URMC-099 facilitates amyloid-ß clearance in a murine model of Alzheimer's disease.

BACKGROUND: The mixed lineage kinase type 3 inhibitor URMC-099 facilitates amyloid-beta (Aß) clearance and degradation in cultured murine microglia. One putative mechanism is an effect of URMC-099 on Aß uptake and degradation. As URMC-099 promotes endolysosomal protein trafficking and reduces Aß microglial pro-inflammatory activities, we assessed whether these responses affect Aß pathobiogenesis. To this end, URMC-099's therapeutic potential, in Aß precursor protein/presenilin-1 (APP/PS1) double-transgenic mice, was investigated in this model of Alzheimer's disease (AD). METHODS: Four-month-old APP/PS1 mice were administered intraperitoneal URMC-099 injections at 10 mg/kg daily for 3 weeks. Brain tissues were examined by biochemical, molecular and immunohistochemical tests. RESULTS: URMC-099 inhibited mitogen-activated protein kinase 3/4-mediated activation and attenuated ß-amyloidosis. Microglial nitric oxide synthase-2 and arginase-1 were co-localized with lysosomal-associated membrane protein 1 (Lamp1) and Aß. Importatly, URMC-099 restored synaptic integrity and hippocampal neurogenesis in APP/PS1 mice. CONCLUSIONS: URMC-099 facilitates Aß clearance in the brain of APP/PS1 mice. The multifaceted immune modulatory and neuroprotective roles of URMC-099 make it an attractive candidate for ameliorating the course of AD. This is buttressed by removal of pathologic Aß species and restoration of the brain's microenvironment during disease.

Granulocyte-macrophage colony-stimulating factor neuroprotective activities in Alzheimer's disease mice.

We investigated the effects of granulocyte-macrophage colony stimulating factor (GM-CSF) on behavioral and pathological outcomes in Alzheimer's disease (AD) and non-transgenic mice. GM-CSF treatment in AD mice reduced brain amyloidosis, increased plasma Aß, and rescued cognitive impairment with increased hippocampal expression of calbindin and synaptophysin and increased levels of doublecortin-positive cells in the dentate gyrus. These data extend GM-CSF pleiotropic neuroprotection mechanisms in AD and include regulatory T cell-mediated immunomodulation of microglial function, Aß clearance, maintenance of synaptic integrity, and induction of neurogenesis. Together these data support further development of GM-CSF as a neuroprotective agent for AD.

Cathepsin B Improves ß-Amyloidosis and Learning and Memory in Models of Alzheimer's Disease.

Amyloid-ß (Aß) precursor protein (APP) metabolism engages neuronal endolysosomal pathways for Aß processing and secretion. In Alzheimer's disease (AD), dysregulation of APP leads to excess Aß and neuronal dysfunction; suggesting that neuronal APP/Aß trafficking can be targeted for therapeutic gain. Cathepsin B (CatB) is a lysosomal cysteine protease that can lower Aß levels. However, whether CatB-modulation of Aß improves learning and memory function deficits in AD is not known. To this end, progenitor neurons were infected with recombinant adenovirus expressing CatB and recovered cell lysates subjected to proteomic analyses. The results demonstrated Lamp1 deregulation and linkages between CatB and the neuronal phagosome network. Hippocampal injections of adeno-associated virus expressing CatB reduced Aß levels, increased Lamp1 and improved learning and memory. The findings were associated with the emergence of c-fos + cells. The results support the idea that CatB can speed Aß metabolism through lysosomal pathways and as such reduce AD-associated memory deficits.

The mixed-lineage kinase 3 inhibitor URMC-099 facilitates microglial amyloid-ß degradation.

