692.29 A Central Nervous System-Penetrant Soluble ... · Disclosures: CJW, JEJ, PG, SJ, SSC, JT,...
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A Central Nervous System-Penetrant Soluble Guanylate Cyclase Stimulator Increased Cerebral Blood Flow and Modulated fMRI-BOLD Responses in Rodents Christopher J. Winrow a , Juli E. Jones a , Peter Germano a , Sarah Jacobson a , Susana S. Correia a , Kim Tang b , Jenny Tobin a , Praveen Kulkarni c , Craig Ferris c , Mark G. Currie a , Rajesh R. Iyengar a and John R. Hadcock a a Ironwood Pharmaceuticals Inc., 301 Binney Street, Cambridge, MA 02142; b Axial Biotherapeutics, 745 Boylston Street, Boston, MA 02115; c Northeastern University College of Science, Boston, MA 02116 www.ironwoodpharma.com Disrupted cerebral blood flow (CBF), diminished endothelial function, and dysregulation of the neurovascular unit are key components of several neurological diseases, and are commonly observed in many dementias. Nitric oxide (NO) signals through soluble guanylate cyclase (sGC) to produce 3',5'-cyclic guanosine monophosphate (cGMP) from guanosine-5'-triphosphate (GTP), resulting in the modulation of a variety of physiological processes including blood flow, inflammation, and endothelial and neuronal function. NO-sGC-cGMP signaling is also impaired across a variety of neurological diseases including Alzheimer’s disease and vascular dementia. Given its critical role in modulating NO signaling, sGC has emerged as a therapeutic target for diseases in which NO-sGC-cGMP signaling is impaired. IWP-247 is an orally bioavailable, central nervous system (CNS)-penetrant, small-molecule sGC stimulator that increases cGMP levels throughout the brain in animal models. Given the importance of altered CBF in neurodegenerative diseases, we evaluated the ability of IWP-247 to modulate cerebral blood flow in rodent studies. CNS Penetrant IWP-247 Increased fMRI-BOLD Signals Introduction IWP-247 Increased CBF as Measured by Cranial Laser Doppler Adapted from Tobin, Zimmer et al. J Pharmacol Exp Ther 2018;365:664-675. Copyright © 2018 The Author(s) Methods • sGC stimulators offer the potential to modulate multiple aspects of CNS diseases. • IWP-247, a CNS-penetrant sGC stimulator, increased regional CBF and activated brain regions involved in attention, learning and memory in rodent studies. • These results support further evaluation of sGC stimulation as a therapeutic target for the treatment of neurological disorders. Regional CBF was measured by cranial laser doppler in anesthetized male Sprague Dawley rats using the Perimed system. Following baseline recording, intravenous administration of the NO synthase inhibitor N(ω)-nitro-L-arginine methyl ester (L-NAME) was delivered to induce a 25% reduction in CBF. Following the reduction in CBF, subcutaneous administration of IWP-247 was evaluated for the ability to reverse the L- NAME-induced deficits in rats. fMRI-BOLD imaging was conducted in 30 awake adult male Sprague Dawley rats following intravenous dosing of vehicle, IWP-247 or a peripherally restricted sGC stimulator. Drug was delivered remotely via a tube connected to the i.v. catheter. Following multi-day acclimation training animals, were scanned using a Bruker Biospec 7.0T/20-cm USR horizontal magnet. Image acquisition, data processing and analysis were conducted as described in the workflow. fMRI BOLD Profile of a Peripheral sGC Stimulator fMRI BOLD Profile of CNS Penetrant IWP-247 Basal laser doppler perfusion monitoring (LDPM) and MAP were collected for 20 minutes prior to i.v. infusion of L-NAME (grey box). L-NAME infusion (3mg/kg i.v.) resulted in a transient increase in CBF, followed by a sustained decrease in signal (~20%) which was reversed by a single s.c. dose of the NO donor SNP (1mg/kg). Data are expressed as a percentage of the baseline values (n=4, Mean±SEM). Summary and Conclusions L-NAME infusion produced a significant reduction in CBF (##p<0.01 vs BL) that was reversed by IWP-247 at 90 minutes post-dose (**p<0.01 vs LNAME). Data show percent baseline values averaged