Direct interrogation of context-dependent GPCR activity with a universal biosensor platform.

G protein-coupled receptors (GPCRs) are the largest family of druggable proteins encoded in the human genome, but progress in understanding and targeting them is hindered by the lack of tools to reliably measure their nuanced behavior in physiologically relevant contexts. Here, we developed a collection of compact ONE vector G-protein Optical (ONE-GO) biosensor constructs as a scalable platform that can be conveniently deployed to measure G-protein activation by virtually any GPCR with high fidelity even when expressed endogenously in primary cells. By characterizing dozens of GPCRs across many cell types like primary cardiovascular cells or neurons, we revealed insights into the molecular basis for G-protein coupling selectivity of GPCRs, pharmacogenomic profiles of anti-psychotics on naturally occurring GPCR variants, and G-protein subtype signaling bias by endogenous GPCRs depending on cell type or upon inducing disease-like states. In summary, this open-source platform makes the direct interrogation of context-dependent GPCR activity broadly accessible.

Direct interrogation of context-dependent GPCR activity with a universal biosensor platform.

G protein-coupled receptors (GPCRs) are the largest family of druggable proteins in the human genome, but progress in understanding and targeting them is hindered by the lack of tools to reliably measure their nuanced behavior in physiologically-relevant contexts. Here, we developed a collection of compact ONE vector G-protein Optical (ONE-GO) biosensor constructs as a scalable platform that can be conveniently deployed to measure G-protein activation by virtually any GPCR with high fidelity even when expressed endogenously in primary cells. By characterizing dozens of GPCRs across many cell types like primary cardiovascular cells or neurons, we revealed new insights into the molecular basis for G-protein coupling selectivity of GPCRs, pharmacogenomic profiles of anti-psychotics on naturally-occurring GPCR variants, and G-protein subtype signaling bias by endogenous GPCRs depending on cell type or upon inducing disease-like states. In summary, this open-source platform makes the direct interrogation of context-dependent GPCR activity broadly accessible.

Enhancement of the therapeutic efficacy of the MAP regimen using thiamine pyrophosphate-decorated albumin nanoclusters in osteosarcoma treatment.

Recent studies on osteosarcoma regimens have mainly focused on modifying the combination of antineoplastic agents rather than enhancing the therapeutic efficacy of each component. Here, an albumin nanocluster (NC)-assisted methotrexate (MTX), doxorubicin (DOX), and cisplatin (MAP) regimen with improved antitumor efficacy is presented. Human serum albumin (HSA) is decorated with thiamine pyrophosphate (TPP) to increase the affinity to the bone tumor microenvironment (TME). MTX or DOX (hydrophobic MAP components) is adsorbed to HSA-TPP via hydrophobic interactions. MTX- or DOX-adsorbed HSA-TPP NCs exhibit 20.8- and 1.64-fold higher binding affinity to hydroxyapatite, respectively, than corresponding HSA NCs, suggesting improved targeting ability to the bone TME via TPP decoration. A modified MAP regimen consisting of MTX- or DOX-adsorbed HSA-TPP NCs and free cisplatin displays a higher synergistic anticancer effect in HOS/MNNG human osteosarcoma cells than conventional MAP. TPP-decorated NCs show 1.53-fold higher tumor accumulation than unmodified NCs in an orthotopic osteosarcoma mouse model, indicating increased bone tumor distribution. As a result, the modified regimen more significantly suppresses tumor growth in vivo than solution-based conventional MAP, suggesting that HSA-TPP NC-assisted MAP may be a promising strategy for osteosarcoma treatment.

Caffeine Produced in Rice Plants Provides Tolerance to Water-Deficit Stress.

