Metrion Biosciences Ion Channel CRO Service Overview: Cardiac safety assays - Metrion Biosciences: the ion channel specialists
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Metrion Biosciences: the ion channel specialists Metrion Biosciences Ion Channel CRO Service Overview: Cardiac safety assays August 2020 www.metrionbiosciences.com
Metrion Biosciences
Company and vision
Metrion Biosciences is a Cambridge, UK-based contract research
organisation specialising in providing a range of high quality ion channel
drug discovery services for our customers
• We aim to provide:
• Exceptional ion channel electrophysiology expertise and services
• High quality, cost-effective compound screening assays
• Detailed characterisation of lead compounds in human cells and ⚫
native tissue
• Confirmation of efficacy in stem cell and other phenotypic models
• Rapid reporting and data interpretation by experienced ion
channel team
• A dedicated, flexible service tailored to each customer’s
requirements
2
Ion channel screening: high quality platforms
High quality electrophysiology
Ion channel drug discovery is reliant on sophisticated screening platforms
• Metrion scientists have decades of accumulated experience utilising such devices
• All of our electrophysiology platforms deliver high quality recordings and data
• Flexible support for assay development, primary target screening, hit confirmation, lead
optimisation and SAR, mechanistic and phenotypic studies
• 384-well platforms available
• QPatch-48 (x2)
• Patchliner (x3)
• Manual patch clamp (x5)
• Multi-Electrode Array and impedance
(native and iPSC neurons and cardiomyocytes)
• FlexStation fluorescence plate reader (x2)
3
Metrion’s ion channel drug discovery services
Ion channel services
Ion channel screening
▪ Generation of biological reagents, such as stable cell lines,
transient transfection and transduction techniques
▪ Assay development
▪ Range of high quality ‘gigaseal’ patch clamp and other ion
channel screening platforms
▪ Validated screening assays against an extensive panel of ion
channel cell lines and native cells
Cardiac safety screening
▪ CiPA-compliant panel of cardiac ion channel screening assays
▪ Phenotypic assays utilising human iPSC-derived cardiomyocytes
▪ In silico assessment of compound pro-arrhythmic liability
Neuroscience services
▪ Screening assays against a panel of voltage-gated, ligand-gated,
inward rectifier and K2P ion channels
▪ Phenotypic assays in native and iPSC neurons to aid the translation
of in vitro neuroscience data to the preclinical stage
4
Cardiac safety screening: overview of CiPA
CiPA: an FDA regulatory proposal
1 2 3 4
Analysis of compound Use in vitro data for in silico Confirm proarrythmia signals Clinical evaluation of
activity in a full panel of modelling to evaluate in human iPSC-derived unanticipated effects
human cardiac ion channel proarrhythmic risk cardiomyocytes
• The Comprehensive in vitro Proarrhythmia Assay (CiPA) initiative is a regulatory proposal
sponsored by global safety bodies such as FDA to improve cardiac safety assessment
• Metrion have established in vitro cell-based assays for CiPA pillars 1 and 3
• Several standard in silico models (Pillar 2) have been validated using Metrion’s QPatch data
• Data from Metrion’s Pillar 1 assays are being used to train in silico models at the FDA
• Metrion data is included in several upcoming CiPA and FDA ion channel publications
Sager et al., (2014); Gintant (2014) 5
CiPA pillar 1: hERG QPatch IC50 assay
hERG QPatch assay
• Metrion Biosciences are a leading provider of high quality hERG screening data
• Optimised hERG assay offers stable recordings with little or no rundown
• Assay is tracked and every screen is benchmarked with positive control (e.g. Verapamil)
• Single hole recordings ensure robust QC of client data
Example current traces Current vs time plot IC50 potency curve
6
CiPA pillar 1: Cav1.2 QPatch assay
hCaV1.2 QPatch assay
• Metrion offers an industry-leading stable and reliable Cav1.2 screening assay
• Choice of voltage protocols (single step, double step, or CiPA step-ramp pulse)
• Three or four-point IC50 format with a minimum double addition of each concentration
• QPatch assay is stable in 0.1% DMSO vehicle, with gigaseal quality and minimal rundown
• Can detect potency of reference compounds from nM to µM range
Cav1.2 reference data
120
N ife d ip in e
100 N im o d ip in e
Compound IC50 (µM)
C is a p r id e
Nifedipine 0.025
% In h ib itio n
80 P im o z id e
C d C l2 Nimodipine 0.204
60
Cisapride 8.52
40 Cav1.2 raw data traces
Pimozide 11.0
20
CdCl2 17.17
0
-1 2 -1 0 -8 -6 -4 -2 0
L o g [C o m p o u n d ] (M )
7
CiPA pillar 1: Nav1.5 QPatch assay
hNaV1.5 QPatch assay
