Kynurenine 3-monooxygenase inhibition in blood ameliorates neurodegeneration.

Metabolites in the kynurenine pathway, generated by tryptophan degradation, are thought to play an important role in neurodegenerative disorders, including Alzheimer's and Huntington's diseases. In these disorders, glutamate receptor-mediated excitotoxicity and free radical formation have been correlated with decreased levels of the neuroprotective metabolite kynurenic acid. Here, we describe the synthesis and characterization of JM6, a small-molecule prodrug inhibitor of kynurenine 3-monooxygenase (KMO). Chronic oral administration of JM6 inhibits KMO in the blood, increasing kynurenic acid levels and reducing extracellular glutamate in the brain. In a transgenic mouse model of Alzheimer's disease, JM6 prevents spatial memory deficits, anxiety-related behavior, and synaptic loss. JM6 also extends life span, prevents synaptic loss, and decreases microglial activation in a mouse model of Huntington's disease. These findings support a critical link between tryptophan metabolism in the blood and neurodegeneration, and they provide a foundation for treatment of neurodegenerative diseases.

Reduced IGF-1 signaling delays age-associated proteotoxicity in mice.

The insulin/insulin growth factor (IGF) signaling (IIS) pathway is a key regulator of aging of worms, flies, mice, and likely humans. Delayed aging by IIS reduction protects the nematode C. elegans from toxicity associated with the aggregation of the Alzheimer's disease-linked human peptide, Abeta. We reduced IGF signaling in Alzheimer's model mice and discovered that these animals are protected from Alzheimer's-like disease symptoms, including reduced behavioral impairment, neuroinflammation, and neuronal loss. This protection is correlated with the hyperaggregation of Abeta leading to tightly packed, ordered plaques, suggesting that one aspect of the protection conferred by reduced IGF signaling is the sequestration of soluble Abeta oligomers into dense aggregates of lower toxicity. These findings indicate that the IGF signaling-regulated mechanism that protects from Abeta toxicity is conserved from worms to mammals and point to the modulation of this signaling pathway as a promising strategy for the development of Alzheimer's disease therapy.

Combined Active Humoral and Cellular Immunization Approaches for the Treatment of Synucleinopathies.

Dementia with Lewy bodies, Parkinson's disease, and Multiple System Atrophy are age-related neurodegenerative disorders characterized by progressive accumulation of α-synuclein (α-syn) and jointly termed synucleinopathies. Currently, no disease-modifying treatments are available for these disorders. Previous preclinical studies demonstrate that active and passive immunizations targeting α-syn partially ameliorate behavioral deficits and α-syn accumulation; however, it is unknown whether combining humoral and cellular immunization might act synergistically to reduce inflammation and improve microglial-mediated α-syn clearance. Since combined delivery of antigen plus rapamycin (RAP) in nanoparticles is known to induce antigen-specific regulatory T cells (Tregs), we adapted this approach to α-syn using the antigen-presenting cell-targeting glucan microparticle (GP) vaccine delivery system. PDGF-α-syn transgenic (tg) male and female mice were immunized with GP-alone, GP-α-syn (active humoral immunization), GP+RAP, or GP+RAP/α-syn (combined active humoral and Treg) and analyzed using neuropathological and biochemical markers. Active immunization resulted in higher serological total IgG, IgG1, and IgG2a anti-α-syn levels. Compared with mice immunized with GP-alone or GP-α-syn, mice vaccinated with GP+RAP or GP+RAP/α-syn displayed increased numbers of CD25-, FoxP3-, and CD4-positive cells in the CNS. GP-α-syn or GP+RAP/α-syn immunizations resulted in a 30-45% reduction in α-syn accumulation, neuroinflammation, and neurodegeneration. Mice immunized with GP+RAP/α-syn further rescued neurons and reduced neuroinflammation. Levels of TGF-ß1 were increased with GP+RAP/α-syn immunization, while levels of TNF-α and IL-6 were reduced. We conclude that the observed effects of GP+RAP/α-syn immunization support the hypothesis that cellular immunization may enhance the effects of active immunotherapy for the treatment of synucleinopathies.SIGNIFICANCE STATEMENT We show that a novel vaccination modality combining an antigen-presenting cell-targeting glucan particle (GP) vaccine delivery system with encapsulated antigen (α-synuclein) + rapamycin (RAP) induced both strong anti-α-synuclein antibody titers and regulatory T cells (Tregs). This vaccine, collectively termed GP+RAP/α-syn, is capable of triggering neuroprotective Treg responses in synucleinopathy models, and the combined vaccine is more effective than the humoral or cellular immunization alone. Together, these results support the further development of this multifunctional vaccine approach for the treatment of synucleinopathies, such as Parkinson's disease, dementia with Lewy bodies, and multiple systems atrophy.

