inhibitor PI3K Inhibitor PKC (c+n) Inhibitor Proteasome Inhibitor RAR/RXR Agonist SR Ca++ ATPase Inhibitor Src Family Inhibitor TNF-alpha Antagonist Vitamin D Receptor Agonist • Dense coverage of biology • Multiple pathway classes detected per assay MechanismClasses BioMAP Assay / Endpoints CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD54/ICAM−1 CD62E/E−Selectin CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR Proliferation SRB Visual CCL2/MCP−1 CCL26/Eotaxin−3 CD106/VCAM−1 CD62P/P−selectin CD87/uPAR SRB VEGFR2 CCL2/MCP−1 CD106/VCAM−1 CD142/TissueFactor CD40 CD62E/E−Selectin CXCL8/IL−8 IL−1alpha M−CSF sPGE2 SRB sTNF−alpha CCL2/MCP−1 CD38 CD40 CD62E/E−Selectin CD69 CXCL8/IL−8 CXCL9/MIG PBMCCytotoxicity Proliferation SRB CD87/uPAR CXCL10/IP−10 CXCL9/MIG HLA−DR IL−1alpha MMP−1 PAI−I SRB TGF−betaI tPA uPA CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR IL−6 LDLR M−CSF Proliferation SerumAmyloidA SRB CD106/VCAM−1 CollagenIII CXCL10/IP−10 CXCL8/IL−8 CXCL9/MIG EGFR M−CSF MMP−1 PAI−I Proliferation_72hr SRB TIMP−1 −1.5 −1.4 −1.3 −1.2 −1.1 −1.0 −0.9 −0.8 −0.7 −0.6 −0.5 −0.4 −0.3 −0.2 −0.1 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 LogRatio Profiles Cyclopamine 40 uM Cyclopamine 13.333 u... Cyclopamine 4.444 uM Cyclopamine 1.482 uM 3C 4H LPS SAg BE3C CASM3C HDF3CGF CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD54/ICAM−1 CD62E/E−Selectin CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR Proliferation SRB Visual CCL2/MCP−1 CCL26/Eotaxin−3 CD106/VCAM−1 CD62P/P−selectin CD87/uPAR SRB VEGFR2 CCL2/MCP−1 CD106/VCAM−1 CD142/TissueFactor CD40 CD62E/E−Selectin CXCL8/IL−8 IL−1alpha M−CSF sPGE2 SRB sTNF−alpha CCL2/MCP−1 CD38 CD40 CD62E/E−Selectin CD69 CXCL8/IL−8 CXCL9/MIG PBMCCytotoxicity Proliferation SRB CD87/uPAR CXCL10/IP−10 CXCL9/MIG HLA−DR IL−1alpha MMP−1 PAI−I SRB TGF−betaI tPA uPA CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR IL−6 LDLR M−CSF Proliferation SerumAmyloidA SRB CD106/VCAM−1 CollagenIII CXCL10/IP−10 CXCL8/IL−8 CXCL9/MIG EGFR M−CSF MMP−1 PAI−I Proliferation_72hr SRB TIMP−1 −1.5 −1.4 −1.3 −1.2 −1.1 −1.0 −0.9 −0.8 −0.7 −0.6 −0.5 −0.4 −0.3 −0.2 −0.1 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 LogRatio Profiles Cyclopamine 40 uM Cyclopamine 13.333 u... Cyclopamine 4.444 uM Cyclopamine 1.482 uM 3C 4H LPS SAg BE3C CASM3C HDF3CGF CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD54/ICAM−1 CD62E/E−Selectin CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR Proliferation SRB Visual CCL2/MCP−1 CCL26/Eotaxin−3 CD106/VCAM−1 CD62P/P−selectin CD87/uPAR SRB VEGFR2 CCL2/MCP−1 CD106/VCAM−1 CD142/TissueFactor CD40 CD62E/E−Selectin CXCL8/IL−8 IL−1alpha M−CSF sPGE2 SRB sTNF−alpha CCL2/MCP−1 CD38 CD40 CD62E/E−Selectin CD69 CXCL8/IL−8 CXCL9/MIG PBMCCytotoxicity Proliferation SRB CD87/uPAR