Alzheimers Disease Project: Neuroimmune responses and therapeutics of alpha-synucleinopathies of the aging population

National Institute On Aging

Investigators

Linked publications, trials & patents

Abstract

The main focus of the Molecular Neuropathology unit (MNU) at Laboratory of Neurogenetics (LNG) is to harness the immune system to better understand the mechanisms of neurodegeneration and to develop therapies for synucleinopathies of the aging population such as DLB, PD and AD. We propose 3 Aims, the first investigating the role of innate immune responses and combinatorial immunotherapy targeting LRRK2, Toll-like receptors, p38, NFAT and the protein aggregates (eg: a-synuclein, Abeta, tau) in PD/DLB; the second assessing downstream pro-inflammatory signaling pathways including MAPK-p38, NFAT and NFkB and the third evaluating the role of aging in T cell mediated adaptive immune cell responses in PD/DLB pathogenesis and for developing immunotherapies for synucleinopathies. During this period, we published 8 manuscripts mostly focusing understanding the pathogenesis of PD/DLB and developing novel pharmacological and anti-sense approaches. Progress for Aim1. In previous studies we identified several immune receptors that mediate neuroinflammation in synucleinopathies, including Toll-like receptor 2 (TLR2) and have shown which species of a-synuclein bind TLR2 and mediates neuroinflammation. We were interested at better characterizing the downstream pathways mediating neurodegeneration by using multiomics approaches. For this purpose we collaboratives with Dr. Jung laboratory in South Korea. First, we establish transcriptomic and epigenomic landscapes of the substantia nigra by profiling 113,207 nuclei obtained from healthy controls and patients with PD. Our multiomics data integration provides cell type annotation of 128,724 cis-regulatory elements (cREs) and uncovers cell type-specific dysregulations in cREs with a strong transcriptional influence on genes implicated in PD. The establishment of high-resolution three-dimensional chromatin contact maps identifies 656 target genes of dysregulated cREs and genetic risk loci, uncovering both potential and known PD risk genes. Notably, these candidate genes exhibit modular gene expression patterns with unique molecular signatures in distinct cell types, highlighting altered molecular mechanisms in immune cells like microglia as well as in oligodendroglial cells (Lee et al, Sci Adv 2023). Together, our single-cell transcriptome and epigenome reveal cell type-specific disruption in transcriptional regulations related to PD that are complementary with studies described for Aim 2. More recently, in collaboration with UCSD we investigated the therapeutical role of downregulating a-synuclein with targeted shRNA delivery into the hippocampus of mouse models of AD and DLB. We showed that downregulation of -syn results in significant reduction in the number of Ab plaques and tau. In addition, mice treated with LV-shRNA -syn had amelioration of abnormal microglial activation and astrocytosis in AD mice. This suggests a link between A and -syn in pathology pointing to a possible therapeutic angle for AD targeting -syn. This paper was published in JAD (Leitao et al JAD 2023). In other studies, in collaboration with the Cookson lab we studied the distribution of p-a-synuclein in control and PD brains using the PLA method (Arlinghaus et al J Park Dis 2023). Progress for Aim 2. We previously showed that once extracellular a-synuclein binds TLR2, (Kim et al Science TM 2020) LRRK2 promoted a neuroinflammatory cascade by selectively phosphorylating and inducing nuclear translocation of the immune transcription factor nuclear factor of activated T cells, cytoplasmic 2 (NFATc2). Our results suggest that modulation of LRRK2 and its downstream signaling mediator NFATc2 might be therapeutic targets for treating synucleinopathies. Next, we investigated the hypothesis that blocking NFATc2 translocation might ameliorate a-synuclein-mediated microglial neurotoxicity. Preliminary studies identified a novel peptide compound (11r VIVIT) that when administered peripherally into -syn tg mice reduced neuroinflammation, restores NFATc2 cytoplasmic localization and ameliorates behavioral deficits. We are currently expanding these studies to include detailed characterization of microglial transcriptomic, phenotypic profiling (M1 vs M2), neuropathological and functional examination. Our results suggest that modulation of LRRK2 and its downstream signaling mediator NFATc2 might be therapeutic targets for treating synucleinopathies. This paper is under revision at Nature Communications. Next, we investigated the role of p38 MAPK as a potential immunomodulatory pathway. We have shown that activation of the p38 MAPK isoform and mislocalization of the p38 MAPK isoform are associated with neuroinflammation and synaptic degeneration in DLB and PD. Therefore, we hypothesized that p38 might be associated with neuronal p38 distribution and synaptic dysfunction in these diseases. To test this hypothesis, we treated in vitro cellular and in vivo mouse models of DLB/PD with SKF-86002, a compound that attenuates inflammation by inhibiting p38/, and then investigated the effects of this compound on p38 and neurodegenerative pathology. We found that inhibition of p38 reduced neuroinflammation and ameliorated synaptic, neurodegenerative, and motor behavioral deficits in transgenic mice overexpressing human -synuclein. Moreover, treatment with SKF-86002 promoted the redistribution of p38 to synapses and reduced the accumulation of -synuclein in mice overexpressing human -synuclein. Treatment with SKF-86002 ameliorated -synuclein-induced neurodegeneration in these neurons only when microglia were pretreated with this compound. These findings provide a mechanistic connection between p38 and p38 as well as a rationale for targeting this pathway in DLB/PD. This study was recently published in Science Translational Medicine (Iba et al Science TM 2023). Progress for Aim 3. In collaboration with the Sen laboratory we showed that T cell infiltration with potential participation of NKT cells play an important role in DLB/PD. Next, we explored the role of aging in the pathogenesis of PD/DLB by analyzing, behavior, neuropathology and transcriptomics and immune responses in young and aged wildtype mice that received intrastriatal injections with a-synuclein preformed fibrils (pff). We found that aged mice showed more extensive accumulation of a-synuclein and behavioral deficits that was associated with greater infiltration of T cells and microgliosis. Distinct inflammatory patterns of gene expression in microglia showed that a-synuclein pff-induced genes networks in young mice (eg: CSF2, TNF, IL1b, IL6) that overlaps with genes differentially expressed in microglia in the aged mice. These results indicate that the aged and more inflamed brain micro-environment directly influences the disease outcome of pathological accumulation of a-syn in age-related chronic diseases such as DLB and PD. Other experiments underway includes investigating the effects of aging and deleting microglia with the PLX3387 (a compound that targets CSF1R) on synucleinopathies in DLB/PD models. Similar studies have been performed in collaboration with the Winkler laboratory (Erlangen University) in another model of synucleinopathy, that is a transgenic model of MSA treated with PLX5622, this study showed that deleting microglia delayed onset of neurological alterations (J Neuroscie 2023). We are also investigating the effects of aging and inflammation in models where T cells and microglia are deleted or where CSF1 and CSF2 are modulated, manuscripts for all these studies are under preparation. Other collaborations at LNG are with Drs. Cookson, Singleton, Traynor and Scholz on the genetic architecture of FTD, DLB and MSA by providing expert neuropathological assessment, animal models and human postmorten brain tissues.