A Non-Canonical p75HER2 Signaling Pathway Underlying Trastuzumab Action and Resistance in Breast Cancer.

Overexpression of HER2 occurs in 25% of breast cancer. Targeting HER2 has proven to be an effective therapeutic strategy for HER2-positive breast cancer. While trastuzumab is the most commonly used HER2 targeting agent, which has significantly improved outcomes, the overall response rate is low. To develop novel therapies to boost trastuzumab efficacy, it is critical to identify the mechanisms underlying trastuzumab action and resistance. We recently showed that the inhibition of breast cancer cell growth by trastuzumab is not through the inhibition of HER2 canonical signaling. Here we report the identification of a novel non-canonical HER2 signaling pathway and its interference by trastuzumab. We showed that HER2 signaled through a non-canonical pathway, regulated intramembrane proteolysis (RIP). In this pathway, HER2 is first cleaved by metalloprotease ADAM10 to produce an extracellular domain (ECD) that is released and the p95HER2 that contains the transmembrane domain (TM) and intracellular domain (ICD). p95HER2, if further cleaved by an intramembrane protease, γ-secretase, produced a soluble ICD p75HER2 with nuclear localization signal (NLS). p75HER2 is phosphorylated and translocated to the nucleus. Nuclear p75HER2 promotes cell proliferation. Trastuzumab targets this non-canonical HER2 pathway via inhibition of the proteolytic cleavage of HER2 by both ADAM10 and γ-secretase. However, p75HER2 pathway also confers resistance to trastuzumab once aberrantly activated. Combination of trastuzumab with ADAM10 and γ-secretase inhibitors completely blocks p75HER2 production in both BT474 and SKBR3 cells. We concluded that HER2 signals through the RIP signaling pathway that promotes cell proliferation and is targeted by trastuzumab. The aberrant HER2 RIP signaling confers resistance to trastuzumab that could be overcome by the application of inhibitors to ADAM10 and γ-secretase.

Multi-modal neuroprotection of Argemone mexicana L. against Alzheimer's disease: In vitro and in silico study.

Argemone mexicana L. is a medicinal plant, but its impact on Alzheimer's disease (AD) is right now undetermined. We intended to investigate the in-vitro anti-AD potential of leaves and flowers of A. mexicana methanol, ethanol, and ethyl extracts and to identify multi-modal anti-AD phytochemicals by computational approaches. Molecular docking of 196 phytochemicals identified three hit phytochemicals (protoberberine, protopine, and codeine) with higher binding affinity and multi-targeting ability toward AChE, BChE, BACE-1, and GSK-3ß. Further MM-GBSA assays confirmed the integrity of these phytochemicals as the hit phytochemicals. However, these phytochemicals demonstrated favorable pharmacokinetics (PK) and drugable properties having no toxicity. Molecular dynamics simulations confirmed the binding strength of the hit phytoconstituents in the active pockets of AChE, BChE, BACE-1, and GSK-3ß with multi-targeting inhibitory activities. All the extracts exhibited dose-dependent antioxidant and anti-cholinesterase activities supporting the in silico results in the context of oxidative stress and cholinergic pathways. Our results offer scientific validation of the anti-AD properties of Argemone mexicana L. and identified protoberberine, protopine, and codeine that could be used for the development of multi-modal inhibitors of AChE, BChE, BACE-1, and GSK-3ß to combat AD. Additional in vivo validation is recommended to ensure a thorough assessment in the present research.

Truncated mini LRP1 transports cargo from luminal to basolateral side across the blood brain barrier.