CXCL10/IP−10 CXCL9/MIG HLA−DR IL−1alpha MMP−1 PAI−I SRB TGF−betaI tPA uPA CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR IL−6 LDLR M−CSF Proliferation SerumAmyloidA SRB CD106/VCAM−1 CollagenIII CXCL10/IP−10 CXCL8/IL−8 CXCL9/MIG EGFR M−CSF MMP−1 PAI−I Proliferation_72hr SRB TIMP−1 CCL2/MCP−1 CD54/ICAM−1 −1.5 −1.4 −1.3 −1.2 −1.1 −1.0 −0.9 −0.8 −0.7 −0.6 −0.5 −0.4 −0.3 −0.2 −0.1 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 LogRatio Profiles Cyclopamine 40 uM Cyclopamine 13.333 u... Cyclopamine 4.444 uM Cyclopamine 1.482 uM 3C 4H LPS SAg BE3C CASM3C HDF3CGF K CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD54/ICAM−1 CD62E/E−Selectin CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR Proliferation SRB Visual CCL2/MCP−1 CCL26/Eotaxin−3 CD106/VCAM−1 CD62P/P−selectin CD87/uPAR SRB VEGFR2 CCL2/MCP−1 CD106/VCAM−1 CD142/TissueFactor CD40 CD62E/E−Selectin CXCL8/IL−8 IL−1alpha M−CSF sPGE2 SRB sTNF−alpha CCL2/MCP−1 CD38 CD40 CD62E/E−Selectin CD69 CXCL8/IL−8 CXCL9/MIG PBMCCytotoxicity Proliferation SRB CD87/uPAR CXCL10/IP−10 CXCL9/MIG HLA−DR IL−1alpha MMP−1 PAI−I SRB TGF−betaI tPA uPA CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR IL−6 LDLR M−CSF Proliferation SerumAmyloidA SRB CD106/VCAM−1 CollagenIII CXCL10/IP−10 CXCL8/IL−8 CXCL9/MIG EGFR M−CSF MMP−1 PAI−I Proliferation_72hr SRB TIMP−1 CCL2/MCP−1 CD54/ICAM−1 −1.5 −1.4 −1.3 −1.2 −1.1 −1.0 −0.9 −0.8 −0.7 −0.6 −0.5 −0.4 −0.3 −0.2 −0.1 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 LogRatio Profiles Cyclopamine 40 uM Cyclopamine 13.333 u... Cyclopamine 4.444 uM Cyclopamine 1.482 uM 3C 4H LPS SAg BE3C CASM3C HDF3CGF K CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD54/ICAM−1 CD62E/E−Selectin CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR Proliferation SRB Visual CCL2/MCP−1 CCL26/Eotaxin−3 CD106/VCAM−1 CD62P/P−selectin CD87/uPAR SRB VEGFR2 CCL2/MCP−1 CD106/VCAM−1 CD142/TissueFactor CD40 CD62E/E−Selectin CXCL8/IL−8 IL−1alpha M−CSF sPGE2 SRB sTNF−alpha CCL2/MCP−1 CD38 CD40 CD62E/E−Selectin CD69 CXCL8/IL−8 CXCL9/MIG PBMCCytotoxicity Proliferation SRB CD87/uPAR CXCL10/IP−10 CXCL9/MIG HLA−DR IL−1alpha MMP−1 PAI−I SRB TGF−betaI tPA uPA CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR IL−6 LDLR M−CSF Proliferation SerumAmyloidA SRB CD106/VCAM−1 CollagenIII CXCL10/IP−10 CXCL8/IL−8 CXCL9/MIG EGFR M−CSF MMP−1 PAI−I Proliferation_72hr SRB TIMP−1 CCL2/MCP−1 CD54/ICAM−1 CXCL10/IP−10 IL−1alpha MMP−9 −1.5 −1.4 −1.3 −1.2 −1.1 −1.0 −0.9 −0.8 −0.7 −0.6 −0.5 −0.4 −0.3 −0.2 −0.1 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 LogRatio Profiles Cyclopamine 40 uM Cyclopamine 13.333 u... Cyclopamine 4.444 uM Cyclopamine 1.482 uM 3C 4H LPS SAg BE3C CASM3C HDF3CGF KF3C CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD54/ICAM−1 CD62E/E−Selectin CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR Proliferation SRB Visual CCL2/MCP−1 CCL26/Eotaxin−3 CD106/VCAM−1 CD62P/P−selectin CD87/uPAR SRB VEGFR2 CCL2/MCP−1 CD106/VCAM−1 CD142/TissueFactor CD40 CD62E/E−Selectin CXCL8/IL−8 IL−1alpha M−CSF sPGE2 SRB sTNF−alpha CCL2/MCP−1 CD38 CD40 CD62E/E−Selectin CD69 CXCL8/IL−8 CXCL9/MIG PBMCCytotoxicity Proliferation SRB CD87/uPAR CXCL10/IP−10 CXCL9/MIG HLA−DR IL−1alpha MMP−1 PAI−I SRB TGF−betaI tPA uPA CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR IL−6 LDLR M−CSF Proliferation SerumAmyloidA SRB CD106/VCAM−1 CollagenIII CXCL10/IP−10 CXCL8/IL−8 CXCL9/MIG EGFR M−CSF MMP−1 PAI−I Proliferation_72hr SRB TIMP−1 CCL2/MCP−1 CD54/ICAM−1 CXCL10/IP−10 IL−1alpha MMP−9 −1.5 −1.4 −1.3 −1.2 −1.1 −1.0 −0.9 −0.8 −0.7 −0.6 −0.5 −0.4 −0.3 −0.2 −0.1 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 LogRatio Profiles Cyclopamine 40 uM Cyclopamine 13.333 u... Cyclopamine 4.444 uM Cyclopamine 1.482 uM 3C 4H LPS SAg BE3C CASM3C HDF3CGF KF3C CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD54/ICAM−1 CD62E/E−Selectin CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR Proliferation SRB Visual CCL2/MCP−1 CCL26/Eotaxin−3 CD106/VCAM−1 CD62P/P−selectin CD87/uPAR SRB VEGFR2 CCL2/MCP−1 CD106/VCAM−1 CD142/TissueFactor CD40 CD62E/E−Selectin CXCL8/IL−8 IL−1alpha M−CSF sPGE2 SRB sTNF−alpha CCL2/MCP−1 CD38 CD40 CD62E/E−Selectin CD69 CXCL8/IL−8 CXCL9/MIG PBMCCytotoxicity Proliferation SRB CD87/uPAR CXCL10/IP−10 CXCL9/MIG HLA−DR IL−1alpha MMP−1 PAI−I SRB TGF−betaI tPA uPA CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR IL−6 LDLR M−CSF Proliferation SerumAmyloidA SRB CD106/VCAM−1 CollagenIII CXCL10/IP−10 CXCL8/IL−8 CXCL9/MIG EGFR M−CSF MMP−1 PAI−I Proliferation_72hr SRB TIMP−1 CCL2/MCP−1 CD54/ICAM−1 CXCL10/IP−10 IL−1alpha MMP−9 SRB TGF−betaI −1.5 −1.4 −1.3 −1.2 −1.1 −1.0 −0.9 −0.8 −0.7 −0.6 −0.5 −0.4 −0.3 −0.2 −0.1 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 LogRatio Profiles Cyclopamine 40 uM Cyclopamine 13.333 u... Cyclopamine 4.444 uM Cyclopamine 1.482 uM 3C 4H LPS SAg BE3C CASM3C HDF3CGF KF3CT CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD54/ICAM−1 CD62E/E−Selectin CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR Proliferation SRB Visual CCL2/MCP−1 CCL26/Eotaxin−3 CD106/VCAM−1 CD62P/P−selectin CD87/uPAR SRB VEGFR2 CCL2/MCP−1 CD106/VCAM−1 CD142/TissueFactor CD40 CD62E/E−Selectin CXCL8/IL−8 IL−1alpha M−CSF sPGE2 SRB sTNF−alpha CCL2/MCP−1 CD38 CD40 CD62E/E−Selectin CD69 CXCL8/IL−8 CXCL9/MIG PBMCCytotoxicity Proliferation SRB CD87/uPAR CXCL10/IP−10 CXCL9/MIG HLA−DR IL−1alpha MMP−1 PAI−I SRB TGF−betaI tPA uPA CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR IL−6 LDLR M−CSF Proliferation SerumAmyloidA SRB CD106/VCAM−1 CollagenIII CXCL10/IP−10 CXCL8/IL−8 CXCL9/MIG EGFR M−CSF MMP−1 PAI−I