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AbstractBackgroundRecent studies showed that neuroinflammation plays a key role in triggering specific neuropsychiatric symptoms (NPS), such as irritability and agitation, in individuals with Alzheimer’s disease (AD). While prior studies showed an association between tau pathology and all NPS domains, the extent to which tau influences each specific NPS domain remains unclear. Here, we aim to investigate the association of tau and NPS domains in the AD continuum. We hypothesize that tau plays a comparatively greater effect on the emergence of psychotic symptoms compared to other NPS domains.MethodWe assessed 56 individuals (21 cognitively unimpaired (CU), 23 MCI, and 12 AD dementia) from the HEAD study who underwent clinical assessments with the Neuropsychiatry Inventory Questionnaire (NPI‐Q) and had positron emission tomography (PET) for amyloid‐ß (Aß) ([18F]AZD4694 or [11C]PiB), and tau tangles ([18F]MK6240) at the same visit. We selected individuals with an NPI‐Q total score =1. Tau SUVR values were tailored with a mask from Braak stages I‐VI, using the inferior cerebellar gray matter as reference region. Leave‐one‐out voxel wise and linear regression tested the association between each NPI‐Q domain and biomarkers accounting for age, sex, cognitive status, and study site.ResultCI individuals had significantly higher NPI‐Q score and Braak VI PET SUVR than CU individuals (Table 1). NPI‐Q score was significantly associated with tau‐PET in the periRolandic and supplementary motor cortex (Figures 1A). Linear regression showed that NPI‐Q associates with tau‐PET in the Braak stage VI (Figure 1B). Leave‐one‐out regression analysis revealed that delusions, motor disturbances, and anxiety contributed most to the association between tau‐PET and NPS (Figure 2A, C). These domains presented a higher magnitude of association compared to each other NPI‐Q domain (Figure 2B). Notably, irritability, agitation, and disinhibition exerted a negative effect to the association, emphasizing that tau may not play a role in the development of these symptoms (Figure 2A, D)ConclusionOur study supports previous evidence suggesting that irritability and agitation may not be triggered by tau, but rather by other pathological process such as neuroinflammation. These findings provide additional rationale for the therapeutics aiming to mitigate irritability and agitation in AD patients.

AbstractBackgroundUtilizing PET amyloid‐beta (Aß) and tau for staging Alzheimer’s Disease (AD) has demonstrated potential in identifying individuals with varying degrees of disease severity, applicable to both clinical trials and practice. However, the diverse binding characteristics of tau tracers pose challenges to the application of this staging across different ligands. In this study, we evaluate a novel staging framework proposed by the AA working group, employing Aß PET and either [18F]MK6240 or [18F]Flortaucipir in individuals participating in a head‐to‐head study of tau PET tracers. This will provide insights into the robustness of this framework when applied across different tau ligands.MethodWe evaluated 154 people (89 CU and 65 CI), with MK6240, Flortaucipir, Aß‐PET, MRI. SUVRs used inferior cerebellar gray as reference. Participants were categorized in Aß positivity(A), using visual read, and tau positivity (T) in medial temporal lobe(MTL) and neocortex(NEO). Tau MTL(TMTL) and NEO positivity were defined as average +3 SD of youngs. NEO moderate(TMOD) and high(THIGH) was defined as below and above median SUVR of NEO‐positive individuals. Individuals were grouped as: A‐T‐, A+T‐, A+TMTL+(early), A+TMOD+(intermediary), and A+THIGH+(advanced).ResultsWe found that the distribution of the number of individuals across the five AD stages was comparable when using both MK6240 and Flortaucipir tracers (Figure 1). However, upon examining the concordance within each group, we found a low concordance of 30% (3/10) between tracers in the early tau stage (A+TMTL+). Concordance increased in the intermediary stage A+TMOD+, with a rate of 65% (9/14), and further increased to 79% (11/14) in the advanced stage (A+THIGH+). The A+T‐ group exhibited the highest concordance between tracer results, at 95% (21/22).ConclusionOur findings indicate that the application of this framework across different populations using different tau tracers can lead to a compatible distribution among groups. However, in a direct comparison, we noted that this did not necessarily result in high concordance between groups, as group shifts were observed that depended on the cutoffs used (data not shown). These results emphasize the necessity for harmonization of tau PET tracers prior to their utilization for stage determination.

