Molecular Libraries ImplementationAnd Imaging Initiative (MLI)

Presentation for the Council of CouncilsFrancis S. Collins, M.D., Ph.D.

March 31, 2008

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Molecular Libraries Initiative: Rationale

Urgent need to determine function of genes, proteins, and pathways

Small molecules are complementary to molecular genetic tools such as siRNASmall molecules generally act on a protein target, therefore most proximate to physiologyCan act as agonists as well as antagonistsReversible in real time

Urgent need to catalyze development of therapeutics for rare and orphan diseases

>6000 rare diseasesGenetic basis of many knownTreatments available for <100

Molecular Libraries Initiative made possible by recent convergent developments

HTOS

CompoundLibraries

Robotic TechnologyHuman

Genome Project

Availability of targets

Availability of screening

Availability of compounds

Public sector screening and chemistry initiative

AssayTechnology

Prob

abili

ty

of s

ucce

ss

Public

Pre-MLI

Cum

ulat

ive

Cos

t

Sector Science

Dedicated Compound Chem-Biol accepted into

Clinical Project Team formed Developme

1 yr 1 yr 1 yr ~ 3 yrs 1 yr 2 yrs ~3 yrs 1.5 yrs IndefiniteIndefinite

nt

Lead Ph I Ph II Ph III Target Regulatory Ph IV-VDevelopment, identification review (Additional Assay Optimization indications, Screening develop- Clinical Trials Safety (HTS or Hit-to- mentmonitoring)otherwise) Probe

How does Molecular Libraries relate to drug development?

Public Sector

Science with MLI

Prob

abili

ty

of s

ucce

ssC

umul

ativ

e C

ost

Ph IV-V(Additional indications, Safety monitoring)

Dedicated Chem-Biol

Project Team formed

Compound accepted into

Clinical Development

Target identification

Assay develop- ment Hit-to-

Probe

Screening (HTS or otherwise)

1 yr 1 yr 1 yr ~ 3 yrs 1 yr 2 yrs ~3 yrs

Ph III (Efficacy and safety in large populations)

Ph II (Dose finding, initial efficacy

in patient pop.)

Ph I (Safety)

Lead Development, Optimization

Indefinite Indefinite1.5 yrs

Regulatory review

How does Molecular Libraries relate to drug development?

Molecular LibrariesScreening Centers

Network (MLSCN)

Compound Repository(MLSMR)

Instrumentation

ChemicalDiversity

AssayDevelopment

PredictiveADMET

Technology Development

Screening/Data Production

Data Analysis/ Dissemination

CheminformaticsResearch Centers

The Molecular Libraries and Imaging Roadmap: An Integrated Initiative

The Molecular Libraries Screening Center Network

Three year pilot phase:

An NIH-academic partnership,

Network of academic initiatives providing improved tools and resources for discovery of chemical probes,

Contains a cooperative of 10 Screening Centers performing probe discovery on a common chemical library (MLSMR) and publishing results to the public chemical biology database, PubChem,

Purpose is to enhance translational progress towards improvements in human health.

MLSCN Center at Columbia University

Emory Chemistry-Biology Center in the MLSCN

Southern Research Molecular Libraries Screening Center (SRMLSC)

San Diego Center for Chemical Genomics(Burnham Institute)

Scripps Research Institute Molecular Screening Center

New Mexico Molecular Libraries Screening Center

The Penn Center for Molecular Discovery

University of Pittsburgh Molecular Libraries Screening Center

Vanderbilt Screening Center for GPCRs, Ion Channels, and Transporters

NIH Chemical Genomics Center (NCGC)

The Molecular Libraries Screening Center Network (MLSCN)

MLSCN Operation

AssayPeer review

Candidate Probe

Optimization Chemistry

Screen Data

Adequate potency and solubility?

YES

NO

Bioassay

Investigator

Optimize Assay

Compound Repository

Advice

MLSMR Compound Collection (260,000 Compounds)

DC = Diversity CompoundsNC = Non-commercialTL-KIN = Kinase Targeted LibraryTL-GPCR = GPCR Targeted LibraryTL-IC = Ion Channel Targeted Library

TL-PRO = Protease Targeted LibraryTL-NUC = Nuclear Receptor Targeted TL-NTP = National Toxicology Program SS = Known BioactivesNP = Natural ProductsDEA = DEA Controlled Substances

MLSMR Collection Growth

Molecular Libraries Compound Collection

Screening Center Goals

Discovery and development of small moleculechemical probes to be used as research tools to interrogate existing and novel biological targets and pathways.

