Homer2 and Homer3 interact with amyloid precursor protein and inhibit Abeta production

The study of Amyloid Precursor Protein (APP) processing has been the focus of considerable interest, since it leads to Aβ peptide generation, the main constituent of neuritic plaques found in brains of Alzheimer's disease patients. Therefore, the identification of novel APP binding partners that regulate Aβ peptide production represents a pharmaceutical target aiming at reducing Αβ pathology. In this study, we provide evidence that Homer2 and Homer3 but not Homer1 proteins interact specifically with APP. Their expression inhibits APP processing and reduces secretion of Aβ peptides. In addition, they decrease the levels of cell surface APP and inhibit maturation of APP and β-secretase (BACE1). The effects of Homer2 and Homer3 on APP trafficking to the cell surface and/or on APP and BACE1 maturation could be part of the mechanism by which the expression of these proteins leads to the significant reduction of Aβ peptide production.     

JNK regulates APP cleavage and degradation in a model of Alzheimer's disease

Secretion of Amyloid-beta peptide (Aβ) circulating oligomers and their aggregate forms derived by processing of beta-amyloid precursor protein (APP) are a key event in Alzheimer's disease (AD).We show that phosphorylation of APP on threonine 668 may play a role in APP metabolism in H4-APP^sw cell line, a degenerative AD model. We proved that JNK plays a fundamental role in this phosphorylation since its specific inhibition, with the JNK inhibitor peptide (D-JNKI1), induced APP degradation and prevented APP phosphorylation at T668. This results in a significant drop of βAPPs, Aβ fragments and Aβ circulating oligomers. Moreover the D-JNKI1 treatment produced a switch in the APP metabolism, since the peptide reduced the rate of the amyloidogenic processing in favour of the non-amyloidogenic one. All together our results suggest an important link between APP metabolism and the JNK pathway and contribute to shed light on the molecular signalling pathway of this disease indicating JNK as an innovative target for AD therapy.     

Vascular endothelial growth factor (VEGF) affects processing of amyloid precursor protein and β-amyloidogenesis in brain slice cultures derived from transgenic Tg2576 mouse brain

The up-regulation of the angiogenic vascular endothelial growth factor (VEGF) in brains of Alzheimer patients in close relationship to β-amyloid (Aβ) plaques, suggests a link of VEGF action and processing of the amyloid precursor protein (APP). To reveal whether VEGF may affect APP processing, brain slices derived from 17-month-old transgenic Tg2576 mice were exposed with 1 ng/ml VEGF for 6, 24, and 72 h, followed by assessing cytosolic and membrane-bound APP expression, level of both soluble and fibrillar Aβ-peptides, as well as activities of α- and β-secretases in brain slice tissue preparations.Treatment of brain slices with VEGF did not significantly affect the expression level of APP, regardless of the exposure time studied. In contrast, VEGF exposure of brain slices for 6 h reduced the formation of soluble, SDS extractable Aβ(1–40) and Aβ(1–42) as compared to brain slice cultures incubated in the absence of any drug, while the fibrillar Aβ peptides did not change significantly. This effect was less pronounced 24 h after VEGF exposure, but was no longer detectable when brain slices were exposed by VEGF for 72 h, which indicates an adaptive response to chronic VEGF exposure. The VEGF-mediated reduction in Aβ formation was accompanied by a transient decrease in β-secretase activity peaking 6 h after VEGF exposure. To reveal whether the VEGF-induced changes in soluble Aβ-level may be due to actions of VEGF on Aβ fibrillogenesis, the fibrillar status of Aβ was examined using the thioflavin-T binding assay. Incubation of Aβ preparations obtained from Tg2576 mouse brain cortex, in the presence of VEGF slightly decreased the fibrillar content with increasing incubation time up to 72 h. The data demonstrate that VEGF may affect APP processing, at least in vitro, suggesting a role of VEGF in the pathogenesis of Alzheimer's disease.     