AbstractBackgroundThe efficacy of Calorie Restriction (CR) in enhancing cognition, promoting healthy aging, and extending lifespan is well‐established. Yet, it remains unclear whether the apolipoprotein E (APOE) genotype, a known modifier for aging and age‐related disorders, influences the beneficial effects of CR in countering aging.MethodsTo investigate this question, we utilized humanized APOE mouse models, which express APOE2, APOE3, or APOE4 alleles systematically (refer to as E2, E3, and E4 mice). These mice were subjected to either ad libitum (AL) feeding or 30% CR feeding, starting from 12 months and continuing up to 20 months of age (N = 11‐20/ genotype /group, mixed sex). We monitored energy expenditure using the Comprehensive Laboratory Animal Monitoring System (CLAMS). Additionally, we evaluated anxiety levels and memory capabilities through various behavioral tests. Finally, we collected brain, blood, and feces samples for transcriptomic, lipidomic, and microbiomic analyses.ResultsCLAMS measurements showed that CR significantly decreased the metabolic rate in E3 and E4 mice, but not E2 mice, during the non‐feeding period of the CR group, compared to the AL group. Furthermore, CR reduced anxiety and enhanced associative memory in E3 and E4 mice relative to the AL group. Brain transcriptomics revealed that CR upregulated the cholesterol synthesis pathway, potentially improving myelination‐related pathways in E3 and E4 mice. This finding was further validated by the increased staining intensity of myelin basic protein (MBP) observed in E3 and E4 mice from an independent cohort. In contrast, these effects were not pronounced in E2 mice. Additionally, while CR’s impact on the brain lipid profile was minor across all APOE genotypes, it significantly altered blood lipid profile of E2 mice, with minimal changes observed in E3 and E4 mice. Finally, in microbiomics, CR led to notable changes in the abundance of Bifidobacterium species in E3 and E4 mice and Lactobacillus species in E2 mice.ConclusionsOur study highlights the role of APOE genotypes in modulating the diverse effects of CR on cognition, brain transcriptomics, lipid metabolism, and the microbiome, providing a crucial foundation for considering APOE genotypes in developing prevention strategies for aging and age‐related disorders.

AbstractBackgroundRecent studies showed that neuroinflammation plays a key role in triggering specific neuropsychiatric symptoms (NPS), such as irritability and agitation, in individuals with Alzheimer’s disease (AD). While prior studies showed an association between tau pathology and all NPS domains, the extent to which tau influences each specific NPS domain remains unclear. Here, we aim to investigate the association of tau and NPS domains in the AD continuum. We hypothesize that tau plays a comparatively greater effect on the emergence of psychotic symptoms compared to other NPS domains.MethodWe assessed 56 individuals (21 cognitively unimpaired (CU), 23 MCI, and 12 AD dementia) from the HEAD study who underwent clinical assessments with the Neuropsychiatry Inventory Questionnaire (NPI‐Q) and had positron emission tomography (PET) for amyloid‐β (Aβ) ([18F]AZD4694 or [11C]PiB), and tau tangles ([18F]MK6240) at the same visit. We selected individuals with an NPI‐Q total score ≥1. Tau SUVR values were tailored with a mask from Braak stages I‐VI, using the inferior cerebellar gray matter as reference region. Leave‐one‐out voxel wise and linear regression tested the association between each NPI‐Q domain and biomarkers accounting for age, sex, cognitive status, and study site.ResultCI individuals had significantly higher NPI‐Q score and Braak VI PET SUVR than CU individuals (Table 1). NPI‐Q score was significantly associated with tau‐PET in the periRolandic and supplementary motor cortex (Figures 1A). Linear regression showed that NPI‐Q associates with tau‐PET in the Braak stage VI (Figure 1B). Leave‐one‐out regression analysis revealed that delusions, motor disturbances, and anxiety contributed most to the association between tau‐PET and NPS (Figure 2A, C). These domains presented a higher magnitude of association compared to each other NPI‐Q domain (Figure 2B). Notably, irritability, agitation, and disinhibition exerted a negative effect to the association, emphasizing that tau may not play a role in the development of these symptoms (Figure 2A, D)ConclusionOur study supports previous evidence suggesting that irritability and agitation may not be triggered by tau, but rather by other pathological process such as neuroinflammation. These findings provide additional rationale for the therapeutics aiming to mitigate irritability and agitation in AD patients.

Abstract Background Temozolomide (TMZ) treatment has demonstrated a variable impact on glioma prognosis. This study examines associations of survival with DNA repair gene germline polymorphisms among glioma patients who did and did not have TMZ treatment. Identifying genetic markers which sensitize tumor cells to TMZ could personalize therapy and improve outcomes. Methods We evaluated TMZ-related survival associations of pathogenic germline SNPs and genetically predicted transcript levels within 34 DNA repair genes among 1504 glioma patients from the University of California San Francisco Adult Glioma Study (UCSF AGS) and Mayo Clinic, whose diagnoses spanned pre- and post-TMZ eras within the major known glioma prognostic molecular subtypes. Results Among those who received TMZ, 5 SNPs were associated with overall survival, but not in those who did not receive TMZ. Only rs2308321-G, in MGMT, was associated with decreased survival (hazard ratio = 1.21, P = .019) for all glioma subtypes. Rs73191162-T (near UNG), rs13076508-C (near PARP3), rs7840433-A (near NEIL2), and rs3130618-A (near MSH5) were only associated with survival and TMZ treatment for certain subtypes, suggesting subtype-specific germline chemosensitization. Genetically predicted elevated expression of PNKP, compared to normal brain expression, was associated with markedly poor survival in TMZ-treated patients with isocitrate dehydrogenase (IDH)-mutant and 1p/19q non-codeleted gliomas (P = .015), with a median difference of over 70 months in overall survival times. Similarly, NEIL2 and TDG expressions were associated with altered TMZ-related survival only among certain subtypes. Conclusions Functional germline alterations within DNA repair genes were associated with TMZ sensitivity, measured by overall survival among adults with glioma. These variants should be evaluated in prospective analyses and functional studies.