Exogenous or endogenous caffeine application confers resistance to diverse biotic stresses in plants. In this study, we demonstrate that endogenous caffeine in caffeine-producing rice (CPR) increases tolerance even to abiotic stresses such as water deficit. Caffeine produced by CPR plants influences the cytosolic Ca2+ ion concentration gradient. We focused on examining the expression of Ca2+-dependent protein kinase genes, a subset of the numerous proteins engaged in abiotic stress signaling. Under normal conditions, CPR plants exhibited increased expressions of seven OsCPKs (OsCPK10, OsCPK12, OsCPK21, OsCPK25, OsCPK26, OsCPK30, and OsCPK31) and biochemical modifications, including antioxidant enzyme (superoxide dismutase, catalase, peroxidase, and ascorbate peroxidase) activity and non-enzymatic antioxidant (ascorbic acid) content. CPR plants exhibited more pronounced gene expression changes and biochemical alterations in response to water-deficit stress. CPR plants revealed increased expressions of 16 OsCPKs (OsCPK1, OsCPK2, OsCPK3, OsCPK4, OsCPK5, OsCPK6, OsCPK9, OsCPK10, OsCPK11, OsCPK12, OsCPK14, OsCPK16, OsCPK18, OsCPK22, OsCPK24, and OsCPK25) and 8 genes (OsbZIP72, OsLEA25, OsNHX1, OsRab16d, OsDREB2B, OsNAC45, OsP5CS, and OsRSUS1) encoding factors related to abiotic stress tolerance. The activity of antioxidant enzymes increased, and non-enzymatic antioxidants accumulated. In addition, a decrease in reactive oxygen species, an accumulation of malondialdehyde, and physiological alterations such as the inhibition of chlorophyll degradation and the protection of photosynthetic machinery were observed. Our results suggest that caffeine is a natural chemical that increases the potential ability of rice to cope with water-deficit stress and provides robust resistance by activating a rapid and comprehensive resistance mechanism in the case of water-deficit stress. The discovery, furthermore, presents a new approach for enhancing crop tolerance to abiotic stress, including water deficit, via the utilization of a specific natural agent.

The QPLEX™ Plus Assay Kit for the Early Clinical Diagnosis of Alzheimer's Disease.

We recently developed a multiplex diagnostic kit, QPLEX™ Alz plus assay kit, which captures amyloid-ß1-40, galectin-3 binding protein, angiotensin-converting enzyme, and periostin simultaneously using microliters of peripheral blood and utilizes an optimized algorithm for screening Alzheimer's disease (AD) by correlating with cerebral amyloid deposition. Owing to the demand for early AD detection, we investigate the potential of our kit for the early clinical diagnosis of AD. A total of 1395 participants were recruited, and their blood samples were analyzed with the QPLEX™ kit. The average of QPLEX™ algorithm values in each group increased gradually in the order of the clinical progression continuum of AD: cognitively normal (0.382 ± 0.150), subjective cognitive decline (0.452 ± 0.130), mild cognitive impairment (0.484 ± 0.129), and AD (0.513 ± 0.136). The algorithm values between each group showed statistically significant differences among groups divided by Mini-Mental State Examination and Clinical Dementia Rating. The QPLEX™ algorithm values could be used to distinguish the clinical continuum of AD or cognitive function. Because blood-based diagnosis is more accessible, convenient, and cost- and time-effective than cerebral spinal fluid or positron emission tomography imaging-based diagnosis, the QPLEX™ kit can potentially be used for health checkups and the early clinical diagnosis of AD.

Detecting GPCR Signals With Optical Biosensors of Gα-GTP in Cell Lines and Primary Cell Cultures.

G protein-coupled receptors (GPCRs) are the largest class of transmembrane receptors and mediate a wide variety of physiological processes. GPCRs respond to a plethora of extracellular ligands and initiate signaling pathways inside cells via heterotrimeric G proteins (Gαßγ). Because of the critical role GPCRs play in regulating biological processes and as pharmacological targets, the availability of tools to measure their signaling activity are of high interest. Live-cell biosensors that detect the activity of G proteins in response to GPCR stimulation have emerged as a powerful approach to investigate GPCR/G protein signaling. Here, we detail methods to monitor G protein activity through direct measurement of GTP-bound Gα subunits using optical biosensors based on bioluminescence resonance energy transfer (BRET). More specifically, this article describes the use of two types of complementary biosensors. The first protocol explains how to use a multicomponent BRET biosensor that relies on expression of exogenous G proteins in cell lines. This protocol yields robust responses that are compatible with endpoint measurements of dose-dependent ligand effects or with kinetic measurements of subsecond resolution. The second protocol describes the implementation of unimolecular biosensors that detect the activation of endogenous G proteins in cell lines expressing exogenous GPCRs or in primary cells upon stimulation of endogenous GPCRs. Overall, using the biosensors as described in this article will help users characterize the mechanisms of action of many pharmacological agents and natural ligands that modulate GPCR and G protein signaling with high precision. © 2023 Wiley Periodicals LLC. Basic Protocol 1: Using bimolecular BRET biosensors to monitor Gα-GTP formation of tagged Gα in live cells Alternate Protocol 1: Measuring GPCR dose-dependent Gα-GTP responses in endpoint format Basic Protocol 2: Using unimolecular BRET biosensors to study endogenous G protein activity Alternate Protocol 2: Using unimolecular BRET biosensors to study endogenous G protein activity in mouse cortical neurons.

Fine-tuning GPCR-mediated neuromodulation by biasing signaling through different G protein subunits.

