Andrew P. Lieberman, MD PhD - US grants

DESCRIPTION (provided by applicant): Nine neurodegenerative diseases are caused by expansions of polyglutamine tracts. Among these disorders is Kennedy's disease, a motor neuronopathy caused by a mutation in the androgen receptor gene. It is not well understood how this mutation causes neuronal degeneration. Lack of Such knowledge is an important problem because it hinders the development of treatment strategies. The laboratory's long-range goal is to understand the pathogenetic mechanisms by which expansions of polyglutamine tracts cause neuronal dysfunction and death. The objective of this application is to determine the molecular basis for the alteration in function that the receptor undergoes as a result of polyglutamine expansion. The central hypothesis is that the mutation alters normal androgen receptor function by affecting acetylation and phosphorylation cascades, and by conferring a toxic gain-of-function that leads to oxidative injury. The rationale for these studies is that understanding the molecular pathways altered by the mutation will yield insights into mechanisms of neurodegeneration, thereby suggesting novel therapeutic targets. The central hypothesis will be tested by pursuing the following two specific aims: 1) Identify the molecular pathways that are dysregulated by the expanded polyglutamine androgen receptor, 2) Establish which of these dysregulated pathways is causally related to cell dysfunction and death. The experimental approach will be to characterize altered androgen receptor function, to modify the activity of acetylation and phosphorylation cascades, and to evaluate the role of iron mediated oxidative injury in cell dysfunction and death. It is my expectation that this award will provide significant protected time to foster my career development as a neuroscientist, and that this scientific approach will yield important data that will serve as the basis for an R01 grant application in years 4 and 5 of this award. We also expect to identify important pathways dysregulated as a result of expanded polyglutamine tracts, and to determine which of these dysregulated pathways contributes to cell dysfunction and death. Such outcomes will have general impact because this new knowledge will suggest novel targets for the treatment of this and related diseases.

[unreadable] DESCRIPTION (provided by applicant): Chronic neurodegenerative diseases are a significant health problem for the elderly, causing profound morbidity and mortality, and generating staggering health care related expenses. Mechanisms that regulate the late onset and severity of phenotype of these disorders are poorly understood. To address this question we have chosen to study Kennedy's disease, a degenerative disorder of motor neurons caused by a CAG/glutamine expansion in the androgen receptor. The laboratory's long-range goal is to understand the mechanisms by which expansions of glutamine tracts cause neuronal dysfunction and death. The objective of this application is to identify pathways that modify toxicity of the mutant androgen receptor, and may therefore impact age of onset and severity of phenotype. Our central hypothesis is that toxicity is modified both by the unfolded protein response (UPR) and by decreased receptor acetylation. The rationale of these studies is that understanding pathways that modify toxicity of the mutant protein will yield insights into the mechanisms of neurodegeneration, thereby suggesting novel therapeutic targets. The central hypothesis will be tested by pursuing the following specific aims: 1) Establish that the UPR is a modifier of polyglutamine toxicity; 2) Determine the extent to which decreased acetylation of the expanded glutamine androgen receptor contributes to misfolding and aggregation; and 3) Determine the extent to which 113 CAG repeats targeted to the mouse androgen receptor gene causes selective dysfunction or degeneration of motor neurons, UPR activation and decreased receptor acetylation in vivo. It is my expectation that receipt of the Beeson Award will enable me to gain critical skills and experience in aging research. At the completion of these studies, we expect to establish that misfolding, aggregation and toxicity of the mutant androgen receptor is modified by the UPR and by pathways that mediate ligand-dependent androgen receptor acetylation. We also expect that our knock-in mouse model will reproduce important aspects of this disorder, including selective motor neuron dysfunction and degeneration. Such outcomes will have general impact by furthering our understanding of disease pathogenesis and by yielding a mouse model that faithfully recapitulates the human disease. [unreadable] [unreadable] [unreadable] [unreadable]