Proliferation_72hr SRB TIMP−1 CCL2/MCP−1 CD54/ICAM−1 CXCL10/IP−10 IL−1alpha MMP−9 SRB TGF−betaI −1.5 −1.4 −1.3 −1.2 −1.1 −1.0 −0.9 −0.8 −0.7 −0.6 −0.5 −0.4 −0.3 −0.2 −0.1 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 LogRatio Profiles Cyclopamine 40 uM Cyclopamine 13.333 u... Cyclopamine 4.444 uM Cyclopamine 1.482 uM 3C 4H LPS SAg BE3C CASM3C HDF3CGF KF3CT CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD54/ICAM−1 CD62E/E−Selectin CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR Proliferation SRB Visual CCL2/MCP−1 CCL26/Eotaxin−3 CD106/VCAM−1 CD62P/P−selectin CD87/uPAR SRB VEGFR2 CCL2/MCP−1 CD106/VCAM−1 CD142/TissueFactor CD40 CD62E/E−Selectin CXCL8/IL−8 IL−1alpha M−CSF sPGE2 SRB sTNF−alpha CCL2/MCP−1 CD38 CD40 CD62E/E−Selectin CD69 CXCL8/IL−8 CXCL9/MIG PBMCCytotoxicity Proliferation SRB CD87/uPAR CXCL10/IP−10 CXCL9/MIG HLA−DR IL−1alpha MMP−1 PAI−I SRB TGF−betaI tPA uPA CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR IL−6 LDLR M−CSF Proliferation SerumAmyloidA SRB CD106/VCAM−1 CollagenIII CXCL10/IP−10 CXCL8/IL−8 CXCL9/MIG EGFR M−CSF MMP−1 PAI−I Proliferation_72hr SRB TIMP−1 CCL2/MCP−1 CD54/ICAM−1 CXCL10/IP−10 IL−1alpha MMP−9 SRB TGF−betaI TIMP−2 −1.5 −1.4 −1.3 −1.2 −1.1 −1.0 −0.9 −0.8 −0.7 −0.6 −0.5 −0.4 −0.3 −0.2 −0.1 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 LogRatio Profiles Cyclopamine 40 uM Cyclopamine 13.333 u... Cyclopamine 4.444 uM Cyclopamine 1.482 uM 3C 4H LPS SAg BE3C CASM3C HDF3CGF KF3CT CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD54/ICAM−1 CD62E/E−Selectin CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR Proliferation SRB Visual CCL2/MCP−1 CCL26/Eotaxin−3 CD106/VCAM−1 CD62P/P−selectin CD87/uPAR SRB VEGFR2 CCL2/MCP−1 CD106/VCAM−1 CD142/TissueFactor CD40 CD62E/E−Selectin CXCL8/IL−8 IL−1alpha M−CSF sPGE2 SRB sTNF−alpha CCL2/MCP−1 CD38 CD40 CD62E/E−Selectin CD69 CXCL8/IL−8 CXCL9/MIG PBMCCytotoxicity Proliferation SRB CD87/uPAR CXCL10/IP−10 CXCL9/MIG HLA−DR IL−1alpha MMP−1 PAI−I SRB TGF−betaI tPA uPA CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR IL−6 LDLR M−CSF Proliferation SerumAmyloidA SRB CD106/VCAM−1 CollagenIII CXCL10/IP−10 CXCL8/IL−8 CXCL9/MIG EGFR M−CSF MMP−1 PAI−I Proliferation_72hr SRB TIMP−1 CCL2/MCP−1 CD54/ICAM−1 CXCL10/IP−10 IL−1alpha MMP−9 SRB TGF−betaI TIMP−2 −1.5 −1.4 −1.3 −1.2 −1.1 −1.0 −0.9 −0.8 −0.7 −0.6 −0.5 −0.4 −0.3 −0.2 −0.1 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 LogRatio Profiles Cyclopamine 40 uM Cyclopamine 13.333 u... Cyclopamine 4.444 uM Cyclopamine 1.482 uM 3C 4H LPS SAg BE3C CASM3C HDF3CGF KF3CT CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD54/ICAM−1 CD62E/E−Selectin CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR Proliferation SRB Visual CCL2/MCP−1 CCL26/Eotaxin−3 CD106/VCAM−1 CD62P/P−selectin CD87/uPAR