- Email: [email protected]
TOUCHING BACE: HOW INHIBITING AMYLOID-β PRODUCTION BY STERIC HINDRANCE WITH A NOVEL IMMUNOTHERAPY IMPROVES MEMORY IN PDAPP MICE
P414
Poster Presentations: Sunday, July 24, 2016
exact mechanism of action for this later effect remains unclear, but growing evidence suggests that soluble Ab oligomers are the toxic species and trigger the perturbation in calcium homeostasis observed in AD. Therefore, we evaluated the ability of aducanumab to target soluble Ab oligomers present in 3 different preparations and hence to normalize calcium homeostasis in primary neuronal cultures as a functional outcome. Methods: Size of Ab species present in the media collected from embryonic Tg2576 neurons in culture (TgCM), synthetic ADDL (Ab-Derived Diffusible Ligand) preparations, or TBS soluble fraction of 22 month old Tg2576 mouse brains, was determined by size exclusion chromatography. TgCM, ADDLs and brain extracts were immunodepleted with aducanumab, and Ab levels in the supernatants and eluates were determined by ELISA. Calcium imaging in indo-1 loaded neuronal cultures was used before and after application of immunodepleted TgCM, ADDLs, and Tg2576 mouse brain extracts. Results: TgCM contains primarily low molecular weight species of Ab. ADDLs and Tg2576 brain extracts contain both low and high molecular weight species of Ab. Aducanumab binds preferentially to high molecular weight species of Ab, as compared to low molecular weight. ADDL preparations and/or Tg2576 mouse brain extracts immunodepleted with aducanumab did not trigger elevations in calcium (calcium overload) when applied to primary neurons. Conclusions: In addition to recognizing and clearing fibrillar Ab aggregated in amyloid plaques, aducanumab targets high molecular weight soluble Ab oligomers, and restores neuronal function by normalizing calcium homeostasis.
P1-034
TOUCHING BACE: HOW INHIBITING AMYLOID-b PRODUCTION BY STERIC HINDRANCE WITH A NOVEL IMMUNOTHERAPY IMPROVES MEMORY IN PDAPP MICE
Charles E. Evans, Rhian S. Thomas, Emma J. Kidd, Mark A. Good, Cardiff University, Cardiff, United Kingdom. Contact e-mail: evansc55@ cardiff.ac.uk Background: Aberrant production or clearance of amyloid-b
(Ab) is associated with cognitive deficits and changes in NMDA receptor function. 2B3, a monoclonal antibody that binds the b-secretase cleavage site of the amyloid precursor protein (APP), inhibits production of Ab through steric hindrance in cell culture. To test the hypothesis that a reduction of Ab production in vivo by 2B3 would benefit cognition and NMDA receptor function, PDAPP transgenic mice were administered 2B3 or vehicle and tested on object-in-place (OiP) recognition memory. As well as amyloid levels, biochemical changes in NMDA receptors were measured ex vivo. Methods: At 17-18 months of age PDAPP mice were administered 2B3 (n¼10), or vehicle (n¼11) and wild-type mice received vehicle (n¼10) or no treatment (n¼11). 2B3 and vehicle were delivered via intracerebroventricular administration using osmotic minipumps for 14 days. OiP recognition memory was tested on treatment days 13-14 in an open arena. Immediately after behavioural tests, the hippocampus from all mice was dissected and snap frozen. Proteins were extracted and quantified using Western blotting and ELISA. Results: PDAPP mice administered 2B3 showed a significant improvement in OiP memory. This improved memory performance was complimented by a significant reduction of bCTF and Ab40, but not Ab42 as determined by ELISA. Western blot analysis further revealed that there was no change in total levels of NMDA receptors. However, a significant reduction in the phosphorylation of the NMDA NR2B sub-
unit was observed ex vivo. Conclusions: Our findings showed a significant improvement of place recognition memory following 2B3 administration in PDAPP mice expressing an APP mutation. Inhibition of APP metabolism by 2B3 was observed ex vivo by reduced levels of APP metabolites bCTF and Ab40. Increased phosphorylation of the NR2B subunit has been implicated in the cognitive deficits observed in AD mouse models. Thus, it is likely that the reduction in NR2B phosphorylation by 2B3 contributed to improved recognition memory. To date, no previous study investigating amyloid-based immunotherapy has reported this effect. Collectively, these results provide evidence that inhibition of APP metabolism by b-secretase is a viable target for immunotherapy and may have potential benefits in the early treatment of AD.
