Michael Paul Murphy - US grants

[unreadable] DESCRIPTION (provided by applicant): Investigation into the genetic causes of the muscular dystrophies has provided unique insights into the mechanisms of several disease processes. Type II Myotonic Dystrophy is caused by a large expansion in the ZNF9 gene, which encodes the single strand RNA binding protein CNBP (Cellular Nucleic Acid Binding Protein). Remarkably, we recently discovered that CNBP is involved in regulating the activity of [unreadable]-secretase, the enzyme that produces the first cleavage event in the generation of the amyloid-[unreadable] peptide (A[unreadable]). The progressive fibrillization and deposition of A[unreadable] is widely believed to be the primary causal factor in the development of Alzheimer's disease. Importantly, the activity and protein levels of the major [unreadable]-secretase enzyme in the brain (BACE1) increase in both Alzheimer's disease and in normal aging. This implicates regulators of BACE1 as potentially critical for the development of Alzheimer's disease, and our preliminary data suggest that CNBP may be one of these regulatory factors. Further, both BACE1 and the related peripheral enzyme BACE2 are both increased in Inclusion Body Myositis, an age-related, degenerative disease of the skeletal musculature considered by some to be a pathological cousin of Alzheimer's disease. Therefore, [unreadable]-secretase activity itself is linked to at least two age-related diseases, making the question of understanding its regulation potentially important for human health. This proposal is designed to determine the role of CNBP as a critical protein for RNA regulation and as a potential mediator of degenerative disease via the regulation of [unreadable]-secretase activity. This project focuses on three specific aims designed to (1) determine the mechanism through which CNBP regulates its targets, (2) determine what the normal function of CNBP is in the cell, and (3) determine what the role of CNBP is in disease. The project will advance through parallel stages of in vitro studies in biochemical and cell culture based model systems, up to studies in animal models of degenerative pathology. Understanding the mechanism through which CNBP operates will inform us about both basic processes in RNA regulation, and how they might go awry. These studies will lay the foundation for future advances into therapeutics to treat these diseases. [unreadable] [unreadable] [unreadable] [unreadable]

DESCRIPTION (provided by applicant): Metabolic disease is linked to obesity - especially mid-life obesity - a persistent problem in our society. An unexplained connection exists between type 2 diabetes mellitus and neurologic disorders, such as vascular dementia (VaD) and Alzheimer's disease (AD). The biological mechanism underlying this link is not known. As our population demographics shift towards an older average age, the potential confluence of obesity, diabetes and age related neurological dysfunction represents a potential public health disaster. The form of dementia afflicting individuals with a history of obesity and diabetes combines vascular pathology, strokes and AD related neuropathology, including amyloid deposits. It is not known if amyloid, either in the brain parenchyma or deposited within the cerebrovasculature, causes the vascular pathology or if some other aspect of diabetes and obesity is reponsible. This proposal grew out of the possibility that this connection may be related to leptin resistance. Leptin is an adipocyte-derived peptide hormone that regulates satiety and hunger via signaling through the leptin receptor, which is expressed throughout the brain. Obese individuals are resistant to leptin. In preliminary experiments, we discovered that leptin is a strong regulator of ?-secretase, particularly PS1 expression, at the mRNA level. To investigate this problem, we created a line of knock-in mice that become rapidly obese and diabetic, and develop amyloid pathology with increasing age. The most remarkable feature of this unique mouse model (db/AD) is that it develops a striking phenotype of cerebrovascular pathology, including aneurysms and strokes, and displays a profound cognitive impairment. This mouse line does not overexpress disease related proteins or use artificial promoter systems, making it an ideal system for the study of how aberant gene regulation in metabolic disease can influence brain pathology. We believe that we have created a mouse model of VaD, an understudied neurological disorder with limited treatment options. We propose to use our unique db/AD model to determine the mechanistic connection between aberrant leptin signaling and the development of aneurysms, strokes, and other aspects of cerebrovascular pathology. We will track the development of pathology longitudinally using state-of-the-art magnetic resonance imaging, combined with two approaches currently in human clinical trials to treat dementia. Since these approaches have been shown to have a good safety profile in human trials, the clinical implications for the outcome of this study are significant. The major strengths of this proposal are the use of a novel mouse model with unique features, and an innovative approach to determine the mechanism driving the development of pathology. This project has the potential to significantly advance our understanding of the major underlying causes of VaD, and has significant implications for the treatment and prevention of age-related cerebrovascular disease.

