New High Conten t Screening Methods Applied to in vitro Toxico logy Analys is

Bette r dec is ions . Earlie r in the game.

Enquiries@cyprotex.com

Anthony D Baxter, CEO

Business overviewA uniquely packaged ADMET solution

The leading specialist Contract Research Organisation (“CRO”) providing ADME-Toxscreening technology and information Evaluates and optimises Absorption, Distribution, Metabolism, Excretion, Toxicity (“ADMET”)

and pharmacokinetic properties of potential drug candidates

A portfolio of technologies to identify potential problems Assisting at the drug discovery stage and to help reduce the attrition of the drug development

process Genuine competitive advantages in different steps of the process

Cyprotex offers highly automated in-vitro and in-silico screening Designed to significantly accelerate and improve the cost-effectiveness of the process from

“hit-to-lead” in drug discovery Enormous global market opportunity – big pharma re-organisations

Commercial success Cloe® integrated product offering (Cloe® Screen, Cloe® Select and Cloe® Predict) Toxicology assays (Cyprotox) CellciphrTM (Cellumen, High content screening)

Company Overview

Founded in 1999. Publicly traded since 2002

Acquired Apredica in Aug 2010

Laboratories in UK (near Manchester) and US (near Boston)

100% focus on ADME-Tox/PK Services

No internal drug discovery programs

Over 500 customers worldwide (typically 50 at any one time)

Exceptional client retention rate

60+ staff

High PhD scientist ratio

Revenue and profitab ility

Group Revenues up 18.4% to £5.92 million (2009: £5.00 million)

UK revenues alone up 7.5% to £5.38 million

Additional Apredicarevenues of £0.54 million (5 months)

Profit/(loss)for the year

Acquis ition of Apredica

Purchase of Apredica meant that we acquired a respected rival ADMET CRO in Boston, MA, USA (August 2010)

Apredica gave us:• World renowned CSO in Dr Katya Tsaioun• Many new assays in customised ADME• Three year head start in in vitro toxicology assays• Access to high content toxicology assays acquired from Cellumen• A bigger share of the US ADME-Tox market• Access to the largest global market for ADME-Tox• Additional highly experienced scientists – further strengthening our team

Prec lin ica l ADME Tox s ervices

ServicesHigh-throughput ADME screening Customised ADME assaysBioanalysis and PK analysisIn silico QSAR and PBPK modelling In vitro Toxicology• Mechanisms of Toxicity• Genotoxicity• Metabolism Related Toxicity• High Content Toxicology

Our ADME As s ays – core bus ines s

AbsorptionCaco-2 PermeabilityMDR1-MDCKPAMPA (Standard and BBB)Transporter Studies (P-gp and BCRP)Dermal AbsorptionIntranasal and Lung Absorption

MetabolismMetabolic Stability (microsomes, hepatocytes, S9, plasma)CYP450 Inhibition (IC50 and Ki)CYP450 Time Dependent InhibitionCYP450 InductionCYP450 Reaction PhenotypingUGT1A1 InhibitionMAO InhibitionMetabolite Profiling

DistributionPlasma Protein BindingBrain Tissue BindingBlood to Plasma RatioMicrosomal Binding

PK and BioanalysisPharmacokinetic AnalysisBioanalytical Method Development and Validation

Physicochemical PropertiesTurbidimetric SolubilityThermodynamic SolubilitypKa DeterminationLipophilicity (CHI, LogP, LogD)Chemical Stability

Our ADME & PK Predic tion Services

PB-PK Modelling• Pharmacokinetic predictor

(Cloe® PK)

• Human intestinal absorption predictor (Cloe® HIA)

• Customized chemistry-specific models

• Animal-to-human in vivo extrapolator

• PK/PD modelling

QSARModel building and predictionNovel auto-QSAR techniques

Current Collaborative ProjectsOSIRIS - Risk assessment of industrial chemicalsTB Alliance – PK predictionIMI – European standards for data and model results access

Our Toxic ity As s ays

General CytotoxicityMTTNeutral RedLDH Release

Mechanisms of ToxicityMitochondrial ToxicityhERG inhibitionHaemolysisPhospholipidosisPhototoxicitySkin IrritationGene Regulation (qRT-PCR)

High Content ToxicologyCytotoxicity Screening Panel (Tier 1)CellCiphrTM Toxicity Profiling (Tier 2)

Metabolism-Related ToxicityReactive Metabolite AssessmentDrug-Drug InteractionsMultispecies Metabolite Profiling

GenotoxicityAMESGreenScreen HCTM

In vitro MNTIn vitro Comet

Why inves t in toxico logy?

