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Measurement of bronchoconstriction in conscious rats: Elegant technique, elusive recordings
Abstracts
Impedance is a recognized methodology for respiratory measurement; however the use of impedance respiratory monitoring in freely moving cynomolgus monkeys has not yet been reported. This study explored impedance respiratory monitoring as a novel implantable telemetry monitoring strategy. A surgically implanted telemetry transmitter with ECG (DII), arterial pressure, physical activity, body temperature, respiratory rate and tidal volume (TV) was characterized. A longitudinal incision was performed lateral to the linea alba. The internal abdominal oblique muscle was separated from the aponeurosis of the transverses abdominis by blunt dissection. A transmitter was inserted between the internal abdominal oblique muscle and the aponeurosis of the transverses abdominis. Impedance electrodes were placed subcutaneously on the lateral aspect of the thorax. After recovery, controlled tidal volumes (20, 40 and 60 mL) were used in 6 anesthetized cynomolgus females to evaluate the effects of body positions on respiratory measurements. All correlations were above 98%. Animals were followed over time to assess both accuracy and stability of telemetric impedance measurements. Slight changes in the impedance signal were noted between the surgery and two weeks post-implant. Impedance remained stable over the following 2-month monitoring period. Circadian rhythms of respiratory rate and TV were correlated with cardiovascular parameters. In conclusion, this new telemetry monitoring modality could be valuable in pharmacological investigations when the use of cynomolgus monkeys is justified.
doi:10.1016/j.vascn.2010.11.015
Poster Number: 12 Board Number: 12 Translational medicine: Can we use publicly available data in correlating preclinical with clinical studies? Ard Teismana, Pieter-Jan Gunsb, Karel Van Ammela, Rob Towarta, Nenad Sarapac, David Gallachera a
Johnson & Johnson EDC, Beerse, Belgium Formerly Johnson & Johnson EDC, Beerse, Belgium and University of Antwerp, Antwerp, Belgium c Formerly Johnson & Johnson Clinical East Coast, Raritan, NJ, United States
e5
suboptimal clinical data diminishes the reputation of preclinical models. Cross company collaboration and improved Phase I study designs may improve this situation.
doi:10.1016/j.vascn.2010.11.016
Poster Number: 13 Board Number: 13 Adaptation of safety pharmacology cardiovascular and respiratory methodology for use on juvenile toxicity studies Kevin Norton, Keith Robinson, Mark Vezina Charles River Laboratories, Senneville, Quebec, Canada The EMEA Guideline on the need for nonclinical testing in juvenile animals on human pharmaceuticals for pediatric indications and FDA Guidance Document. Nonclinical safety evaluation of pediatric drug products; require the conduct of juvenile toxicity studies intended to bridge the developmental toxicity studies adult toxicity studies. As per the EMEA guidance document, the objective of such studies is to assess “potentially different safety profiles from those seen in adults.” With new chemical entities being specifically evaluated to assess effects on overall growth of the organ systems that develop postnatally (e.g., skeletal, renal, lung, neurological, immunologic and reproductive systems). We have looked at incorporation of traditional assessments of safety pharmacology core battery assessments for CNS, respiratory and cardiovascular studies in juvenile toxicity studies. We demonstrate the use of a modified functional observation battery and whole body plethysmography to assess effects on CNS and respiratory parameters, respectively, in rats between day 10 and sexual maturation (approximately day 50). Whilst beagle dogs approximately 3 to 4 month of age were instrumented with DSI telemetry transmitters to assess cardiovascular effect. The data reported illustrate that traditional safety pharmacology can be adapted to meet the growing requirements for juvenile toxicity studies.