• Metrion offers standard ‘peak’ Nav1.5 assay, as well as a ‘late’ DKPQ LQT3 mutant assay
• Standard format of core panel assay is a 1Hz ten pulse train format
• Can also be run at higher frequencies (2 - 5Hz) to mimic cardiac disease
• Four-point double addition IC50 format reports Pulse1 and Pulse10 potency
• Simultaneously extract tonic and phasic block of test compounds
• QPatch assay is stable in 0.1% DMSO vehicle with gigaseal quality
• Assay is pharmacologically validated with a toolbox of reference compounds
• Amitriptyline is the use-dependent positive control (P1=15 µM, P10=7 µM)
Nav1.5 pulse train protocol DMSO stability Amitriptyline IC50
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CiPA pillar 1: Late Nav1.5 assay
hNaV1.5 ‘late’ assay
• Metrion also offers a ‘late’ Nav1.5 assay using ATX-II opener or DKPQ LQT3 cell line
• Recent CiPA data suggests that cardiac risk is better predicted using ‘late’ Nav1.5 data
• Standard assay uses ATX-II to pharmacologically induce sustained ‘late’ currents (A)
• Metrion also created an LQT3 mutant cell line assay to avoid non-specific effects of ATX-II (B)
• Four-point double addition IC50 format reports Peak and Late current potency
• Simultaneously extract cardiac risk MoA profile of test compounds
• Metrion is the only CRO that offers a physiological LQT3 ‘late’ Nav1.5 assay
• ‘Late’ Nav1.5 assay is pharmacologically validated with reference compounds (C)
Nav1.5 + ATX-II LQT3 DKPQ late Nav1.5 Pharmacology
9
Cardiac safety screening: CiPA+ panel and in silico APs
Cardiac safety assays
1. Comprehensive core and CiPA cardiac ion channel panels 2. In silico AP modelling
• Metrion have validated assays for core (hERG, Nav1.5 and Cav1.2 ) and full CiPA cardiac panels
• We are the only CRO with high quality automated ‘late’ Nav1.5 and dynamic hERG assays (CiPA+)
• High quality in vitro cardiac channel IC50’s are suitable for use in in silico action potential models
• Metrion is the only CRO offering access to official FDA in silico AP model (dynamic O’Hara-Rudy, ORd)
10CiPA cardiac panel 3: human iPSC cardiomyocytes
Cardiac safety assays
A: iPSC cardiomyocyte B: Phenotypic iPSC cardiotoxicity assays C: Chronic iPSC cardiotoxicity assay
electrophysiology
i ii Doxorubicin
Dofetilide
• Human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) offer an accessible
and validated set of reagents for translational and predictive cardiac safety testing
• Metrion have a fully validated set of acute toxicity assays using commercial iPSC-CM reagents
• Manual patch voltage clamp (ionic currents) and current clamp (AP firing and arrhythmia) (A)
• MEA (Bi) and impedance (Bii) platforms measure proarrhythmic liability and contractility readouts
• Chronic cardiotoxicity assays are also available on the impedance platform (C)
11Guideline prices for core cardiac electrophysiology assays
QPatch assays
• Standard prices for ‘core’ panel cardiac profiling (hERG, hNav1.5, and hCav1.2):
Price per compound based upon
Ion Channel Platform Assay format number of compounds/batch
2 3 5 10 ≥ 20
hERG £520 £465 £440 £410 £380
Nav1.5 Four-point £520 £465 £440 £410 £380
QPatch
IC50 (n=3)
Cav1.2 £660 £600 £575 £550 £500
• Turnaround times are 2-3 weeks from compound receipt
• Potency data for a positive control reference compound will also be included
for each core panel ion channel investigated
Confidential, not for distribution
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*All prices are in GBP, ex VATMetrion Biosciences: strength and expertise
In summary
• Extensive and proven drug discovery expertise
• Expert provider of ion channel screening assays
• Proven record of delivering high quality data, on-time and on-budget
• Staff have been involved with multiple, successful integrated drug discovery programmes
• Wealth of ion channel pharmacology and screening know-how
• Highly experienced with voltage-gated and ligand-gated ion channel families
• Detailed understanding of modulator MOA, compound class issues and safety profiles
• Screening cascade experience from target validation to HTS and SAR
• Translation of ion channel expertise to phenotypic neuronal and cardiac assays
• Automated patch clamp experts
• Over fifteen years of experience operating a range of gigaseal APC platforms
• Trusted real-world drug discovery users working closely with all APC platform vendors
• Published a number of cell line and assay validation application notes
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www.metrionbiosciences.comMetrion Biosciences: the ion channel specialists For science, business or customer enquiries, please contact: info@metrionbiosciences.com www.metrionbiosciences.com
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