NitroSynapsin for the treatment of neurological manifestations of tuberous sclerosis complex in a rodent model.

Tuberous sclerosis (TSC) is an autosomal dominant disorder caused by heterozygous mutations in the TSC1 or TSC2 gene. TSC is often associated with neurological, cognitive, and behavioral deficits. TSC patients also express co-morbidity with anxiety and mood disorders. The mechanism of pathogenesis in TSC is not entirely clear, but TSC-related neurological symptoms are accompanied by excessive glutamatergic activity and altered synaptic spine structures. To address whether extrasynaptic (e)NMDA-type glutamate receptor (NMDAR) antagonists, as opposed to antagonists that block physiological phasic synaptic activity, can ameliorate the synaptic and behavioral features of this disease, we utilized the Tsc2+/- mouse model of TSC to measure biochemical, electrophysiological, histological, and behavioral parameters in the mice. We found that antagonists that preferentially block tonic activity as found at eNMDARs, particularly the newer drug NitroSynapsin, provide biological and statistically significant improvement in Tsc2+/- phenotypes. Accompanying this improvement was correction of activity in the p38 MAPK-TSC-Rheb-mTORC1-S6K1 pathway. Deficits in hippocampal long-term potentiation (LTP), histological loss of synapses, and behavioral fear conditioning in Tsc2+/- mice were all improved after treatment with NitroSynapsin. Taken together, these results suggest that amelioration of excessive excitation, by limiting aberrant eNMDAR activity, may represent a novel treatment approach for TSC.

Systemic peptide mediated delivery of an siRNA targeting α-syn in the CNS ameliorates the neurodegenerative process in a transgenic model of Lewy body disease.

Neurodegenerative disorders of the aging population are characterized by progressive accumulation of neuronal proteins such as α-synuclein (α-syn) in Parkinson's Disease (PD) and Amyloid ß (Aß) and Tau in Alzheimer's disease (AD) for which no treatments are currently available. The ability to regulate the expression at the gene transcription level would be beneficial for reducing the accumulation of these proteins or regulating expression levels of other genes in the CNS. Short interfering RNA molecules can bind specifically to target RNAs and deliver them for degradation. This approach has shown promise therapeutically in vitro and in vivo in mouse models of PD and AD and other neurological disorders; however, delivery of the siRNA to the CNS in vivo has been achieved primarily through intra-cerebral or intra-thecal injections that may be less amenable for clinical translation; therefore, alternative approaches for delivery of siRNAs to the brain is needed. Recently, we described a small peptide from the envelope protein of the rabies virus (C2-9r) that was utilized to deliver an siRNA targeting α-syn across the blood brain barrier (BBB) following intravenous injection. This approach showed reduced expression of α-syn and neuroprotection in a toxic mouse model of PD. However, since receptor-mediated delivery is potentially saturable, each allowing the delivery of a limited number of molecules, we identified an alternative peptide for the transport of nucleotides across the BBB based on the apolipoprotein B (apoB) protein targeted to the family of low-density lipoprotein receptors (LDL-R). We used an 11-amino acid sequence from the apoB protein (ApoB11) that, when coupled with a 9-amino acid arginine linker, can transport siRNAs across the BBB to neuronal and glial cells. To examine the value of this peptide mediated oligonucleotide delivery system for PD, we delivered an siRNA targeting the α-syn (siα-syn) in a transgenic mouse model of PD. We found that ApoB11 was effective (comparable to C2-9r) at mediating the delivery of siα-syn into the CNS, co-localized to neurons and glial cells and reduced levels of α-syn protein translation and accumulation. Delivery of ApoB11/siα-syn was accompanied by protection from degeneration of selected neuronal populations in the neocortex, limbic system and striato-nigral system and reduced neuro-inflammation. Taken together, these results suggest that systemic delivery of oligonucleotides targeting α-syn using ApoB11 might be an interesting alternative strategy worth considering for the experimental treatment of synucleinopathies.