The MADRC Neuropathology Core (NPC) provides systematic and standardized collection, pathologic characterization, storage, and distribution of human post-mortem material appropriate for the research requirements of investigators affiliated with the MADRC. The overall goal of the NPC is to facilitate research by MADRC and other investigators probling the dementias by providing access to human post-mortem material that is both clinically and pathologically well characterized using state-of-art techniques. NPC services support the needs of local researchers, as well as cooperative research across ADCs. Working in tandem with the Clinical Core, the NPC assists subjects and families of subjects followed by the Clinical Core with pre-arrangements for brain donatioa The NPC is responsible for rapid collection of post-mortem material, standardized preparation of fixed and frozen tissues for subsequent pathologic characterization and distribution to investigators, and storage of fixed and frozen material in brain bank. The NPC performs extensive pathologic characterization of harvested post-mortem material, distributes material to investigators, transmits data to the Data Management and Statistics Core, prepares post-mortem human material for investigators, assists investigators with use of human post-mortem material, and assists investigators with special staining methods. In a new initiative, the NPC is also assisting investigators with histologic evaluation of animal models of dementias. The NPC interacts extensively with other components of the MADRC. The NPC depends on the Administrative Core for crucial adminstrative oversight and support. The NPC works closely with the Clinical Core, Education Core, and Data Management and Statistics Core for subject recruitment, subject characterization, and data archiving. The NPC serves all projects, providing key histology services for Project 1, and crucial pathologic information for Projects 2 and 3. The NPC is a major resource for study of dementia on our campus and has been used by a number of investigators at other centers to further investigations of dementias and other neurodegenerations. The NPC will adhere to standard neuropathology procedures being developed by a neuropathology working group once those procedures have been issued.

[unreadable] DESCRIPTION (provided by applicant): The sphingolipid storage diseases are a group of ~40 genetically distinct disorders that occur with a collective frequency of 1 in 8000 live births and are often associated with devastating neurodegeneration. Among these diseases is Niemann-Pick C, an autosomal recessive disorder of lipid trafficking that produces cognitive impairment, ataxia and death, often in childhood. Most cases of this disease are caused by loss of function mutations in the Npc1 gene. Data from several laboratories demonstrate that the NPC1 protein functions in late endosome lipid sorting and vesicular trafficking, and may act as a transmembrane efflux pump. Despite these significant advances in our understanding of NPC1 function, it remains unknown how mutations in this gene cause severe neuropathology. The objective of this proposal is to develop and characterize a conditional null mutant of the mouse Npc1 gene. We expect this model will be an important resource for the neuroscience community, enabling studies characterizing the mechanism of neurodegeneration. Our central hypothesis is that conditional deletion of Npc1 exon 9 will produce a null phenotype. This hypothesis is based on the observation that deletion of exon 9 will cause a frame-shift mutation, and is expected to yield a null phenotype similar to that produced by an insertional mutation in exon 9 present in the existing constitutive Npc1 null mutant. We will test our hypothesis by characterizing mice in which loxP sites flank exon 9 of the mouse Npc1 gene. These animals will be crossed with an established mouse line expressing Cre recombinase as a transgene in oocytes, thereby causing global deletion of the floxed Npc1 allele. We are particularly well prepared to undertake the proposed research because we recently established two independent mouse lines in which one copy of the Npc1 gene has loxP sites flanking exon 9. Biochemical, histologic and behavioral approaches will be used to characterize Npc1 conditional null mice, and these animals will be made available to the neuroscience community. The relevance of the proposed studies to public health is that this animal model will be a unique resource to study the neuropathology of Niemann-Pick C and to characterize the normal function of the NPC1 protein in vivo. [unreadable] [unreadable] [unreadable]

