David L. Brody - US grants

[unreadable] DESCRIPTION (provided by applicant): The candidate is an MD/PhD trained clinical neurologist whose career goal is to investigate genetic causes of susceptibility to traumatic brain injury (TBI), with emphasis in dementia and Alzheimer's disease (AD). The proposed period of mentored scientific training in the laboratory of Dr. David Holtzman at Washington University will allow the candidate develop the scientific skills to become an independent investigator. The central hypotheses that will be tested during the proposed project are as follows: 1) TBI causes increased production and/or decreased clearance of the amyloid-beta peptide (A-beta) in the brain. 2) The resulting increase in A-beta concentration contributes significantly to both acute cognitive impairment and the chronic pathology of AD in humans and in transgenic mice. 3) Blocking A-beta production or enhancing its clearance will reduce cognitive and pathological sequelae of TBI. 4) The increased vulnerability to TBI and increased risk of AD conferred by the apolipoprotein E e4 genotype (APOE4) occurs largely via apoE's effects on A-beta handling and metabolism. 5) Preventing TBI related changes in A-beta will lessen the adverse outcomes conferred by APOE4 after TBI. The candidate proposes to test these hypotheses using double transgenic mice that produce both apoE and a mutant amyloid precursor protein, and hence deposit A-beta in Alzheimer's disease-like pathological patterns. Experimental TBI will be performed on these mice in collaboration with Drs. Tracy McIntosh and Phil Bayly. Cognitive outcomes will be assessed using the Morris water maze and other behavioral tasks, and histological analysis of TBl-induced lesions and AD-like pathological changes will be made. Interventions will include administration of an anti-A-beta antibody that enhances A-beta clearance from the brain and a gamma secretase inhibitor that blocks A-beta production. Mechanistic studies of A-beta concentrations measured in brain homogenates and in vivo using microdialysis will be performed to gain insight into the underlying mechanisms involved. This work may lead to improved therapeutic options for patients with TBI in the future. [unreadable] [unreadable]

DESCRIPTION (provided by applicant): Traumatic brain injury (TBI) is a major cause of death and disability, especially among young people. Pathological processes involving the amyloid-beta (A[unreadable]) peptide and tau protein are accelerated by acute TBI. Moderate to severe TBI substantially increases the risk of dementia of the Alzheimer type later in life. A[unreadable] deposition is a major pathological hallmark of Alzheimer disease (AD) and pathological tau abnormalities are seen in AD and many other neurodegenerative conditions. These A[unreadable] and tau abnormalities may also contribute to short-term adverse outcomes, as some forms of these proteins can have immediately neurotoxic effects. Additional studies have found that A[unreadable] can cause or accelerate tau pathology under some conditions, thus A[unreadable] may play a central role in both pathologies. However, the mechanisms underlying A[unreadable] and tau abnormalities following TBI are incompletely understood, in large part due to the lack of an appropriate small animal model. Preliminary results indicate that experimental controlled cortical impact TBI in 3xTg-AD mice results in intra-axonal A[unreadable] deposition in the injured fimbria and accelerated tau pathology in the contralateral hippocampus within 24 hours of injury. 3xTg-AD mice produce both human A[unreadable] and human tau with mutations that accelerate pathology, but in our experiments they were injured at young ages before significant neuropathological abnormalities are normally present. The long-term objective of the proposed project is to test the central hypothesis that A[unreadable] is a key mediator of adverse behavioral and neuropathological outcomes following TBI. The specific aims are 1) to determine the mechanisms responsible for accelerated A[unreadable] deposition following TBI in the 3xTg-AD mouse model, 2) to determine if TBI-induced A[unreadable] overproduction plays a causal role in tau hyperphosphorylation and accumulation, and 3) to determine the effects of TBI-induced acute A[unreadable] accumulation on sub-acute outcomes including behavioral performance axonal injury, neuronal cell loss, and tau pathology. Methods used will include a) colocalization studies of amyloid precursor protein (APP) and A[unreadable] with several proteolytic enzymes thay may cleave APP to produce A[unreadable], b) pharmacological or genetic inhibition of several potential A[unreadable] producing enzymes, c) analysis of tau pathology in several lines of transgenic mice with and without human A[unreadable], d) assessment of behavioral performance in mice treated with A[unreadable] -targeted therapeutics or with genetic manipulations that eliminate A[unreadable] production. If successful, these experiments will result in an improved understanding of the mechanisms underlying A[unreadable] and tau abnormalities following TBI. This may facilitate development of therapeutic strategies targeting A[unreadable] and tau, which could ultimately improve outcomes and reduce the subsequent risk of dementia and adverse cognitive outcomes in TBI patients. PUBLIC HEALTH RELEVANCE: Traumatic brain injury- such as from motor vehicle accidents, falls, assault, or military operations- is a common cause of impaired thinking and memory immediately after the injury and increases the chances of developing Alzheimer's disease later in life. The goal of this project is to use genetically engineered mice to improve our understanding of the causes of these thinking and memory problems and increased risk of Alzheimer's disease. If successful, the results of the project may point to ways to develop effective treatments to prevent or reverse these problems in the future.