DESCRIPTION (provided by applicant): Alzheimer's disease (AD) is the most common age-related neurodegenerative disease, currently affecting more than five million people in the U.S.A. The vast majority of AD cases are sporadic, with no clear etiology. It is well-documented that lifestyle and overall health play a role in the development of the disease. A connection exists between risk for AD and Type 2 Diabetes Mellitus (T2DM), a metabolic disease that is linked to obesity. The mechanism that underlies this epidemiological connection is unknown. Recent neuropathology studies have shown that AD cases that have a history of T2DM and obesity have substantially more cerebrovascular pathology, but have either the same or lesser amounts of neuritic plaques and neurofibrillary tangles. It may be that changes in the cerebrovasculature alter the threshold for the development of AD, or that individuals with AD and a history of obesity represent cases of mixed AD and vascular dementia (VaD). This proposal considers the possibility that these elements are connected through defective leptin signaling, since individuals with a history of obesity and diabetes are leptin resistant. Leptin is an adipocyte-derived peptide hormone that regulates satiety and hunger via signaling through the leptin receptor (LepR), and these receptors are expressed throughout the brain. To explore the connection between leptin resistance, obesity and AD, we created a unique mouse line (db/AD) that combines features of these pathophysiologic states. These mice are resistant to leptin (the LepR is inactivated), become rapidly obese and diabetic, and develop AD-related neuropathology. These mice also develop a striking phenotype of cerebrovascular pathology, and display a profound cognitive impairment. In recent years, several studies have linked leptin with cognitive function. It has even been suggested that leptin treatment may be a viable therapeutic approach towards alleviating age-related cognitive decline. This project proposes to directly test the role of leptin in cognitive function in our novel model by selectively replacing he defective LepR in the brain with a functional version via adeno-associated virus, thereby restoring leptin sensitivity. Preliminary tests indicate that we are able to replace the receptor without affecting the hypothalamus, and the animals remain hyperphagic and retain their metabolic disease phenotype. We therefore believe that this approach will allow us to separate the effects of leptin on the development of neurologic dysfunction from the many complex problems associated with obesity and systemic metabolic disease. If leptin resistance is the cause of the cognitive dysfunction in the db/AD mice, then restoring leptin sensitivity will improve function. This project, utilizing a novel and innovative mouse model, has the potential to significantly advance our understanding of the association between obesity, diabetes, and dementia. If there is a clear connection between leptin, leptin resistance and neuropathology, patients may one day benefit from preventative of therapeutic strategies targeting leptin pathways.

DESCRIPTION (provided by applicant): Environmental exposure to lead (Pb) has been linked to risk of late-onset Alzheimer's disease (AD) and dementia. Although Pb has long been known as a neurotoxic agent in children, a recent and growing body of both toxicological and epidemiological research indicates that cumulative environmental Pb exposure is toxic to adults as well, and may be a significant contributor to age-related neurologic dysfunction. The biological mechanism underlying this link is not known. It has been proposed based on a limited number of animal studies that the linkage is through epigenetic changes in the methylation state of DNA, although evidence for this mechanism in human disease has been lacking. This proposal considers another possibility, that the linkage is through a combination of Pb-driven neuropathologic change, and that cerebrovascular pathology is the major contributor to this form of neurologic dysfunction. The amount of cerebrovascular pathology is a significant co-morbidity in all forms of age-related dementia. Most individuals with AD have some degree of comorbid cerebrovascular pathology, although individuals with a history of obesity and T2DM have substantial amounts of this pathology. To investigate this problem, we created a line of knock-in mice that co-develops amyloid pathology and cerebrovascular abnormalities with increasing age. The most remarkable feature of this novel mouse model (db/AD) is that it develops a striking phenotype of cerebrovascular pathology, including aneurysms and strokes, and also displays a profound cognitive impairment. We believe that we have created an innovative model of AD with significant cerebrovascular disease, an understudied variant with limited treatment options. The unique db/AD mouse does not overexpress disease related proteins or use artificial promoter systems, making it an ideal system for the study of how aberrant gene regulation in disease can influence brain pathology. This presents an unparalleled opportunity to study, and potentially dissociate the role of Pb exposure at different times in an animal's lifespa and gauge the ultimate impact on neuropathology. This proposal thus seeks to answer three key questions relating to Pb exposure and neurologic disease that occurs later in life. First, is early life Pb exposure more damaging than exposure as an adult? Second, can Pb cause late-life cognitive dysfunction by increasing cerebrovascular pathology, such as strokes, through increased hypertension, a well-known outcome of Pb exposure? Finally, how much does Pb exposure affect the development of AD-related pathology by affecting the expression of AD related genes? We believe that our mouse model is uniquely suited to answering these questions. A major innovative feature of this proposal is the use of a novel mouse model with unique features, exhibiting significant age associated AD-related amyloid deposition, cerebrovascular pathology, and cognitive dysfunction. This project not only has clear implications for the prevention and treatment of age-related cerebrovascular disease, but also has the potential to advance our understanding of the major underlying causes of cerebrovascular disease as comorbidity in the AD brain.