G-protein-coupled receptors (GPCRs) mediate neuromodulation through the activation of heterotrimeric G proteins (Gαßγ). Classical models depict that G protein activation leads to a one-to-one formation of Gα-GTP and Gßγ species. Each of these species propagates signaling by independently acting on effectors, but the mechanisms by which response fidelity is ensured by coordinating Gα and Gßγ responses remain unknown. Here, we reveal a paradigm of G protein regulation whereby the neuronal protein GINIP (Gα inhibitory interacting protein) biases inhibitory GPCR responses to favor Gßγ over Gα signaling. Tight binding of GINIP to Gαi-GTP precludes its association with effectors (adenylyl cyclase) and, simultaneously, with regulator-of-G-protein-signaling (RGS) proteins that accelerate deactivation. As a consequence, Gαi-GTP signaling is dampened, whereas Gßγ signaling is enhanced. We show that this mechanism is essential to prevent the imbalances of neurotransmission that underlie increased seizure susceptibility in mice. Our findings reveal an additional layer of regulation within a quintessential mechanism of signal transduction that sets the tone of neurotransmission.

Small-molecule targeting of GPCR-independent noncanonical G-protein signaling in cancer.

Activation of heterotrimeric G-proteins (Gαßγ) by G-protein-coupled receptors (GPCRs) is a quintessential mechanism of cell signaling widely targeted by clinically approved drugs. However, it has become evident that heterotrimeric G-proteins can also be activated via GPCR-independent mechanisms that remain untapped as pharmacological targets. GIV/Girdin has emerged as a prototypical non-GPCR activator of G proteins that promotes cancer metastasis. Here, we introduce IGGi-11, a first-in-class small-molecule inhibitor of noncanonical activation of heterotrimeric G-protein signaling. IGGi-11 binding to G-protein α-subunits (Gαi) specifically disrupted their engagement with GIV/Girdin, thereby blocking noncanonical G-protein signaling in tumor cells and inhibiting proinvasive traits of metastatic cancer cells. In contrast, IGGi-11 did not interfere with canonical G-protein signaling mechanisms triggered by GPCRs. By revealing that small molecules can selectively disable noncanonical mechanisms of G-protein activation dysregulated in disease, these findings warrant the exploration of therapeutic modalities in G-protein signaling that go beyond targeting GPCRs.

Sex differences in the progression of glucose metabolism dysfunction in Alzheimer's disease.

Alzheimer's disease (AD) is a common neurodegenerative disease characterized by amyloid plaques and impaired brain metabolism. Because women have a higher prevalence of AD than men, sex differences are of great interest. Using cross-sectional and longitudinal data, we showed sex-dependent metabolic dysregulations in the brains of AD patients. Cohort 1 (South Korean, n = 181) underwent Pittsburgh compound B-PET, fluorodeoxyglucose-PET, magnetic resonance imaging, and blood biomarker (plasma tau and beta-amyloid 42 and 40) measurements at baseline and two-year follow-ups. Transcriptome analysis of data from Cohorts 2 and 3 (European, n = 78; Singaporean, n = 18) revealed sex differences in AD-related alterations in brain metabolism. In women (but not in men), all imaging indicators displayed consistent correlation curves with AD progression. At the two-year follow-up, clear brain metabolic impairment was revealed only in women, and the plasma beta-amyloid 42/40 ratio was a possible biomarker for brain metabolism in women. Furthermore, our transcriptome analysis revealed sex differences in transcriptomes and metabolism in the brains of AD patients as well as a molecular network of 25 female-specific glucose metabolic genes (FGGs). We discovered four key-attractor FGG genes (ALDOA, ENO2, PRKACB, and PPP2R5D) that were associated with amyloid/tau-related genes (APP, MAPT, BACE1, and BACE2). Furthermore, these genes successfully distinguished amyloid positivity in women. Understanding sex differences in the pathogenesis of AD and considering these differences will improve development of effective diagnostics and therapeutic treatments for AD.

Small-molecule targeting of GPCR-independent non-canonical G protein signaling inhibits cancer progression.

Activation of heterotrimeric G-proteins (Gαßγ) by G-protein-coupled receptors (GPCRs) is a quintessential mechanism of cell signaling widely targeted by clinically-approved drugs. However, it has become evident that heterotrimeric G-proteins can also be activated via GPCR-independent mechanisms that remain untapped as pharmacological targets. GIV/Girdin has emerged as a prototypical non-GPCR activator of G proteins that promotes cancer metastasis. Here, we introduce IGGi-11, a first-in-class smallmolecule inhibitor of non-canonical activation of heterotrimeric G-protein signaling. IGGi-11 binding to G-protein α-subunits (Gαi) specifically disrupted their engagement with GIV/Girdin, thereby blocking non-canonical G-protein signaling in tumor cells, and inhibiting pro-invasive traits of metastatic cancer cells in vitro and in mice. In contrast, IGGi-11 did not interfere with canonical G-protein signaling mechanisms triggered by GPCRs. By revealing that small molecules can selectively disable non-canonical mechanisms of G-protein activation dysregulated in disease, these findings warrant the exploration of therapeutic modalities in G-protein signaling that go beyond targeting GPCRs.