SRB VEGFR2 CCL2/MCP−1 CD106/VCAM−1 CD142/TissueFactor CD40 CD62E/E−Selectin CXCL8/IL−8 IL−1alpha M−CSF sPGE2 SRB sTNF−alpha CCL2/MCP−1 CD38 CD40 CD62E/E−Selectin CD69 CXCL8/IL−8 CXCL9/MIG PBMCCytotoxicity Proliferation SRB CD87/uPAR CXCL10/IP−10 CXCL9/MIG HLA−DR IL−1alpha MMP−1 PAI−I SRB TGF−betaI tPA uPA CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR IL−6 LDLR M−CSF Proliferation SerumAmyloidA SRB CD106/VCAM−1 CollagenIII CXCL10/IP−10 CXCL8/IL−8 CXCL9/MIG EGFR M−CSF MMP−1 PAI−I Proliferation_72hr SRB TIMP−1 CCL2/MCP−1 CD54/ICAM−1 CXCL10/IP−10 IL−1alpha MMP−9 SRB TGF−betaI TIMP−2 uPA −1.5 −1.4 −1.3 −1.2 −1.1 −1.0 −0.9 −0.8 −0.7 −0.6 −0.5 −0.4 −0.3 −0.2 −0.1 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 LogRatio Profiles Cyclopamine 40 uM Cyclopamine 13.333 u... Cyclopamine 4.444 uM Cyclopamine 1.482 uM 3C 4H LPS SAg BE3C CASM3C HDF3CGF KF3CT CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD54/ICAM−1 CD62E/E−Selectin CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR Proliferation SRB Visual CCL2/MCP−1 CCL26/Eotaxin−3 CD106/VCAM−1 CD62P/P−selectin CD87/uPAR SRB VEGFR2 CCL2/MCP−1 CD106/VCAM−1 CD142/TissueFactor CD40 CD62E/E−Selectin CXCL8/IL−8 IL−1alpha M−CSF sPGE2 SRB sTNF−alpha CCL2/MCP−1 CD38 CD40 CD62E/E−Selectin CD69 CXCL8/IL−8 CXCL9/MIG PBMCCytotoxicity Proliferation SRB CD87/uPAR CXCL10/IP−10 CXCL9/MIG HLA−DR IL−1alpha MMP−1 PAI−I SRB TGF−betaI tPA uPA CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR IL−6 LDLR M−CSF Proliferation SerumAmyloidA SRB CD106/VCAM−1 CollagenIII CXCL10/IP−10 CXCL8/IL−8 CXCL9/MIG EGFR M−CSF MMP−1 PAI−I Proliferation_72hr SRB TIMP−1 CCL2/MCP−1 CD54/ICAM−1 CXCL10/IP−10 IL−1alpha MMP−9 SRB TGF−betaI TIMP−2 uPA −1.5 −1.4 −1.3 −1.2 −1.1 −1.0 −0.9 −0.8 −0.7 −0.6 −0.5 −0.4 −0.3 −0.2 −0.1 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 LogRatio Profiles Cyclopamine 40 uM Cyclopamine 13.333 u... Cyclopamine 4.444 uM Cyclopamine 1.482 uM 3C 4H LPS SAg BE3C CASM3C HDF3CGF KF3CT CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD54/ICAM−1 CD62E/E−Selectin CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR Proliferation SRB Visual CCL2/MCP−1 CCL26/Eotaxin−3 CD106/VCAM−1 CD62P/P−selectin CD87/uPAR SRB VEGFR2 CCL2/MCP−1 CD106/VCAM−1 CD142/TissueFactor CD40 CD62E/E−Selectin CXCL8/IL−8 IL−1alpha M−CSF sPGE2 SRB sTNF−alpha CCL2/MCP−1 CD38 CD40 CD62E/E−Selectin CD69 CXCL8/IL−8 CXCL9/MIG PBMCCytotoxicity Proliferation SRB CD87/uPAR CXCL10/IP−10 CXCL9/MIG HLA−DR IL−1alpha MMP−1 PAI−I SRB TGF−betaI tPA uPA CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR IL−6 LDLR M−CSF Proliferation SerumAmyloidA SRB CD106/VCAM−1 CollagenIII CXCL10/IP−10 CXCL8/IL−8 CXCL9/MIG EGFR M−CSF MMP−1 PAI−I Proliferation_72hr SRB TIMP−1 CCL2/MCP−1 CD54/ICAM−1 CXCL10/IP−10 IL−1alpha MMP−9 SRB TGF−betaI