P4-304: Therapeutic effects of a novel phytochemical formulation phyllsarone: 2H in animal models...
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tia. In epidemiological studies the presence of diabetes increased the risk of Alzheimer’s disease (AD) by 50-100% as well. The effect seems largely independent of other, so-called vascular risk factors. Methods: Literature search has been conducted to unravel the mechanisms mediating the association between DM and AD. Results: Putative hypotheses explaining this association comprise chronic brain hypoperfusion due to macro- and microangiopathy, the toxic effects of hyperglycaemia itself (damage to the blood-brain barrier), and the mediating role of insulin. Brain has for long been considered an insulin-independent organ; the discovery of insulin and its receptors in the central nervous system falsified this theory. Physiologic concentrations of insulin exert a beneficial effect on cognition. On the other hand, the risk of AD is significantly increased by the state of central (brain) hypoinsulinaemia, even in people not suffering from DM. Brain insulin deficiency can be secondary to either too low a concentration of insulin in the periphery or - on the contrary - the state of peripheral hyperinsulin- aemia, usually as a result of insulin resistance, a typical phenomenon in DM. Several mechanisms mediate between the state of central hypoinsu- linaemia and the acceleration of Alzheimer pathology generation: decline of glucose utilization, particularly in the hippocampus and enthorinal cortex; increased oxidative stress associated with the synthesis of advanced glycation endproducts (AGE); increased tau protein phosphorylation and neurofibrillary tangle formation; increased aggregation of beta-amyloid protein secondary to the insulin-degrading enzyme (IDE) inhibition. Ther- apeutic strategies targeted at restoring the balance in insulin metabolism in AD - applying nasal insulin or using thiazolidinedions - are currently in the phase of clinical trials. Conclusions: The role of central insulin distur- bances in AD etiologty seems underestimated. Insulin deficiency in the brain is involved in multiple mechanisms associated with AD development and progression. P4-302 LIPOPROTEIN-INDUCED CHROMOSOME MISSEGREGATION IN VITRO AND IN VIVO: IMPLICATIONS FOR CARDIOVASCULAR AND ALZHEIMER’S DISEASE Antoneta Granic 1,2 , Huntington Potter 1,2 , 1 Johnnie B. Byrd Sr. Alzheimer’s Center and Research Institute, Tampa, FL, USA; 2 Molecular Medicine and Suncoast Alzheimer’s & Gerontology Ctr., Tampa, FL, USA. Contact e-mail: [email protected] Background: An increasing body of evidence links the complex neurop- athy of Alzheimer’s disease (AD) to cardiovascular risk factors and ath- erosclerosis in older adults. High cholesterol may be a common feature linking the two diseases. Elevated low density lipoprotein (LDL) choles- terol is an important risk factor for the development of atherosclerosis and for AD. AD is characterized by the abnormal accumulation of Abeta in the brain and cerebral blood vessels. The production of Abeta is promoted by high cholesterol and reduced by low cholesterol levels. Thus, cholesterol homeostasis might be directly involved in amyloid plaque formation. The development of chromosomal instability has been implicated in both AD and atherosclerosis. Analysis of atherosclerotic plaques by X inactivation and FISH analysis indicate clonal hyperproliferation of smooth muscle cells with trisomy 7 and monosomy 11 as the most predominant chromo- somal instability. Thus, the development of atherosclerotic lesions may be similar to the development of benign aneuploidy tumors. Likewise, AD patients also harbor aneuploid cells in their skin, blood and brain. Thus, we hypothesize that high levels of lipoprotein, a common factor in both diseases, might cause chromosomal instability and mitotic defects. Meth- ods: hTERT-HME1 cells, human primary aortic smooth muscle cells, normal human fibroblasts and fibroblasts harboring mutated LDL receptor genes were treated with 20 ug/ml oxidized LDL, LDL or HDL or 4 ug/ml water soluble cholesterol and analyzed by karyotype and FISH analysis for chromosomes 12, 21 and 7. Results: Lipoproteins and cholesterol induce chromosome missegregation within 48 h in all normal cell types studied. However, human