over a 5 minute sampling period for each timepoint (n=3, Mean±SEM). Thalamus Reticular Activating System Hippocampus Image courtesy of Perimed (www.perimed-instruments.com) 0 20 40 60 80 40 60 80 100 120 140 Time (min) Percent Baseline MAP CBF L-NAME (3mg/kg/min; i.v.) BL LNAME IWP247 (30min) IWP247 (60min) IWP247 (90min) 0 50 100 150 Percent Baseline ** ## Change Over Time in CA1 Activity IWP-247 Potentially Addresses Multiple Aspects of CNS Diseases Characterization of sGC Stimulators Disclosures: CJW, JEJ, PG, SJ, SSC, JT, MGC, RRI and JRH are full time employees of Ironwood Pharmaceuticals and may own stock or stock options. CF has a financial interest in Animal Imaging Research, the company that makes the awake rat imaging system. KT is employed by Axial Biotherapeutics. SNP NO Donor (1mg/kg; s.c.) 3D representation of brain activation by IWP-247 compared to vehicle. Highlighted regions represent increased BOLD signal. Thalamus is highlighted in purple. Cortical/Hippocampal complex is highlighted in yellow. Brainstem is highlighted in green. 3D representation of brain activation by peripheral sGC stimulator compared to vehicle. Regions highlighted in yellow represent an increased BOLD signal relative to vehicle. * BLQ – Below the Limit of Quantitation (1 nM) vehicle IWP-247 Basal laser doppler perfusion monitoring (LDPM) and MAP were collected for 15 minutes prior to i.v. administration of IWP-247 (1mg/kg) or vehicle. Treatment with IWP-247 resulted in a significant increase in CBF compared to vehicle control (*p<0.05, **p<0.01). MAP was also significantly decreased compared to vehicle (#p<0.05, ##p<0.01, ####p<0.0001). Data are expressed as a percentage of the baseline values (n=4, Mean±SEM). 0 10 20 30 40 50 60 -40 -20 0 20 40 -30 -20 -10 0 10 Time (min) CBF ∆ VEH MAP ∆ VEH CBF MAP iv bolus * ** * ## #### # # # # 692.29 HHH34 Laser Doppler Monitoring of Cerebral Blood Flow fMRI BOLD Workflow – Image Capture and Analysis Effects of IWP-247 and Peripheral sGC Stimulator on Mean Arterial Pressure (MAP) IWP-247: Plasma and CNS Exposure at 30 minutes Peripheral sGC Stimulator: Plasma and CNS Exposure at 30 minutes L-NAME Reduced CBF and Increased MAP IWP-247 Restored L-NAME Induced Reduction of CBF IWP-247 Increased CBF in Normal Rats
Transcript of 692.29 A Central Nervous System-Penetrant Soluble ... · Disclosures: CJW, JEJ, PG, SJ, SSC, JT,...
A Central Nervous System-Penetrant Soluble Guanylate Cyclase Stimulator Increased Cerebral Blood Flow and Modulated fMRI-BOLD Responses in Rodents
Christopher J. Winrowa, Juli E. Jonesa, Peter Germanoa, Sarah Jacobsona, Susana S. Correiaa, Kim Tangb, Jenny Tobina, Praveen Kulkarnic, Craig Ferrisc, Mark G. Curriea, Rajesh R. Iyengara and John R. Hadcocka
aIronwood Pharmaceuticals Inc., 301 Binney Street, Cambridge, MA 02142; bAxial Biotherapeutics, 745 Boylston Street, Boston, MA 02115; cNortheastern University College of Science, Boston, MA 02116 www.ironwoodpharma.com
Disrupted cerebral blood flow (CBF), diminished endothelial function, and dysregulationof the neurovascular unit are key components of several neurological diseases, and arecommonly observed in many dementias. Nitric oxide (NO) signals through solubleguanylate cyclase (sGC) to produce 3',5'-cyclic guanosine monophosphate (cGMP)from guanosine-5'-triphosphate (GTP), resulting in the modulation of a variety ofphysiological processes including blood flow, inflammation, and endothelial andneuronal function. NO-sGC-cGMP signaling is also impaired across a variety ofneurological diseases including Alzheimer’s disease and vascular dementia. Given itscritical role in modulating NO signaling, sGC has emerged as a therapeutic target fordiseases in which NO-sGC-cGMP signaling is impaired. IWP-247 is an orallybioavailable, central nervous system (CNS)-penetrant, small-molecule sGC stimulatorthat increases cGMP levels throughout the brain in animal models. Given theimportance of altered CBF in neurodegenerative diseases, we evaluated the ability ofIWP-247 to modulate cerebral blood flow in rodent studies.