DESCRIPTION (provided by applicant): Traumatic brain injury (TBI) has been a major cause of death and threat to optimal brain function throughout the entire history of human existence. Delayed secondary injury processes that could be reversed or prevented are important therapeutic targets for TBI. The focus of this project is on secondary injury processes involving the protein tau. Abnormal aggregations of tau have been detected hours to days after injury in the brains of many TBI patients. Tau may be involved in the early development of dementia similar to Alzheimer's disease, and tau is the hallmark of chronic traumatic encephalopathy seen in boxers, football players, and military personnel with TBIs. Importantly, an up-to-date definitio of the spectrum of tau pathology now includes not only tau that is visible under the microscope, but also small clusters of tau called oligomers and aggregated forms of tau that can spread through the brain by triggering additional tau aggregation. However, the mechanisms underlying tau pathologies following TBI are not understood, in large part due to the lack until recently of a appropriate small animal model. To address this, we developed the first transgenic mouse model which recapitulates many aspects of tau pathology following experimental TBI. 1) For our first aim, we now propose to test the hypothesis that tau oligomers and spreading tau aggregation contribute to delayed brain degeneration following TBI. These studies will be performed in collaboration with Dr. Rakez Kayed at the University of Texas, Galveston and Dr. Marc Diamond at Washington University, two researchers working at the cutting edge of tau biology. 2) Intriguingly, treatment with an inhibitor of an enzyme called c-jun N-terminal kinase (JNK) before injury reduced TBI-related tau pathology in the brain. For our second aim, we propose to test whether treatment with a JNK inhibitor at therapeutically realistic times after injury blocks tau pathology and improves outcomes in mice. We will assess both short term and longer-term pathological and behavioral outcomes, including innovative tests of social behavior and mood regulation in mice. 3) There are three types of JNK in the brain called JNK1, JNK2 and JNK3. Mice without JNK1 or JNK2 have problems with immune system function whereas mice without JNK3 are actually protected from other types of brain insults. Our hypothesis is that JNK3 is playing a key role and that JNK3 would make a better and safer target for new therapeutics than nonselective JNK inhibitors. We propose for our third aim to create antisense oligonucleotides specifically targeting each type of JNK. We will treat mice with these antisense oligonucleotides and determine whether this reduces tau pathology and improves outcomes following TBI. These antisense oligonucleotides have the potential to be used in human TBI patients, as there are two antisense therapeutics already approved and approximately 35 more in various stages of human clinical trials. Our collaborators, Dr. T.M. Miller at Washington University and Dr. Eric Swayze of Isis Pharmaceuticals are among the world's experts in antisense treatments. The broad, long term goals are to uncover the mechanisms leading to secondary injury processes involving tau and develop therapeutics to block them, with the hope that this would improve outcomes following TBI. The worldwide interest in athletes and military personnel with tau pathology and chronic traumatic encephalopathy underscores the urgency of this line of investigation.

PROJECT SUMMARY Clinical dementia of the Alzheimer's type (DAT) is the most common dementing disorder in the elderly. DAT is most tightly correlated with synaptic loss in vulnerable brain regions, which has led to the hypothesis that loss of synaptic terminals is a key event in early cognitive decline. To date, the mechanisms of synaptic loss in DAT remain unknown. Previous researchers have assumed that specific molecular entities such as tau or amyloid-beta (A?) are responsible for synaptic degeneration in DAT, but have not performed unbiased screens. Our primary hypothesis is there are as-yet- uncharacterized synaptotoxic substances in brain lysates from patients with DAT that are not present in brain lysates from high pathology, age matched, non-demented controls. We further hypothesize these substances cause synapses to deteriorate in the early phase of DAT which eventually leads to neuronal cell death and severe cognitive decline. To test our hypothesis, we have recently developed a novel, unbiased, high content screening approach using live primary hippocampal neurons from mice genetically manipulated to express a red fluorescent pre-synaptic tag and a green fluorescent post-synaptic tag. This new approach allows us to measure the synapse gain/loss of cultured live neurons serially over several days in a highly sensitive and fully quantitative fashion. We therefore propose to use our novel, unbiased screening approach to identify as-yet-uncharacterized synaptotoxic substances in brain lysates from patients with DAT and to further characterize the synaptotoxic activity of the newly-identified substances. Specific aims of the proposed studies are: 1) To determine whether there are synaptotoxic substances in aqueous brain lysates from patients with DAT using our novel live cell screening approach for synaptic toxicity, as well as how consistent the toxicity is from patient to patient; 2) To determine the specificity of the toxic entities from patients with DAT by assessing brain lysates from non-demented control subjects with A? and tau pathology, and normal control subjects; 3) To use our live neuron screening of fractionated aqueous brain lysates from patients with DAT for synaptic toxicity to guide purification of the most potent and specific toxic entities, and then identify the previously uncharacterized synaptotoxic substances in patients with DAT. If successful, we expect to identify a list of previously-uncharacterized synaptotoxic proteins, peptides or other substances specific to patients with DAT. Once the synaptotoxic activities of these substances have been characterized, we will target these novel synaptotoxic agents as new therapeutic targets and perform a drug screening for new candidate therapeutic molecules. The results of our proposed experiments may fundamentally advance knowledge of DAT as well as improve the effectiveness of DAT treatment by identifying targets for new candidate therapeutics.