The role of the low-density lipoprotein receptor-related protein (LRP1) in Alzheimer's A beta generation: development of a cell-based model system

The clearance and degradation of extracellular Aβ is critical for regulating β-amyloid deposition, a major hallmark of brains of patients with Aβ in Alzheimer’s Disease. The low-density lipoprotein receptor-related protein, LRP1, is a large endocytic receptor that significantly contributes to the balance between degradation and production of Aβ. An extracellular portion of the LRP, known as the cluster II region can bind to the secreted form of APP (sAPP-KPI). We show here that a GST fusion protein containing the cluster II region of LRP can be used as a ‘mini-receptor’ that specifically binds to sAPP-KPI from conditioned cultured medium. The binding between the GST-LRP-cluster II fusion protein and sAPP-KPI can be inhibited with the strong binding ligand of LRP1, called receptor-associated protein (RAP). Furthermore, a cell-based in vitro assay system has been developed to monitor the production of total Aβ and Aβ_1–42 in the presence and absence of RAP in Chinese hamster ovary (CHO) cell lines both deficient in LRP and expressing LRP. A 3-day treatment of the L2 (CHO cells deficient in LRP and overexpressing APP751) and L3 (CHO cells expressing LRP and overexpressing APP751) cell lines with RAP showed a decrease in total Aβ and, interestingly, also a decrease in the ratio of Aβ₄₂/Aβ_total. This cell-based model system and LRP-cluster II mini-receptor will be very useful for screening novel compounds that can reduce Aβ accumulation by inhibiting binding of APP-KPI to LRP1.     

FAD Mutations in Presenilin-1 or Amyloid Precursor Protein Decrease the Efficacy of a γ-Secretase Inhibitor: Evidence for Direct Involvement of PS1 in the γ-Secretase Cleavage Complex

To investigate the mechanism of regulation of Aß production by familial Alzheimer's disease (FAD)-linked presenilin 1 (PS1), we used a cell-free system that allows de novo Aß generation to examine whether PS1 participates directly in the γ-secretase reaction. Optimal Aß generation in vitro was achieved at mildly acidic pH and could be inhibited by the aspartyl protease inhibitor pepstatin A, consistent with the suggestion that γ-secretase is an aspartyl protease. Dominant negative mutations of the critical transmembrane aspartates in PS1 or full deletion of PS1 did not alter the maturation of APP in the secretory pathway. Instead, PS1 had a direct effect on the inhibition of Aß production by a designed peptidomimetic inhibitor: the inhibition was significantly less effective in cells expressing FAD-causing mutations in either APP or PS1 than in cells expressing the wild-type proteins. Taken together, these findings suggest that PS1 participates physically in a complex with APP during the γ-secretase cleavage event.     

X11α haploinsufficiency enhances Aβ amyloid deposition in Alzheimer's disease transgenic mice

The neuronal adaptor protein X11α/mint-1/APBA-1 binds to the cytoplasmic domain of the amyloid precursor protein (APP) to modulate its trafficking and metabolism. We investigated the consequences of reducing X11α in a mouse model of Alzheimer's disease (AD). We crossed hAPPswe/PS-1ΔE9 transgenic (AD tg) mice with X11α heterozygous knockout mice in which X11α expression is reduced by approximately 50%. The APP C-terminal fragments C99 and C83, as well as soluble Aβ40 and Aβ42, were increased significantly in brain of X11α haploinsufficient mice. Aβ/amyloid plaque burden also increased significantly in the hippocampus and cortex of one year old AD tg/X11α (+/−) mice compared to AD tg mice. In contrast, the levels of sAPPα and sAPPβ were not altered significantly in AD tg/X11α (+/−) mice. The increased neuropathological indices of AD in mice expressing reduced X11α suggest a normal suppressor role for X11α on CNS Aβ/amyloid deposition.     