b
Criticism of preclinical models is based on their apparent lack of reliability to predict clinical events. In the cardiovascular (CV) safety area blockade of the IKr channel can lead to QT prolongation, which in turn may lead to TdP. Although we know that drug-induced TdP may be caused by block of IKr, we also know that some IKr blockers do not necessarily induce QT-prolongation/TdP. To answer the question “how good are current preclinical models?” one needs to review both preclinical and clinical data in a wide range of chemical and pharmacological classes. The majority of clinical data is not in the public domain. Within this project we compared both J&J internal and publicly available clinical CVdata with results of preclinical models. We evaluated several different clinical data sources and identified 80 clinical reports on 73 compounds with different levels of CV data. The data were collected in a data warehouse and compared with preclinical data. Due to the different levels of detail and large variability of designs etc., the actual number of compounds that could be used was significantly reduced. Preliminary analysis shows that TQT-data tend to have the lowest variability, and that these should preferably be used for comparison with preclinical ECG-models. Unfortunately, little of this high quality data is available in the public domain, as many compounds will not reach the clinical stage. Using
doi:10.1016/j.vascn.2010.11.017
Poster Number: 14 Board Number: 14 Measurement of bronchoconstriction in conscious rats: Elegant technique, elusive recordings Lorna C. Ewarta, Katherine Elliottb, Michael Haleyb, Tom Hollandb, Sally-Ann Rickettsc, Alan McCarthyb, Lisa Williamsb, Jean-Pierre Valentinb a
AstraZeneca, Macclesfield, Cheshire, United Kingdom Safety Assessment, AstraZeneca, Macclesfield, Cheshire, United Kingdom c DECS, AstraZeneca, Macclesfield, Cheshire, United Kingdom b
Placement of a pressure-sensitive catheter radio-telemetry device, beneath the pleural surface, enables assessment of bronchoconstriction in parallel with changes in rate and depth of breathing (Murphy et al. 1998). Here, we assess this approach using an infusion of methacholine (MCh) up to 30 μg/kg/min or vehicle (saline) via the tail vein in adapted head-out plethysmography (HOP) chambers. Resistance measured by the telemetric method in conscious male rats (18 Han Wistar, 10 Sprague Dawley) was compared to resistance
e6
Abstracts
measured by an oesophageal catheter in anaesthetised rats (32 Sprague Dawley). Surgical placement of the pressure-sensitive catheter was successful in 60% of rats as confirmed by a strong, physiological signal and high-resolution computerised tomography or histology. MCh dose dependently increased resistance in 2 telemetered rats. No apparent pharmacological response was observed in the other dosed rats due to large pressure signal variability. Baseline resistance approximately doubled in all anaesthetised rats (n = 32) given 10 μg/kg/min MCh. Variability in the telemetry signal was associated with the posture adopted by the rat or the degree of movement within the HOP chamber, especially during dosing. Despite widespread consultation, extensive evaluation of habituation protocols and confirmation of catheter tip placement, we were not able to obtain reliable broncoconstrictor responses. Given the challenges associated with setting up this model, we would favour assessment of bronchoconstriction in anaesthetised rats.
doi:10.1016/j.vascn.2010.11.018
Poster Number: 15 Board Number: 15 The use of whole body plethysmography to evaluate drug-induced changes in respiratory disorders: Sleep apneas, cough and bronchospasm Eric Delpy, Fabrice Infanti, Mathilde Loiseau, Christophe Drieu La Rochelle Biotrial Pharmacology, Rennes, France Whole body plethysmography (WBP) is widely used in safety pharmacology to investigate in conscious animals potential undesirable pharmacodynamic effects of a substance on respiratory function. Proper characterisation of a breathing pattern includes measurements of respiratory rate, tidal and minute volume, inspiratory and expiratory times and peak flows. The aim of this study is to show that WBP could also be used to study drug-induced changes in respiratory disorders such as apneas, cough or bronchospasm. Spontaneous and post-sight sleep apneas were investigated in rats. Apneas were recorded from 30 min post dosing (time required for rats to fall asleep) and over 5 hours. In saline-treated animals, the number of apneas with a duration of more than 2.5 s was 34 ± 3 and the total duration was 117 ± 12 s. Diazepam (5 mg/kg, i.p.) decreased both the number (-20%) and the duration (-38%) of apneas. Cough was investigated in guinea-pigs and was induced by a 10-min citric acid inhalation. The number of coughs was counted by a trained experimenter based on characteristic animal posture, sound produced and respiratory signal. The number of coughs in saline-treated animals was 25 ± 2. Baclofen (0.1, 0.3 and 1 mg/kg s.c.) and codeine (10, 30 and 100 mg/kg p.o.) dose-dependently decreased the number of coughs. Bronchospasm was investigated both in ovalbuminsensitised and non-sensitised guinea-pigs; bronchoconstriction was evaluated by the calculation of enhanced pause (Penh). In sensitised animals, ovalbumin (1-100 μg/kg i.v.) dose-dependently increased Penh. In non-sensitised animals, histamine (0.3-30 μg/kg i.v.) dosedependently increased Penh and this effect was inhibited by mepyramine. These results show that WBP is appropriate to evaluate the effects of new chemical entities on respiratory disorders such as sleep apneas, cough and bronchospasm.
