Cardiac Safety Screening 2009
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Cardiac Safety Screening 2009

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Cardiac Safety Screening 2009 Cardiac Safety Screening 2009 Presentation Transcript

  • Cardiac Safety Testing Services
  • Why Cardiac Safety Testing? S7B Guidance Objective − Identify potential of a test substance or its metabolites to delay ventricular repolarization. − Relate extent of delayed ventricular repolarization to concentrations of a test substance and its metabolites. Components that contribute to an assessment of cardiac risk − In Vitro IKr Assay – effect on native or expressed IKr (such as hERG) − In Vivo QT Assay – effect on QT interval – this assay can be designed to meet the objective of both the ICH S7A (cardiovascular core battery) and S7B
  • Ion Channels that Contribute to Cardiac Action Potential voltage K+ Channel Block time Ca2+ Channel Block Na+ Channel Block Normal AP Inhibition of cardiac ion currents can alter QT interval, cause arrhythmia or death Discovery Safety Testing: Detect cardiac safety liabilities early.
  • Cardiac Safety Testing Services Radioligand Binding Assays (Pharmacology, Taiwan) − Ideal for early hit selection profiling Information Value Cellular patch clamp assays (Pharmacology, Bothell) − Automated Patch Clamp (APC) with PatchXpress – cost-effective technology providing a medium-throughput method usually used in discovery stage after hit selection Tissue Assays (Pharmacology, Taiwan) − L-Type Calcium channel Atrial Inotropy In Vivo Assays – (Pharmacology, Taiwan and DSA, Lyon) − - Cardiovascular, QTc Interval 4
  • Ion Channel Testing Services Services at Bothell Site Non-GLP (10 day TAT) - Cardiac – hERG (Kv11.1) – hNav1.5 – hCav1.2 - Immuno-suppression and Obesity – hKv1.3 5
  • Functional Ion Channel Services in Bothell Discovery Safety Testing Fail Early Crucial attributes for discovery assays: Fast 10-day Turnaround time on Automated Patch Clamp (PatchXpress) assays Medium- to High-throughput capacity Automated Patch Clamp (PatchXpress) assays are medium-throughput at modest cost compared to conventional patch clamp Predictive of clinical outcome Over-expressed human cardiac ion channels that play central role in AP hKv11.1 (hERG) hNav1.5 hCav1.2 Assays validated against drugs with known cardiac liabilities or effects; compare favorably with conventional patch clamp
  • hERG Validation (PatchXpress)
  • hERG Validation (PatchXpress) hERG validation studies with control compounds 100 Astemizole Cisapride Percent of Control 75 E4031 Haloperidol Ketoconozole 50 Pimozide Quinidine 25 Risperidone Terfenadine 0 Verapamil -9 -8 -7 -6 -5 -4 Moxifloxacine Log Compound [uM] Assay comparison IC 50 (nM) Patch Express Conventional Patch Clamp Radioligand Binding Astemizole 13 + 2 26 15 Cisapride* 13 + 1 45 158 E4031 32 + 4 18-36 75 Haloperidol 90 + 21 93 250 Ketoconozole 2,340 + 250 1920-4700 19050 Moxifloxacine 39,167 + 4,061 41,000-170,000 NA Pimozide 12 + 4 18 53 Quinidine 760 + 91 300-1000 13155 Risperidone 379 + 59 394 5865 Terfenadine 38 + 7 28-56 127 Verapamil 535 + 46 140-830 5600 * - IC50 curve for Cisapride is not representative of current data.
  • hERG Validation (PatchXpress) Comparison of PatchXpress data to conventional patch clamp (Literature values)
  • Nav1.5 Validation (PatchXpress) -10 mV Voltage Clamp Protocol -120 mV A 1 nA 3 ms 1 -75 -50 -25 25 50 75 B -1 0.5 nA -2 5 ms -3
  • Nav1.5 Validation (PatchXpress) hNav 1.5 validation studies with 100 control compounds Rank Order of Potency IC50 (uM) Dibucaine 0.149 + 0.054 75 Imipramine 0.676 + 0.061 Percent of Control Terfenadine 0.996 + 0.22 50 TTX 6.6 + 1.3 Flecainide 7.58 + 1.3 Veratridine 7.6 + 1.7 25 Quinidine 18.8 + 4.6 Lidocaine 41.4 + 6.5 0 -8 -7 -6 -5 -4 -3 Log Compound [uM]
  • Nav1.5 Validation (PatchXpress) PX pIC50 vs. RBA pIC50 8.00 y = 0.9863x + 0.2804 2 R = 0.8216 7.00 Dibucaine PatchXpress (PX) pIC50 (nM) Imipramine 6.00 Terfenadine Propranolol 5.00 Veratridine Flecanide Lidocaine 4.00 3.00 3.00 4.00 5.00 6.00 7.00 8.00 Radioligand Binding Assay (RBA) pIC50 (nM)
  • Cav1.2 Validation (PatchXpress) Cardiac Ca2+ channel hCav1.2 +10 mV -40 mV -80 mV 2 nA 10 ms 1 Normalized Current .8 .6 .4 .2 0.1 nA 0.2 nA 10 ms -40 -20 0 20 40 60 Test pulse (mV)
  • Cav1.2 Validation (PatchXpress) hCav 1.2 validation studies with control compounds IC50 (µM) µ 100 Bepridil 1.19 0.87 Percent of Control Nitrendipine 75 Nicardipine 0.114 Nifedipine 0.138 50 Lacidipine 0.177 25 Diltiazem 36.5 Nimodipine 0.824 0 Isradipine 0.0535 -9 -8 -7 -6 -5 -4 -3 Log Compound [uM]
  • Cav1.2 Validation (PatchXpress) IC50 correlation between the PatchXpress 7000A vs. referenced conventional patch-clamp results. Conventional pIC50 vs. PX pIC50 8.5 Spearman r = 0.90, p <0.01 8.0 7.5 Nitrendipine PX log pIC50(nM) 7.0 Isradipine Nifedipine 6.5 Nicardipine Lacidipine 6.0 Nimodipine Bepridil 5.5 5.0 4.5 Diltiazem 4.0 4.0 4.5 5.0 5.5 6.0 6.5 7.0 7.5 8.0 8.5 Conventional log pIC50(nM)