CNS Penetrant IWP-247 Increased fMRI-BOLD SignalsIntroduction IWP-247 Increased CBF as Measured by Cranial Laser Doppler
Adapted from Tobin, Zimmer et al. J Pharmacol Exp Ther2018;365:664-675. Copyright © 2018 The Author(s)
Methods
• sGC stimulators offer the potential to modulate multiple aspects of CNS diseases.
• IWP-247, a CNS-penetrant sGC stimulator, increased regional CBF and activated brain regions involved in attention, learning and memory in rodent studies.
• These results support further evaluation of sGC stimulation as a therapeutic target for the treatment of neurological disorders.
Regional CBF was measured by cranial laser doppler in anesthetized male SpragueDawley rats using the Perimed system. Following baseline recording, intravenousadministration of the NO synthase inhibitor N(ω)-nitro-L-arginine methyl ester (L-NAME)was delivered to induce a 25% reduction in CBF. Following the reduction in CBF,subcutaneous administration of IWP-247 was evaluated for the ability to reverse the L-NAME-induced deficits in rats.
fMRI-BOLD imaging was conducted in 30 awake adult male Sprague Dawley ratsfollowing intravenous dosing of vehicle, IWP-247 or a peripherally restricted sGCstimulator. Drug was delivered remotely via a tube connected to the i.v. catheter.Following multi-day acclimation training animals, were scanned using a Bruker Biospec7.0T/20-cm USR horizontal magnet. Image acquisition, data processing and analysiswere conducted as described in the workflow.
fMRI BOLD Profile of a Peripheral sGC Stimulator
fMRI BOLD Profile of CNS Penetrant IWP-247
Basal laser doppler perfusion monitoring (LDPM) andMAP were collected for 20 minutes prior to i.v.infusion of L-NAME (grey box). L-NAME infusion(3mg/kg i.v.) resulted in a transient increase in CBF,followed by a sustained decrease in signal (~20%)which was reversed by a single s.c. dose of the NOdonor SNP (1mg/kg). Data are expressed as apercentage of the baseline values (n=4, Mean±SEM).
Summary and Conclusions
L-NAME infusion produced a significant reduction inCBF (##p<0.01 vs BL) that was reversed by IWP-247at 90 minutes post-dose (**p<0.01 vs LNAME). Datashow percent baseline values averaged over a 5minute sampling period for each timepoint (n=3,Mean±SEM).
Thalamus
Reticular Activating System
Hippocampus
Image courtesy of Perimed (www.perimed-instruments.com)
0 2 0 4 0 6 0 8 04 0
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Change Over Time in CA1 Activity
IWP-247 Potentially Addresses Multiple Aspects of CNS Diseases
Characterization of sGC Stimulators
Disclosures: CJW, JEJ, PG, SJ, SSC, JT, MGC, RRI and JRH are full time employees of Ironwood Pharmaceuticals and may own stock or stock options. CF has a financial interest in Animal Imaging Research, the company that makes the awake rat imaging system. KT is employed by Axial Biotherapeutics.
SNP NO Donor(1mg/kg; s.c.)
3D representation of brain activation by IWP-247 compared to vehicle. Highlighted regions represent increased BOLD signal. Thalamus is highlighted in purple. Cortical/Hippocampal complex is highlighted in yellow. Brainstem is highlighted in green.
3D representation of brain activation by peripheral sGC stimulator compared to vehicle. Regions highlighted in yellow represent an increased BOLD signal relative to vehicle.
* BLQ – Below the Limit of Quantitation (1 nM)
vehicle
IWP-247
Basal laser doppler perfusion monitoring (LDPM) andMAP were collected for 15 minutes prior to i.v.administration of IWP-247 (1mg/kg) or vehicle.Treatment with IWP-247 resulted in a significantincrease in CBF compared to vehicle control(*p<0.05, **p<0.01). MAP was also significantlydecreased compared to vehicle (#p<0.05, ##p<0.01,####p<0.0001). Data are expressed as a percentageof the baseline values (n=4, Mean±SEM).
0 1 0 2 0 3 0 4 0 5 0 6 0-4 0
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692.29HHH34
Laser Doppler Monitoring of Cerebral Blood FlowfMRI BOLD Workflow – Image Capture and Analysis
Effects of IWP-247 and Peripheral sGC Stimulator on Mean Arterial Pressure (MAP)
IWP-247:Plasma and CNS Exposure at 30 minutes
Peripheral sGC Stimulator:Plasma and CNS Exposure at 30 minutes
L-NAME Reduced CBF and Increased MAP IWP-247 Restored L-NAME Induced Reduction of CBF
IWP-247 Increased CBF in Normal Rats