P1-035
NO/CGMP/CREB-BASED SYNAPTIC REPAIR AS A MULTIFUNCTIONAL THERAPEUTIC APPROACH TO DISEASE MODIFICATION IN MULTIPLE FAMILIAL MOUSE MODELS AND A NOVEL SPORADIC MODEL OF ALZHEIMER’S DISEASE
Manel Ben Aissa1, Sue H. Lee1, Jia Luo1, Ottavio Arancio2, Mary Jo LaDu3, Brian M. Bennett4, David R. Pepperberg3, Gregory R. Thatcher1, 1University of Illinois at Chicago, College of Pharmacy, Chicago, IL, USA; 2Columbia University, New York, NY, USA; 3University of Illinois at Chicago, Chicago, IL, USA; 4Queen’s University, School of Medicine, Kingston, ON, Canada. Contact e-mail: [email protected] Background: The search for Alzheimer’s disease (AD) therapeutics
singularly targeting amyloid-b and largely driven by familial AD transgenic mouse models is yet to succeed. Mindful of the multifactorial contributions to AD, a clinical drug with known neuroprotective, anti-inflammatory, and anticonvulsant actions was redesigned to address synaptic dysfunction by reactivating CREB via enhanced NO/cGMP signaling. Methods: In four diverse AD mouse models, including (EFAD) mice carrying human (h)-APO4 and a novel sporadic AD model, the treatment outcomes of the redesigned drug (NMZ) on synaptic markers, neuropathology, and cognitive deficits were assayed using behavioral, histological, and biochemical methods. Results: NMZ treatment lowered the neurotoxic forms of Ab in both APP/PS1 and 3xTg transgenic mice. Incorporation of (h)-APO4 the major genetic AD risk factor, in EFAD mice, did not mute the beneficial drug effects. In a novel sporadic mouse model that manifests ADlike pathology caused by accelerated oxidative stress, in the absence of any familial AD mutation, NMZ attenuated hallmark AD pathology, and restored biomarkers of synaptic and neuronal function. Our system pharmacology approach restored long-term potentiation (LTP) and memory deficits in all animals with robust cognitive deficits. Conclusions: Our findings demonstrate that NMZ restored CREB activation and effectively improved memory and reduced multiple AD pathological features in diverse mouse models of familial AD and a novel model not dependent on rare human genetic factors. In the search for disease-modifying AD therapeutics, targeting CREB and multiple systems associated with mixed pathology dementia and sporadic AD is highly promising. P1-036
DISSOCIATING ABERRANT MEMORY PROCESSES IN MOUSE MODELS OF ALZHEIMER’S DISEASE AND DOWN SYNDROME
Jessica H. Hall1, Mark A. Good1, Elizabeth Fisher2, Frances K. Wiseman2, John L. Harwood1, Victor L. Tybulewicz3, 1Cardiff University, Cardiff, United Kingdom; 2University College London, London, United Kingdom;
Poster Presentations: Sunday, July 24, 2016
exact mechanism of action for this later effect remains unclear, but growing evidence suggests that soluble Ab oligomers are the toxic species and trigger the perturbation in calcium homeostasis observed in AD. Therefore, we evaluated the ability of aducanumab to target soluble Ab oligomers present in 3 different preparations and hence to normalize calcium homeostasis in primary neuronal cultures as a functional outcome. Methods: Size of Ab species present in the media collected from embryonic Tg2576 neurons in culture (TgCM), synthetic ADDL (Ab-Derived Diffusible Ligand) preparations, or TBS soluble fraction of 22 month old Tg2576 mouse brains, was determined by size exclusion chromatography. TgCM, ADDLs and brain extracts were immunodepleted with aducanumab, and Ab levels in the supernatants and eluates were determined by ELISA. Calcium imaging in indo-1 loaded neuronal cultures was used before and after application of immunodepleted TgCM, ADDLs, and Tg2576 mouse brain extracts. Results: TgCM contains primarily low molecular weight species of Ab. ADDLs and Tg2576 brain extracts contain both low and high molecular weight species of Ab. Aducanumab binds preferentially to high molecular weight species of Ab, as compared to low molecular weight. ADDL preparations and/or Tg2576 mouse brain extracts immunodepleted with aducanumab did not trigger elevations in calcium (calcium overload) when applied to primary neurons. Conclusions: In addition to recognizing and clearing fibrillar Ab aggregated in amyloid plaques, aducanumab targets high molecular weight soluble Ab oligomers, and restores neuronal function by normalizing calcium homeostasis.
