SBS conference update

1. SBS conference update

2. The search for the cure • Involves 3 major players: • The Target or miscreant • The Compound • The Assay • And several disciplines: • Biologist • Chemist, Cheminformatician • Bioinformatician • Information technologist • Biophysicist

3. Notable talks • Assays and automation by Ricardo Macarron(Glaxo Smithkline) • High throughput screening (HTS) timeline: • 1986: HTS birth • 1987: SPA (Scintillation proximity assay) • 1993: Haystack, first HTS robot • 1994: SBS is founded • 1995: FLIPR • 1996: 384 well-plate • 1997: Lipinski’s rule of five. Rule of thumb to evaluate druglikeness • 2005: NIH roadmap; HTS in academia • Drugs represent 10% of healthcare costs!!

4. Notable talks • 2. Oncology drug modifier screens in live cells by Hakim Djaballah(Memorial Sloan-Kettering) • Highly optimistic; used personalized medicine approach • Tumor cells from an relapsed/refractory acute myeloid leukemia (AML) patient screened against established chemotherapeutic and novel compounds • Patient cells sensitive to 6-thioguanine in screening (IC50=70 nM). • Same drug was administered to patient, who responded and was cancer-free for 8 months with no bone marrow transplants

5. 3. Promiscuity and PubChem by CristianBologa, (University of New Mexico) • He is analyzing the compounds in MLSMR to see what makes one a promiscuous binder vs. a selective probe. • He has scaffold bioactivity classes: 1) Not active (yet), 2) selective-on multiple targets from same family and, 3) privileged and 4) promiscuous-active on everything. • He analyzed 417 assays which exclude the physico-chemical profiling assays, but include the cytotoxic assays. He generated assay statistics about Screening center, Organism, Format, Assay type, Target, and Detection method.  • He also analyzed assays by 1) spectroscopic profiling, 2) Redox cycling Profiling, 3) Aggregate profiling, etc. • In the future, he wants to look at “scaffold vs. substrate filters, Promiscuity patterns for different compounds”, and so on.

6. 4. Natural product inspired strategies for hit-finding activitiesby Jean-Yves Ortholand, EDELRIS, France. Registered compounds: 50 million (in CAS registry 2010) Enumerated number: 109   -up to 30 heavy atoms Potentially: 1020-1060 29 providers with 30% redundancy! Known chemical space: 50 ml worth of sand Total chemical space: 5 Km long, 400 m wide, 50 m deep Natural products (NP): 1.Years of validity  2. Passed evolution Hence looking into 3 types: NP-derived, NP-inspired and NP-like NP-inspired and NP-like: tractability, analogability, addressing new targets. • Chirality: 3D shape • Rings and scaffolds: 83% of rings in NP are not commercially available (1-3 aromatic rings) • Specific atom proportion: Higher O and N content. In alcohols, amines, amides • Complexity: this increases binding affinity Dioxaspiro motif: A potential scaffold found in many NP

7. Public databases (Steve Bryant): Largest repository of small molecule screening data. Created in 2003; went public in 2004. Has 115 contributing organizations; ~70 million substances. ChemBank Psychoactive drug screening program (PDSP) EU Open Screen

8. Problems with PubChem data • Very generic data repository: Flexible to put data in, difficult to retrieve and analyze • Few annotations of assay technologies and the underlying biology • No standardization of data • Input data format varies based on the depositor: some assays lack enough information for further analysis

9. 2. ChemBank (Joshua Bittker): A collaboration between the Chemical Biology Program and Platform at the Broad Institute of Harvard and MIT. • ChemBank used to store data from assays performed at the Broad Institute screening center. • Has two components: the Public and Data sharing agreement- accessible only to the Harvard community and their collaborators. • They are annotating PubChem compounds using a Java-based tool (Calypso?). Their search page contains these columns : • Activity Outcome: Probe, Active, Inactive, Unspecified, inconclusive, Discrepant, Untested, Collapsed • BioAssay type: Primary, Confirmatory, Summary, Other • Summary Data Table: Shows the compound cluster of the input assays

10. 3. Psychoactive drug screening program (PDSP, Vincent Setola) • Provides screening of novel psychoactive compounds for pharmacological and functional activity at cloned human or rodent CNS receptors, channels, and transporters. • Targeting the Recepterome: GPCRs that account for ~5% of human genome; have 7 transmembrane domains; 50% of drugs in the market target GPCRs. • Valvular heart disease: A serious side-effect of some drugs that cause fibrosis, valve leakage and death. Drugs such as fenfluramine (anorexigen), pergolide and cabergoline (Parkinson’s drugs) act on 5-hydroxytryptamine (HT)2B receptors on the heart. • Recent screen identified 27 of the 2200 FDA approved drugs to be 5-HT2B receptor agonists!