Age-related neurologic disease is a significant and growing burden on our society. Although the largest share of research effort has typically been devoted to the common neurodegenerative illnesses (such as Alzheimer's disease, or AD), the reality is that nearly all cases of neurodegenerative disease possess elements of mixed pathology. Individuals diagnosed with AD frequently harbor neuropathologic hallmarks common in other diseases. For example, tau pathology is also found in some forms of frontotemporal dementia, ALS, and other forms of neurodegenerative disease, and is also believed to be a key form of neuropathology that develops following traumatic brain injury. Cerebrovascular disease (CVD) is abundant in individuals with a history of obesity (and type 2 diabetes, or T2D), which have a well known elevated risk of dementia. In general, it is actually quite rare to identify AD cases lacking elements of co-morbid cerebrovascular pathology. It is unclear as to whether these elements of pathology contribute to dementia in an additive or synergistic manner. In recent studies in our lab, we have observed an intriguing relationship between various aspects of neuropathology that could potentially connect to AD and CVD. We have identified a membrane ion exchanger, NHE1 (SLC9A1), as potentially involved in both pathologic processes. NHE1 has been shown to be involved with neuronal injury, and tau pathology as our preliminary data indicates. This project seeks to determine whether the function of this exchanger is important for either, or both, of these pathologies. This project combines both genetic and pharmacologic approaches to explore this exciting new target that has not previously been examined as a major player in age-related neurodegenerative disease. In specific aim 1 (SA1), we will investigate the mechanism of the membrane ion exchanger NHE1 in a unique mouse model combining AD- and CVD-related pathology, using a highly specific pharmacologic agent. In SA2, we will investigate how this membrane ion exchanger drives the formation of tau pathology, by over expressing tau on a background of NHE1 genetic reduction. Efficacy will be determined using a range of immunohistochemical, molecular, and biochemical markers of pathology, as well as gauging changes in cognitive function. We hypothesize that NHE1 will be responsible for multiple aspects of neurodegenerative disease pathology, and that interfering with its activity will ameliorate these problems, and will alleviate cognitive dysfunction. This is a highly innovative hypothesis that, to our knowledge, has not been previously explored. Another strength of this proposal is the use of a novel mouse model with unique features. This project has the capacity to significantly improve our understanding of co-morbid neuropathologies, and could have significant implications for the treatment and prevention of age-related neurodegenerative disease.

