vitro Toxicology Analysis

Transcription

1 Anthony D Baxter, CEO New High Content Screening Methods Applied to in vitro Toxicology Analysis Better decisions. Earlier in the game. Enquiries@cyprotex.com

2 Business overview A uniquely packaged ADMET solution The leading specialist Contract Research Organisation ( CRO ) providing ADME-Tox screening 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)

3 Company Overview Founded in 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

4 Revenue and profitability Group Revenues up 18.4% to 5.92 million (2009: 5.00 million) UK revenues alone up 7.5% to 5.38 million Additional Apredica revenues of 0.54 million (5 months) Profit/(loss)for the year

5 Acquisition 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

6 Preclinical ADME To x services Services High-throughput ADME screening Customised ADME assays Bioanalysis and PK analysis In silico QSAR and PBPK modelling In vitro Toxicology Mechanisms of Toxicity Genotoxicity Metabolism Related Toxicity High Content Toxicology

7 Our ADME As s ays core business Absorption Caco-2 Permeability MDR1-MDCK PAMPA (Standard and BBB) Transporter Studies (P-gp and BCRP) Dermal Absorption Intranasal and Lung Absorption Metabolism Metabolic Stability (microsomes, hepatocytes, S9, plasma) CYP450 Inhibition (IC 50 and K i ) CYP450 Time Dependent Inhibition CYP450 Induction CYP450 Reaction Phenotyping UGT1A1 Inhibition MAO Inhibition Metabolite Profiling Distribution Plasma Protein Binding Brain Tissue Binding Blood to Plasma Ratio Microsomal Binding PK and Bioanalysis Pharmacokinetic Analysis Bioanalytical Method Development and Validation Physicochemical Properties Turbidimetric Solubility Thermodynamic Solubility pka Determination Lipophilicity (CHI, LogP, LogD) Chemical Stability

8 Our ADME & PK Prediction 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 QSAR Model building and prediction Novel auto-qsar techniques Current Collaborative Projects OSIRIS - Risk assessment of industrial chemicals TB Alliance PK prediction IMI European standards for data and model results access

9 Our Toxicity As s ays General Cytotoxicity MTT Neutral Red LDH Release Metabolism-Related Toxicity Reactive Metabolite Assessment Drug-Drug Interactions Multispecies Metabolite Profiling Mechanisms of Toxicity Mitochondrial Toxicity herg inhibition Haemolysis Phospholipidosis Phototoxicity Skin Irritation Gene Regulation (qrt-pcr) Genotoxicity AMES GreenScreen HC TM In vitro MNT In vitro Comet High Content Toxicology Cytotoxicity Screening Panel (Tier 1) CellCiphr TM Toxicity Profiling (Tier 2)

10 Why invest in toxicology? Paracelsus (Phillippus Aureolus Theophrastus Bombastus von Hohenheim) Switzerland The Grandfather of toxicology 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."

11 Why invest in toxicology? 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 Attrition (%) How compounds fail in the clinic Kola & Landis (2004)! Clinical Safety Efficacy Formulation ADME-PK Commercial Criterion Toxicology Cost of Goods Other

12 Toxicity is a major reason for drug attrition 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 Toxicity is still a late-stage issue for many companies

13 Why High Content Toxicology? Often drug toxicity is combination of multiple mechanisms A single experimental approach may not be predictive of the complex steps involved in toxicological effects HCS captures multiple mechanistic parameters which cover a wide spectrum of cytopathological changes Parameters can be measured in same cell populations in the same well, reducing inter-assay differences

14 Two tiers of toxicity profiling Profiling: Predict Preclinical Outcome Tier 1: Screening General Cytotoxicity Assay Sets Mechanism-specific Assay Sets Tier 2: Full profiling Organ-specific Panel Profiles

15 Rationale of Selected Parameters: Tier 1 O Brien et al, Arch. Toxicol (2006) 80: : Pfizer Assay Sensitivity Specificity Cell Viability Membrane Integrity Apoptosis (Caspase 3) Protein Synthesis Oxidative Stress (superoxide induction) Oxidative Stress (glutathione depletion) DNA Synthesis 10% 2% 5% 4% 1% 19% 10% 92% 99% 95% 97% 97% 85% 92% Cell Viability or GSH or DNA synthesis Regulatory animal toxicity testing 25% 52% 83% - Cellomics Quad Probe Assay (Tier 1) 93% 98% 611 compounds (retrospective) 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%)