11. 4. EU Open screen (Simone Graber) www.eu-openscreen.eu European Strategy Forum on Research Infrastructures (ESFRI)- initiated in 2002; European Chemical Biology Database: (ECBD); Chembionet (list of German companies) An association of high throughput screening (HTS) centers that offer chemical resources for hit discovery and optimization, bio- and cheminformatics support, and a publicly accessible database. A central facility will make available a large collection of diverse compounds representing the chemical knowledge of Europe

12. For data to be easily retrieved, analyzed, compared and integrated, standard formats and controlled vocabularies are necessary • Existing PubChem data lacks this organization • Our group is creating controlled vocabulary and annotating PubChem bioassays Why annotate?

13. What do we need to describe? • BioAssay specifications • BioAssay relationships

14. Components and Specifications • Format: The assay system used: cell-based or cell free, treatments, media, etc. • Perturbagen: Chemical compounds, siRNAs, etc tested by the assay for their ability to produce a measurable change. • Purpose: Abstract concept representing the reasons for conducting the assay. • Technology: The underlying method used to determine the action of the perturbagen. • MetaTarget: The presumed subject of the perturbagen: protein, gene, or RNA target, a process, a signaling pathway, phenotype, etc. • Endpoint: A metric used by the assay to quantitate or qualify activity: IC50, response at defined conc.

16. Other 1% Organism Based 10% Assay Formats in PubChem Formats of 1924 assays Biochemical 43% Cell Based 46 %

17. Bacterium 4% Cell Based Eukaryote 96% Pathogenic organisms 11% Sub-cellular organelles 3 % Other 6 % Other 1% Biochemical Organism Based Yeast 42 % Purified protein 96 % Mouse 41%

18. Figure xx:Assay formats of the PubChem bioassays. Majority of the assays were either cell based (888) or biochemical (822), followed by a good number of organism based assays (202) and a few in the others category (12), which include viruses and ex vivo tissues, A. The cell based assays were predominantly performed on eukaryotic cells (853) and a smaller proportion on pathogenic bacterial cells (35), B. Most of the biochemical assays were performed with purified protein samples (785) and a smaller proportion with sub-cellular components (26), including cell lysate, cell membrane, nuclear extract, microsomes, or mitochondria and others (11), which include tissue homogenate, yeast extract, and plasma, C. Among the organism based assays, mouse (83) and yeast (85) were the preferred species for the compound testing, followed by other model organisms (11), namely, C. elegans, Drosophila melanogaster, sea urchin and zebrafish, and pathogenic organisms (23), namely, Leishmania, Plasmodium falciparum, Salmonella typhi and Trypanosomacruzi, D.

19. Advantages of Label-Free Assays: • Direct measurement of binding interactions independent of functional activity • Direct measurement of functional activity without modification of the binding partners with labels • Measurement of binding interactions where natural ligand is unknown • Rapid assay development • Cell-Based GPCR assays can be conducted without the need for engineering cell-lines to over-express given receptors • Ability to work with intractable targets where no commercially available assay exists • Noninvasive measurement enables acquiring both real-time kinetic and endpoint measurements

20. Assay Technology • Technology category: the nature of the experimental technique used to measure the effect of the perturbagen on the target • Binding reporter technology • Immunoassay • Energy transfer • Luminescence proximity • Rotational motion • Optical sensor • Enzyme reporter technology • ATP-coupled enzyme • Luciferin-coupled enzyme • Chaperone-coupled enzyme • Inducible reporter technology • Beta lactamase reporter • Luciferase reporter • LacZ reporter • GFP reporter • Viability reporter technology • Caspase activity • Dehydrogenase activity • ATP content, luciferase coupled • Membrane content • Nucleic acid content • Protease activity • Mitochondrial succinate dehydrogenase

21. Redistribution reporter technology • Calcium redistribution • cAMP redistribution • GFP tag redistribution • Metal ion redistribution • Label free/Affinity based technologies • Impedance • Optical sensor • Thermal shift • NMR • X-ray crystallography • Mass spectrometry • DNA encoding • Acoustic technology • Automated microscopy • Fiber optic waveguide measurement • Light scattering (flow cytometry)