doi:10.1016/j.vascn.2010.11.019
Poster Number: 16 Board Number: 16 Orthostatic hypotension can be investigated in conscious dogs as part of standard telemetry studies Nicola Smith, Matt Skinner, Pierre Lainee, Jackie Moors, Stewart Brown, Amy Deaville, Jean-Pierre Valentin AstraZeneca R&D, Macclesfield, Cheshire, United Kingdom Despite being a common cardiovascular adverse event in Phase 1 clinical trials, orthostatic hypotension (OH) is not routinely investigated during standard preclinical studies. The aim of this work was to validate a dog tilt model designed to assess OH. Four to six dogs implanted with DSI telemetry transmitters were orally dosed on separate occasions with Prazosin (0.01, 0.03 and 0.1 mg/kg), Captopril (3, 30 and 100 mg/kg), Verapamil (1, 5 and 15 mg/kg), Atenolol (1, 3 and 10 mg/kg) or vehicle. Heart Rate and systolic blood pressure (SBP) were recorded while the dogs were exposed to a 1-minute head-up postural change of approximately 90° to induce OH at set timepoints pre and post-dose. Under control conditions, orthostatic tilt elicited an increase in SBP maintained for the duration of the tilt and an initial transient tachycardia. Prazosin and Captopril caused a decrease in the tilt-evoked increase in SBP (-75% and –30% vs vehicle, respectively), and increased tachycardia (27% and 57% vs vehicle, respectively). Verapamil caused an increase in the tilt-evoked tachycardia (176% vs vehicle) with minimal effect on the SBP response. Atenolol caused a decrease in tilt-evoked tachycardia (-69% vs vehicle) without affecting the SBP response. (% Values show max. change). Results correlate with the potency for these reference compounds to induce OH in man and demonstrate that the dog tilt is sensitive enough to identify potential OH inducers. These tests can be included in the ICH S7A/B telemetry designs to refine CV preclinical assessments prior to the first clinical trials.
doi:10.1016/j.vascn.2010.11.020
Poster Number: 17 Board Number: 17
Using the conscious telemetered rat for early in vivo ECG safety studies: QT interval measurement and correction Oladipupo Adeyemi, Sonia Roberts, Jolie Harris, Mark Dewhurst Pfizer Global Research & Development, Sandwich, United Kingdom The QT interval of the ECG represents the time from the start of ventricular depolarisation to the end of repolarisation. QT varies inversely with heart rate (HR) in dogs, primates and man; however, this relationship was previously reported to be absent in rats1. In addition, the lack of IKr-channel function in repolarisation in rats, has limited the interest in ECG measurements from this species. The aim of this study was to investigate further the utility of QT interval measurements in conscious telemetered rats. QT was measured from the Q-point to the end of the ST-complex in untreated male Sprague-Dawley rats (n = 7, 450-650 g) over 23 hrs. QT was inversely correlated to HR (r2 = 0.87), and directly correlated to RR (r2 = 0.89). QTc was calculated using the slope of the linear regression line to correct to a HR of 350 bpm. Other methods of correction (Bazett and Fridericia) were also investigated; however, these did not correct for changes in HR over the range studied. The ECG effects of two compounds, PF-x and cisapride (3, 10 and 30 mg/kg p.o.) were