Effects of single and combined immunotherapy approach targeting amyloid ß protein and α-synuclein in a dementia with Lewy bodies-like model.

INTRODUCTION: Immunotherapeutic approaches targeting amyloid ß (Aß) protein and tau in Alzheimer's disease and α-synuclein (α-syn) in Parkinson's disease are being developed for treating dementia with Lewy bodies. However, it is unknown if single or combined immunotherapies targeting Aß and/or α-syn may be effective. METHODS: Amyloid precursor protein/α-syn tg mice were immunized with AFFITOPEs® (AFF) peptides specific to Aß (AD02) or α-syn (PD-AFF1) and the combination. RESULTS: AD02 more effectively reduced Aß and pTau burden; however, the combination exhibited some additive effects. Both AD02 and PD-AFF1 effectively reduced α-syn, ameliorated degeneration of pyramidal neurons, and reduced neuroinflammation. PD-AFF1 more effectively ameliorated cholinergic and dopaminergic fiber loss; the combined immunization displayed additive effects. AD02 more effectively improved buried pellet test behavior, whereas PD-AFF1 more effectively improved horizontal beam test; the combined immunization displayed additive effects. DISCUSSION: Specific active immunotherapy targeting Aß and/or α-syn may be of potential interest for the treatment of dementia with Lewy bodies.

α-Synuclein interferes with the ESCRT-III complex contributing to the pathogenesis of Lewy body disease.

α-Synuclein (α-syn) has been implicated in neurological disorders with parkinsonism, including Parkinson's disease and Dementia with Lewy body. Recent studies have shown α-syn oligomers released from neurons can propagate from cell-to-cell in a prion-like fashion exacerbating neurodegeneration. In this study, we examined the role of the endosomal sorting complex required for transport (ESCRT) pathway on the propagation of α-syn. α-syn, which is transported via the ESCRT pathway through multivesicular bodies for degradation, can also target the degradation of the ESCRT protein-charged multivesicular body protein (CHMP2B), thus generating a roadblock of endocytosed α-syn. Disruption of the ESCRT transport system also resulted in increased exocytosis of α-syn thus potentially increasing cell-to-cell propagation of synuclein. Conversely, delivery of a lentiviral vector overexpressing CHMP2B rescued the neurodegeneration in α-syn transgenic mice. Better understanding of the mechanisms of intracellular trafficking of α-syn might be important for understanding the pathogenesis and developing new treatments for synucleinopathies.

Glucocerebrosidase modulates cognitive and motor activities in murine models of Parkinson's disease.

