Franz Hefti - US grants

This proposal will test the hypothesis that nerve growth factor (NGF) is a neurotrophic factor for cholinergic neurons of the forebrain. In an attempt to characterize the actions of NGF on these cells, the P.I. has found that NGF promotes survival of cholinergic septo-hippocampal neurons of adult rats after destruction of their processes by transection of the fimbria. Furthermore, preliminary findings indicate that NGF stimulates survival and fiber growth of cholinergic neurons in culture. Planned experiments aim at thoroughly investigating these previously undiscovered effects of NGF and at characterizing the conditions necessary for them to occur. Studies will be carried out in vivo, on young and aged adult rats with lesions of the septo- hippocampal cholinergic pathway, and in vitro, on cultured cholinergic neurons from fetal rat brains. In addition, an immunohistochemical method has recently been developed by the P.I. for the visualization of NGF receptors in the rat brain. Findings obtained so far, and which were limited to telen- and diencephalon, indicate that NGF receptors are selectively located on forebrain cholinergic neurons. Planned experiments aim at substantiating this selective localization of NGF receptors and at differentiating between a localization in cholinergic cells and afferent terminals. Furthermore, it is proposed to map NGF receptors in the entire rat brain. The proposed studies will help to establish whether NGF is a neurotrophic factor for forebrain cholinergic neurons. Furthermore, the studies have implications for Alzheimer's disease which is associated with a selective loss of cholinergic neurons in the basal forebrain. The proposed studies concern survival and function of a neuronal population selectively affected in Alzheimer's disease and the effects of a neurotrophic factor for these neurons.

Dissociated cultures of neurons obtained from various rat brain regions will be employed to study the effects of the newly discovered neurotrophic factors, brain derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3) and leukemia inhibitory factor (LIF) on cholinergic and dopaminergic biochemical differentiation. Observed effects will be compared to those of nerve growth factor (NGF). In vivo experiments in which the neurotrophic factors are injected into specific brain areas will be done to determine whether the effects observed in culture are biologically relevant.

Understanding underlying alteration in brain function is a prerequisite for the amelioration of age-related behavioral impairments and age-related neurodegenerative diseases which represent major problems for advanced societies. Research during the past decade has revealed that age-related morphological and physiological changes in neuron function can not be generalized to all neurons in the central nervous system but are brain region and cell type specific. In addition, recent studies have suggested that some neural functions successfully maintained with age may depend on the ability of neurons to preserve or repair their neural circuitry in response to naturally occurring cell loss, deafferentation or neurotransmitter deficits. Understanding these cellular mechanisms will provide a basis for the development of future therapeutic strategies aimed a amelioration of age-related neurodegenerative diseases. The proposed program project aims at contributing to such progress by providing information on age-related changes in basal ganglia function of the mammalian brain. Basal ganglia play a central role in motor function and are also involved in functions related to cognitive and emotional behavior. Disturbances in these behaviors are a clinical feature in some, but no all, elderly individuals and neuropathological changes in the basal ganglia are the hallmark of several neurodegenerative diseases, including Huntington's and Parkinson's disease. Emphasis is given to dopaminergic neurons whose degeneration is responsible for the majority of parkinsonian symptoms. The study of plasticity, neuronal death and age-related functional changes represent the main area of focus of the program project. Plasticity is a normal feature of a developing nervous system. While less frequent in the adult and aging nervous system, plasticity can occur in response to age- or disease-induced degenerative processes. The program project aims at establishing the extent of changes in plasticity of basal ganglia neurons induced by aging and injury. Neuronal atrophy an degeneration of selected neuronal populations accompanies aging and selective neuronal death is a key feature of neurodegenerative diseases. Several specific aims of the program project are directed at understanding molecular mechanisms playing a key role in determining the consequences of aging or of experimental injury to the brain. Attempts to prevent neuronal degeneration by the pharmacological use of protein growth factors or compounds altering transmitter functions is a central theme in the program project. Functional adaptations to aging have been partly characterized for dopaminergic but not other basal ganglia neurons. A specific aim of this program project is to study the molecular basis of selected age-related functional changes in the dopaminergic system. An entire project aims at providing the first thorough description of age-related changes in electrophysiological behavior of basal ganglia neurons. Preliminary findings reported in this application suggest that aging does not generally produce neuronal atrophy or impair plasticity and functional adaptation of basal ganglia neurons. The possibility that basal ganglia undergo unique age-related changes retaining the ability for plasticity and functional adaptation represents a key hypothesis to be tested. The proposed experiments will help to understand the molecular basis of plasticity and adaptation.

