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INTRODUCTION: A noncoding variant (rs35349669) within INPP5D, a lipid and protein phosphatase restricted to microglia in the brain, is linked to increased susceptibility to Alzheimer's disease (AD). While Inpp5d is well‐studied in amyloid pathology, its role in tau pathology remains unclear. METHODS: PS19 Tauopathy mice were crossed with Inpp5d‐haplodeficient (Inpp5d+/−) mice to examine the impact of Inpp5d in tau pathology. RESULTS: Increased INPP5D expression correlated positively with phospho‐Tau AT8 in PS19 mice. Inpp5d haplodeficiency mitigated hyperphosphorylated tau levels (AT8, AT180, AT100, and PHF1) and motor deficits in PS19 mice. Transcriptomic analysis revealed an up‐regulation of genes associated with immune response and cell migration. DISCUSSION: Our findings define an association between INPP5D expression and tau pathology in PS19 mice. Alleviation in hyperphosphorylated tau, motor deficits, and transcriptomics changes in haplodeficient‐Inpp5d PS19 mice indicate that modulation in INPP5D expression may provide therapeutic potential for mitigating tau pathology and improving motor deficits. Highlights: The impact of Inpp5d in the context of tau pathology was studied in the PS19 mouse model.INPP5D expression is associated with tau pathology.Reduced Inpp5d expression in PS19 mice improved motor functions and decreased total and phospho‐Tau levels.Inpp5d haplodeficiency in PS19 mice modulates gene expression patterns linked to immune response and cell migration.These data suggest that inhibition of Inpp5d may be a therapeutic approach in tauopathies. [ABSTRACT FROM AUTHOR]

Abstract Recent genetic studies on Alzheimer’s disease (AD) have brought microglia under the spotlight, as loci associated with AD risk are enriched in genes expressed in microglia. Several of these genes have been recognized for their central roles in microglial functions. Increasing evidence suggests that SHIP1, the protein encoded by the AD-associated gene INPP5D, is an important regulator of microglial phagocytosis and immune response. A recent study from our group identified SHIP1 as a negative regulator of the NLRP3 inflammasome in human iPSC-derived microglial cells (iMGs). In addition, we found evidence for a connection between SHIP1 activity and inflammasome activation in the AD brain. The NLRP3 inflammasome is a multiprotein complex that induces the secretion of pro-inflammatory cytokines as part of innate immune responses against pathogens and endogenous damage signals. Previously published studies have suggested that the NLRP3 inflammasome is activated in AD and contributes to AD-related pathology. Here, we provide an overview of the current understanding of the microglial NLRP3 inflammasome in the context of AD-related inflammation. We then review the known intracellular functions of SHIP1, including its role in phosphoinositide signaling, interactions with microglial phagocytic receptors such as TREM2 and evidence for its intersection with NLRP3 inflammasome signaling. Through rigorous examination of the intricate connections between microglial signaling pathways across several experimental systems and postmortem analyses, the field will be better equipped to tailor newly emerging therapeutic strategies targeting microglia in neurodegenerative diseases.

Recent genetic studies on Alzheimer's disease (AD) have brought microglia under the spotlight, as loci associated with AD risk are enriched in genes expressed in microglia. Several of these genes have been recognized for their central roles in microglial functions. Increasing evidence suggests that SHIP1, the protein encoded by the AD-associated gene INPP5D, is an important regulator of microglial phagocytosis and immune response. A recent study from our group identified SHIP1 as a negative regulator of the NLRP3 inflammasome in human iPSC-derived microglial cells (iMGs). In addition, we found evidence for a connection between SHIP1 activity and inflammasome activation in the AD brain. The NLRP3 inflammasome is a multiprotein complex that induces the secretion of pro-inflammatory cytokines as part of innate immune responses against pathogens and endogenous damage signals. Previously published studies have suggested that the NLRP3 inflammasome is activated in AD and contributes to AD-related pathology. Here, we provide an overview of the current understanding of the microglial NLRP3 inflammasome in the context of AD-related inflammation. We then review the known intracellular functions of SHIP1, including its role in phosphoinositide signaling, interactions with microglial phagocytic receptors such as TREM2 and evidence for its intersection with NLRP3 inflammasome signaling. Through rigorous examination of the intricate connections between microglial signaling pathways across several experimental systems and postmortem analyses, the field will be better equipped to tailor newly emerging therapeutic strategies targeting microglia in neurodegenerative diseases. [ABSTRACT FROM AUTHOR]