AbstractMacitentan is a dual endothelin receptor antagonist (ERA) approved for treating pulmonary arterial hypertension (PAH). SOPRANO evaluated the efficacy and safety of macitentan versus placebo in pulmonary hypertension (PH) patients after left ventricular assist device (LVAD) implantation. SOPRANO was a phase 2, multicenter, double‐blind, randomized, placebo‐controlled, parallel‐group study. Patients with an LVAD implanted within the prior 90 days who had persistent PH (i.e., mean pulmonary arterial pressure ≥25 mmHg, pulmonary artery wedge pressure [PAWP] ≤18 mmHg, and pulmonary vascular resistance [PVR] >3 Wood units [WU]) were randomized (1:1) to macitentan 10 mg or placebo once daily for 12 weeks. The primary endpoint was change in PVR. Secondary endpoints included change in right‐heart catheterization hemodynamic variables, N‐terminal prohormone of brain natriuretic peptide levels, World Health Organization functional class, and safety/tolerability. Fifty‐seven patients were randomized to macitentan (n = 28) or placebo (n = 29). A statistically significant reduction in PVR from baseline to Week 12 was observed with macitentan versus placebo (placebo‐corrected geometric mean ratio, 0.74; 95% confidence interval, 0.58–0.94; p = .0158). No statistically significant differences were observed in secondary endpoints. In a post‐hoc analysis, 66.7% of patients receiving macitentan achieved PVR <3 WU versus 40.0% receiving placebo (p = .0383). Macitentan was generally well tolerated; adverse events were consistent with those in previous PAH studies with macitentan. In conclusion, macitentan showed promising tolerability and significantly reduced PVR in PH patients with persistently elevated PVR after LVAD implantation. ClinicalTrials. gov identifier: NCT02554903.

Abstract Background The majority of missense variants in clinical genetic tests are classified as variants of uncertain significance. Prior research has shown that the deleterious effects and the subsequent molecular consequence of variants are often conserved among paralogous protein sequences within a gene family. Here, we systematically quantified on an exome-wide scale if the existence of pathogenic variants in paralogous genes at a conserved position could serve as evidence for the pathogenicity of a new variant. For the gene family of voltage-gated sodium channels where variants and expert-curated clinical phenotypes were available, we also assessed whether phenotype patterns of multiple disorders for each gene were also conserved across variant positions within the gene family. Methods We developed a framework that assesses the presence of pathogenic missense variants located in conserved residues across paralogous genes. We systematically mapped 2.5 million pathogenic and general population variants from the ClinVar, HGMD, and gnomAD databases onto a total of 9,990 genes and aligned them by gene families. We evaluated the quantity of classifiable amino acids by utilizing pathogenic variants identified in databases alone and then compared this assessment to the inclusion of paralogous pathogenic variants. We validated and quantified the evidence of conserved pathogenic paralogous variants in variant pathogenicity classification. Results Considering conserved pathogenic variants in paralogous genes, increased the number of classifiable variants 2.8-fold across the exome, compared to pathogenic variants in the gene of interest alone. The presence of a pathogenic variant in a paralogous gene is associated with a positive likelihood ratio of 8.32 for variant pathogenicity. The likelihood ratio was gene family-specific. Across ten genes encoding voltage-gated sodium channels and 22 expert-curated disorders, we identified cross-paralog correlated phenotypes based on 3D structure spatial position. For example, the established loss-of-function disorders SCN1A-associated Dravet syndrome, SCN2A-associated autism, SCN5A-associated Brugarda Syndrome, and SCN8A-associated neurodevelopmental disorder without seizures were correlated in their spatial variant position on structure. Finally, we show that phenotype integration in paralog variant selection improves variant classification. Conclusion Our results show that paralogous variants, in particular with phenotype information can enhance our understanding of variant effects.