Mutations in GBA1, the gene encoding glucocerebrosidase, are associated with an enhanced risk of developing synucleinopathies such as Parkinson's disease (PD) and dementia with Lewy bodies. A higher prevalence and increased severity of motor and non-motor symptoms is observed in PD patients harboring mutant GBA1 alleles, suggesting a link between the gene or gene product and disease development. Interestingly, PD patients without mutations in GBA1 also exhibit lower levels of glucocerebrosidase activity in the central nervous system (CNS), implicating this lysosomal enzyme in disease pathogenesis. Here, we investigated whether modulation of glucocerebrosidase activity in murine models of synucleinopathy (expressing wild type Gba1) affected α-synuclein accumulation and behavioral phenotypes. Partial inhibition of glucocerebrosidase activity in PrP-A53T-SNCA mice using the covalent inhibitor conduritol-B-epoxide induced a profound increase in soluble α-synuclein in the CNS and exacerbated cognitive and motor deficits. Conversely, augmenting glucocerebrosidase activity in the Thy1-SNCA mouse model of PD delayed the progression of synucleinopathy. Adeno-associated virus-mediated expression of glucocerebrosidase in the Thy1-SNCA mouse striatum led to decrease in the levels of the proteinase K-resistant fraction of α-synuclein, amelioration of behavioral aberrations and protection from loss of striatal dopaminergic markers. These data indicate that increasing glucocerebrosidase activity can influence α-synuclein homeostasis, thereby reducing the progression of synucleinopathies. This study provides robust in vivo evidence that augmentation of CNS glucocerebrosidase activity is a potential therapeutic strategy for PD, regardless of the mutation status of GBA1.

Selective targeting of 3 repeat Tau with brain penetrating single chain antibodies for the treatment of neurodegenerative disorders.

Alzheimer's disease (AD) is the most common form of dementia in the elderly affecting more than 5 million people in the U.S. AD is characterized by the accumulation of ß-amyloid (Aß) and Tau in the brain, and is manifested by severe impairments in memory and cognition. Therefore, removing tau pathology has become one of the main therapeutic goals for the treatment of AD. Tau (tubulin-associated unit) is a major neuronal cytoskeletal protein found in the CNS encoded by the gene MAPT. Alternative splicing generates two major isoforms of tau containing either 3 or 4 repeat (R) segments. These 3R or 4RTau species are differentially expressed in neurodegenerative diseases. Previous studies have been focused on reducing Tau accumulation with antibodies against total Tau, 4RTau or phosphorylated isoforms. Here, we developed a brain penetrating, single chain antibody that specifically recognizes a pathogenic 3RTau. This single chain antibody was modified by the addition of a fragment of the apoB protein to facilitate trafficking into the brain, once in the CNS these antibody fragments reduced the accumulation of 3RTau and related deficits in a transgenic mouse model of tauopathy. NMR studies showed that the single chain antibody recognized an epitope at aa 40-62 of 3RTau. This single chain antibody reduced 3RTau transmission and facilitated the clearance of Tau via the endosomal-lysosomal pathway. Together, these results suggest that targeting 3RTau with highly specific, brain penetrating, single chain antibodies might be of potential value for the treatment of tauopathies such as Pick's Disease.

Reducing Endogenous α-Synuclein Mitigates the Degeneration of Selective Neuronal Populations in an Alzheimer's Disease Transgenic Mouse Model.

UNLABELLED: Alzheimer's disease (AD) is characterized by the progressive accumulation of amyloid ß (Aß) and microtubule associate protein tau, leading to the selective degeneration of neurons in the neocortex, limbic system, and nucleus basalis, among others. Recent studies have shown that α-synuclein (α-syn) also accumulates in the brains of patients with AD and interacts with Aß and tau, forming toxic hetero-oligomers. Although the involvement of α-syn has been investigated extensively in Lewy body disease, less is known about the role of this synaptic protein in AD. Here, we found that reducing endogenous α-syn in an APP transgenic mouse model of AD prevented the degeneration of cholinergic neurons, ameliorated corresponding deficits, and recovered the levels of Rab3a and Rab5 proteins involved in intracellular transport and sorting of nerve growth factor and brain-derived neurotrophic factor. Together, these results suggest that α-syn might participate in mechanisms of vulnerability of selected neuronal populations in AD and that reducing α-syn might be a potential approach to protecting these populations from the toxic effects of Aß. SIGNIFICANCE STATEMENT: Reducing endogenous α-synuclein (α-syn) in an APP transgenic mouse model of Alzheimer's disease (AD) prevented the degeneration of cholinergic neurons, ameliorated corresponding deficits, and recovered the levels of Rab3a and Rab5 proteins involved in intracellular transport and sorting of nerve growth factor and brain-derived neurotrophic factor. These results suggest that α-syn might participate in mechanisms of vulnerability of selected neuronal populations in AD and that reducing α-syn might be a potential approach to protecting these populations from the toxic effects of amyloid ß.