BACKGROUND: The most crucial area to focus on when thinking of novel pathways for drug delivery into the CNS is the blood brain barrier (BBB). A number of nanoparticulate formulations have been shown in earlier research to target receptors at the BBB and transport therapeutics into the CNS. However, no mechanism for CNS entrance and movement throughout the CNS parenchyma has been proposed yet. Here, the truncated mini low-density lipoprotein receptor-related protein 1 mLRP1_DIV* was presented as blood to brain transport carrier, exemplified by antibodies and immunoliposomes using a systematic approach to screen the receptor and its ligands' route across endothelial cells in vitro. METHODS: The use of mLRP1_DIV* as liposomal carrier into the CNS was validated based on internalization and transport assays across an in vitro model of the BBB using hcMEC/D3 and bEnd.3 cells. Trafficking routes of mLRP1_DIV* and corresponding cargo across endothelial cells were analyzed using immunofluorescence. Modulation of γ-secretase activity by immunoliposomes loaded with the γ-secretase modulator BB25 was investigated in co-cultures of bEnd.3 mLRP1_DIV* cells and CHO cells overexpressing human amyloid precursor protein (APP) and presenilin 1 (PSEN1). RESULTS: We showed that while expressed in vitro, mLRP1_DIV* transports both, antibodies and functionalized immunoliposomes from luminal to basolateral side across an in vitro model of the BBB, followed by their mLRP1_DIV* dependent release of the cargo. Importantly, functionalized liposomes loaded with the γ-secretase modulator BB25 were demonstrated to effectively reduce toxic Aß42 peptide levels after mLRP1_DIV* mediated transport across a co-cultured endothelial monolayer. CONCLUSION: Together, the data strongly suggest mLRP1_DIV* as a promising tool for drug delivery into the CNS, as it allows a straight transport of cargo from luminal to abluminal side across an endothelial monolayer and it's release into brain parenchyma in vitro, where it exhibits its intended therapeutic effect.

Simple improvements in vector design afford substantial gains in AAV delivery of aggregation-slowing Aß variants.

Adeno-associated virus (AAV) gene therapy for neurological disease has gained traction due to stunning advances in capsid evolution for CNS targeting. With AAV brain delivery now in focus, conventional improvements in viral expression vectors offer a complementary route for optimizing gene delivery. We previously introduced a novel AAV gene therapy to slow amyloid aggregation in the brain based on neuronal release of an Aß sequence variant that inhibited fibrilization of wild-type Aß. Here we explore three coding elements of the virally delivered DNA plasmid in an effort to maximize the production of therapeutic peptide in the brain. We demonstrate that simply replacing the Gaussia luciferase signal peptide with the mouse immunoglobulin heavy chain signal peptide increased release of variant Aß by ∼5-fold. Sequence modifications within the expressed minigene further increased peptide release by promoting γ-secretase cleavage. Addition of a cytosolic fusion tag compatible with γ-secretase interaction allowed viral transduction to be tracked by immunostaining, independent from the variant Aß peptide. Collectively these construct modifications increased neuronal production of therapeutic peptide by 10-fold upon intracranial AAV injection of neonatal mice. These findings demonstrate that modest changes in expression vector design can yield substantial gains in AAV efficiency for therapeutic applications.

Inhibition of Amyloid ß Accumulation by Protease-Digested Whitebait (Shirasu) in a Murine Model of Alzheimer's Disease.

The number of people with dementia is increasing annually worldwide. Alzheimer's disease (AD), which accounts for the highest percentage of dementia-causing diseases, remains difficult to cure, and prevention of its onset is important. We aimed to discover new AD-preventive ingredients and investigate the inhibitory effects of ten different species of seafood digests prepared by protease treatment on ß-secretase 1 (BACE1) activity. Substantial inhibition of BACE1 activity was observed in five species of seafood, and protease-digested whitebait (WPD) showed the highest inhibitory effect among the ten marine samples. We further examined the potential of WPD as an AD preventive component using a familial AD strain (5xFAD) murine model. The intraperitoneal administration of WPD for 28 days substantially decreased the insoluble amyloid ß1-42 content and the expression of glial fibrillary acidic protein, a marker of astrogliosis, in the cerebral cortex of the 5xFAD mice. These results strongly suggest that WPD is a novel functional food-derived ingredient with preventive effects against AD.

Neutrophil extracellular traps activate Notch-γ-secretase signaling in hidradenitis suppurativa.