7. PROJECT SUMMARY/ABSTRACT This is an application for a T32 training grant titled ?Training in Translational Research in Alzheimer's Disease and Related Dementias (TRIAD)? to support 4 predoctoral and 4 postdoctoral fellows. The overall goal of the proposed program is to provide cross-disciplinary training from bench to bedside, to produce a new translational workforce that is critically needed to develop and advance effective treatments for Alzheimer's disease and related dementias. Trainees will receive rigorous and innovative translational research education and training through a 3-cluster multidisciplinary and integrative mentoring program that spans the discovery continuum from molecular/biochemical methods, preclinical translational approaches and clinical research. The thematic focus is on risk factors for Alzheimer's disease and related dementias; these risk factors include cerebrovascular disease, neuroinflammation, Down syndrome, and traumatic brain injury. The training program has 18 faculty mentors with established and extensive interactions and collaborations, and successful track records in training. The faculty mentors encompass diverse areas of expertise from cell & molecular biology, genetics, and data science, through preclinical model systems, therapeutic development, neuroimaging, neuropathology, clinical trials, and longitudinal clinical studies. The majority (13) of these mentors are housed within the University of Kentucky Sanders Brown Center on Aging, which includes a long-standing NIA-funded Alzheimer's Disease Center. All faculty with primary appointments in a Center also have academic appointments in other basic science or clinical departments at the University of Kentucky. Predoctoral trainees will be recruited from our Integrated Biomedical Sciences graduate program and from the MD/PhD program in the College of Medicine, and postdoctoral trainees through targeted advertisement and recommendations from colleagues and other Alzheimer's Disease Centers. In addition to their individualized research program and career development plan, each group of trainees will participate in a series of educational experiences that include a dedicated AD101 course in Alzheimer's disease and related dementias, seminars describing research and clinic ethics, monthly research and career development seminars, journal clubs, Training the Trainers workshops, and grant writing workshops. Each year trainees will be expected to present their research at the annual Markesbery Symposium on Aging and Dementia. Postdoctoral trainees will be provided with additional training opportunities, including: 1) a short off-site externship at another laboratory that conducts Alzheimer's disease research and 2) presentations at the annual College of Medicine Postdoctoral Poster Session. Quantitative and qualitative metrics for evaluation of trainees and the training program will assure a high-quality and effective training experience. The outcomes of our program are intended to nurture highly innovative, well-trained scholars who are dedicated and passionate about finding new approaches to prevent or slow Alzheimer's disease and related dementias.

Chronic sleep disruption, resulting from work schedules, noise exposure, family obligations, sleep disorders, or lifestyle choices, is a pervasive feature of contemporary life. Sleep problems affect up to 40% of AD patients, may precede cognitive impairments by more than a decade, and worsen as the disease progresses. As well as affecting mood and well-being, sleep disruption may drive the development of AD neuropathology for instance, by reducing clearance of amyloid-? (A?) and by promoting a neurotoxic proinflammatory state involving astrocytes and microglia. Sleep disruption can include reduced total sleep (sleep restriction [SR]), loss of deep sleep (also known as slow-wave sleep [SWS], marked by large amplitude, low frequency electrical activity), and fragmentation of sleep (SF) into shorter bouts. Fragmentation of the daily sleep-wake rhythm is associated with greater risk of incident AD and earlier cognitive decline in older humans. In spite of these correlative studies, whether or how chronic SF impacts the progression of AD has not been experimentally investigated. SF may be a better model of the sleep disruption associated with AD than the traditional approach of SR. Our studies of AD mouse models show that spontaneously occurring SF is associated with more severe A? accumulation and that experimentally-induced SF leads to A? accumulation and neuroinflammation. Besides SF, loss of SWS may exacerbate AD, and improving SWS may be beneficial in mild cognitive impairment (MCI) or even in AD. Since sleep disruption adversely affects the development of AD-related neuropathology, it is surprising that sleep enhancement (SE) strategies to consolidate sleep and increase SWS have not been adequately explored to slow or reverse these effects. Our overall working hypothesis is that a change in the quality of sleep, especially sleep fragmentation and loss of SWS, is more important than the quantity of sleep. Further, we hypothesize that the mechanism underlying these effects is primarily neuroinflammation, at least in part mediated by A? peptide deposition. We will use a unique, well-characterized mouse model, that exhibits AD-related A? pathology, neuroinflammation, and cognitive deficits. This project has three specific aims: (1) that SF will accelerate (and SE decelerate) AD progression; (2) that increases in A? accumulation mediates SF-induced neuroinflammation, neuropathology, and cognitive decline; and (3) that increases in neuroinflammation mediate SF-induced neuropathology and cognitive decline. We will use multiple novel approaches, including thermoneutral temperature manipulation, and a unique anti-inflammatory compound that has recently entered early stage clinical trials. Thus, these studies will elucidate the underlying mechanisms by which sleep disruption is linked to AD and will lay the groundwork for new therapeutic strategies.