Systemic Central Nervous System (CNS)-targeted Delivery of Neuropeptide Y (NPY) Reduces Neurodegeneration and Increases Neural Precursor Cell Proliferation in a Mouse Model of Alzheimer Disease.

Neuropeptide Y (NPY) is one of the most abundant protein transmitters in the central nervous system with roles in a variety of biological functions including: food intake, cardiovascular regulation, cognition, seizure activity, circadian rhythms, and neurogenesis. Reduced NPY and NPY receptor expression is associated with numerous neurodegenerative disorders including Alzheimer disease (AD). To determine whether replacement of NPY could ameliorate some of the neurodegenerative and behavioral pathology associated with AD, we generated a lentiviral vector expressing NPY fused to a brain transport peptide (apoB) for widespread CNS delivery in an APP-transgenic (tg) mouse model of AD. The recombinant NPY-apoB effectively reversed neurodegenerative pathology and behavioral deficits although it had no effect on accumulation of Aß. The subgranular zone of the hippocampus showed a significant increase in proliferation of neural precursor cells without further differentiation into neurons. The neuroprotective and neurogenic effects of NPY-apoB appeared to involve signaling via ERK and Akt through the NPY R1 and NPY R2 receptors. Thus, widespread CNS-targeted delivery of NPY appears to be effective at reversing the neuronal and glial pathology associated with Aß accumulation while also increasing NPC proliferation. Overall, increased delivery of NPY to the CNS for AD might be an effective therapy especially if combined with an anti-Aß therapeutic.

Differential effects of immunotherapy with antibodies targeting α-synuclein oligomers and fibrils in a transgenic model of synucleinopathy.

Disorders with progressive accumulation of α-synuclein (α-syn) are a common cause of dementia and parkinsonism in the aging population. Accumulation and propagation of α-syn play a role in the pathogenesis of these disorders. Previous studies have shown that immunization with antibodies that recognize C-terminus of α-syn reduces the intra-neuronal accumulation of α-syn and related deficits in transgenic models of synucleinopathy. These studies employed antibodies that recognize epitopes within monomeric and aggregated α-syn that were generated through active immunization or administered via passive immunization. However, it is possible that more specific effects might be achieved with antibodies recognizing selective species of the α-syn aggregates. In this respect we recently developed antibodies that differentially recognized various oligomers (Syn-O1, -O2, and -O4) and fibrilar (Syn-F1 and -F2) forms of α-syn. For this purpose wild-type α-syn transgenic (line 61) mice were immunized with these 5 different antibodies and neuropathologically and biochemically analyzed to determine which was most effective at reducing α-syn accumulation and related deficits. We found that Syn-O1, -O4 and -F1 antibodies were most effective at reducing accumulation of α-syn oligomers in multiple brain regions and at preventing neurodegeneration. Together this study supports the notion that selective antibodies against α-syn might be suitable for development new treatments for synucleinopathies such as PD and DLB.

The anticancer drug sunitinib promotes autophagyand protects from neurotoxicity in an HIV-1 Tat model of neurodegeneration.