DESCRIPTION (provided by applicant): Neurodegenerative disorders associated with protein aggregation include nine untreatable diseases caused by CAG/glutamine tract (polyQ) expansions. One of these disorders, spinobulbar muscular atrophy (SBMA), is characterized by degeneration of lower motor neurons and is caused by a mutation in the androgen receptor (AR) gene. The mutant protein undergoes hormone-dependent nuclear translocation, unfolding and oligomerization, steps that are critical to toxicity and to the development of progressive proximal limb and bulbar muscle weakness in men. Although the disease causing mutation was identified two decades ago, treatments available to SBMA patients remain largely supportive. Furthermore, the cellular pathways that degrade the mutant protein remain incompletely defined, and this lack of knowledge hinders the development of disease-modifying therapies. The objective of this application is to define the role of the Hsp90-based chaperone machinery in the protein quality control decisions that govern polyQ AR degradation. Our central hypothesis is that Hsp70 and Hsp90 have essentially opposing roles in the triage of the polyQ AR, in that Hsp70 promotes substrate ubiquitination whereas Hsp90 inhibits ubiquitination. This hypothesis springs from our preliminary data showing that association with Hsp90 stabilizes the polyQ AR, while unfolding of the mutant protein leads to ubiquitination by Hsp70-dependent E3 ligases. Here we will use genetic and pharmacological tools to define the consequences of allosterically activating Hsp70-dependent ubiquitination. Additionally, as our data point to contributions from both skeletal muscle and motor neurons to the disease phenotype, we will use genetic approaches to determine the extent to which toxicity at each site must be targeted to achieve beneficial therapeutic effects. The rationale of the proposed work is that establishing the mechanisms that regulate polyQ AR degradation will identify targets that can be exploited by the development of new therapies. Genetic and biochemical approaches will be used to determine the extent to which allosteric activators of Hsp70 promote clearance of the polyQ AR (Aim 1), to identify critical sties of polyQ AR toxicity in SBMA mice (Aim 2), and to establish the effects of novel, small molecule allosteric activators of Hsp70 in SBMA mice (Aim 3). These studies are expected to have a significant positive impact by defining pathways that limit SBMA toxicity while providing proof-of-concept data supporting a new therapeutic approach. As Hsp70 also regulates quality control decisions governing the turnover of other mutant proteins that cause neurodegeneration, we expect that the approaches defined here will inform therapeutic strategies that will be broadly applicable.

Abstract The lysosomal storage diseases are a group of ~50 genetically distinct disorders that result from inherited deficiencies of lysosomal hydrolytic activities or lipid transport. Among this group is Niemann-Pick type C disease, an autosomal recessive disorder for which there is no effective treatment. Niemann-Pick C patients exhibit a clinically heterogeneous phenotype characterized by severe, progressive neurodegeneration that is usually fatal in childhood. Most cases are caused by loss-of-function mutations in the NPC1 gene, resulting in disrupted intracellular trafficking of cholesterol and glycosphingolipids. Although disease-causing mutations were identified almost two decades ago, it remains unknown how the resulting defects of lipid trafficking lead to the severe neurological disease that is characteristic of this disorder. This lack of knowledge hinders the identification of specific targets for developing disease-modifying therapies. The objective of this application is to identify mechanisms leading to neurodegeneration and to define cellular pathways where interventions could result in effective treatments. Our central hypothesis is that the disruption of cellular quality control pathways caused by Npc1 deficiency underlies neurodegeneration. This hypothesis springs from our analysis of patient fibroblasts and mice with a conditional null allele of the Npc1 gene generated in our lab. These studies and results in the literature revealed impairments of cellular proteostasis, including abnormalities in autophagy, that result in the accumulation of ubiquitinated proteins and fragmented mitochondria, particularly within neurons and in CNS regions of selective vulnerability. Our studies also build on our preliminary data demonstrating unexpected contributions of oligodendrocytes to neuronal degeneration in the mutant brain, suggesting impaired support of neurons by glia. These findings are complemented by recent work characterizing a new mouse model of disease that expresses Npc1 I1061T, the most prevalent disease-causing mutation. Behavioral, histological, biochemical, cell biological and genetic approaches will be used to characterize alterations in autophagy in Npc1 deficient neurons (Aim 1), establish the contribution of altered energy metabolism to axonal pathology and neuron loss (Aim 2), and identify critical components of the machinery that regulates degradation of Npc1 I1061T (Aim 3). These studies are expected to have an important positive impact by defining mechanisms through which Npc1 deficiency leads to progressive neurodegeneration and by identifying potential therapeutic targets. Furthermore, we expect that shared mechanisms mediate toxicity in several lipid storage diseases, suggesting that advances here will impact our understanding and treatment approaches to genetically distinct lysosomal storage disorders.