16 High Content Screening Technology for Cellular Toxicity Assessment State-of-the-art Thermo ArrayScan VTI Automated fluorescence imaging and cellular analysis Multi-parametric indicators of cell toxicity Detection of cell death and mechanisms of cell death

17 Screening: Validated channels/colours Customization is possible! Measuring other features is possible if reagents are commercially available nm

18 Cell Health/Death Signalling

19 High Content Toxicology: Tier 1 Screening Panel 24hr exposure of HepG2 cells Blue Green Red Far Red Nuclear morphology 4x Cell permeability Mitochondrial potential Cytochrome c Control: 0.2% DMSO 0.2% DMSO 0.2% DMSO 0.2% DMSO Treated: 4x 1µM Paclitaxel Cell loss Nuclear area DNA structure 1µM Cerivastatin Increased cell permeability 1µM Paclitaxel Observations from single cell population Mitochondrial mass Mitochondrial potential loss 10µM Amiodarone Cytochrome C release

20 High Content Toxicology CellCiphr TM Organ Specific Toxicity Panels (Tier 2) Cytotoxicity Profiling HepG2 cells (Human) Human cell line Insights into toxicity to cycling cells Hepatotoxicity Profiling Rat Primary Hepatocytes Primary cells with metabolic capacity Investigate hepatocyte-specific toxicities HepaRG (in development) Human cell line Metabolic competency Co-development opportunities Cardiotoxicity Profiling H9C2 cardiomyocytes (Rat) Hypertrophy and cardiomyocyte-specific toxicities Hepatocytes Cardiomyocytes Animal & Human cell models

21 High Content Toxicology CellCiphr TM : Tier 2 Toxicity Panels Cytotoxicity Profiling - HepG2 Cell Loss Nuclear Size DNA Damage Mitochondrial Potential Cell Cycle Arrest Cytoskeletal Disruption Oxidative Stress Mitochondrial Mass Mitosis Marker Stress Kinase Activation Hepatotoxicity Profiling 1 o Rat Cell Loss Nuclear Size DNA Damage Mitochondrial Potential Apoptosis Steatosis DNA Fragmentation Phospholipidosis Cardiotoxicity Profiling H9C2 Cell Loss Nuclear Size DNA Damage Mitochondrial Potential Apoptosis Steatosis Hypertrophy Reactive Oxygen

22 CellCiphr CSB Tier 2 HepG2 Panel DMSO Cytotoxicity Profiling - HepG2 Cell Loss Nuclear Size DNA Damage Mitochondrial Potential Cell Cycle Arrest Cytoskeletal Disruption Oxidative Stress Mitochondrial Mass Mitosis Marker Stress Kinase Activation DMSO 0.2 μm paclitaxel HepG2 cells. Blue nuclei Green microtubule stability marker Red mitochondrial function marker Magenta mitotic arrest marker.

23 CellCiphr CSB Tier 2 Primary Rat Hepatocyte Panel Hepatotoxicity Profiling 1 o Rat Cell Loss Nuclear Size DNA Damage Mitochondrial Potential Apoptosis DNA Fragmentation Phospholipidosis Steatosis Cell loss & Nuclear Size Apoptosis Phospholipidosis DNA Damage Mitochondrial Potential Steatosis

24 Tier 2: High Content Toxicology CellCiphr TM Database Source Data Sources Commercial Libraries, FDA, EPA, and Cellumen Database 580 Reference Cpds PharmaPendium EPA, FDA, others Customer Proprietary Compounds Physical Properties Safety Toxicology Customer Cpds Safety Alerts CellCiphr Analysis Rank Order Similarity Profiles

25 Summary Cyprotex is an ADME-Tox Expert Cyprotex s Services Routine, 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

26 Cyprotex 15 Beech Lane Macclesfield Cheshire SK10 2DR UK Tel (UK): +44 (0) Tel (US): Website: Apredica, a Cyprotex company 313 Pleasant Street Watertown, MA USA