Amyloid beta, mitochondrial structural and functional dynamics in Alzheimer's disease

Mitochondria are the major source of energy for the normal functioning of brain cells. Increasing evidence suggests that the amyloid precursor protein (APP) and amyloid beta (Aβ) accumulate in mitochondrial membranes, cause mitochondrial structural and functional damage, and prevent neurons from functioning normally. Oligomeric Aβ is reported to induce intracellular Ca²⁺ levels and to promote the excess accumulation of intracellular Ca²⁺ into mitochondria, to induce the mitochondrial permeability transition pore to open, and to damage mitochondrial structure. Based on recent gene expression studies of APP transgenic mice and AD postmortem brains, and APP/Aβ and mitochondrial structural studies, we propose that the overexpression of APP and the increased production of Aβ may cause structural changes of mitochondria, including an increase in the production of defective mitochondria, a decrease in mitochondrial trafficking, and the alteration of mitochondrial dynamics in neurons affected by AD. This article discusses some critical issues of APP/Aβ associated with mitochondria, mitochondrial structural and functional damage, and abnormal intracellular calcium regulation in neurons from AD patients. This article also discusses the link between Aβ and impaired mitochondrial dynamics in AD.     

Presenilin/γ-Secretase Activity Is Located in Acidic Compartments of Live Neurons

Presenilin (PSEN)/γ-secretase is a protease complex responsible for the proteolytic processing of numerous substrates. These substrates include the amyloid precursor protein (APP), the cleavage of which by γ-secretase results in the production of β-amyloid (Aβ) peptides. However, exactly where within the neuron γ-secretase processes APP C99 to generate Aβ and APP intracellular domain (AICD) is still not fully understood. Here, we employ novel Förster resonance energy transfer (FRET)-based multiplexed imaging assays to directly “visualize” the subcellular compartment(s) in which γ-secretase primarily cleaves C99 in mouse cortex primary neurons (from both male and female embryos). Our results demonstrate that γ-secretase processes C99 mainly in LysoTracker-positive low-pH compartments. Using a new immunostaining protocol which distinguishes Aβ from C99, we also show that intracellular Aβ is significantly accumulated in the same subcellular loci. Furthermore, we found functional correlation between the endo-lysosomal pH and cellular γ-secretase activity. Taken together, our findings are consistent with Aβ being produced from C99 by γ-secretase within acidic compartments such as lysosomes and late endosomes in living neurons.SIGNIFICANCE STATEMENT Alzheimer''s disease (AD) genetics and histopathology highlight the importance of amyloid precursor protein (APP) processing by γ-secretase in pathogenesis. For the first time, this study has enabled us to directly “visualize” that γ-secretase processes C99 mainly in acidic compartments such as late endosomes and lysosomes in live neurons. Furthermore, we uncovered that intracellular β-amyloid (Aβ) is significantly accumulated in the same subcellular loci. Emerging evidence proposes the great importance of the endo-lysosomal pathway in mechanisms of misfolded proteins propagation (e.g., Tau, α-Syn). Therefore, the predominant processing of C99 and enrichment of Aβ in late endosomes and lysosomes may be critical events in the molecular cascade leading to AD.     

Potentially amyloidogenic fragment of 50 kDa and intracellular processing of amyloid precursor protein in cells cultured under leupeptin

The principal neuropathological feature of Alzheimer's disease is extracellular deposition of 4-kDa proteinous fragment, designated as β-amyloid peptides (β/A4 peptides) derived by proteolytic cleavage from amyloid precursor protein (APP), a large cell-surface receptor-like protein. There has been evidence that APP is proteolytically degraded in the secretory and endosomal/lysosomal pathways. The pathway in which APP is cleaved to generate β/A4 peptides is still not identified. To clarify the intracellular processing of APP into the generation of β/A4 peptides, we detected and characterized potentially amyloidogenic or non-amyloidogenic fragments using newly established monoclonal and polyclonal antibodies in the cultured cells with or without leupeptin, potent lysosomal protease inhibitor of lysosome. APP fragments of 50 and 20 kDa containing full-length β/A4 peptides were identified in the cultured cells. Immunoblot analysis, biochemical study for specific marker enzyme activity of the fractions obtained from subcellular fractionation, sucrose density gradient centrifugation indicated that the 50-kDa APP fragment was produced in the compartment closely related to endosomal/lysosomal system. Our data suggest that the endosomal/lysosomal pathway is involved in the processing and generation of β/A4 peptides.     