DESCRIPTION (provided by applicant): Niemann-Pick C disease is an autosomal recessive lysosomal storage disorder for which there is no effective treatment. Patients with this disease exhibit a clinically heterogeneous phenotype characterized by severe, progressive neurological impairment that is usually fatal in childhood. Most cases are caused by loss-of- function mutations in the NPC1 gene, resulting in impaired intracellular trafficking of cholesterol and glycosphingolipids. Disease causing mutations were first identified over a decade ago and the role of NPC1 in cholesterol trafficking is emerging from elegant cell biology studies. Nonetheless, strategies to promote normal intracellular lipid transport have been elusive, and to date, the severe, progressive neurological impairment characteristic of Niemann-Pick C disease remains untreatable. The objective of this application is to identify cellular pathways where interventions could result in effective treatments. Our central hypothesis is that modulating the protein homeostasis network will enable proper folding and trafficking of some NPC1 mutants to produce a functional recovery. This hypothesis springs from our analysis of patient fibroblasts in which the NPC1 gene carries disease-causing missense mutations. Our preliminary studies show that modulating ER calcium levels by treatment with ryanodine receptor antagonists produces a striking recovery of functional NPC1. Further, we demonstrate that patient fibroblasts can be reprogrammed into induced pluripotent stem cells (iPSCs) to obtain patient-specific neurons (hereafter referred to as induced neurons), thereby generating a model system to test biologically active small molecules on the target cells that are critical to neurodegeneration. Cell biological, pharmacological, biochemical and genetic approaches will be used to determine the extent to which modulating ER calcium levels promotes the recovery of functional NPC1 in patient fibroblasts (Aim 1), and to establish that targeting ER protein homeostasis pathways ameliorates the effects of NPC1 deficiency in patient-specific induced neurons (Aim 2). These studies are expected to have an important positive impact by providing proof-of-concept evidence in support of a novel treatment strategy for patients with missense mutations in the NPC1 gene, and demonstrate the utility of iPSC-derived neurons as a model system for therapeutic development.

DESCRIPTION (provided by applicant): The neurodegenerative disorders characterized by protein aggregation include nine untreatable diseases caused by CAG/glutamine tract expansions. One of these polyglutamine (polyQ) diseases, spinobulbar muscular atrophy (SBMA), is a degenerative disorder of lower motor neurons caused by a mutation in the androgen receptor (AR) gene. The mutant protein undergoes hormone-dependent nuclear translocation, unfolding and oligomerization, steps that are critical to toxicity and to the development of progressive proximal limb and bulbar muscle weakness in men. Although the disease causing mutation was identified about two decades ago, mechanisms that are central to the pathogenesis of SBMA remain poorly understood and available therapies are largely supportive. Recent studies demonstrate that post-translational modifications of the AR triggered by ligand influence toxicity. Our laboratory has shown that conjugation of the polyQ AR by SUMO (small ubiquitin-like modifier) impairs ligand-dependent oligomerization of the mutant protein. However, the extent to which this modification alters the disease phenotype in vivo is currently unknown. The objective of this application is to determine the extent to which SUMOylation of the polyQ AR affects SBMA pathogenesis. Our central hypothesis is that SUMOylation of the polyQ AR diminishes neuromuscular toxicity in SBMA. This hypothesis springs from our own preliminary data demonstrating that SUMOylation decreases the levels of soluble AR oligomers and aggregates in cellular models of SBMA. Here we will use gene targeting to generate knock-in mice expressing a SUMO resistant polyQ AR to determine the extent to which this pathway affects disease pathogenesis. The rationale of the proposed work is that defining the role of the SUMOylation pathway in disease will yield insights into pathogenic mechanisms and accelerate the discovery of targets for disease- modifying therapies. Genetic and biochemical approaches will be combined with characterization of mouse behavioral and neuropathological changes to establish the effects of polyQ AR SUMOylation in a knock-in mouse model of SBMA. These studies are expected to have a significant positive impact by defining the role of SUMOylation in SBMA and thereby identifying potential therapeutic targets. As several neurodegenerative disease-causing proteins are targeted by SUMO, we anticipate that our findings will also serve as a paradigm for understanding the effects of SUMOylation on the phenotype of these related disorders.