Generate comprehensive datasets in PubChem for ML compounds

Progress on Awarded HTS Assays 2005-7

Center Assignments Assays Screened in Chemistry ProbesBurnham 19 33 22 5 15Columbia 9 18 12 4 2Emory 15 20 17 6 2NCGC 24 33 31 5 19Upenn 10 20 13 3 8Pittsburgh 13 1  4 8 4 2Scripps 12 31 30 3 7SRI 21 28 20 7 0New Mexico 10 21 16 5 2Vanderbilt 13 16 10 4 1Totals 146 234 179 44 35

Broad Range of Assay Assignments

Research Field of Awarded HTS Assays 2005-7

Mol & Cell Biol CancerInfectious Neuro Diabetes-Metabolic-Endocrine InflammationCardiovascular-hemato AgingKidney Bone-Muscle-SkinCompound Profiling

Target Class of Awarded HTS Assays 2005-7

Enzyme Cell pathwayProtein-Protein Interaction GPCRIon Channel TransporterCell viability Zebrafish assayNuclear Receptor Protein-Nucleotide interactionProtein conformation NMR /fragmentCompound Profiling

Growth In PubChem Substances / Compounds

Growth In PubChem BioAssays

Growth in PubChem Users per Day

Two Examples of Projects

1. Emory University Molecular Libraries Screening Center, Ray Dingledine

Inhibitor of measles virus RNA polymeraseAssay from Richard Plemper, Emory University

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Measles virus (MV) inhibitor

Despite the existence of a vaccine, MV remains among the most lethal human pathogens and accounts for approximately 500,000 deaths annually (WHO)

Novel antivirals against MV are desired to control local outbreaks.

Why?Herd immunity is below protective levels in developing countriesDecreased vaccination compliance in parts of the developed worldNo therapeutic strategy for management of cases of severe measles

Richard Plemper & Jim Snyder

MV-eGFP control

GFP

Phase

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Development of a robust HTS assay

Cellular assay based on recombinant MV, with eGFP added to the amino terminus of the viral glycoprotein34,000 compounds assessed in a first-pass screenAS-48 is a known positive control

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Compound #16677 shows nanomolar activity against 8 primary MV isolates

IC50 concentrations against live MV range from 31 to 245 nMCompounds shown to not inhibit viral entry or host protein synthesisTarget narrowed down to the RNA polymerase complex

MV escape mutants and directed mutagenesis identify the target

I->V N->T S->AH->YIIIIII

S->PN->TS->P

E->G

Natural escape mutants paramyxovirus polymerase

765-R I A S L V Q G D N Q

L

T I A-778

PA A

GDNQ motif is the active center for phosphodiesterase bond formation

Site-directed mutagenesis of polymerase domain confirms IDIII

16677

Hit to probe development

Iterative rounds of analog generation and cell-based bioassayApprox. 150 analogs created in increasingly focused libraries

First measles virus probe

Mean EC50 = 3.8 nM(n=5 experiments)

Cell toxicity >300 μM

Mechanism: viral polymerase inhibitor

Shows in vivo efficacy in rats against intranasal infection with MV

1E-6 1E-5 1E-4 1E-3 0.01 0.1 1 100

25

50

75

100

10,000

100,000

1,000,000)lortn c

o %,

/ml

fup (etrs

tiu

s vi

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eae

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μconcentration of 14d probe ( M)

spe

rimen

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/ml

fup(e in

5 e

xrit

ial t

init

EC50 = 0.6 nM

SO O

NH

O

NN

CF3

MeN

CID 16122506

Two Examples of Projects

1. Emory University Molecular Libraries Screening Center, Ray Dingledine

Inhibitor of measles virus RNA polymeraseAssay from Richard Plemper, Emory University

2. NIH Chemical Genomics Center, Chris AustinInhibitors of S. mansoni peroxiredoxinsAssay from David Williams, Illinois State University

WHO/TDR

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Developing drugs for Schistosomiasis

Parasitic disease that affects 250 million people, mostly in Africa Dr. David Williams at Illinois State University identified potential new targetThe NIH Chemical Genomics Center and Dr. Williams worked together to successfully identify targeted chemicals that provide a starting point for new drugs

Targeted Redox Pathway

Inhibition of S. mansoni peroxiredoxin would prevent worm degradation of hydrogen peroxide and kill schistosomes

O2 O2*- H2 O2 H2 O

OH* / OCl- Schistosome death

Superoxide dismutasee-

CatalaseGlutathione peroxidasePeroxiredoxin

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Quantitative HTS

70,000 compounds at 7 concentrations (qHTS)Dose-response curve for all compounds (PNAS 103, 11473-8 (2006))~10,000,000 data points (16 Time-Point Reads)31 hours of robot time

Results: 100 compounds with IC50 < 40 µM71 compounds6 different structural classes

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5

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Control Treatment 1 Treatment 2 Treatment 3

Tota

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f wor

ms

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NCGC1597 Inhibitor of Schistosoma mansoni Peroxiredoxins

NCGC1597 was administered by intraperitoneal injection at 10 mg/kg for 5 days at different points during the development of Schistosoma mansoni in the mouse.

Nature Medicine, March 2008

ML Production Phase, starting summer 2008

Three different types of CentersComprehensive Screening CentersSpecialized Screening CentersSpecialized Chemistry Centers

With advent of Chemistry Centers, increased emphasis on network communication

Each Center may have center-defined research

Screening Centers will provide outreach and greater support for assay implementation

Screening Centers will be expected to run all primary and secondary screening assays

http://mli.nih.gov/