BACKGROUND: Hidradenitis suppurativa (HS) is an inflammatory chronic skin disorder of unknown etiology characterized by inflamed abscess-like nodules and boils resulting in sinus tract formation, tissue scarring, and massive infiltration of neutrophils. Multiple lines of evidence have highlighted the potential association between alterations in the Notch pathway and HS pathogenesis, but the mechanisms remain incompletely characterized. OBJECTIVE: We aimed to elucidate the role of neutrophil extracellular traps in Notch-γ-secretase signaling. METHODS: Twenty-six HS lesional tissues, primary HS macrophages, and skin fibroblasts were interrogated by quantitative PCR, Western blot, and ELISA analyses. γ-Secretase and TNF-α converting enzyme activities were measured in HS skin lesions, macrophages, and skin fibroblasts. Immunofluorescence and RNAscope analyses were performed in HS and control skin. RESULTS: A prominent presence of Notch ligands, Delta-like ligand 4 (DLL4), and Jagged (JAG) 2 were detected at the protein and mRNA levels in HS skin lesion compared to control. Levels of DLL4, JAG1, citrullinated histone H3 DNA, and γ-secretase activity correlated with HS disease severity. Additionally, significantly elevated levels of Notch ligands and γ-secretase activity were found in dissected sinus tracts compared to the rest of HS tissue. Immunofluorescence microscopy of HS skin lesions revealed activation of Notch-1 signaling in macrophages and skin fibroblasts. Neutrophil extracellular traps (NETs) purified from HS patients displayed elevated levels of DLL4. HS NETs activated the Notch pathway in macrophages and dermal fibroblasts isolated from HS patients. HS skin fibroblasts displayed elevated levels of CD90 and DPP4 in association with increased migratory capacity and Notch activation. Inhibition of Notch decreased migratory capacity and profibrotic markers in HS fibroblasts. CONCLUSION: These data support a pathogenic connection between NETs, Notch-γ-secretase activation, and the release of profibrotic molecules that promote dysregulation of macrophages and skin fibroblasts in HS. Unveiling the relevance of these molecular events not only expands our understanding of HS but also opens new venues for the development of targeted therapies to address the fibrotic complications of advanced stages of HS.

Lactate increases ADAM10 activity and reduces BACE1 activity in mouse brain.

The accumulation and aggregation of beta-amyloid (Aß) peptides contributes to neuronal dysfunction and death. These Aß peptides originate from a transmembrane protein known as amyloid precursor protein (APP), which can be processed via two competing pathways. Alpha-secretase (ADAM10) cleavage is thought to be neuroprotective while beta-secretase (BACE1) cleavage results in the production of Aß. Aerobic exercise reduces BACE1 activity, but the mechanisms involved are unknown though several exercise-induced mediators such as lactate may be involved. The current study examined whether systemic lactate can alter APP processing and BACE1 and ADAM10 activity. Mice were randomly assigned to one of four groups (n = 10 per group): (1) sedentary; (2) lactate-injection (1.0 g kg-1 body mass); (3) exercise; and (4) exercise and oxamate (lactate dehydrogenase inhibitor; 750 mg kg-1 body mass). Two hours following intervention, the hippocampus and prefrontal cortex (PFC) were collected. In the PFC lactate-injection and exercise resulted in higher ADAM10 activity compared to sedentary (exercise P = 0.0215, lactate P = 0.0038), in the hippocampus lactate-injection was higher compared to sedentary (lactate P = 0.011), and this was absent in the presence of oxamate. Hippocampal BACE1 activity was lower in the lactate group compared to the exercise group (P = 0.01). Oxamate resulted in higher BACE1 protein content compared to sedentary in the PFC (vs. sedentary P = 0.048). These findings suggest that lactate is important for regulating ADAM10 activity and thereby shifts APP processing away from Aß production. KEY POINTS: Exercise is known to alter the processing of amyloid precursor protein by reducing the activity of the rate-limiting enzyme BACE1 and increasing the activity of ADAM10. It is thought that exercise-induced factors are responsible for these enzymatic changes. This study examined if lactate accumulation plays a role in this process. Mice were assigned to one of four groups: sedentary, lactate, exercise and exercise + lactate. The findings demonstrate that lactate accumulation alters brain BACE1 and ADAM10 and shifts amyloid precursor protein processing away from beta-amyloid production.

Development of capillary dysfunction in Alzheimer's disease.

Alzheimer's disease (AD) is currently considered the major cause of cognitive impairment in older adults. This explains the close attention to the issue of AD research. The pathomorphological basis of the disease is a neurodegenerative process, the early stages of which are formed in the hippocampus and the morphofunctionally deep parts of the temporal lobes of the brain closely related to it. Several hypotheses have been advanced concerning the causes of neurodegeneration: the amyloid hypothesis, the calcium homeostasis impairment hypothesis, the inflammatory hypothesis, and the prion hypothesis. However, these hypotheses cannot explain the early stages of the pathogenesis of neurodegenerative diseases, in particular Alzheimer's disease. This health problem requires further comprehensive study of available data, as well as additional investigations to determine the nature of such a process. In this review, the data on microcirculatory disorders in the capillaries of the hippocampus and mediobasal structures of the temporal lobes of the brain, which may be an initiating factor that triggers neurodegenerative events, are analyzed.