Introduction: Mutations in INPP5D, which encodes for the SH2‐domain‐containing inositol phosphatase SHIP‐1, have recently been linked to an increased risk of developing late‐onset Alzheimer's disease. While INPP5D expression is almost exclusively restricted to microglia in the brain, little is known regarding how SHIP‐1 affects neurobiology or neurodegenerative disease pathogenesis. Methods: We generated and investigated 5xFAD Inpp5dfl/flCx3cr1Ert2Cre mice to ascertain the function of microglial SHIP‐1 signaling in response to amyloid beta (Aβ)‐mediated pathology. Results: SHIP‐1 deletion in microglia led to substantially enhanced recruitment of microglia to Aβ plaques, altered microglial gene expression, and marked improvements in neuronal health. Further, SHIP‐1 loss enhanced microglial plaque containment and Aβ engulfment when compared to microglia from Cre‐negative 5xFAD Inpp5dfl/fl littermate controls. Discussion: These results define SHIP‐1 as a pivotal regulator of microglial responses during Aβ‐driven neurological disease and suggest that targeting SHIP‐1 may offer a promising strategy to treat Alzheimer's disease. Highlights: Inpp5d deficiency in microglia increases plaque‐associated microglia numbers.Loss of Inpp5d induces activation and phagocytosis transcriptional pathways.Plaque encapsulation and engulfment by microglia are enhanced with Inpp5d deletion.Genetic ablation of Inpp5d protects against plaque‐induced neuronal dystrophy. [ABSTRACT FROM AUTHOR]

INTRODUCTION: The risk of developing Alzheimer's disease is associated with genes involved in microglial function. Inositol polyphosphate‐5‐phosphatase (INPP5D), which encodes Src homology 2 (SH2) domain–containing inositol polyphosphate 5‐phosphatase 1 (SHIP1), is a risk gene expressed in microglia. Because SHIP1 binds receptor immunoreceptor tyrosine‐based inhibitory motifs (ITIMs), competes with kinases, and converts PI(3,4,5)P3 to PI(3,4)P2, it is a negative regulator of microglia function. Validated inhibitors are needed to evaluate SHIP1 as a potential therapeutic target. METHODS: We identified inhibitors and screened the enzymatic domain of SHIP1. A protein construct containing two domains was used to evaluate enzyme inhibitor potency and selectivity versus SHIP2. Inhibitors were tested against a construct containing all ordered domains of the human and mouse proteins. A cellular thermal shift assay (CETSA) provided evidence of target engagement in cells. Phospho‐AKT levels provided further evidence of on‐target pharmacology. A high‐content imaging assay was used to study the pharmacology of SHIP1 inhibition while monitoring cell health. Physicochemical and absorption, distribution, metabolism, and excretion (ADME) properties were evaluated to select a compound suitable for in vivo studies. RESULTS: SHIP1 inhibitors displayed a remarkable array of activities and cellular pharmacology. Inhibitory potency was dependent on the protein construct used to assess enzymatic activity. Some inhibitors failed to engage the target in cells. Inhibitors that were active in the CETSA consistently destabilized the protein and reduced pAKT levels. Many SHIP1 inhibitors were cytotoxic either at high concentration due to cell stress or they potently induced cell death depending on the compound and cell type. One compound activated microglia, inducing phagocytosis at concentrations that did not result in significant cell death. A pharmacokinetic study demonstrated brain exposures in mice upon oral administration. DISCUSSION: 3‐((2,4‐Dichlorobenzyl)oxy)‐5‐(1‐(piperidin‐4‐yl)‐1H‐pyrazol‐4‐yl) pyridine activated primary mouse microglia and demonstrated exposures in mouse brain upon oral dosing. Although this compound is our recommended chemical probe for investigating the pharmacology of SHIP1 inhibition at this time, further optimization is required for clinical studies. Highlights: Cellular thermal shift assay (CETSA) and signaling (pAKT) assays were developed to provide evidence of src homology 2 (SH2) domain‐contaning inositol phosphatase 1 (SHIP1) target engagement and on‐target activity in cellular assays.A phenotypic high‐content imaging assay with simultaneous measures of phagocytosis, cell number, and nuclear intensity was developed to explore cellular pharmacology and monitor cell health.SHIP1 inhibitors demonstrate a wide range of activity and cellular pharmacology, and many reported inhibitors are cytotoxic.The chemical probe 3‐((2,4‐dichlorobenzyl)oxy)−5‐(1‐(piperidin‐4‐yl)−1H‐pyrazol‐4‐yl) pyridine is recommended to explore SHIP1 pharmacology. [ABSTRACT FROM AUTHOR]