Alle Ding' sind Gift, und nichts ohn' Gift; allein die Dosis macht, daß ein Ding kein Gift ist.

"All things are poison and nothing is without poison, only the dose permits something not to be poisonous."

Paracelsus (Phillippus Aureolus Theophrastus Bombastus von Hohenheim)

1493 – 1541 Switzerland

The Grandfather of toxicology

Why inves t in toxico logy?

In 1990s drug metabolism activities increasingly moved earlier, from development to discovery

Led to lower attrition in the clinic due to DMPK problems

Success in early ADME screening means that toxicology is now the ‘enemy of successful drug discovery’

Kola & Landis (2004)

Clinica

l Safe

ty

Efficac

y

Formulat

ion

ADME-PK

Commercial

Toxicology

Cost of G

oodsOther

0

10

20

30

40

5019912000

Criterion

Attr

ition

(%)

How compounds fail in the clinic

!

Toxic ity is a major reas on for drug a ttrition

Over 40% of all drugs fail preclinical or clinical testing (2010 data) because of apparent or suspected drug toxicities

Cost of toxicity-related drug failures is ~$2 billion per year

DM&D Market Analysis Report, 2003

Conventional animal toxicity studies fail to predict idiosyncratic human hepatotoxicity

http://www.fda.gov/cder/livertox/stateArt.htm

Toxicity is still a late-stage issue for many companies

Why High Content Toxicology?

Often drug toxicity is combination of multiple mechanismsA single experimental approach may not be predictive of the complex steps involved in toxicological effectsHCS captures multiple mechanistic parameters which cover a wide spectrum of cytopathological changesParameters can be measured in same cell populations in the same well, reducing inter-assay differences

Two tiers of toxicity profilingProfiling: Predict Preclinical Outcome

Organ-specific Panel Profiles

General Cytotoxicity Assay Sets

Mechanism-specific Assay Sets

Tier 1: Screening

Tier 2: Full profiling

Rationale of Selected Parameters: Tier 1

O’Brien et al, Arch. Toxicol (2006) 80:580-604: Pfizer

Assay Sensitivity Specificity

Cell ViabilityMembrane IntegrityApoptosis (Caspase 3)Protein SynthesisOxidative Stress (superoxide induction)Oxidative Stress (glutathione depletion)DNA Synthesis

Cell Viability or GSH or DNA synthesisRegulatory animal toxicity testing

10% 2%5%4%1%

19%10%

25%52%

92%99%95%97%97%85%92%

83%-

611 compounds (retrospective)

Cellomics “Quad Probe Assay (Tier 1)” 93% 98%

Sensitivity = % of hepatotoxic compounds detected i.e. 1:10 known hepatotoxicants in test set detected (10%)Specificity = % of correctly identified hepatotoxic compounds i.e. 9:10 positive compounds are hepatotoxicants (90%)

High Content Screening Technology for Cellu la r Toxic ity As s es s ment

State-of-the-art Thermo ArrayScan VTIAutomated fluorescence imaging and cellular analysisMulti-parametric indicators of cell toxicityDetection of cell death and mechanisms of cell death