Furin, ADAM, and γ-secretase: Core regulatory targets in the Notch pathway and the therapeutic potential for breast cancer.

The activation of the Notch pathway promotes the occurrence and progression of breast cancer. The Notch signal plays different roles in different molecular subtypes of breast cancer. In estrogen receptor-positive (ER+) breast cancer, the Notch pathway regulates the activity of estrogen receptors. In human epidermal growth factor receptor 2-positive (HER2+) breast cancer, crosstalk between Notch and HER2 enhances HER2 signal expression. In triple-negative breast cancer (TNBC), Notch pathway activation is closely linked to tumor invasion and drug resistance. This article offers a comprehensive review of the structural domains, biological functions, and key targets of Notch with a specific focus on the roles of Furin protease, ADAM metalloprotease, and γ-secretase in breast cancer and their potential as therapeutic targets. We discuss the functions and mutual regulatory mechanisms of these proteinases in the Notch pathway as well as other potential targets in the Notch pathway, such as the glycosylation process and key transcription factors. This article also introduces new approaches in the treatment of breast cancer, with a special focus on the molecular characteristics and treatment response differences of different subtypes. We propose that the core regulatory molecules of the Notch pathway may become key targets for development of personalized treatment, which may significantly improve treatment outcomes and prognosis for patients with breast cancer.

Synthesis and Evaluation of Chloride-Substituted Ramalin Derivatives for Alzheimer's Disease Treatment.

Alzheimer's disease (AD) is a progressive neurodegenerative disorder marked by the accumulation of amyloid-beta plaques and hyperphosphorylated tau proteins, leading to cognitive decline and neuronal death. However, despite extensive research, there are still no effective treatments for this condition. In this study, a series of chloride-substituted Ramalin derivatives is synthesized to optimize their antioxidant, anti-inflammatory, and their potential to target key pathological features of Alzheimer's disease. The effect of the chloride position on these properties is investigated, specifically examining the potential of these derivatives to inhibit tau aggregation and beta-site amyloid precursor protein cleaving enzyme 1 (BACE-1) activity. Our findings demonstrate that several derivatives, particularly RA-3Cl, RA-4Cl, RA-26Cl, RA-34Cl, and RA-35Cl, significantly inhibit tau aggregation with inhibition rates of approximately 50%. For BACE-1 inhibition, Ramalin and RA-4Cl also significantly decrease BACE-1 expression in N2a cells by 40% and 38%, respectively, while RA-23Cl and RA-24Cl showed inhibition rates of 30% and 35% in SH-SY5Y cells. These results suggest that chloride-substituted Ramalin derivatives possess promising multifunctional properties for AD treatment, warranting further investigation and optimization for clinical applications.

Air pollution amyloidogenesis is attenuated by the gamma-secretase modulator GSM-15606.

INTRODUCTION: Chronic air pollution (AirPoll) is associated with accelerated cognitive decline and risk of Alzheimer's disease (AD). Correspondingly, wild-type and AD-transgenic rodents exposed to AirPoll have increased amyloid peptides and behavioral impairments. METHODS: We examined the γ-secretase modulator GSM-15606 for potential AirPoll protection by its attenuating of amyloid beta (Aß)42 peptide production. Male and female wild-type mice were fed GSM-15606 during an 8-week inhalation exposure to AirPoll subfractions, ambient nanoparticulate matter (nPM), and diesel exhaust particles (DEP). RESULTS: GSM-15606 decreased Aß42 during nPM and DEP exposure without changing beta- or gamma-secretase activity or BACE1 and PS1 protein levels. DEP increased lateral ventricle volume by 25%. DISCUSSION: These enzyme responses are relevant to AD drug treatments, as well as to the physiological functions of the Aß42 peptide. GSM-15606 attenuation of Aß42 may benefit human exposure to AirPoll. HIGHLIGHTS: Gamma-secretase modulator (GSM-15606) attenuates the amyloidogenic amyloid beta (Aß)42 peptide during exposure to air pollution, which may be a mechanism by which air pollution increases Alzheimer's disease (AD) risk. AD drug treatments may also consider Aß homeostasis among the chronic effects of GSM-15606 and other amyloid reduction treatments on secretase enzymes.