TIMP−2 uPA −1.5 −1.4 −1.3 −1.2 −1.1 −1.0 −0.9 −0.8 −0.7 −0.6 −0.5 −0.4 −0.3 −0.2 −0.1 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 LogRatio Profiles Cyclopamine 40 uM Cyclopamine 13.333 u... Cyclopamine 4.444 uM Cyclopamine 1.482 uM 3C 4H LPS SAg BE3C CASM3C HDF3CGF KF3CT CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD54/ICAM−1 CD62E/E−Selectin CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR Proliferation SRB Visual CCL2/MCP−1 CCL26/Eotaxin−3 CD106/VCAM−1 CD62P/P−selectin CD87/uPAR SRB VEGFR2 CCL2/MCP−1 CD106/VCAM−1 CD142/TissueFactor CD40 CD62E/E−Selectin CXCL8/IL−8 IL−1alpha M−CSF sPGE2 SRB sTNF−alpha CCL2/MCP−1 CD38 CD40 CD62E/E−Selectin CD69 CXCL8/IL−8 CXCL9/MIG PBMCCytotoxicity Proliferation SRB CD87/uPAR CXCL10/IP−10 CXCL9/MIG HLA−DR IL−1alpha MMP−1 PAI−I SRB TGF−betaI tPA uPA CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR IL−6 LDLR M−CSF Proliferation SerumAmyloidA SRB CD106/VCAM−1 CollagenIII CXCL10/IP−10 CXCL8/IL−8 CXCL9/MIG EGFR M−CSF MMP−1 PAI−I Proliferation_72hr SRB TIMP−1 CCL2/MCP−1 CD54/ICAM−1 CXCL10/IP−10 IL−1alpha MMP−9 SRB TGF−betaI TIMP−2 uPA −1.5 −1.4 −1.3 −1.2 −1.1 −1.0 −0.9 −0.8 −0.7 −0.6 −0.5 −0.4 −0.3 −0.2 −0.1 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 LogRatio Profiles Cyclopamine 40 uM Cyclopamine 13.333 u... Cyclopamine 4.444 uM Cyclopamine 1.482 uM 3C 4H LPS SAg BE3C CASM3C HDF3CGF KF3CT CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD54/ICAM−1 CD62E/E−Selectin CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR Proliferation SRB Visual CCL2/MCP−1 CCL26/Eotaxin−3 CD106/VCAM−1 CD62P/P−selectin CD87/uPAR SRB VEGFR2 CCL2/MCP−1 CD106/VCAM−1 CD142/TissueFactor CD40 CD62E/E−Selectin CXCL8/IL−8 IL−1alpha M−CSF sPGE2 SRB sTNF−alpha CCL2/MCP−1 CD38 CD40 CD62E/E−Selectin CD69 CXCL8/IL−8 CXCL9/MIG PBMCCytotoxicity Proliferation SRB CD87/uPAR CXCL10/IP−10 CXCL9/MIG HLA−DR IL−1alpha MMP−1 PAI−I SRB TGF−betaI tPA uPA CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR IL−6 LDLR M−CSF Proliferation SerumAmyloidA SRB CD106/VCAM−1 CollagenIII CXCL10/IP−10 CXCL8/IL−8 CXCL9/MIG EGFR M−CSF MMP−1 PAI−I Proliferation_72hr SRB TIMP−1 CCL2/MCP−1 CD54/ICAM−1 CXCL10/IP−10 IL−1alpha MMP−9 SRB TGF−betaI TIMP−2 uPA −1.5 −1.4 −1.3 −1.2 −1.1 −1.0 −0.9 −0.8 −0.7 −0.6 −0.5 −0.4 −0.3 −0.2 −0.1 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 LogRatio Profiles Cyclopamine 40 uM Cyclopamine 13.333 u... Cyclopamine 4.444 uM Cyclopamine 1.482 uM 3C 4H LPS SAg BE3C CASM3C HDF3CGF KF3CT CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD54/ICAM−1 CD62E/E−Selectin CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR Proliferation SRB Visual CCL2/MCP−1 CCL26/Eotaxin−3 CD106/VCAM−1 CD62P/P−selectin CD87/uPAR SRB VEGFR2 CCL2/MCP−1 CD106/VCAM−1 CD142/TissueFactor CD40 CD62E/E−Selectin