? DESCRIPTION (provided by applicant): Spinobulbar muscular atrophy (SBMA) is an untreatable degenerative disorder of motor neurons and skeletal muscle caused by a CAG/glutamine tract expansion in the androgen receptor (AR) gene. The polyglutamine AR (polyQ AR) undergoes hormone-dependent nuclear translocation and unfolding, steps that are essential to toxicity and to the development of progressive muscle weakness in men. Although the disease causing mutation was identified over two decades ago, mechanisms central to SBMA pathogenesis remain poorly understood. As an alternative to focusing on downstream pathways disrupted by the polyQ AR, here we propose to test a novel therapeutic strategy using antisense oligonucleotides (ASO) to knock-down expression of the mutant gene. The objective of this application is to complete preclinical studies in a knock-in mouse model of SBMA to establish the safety and efficacy of peripherally delivered ASO. Our central hypothesis is that toxicity of the polyQ AR in the periphery is an important contributor to disease pathogenesis and an attractive therapeutic target. This hypothesis, which is distinct from efforts in the field aime at targeting toxic effects within the CNS, is based on our preliminary findings using ASO that suppress AR gene expression in the periphery but not CNS following subcutaneous administration to mice. These ASO were developed in an on-going academic-industrial partnership with Isis Pharmaceuticals, a company that specializes in ASO production. Working with Isis, we found that peripheral gene suppression rescues deficits in skeletal muscle mass, muscle fiber size, and lifespan in a knock-in mouse model of SBMA developed by our laboratory. These data point to an important role of peripheral polyQ AR in disease and suggest a novel route to therapy. Using phenotypic, histological, genetic and biochemical analyses we will establish the optimal ASO delivery route, dose and time course to ameliorate disease in SBMA knock-in mice (Aim 1), and determine the long-term effects of peripheral ASO therapy (Aim 2). If successful, these studies will provide essential efficacy data in a preclinical mouse model, which will then proceed to lead optimization and IND-enabling studies in the next stage of this project, as a prelude to a clinical trial in human SBMA patients.

CORE D: NEUROPATHOLOGY CORE Abstract The Neuropathology Core (NC) will capitalize on the longstanding, actively recruiting University of Michigan (UM) Brain Bank, several decades of patient clinical characterization by Alzheimer?s Disease (AD) researchers at UM, and the substantial resources of the UM Protein Folding Diseases (PFD) Initiative to advance the Michigan ADCC's central theme: to identify, understand and modulate the non-? amyloid factors contributing to brain dysfunction and neurodegeneration in AD and related dementias. Studies of these diseases will be facilitated by integrating NC, Clinical Core and Data Management and Statistical Core activities to collect and properly characterize post-mortem material and associated antemortem biospecimens from ADCC participants with no neurologic disease and those with varied stages of AD and related neurodegenerative disorders. The NC will follow the 2014 NIA Biospecimen Task Force best practice guidelines for the evaluation, banking and distribution of brain tissue, DNA, and plasma. These essential resources, along with access to NC expertise and technologies, will be available for investigators at UM, our regional partner institutions Michigan State University and Wayne State University, and AD Centers across the country. Furthermore, by leveraging resources of the PFD Initiative and the diverse expertise of NC investigators in neurodegenerative proteinopathies, the NC will be uniquely positioned to facilitate innovative research into the pathologic mechanisms underlying protein dysfunction in dementia. To accomplish these goals and tasks, the NC will pursue the following aims: 1) grow and maintain a brain bank with well-characterized frozen and fixed tissue, 2) provide accurate, detailed, and standardized neuropathological evaluation, 3) distribute banked tissue and biospecimens, 4) contribute neuropathologic data to NACC, 5) support researchers studying AD and related dementias, and 6) actively educate trainees about the neuropathologic assessment of dementia. Through these aims, the NC will greatly facilitate basic and translational studies of the dementias, expand the research community actively engaged in this work, and help build new collaborations for future discoveries.