Impedance is a recognized methodology for respiratory measurement; however the use of impedance respiratory monitoring in freely moving cynomolgus monkeys has not yet been reported. This study explored impedance respiratory monitoring as a novel implantable telemetry monitoring strategy. A surgically implanted telemetry transmitter with ECG (DII), arterial pressure, physical activity, body temperature, respiratory rate and tidal volume (TV) was characterized. A longitudinal incision was performed lateral to the linea alba. The internal abdominal oblique muscle was separated from the aponeurosis of the transverses abdominis by blunt dissection. A transmitter was inserted between the internal abdominal oblique muscle and the aponeurosis of the transverses abdominis. Impedance electrodes were placed subcutaneously on the lateral aspect of the thorax. After recovery, controlled tidal volumes (20, 40 and 60 mL) were used in 6 anesthetized cynomolgus females to evaluate the effects of body positions on respiratory measurements. All correlations were above 98%. Animals were followed over time to assess both accuracy and stability of telemetric impedance measurements. Slight changes in the impedance signal were noted between the surgery and two weeks post-implant. Impedance remained stable over the following 2-month monitoring period. Circadian rhythms of respiratory rate and TV were correlated with cardiovascular parameters. In conclusion, this new telemetry monitoring modality could be valuable in pharmacological investigations when the use of cynomolgus monkeys is justified.
doi:10.1016/j.vascn.2010.11.015
Poster Number: 12 Board Number: 12 Translational medicine: Can we use publicly available data in correlating preclinical with clinical studies? Ard Teismana, Pieter-Jan Gunsb, Karel Van Ammela, Rob Towarta, Nenad Sarapac, David Gallachera a
Johnson & Johnson EDC, Beerse, Belgium Formerly Johnson & Johnson EDC, Beerse, Belgium and University of Antwerp, Antwerp, Belgium c Formerly Johnson & Johnson Clinical East Coast, Raritan, NJ, United States
e5
suboptimal clinical data diminishes the reputation of preclinical models. Cross company collaboration and improved Phase I study designs may improve this situation.
doi:10.1016/j.vascn.2010.11.016
Poster Number: 13 Board Number: 13 Adaptation of safety pharmacology cardiovascular and respiratory methodology for use on juvenile toxicity studies Kevin Norton, Keith Robinson, Mark Vezina Charles River Laboratories, Senneville, Quebec, Canada The EMEA Guideline on the need for nonclinical testing in juvenile animals on human pharmaceuticals for pediatric indications and FDA Guidance Document. Nonclinical safety evaluation of pediatric drug products; require the conduct of juvenile toxicity studies intended to bridge the developmental toxicity studies adult toxicity studies. As per the EMEA guidance document, the objective of such studies is to assess “potentially different safety profiles from those seen in adults.” With new chemical entities being specifically evaluated to assess effects on overall growth of the organ systems that develop postnatally (e.g., skeletal, renal, lung, neurological, immunologic and reproductive systems). We have looked at incorporation of traditional assessments of safety pharmacology core battery assessments for CNS, respiratory and cardiovascular studies in juvenile toxicity studies. We demonstrate the use of a modified functional observation battery and whole body plethysmography to assess effects on CNS and respiratory parameters, respectively, in rats between day 10 and sexual maturation (approximately day 50). Whilst beagle dogs approximately 3 to 4 month of age were instrumented with DSI telemetry transmitters to assess cardiovascular effect. The data reported illustrate that traditional safety pharmacology can be adapted to meet the growing requirements for juvenile toxicity studies.