Despite the success of antiretroviral therapies to control systemic HIV-1 infection, the prevalence of HIV-associated neurocognitive disorders (HANDs) has not decreased among aging patients with HIV. Autophagy pathway alterations, triggered by HIV-1 proteins including gp120, Tat, and Nef, might contribute to the neurodegenerative process in aging patients with HAND. Although no treatments are currently available to manage HAND, we have previously shown that sunitinib, an anticancer drug that blocks receptor tyrosine-kinase and cyclin kinase pathways, might be of interest. Studies in cancer models suggest that sunitinib might also modulate autophagy, which is dysregulated in our models of Tat-induced neurotoxicity. We evaluated the efficacy of sunitinib to promote autophagy in the CNS and ameliorate neurodegeneration using LC3-GFP-expressing neuronal cells challenged with low concentrations of Tat and using inducible Tat transgenic mice. In neuronal cultures challenged with low levels of Tat, sunitinib increased markers of autophagy such as LC3-II and reduced p62 accumulation in a dose-dependent manner. In vivo, sunitinib treatment restored LC3-II, p62, and endophilin B1 (EndoB1) levels in doxycycline-induced Tat transgenic mice. Moreover, in these animals, sunitinib reduced the hyperactivation of CDK5, tau hyperphosphorylation, and p35 cleavage to p25. Restoration of CDK5 and autophagy were associated with reduced neurodegeneration and behavioral alterations. Alterations in autophagy in the Tat tg mice were associated with reduced levels of a CDK5 substrate, EndoB1, and levels of total EndoB1 were normalized by sunitinib treatment. We conclude that sunitinib might ameliorate Tat-mediated autophagy alterations and may decrease neurodegeneration in aging patients with HAND.

Distinct Pattern of Microgliosis in the Olfactory Bulb of Neurodegenerative Proteinopathies.

The olfactory bulb (OB) shows early neuropathological hallmarks in numerous neurodegenerative diseases, for example, in Alzheimer's disease (AD) and Parkinson's disease (PD). The glomerular and granular cell layer of the OB is characterized by preserved cellular plasticity in the adult brain. In turn, alterations of this cellular plasticity are related to neuroinflammation such as microglia activation, implicated in the pathogenesis of AD and PD, as well as frontotemporal lobe degeneration (FTLD). To determine microglia proliferation and activation we analyzed ionized calcium binding adaptor molecule 1 (Iba1) expressing microglia in the glomerular and granular cell layer, and the olfactory tract of the OB from patients with AD, PD dementia/dementia with Lewy bodies (PDD/DLB), and FTLD compared to age-matched controls. The number of Iba1 and CD68 positive microglia associated with enlarged amoeboid microglia was increased particularly in AD, to a lesser extent in FTLD and PDD/DLB as well, while the proportion of proliferating microglia was not altered. In addition, cells expressing the immature neuronal marker polysialylated neural cell adhesion molecule (PSA-NCAM) were increased in the glomerular layer of PDD/DLB and FTLD cases only. These findings provide novel and detailed insights into differential levels of microglia activation in the OB of neurodegenerative diseases.

HIV-1 Tat alters neuronal autophagy by modulating autophagosome fusion to the lysosome: implications for HIV-associated neurocognitive disorders.

