Juliane Schelle

Persistence of Aβ seeds in APP null mouse brain.

Cerebral β-amyloidosis is induced by inoculation of Aβ seeds into APP transgenic mice, but not into App−/− (APP null) mice. We found that brain extracts from APP null mice that had been inoculated with Aβ seeds up to 6 months previously still induced β-amyloidosis in APP transgenic hosts following secondary transmission. Thus, Aβ seeds can persist in the brain for months, and they regain propagative and pathogenic activity in the presence of host Aβ.

Pharmacological BACE1 and BACE2 inhibition induces hair depigmentation by inhibiting PMEL17 processing in mice

Melanocytes of the hair follicle produce melanin and are essential in determining the differences in hair color. Pigment cell-specific MELanocyte Protein (PMEL17) plays a crucial role in melanogenesis. One of the critical steps is the amyloid-like functional oligomerization of PMEL17. Beta Site APP Cleaving Enzyme-2 (BACE2) and γ-secretase have been shown to be key players in generating the proteolytic fragments of PMEL17. The β-secretase (BACE1) is responsible for the generation of amyloid-β (Aβ) fragments in the brain and is therefore proposed as a therapeutic target for Alzheimer’s disease (AD). Currently BACE1 inhibitors, most of which lack selectivity over BACE2, have demonstrated efficacious reduction of amyloid-β peptides in animals and the CSF of humans. BACE2 knock-out mice have a deficiency in PMEL17 proteolytic processing leading to impaired melanin storage and hair depigmentation. Here, we confirm BACE2-mediated inhibition of PMEL17 proteolytic processing in vitro in mouse and human melanocytes. Furthermore, we show that wildtype as well as bace2+/− and bace2−/− mice treated with a potent dual BACE1/BACE2 inhibitor NB-360 display dose-dependent appearance of irreversibly depigmented hair. Retinal pigmented epithelium showed no morphological changes. Our data demonstrates that BACE2 as well as additional BACE1 inhibition affects melanosome maturation and induces hair depigmentation in mice.

Increased CSF Aβ during the very early phase of cerebral Aβ deposition in mouse models

Abnormalities in brains of Alzheimers disease (AD) patients are thought to start long before the first clinical symptoms emerge. The identification of affected individuals at this ‘preclinical AD’ stage relies on biomarkers such as decreased levels of the amyloid‐β peptide (Aβ) in the cerebrospinal fluid (CSF) and positive amyloid positron emission tomography scans. However, there is little information on the longitudinal dynamics of CSF biomarkers, especially in the earliest disease stages when therapeutic interventions are likely most effective. To this end, we have studied CSF Aβ changes in three Aβ precursor protein transgenic mouse models, focusing our analysis on the initial Aβ deposition, which differs significantly among the models studied. Remarkably, while we confirmed the CSF Aβ decrease during the extended course of brain Aβ deposition, a 20–30% increase in CSF Aβ40 and Aβ42 was found around the time of the first Aβ plaque appearance in all models. The biphasic nature of this observed biomarker changes stresses the need for longitudinal biomarker studies in the clinical setting and the search for new ‘preclinical AD’ biomarkers at even earlier disease stages, by using both mice and human samples. Ultimately, our findings may open new perspectives in identifying subjects at risk for AD significantly earlier, and in improving the stratification of patients for preventive treatment strategies.

The Prion-Like Properties of Amyloid-β Assemblies: Implications for Alzheimer's Disease

Since the discovery that prion diseases can be transmitted to experimental animals by inoculation with afflicted brain matter, researchers have speculated that the brains of patients suffering from other neurodegenerative diseases might also harbor causative agents with transmissible properties. Foremost among these disorders is Alzheimers disease (AD), the most common cause of dementia in the elderly. A growing body of research supports the concept that the pathogenesis of AD is initiated and sustained by the endogenous, seeded misfolding and aggregation of the protein fragment amyloid-β (Aβ). At the molecular level, this mechanism of nucleated protein self-assembly is virtually identical to that of prions consisting of the prion protein (PrP). The formation, propagation, and spread of Aβ seeds within the brain can thus be considered a fundamental feature of AD pathogenesis.

Aβ seeding potency peaks in the early stages of cerebral β‐amyloidosis

Little is known about the extent to which pathogenic factors drive the development of Alzheimers disease (AD) at different stages of the long preclinical and clinical phases. Given that the aggregation of the β‐amyloid peptide (Aβ) is an important factor in AD pathogenesis, we asked whether Aβ seeds from brain extracts of mice at different stages of amyloid deposition differ in their biological activity. Specifically, we assessed the effect of age on Aβ seeding activity in two mouse models of cerebral Aβ amyloidosis (APPPS1 and APP23) with different ages of onset and rates of progression of Aβ deposition. Brain extracts from these mice were serially diluted and inoculated into host mice. Strikingly, the seeding activity (seeding dose SD50) in extracts from donor mice of both models reached a plateau relatively early in the amyloidogenic process. When normalized to total brain Aβ, the resulting specific seeding activity sharply peaked at the initial phase of Aβ deposition, which in turn is characterized by a temporary several‐fold increase in the Aβ42/Aβ40 ratio. At all stages, the specific seeding activity of the APPPS1 extract was higher compared to that of APP23 brain extract, consistent with a more important contribution of Aβ42 than Aβ40 to seed activity. Our findings indicate that the Aβ seeding potency is greatest early in the pathogenic cascade and diminishes as Aβ increasingly accumulates in brain. The present results provide experimental support for directing anti‐Aβ therapeutics to the earliest stage of the pathogenic cascade, preferably before the onset of amyloid deposition.