b
Criticism of preclinical models is based on their apparent lack of reliability to predict clinical events. In the cardiovascular (CV) safety area blockade of the IKr channel can lead to QT prolongation, which in turn may lead to TdP. Although we know that drug-induced TdP may be caused by block of IKr, we also know that some IKr blockers do not necessarily induce QT-prolongation/TdP. To answer the question “how good are current preclinical models?” one needs to review both preclinical and clinical data in a wide range of chemical and pharmacological classes. The majority of clinical data is not in the public domain. Within this project we compared both J&J internal and publicly available clinical CVdata with results of preclinical models. We evaluated several different clinical data sources and identified 80 clinical reports on 73 compounds with different levels of CV data. The data were collected in a data warehouse and compared with preclinical data. Due to the different levels of detail and large variability of designs etc., the actual number of compounds that could be used was significantly reduced. Preliminary analysis shows that TQT-data tend to have the lowest variability, and that these should preferably be used for comparison with preclinical ECG-models. Unfortunately, little of this high quality data is available in the public domain, as many compounds will not reach the clinical stage. Using
doi:10.1016/j.vascn.2010.11.017
Poster Number: 14 Board Number: 14 Measurement of bronchoconstriction in conscious rats: Elegant technique, elusive recordings Lorna C. Ewarta, Katherine Elliottb, Michael Haleyb, Tom Hollandb, Sally-Ann Rickettsc, Alan McCarthyb, Lisa Williamsb, Jean-Pierre Valentinb a
AstraZeneca, Macclesfield, Cheshire, United Kingdom Safety Assessment, AstraZeneca, Macclesfield, Cheshire, United Kingdom c DECS, AstraZeneca, Macclesfield, Cheshire, United Kingdom b
Placement of a pressure-sensitive catheter radio-telemetry device, beneath the pleural surface, enables assessment of bronchoconstriction in parallel with changes in rate and depth of breathing (Murphy et al. 1998). Here, we assess this approach using an infusion of methacholine (MCh) up to 30 μg/kg/min or vehicle (saline) via the tail vein in adapted head-out plethysmography (HOP) chambers. Resistance measured by the telemetric method in conscious male rats (18 Han Wistar, 10 Sprague Dawley) was compared to resistance
e6
Abstracts
measured by an oesophageal catheter in anaesthetised rats (32 Sprague Dawley). Surgical placement of the pressure-sensitive catheter was successful in 60% of rats as confirmed by a strong, physiological signal and high-resolution computerised tomography or histology. MCh dose dependently increased resistance in 2 telemetered rats. No apparent pharmacological response was observed in the other dosed rats due to large pressure signal variability. Baseline resistance approximately doubled in all anaesthetised rats (n = 32) given 10 μg/kg/min MCh. Variability in the telemetry signal was associated with the posture adopted by the rat or the degree of movement within the HOP chamber, especially during dosing. Despite widespread consultation, extensive evaluation of habituation protocols and confirmation of catheter tip placement, we were not able to obtain reliable broncoconstrictor responses. Given the challenges associated with setting up this model, we would favour assessment of bronchoconstriction in anaesthetised rats.
doi:10.1016/j.vascn.2010.11.018
Poster Number: 15 Board Number: 15 The use of whole body plethysmography to evaluate drug-induced changes in respiratory disorders: Sleep apneas, cough and bronchospasm Eric Delpy, Fabrice Infanti, Mathilde Loiseau, Christophe Drieu La Rochelle Biotrial Pharmacology, Rennes, France Whole body plethysmography (WBP) is widely used in safety pharmacology to investigate in conscious animals potential undesirable pharmacodynamic effects of a substance on respiratory function. Proper characterisation of a breathing pattern includes measurements of respiratory rate, tidal and minute volume, inspiratory and expiratory times and peak flows. The aim of this study is to show that WBP could also be used to study drug-induced changes in respiratory disorders such as apneas, cough or bronchospasm. Spontaneous and post-sight sleep apneas were investigated in rats. Apneas were recorded from 30 min post dosing (time required for rats to fall asleep) and over 5 hours. In saline-treated animals, the number of apneas with a duration of more than 2.5 s was 34 ± 3 and the total duration was 117 ± 12 s. Diazepam (5 mg/kg, i.p.) decreased both the number (-20%) and the duration (-38%) of apneas. Cough was investigated in guinea-pigs and was induced by a 10-min citric acid inhalation. The number of coughs was counted by a trained experimenter based on characteristic animal posture, sound produced and respiratory signal. The number of coughs in saline-treated animals was 25 ± 2. Baclofen (0.1, 0.3 and 1 mg/kg s.c.) and codeine (10, 30 and 100 mg/kg p.o.) dose-dependently decreased the number of coughs. Bronchospasm was investigated both in ovalbuminsensitised and non-sensitised guinea-pigs; bronchoconstriction was evaluated by the calculation of enhanced pause (Penh). In sensitised animals, ovalbumin (1-100 μg/kg i.v.) dose-dependently increased Penh. In non-sensitised animals, histamine (0.3-30 μg/kg i.v.) dosedependently increased Penh and this effect was inhibited by mepyramine. These results show that WBP is appropriate to evaluate the effects of new chemical entities on respiratory disorders such as sleep apneas, cough and bronchospasm.