Antiretroviral therapy has increased the life span of HIV+ individuals; however, HIV-associated neurocognitive disorder (HAND) occurrence is increasing in aging HIV patients. Previous studies suggest HIV infection alters autophagy function in the aging CNS and HIV-1 proteins affect autophagy in monocyte-derived cells. Despite these findings, the mechanisms leading to dysregulated autophagy in the CNS remain unclear. Here we sought to determine how HIV Tat dysregulates autophagy in neurons. Tat caused a dose-dependent decrease in autophagosome markers, microtubule-associated protein-1 light chain ß II (LC3II), and sequestosome 1(SQSTM1), in a membrane-enriched fraction, suggesting Tat increases autophagic degradation. Bafilomycin A1 increased autophagosome number, LC3II, and SQSTM1 accumulation; Tat cotreatment diminished this effect. Tat had no effect when 3-methyladenine or knockdown of beclin 1 blocked early stages of autophagy. Tat increased numbers of LC3 puncta and resulted in the formation of abnormal autophagosomes in vitro. Likewise, in vivo studies in GFAP-Tat tg mice showed increased autophagosome accumulation in neurons, altered LC3II levels, and neurodegeneration. These effects were reversed by rapamycin treatment. Tat colocalized with autophagosome and lysosomal markers and enhanced the colocalization of autophagosome with lysosome markers. Furthermore, co-IP studies showed that Tat interacts with lysosomal-associated membrane protein 2A (LAMP2A) in vitro and in vivo, and LAMP2A overexpression reduces Tat-induced neurotoxicity. Hence, Tat protein may induce autophagosome and lysosome fusion through interaction with LAMP2A leading to abnormal neuronal autophagy function and dysregulated degradation of critical intracellular components. Therapies targeting Tat-mediated autophagy alterations may decrease neurodegeneration in aging patients with HAND.

HIV alters neuronal mitochondrial fission/fusion in the brain during HIV-associated neurocognitive disorders.

HIV-associated neurocognitive disorders (HAND) still occur in approximately 50% of HIV patients, and therapies to combat HAND progression are urgently needed. HIV proteins are released from infected cells and cause neuronal damage, possibly through mitochondrial abnormalities. Altered mitochondrial fission and fusion is implicated in several neurodegenerative disorders. Here, we hypothesized that mitochondrial fission/fusion may be dysregulated in neurons during HAND. We have identified decreased mitochondrial fission protein (dynamin 1-like; DNM1L) in frontal cortex tissues of HAND donors, along with enlarged and elongated mitochondria localized to the soma of damaged neurons. Similar pathology was observed in the brains of GFAP-gp120 tg mice. In vitro, recombinant gp120 decreased total and active DNM1L levels, reduced the level of Mitotracker staining, and increased extracellular acidification rate (ECAR) in primary neurons. DNM1L knockdown enhanced the effects of gp120 as measured by reduced Mitotracker signal in the treated cells. Interestingly, overexpression of DNM1L increased the level of Mitotracker staining in primary rat neurons and reduced neuroinflammation and neurodegeneration in the GFAP-gp120-tg mice. These data suggest that mitochondrial biogenesis dynamics are shifted towards mitochondrial fusion in brains of HAND patients and this may be due to gp120-induced reduction in DNM1L activity. Promoting mitochondrial fission during HIV infection of the CNS may restore mitochondrial biogenesis and prevent neurodegeneration.

Neuroprotective effects of the immunomodulatory drug FK506 in a model of HIV1-gp120 neurotoxicity.

BACKGROUND: HIV-associated neurocognitive disorders (HAND) continue to be a common morbidity associated with chronic HIV infection. It has been shown that HIV proteins (e.g., gp120) released from infected microglial/macrophage cells can cause neuronal damage by triggering inflammation and oxidative stress, activating aberrant kinase pathways, and by disrupting mitochondrial function and biogenesis. Previous studies have shown that FK506, an immunophilin ligand that modulates inflammation and mitochondrial function and inhibits calcineurin, is capable of rescuing the neurodegenerative pathology in models of Parkinson's disease, Alzheimer's disease, and Huntington's disease. In this context, the main objective of this study was to evaluate if FK506 could rescue the neuronal degeneration and mitochondrial alterations in a transgenic (tg) animal model of HIV1-gp120 neurotoxicity. METHODS: GFAP-gp120 tg mice were treated with FK506 and analyzed for neuropathology, behavior, mitochondrial markers, and calcium flux by two-photon microscopy. RESULTS: We found that FK506 reduced the neuronal cell loss and neuro-inflammation in the gp120 tg mice. Moreover, while vehicle-treated gp120 tg mice displayed damaged mitochondria and increased neuro-inflammatory markers, FK506 rescued the morphological mitochondrial alterations and neuro-inflammation while increasing levels of optic atrophy 1 and mitofusin 1. By two-photon microscopy, calcium levels were not affected in the gp120 tg mice and no effects of FK506 were detected. However, at a functional level, FK506 ameliorated the gp120 tg mice hyperactivity in the open field. CONCLUSIONS: Together, these results suggest that FK506 might be potentially neuroprotective in patients with HAND by mitigating inflammation and mitochondrial alterations.