Prevention of tau increase in cerebrospinal fluid of APP transgenic mice suggests downstream effect of BACE1 inhibition

The inhibition of the β‐site amyloid precursor protein‐cleaving enzyme 1 (BACE1) is a main therapeutic approach for the treatment of Alzheimers disease (AD). We previously reported an age‐related increase of tau protein in the cerebrospinal fluid (CSF) of amyloid β (Aβ) precursor protein (APP) transgenic mice.

NEUROFILAMENT LIGHT LEVELS IN THE CEREBROSPINAL FLUID ARE UNCOUPLED OF AMYLOIDOSIS IN LATE STAGE PATHOGENESIS OF APP TRANSGENIC MICE

P1-115 INVIVO PROPAGATIONOFSYNTHETICAb CONFORMATIONAL STRAINS Rodrigo Morales, Carlos Kramm-Barria, Ruben GomezGutierrez, Robert Tycko, Claudio Soto, The University of Texas Health Science Center at Houston, Houston, TX, USA; Dept. Neurology, The University of Texas Health Science Center at Houston, Houston, TX, USA; National Institutes of Health, Bethesda, MD, USA. Contact e-mail: rodrigo.moralesloyola@uth. tmc.edu

PREVENTION OF CEREBRAL AMYLOID ANGIOPATHY IN A MOUSE MODEL OF HEREDITARY CEREBRAL HEMORRAGE WITH AMYLOIDOSIS-DUTCH TYPE

protofibrils were readily taken up in human AD-derived microglia in an FcgR-independent manner, while BAN2401 facilitated further uptake of protofibrils in FcgR-mediated fashion. BAN2401 facilitated protofibril uptake in EOC-20 microglia cells was blocked with an FcgR blocker or F(ab’)2 fragments (EC501⁄4257661 ng/mL; corrected for background uptake). Conclusions:Ab can be taken up by microglia via a non-specific pattern-recognizing receptor pathway or through an FcgR-mediated process. Previous work has demonstrated that BAN2401 facilitates protofibril clearance, and this clearance was inhibited by an FcgR blocker to levels lower than those seen in the absence of BAN2401, suggesting a shift from non-specific uptake toward microglial activation and FcgR-dependent microglial phagocytosis. The present studies isolate the FcgR-dependent component of BAN2401-mediated protofibril uptake and provide further support for antibody dependent cellular phagocytosis of Ab protofibrils as the primary mechanism of action for BAN2401.

SUPPRESSION OF TAU INCREASE IN CEREBROSPINAL FLUID OF APP TRANSGENIC MICE PROVIDES EVIDENCE FOR DOWNSTREAM EFFECT OF BACE1 INHIBITION

SAMP8/SAMR1. Results: The level of cortical p-tau396 of 2months-old SAMP8 was significantly lower than SAMR1, while p-tau262 level of 2-months-old SAMP8 was comparable, and these levels increased upto 10 months-old SAMP8. The pattern of pGSK3b expression showed a negative correlation with p-tau396. In the hippocampus, p-tau396 and p-tau262 were observed to increase as SAMP8 aged, as compared to the levels of age-matched SAMR1. The early activation of cortical AMPK of 2-months-old SAMP8 disappeared in 5-monthsand 10-months-old mice, as compared to SAMR1; however, we could not observed the activation of AMPK in the hippocampus of SAMP8 across all ages. The level of Sirt1 in the cortex of 2-months-old SAMP8 was significantly lower than SAMR1, which persisted until 5-months-old and disappeared at 10-months-old, compared to the levels of SAMR1. The level of insulin receptor substrate-1 (IRS-1) expression in the cortex of SAMP8 was significantly lower than SAMR1 across all ages. In the hippocampus, the level of Sirt1, but not IRS-1, was lower in 2-months-old SAMP8 than SAMR1; however, the levels of Sirt1 in 10-months-old SAMP8 were comparable. However, when we treated metformin to activate AMPK, the AMPK-mediated regulation of p-tau396 in SAMP8 was not clear. Conclusions: Our data showed that the differential and possibly indirect regulation of tau phosphorylation by AMPK in cerebral cortex of SAMP8 ADmodel. However, further investigations to clarify precise mechanism of AMPK in tau phosphorylation and energy metabolism in the cerebral cortex are needed.