fibroblasts harboring mutated LDL receptor genes did not develop aneuploidy. These results indicate that functional LDLR is re- quired for the aneugenic effect of LDL cholesterol in vitro. To further test the hypothesis that elevated levels of LDL cholesterol contributes to the accumulation of aneuploid cells, we are investigating peripheral blood samples from cognitively intact subjects with high, borderline high and low levels of cholesterol by FISH analysis. Conclusions: Together, these results suggest a novel mechanism by which cholesterol promotes both atherosclerosis and AD by inducing chromosome missegregation and ac- cumulation of aneuploidy cells. P4-303 SORAFENIB AS A THERAPEUTIC AGENT AGAINST ALZHEIMER’S DISEASE Valentina Echeverria Moran 1 , Joyonna Gamble-George 1 , Sarah Burgess 1 , Alexander Dickson 2 , Malgorzata Mamcarz 3 , Gary W. Arendash 3 , 1 Bay Pines VA Healthcare System, Bay Pines, FL, FL, USA; 2 Johnnie B. Byrd, Sr. Alzheimer’s Center & Research Institute, Tampa, FL, USA; 3 Johnnie B. Byrd, Sr. Alzheimer’s Center & Research Institute, Tampa, FL, USA. Contact e-mail: [email protected] Background: Alzheimer’s disease (AD) is a neurodegenerative condition that is characterized by neuronal and synaptic loss and increasing deteri- oration of higher-order cognitive functions. For the past few years we have been studying the usefulness of pharmacological inhibition of cell signal- ing factors that promote synthesis of amyloid (A) peptide and neuroin- flammation. Specifically, we have been investigating therapeutic strategies to reduce neuronal loss and to restore or ameliorate cognitive functions in AD. We have shown that the inactive form of cRaf-1 is upregulated in human AD brains and in the hippocampus and cortex of AD mice express- ing the Swedish familial AD mutation. We also reported that the inhibition of Raf kinase activity protects cortical neurons against A toxicity in vitro and that the treatment of APPswe mice, for two months with the oral Raf inhibitor Bay-43-900 (Sorafenib), restores working memory as analyzed using the interference test. Methods: We analyzed by western blot the expression of several neuroinflammatory factors and the APP levels in the hippocampus and cortex of mice after two months of treatment with Sorafenib. Results: The neurochemical analysis of brain tissues from APPswe mice and wild type age matched control mice treated and un- treated with Sorafenib showed that APPswe mice present a decrease in the levels of the inhibitor of NFB, IB-. Sorafenib was able to normalize IB- expression to the level found in age-matched wild type control littermate mice. It has been suggested that the inhibition of NFB mitigates the synthesis of A and its toxic effect by controlling the expression of APP, and the gamma and beta secretases. Consistent with this idea, we found that the increase of IB- is accompanied by a reduction in APP levels in hippocampus and cortex of APPswe mice. Conclusions: We postulate that Sorafenib improves memory and protects cortical cells against amyloid toxicity by decreasing neuroinflammation in the brain. P4-304 THERAPEUTIC EFFECTS OF A NOVEL PHYTOCHEMICAL FORMULATION PHYLLSARONE: 2H IN ANIMAL MODELS RELEVANT TO ALZHEIMER’S DISEASE Hanumanthachar Joshi 1,2 , Milind Parle 3 , Chandrashekhar Disale 1 , 1 SET College of Pharmacy, Dharwad, India; 2 Division of Neuropharmacology, Dept. of Postgraduate Studies and Research, SET College of Pharmacy, Dharwad, India; 3 Division of Pharmacology, Dept. of Pharm Sciences, Guru Jambheshwar University of Science and Technology,, Hisar, Haryana,, India. Contact e-mail: [email protected] Background: There has been a steady rise in the number of patients suffering from Alzheimer’s disease (AD) all over the world. Alzheimer’s disease is a genetically heterogeneous neurodegenerative disorder, which is slow in onset but relentless in progress. It is the most common form of onset of adult dementia and attention deficit disorders. Newer drugs are continually introduced into the market with little efficacy. Hence it is worthwhile to explore the utility of traditional medicines in treatment of T760 Poster Presentations P4:
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Transcript of P4-304: Therapeutic effects of a novel phytochemical formulation phyllsarone: 2H in animal models...
