Activation of microglia by classical inflammatory mediators can convert astrocytes to a neurotoxic A1 phenotype in a variety of neurological diseases1,2. Development of agents that could inhibit the formation of A1 reactive astrocytes could be used to treat these diseases for which there are no disease modifying therapies. Glucagon-like peptide-1 receptor (GLP-1R) agonists have been touted as potential neuroprotective agents for neurologic disorders such as Alzheimer’s disease (AD) and Parkinson’s disease (PD)3-13. The mechanisms by which GLP-1R agonists are neuroprotective are not known. Here we show that a potent, brain penetrant long acting GLP-1R agonist NLY01 protects against the loss of dopamine neurons and behavioral deficits in the α-synuclein preformed fibril (α-syn PFF) model of sporadic PD14,15. NLY01 also prolongs the life and reduces the behavioral deficits and neuropathological abnormalities in the human A53T α-synuclein (hA53T) transgenic (Tg) model of α-synucleinopathy induced neurodegeneration16. We found that NLY01 is a potent GLP-1R agonist with favorable properties that is neuroprotective via the direct prevention of microglial mediated conversion of astrocytes to an A1 neurotoxic phenotype. In light of NLY01 favorable properties it should be evaluated in the treatment of PD and related neurologic disorders characterized by microglial activation.

INTRODUCTION Parkinson’s disease (PD) is the second most common neurodegenerative disorder that leads to slowness of movement, tremor, rigidity and in the later stages of PD, cognitive impairment. Pathologically PD is characterized by the accumulation of α-synuclein in Lewy bodies and neurites. There is degeneration of neurons throughout the nervous system with the degeneration of dopamine neurons in the substantia nigra pars compacta leading to the major symptoms of PD. RATIONALE In the brains of PD patients, pathologic α-synuclein seems to spread from cell-to-cell via self-amplification, propagation, and transmission in a stereotypical and topographical pattern among neighboring cells and/or anatomically connected brain regions. The spread or transmission of pathologic α-synuclein is emerging as potentially important driver of PD pathogenesis. The underlying mechanisms and molecular entities responsible for the transmission of pathologic α-synuclein from cell-to-to cell are not known, but the entry of pathologic α-synuclein into neurons is thought to occur, in part through an active clathrin-dependent endocytic process. RESULTS Using recombinant α-synuclein pre-formed fibrils (PFF) as a model system to study the transmission of misfolded α-synuclein from neuron to neuron, we screened a library encoding transmembrane proteins for α-synuclein-biotin PFF binding candidates via detection by streptavidin-AP (alkaline phosphatase) staining. Three positive clones were identified that bind α-synuclein PFF and include lymphocyte-activation gene 3 (LAG3), neurexin 1β and amyloid beta precursor-like protein 1 (APLP1). Of these three transmembrane proteins, LAG3 demonstrated the highest ratio of selectivity for α-synuclein PFF over the α-synuclein monomer. α-Synuclein PFF binds to LAG3 in a saturable manner (Kd = 77 nM), while the α-synuclein monomer does not bind to LAG3. Co-immunoprecipitation also suggests that pathological α-synuclein PFF specifically binds to LAG3. Tau PFF, β-amyloid oligomer and β-amyloid PFF do not bind LAG3 indicating that LAG3 is specific for α-synuclein PFF. The internalization of α-synuclein PFF involves LAG3 since deletion of LAG3 reduces the endocytosis of α-synuclein PFF. LAG3 colocalizes with the endosomal GTPases, Rab5 and Rab7 and co-endocytoses with pathologic α-synuclein. Neuron-to-neuron transmission of pathologic α-synuclein and the accompanying pathology and neurotoxicity is substantially attenuated by deletion of LAG3 or by LAG3 antibodies. The lack of LAG3 also substantially delayed α-synuclein PFF induced loss of dopamine neurons, as well as biochemical and behavioral deficits in vivo. CONCLUSION We discovered that pathologic α-synuclein transmission and toxicity is initiated by binding to LAG3 and that neuron-to-neuron transmission of pathological α-synuclein involves the endocytosis of exogenous α-synuclein PFF by the engagement of LAG3 on neurons. Depletion of LAG3 or antibodies to LAG3 substantially reduce the pathology set in motion by the transmission of pathologic α-...

The pathologic accumulation and aggregation of α-synuclein (α-syn) underlies Parkinson’s disease (PD). The molecular mechanisms by which pathologic α-syn causes neurodegeneration in PD is not known. Here we found that pathologic α-syn activates poly (ADP-ribose) (PAR) polymerase-1 (PARP-1) and PAR generation accelerates the formation of pathologic α-syn resulting in cell death via parthanatos. PARP inhibitors or genetic deletion of PARP-1 prevented pathologic α-syn toxicity. In a feed-forward loop, PAR converted pathologic α-syn to a more toxic strain. PAR levels were increased in the cerebral spinal fluid and brains of PD patients suggesting that PARP activation plays a role in PD pathogenesis. Thus, strategies aimed at inhibiting PARP-1 activation could hold promise as a disease modifying therapy to prevent the loss of dopamine (DA) neurons in PD.