Residues at P2-P1 positions of - and ζ-cleavage sites are important in formation of β-amyloid peptide

Most of the Alzheimer's disease (AD)-linked mutations in amyloid precursor protein (APP), which cause abnormal production of β-amyloid (Aβ), are localized at the major β-secretase-and γ-secretase cleavage sites. In this study, using an APP-knockout mouse neuronal cell line, our data demonstrated that at the P2-P1 positions of the -cleavage site at Aβ49 and the ζ-cleavage site at Aβ46, aromatic amino acids caused a strong reduction in total Aβ. On the other hand, residues with a long side chain caused a decrease in Aβ₄₀ and a concomitant increase in Aβ₄₂ and Aβ₃₈. These findings indicate that the structures of the substituting residues at these key positions strongly determine the efficiency and preference of γ-secretase-mediated APP processing, which determines the ratio of different secreted Aβ species, a crucial factor in the disease development. Our findings provide new insight into the mechanisms of γ-secretase-mediated APP processing and, specifically, into why most AD-linked APP mutations are localized at major γ-secretase cleavage sites. This information may contribute to the development of methods of prevention and treatment of Alzheimer's disease aimed at modulating γ-secretase activity.     

Co-expression of β-amyloid precursor protein (βAPP) and apolipoprotein E in cell culture: analysis of βAPP processing

Apolipoprotein E (ApoE) is the major genetic risk factor for Alzheimer's disease (AD). The ApoE4 allele is associated with earlier disease onset and greater cerebral deposition of the amyloid beta peptide (Aβ), the major constituent of senile (amyloid) plaques. The molecular mechanism underlying these effects of ApoE4 remains unclear; ApoE alleles could have different influences on Aβ production, extracellular aggregation, or clearance. Because the missense mutations on chromosomes 14 and 21 that cause familial forms of AD appear to lead to increased secretion of Aβ, it is important to determine whether ApoE4 has a similar effect. Here, we have examined the effects of all three ApoE alleles on the processing of βAPP and the secretion of Aβ in intact cells. We established neural (HS683 human glioma) and non-neural (Chinese hamster ovary) cell culture systems that constitutively secrete both ApoE and Aβ at concentrations like those in human cerebrospinal fluid. βAPP metabolites, generated in the presence of each ApoE allele, were analysed and quantified by two methods: immunoprecipitation and phosphorimaging, and ELISA. We detected no consistent allele-specific effects of ApoE on βAPP processing in either cell type. Our data suggest that the higher amyloid burden found in AD subjects expressing ApoE4 is not due to increased amyloidogenic processing of βAPP, in contrast to findings in AD linked to chromosome 14 or 21. These co-expressing cell lines will be useful in the further search for the effects of ApoE on Aβ aggregation or clearance under physiologically relevant conditions.     

Distribution of β-amyloid and amyloid precursor protein in the brain of spawning (senescent) salmon: a natural, brain-aging model