tia. In epidemiological studies the presence of diabetes increased the risk ofAlzheimer’s disease (AD) by 50-100% as well. The effect seems largelyindependent of other, so-called vascular risk factors. Methods: Literaturesearch has been conducted to unravel the mechanisms mediating theassociation between DM and AD. Results: Putative hypotheses explainingthis association comprise chronic brain hypoperfusion due to macro- andmicroangiopathy, the toxic effects of hyperglycaemia itself (damage to theblood-brain barrier), and the mediating role of insulin. Brain has for longbeen considered an insulin-independent organ; the discovery of insulin andits receptors in the central nervous system falsified this theory. Physiologicconcentrations of insulin exert a beneficial effect on cognition. On the otherhand, the risk of AD is significantly increased by the state of central (brain)hypoinsulinaemia, even in people not suffering from DM. Brain insulindeficiency can be secondary to either too low a concentration of insulin inthe periphery or - on the contrary - the state of peripheral hyperinsulin-aemia, usually as a result of insulin resistance, a typical phenomenon inDM. Several mechanisms mediate between the state of central hypoinsu-linaemia and the acceleration of Alzheimer pathology generation: declineof glucose utilization, particularly in the hippocampus and enthorinalcortex; increased oxidative stress associated with the synthesis of advancedglycation endproducts (AGE); increased tau protein phosphorylation andneurofibrillary tangle formation; increased aggregation of beta-amyloidprotein secondary to the insulin-degrading enzyme (IDE) inhibition. Ther-apeutic strategies targeted at restoring the balance in insulin metabolism inAD - applying nasal insulin or using thiazolidinedions - are currently in thephase of clinical trials. Conclusions: The role of central insulin distur-bances in AD etiologty seems underestimated. Insulin deficiency in thebrain is involved in multiple mechanisms associated with AD developmentand progression.
P4-302 LIPOPROTEIN-INDUCED CHROMOSOMEMISSEGREGATION IN VITRO AND IN VIVO:IMPLICATIONS FOR CARDIOVASCULAR ANDALZHEIMER’S DISEASE
Antoneta Granic1,2, Huntington Potter1,2, 1Johnnie B. Byrd Sr.Alzheimer’s Center and Research Institute, Tampa, FL, USA;2Molecular Medicine and Suncoast Alzheimer’s & Gerontology Ctr.,Tampa, FL, USA. Contact e-mail: [email protected]
Background: An increasing body of evidence links the complex neurop-athy of Alzheimer’s disease (AD) to cardiovascular risk factors and ath-erosclerosis in older adults. High cholesterol may be a common featurelinking the two diseases. Elevated low density lipoprotein (LDL) choles-terol is an important risk factor for the development of atherosclerosis andfor AD. AD is characterized by the abnormal accumulation of Abeta in thebrain and cerebral blood vessels. The production of Abeta is promoted byhigh cholesterol and reduced by low cholesterol levels. Thus, cholesterolhomeostasis might be directly involved in amyloid plaque formation. Thedevelopment of chromosomal instability has been implicated in both ADand atherosclerosis. Analysis of atherosclerotic plaques by X inactivationand FISH analysis indicate clonal hyperproliferation of smooth musclecells with trisomy 7 and monosomy 11 as the most predominant chromo-somal instability. Thus, the development of atherosclerotic lesions may besimilar to the development of benign aneuploidy tumors. Likewise, ADpatients also harbor aneuploid cells in their skin, blood and brain. Thus, wehypothesize that high levels of lipoprotein, a common factor in bothdiseases, might cause chromosomal instability and mitotic defects. Meth-ods: hTERT-HME1 cells, human primary aortic smooth muscle cells,normal human fibroblasts and fibroblasts harboring mutated LDL receptorgenes were treated with 20 ug/ml oxidized LDL, LDL or HDL or 4 ug/mlwater soluble cholesterol and analyzed by karyotype and FISH analysis forchromosomes 12, 21 and 7. Results: Lipoproteins and cholesterol inducechromosome missegregation within 48 h in all normal cell types studied.However, human fibroblasts harboring mutated LDL receptor genes did notdevelop aneuploidy. These results indicate that functional LDLR is re-quired for the aneugenic effect of LDL cholesterol in vitro. To further test
the hypothesis that elevated levels of LDL cholesterol contributes to theaccumulation of aneuploid cells, we are investigating peripheral bloodsamples from cognitively intact subjects with high, borderline high and lowlevels of cholesterol by FISH analysis. Conclusions: Together, theseresults suggest a novel mechanism by which cholesterol promotes bothatherosclerosis and AD by inducing chromosome missegregation and ac-cumulation of aneuploidy cells.