Abstract The Training Program in Translational Research offers an interdisciplinary program of study and research that prepares graduate students for successful careers at the interface between basic biomedical science and clinical medicine. This program is designed for predoctoral Ph.D. students and aims to address the widely-recognized shortage of rigorously trained scientists who can successfully work together with medical professionals to bridge the gap between basic science and clinical practice. The program is centered in the University of Michigan School of Medicine, with participating faculty members affiliated with a twelve departments (7 clinical and 5 basic science). Additional faculty mentors have their primary appointments in the School of Pharmacy and School of Dentistry. These faculty members were selected because of their commitment to graduate education and expertise in translational research. Based on the long- standing interests of these research preceptors, scientific projects will be focused on areas of excellence in cancer biology, development, epigenetics, aging, immunology, neuroscience and experimental therapeutics & biomarkers. This broad spectrum of research activities provides a wide-range of exciting opportunities for students to work on translational research projects. The educational experience will be enhanced and supported by integration of dual-mentors with corresponding expertise in the basic and clinical aspects of the research project. The curriculum includes innovative coursework in translational research and student participation in clinical rotations, interdisciplinary conferences, tumor boards, and grand rounds. These activities will be complemented by a T32-based seminar series and journal club and an annual retreat. The program will be administered by co-directors who are experienced in, and passionate about, graduate student education and translational research. The program will benefit from the advice and oversight of a steering committee and an external advisory board, both of which will assist the co-directors in running and evaluating the program. Graduates from this program will have the skills and knowledge to undertake an independent career that features translational multidisciplinary research. In summary, the strengths of this training program are the direct link between the basic and clinical sciences, the collegial and successful preceptors, the experienced program leadership, and the rich academic research and clinical environment at the University of Michigan.

Niemann-Pick disease type C is an invariably fatal autosomal recessive lipid storage disease. Patients develop a clinically heterogeneous phenotype that includes progressive neurodegeneration and early death. Disease is commonly caused by loss-of-function mutations in the NPC1 gene (95% of cases), encoding a multipass transmembrane glycoprotein required for exporting unesterified cholesterol from late endosomes and lysosomes. The most common disease-causing mutation (~20% of cases) is an isoleucine to threonine substitution at position 1061 (I1061T). I1061T NPC1 misfolds in the endoplasmic reticulum (ER) and is rapidly degraded by the proteasome and ER-autophagy. Importantly, transient over-expression of I1061T in vitro or treatment of I1061T NPC1 patient fibroblasts with ryanodine receptor antagonists drives trafficking of mutant NPC1 to the lysosome where it is still functional. These observations have spurred interest in developing proteostasis modulators to treat disease. These efforts have relied on gene targeted mice in which the I1061T mutation was inserted into the mouse Npc1 gene to test therapeutic strategies. However, our preliminary data indicate that there are marked differences in how the human and mouse I1061T NPC1 proteins are handled by the cellular quality control machinery. These previously unappreciated differences underscore the critical need to develop a new model system that reliably reproduces human I1061T NPC1 proteostasis. The rationale for this project is that developing humanized I1061T NPC1 mice will enable in vivo testing of proteostatic therapeutics for Niemann-Pick C. To attain the overall objective of this application, we will use genetic and biochemical approaches to pursue the following specific aim: Develop and characterize humanized I1061T NPC1 mice. We expect that targeting the mouse Npc1 gene to express human I1061T NPC1 protein will generate a robust model of disease, in which misfolding and trafficking of the mutant NPC1 protein closely mimics the behavior of human I1061T NPC1 as occurs in Niemann-Pick C patients.