Reducing C-terminal-truncated alpha-synuclein by immunotherapy attenuates neurodegeneration and propagation in Parkinson's disease-like models.

Parkinson's disease (PD) and dementia with Lewy bodies (DLB) are common neurodegenerative disorders of the aging population, characterized by progressive and abnormal accumulation of α-synuclein (α-syn). Recent studies have shown that C-terminus (CT) truncation and propagation of α-syn play a role in the pathogenesis of PD/DLB. Therefore, we explored the effect of passive immunization against the CT of α-syn in the mThy1-α-syn transgenic (tg) mouse model, which resembles the striato-nigral and motor deficits of PD. Mice were immunized with the new monoclonal antibodies 1H7, 5C1, or 5D12, all directed against the CT of α-syn. CT α-syn antibodies attenuated synaptic and axonal pathology, reduced the accumulation of CT-truncated α-syn (CT-α-syn) in axons, rescued the loss of tyrosine hydroxylase fibers in striatum, and improved motor and memory deficits. Among them, 1H7 and 5C1 were most effective at decreasing levels of CT-α-syn and higher-molecular-weight aggregates. Furthermore, in vitro studies showed that preincubation of recombinant α-syn with 1H7 and 5C1 prevented CT cleavage of α-syn. In a cell-based system, CT antibodies reduced cell-to-cell propagation of full-length α-syn, but not of the CT-α-syn that lacked the 118-126 aa recognition site needed for antibody binding. Furthermore, the results obtained after lentiviral expression of α-syn suggest that antibodies might be blocking the extracellular truncation of α-syn by calpain-1. Together, these results demonstrate that antibodies against the CT of α-syn reduce levels of CT-truncated fragments of the protein and its propagation, thus ameliorating PD-like pathology and improving behavioral and motor functions in a mouse model of this disease.

A neuroprotective brain-penetrating endopeptidase fusion protein ameliorates Alzheimer disease pathology and restores neurogenesis.

Alzheimer disease (AD) is characterized by widespread neurodegeneration throughout the association cortex and limbic system, deposition of amyloid-ß peptide (Aß) in the neuropil and around the blood vessels, and formation of neurofibrillary tangles. The endopeptidase neprilysin has been successfully used to reduce the accumulation of Aß following intracranial viral vector delivery or ex vivo manipulated intracranial delivery. These therapies have relied on direct injections into the brain, whereas a clinically desirable therapy would involve i.v. infusion of a recombinant enzyme. We previously characterized a recombinant neprilysin that contained a 38-amino acid brain-targeting domain. Recombinant cell lines have been generated expressing this brain-targeted enzyme (ASN12). In this report, we characterize the ASN12 recombinant protein for pharmacology in a mouse as well as efficacy in two APPtg mouse models of AD. The recombinant ASN12 transited to the brain with a t½ of 24 h and accumulated to 1.7% of injected dose at 24 h following i.v. delivery. We examined pharmacodynamics in the tg2576 APPtg mouse with the prion promoter APP695 SWE mutation and in the Line41 mThy1 APP751 mutation mouse. Treatment of either APPtg mouse resulted in reduced Aß, increased neuronal synapses, and improved learning and memory. In addition, the Line41 APPtg mice showed increased levels of C-terminal neuropeptide Y fragments and increased neurogenesis. These results suggest that the recombinant brain-targeted neprilysin, ASN12, may be an effective treatment for AD and warrant further investigation in clinical trials.