Screening: Validated channels/colours

Customization is possible!Measuring other features is possible if reagents are commercially available

nm

Cell Health/Death Signalling

High Content Toxicology: Tier 1 Screening Panel

Cell lossNuclear areaDNA structure

1µM Paclitaxel

Nuclear morphology Cell permeability

1µM Cerivastatin

Mitochondrial potential

1µM Paclitaxel

0.2% DMSO

Cytochrome c

10µM Amiodarone

0.2% DMSO0.2% DMSO 0.2% DMSO

4x

4x

1µM Paclitaxel

Nuclear morphology Cell permeability

1µM Cerivastatin

Mitochondrial potential

1µM Paclitaxel

0.2% DMSO

Cytochrome c

10µM Amiodarone

0.2% DMSO0.2% DMSO 0.2% DMSO

4x

4x

24hr exposure of HepG2 cells

Increased cell permeability

Mitochondrial massMitochondrial potential loss

Cytochrome C release

Observations from single cell population

Control:

Treated:

Blue Green Red Far Red

High Content Toxicology CellCiphrTM Organ Specific Toxicity Panels (Tier 2)

Cytotoxicity ProfilingHepG2 cells (Human)

Human cell lineInsights into toxicity to cycling cells

Hepatotoxicity ProfilingRat Primary Hepatocytes

Primary cells with metabolic capacityInvestigate hepatocyte-specific toxicities

HepaRG (in development)Human cell lineMetabolic competencyCo-development opportunities

Cardiotoxicity ProfilingH9C2 cardiomyocytes (Rat)

Hypertrophy and cardiomyocyte-specific toxicities

Hepatocytes

Cardiom

yocytes

Animal & Human cell models

High Content Toxicology CellCiphrTM: Tier 2 Toxicity Panels

Cell Loss Nuclear Size DNA DamageMitochondrial Potential

Cell Cycle Arrest Cytoskeletal DisruptionOxidative Stress Mitochondrial MassMitosis Marker Stress Kinase Activation

Cytotoxicity Profiling -HepG2

CardiotoxicityProfiling – H9C2Cell Loss Nuclear Size DNA Damage Mitochondrial Potential

Apoptosis Steatosis

Hypertrophy Reactive Oxygen

Hepatotoxicity Profiling – 1o RatCell Loss Nuclear Size DNA Damage Mitochondrial Potential

ApoptosisSteatosis

DNA Fragmentation Phospholipidosis

CellCiphr® CSB™ Tier 2 HepG2 Panel

DMSO

HepG2 cells. Blue – nucleiGreen – microtubule stability markerRed – mitochondrial function markerMagenta – mitotic arrest marker.

0.2 μM paclitaxelDMSO

Cell Loss Nuclear Size DNA DamageMitochondrial PotentialCell Cycle Arrest Cytoskeletal DisruptionOxidative Stress Mitochondrial MassMitosis Marker Stress Kinase Activation

Cytotoxicity Profiling -HepG2

CellCiphr® CSB™ Tier 2 Primary Rat Hepatocyte Panel

Phospholipidosis

Cell loss & Nuclear Size

Apoptosis

DNA Damage

Mitochondrial Potential

Steatosis

Hepatotoxicity Profiling – 1o RatCell Loss Nuclear Size DNA Damage Mitochondrial PotentialApoptosisDNA Fragmentation PhospholipidosisSteatosis

Customer Cpds

Tier 2: High Content Toxicology CellCiphrTM

Database Source

Safety Toxicology

Physical Properties

580 Reference CpdsCommercial Libraries, FDA, EPA, and Cellumen

CellCiphr Analysis

PharmaPendiumEPA, FDA, others

Customer Proprietary Compounds

Safety Alerts

Rank Order

Similarity Profiles

Data Sources Database

Summary

Cyprotex is an ADME-Tox Expert

Cyprotex’s ServicesRoutine, custom & novel ADME-Tox assays

Process automation consultancy

Bioanalysis and PK analysis

In silico PBPK modelling

Cyprotex Advantage• Experienced technical staff

• Superior client communication

• Collaborative, flexible approach

Unrivaled process, speed, and quality

Cyprotex15 Beech LaneMacclesfieldCheshireSK10 2DRUK

Apredica , a Cyprotex company313 Pleasant StreetWatertown, MA 02472USA

Tel (UK): +44 (0) 1625 505115

Tel (US): +1-617-923-1466

Email: sales@cyprotex.com

Website: www.cyprotex.com