Neuroprotective Effects of Phenolic Constituents from Drynariae Rhizoma.

This study aimed to provide scientific data on the anti-Alzheimer's disease (AD) effects of phenolic compounds from Drynariae Rhizoma (DR) extract using a multi-component approach. Screening of DR extracts, fractions, and the ten phenolic compounds isolated from DR against the key AD-related enzymes acetylcholinesterase (AChE), butyrylcholinesterase (BChE), ß-site amyloid precursor protein cleaving enzyme 1 (BACE1), and monoamine oxidase-B (MAO-B) confirmed their significant inhibitory activities. The DR extract was confirmed to have BACE1-inhibitory activity, and the ethyl acetate and butanol fractions were found to inhibit all AD-related enzymes, including BACE1, AChE, BChE, and MAO-B. Among the isolated phenolic compounds, compounds (2) caffeic acid 4-O-ß-D-glucopyranoside, (6) kaempferol 3-O-rhamnoside 7-O-glucoside, (7) kaempferol 3-o-b-d-glucopyranoside-7-o-a-L-arabinofuranoside, (8) neoeriocitrin, (9) naringin, and (10) hesperidin significantly suppressed AD-related enzymes. Notably, compounds 2 and 8 reduced soluble Amyloid Precursor Protein ß (sAPPß) and ß-secretase expression by over 45% at a concentration of 1.0 µM. In the thioflavin T assay, compounds 6 and 7 decreased Aß aggregation by approximately 40% and 80%, respectively, and degraded preformed Aß aggregates. This study provides robust evidence regarding the potential of DR as a natural therapeutic agent for AD, highlighting specific compounds that may contribute to its efficacy.

The reciprocal relationship between amyloid precursor protein and mitochondrial function.

Amyloid precursor protein (APP), secretase enzymes, and amyloid beta (Aß) have been extensively studied in the context of Alzheimer's disease (AD). Despite this, the function of these proteins and their metabolism is not understood. APP, secretase enzymes, and APP processing products (Aß and C-terminal fragments) localize to endosomes, mitochondria, endoplasmic reticulum (ER), and mitochondrial/ER contact sites. Studies implicate significant relationships between APP, secretase enzyme function, APP metabolism, and mitochondrial function. Mitochondrial dysfunction is a key pathological hallmark of AD and is intricately linked to proteostasis. Here, we review studies examining potential functions of APP, secretase enzymes, and APP metabolites in the context of mitochondrial function and bioenergetics. We discuss implications and limitations of studies and highlight knowledge gaps that remain in the field.

Harnessing the Therapeutic Potential of Peptides for Synergistic Treatment of Alzheimer's Disease by Targeting Aß Aggregation, Metal-Mediated Aß Aggregation, Cholinesterase, Tau Degradation, and Oxidative Stress.

Alzheimer's disease (AD) is a progressive multifaceted neurodegenerative disease and remains a formidable global health challenge. The current medication for AD gives symptomatic relief and, thus, urges us to look for alternative disease-modifying therapies based on a multitarget directed approach. Looking at the remarkable progress made in peptide drug development in the last decade and the benefits associated with peptides, they offer valuable chemotypes [multitarget directed ligands (MTDLs)] as AD therapeutics. This review recapitulates the current developments made in harnessing peptides as MTDLs in combating AD by targeting multiple key pathways involved in the disease's progression. The peptides hold immense potential and represent a convincing avenue in the pursuit of novel AD therapeutics. While hurdles remain, ongoing research offers hope that peptides may eventually provide a multifaceted approach to combat AD.

Evidence of γ-secretase complex involved in the regulation of intramembrane proteolysis in Entamoeba histolytica.