ABSTRACT Spinal and bulbar muscular atrophy (SBMA) is a degenerative disorder of the neuromuscular system caused by a CAG/glutamine tract expansion in the androgen receptor (AR) gene. The polyglutamine AR (polyQ AR) undergoes hormone-dependent nuclear translocation and unfolding, steps that are essential to toxicity and to the development of progressive muscle weakness in men. Although it was long considered that lower motor neurons are the primary targets of degeneration in SBMA, recent studies from our laboratory and others have established the importance of peripheral polyQ AR expression in disease. This work highlights a central contribution of polyQ AR expression in skeletal muscle to weakness and atrophy. Based on these findings, we have developed an innovative model of SBMA pathogenesis in which degeneration of the neuromuscular system begins with toxic effects in skeletal muscle and progresses over time to involve spinal motor neuron degeneration. Here, we propose to test this model of disease pathogenesis in gene targeted mice (AR113Q mice) expressing polyQ AR at endogenous levels and in appropriate cell types. The objective of this application is to experimentally test our novel model of disease pathogenesis. This model forms our central hypothesis and is supported by a rigorous foundation of published and preliminary data. Here, we will use two complementary approaches to test our central hypothesis: First, we will use antisense oligonucleotides (ASOs) to knock-down expression of polyQ AR selectively in peripheral tissues or CNS of symptomatic AR113Q mice and determine effects on late onset motor neuron degeneration. Second, we will restore function of a critical transcriptional regulator in skeletal muscle that contributes to SBMA skeletal muscle atrophy and then determine the extent to which this influences the AR113Q phenotype, including motor neuron degeneration. These aims will be complemented by studies designed to leverage the endogenous cellular machinery that regulates polyQ AR degradation in order to eliminate proteotoxicity in disease relevant human cells. We will use biochemical, genetic, and histological assays to establish beneficial effects of AR targeted ASOs administered to symptomatic AR113Q mice (Aim 1), determine the extent to which increased MEF2 function rescues the AR113Q phenotype (Aim 2), and establish effects of targeting the Hsp90/Hsp70 chaperone machinery in SBMA models (Aim 3). These studies are expected to experimentally test our proposed model of disease pathogenesis and provide a strong foundation for developing targeted therapies to treat SBMA patients.

ABSTRACT Niemann-Pick disease type C (NPC) is an invariably fatal autosomal recessive lipid storage disorder affecting all ages. Patients develop a clinically heterogeneous phenotype that includes severe, progressive neurodegeneration, hepatomegaly, and early death. NPC is commonly caused by loss-of-function mutations in the NPC1 gene (95% of cases), encoding a multipass transmembrane glycoprotein required for exporting unesterified cholesterol from late endosomes and lysosomes. Despite our emerging understanding of the role of NPC1 in intracellular cholesterol trafficking, a diagnosis of NPC remains particularly bleak. There are currently no FDA-approved disease modifying therapies and patients most often die in childhood, reflecting both gaps in our current knowledge of disease pathogenesis and a significant unmet medical need. Our long-term goal is to contribute toward the development of disease-modifying therapies for NPC patients. The next step in attaining this goal is to pursue the overall objective of this application: to define critical targets in CNS disease pathogenesis that can be exploited by drug development efforts. Our central hypothesis is that NPC1 deficiency causes toxicity in both neurons and oligodendrocytes that underlies NPC neuropathology. Moreover, we hypothesize that this toxicity can be rescued by novel therapeutic strategies aimed at reducing the intracellular lipid storage that is characteristic of the disease or by correcting the misfolding of mutant NPC1 protein. These notions are based upon robust preliminary data supporting our model of NPC pathogenesis and the use of innovative therapeutic approaches to rescue disease phenotypes. We will use genetic, biochemical, histological, and phenotypic analyses to: establish the extent to which neuronal lipid storage and toxicity are rescued by optimized synthetic HDL nanoparticles (Aim 1); determine the role of oligodendrocyte lineage cells in NPC neuropathology (Aim 2); and establish effects of proteostasis regulators in humanized NPC1 model systems (Aim 3). These studies are expected to establish that targeting intracellular lipid storage using optimized sHDLs and modulating mutant NPC1 proteostasis will ameliorate disease phenotypes. Moreover, we expect to demonstrate an important, yet under-studied role for oligodendrocyte lineage cells in NPC neuropathology.