P4-303 SORAFENIB AS A THERAPEUTIC AGENTAGAINST ALZHEIMER’S DISEASE
Valentina Echeverria Moran1, Joyonna Gamble-George1,Sarah Burgess1, Alexander Dickson2, Malgorzata Mamcarz3,Gary W. Arendash3, 1Bay Pines VA Healthcare System, Bay Pines, FL,FL, USA; 2Johnnie B. Byrd, Sr. Alzheimer’s Center & ResearchInstitute, Tampa, FL, USA; 3Johnnie B. Byrd, Sr. Alzheimer’s Center &Research Institute, Tampa, FL, USA. Contact e-mail:[email protected]
Background: Alzheimer’s disease (AD) is a neurodegenerative conditionthat is characterized by neuronal and synaptic loss and increasing deteri-oration of higher-order cognitive functions. For the past few years we havebeen studying the usefulness of pharmacological inhibition of cell signal-ing factors that promote synthesis of amyloid � (A�) peptide and neuroin-flammation. Specifically, we have been investigating therapeutic strategiesto reduce neuronal loss and to restore or ameliorate cognitive functions inAD. We have shown that the inactive form of cRaf-1 is upregulated inhuman AD brains and in the hippocampus and cortex of AD mice express-ing the Swedish familial AD mutation. We also reported that the inhibitionof Raf kinase activity protects cortical neurons against A� toxicity in vitroand that the treatment of APPswe mice, for two months with the oral Rafinhibitor Bay-43-900 (Sorafenib), restores working memory as analyzedusing the interference test. Methods: We analyzed by western blot theexpression of several neuroinflammatory factors and the APP levels in thehippocampus and cortex of mice after two months of treatment withSorafenib. Results: The neurochemical analysis of brain tissues fromAPPswe mice and wild type age matched control mice treated and un-treated with Sorafenib showed that APPswe mice present a decrease in thelevels of the inhibitor of NFB, IB-�. Sorafenib was able to normalizeIB-� expression to the level found in age-matched wild type controllittermate mice. It has been suggested that the inhibition of NFB mitigatesthe synthesis of A� and its toxic effect by controlling the expression ofAPP, and the gamma and beta secretases. Consistent with this idea, wefound that the increase of IB-� is accompanied by a reduction in APPlevels in hippocampus and cortex of APPswe mice. Conclusions: Wepostulate that Sorafenib improves memory and protects cortical cellsagainst amyloid toxicity by decreasing neuroinflammation in the brain.