Presenilins (PSNs) are multifunctional membrane proteins involved in signal transduction, lysosomal acidification, and certain physiological processes related to mitochondria. The aspartic protease activity of PSN and the formation of a γ-secretase complex with other subunits such as nicastrin (NCT) are required for the biological functions. Although PSN is widely conserved in eukaryotes, most studies on PSN were conducted in metazoans. Homologous genes for PSN and NCT (EhPSN and EhNCT, respectively) are encoded in the genome of Entamoeba histolytica, however, their functions remain unknown. In this study, we showed that EhPSN and EhNCT form a complex on the cell membrane, demonstrating that the parasite possesses γ-secretase. The predicted structure of EhPSN was similar to the human homolog, demonstrated by the crystal structure, and phylogenetic analysis indicated good conservation between EhPSN and human PSN, supporting the premise that EhPSN functions as a subunit of γ-secretase. By contrast, EhNCT appears to have undergone remarkable structural changes during its evolution. Blue native-polyacrylamide gel electrophoresis combined with western blotting indicated that a 150-kDa single band contains both EhPSN (estimated molecular size: 47-kDa) and EhNCT (64-kDa), suggesting that the complex also contains other unknown components or post-translational modifications. Coimmunoprecipitation from amebic lysates also confirmed that EhPSN and EhNCT formed a complex. Indirect immunofluorescence analysis revealed that the complex localized to the plasma membrane. Moreover, EhPSN exhibited protease activity, which was suppressed by a γ-secretase inhibitor. This is the first report of a γ-secretase complex in protozoan parasites.

Eta-secretase-like processing of the amyloid precursor protein (APP) by the rhomboid protease RHBDL4.

The amyloid precursor protein (APP) is a key protein in Alzheimer's disease synthesized in the endoplasmic reticulum (ER) and translocated to the plasma membrane where it undergoes proteolytic cleavages by several proteases. Conversely, to other known proteases, we previously elucidated rhomboid protease RHBDL4 as a novel APP processing enzyme where several cleavages likely occur already in the ER. Interestingly, the pattern of RHBDL4-derived large APP C-terminal fragments resembles those generated by the η-secretase or MT5-MMP, which was described to generate so-called Aη fragments. The similarity in large APP C-terminal fragments between both proteases raised the question of whether RHBDL4 may contribute to η-secretase activity and Aη-like fragments. Here, we identified two cleavage sites of RHBDL4 in APP by mass spectrometry, which, intriguingly, lie in close proximity to the MT5-MMP cleavage sites. Indeed, we observed that RHBDL4 generates Aη-like fragments in vitro without contributions of α-, ß-, or γ-secretases. Such Aη-like fragments are likely generated in the ER since RHBDL4-derived APP-C-terminal fragments do not reach the cell surface. Inherited, familial APP mutations appear to not affect this processing pathway. In RHBDL4 knockout mice, we observed increased cerebral full-length APP in comparison to wild type (WT) in support of RHBDL4 being a physiologically relevant protease for APP. Furthermore, we found secreted Aη fragments in dissociated mixed cortical cultures from WT mice, however significantly fewer Aη fragments in RHBDL4 knockout cultures. Our data underscores that RHBDL4 contributes to the η-secretease-like processing of APP and that RHBDL4 is a physiologically relevant protease for APP.

Natural Bioactive Compounds from Macroalgae and Microalgae for the Treatment of Alzheimer's Disease: A Review.

Neuroinflammation, toxic protein aggregation, oxidative stress, and mitochondrial dysfunction are key pathways in neurodegenerative diseases like Alzheimer's disease (AD). Targeting these mechanisms with antioxidants, anti-inflammatory compounds, and inhibitors of Aß formation and aggregation is crucial for treatment. Marine algae are rich sources of bioactive compounds, including carbohydrates, phenolics, fatty acids, phycobiliproteins, carotenoids, fatty acids, and vitamins. In recent years, they have attracted interest from the pharmaceutical and nutraceutical industries due to their exceptional biological activities, which include anti-inflammation, antioxidant, anticancer, and anti-apoptosis properties. Multiple lines of evidence have unveiled the potential neuroprotective effects of these multifunctional algal compounds for application in treating and managing AD. This article will provide insight into the molecular mechanisms underlying the neuroprotective effects of bioactive compounds derived from algae based on in vitro and in vivo models of neuroinflammation and AD. We will also discuss their potential as disease-modifying and symptomatic treatment strategies for AD.

Alzheimer's disease linked Aß42 exerts product feedback inhibition on γ-secretase impairing downstream cell signaling.