CXCL8/IL−8 IL−1alpha M−CSF sPGE2 SRB sTNF−alpha CCL2/MCP−1 CD38 CD40 CD62E/E−Selectin CD69 CXCL8/IL−8 CXCL9/MIG PBMCCytotoxicity Proliferation SRB CD87/uPAR CXCL10/IP−10 CXCL9/MIG HLA−DR IL−1alpha MMP−1 PAI−I SRB TGF−betaI tPA uPA CCL2/MCP−1 CD106/VCAM−1 CD141/Thrombomodulin CD142/TissueFactor CD87/uPAR CXCL8/IL−8 CXCL9/MIG HLA−DR IL−6 LDLR M−CSF Proliferation SerumAmyloidA SRB CD106/VCAM−1 CollagenIII CXCL10/IP−10 CXCL8/IL−8 CXCL9/MIG EGFR M−CSF MMP−1 PAI−I Proliferation_72hr SRB TIMP−1 CCL2/MCP−1 CD54/ICAM−1 CXCL10/IP−10 IL−1alpha MMP−9 SRB TGF−betaI TIMP−2 uPA −1.5 −1.4 −1.3 −1.2 −1.1 −1.0 −0.9 −0.8 −0.7 −0.6 −0.5 −0.4 −0.3 −0.2 −0.1 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 LogRatio Profiles Cyclopamine 40 uM Cyclopamine 13.333 u... Cyclopamine 4.444 uM Cyclopamine 1.482 uM 3C 4H LPS SAg BE3C CASM3C HDF3CGF KF3CT 26 VCAM
- 27. An Adverse Outcome Pathway for Skin Rash MIE Key Event Adverse Outcome Inhibition of p38 MAPK Upregulation of VCAM Skin Rash MIE Inhibition of MEK Inflammatory Cell Recruitment? Key Event Key Event JNK Pathway Activation? Molecular Initiating Event Clinical Effect HDF3CGF In vitro disease model
- 28. • Phenotypic data sets from primary human cell and co- culture models can be used to classify mechanisms of action - Assays are sufficiently reproducible - Mechanisms are distinguishable • Applications - Screening & Library characterization - Triage of discovery program hits - Outcome pathway knowledge - Phenotypic drug discovery Summary 28
- 29. • Using biological systems to discover new drugs - Target agnostic approach • Neoclassic Drug Discovery - The combination of using biologically complex model systems & high throughput approaches (JAL & EB, 2013) - Screening assays that are extraordinarily well characterized • Tool compounds • Omics and genetic technologies - Integration of target-based and phenotypic drug discovery • Please attend: - Phenotypic Drug Discovery SIG, Wednesday 8:00 AM Phenotypic Drug Discovery 29
- 30. DiscoveRx Solutions: an Integrated Portfolio HDAC/HMT DepthofCoverage 100% 0% 284 389 21 23 40 GPCRs Kinases NHRs Pathways Bromodomains Primary Human Cell Profiling 1,118 assays covering 741 druggable targets 30
- 31. • BioSeek - Mark A. Polokoff - Alison O’Mahony - Jian Yang - Antal Berenyi Acknowledgements • EPA - Keith Houck - Nicole Kleinstreuer 31
- 32. BioSeek, A Division of DiscoveRx 310 Utah, Suite 100 South San Francisco, CA 94030 650-416-7600 Ellen L. Berg, PhD eberg@bioseekinc.com www.biomapsystems.com CONTACTS
Editor's Notes
- Depth of coverage in all target classes, with the largest menus of any provider in both GPCRs and Kinases unparalleled in the industry.Focus on expanding our pathway signaling portfolio and several additional technology platforms under development in R&D.