P4-304 THERAPEUTIC EFFECTS OF A NOVELPHYTOCHEMICAL FORMULATIONPHYLLSARONE: 2H IN ANIMAL MODELSRELEVANT TO ALZHEIMER’S DISEASE
Hanumanthachar Joshi1,2, Milind Parle3, Chandrashekhar Disale1,1SET College of Pharmacy, Dharwad, India; 2Division ofNeuropharmacology, Dept. of Postgraduate Studies and Research, SETCollege of Pharmacy, Dharwad, India; 3Division of Pharmacology,Dept. of Pharm Sciences, Guru Jambheshwar University of Science andTechnology,, Hisar, Haryana,, India. Contact e-mail:[email protected]
Background: There has been a steady rise in the number of patientssuffering from Alzheimer’s disease (AD) all over the world. Alzheimer’sdisease is a genetically heterogeneous neurodegenerative disorder, which isslow in onset but relentless in progress. It is the most common form ofonset of adult dementia and attention deficit disorders. Newer drugs arecontinually introduced into the market with little efficacy. Hence it isworthwhile to explore the utility of traditional medicines in treatment of
T760 Poster Presentations P4:
cognitive disorders. The Indian system of medicine, Ayurveda has been inpractice since 5000 years. Objective: The main goal of the studies reportedhere is to investigate the cerebro-protective and memory improving poten-tials of PHYLLSARONE-2H. Methods: �-amyloid, scopolamine, ibotenicacid, CO2 and aging induced amnesia, KCN and Carotid artery ligation-induced hypoxia were the experimental models. Basal forebrain lesioninduced decrease in cerebral Ach and ChAT activity were assessed, Con-centrations of Norepinerphrine, Epinephrine, Dopamine, 5-HT in cerebralcortex, cerebellum, hypothalamus, hippocampus, pons and corpus striatumwere measured by HPLC analysis. Neuromorphology, contents of MDA,NO, activities of SOA and CAT were also Measured. Exteroceptive modelssuch as Water maze, Hebb william’s maze, Radial arm maze were used toassess acquistion and retention. Results: Neuromorphology, contents ofMDA, NO, activities of SOA and CAT were Measured which indicatedthat PHYLLSARONE-2H significantly protected the experimental animalsfrom hypoxia, stress, amnesia and neurodegeneration. They inhibited KCNand Carotid artery ligation induced hypoxia, reversed amnesia and neuro-degeneration induced by �-amyloid, scopolamine, ibotenic acid, CO2 andbrain aging, produced normalizing action on discrete regions of brain andcontrolled alterations in neurotransmitter levels due to neurodegeneration.They also decreased ChAT activity in the parietal cortex and inhibiteddecrease in Ach levels in both parietal and frontal cortex in amnesicrats.Neurohistochemistry revealed that both PHYLLSARONE-2H pro-foundly reduced the neurodegeneration in the experimental animals. It alsoimproved acquistion and retention capacities of the animals. Conclusions:PHYLLSARONE-2H, profoundly reduced the neurodegeneration, improvedlearning and memory in the experimental animals. Hence PHYLLSA-RONE-2H can be of beneficial in the treatment of dementia of Alzheimer’stype and other neurodegenerative disorders.
P4-305 ALLOSTERIC POTENTIATION OF THE M1MUSCARINIC RECEPTOR PROVIDESUNPRECEDENTED SELECTIVITY AND A NOVELTHERAPEUTIC STRATEGY FOR THETREATMENT OF ALZHEIMER’S DISEASE
William J. Ray, Matthew Seager, Lei Ma, Marion Wittmann,Krista Getty, Michael Marlatt, Ming-Chih Crouthamel, Guoxin Wu,Sethu Sankaranarayananan, Adam Simon, Maryann Burno, Keith Jones,Valerie Kuzmick Graufields, Denise Bickel, Diane Posavec,Jacquelynn Cook, Lone Veng, Scott Kuduk, Cyrille Sur, William Shipe,Craig Lindsley, Gene Kinney, Danette Pascarella, Marlene Jacobson,Guy Seabrook, Merck Research Labs, West Point, PA, USA. Contacte-mail: [email protected]
Background: Alzheimer’s disease (AD) patients suffer from an inexorableloss of cognitive function due in part to the degeneration of the basalforebrain cholinergic projection neurons. Restoring signalling through thepost-synaptic muscarinic acetylcholine receptors that mediate cholinergicsignalling is thus a promising therapeutic strategy for the symptomaticmanagement of AD. Moreover muscarinic receptor agonists reduce A�42in animal models, suggesting that this class of compounds could addressthe underlying disease pathology. Unfortunately, multiple non-selectivemuscarinic receptor agonists have been tested in AD patients and were nottolerated due to unwanted peripheral cholinergic stimulation. Improvedselectivity for M1, the most prominently expressed post-synaptic musca-rinic receptor in the cortex and hippocampus, could address these tolera-bility issues, however the high conservation of the acetylcholine bindingsite between receptor subtypes has precluded the discovery of selectiveagonists. Objective: To achieve selectivity for M1 by discovering andexploiting an allosteric ligand binding site. Methods: We screened achemical library for positive allosteric modulators of the M1 receptor andassessed chemical leads for selectivity in vitro and efficacy in rodent ADmodels in vivo. Results: We identified Benzyl Quinolone Carboxylic Acid(BQCA), a small molecule potentiator selective for M1. In CHO cellsexpressing recombinant human receptor, BQCA sensitizes M1 to acetyl-choline 83.9-fold (inflection point � 449 nM), while having no potentia-
tion, agonist, or antagonist effect on M2, M3, or M4 receptors up to 100�M. In the mouse contextual fear conditioning model of episodic-likememory, BQCA fully reversed the cognitive impairment caused by thenon-selective muscarinic antagonist scopolamine, demonstrating a criticalrole for M1 in this type of memory. Importantly BQCA does not showsigns of unwanted peripheral cholinergic stimulation at doses that producecentral physiological responses. In vivo BQCA and other muscarinic ago-nists can specifically reduce cortical A�42 in some experiments. However,this effect is likely complex and indirect, since it does not occur in culturedneurons and is not consistently observed across species and disease models.Conclusions: BQCA is a highly selective pharmacological reagent forunderstanding the normal physiology of the M1 receptor and its potentialas a therapeutic target for AD.