Introduction: The inositol polyphosphate‐5‐phosphatase D (INPP5D) gene encodes a dual‐specificity phosphatase that can dephosphorylate both phospholipids and phosphoproteins. Single nucleotide polymorphisms in INPP5D impact risk for developing late onset sporadic Alzheimer's disease (LOAD). Methods: To assess the consequences of inducible Inpp5d knockdown in microglia of APPKM670/671NL/PSEN1Δexon9 (PSAPP) mice, we injected 3‐month‐old Inpp5dfl/fl/Cx3cr1CreER/+ and PSAPP/Inpp5dfl/fl/Cx3cr1CreER/+ mice with either tamoxifen (TAM) or corn oil (CO) to induce recombination. Results: At age 6 months, we found that the percent area of 6E10+ deposits and plaque‐associated microglia in Inpp5d knockdown mice were increased compared to controls. Spatial transcriptomics identified a plaque‐specific expression profile that was extensively altered by Inpp5d knockdown. Discussion: These results demonstrate that conditional Inpp5d downregulation in the PSAPP mouse increases plaque burden and recruitment of microglia to plaques. Spatial transcriptomics highlighted an extended gene expression signature associated with plaques and identified CST7 (cystatin F) as a novel marker of plaques. Highlights: Inpp5d knockdown increases plaque burden and plaque‐associated microglia number.Spatial transcriptomics identifies an expanded plaque‐specific gene expression profile.Plaque‐induced gene expression is altered by Inpp5d knockdown in microglia.Our plaque‐associated gene signature overlaps with human Alzheimer's disease gene networks. [ABSTRACT FROM AUTHOR]

The single nucleotide polymorphisms rs35349669 and rs10933431 within Inositol Polyphosphate-5-Phosphatase D (INPP5D) are strongly associated with Alzheimer's Disease risk. To better understand INPP5D expression in the brain, we investigated INPP5D isoform expression as a function of rs35349669 and rs10933431, as well as Alzheimer's disease neuropathology, by qPCR and isoform-specific primers. In addition, INPP5D allelic expression imbalance was evaluated relative to rs1141328 within exon 1. Expression of INPP5D isoforms associated with transcription start sites in exon 1 and intron 14 was increased in individuals with high Alzheimer's disease neuropathology. In addition, a novel variant with 47bp lacking from exon 12 increased expression in Alzheimer's Disease brains, accounting for 13% of total INPP5D expression, and was found to undergo nonsense-mediated decay. Although inter-individual variation obscured a possible polymorphism effect on INPP5D isoform expression as measured by qPCR, rs35349669 was associated with rs1141328 allelic expression imbalance, suggesting that rs35349669 is significantly associated with full-length INPP5D isoform expression. In summary, expression of INPP5D isoforms with start sites in exon 1 and intron 14 are increased in brains with high Alzheimer's Disease neuropathology, a novel isoform lacking the phosphatase domain was significantly increased with the disease, and the polymorphism rs35349669 correlates with allele-specific full-length INPP5D expression. [ABSTRACT FROM AUTHOR]