Background: D-flow’s impact on metabolic reprogramming, particularly glycolysis, needs reevaluation due to its conflicting role in atherosclerosis. We show that d-flow reduces LATS1/2 expression, leading to both EC proliferation and senescence. However, the exact molecular mechanisms remain elusive. Methods: We utilized EC-specific Lats1 +/- /Lats2 -/- knockout (LATS1/2-EKO) mice in a partial left carotid ligation (PLCL) model under hypercholesterolemia with AAV-PCSK9 injection to mimic d-flow. Anti-CD38 antibody (Ab68) and IgG2a control were administered intraperitoneally every 5 days at a dose of 5 mg/kg starting one day before PLCL. Plaque characterization was performed using imaging mass cytometry and sequential immunofluorescence (COMET TM ), integrating spatial metabolite data from Bruker timsTOF Flex Mass Spectrometer, and spatial single-cell metabolite data via the Visiopharm platform. Results: EC LATS1/2 depletion induced a unique atherothrombosis lesion with induced EC proliferation and senescence-associated secretory phenotype (SASP). Utilizing spatial single-cell metabolomics, we cataloged 134 distinct metabolites within the ECs. Subsequent overrepresentation analysis revealed that pathways related to glutamate and the citric acid cycle were enhanced in LATS1/2-EKO mice. Specifically, CD38-dependent downregulation of the pyruvate dehydrogenase (PDH) substrate lipoamide decreased PDH enzymatic activity, linking to CD38 to senescence induction. This inhibition led to a metabolic shift towards YAP-dependent glutaminolysis, and upregulated the citric acid cycle. The use of a glutaminase 1 inhibitor (CB-839) significantly curtailed senescence-associated stemness (SAS), underscoring the pivotal role of CD38 in driving glutaminolysis that contributes to d-flow-induced SAS. Moreover, the application of Ab68 but not IgG2a control decreased atherothrombosis formation following PLCL in LATS1/2-EKO mice. However, the effectiveness of Ab68 in attenuating atherosclerosis in wild-type mice was less pronounced. Conclusions: In the presence of d-flow, upregulated CD38, resulting from reduced LATS1/2 expression, triggers EC responses characterized by increased glutaminolysis and the emergence of an EC SAS phenotype. These processes play a crucial role in a unique form of atherothrombosis. Additionally, CD38’s impact on reducing PDH activity, along with YAP and CD38-mediated enhancement of glutaminolysis, constitutes critical pathways for inducing EC SAS by d-flow.

Background: Atherosclerotic plaques, especially high-risk ones, form in blood vessel regions exposed to d-flow. EC dysfunction induced by d-flow likely contributes to plaque development. In contrast, regions with laminar flow (l-flow) are less susceptible to plaque formation. Although the Hippo pathway is implicated in mechano-transduction, the exact role and molecular mechanisms of LATS1/2 in response to d-flow remain incompletely understood. Methods: We employed endothelial cell (EC)-specific Lats1 and Lats2 knockout mice within a partial left carotid ligation (PLCL) model to simulate disturbed flow conditions. We characterized plaques using imaging mass cytometry (IMC) and sequential immunofluorescence via COMET TM . Results: We investigated the effects of d-flow on YAP activity and LATS1/2 expression. While both l-flow and d-flow activate YAP activity, only d-flow leads to decreased LATS1/2 expression. We induced Lats1/2 deletion in tamoxifen-inducible Lats1 -/- /Lats2 -/- EKO mice. Within 14 days, all mice (27/27) died due to severe systemic edema and increased vascular permeability. In Lats1 +/- /Lats2 -/- EKO mice (LATS1/2-EKO), we observed atherothrombotic lesions characterized by increased EC proliferation and a senescence-associated secretory phenotype (SASP) in vivo . Mechanistically, d-flow reduces LATS1/2 expression, leading to increased CD38 expression. CD38 triggers not only SASP, but also senescence-associated stemness (SAS) via NAD + depletion, dependent on Lamin A but independent of YAP. We also explored the relationship between CD38 and Ki67 in plaques in vivo. We calculated the logarithm of the Ki67:CD38 expression ratio at the single-cell level using IMC/COMET data. Based on the trimodal density distribution of this parameter, we found that in one group, CD38 and Ki67 were linearly co-expressed, and the percentage of LATS1/2-depleted cells in this group exceeded that of wild type (WT) cells, suggesting that CD38’s pro-proliferative effects dominate under LATS1/2 depletion conditions, despite its role in inducing senescence. Lastly, inhibiting CD38 mitigated d EC SAS/SASP, and atherothrombosis under LATS1/2 depletion conditions. Conclusions: Reduced LATS1/2-mediated CD38 expression in response to d-flow leads to an EC response marked by increased SASP/SAS, ultimately contributing to atherothrombosis. While this phenomenon is common in human advanced coronary atherosclerosis, it is not observed in commonly used mouse models of atherosclerosis.