doi:10.1016/j.vascn.2010.11.019
Poster Number: 16 Board Number: 16 Orthostatic hypotension can be investigated in conscious dogs as part of standard telemetry studies Nicola Smith, Matt Skinner, Pierre Lainee, Jackie Moors, Stewart Brown, Amy Deaville, Jean-Pierre Valentin AstraZeneca R&D, Macclesfield, Cheshire, United Kingdom Despite being a common cardiovascular adverse event in Phase 1 clinical trials, orthostatic hypotension (OH) is not routinely investigated during standard preclinical studies. The aim of this work was to validate a dog tilt model designed to assess OH. Four to six dogs implanted with DSI telemetry transmitters were orally dosed on separate occasions with Prazosin (0.01, 0.03 and 0.1 mg/kg), Captopril (3, 30 and 100 mg/kg), Verapamil (1, 5 and 15 mg/kg), Atenolol (1, 3 and 10 mg/kg) or vehicle. Heart Rate and systolic blood pressure (SBP) were recorded while the dogs were exposed to a 1-minute head-up postural change of approximately 90° to induce OH at set timepoints pre and post-dose. Under control conditions, orthostatic tilt elicited an increase in SBP maintained for the duration of the tilt and an initial transient tachycardia. Prazosin and Captopril caused a decrease in the tilt-evoked increase in SBP (-75% and –30% vs vehicle, respectively), and increased tachycardia (27% and 57% vs vehicle, respectively). Verapamil caused an increase in the tilt-evoked tachycardia (176% vs vehicle) with minimal effect on the SBP response. Atenolol caused a decrease in tilt-evoked tachycardia (-69% vs vehicle) without affecting the SBP response. (% Values show max. change). Results correlate with the potency for these reference compounds to induce OH in man and demonstrate that the dog tilt is sensitive enough to identify potential OH inducers. These tests can be included in the ICH S7A/B telemetry designs to refine CV preclinical assessments prior to the first clinical trials.
doi:10.1016/j.vascn.2010.11.020
Poster Number: 17 Board Number: 17
Using the conscious telemetered rat for early in vivo ECG safety studies: QT interval measurement and correction Oladipupo Adeyemi, Sonia Roberts, Jolie Harris, Mark Dewhurst Pfizer Global Research & Development, Sandwich, United Kingdom The QT interval of the ECG represents the time from the start of ventricular depolarisation to the end of repolarisation. QT varies inversely with heart rate (HR) in dogs, primates and man; however, this relationship was previously reported to be absent in rats1. In addition, the lack of IKr-channel function in repolarisation in rats, has limited the interest in ECG measurements from this species. The aim of this study was to investigate further the utility of QT interval measurements in conscious telemetered rats. QT was measured from the Q-point to the end of the ST-complex in untreated male Sprague-Dawley rats (n = 7, 450-650 g) over 23 hrs. QT was inversely correlated to HR (r2 = 0.87), and directly correlated to RR (r2 = 0.89). QTc was calculated using the slope of the linear regression line to correct to a HR of 350 bpm. Other methods of correction (Bazett and Fridericia) were also investigated; however, these did not correct for changes in HR over the range studied. The ECG effects of two compounds, PF-x and cisapride (3, 10 and 30 mg/kg p.o.) were