Brain amyloid precursor protein (APP), a normal constituent of neurons, glial cells and cerebrospinal fluid, has several proposed functions (e.g., in neuronal growth and survival). It appears, however, that altered processing of APP is an initial or downstream step in the neuropathology of brain aging, Alzheimer's disease (AD), and Down's syndrome (DS). Some studies suggest that proteolytic cleavage of APP, producing β-amyloid (Aβ_1–42), could have neurotoxic or neuroprotective effects. In this study, we utilized antibodies to human APP₆₉₅ and Aβ_1–42, and Congo red staining, to search for amyloid deposition in the brain of semelparous spawning kokanee salmon (Oncorhynchus nerka kennerlyi). Intracellular APP₆₉₅ immunoreactivity (APP-ir) was observed in brain regions involved in gustation (glomerulosus complex), olfaction (putative hippocampus, olfactory bulb), vision (optic tectum), the stress response (nucleus preopticus and nucleus lateralis tuberis), reproductive behavior (nucleus preopticus magnocellularis, nucleus preopticus periventricularis, ventral telencephalon), and coordination (cerebellum). Intra- and extra-neuronal Aβ_1–42 immunoreactivity (Aβ-ir) were present in all APP-ir regions except the nucleus lateralis tuberis and Purkinje cells of the cerebellum (coordination). Thus, the relationship between APP and Aβ deposition during brain aging could shed light on the processing of APP into Aβ, neurodegeneration, and possible protection of neurons that are functioning in spawning but senescent salmon. Pacific salmon, with their predictable and synchronized life history, could provide research options not available with the existing models for studies of brain aging and amyloidosis.     

Advances in the cellular and molecular biology of the beta-amyloid protein in Alzheimer's disease

Alzheimer’s disease (AD) is a progressive senile dementia characterized by deposition of a 4 kDa peptide of 39–42 residues known as amyloid beta-peptide (Aβ) in the form of senile plaques and the microtubule associated protein tau as paired helical filaments. Genetic studies have identified mutations in the Aβ precursor protein (APP) as the key triggers for the pathogenesis of AD. Other genes such as presenilins 1 and 2 (PS1/2) and apolipoprotein E (APOE) also play a critical role in increased Aβ deposition. Several biochemical and molecular studies using transfected cells and transgenic animals point to mechanisms by which Aβ is generated and aggregated to trigger the neurodegeneration that may cause AD. Three important enzymes collectively known as “secretases” participate in APP processing. An enzymatic activity, β-secretase, cleaves APP on the amino side of Aβ producing a large secreted derivative, sAPPβ, and an Aβ-bearing membrane-associated C-terminal derivative, CTFβ, which is subsequently cleaved by the second activity, γ-secretase, to release Aβ. Alternatively, a third activity, α-secretase, cleaves APP within Aβ to the secreted derivative sAPPα and membrane-associated CTFα. The predominant secreted APP derivative is sAPPα in most cell-types. Most of the secreted Aβ is 40 residues long (Aβ40) although a small percentage is 42 residues in length (Aβ42). However, the longer Aβ42 aggregates more readily and was therefore considered to be the pathologically important form. Advances in our understanding of APP processing, trafficking, and turnover will pave the way for better drug discovery for the eventual treatment of AD. In addition, APP gene regulation and its interaction with other proteins may provide useful drug targets for AD. The emerging knowledge related to the normal function of APP will help in determining whether or not the AD associated changes in APP metabolism affect its function. The present review summarizes our current understanding of APP metabolism and function and their relationship to other proteins involved in AD.     

The beta amyloid peptide can act as a modular aggregation domain

Although there is compelling evidence that the β amyloid peptide (Aβ) can be centrally involved in Alzheimer's disease, the natural role (if any) of this peptide remains unclear. Here we use green fluorescent protein (GFP) fusions to demonstrate that the Aβ sequence, like prion domains, can act as a modular aggregation domain when terminally appended to a normally soluble protein. We find that a single amino acid substitution (Leu¹⁷ to Pro) in the β peptide sequence can abolish this cis capacity to induce aggregation. Introduction of this substitution into full-length APP (i.e., a Leu⁶¹³Pro substitution in APP695) alters the processing of APP leading to the accumulation of the C99 C-terminal fragment (CTF). We suggest that in at least some aggregation disease-related proteins the presence of an aggregation domain is not “accidental”, but reflects a selected role of these domains in modulating the trafficking or metabolism of the parental protein.     