P4-306 CHARACTERIZATION OF SEROTONIN 5HT6RECEPTOR ANTAGONISTS AS PUTATIVEDRUGS FOR AGE-RELATED MILD COGNITIVEIMPAIRMENT AND ALZHEIMER’S DISEASE
Wayne B. Rowe, Partick M. Callahan, Cathleen Hsu,Rachael DiSomma, Ashok Tehim, David A. Lowe, MemoryPharmaceuticals, Montvale, NJ, USA. Contact e-mail:[email protected]
Background: Serotonin 5HT6 receptors are almost exclusively located inthe CNS with high expression in the hippocampus, cerebral cortex, nucleusaccumbens and striatum. The presence of this receptor in brain areasknown to be involved in learning and memory, coupled with the findingthat 5HT6 receptor antagonists stimulate acetylcholine and glutamate re-lease, suggests an important role in cognition and restoration of cognitiveimpairment. Methods: Behavioral data using a variety of cognitive tasksstrongly supports this contention. Results: Improved episodic memoryperformance has been observed in the novel object recognition task inyoung rats following a 48 hour delay with several 5HT6 antagonistsincluding MEM 34551. In the aged-impaired rat Morris water maze task,significant improvements in spatial memory performance has been ob-served with several 5HT6 receptor antagonists. Similar findings were alsoobserved in the passive avoidance task using aged animals. These laterstudies support a role for 5HT6 receptor antagonists in enhancing cognitivedeficits associated with age-related mild cognitive impairment (MCI) andAlzheimer’s disease. Interestingly, 5HT6 receptor density was shown to bevery low in a variety of mouse strains including the C57/BL6 mouse. Wetested several 5HT6 antagonist compounds in our aged-impaired C57/BL6mouse Morris water maze task and saw no cognitive restoration effects,thus supporting the neuroanatomical observations. Conclusions: Takentogether, these data support a role for 5HT6 receptor antagonists in learningand memory processing and highlight their potential as therapeutic agentsin MCI and Alzheimer’s disease patients.
P4-307 PN401 TREATMENT IMPROVES MEMORY INTg2576 AND TAPP MODELS OF ALZHEIMER’SDISEASE
Joel A. Saydoff, Ana Olariu, Zhongyi Hu, Jin Sheng, Jiong Pei, Qin Li,Rolando Garcia, Denise Brenneman, Reid W. von Borstel, WellstatTherapeutics, Gaithersburg, MD, USA. Contact e-mail:[email protected]
Background: PN401 is an orally bioavailable prodrug of uridine thatenables delivery of therapeutic concentrations of 20 - 200 �M plasmauridine in humans. PN401 has shown neuroprotective activity in models ofParkinson’s disease, Huntington’s disease in addition to models of strokeand neuroinflammation. Alzheimer’s disease (AD) has been also reportedto be associated with age-related mitochondrial dysfunction, amyloidplaques, neurofibrillary tangles, oxidative stress and neuroinflammation.Uridine and uridine precursors have been shown to improve memory.Methods: In this study we have tested the effect of PN401 administered inchow on memory in two transgenic animal models of AD. Human neuro-
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