P1-034
TOUCHING BACE: HOW INHIBITING AMYLOID-b PRODUCTION BY STERIC HINDRANCE WITH A NOVEL IMMUNOTHERAPY IMPROVES MEMORY IN PDAPP MICE
Charles E. Evans, Rhian S. Thomas, Emma J. Kidd, Mark A. Good, Cardiff University, Cardiff, United Kingdom. Contact e-mail: evansc55@ cardiff.ac.uk Background: Aberrant production or clearance of amyloid-b
(Ab) is associated with cognitive deficits and changes in NMDA receptor function. 2B3, a monoclonal antibody that binds the b-secretase cleavage site of the amyloid precursor protein (APP), inhibits production of Ab through steric hindrance in cell culture. To test the hypothesis that a reduction of Ab production in vivo by 2B3 would benefit cognition and NMDA receptor function, PDAPP transgenic mice were administered 2B3 or vehicle and tested on object-in-place (OiP) recognition memory. As well as amyloid levels, biochemical changes in NMDA receptors were measured ex vivo. Methods: At 17-18 months of age PDAPP mice were administered 2B3 (n¼10), or vehicle (n¼11) and wild-type mice received vehicle (n¼10) or no treatment (n¼11). 2B3 and vehicle were delivered via intracerebroventricular administration using osmotic minipumps for 14 days. OiP recognition memory was tested on treatment days 13-14 in an open arena. Immediately after behavioural tests, the hippocampus from all mice was dissected and snap frozen. Proteins were extracted and quantified using Western blotting and ELISA. Results: PDAPP mice administered 2B3 showed a significant improvement in OiP memory. This improved memory performance was complimented by a significant reduction of bCTF and Ab40, but not Ab42 as determined by ELISA. Western blot analysis further revealed that there was no change in total levels of NMDA receptors. However, a significant reduction in the phosphorylation of the NMDA NR2B sub-
unit was observed ex vivo. Conclusions: Our findings showed a significant improvement of place recognition memory following 2B3 administration in PDAPP mice expressing an APP mutation. Inhibition of APP metabolism by 2B3 was observed ex vivo by reduced levels of APP metabolites bCTF and Ab40. Increased phosphorylation of the NR2B subunit has been implicated in the cognitive deficits observed in AD mouse models. Thus, it is likely that the reduction in NR2B phosphorylation by 2B3 contributed to improved recognition memory. To date, no previous study investigating amyloid-based immunotherapy has reported this effect. Collectively, these results provide evidence that inhibition of APP metabolism by b-secretase is a viable target for immunotherapy and may have potential benefits in the early treatment of AD.
P1-035
NO/CGMP/CREB-BASED SYNAPTIC REPAIR AS A MULTIFUNCTIONAL THERAPEUTIC APPROACH TO DISEASE MODIFICATION IN MULTIPLE FAMILIAL MOUSE MODELS AND A NOVEL SPORADIC MODEL OF ALZHEIMER’S DISEASE
Manel Ben Aissa1, Sue H. Lee1, Jia Luo1, Ottavio Arancio2, Mary Jo LaDu3, Brian M. Bennett4, David R. Pepperberg3, Gregory R. Thatcher1, 1University of Illinois at Chicago, College of Pharmacy, Chicago, IL, USA; 2Columbia University, New York, NY, USA; 3University of Illinois at Chicago, Chicago, IL, USA; 4Queen’s University, School of Medicine, Kingston, ON, Canada. Contact e-mail: [email protected] Background: The search for Alzheimer’s disease (AD) therapeutics
singularly targeting amyloid-b and largely driven by familial AD transgenic mouse models is yet to succeed. Mindful of the multifactorial contributions to AD, a clinical drug with known neuroprotective, anti-inflammatory, and anticonvulsant actions was redesigned to address synaptic dysfunction by reactivating CREB via enhanced NO/cGMP signaling. Methods: In four diverse AD mouse models, including (EFAD) mice carrying human (h)-APO4 and a novel sporadic AD model, the treatment outcomes of the redesigned drug (NMZ) on synaptic markers, neuropathology, and cognitive deficits were assayed using behavioral, histological, and biochemical methods. Results: NMZ treatment lowered the neurotoxic forms of Ab in both APP/PS1 and 3xTg transgenic mice. Incorporation of (h)-APO4 the major genetic AD risk factor, in EFAD mice, did not mute the beneficial drug effects. In a novel sporadic mouse model that manifests ADlike pathology caused by accelerated oxidative stress, in the absence of any familial AD mutation, NMZ attenuated hallmark AD pathology, and restored biomarkers of synaptic and neuronal function. Our system pharmacology approach restored long-term potentiation (LTP) and memory deficits in all animals with robust cognitive deficits. Conclusions: Our findings demonstrate that NMZ restored CREB activation and effectively improved memory and reduced multiple AD pathological features in diverse mouse models of familial AD and a novel model not dependent on rare human genetic factors. In the search for disease-modifying AD therapeutics, targeting CREB and multiple systems associated with mixed pathology dementia and sporadic AD is highly promising. P1-036
DISSOCIATING ABERRANT MEMORY PROCESSES IN MOUSE MODELS OF ALZHEIMER’S DISEASE AND DOWN SYNDROME
Jessica H. Hall1, Mark A. Good1, Elizabeth Fisher2, Frances K. Wiseman2, John L. Harwood1, Victor L. Tybulewicz3, 1Cardiff University, Cardiff, United Kingdom; 2University College London, London, United Kingdom;