S-Adenosylhomocysteine increases β-amyloid formation in BV-2 microglial cells by increased expressions of β-amyloid precursor protein and presenilin 1 and by hypomethylation of these gene promoters

S-Adenosylhomocysteine (SAH) has been implicated as a risk factor for neurodegenerative diseases such as Alzheimer's disease. As SAH is a potent inhibitor of all cellular methyltransferases, we herein examined the hypothesis that SAH may increase the formation of amyloid β-peptide (Aβ) in BV-2 mouse microglial cells through hypomethylation of presenilin 1 protein (PS1) and β-site amyloid precursor protein cleaving enzyme 1 (BACE1), both of which cleave Aβ precursor protein (APP) to form Aβ. The results showed that SAH increased Aβ protein formation in a concentration-dependent manner (10–500 nM), and this effect of SAH was accompanied by significantly increased expression of APP and PS1 proteins, although SAH only significantly increased the expression of BACE1 at the highest concentration used (500 nM). SAH (500 nM) markedly induced hypomethylation of APP and PS1 gene promoters. Incubation of cells with 5′-azc (20 μM), also an inhibitor of DNA methyltransferases enhanced Aβ protein expression and APP and PS1 gene promoters hypomethylation. By contrast, pre-incubation of cells with betaine (1.0 mM), 30 min followed by incubation with SAH (500 nM) or 5′-azc (20 μM) for 24 h markedly prevented the expression of Aβ protein (by 50%, P < 0.05) and the gene promoter hypomethylation of APP and PS1. Taken together, this study demonstrates that SAH increases the production of Aβ in BV-2 cells possibly by increased expression of APP and induction of hypomethylation of APP and PS1 gene promoters.     

The role of low-density receptor-related protein 1 (LRP1) as a competitive substrate of the amyloid precursor protein (APP) for BACE1

Cleavage of APP by BACE1 is the first proteolytic step in the production of amyloid-beta (Aβ), which accumulates in senile plaques in Alzheimer's disease. Through its interaction with APP, the low-density receptor-related protein 1 (LRP1) enhances APP internalization. Recently, BACE1 has been shown to interact with and cleave the light chain (lc) of LRP1. Since LRP1 is known to compete with APP for cleavage by gamma-secretase, we tested the hypothesis that LRP1 also acts as a competitive substrate for β-secretase. We found that the increase in secreted APP (sAPP) mediated by over-expression of BACE1 in APP-transfected cells could be decreased by simultaneous LRP1 over-expression. Analysis by multi-spot ELISA revealed that this is due to a decrease in sAPPβ, but not sAPPα. Interaction between APP and BACE1, as measured by immunoprecipitation and fluorescence lifetime assays, was impaired by LRP1 over-expression. We also demonstrate that APP over-expression leads to decreased LRP1 association with and cleavage by BACE1. In conclusion, our data suggest that - in addition to its role in APP trafficking - LRP1 affects APP processing by competing for cleavage by BACE1.     

Overexpression of wild-type human APP in mice causes cognitive deficits and pathological features unrelated to Aβ levels

Transgenic mice expressing mutant human amyloid precursor protein (APP) develop an age-dependent amyloid pathology and memory deficits, but no overt neuronal loss. Here, in mice overexpressing wild-type human APP (hAPP_wt) we found an early memory impairment, particularly in the water maze and to a lesser extent in the object recognition task, but β-amyloid peptide (Aβ₄₂) was barely detectable in the hippocampus. In these mice, hAPP processing was basically non-amyloidogenic, with high levels of APP carboxy-terminal fragments, C83 and APP intracellular domain. A tau pathology with an early increase in the levels of phosphorylated tau in the hippocampus, a likely consequence of enhanced ERK1/2 activation, was also observed. Furthermore, these mice presented a loss of synapse-associated proteins: PSD95, AMPA and NMDA receptor subunits and phosphorylated CaMKII. Importantly, signs of neurodegeneration were found in the hippocampal CA1 subfield and in the entorhinal cortex that were associated to a marked loss of MAP2 immunoreactivity. Conversely, in mice expressing mutant hAPP, high levels of Aβ₄₂ were found in the hippocampus, but no signs of neurodegeneration were apparent. The results support the notion of Aβ-independent pathogenic pathways in Alzheimer's disease.     