The vascular hypothesis of Alzheimer's disease (AD) has proposed the involvement of brain hypoperfusion in AD pathogenesis, where cognitive decline and dysfunction result from dwindling cerebral blood flow (CBF). Based on the vascular hypothesis of Alzheimer's disease, we focused on exploring how genetic factors influence AD pathogenesis via the cerebrovascular system. To investigate the role of CBF endophenotypes in AD pathogenesis, we performed a targeted genetic analysis of 258 subjects from the Alzheimer's Disease Neuroimaging Initiative cohort to examine associations between 4033 single-nucleotide polymorphisms of 24 AD genes and CBF measures in 4 brain regions. A novel association with CBF measure in the left angular gyrus was identified in an INPP5D single-nucleotide polymorphism (i.e., rs61068452; p = 1.48E-7; corrected p = 2.39E-3). The gene-based analysis discovered both INPP5D and CD2AP associated with the left angular gyrus CBF. Further analyses on nonoverlapping samples revealed that rs61068452-G was associated with lower CSF t-tau/Aβ 1–42 ratio. Our findings suggest a protective role of rs61068452-G in an AD-relevant cerebrovascular endophenotype, which has the potential to provide novel insights for better mechanistic understanding of AD. • We performed a targeted genetic study of CBF phenotypes captured by arterial spin labeling (ASL) perfusion magnetic resonance imaging (pMRI). • We discovered a novel locus in INPP5D (rs61068452) significantly associated with the CBF measure in the L-AG. • The identified genetic variation was also associated with several AD biomarkers. • This work is among the first genetic association studies of CBF in AD, and our finding has the potential to help understand AD molecular mechanism. [ABSTRACT FROM AUTHOR]

Alzheimer's disease (AD) is a progressive neurodegenerative disorder characterized by cognitive decline, robust microgliosis, neuroinflammation, and neuronal loss. Genome-wide association studies recently highlighted a prominent role for microglia in late-onset AD (LOAD). Specifically, inositol polyphosphate-5-phosphatase (INPP5D), also known as SHIP1, is selectively expressed in brain microglia and has been reported to be associated with LOAD. Although INPP5D is likely a crucial player in AD pathophysiology, its role in disease onset and progression remains unclear. We performed differential gene expression analysis to investigate INPP5D expression in AD and its association with plaque density and microglial markers using transcriptomic (RNA-Seq) data from the Accelerating Medicines Partnership for Alzheimer's Disease (AMP-AD) cohort. We also performed quantitative real-time PCR, immunoblotting, and immunofluorescence assays to assess INPP5D expression in the 5xFAD amyloid mouse model. Differential gene expression analysis found that INPP5D expression was upregulated in LOAD and positively correlated with amyloid plaque density. In addition, in 5xFAD mice, Inpp5d expression increased as the disease progressed, and selectively in plaque-associated microglia. Increased Inpp5d expression levels in 5xFAD mice were abolished entirely by depleting microglia with the colony-stimulating factor receptor-1 antagonist PLX5622. Our findings show that INPP5D expression increases as AD progresses, predominantly in plaque-associated microglia. Importantly, we provide the first evidence that increased INPP5D expression might be a risk factor in AD, highlighting INPP5D as a potential therapeutic target. Moreover, we have shown that the 5xFAD mouse model is appropriate for studying INPP5D in AD. • INPP5D was positively associated with amyloid plaque density in the human brain. • In the 5xFAD mouse model, Inpp5d increased in a disease-progression-dependent manner • Inpp5d was selectively expressed in plaque-associated microglia. • The increased Inpp5d expression levels were attenuated following microglia depletion. [ABSTRACT FROM AUTHOR]