BACKGROUND: Amyloid-ß (Aß)-stimulated microglial inflammatory responses engage mitogen-activated protein kinase (MAPK) pathways in Alzheimer's disease (AD). Mixed-lineage kinases (MLKs) regulate upstream MAPK signaling that include p38 MAPK and c-Jun amino-terminal kinase (JNK). However, whether MLK-MAPK pathways affect Aß-mediated neuroinflammation is unknown. To this end, we investigated if URMC-099, a brain-penetrant small-molecule MLK type 3 inhibitor, can modulate Aß trafficking and processing required for generating AD-associated microglial inflammatory responses. METHODS: Aß1-42 (Aß42) and/or URMC-099-treated murine microglia were investigated for phosphorylated mitogen-activated protein kinase kinase (MKK)3, MKK4 (p-MKK3, p-MKK4), p38 (p-p38), and JNK (p-JNK). These pathways were studied in tandem with the expression of the pro-inflammatory cytokines interleukin (IL)-1ß, IL-6, and tumor necrosis factor (TNF)-α. Gene expression of the anti-inflammatory cytokines, IL-4 and IL-13, was evaluated by real-time quantitative polymerase chain reaction. Aß uptake and expression of scavenger receptors were measured. Protein trafficking was assessed by measures of endolysosomal markers using confocal microscopy. RESULTS: Aß42-mediated microglial activation pathways were shown by phosphorylation of MKK3, MKK4, p38, and JNK and by expression of IL-1ß, IL-6, and TNF-α. URMC-099 modulated microglial inflammatory responses with induction of IL-4 and IL-13. Phagocytosis of Aß42 was facilitated by URMC-099 with up-regulation of scavenger receptors. Co-localization of Aß and endolysosomal markers associated with enhanced Aß42 degradation was observed. CONCLUSIONS: URMC-099 reduced microglial inflammatory responses and facilitated phagolysosomal trafficking with associated Aß degradation. These data demonstrate a new immunomodulatory role for URMC-099 to inhibit MLK and to induce microglial anti-inflammatory responses. Thus, URMC-099 may be developed further as a novel disease-modifying AD therapy.

Presenilin-1 familial Alzheimer's disease mutation alters hippocampal neurogenesis and memory function in CCL2 null mice.

Aberrations in hippocampal neurogenesis are associated with learning and memory, synaptic plasticity and neurodegeneration in Alzheimer's disease (AD). However, the linkage between them, ß-amyloidosis and neuroinflammation is not well understood. To this end, we generated a mouse overexpressing familial AD (FAD) mutant human presenilin-1 (PS1) crossed with a knockout (KO) of the CC-chemokine ligand 2 (CCL2) gene. The PS1/CCL2KO mice developed robust age-dependent deficits in hippocampal neurogenesis associated with impairments in learning and memory, synaptic plasticity and long-term potentiation. Neurogliogenesis gene profiling supported ß-amyloid independent pathways for FAD-associated deficits in hippocampal neurogenesis. We conclude that these PS1/CCL2KO mice are suitable for studies linking host genetics, immunity and hippocampal function.

A new quantitative rating scale for dyskinesia in nonhuman primates.

The aim of this study was to develop a quantitative scale to assess levodopa-induced dyskinesias (LIDs) in nonhuman primates using a video-based scoring system [Quantitative Dyskinesia Scale (QDS)]. Six macaques with stable Parkinsonism and LID were used for tests of the new QDS, in comparison with our current standardized scale (Drug-Related Side effects), which provides a classic subjective measurement of dyskinesia. QDS scoring is based on systematic movement counts in time frames, using videotape recordings. For both scales, body segments scored included each extremity, the trunk, the neck, and the face, and raters were blinded to L-dopa treatments. Comparison of the two scales revealed that their scores are highly correlated with and are parallel to the L-dopa pharmacokinetic profile, although the QDS provided significantly more quantifiable measurements. This remained the case after separating animals into groups of mild and severe dyskinesias. Inter-rater reliability for application of the QDS was confirmed from scores obtained by three examiners. We conclude that the QDS is a quantitative tool for reliably scoring LID in parkinsonian monkeys at all levels of severity of dyskinesia. The application of this new standard for scoring LID in primates will allow for more precise measurements of the effects of experimental treatments and will improve the quality of results obtained in translational studies.