Amyloid precursor protein and membrane phospholipids in primary cortical neurons increase with development, or after exposure to nerve growth factor or Aβ1–40

We examined the relationships between membrane phospholipid levels, the secretion and expression of the amyloid precursor protein (APP), and the responses of both to nerve growth factor (NGF), Aβ_1–40 or Aβ_40–1 in developing cortical neurons cultured from rat embryos. Neuronal membrane phospholipid levels per cell, and phosphatidylcholine, phosphatidylserine, phosphatidylinositol and phosphatidylethanolamine increased individually between the first and seventh days of culturing. The amounts of APP holoprotein and APP mRNAs in the cells, as well as the amounts of soluble APP (APPs) secreted by them, also increased during neuronal development in vitro. The increases in APPs exceeded the increases in APP which, in turn, exceed those in phospholipid levels. The levels of APP holoprotein, but not of phospholipids, increased when neurons were grown in a choline-free medium, suggesting that increases in APP are not sufficient to stimulate changes in membrane phospholipids. Treatment of neuron cultures for four days with NGF or Aβ_1–40, but not with Aβ_40–1, dose-dependently increased membrane phospholipids, tau and GAP-43, as well as APP holoprotein and secreted APPs. These results indicate that agents, like NGF or Aβ_1–40, which enhance membrane phospholipid levels may promote neurite formation, APP expression and APPs secretion in primary neuronal cultures.     

Functional role of lipoprotein receptors in Alzheimer's disease

The LDL receptor gene family constitutes a class of structurally closely related cell surface receptors fulfilling diverse functions in different organs, tissues, and cell types. The LDL receptor is the prototype of this family, which also includes the VLDLR, ApoER2/LRP8, LRP1 and LRP1B, as well as Megalin/GP330, SorLA/LR11, LRP5, LRP6 and MEGF7. Recently several lines of evidence have positioned the LDL receptor gene family as one of the key players in Alzheimer's disease (AD) research. Initially this receptor family was of high interest due to its key function in cholesterol/apolipoprotein E (ApoE) uptake, with the epsilon4 allele of ApoE as the strongest genetic risk factor for late-onset AD. It has been established that the cholesterol metabolism of the cell has a strong impact on the production of Abeta, the major component of the plaques found in the brain of AD-patients. The original report that soluble amyloid precursor protein (APP) containing the kunitz proteinase inhibitor (KPI) domain might act as a ligand for LRP1 led to a complex investigation of the interaction of both proteins and their potential function in AD development. Meanwhile, it has been demonstrated that LRP1 might bind to APP independent of the KPI domain in APP. This APP - LRP1 interaction is facilitated through a trimeric complex of APP-FE65-LRP1, which has a functional role in APP processing. Along with LRP1, APP is transported from the early secretory compartments to the cell surface and subsequently internalised into the endosomal / lysosomal compartments. Recent investigations indicate that ApoER2 and SorLA fulfil a similar role in shifting APP localisation in the cell, which affects APP processing and the production of the APP derived amyloid beta-peptide (Abeta). In addition to the effect of lipoprotein receptors on APP processing and Abeta production, LRP1 has been shown to bind Abeta directly or indirectly through Abeta-lactoferrin, Abeta-alpha2M and Abeta-ApoE complexes in vitro and in vivo. Based on these observations two LRP1 mediated clearance mechanisms of Abeta are proposed to play a crucial role in the prevention of AD: either Abeta-uptake into a cell with its subsequent degradation or its transport out of the brain over the blood brain barrier into the periphery. Following this export Abeta is degraded in the liver, where LRP1 potentially conducts the removal of Abeta from the blood stream. Although the involvement of LDLR family members in AD is not yet fully understood it becomes clear that they can directly affect APP production, Abeta-clearance and Abeta-transport over the blood brain barrier.     

Discovery of a Novel Pharmacological and Structural Class of Gamma Secretase Modulators Derived from the Extract of Actaea racemosa

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