Alzheimer's disease (AD) is associated with degenerative changes and loss of various populations of neurons in the brain. Since neurotrophic factors regulate neuronal survival and maintenance of function, such factors may provide avenues for the development of effective treatment for AD. By preventing or attenuating neuronal degeneration, neurotrophic factors may slow down or stop the progression of the disease and associated cognitive disturbances. Based on its ability to prevent degenerative changes of cholinergic neurons induced by experimental lesions in rats and monkeys, nerve growth factor (NGF) is presently under consideration as experimental treatment for AD. One specific aim of this application, related to NGF's long-term effects and the feasibility of slow-releasing implants, will provide additional information valuable for clinical use of NGF. However, the value of NGF in the treatment of AD is limited by its specificity for forebrain cholinergic neurons. The recent discovery of homologs of NGF, brain-derived neurotrophic factor (BDNF) and neurotrophin-3 (NT-3), established the family of neurotrophins. These neurotrophins affect a variety of peripheral and central neurons and their effects overlap for some neuronal populations but differ for others. Recent findings, which provide the basis for this application, indicate that posttranslational modifications of the neurotrophins influence their biological activities and their specificity to act on subsets of neuronal populations. Therefore, it is proposed to prepare modified molecular forms of the neurotrophins either by recombinant expression or by specific protease treatment. Biological effects of these modified neurotrophins will then be characterized initially in neurotrophin receptor assays and in cell culture assays.Biologically active modified neurotrophins selected by the in vitro assays will then be tested in vivo in rat brain lesion models mimicking selective degenerative changes occurring in AD brains. These modified neurotrophins will be tested for their ability to counteract molecular, morphological, and behavioral deficits induced by the experimental lesions. Additional studies, using immunohistochemistry and retrograde transport methodologies, aim at identifying previously unknown neurotrophin-responsive cell populations. In summary, this program project aims at identifying modified neurotrophins able to specifically affect survival and function of neuronal populations degenerating in Alzheimer's disease. It is hoped that the studies will lead to the discovery of modified neurotrophins with either restricted or broadened biological specificity when compared with their natural counterparts and in the identification of molecules most promising for further development as therapeutics in Alzheimer's disease. The program project joins established academic and industrial scientists with a record of previous collaboration in the pharmacological exploitation of neurotrophic factors for developing effective treatment for AD and cognitive diseases.

Summary: Alzheimer's disease (AD) is associated with degeneration changes and loss of various specific populations of neurons in the brain. Since neurotrophic factors regulate survival and differentiated functions of neurons, they may provide avenues for the development of effective treatment for AD. By preventing or attenuating neuronal degeneration, neurotrophic factors may slow down or stop the progression of the disease and associated cognitive disturbances. Based on its ability to prevent degenerative changes of cholinergic neurons induced by experimental lesions in rats and monkeys, nerve growth factor (NGF) is presently considered as experimental treatment for AD. The value of NGF in the treatment of AD is limited by its selectivity for forebrain cholinergic neurons. NGF homologs, brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3) and neurotrophin-4/5 (NT-4/5), forming the family of neurotrophins, affect many populations of brain neurons. The analysis of their neurotrophic actions is ongoing. They appear to have different, but partially overlapping spectra of activity on select populations of central neurons. Aiming at identifying the most useful neurotrophin molecules, we propose to prepare neurotrophins, neurotrophin mutants, modified neurotrophins, as well as novel neurotrophic factors for this program project. The new molecules will be characterized initially in receptor assays and in cell culture assays. Biologically active molecules selected by the in vitro assays will then be tested in vivo in rat brain lesion models mimicking degenerative changes occurring in AD brains. Models of neurodegeneration to be used include septo-hippocampal cholinergic and GABAergic neurons after fimbrial transection, cortico- hippocampal neurons after transections of the angular bundle, and thalamo-cortical neurons after cingulotomy. Two additional projects are proposed. To assess the therapeutic potential of neurotrophins, we plan to measure the responsiveness of brain tissue to neurotrophins using a novel ex vivo assay. We propose to further analyze the therapeutic potential of the small molecules K-252a and K-252b, compounds earlier shown to potentiate the action of NT-3. It is hoped that the studies will lead to the identification of neurotrophin variants, other neurotrophic factors or small molecules most useful for further development as therapeutics in Alzheimer's disease. The program project joins established academic and industrial scientists with a record of previous collaboration in the pharmacological exploitation of neurotrophic factors for developing effective treatment for AD and cognitive diseases.

DESCRIPTION (provided by applicant): Dementia, with its most prevalent subforms Alzheimer's disease (AD), dementia with Lewy bodies (DLB), and vascular dementia, is a very substantial medical and societal problem. The emergence of many drugs and drug candidates that interfere with the disease mechanism of specific types of dementia makes it crucial to distinguish early between the various forms. Useful diagnostic tools will make it possible to direct patients to the most appropriate drug regimen, to avoid exposure to potentially harmful drugs, and to monitor the efficacy of disease-modifying drugs. Brain imaging techniques provide a non-invasive tool for differential diagnosis and monitoring of disease progression of dementia. We and others have been developing radiopharmaceuticals that allow detection of A amyloid deposits typical for AD. Our efforts have focused on 18F-labeled compounds for PET imaging, with the goal that the imaging diagnostics can be made widely available at minimal costs. We now propose the clinical development of a new radioligand for the differential diagnosis of DLB from AD as well as the monitoring of DLB disease progression. 18F-AV-133 is a highly selective ligand for the vesicular monoamine transporter VMAT2, believed to be the best marker for the functional integrity of dopaminergic neurons which are known to degenerate in DLB. Our proposal tests that hypothesis that imaging of VMAT2 provides a useful biomarker for dopaminergic neuron degeneration, which can be synergistically combined with amyloid plaque imaging to generate a powerful diagnostic work up for classification of dementia patients. We propose to conduct two clinical studies to establish safety and proof-of-concept for 18F-AV-133 as a diagnostic imaging agent for DLB. PUBLIC HEALTH RELEVANCE: Successful completion of the proposed phase I studies will provide a key step towards the provision of diagnostic tools for the clinical diagnosis and monitoring of disease progression of dementia with Lewy bodies (DLB), Alzheimer's disease (AD) and dementias in general. The studies will identify a diagnostic tool to allow the physicians to direct patients towards the most appropriate drug regimen and prevent that the patients are exposed to potentially harmful drugs. Furthermore, the resulting diagnostic tools will help significantly in the development of disease-modifying drugs, by making it possible to monitor treatment efficacy at the molecular level in vivo. Such a diagnostic tool will represent a significant technological and medical benefit to society.