Amyloid ß (Aß) peptides accumulating in the brain are proposed to trigger Alzheimer's disease (AD). However, molecular cascades underlying their toxicity are poorly defined. Here, we explored a novel hypothesis for Aß42 toxicity that arises from its proven affinity for γ-secretases. We hypothesized that the reported increases in Aß42, particularly in the endolysosomal compartment, promote the establishment of a product feedback inhibitory mechanism on γ-secretases, and thereby impair downstream signaling events. We conducted kinetic analyses of γ-secretase activity in cell-free systems in the presence of Aß, as well as cell-based and ex vivo assays in neuronal cell lines, neurons, and brain synaptosomes to assess the impact of Aß on γ-secretases. We show that human Aß42 peptides, but neither murine Aß42 nor human Aß17-42 (p3), inhibit γ-secretases and trigger accumulation of unprocessed substrates in neurons, including C-terminal fragments (CTFs) of APP, p75, and pan-cadherin. Moreover, Aß42 treatment dysregulated cellular homeostasis, as shown by the induction of p75-dependent neuronal death in two distinct cellular systems. Our findings raise the possibility that pathological elevations in Aß42 contribute to cellular toxicity via the γ-secretase inhibition, and provide a novel conceptual framework to address Aß toxicity in the context of γ-secretase-dependent homeostatic signaling.

Pharmacophore-based virtual screening of commercial databases against ß-secretase 1 for drug development against Alzheimer's disease.

ß-secretase 1, one of the most important proteins, is an aspartate protease. This membrane-associated protein is used for treating Alzheimer's disease (AD). Several inhibitors have been pursued against ß-secretase 1, but they still have not resulted effectively. Virtual screening based on pharmacophores has been shown to be useful for lead optimization and hit identification in the preliminary phase of developing a new drug. Here, we screen the commercially available databases to find the hits against ß-secretase 1 for drug discovery against AD. Virtual screening for 200,000 compounds was done using the database from the Vitas-M Laboratory. The phase screen score was utilized to assess the screened hits. Molecular docking was performed on compounds with phase scores >1.9. According to the study, the 66H ligand of the crystal structure has the maximum performance against ß-secretase 1. The redocking of the co-crystal ligand showed that the docked ligand was seamlessly united with the crystal structure. The reference complex had three hydrogen bonds with Asp93, Asp289, and Gly291; one van der Waals interaction with Gly74; and three hydrophobic interactions. After equilibration, the RMSD of the reference compound sustained a value of ∼1.5 Å until 30 ns and then boosted to 2.5 Å. On comparison, the RMSD of the S1 complex steadily increased to ∼2.5 Å at 15 ns, displayed slight aberrations at approximately ∼2.5-3 Å until 80 ns, and then achieved steadiness toward the end of the simulation. The arrangements of proteins stayed condensed during the mockup when bonded to these complexes as stable Rg values showed. Furthermore, the MM/GBSA technique was employed to analyze both compounds' total binding free energies (ΔGtotal). Our research study provides a new understanding of using 66H as anti-ß-secretase 1 for drug development against AD.

APP fragment controls both ionotropic and non-ionotropic signaling of NMDA receptors.

NMDA receptors (NMDARs) are ionotropic receptors crucial for brain information processing. Yet, evidence also supports an ion-flux-independent signaling mode mediating synaptic long-term depression (LTD) and spine shrinkage. Here, we identify AETA (Aη), an amyloid-ß precursor protein (APP) cleavage product, as an NMDAR modulator with the unique dual regulatory capacity to impact both signaling modes. AETA inhibits ionotropic NMDAR activity by competing with the co-agonist and induces an intracellular conformational modification of GluN1 subunits. This favors non-ionotropic NMDAR signaling leading to enhanced LTD and favors spine shrinkage. Endogenously, AETA production is increased by in vivo chemogenetically induced neuronal activity. Genetic deletion of AETA production alters NMDAR transmission and prevents LTD, phenotypes rescued by acute exogenous AETA application. This genetic deletion also impairs contextual fear memory. Our findings demonstrate AETA-dependent NMDAR activation (ADNA), characterizing AETA as a unique type of endogenous NMDAR modulator that exerts bidirectional control over NMDAR signaling and associated information processing.