CSRC/HESI/SPS/FDA Think Tank. Ion Channel Testing Approach Najah Abi Gerges / Bernard Fermini December 11, 2014

Transcription

1 CSRC/HESI/SPS/FDA Think Tank Ion Channel Testing Approach Najah Abi Gerges / Bernard Fermini December 11, 2014

2 Objectives of ICWG Address important questions related to best practices including: 1. Which ion channels should be selected to support in silico working group (ISWG) AP modelling efforts. 2. What properties should be studied (IC 50 determination, kinetics, rate/use/voltage dependence, etc )? 3. What requirements are needed to deliver robust, reliable and reproducible ion channel data in a high throughput screening (HTS) environment. 2

3 Pre-CiPA state of Ion Channel testing Need for standardization Current pre-clinical guidelines (S7B) are just that: guidance and not dogma. Allows flexibility in the testing procedure. As a result, safety profiling is quite different from company to company despite following the same guidelines. There are a variety of automated patch clamp systems available to test for herg channel activity leading to variability in the performance of these systems and variability in the quality of data generated. The lack of standardization in the protocols used within the industry and in the academic environment contributes to additional uncertainty in the data. Current cardiac ion channel safety screening strategies focus on conductance block in an effort to define potency (IC 50 ). However; kinetic of drug block can influence use- and voltage- dependency. Evaluation of kinetic of block may lead to more complex but more predictive in silico models. It is therefore important to establish best practices for IC studies used to characterize drug effects (voltage protocols, temperature, reagents, etc ) so that consistent electrophysiological data may be obtained and contribute to better prediction of proarrhythmic risk. 3

4 Sample of electrophysiology devices for IC screening PatchXpress QPatch Patchliner SyncroPatch 384E IW Quattro IonFlux SyncroPatch 96 Conventional patch Barracuda CytoPatch Port-a-Patch Qube 384 4

5 ICWG Deliverables (1) ICWG conducted and completed a survey with the purpose of collecting frequency/type data on the commonly used ion channels (ICs) in the various laboratories. ICWG identified 7 ICs of interest: 1. Selected outward currents: I Kr (herg), I Ks (KvLQT1/KCNE1), I to (Kv4.3) 2. Selected inward currents: I K1 (Kir2.1), I Ca-L (Cav1.2), I Na (Nav1.5; peak and late). Action potential morphology and duration are controlled by the complex interplay between inward and outward currents. ICWG conducted > 25 Teleconferences in

6 Pre-CiPA state of Ion Channel testing Cardiac ion channels screened for safety (n=65)

7 Pre-CiPA state of Ion Channel testing Temperature at which assays are performed (n=54) Room Temperature Physiologic Temperature Other 5 0 7

8 Pre-CiPA state of Ion Channel testing Ion channels believed essential for in silico modelling (n=7)

9 ICWG Deliverables (2) ICWG designed and delivered two sets of protocols for all 7 ICs 1. Dynamic block protocols that provide data on affinity, kinetics and state-dependence of drug-channel interaction. To be used by ISWG. Dofetilide, cisapride and verapamil data generated for the herg channel and provided to ISWG. Data generated at RT and 37ºC Work in progress 2. Physiological protocols that provide data on IC 50, voltage / rate / use dependence, activation / inactivation / deactivation rates. To be used in FDA sponsored study (David Strauss Lead). Data to be generated at RT and 37 C. Pacing rates: 0.1 and 1 Hz Some data obtained using Action Potential voltage protocol Work in progress 9

10 Dynamic Block Protocols: Effects of Dofetilide on herg 22 ± 1ºC 37 ± 1ºC 100 nm Dofetilide 100 nm Dofetilide b b a d a d 10

11 Dynamic Block Protocols: Effects of Dofetilide on herg 22 ± 1ºC 37 ± 1ºC a b d normalized current nM 100nM 300nM = 8456 ms = 5666 ms 0.0 = NA prepulse duration (ms) normalized current nM 100nM 300nM = ms = 2458 ms = 1026 ms prepulse duration (ms) Data provided by Zenas Technologies 11

12 Physiological Protocols: some examples I to (Kv4.3); I Ks (KvLQT1); I Na (Nav1.5) 12

13 Physiological Protocol Parameters All channels ICWG Final Protocols Protocol HP (mv) HP (ms) Ramp to (mv) Ramp (ms) Ramp (V/S) Step1 (mv) Step 1 (ms) Step 2 (mv) Step 2 (ms) Ramp to (mv) Ramp (ms) Ramp (V/S) Frequency (Hz) IKr (herg) TBD IK1 (Kir2.1) TBD IKs (KvLQT1/KCNE1) TBD Ito (Kv4.3) TBD INa peak (Nav1.5) TBD INa late (Nav1.5) TBD ICa-L (Cav1.2) TBD 13

14 ICWG Next Steps Work closely with ISWG and FDA. Provide feedback and scientific input/support. Revise/modify/optimize protocols, as required. Coordinate the generation of additional ion channel pharmacology from CiPA list using standardized protocols (IC 50 + dynamic block) Adapt protocols to HT screening environment. Publish the work. 14

15 CiPA/SPS Ion Channel Working Group (ICWG) Membership Najah Abi Gerges Khuram Chaudray Krystle Correll William Crumb Bruce Damiano Gul Erdemli Bernard Fermini Gary Gintant Jules C Hancox John Imredy John Koerner Jim Kramer Derek J Leishman Paul Levesque Anders Lindqvist Stanley Nattel Carlos Obejero-Paz David Rampe Gail Robertson Kowei Sawada David Strauss Jamie Vandenberg Hugo Vargas Affiliation AstraZeneca (co-chair) GSK SPS Zenas technologies Janssen Novartis Pfizer Inc (co-chair) AbbVie Bristol University Merck FDA Chantest Lilly BMS Sophion Montreal Heart Institute Chantest Sanofi Wisconsin University Eisai FDA Victor Chang Cardiac Research Institute Amgen 15