Association of B cell profile and receptor repertoire with the progression of Alzheimer's disease.

Alzheimer's disease (AD) is the most prevalent type of dementia. Reports have revealed that the peripheral immune system is linked to neuropathology; however, little is known about the contribution of B lymphocytes in AD. For this longitudinal study, 133 participants are included at baseline and second-year follow-up. Also, we analyze B cell receptor (BCR) repertoire data generated from a public dataset of three normal and 10 AD samples and perform BCR repertoire profiling and pairwise sharing analysis. As a result, longitudinal increase in B lymphocytes is associated with increased cerebral amyloid deposition and hyperactivates induced pluripotent stem cell-derived microglia with loss-of-function for beta-amyloid clearance. Patients with AD share similar class-switched BCR sequences with identical isotypes, despite the high somatic hypermutation rate. Thus, BCR repertoire profiling can lead to the development of individualized immune-based therapeutics and treatment. We provide evidence of both quantitative and qualitative changes in B lymphocytes during AD pathogenesis.

Multi-Omics-Based Autophagy-Related Untypical Subtypes in Patients with Cerebral Amyloid Pathology.

Recent multi-omics analyses paved the way for a comprehensive understanding of pathological processes. However, only few studies have explored Alzheimer's disease (AD) despite the possibility of biological subtypes within these patients. For this study, unsupervised classification of four datasets (genetics, miRNA transcriptomics, proteomics, and blood-based biomarkers) using Multi-Omics Factor Analysis+ (MOFA+), along with systems-biological approaches following various downstream analyses are performed. New subgroups within 170 patients with cerebral amyloid pathology (Aß+) are revealed and the features of them are identified based on the top-rated targets constructing multi-omics factors of both whole (M-TPAD) and immune-focused models (M-IPAD). The authors explored the characteristics of subtypes and possible key-drivers for AD pathogenesis. Further in-depth studies showed that these subtypes are associated with longitudinal brain changes and autophagy pathways are main contributors. The significance of autophagy or clustering tendency is validated in peripheral blood mononuclear cells (PBMCs; n = 120 including 30 Aß- and 90 Aß+), induced pluripotent stem cell-derived human brain organoids/microglia (n = 12 including 5 Aß-, 5 Aß+, and CRISPR-Cas9 apolipoprotein isogenic lines), and human brain transcriptome (n = 78). Collectively, this study provides a strategy for precision medicine therapy and drug development for AD using integrative multi-omics analysis and network modelling.

Neurotoxicity of phenylalanine on human iPSC-derived cerebral organoids.

Phenylketonuria (PKU) is a common genetic metabolic disorder that causes phenylalanine accumulation in the blood. The most serious symptoms are related to the brain, as intellectual disability, seizure, and microcephaly are commonly found in poorly treated PKU patients and the babies of maternal PKU. However, the mechanism of hyperphenylalaninemia on human neurodevelopment is still unclear. Here we utilized human induced pluripotent stem cell (iPSC)-derived cerebral organoids to investigate the neurotoxicity of hyperphenylalaninemia. Cerebral organoids at days 40 or 100 were treated with different concentrations of phenylalanine for 5 days. After phenylalanine treatments, the cerebral organoids displayed alterations in organoid size, induction of apoptosis, and depletion of neural progenitor cells. However, phenylalanine did not have an impact on neurons and glia, including astrocytes, immature oligodendrocytes, and mature oligodendrocytes. Remarkably, a reduction in the thickness of the cortical rosettes and a decrease in myelination at the intermediate zone were inspected with the elevated phenylalanine concentrations. RNA-seq of phenylalanine-treated organoids revealed that gene sets related to apoptosis, p53 signaling pathway, and TNF signaling pathway via NF-kB were enriched in upregulated genes, while those related to cell cycle and amino acid metabolism were enriched in downregulated genes. In addition, there were several microcephaly disease genes, such as ASPM, LMNB1, and CENPE, ranked at the top of the downregulated genes. These findings indicate that phenylalanine exposure may contribute to microcephaly, abnormal cortical expansion, and myelination lesions in the developing human brain.

Application of QPLEXTM biomarkers in cognitively normal individuals across a broad age range and diverse regions with cerebral amyloid deposition.

The deposition of beta-amyloid (Aß) in the brain precedes the onset of symptoms such as cognitive impairment in Alzheimer's disease (AD); therefore, the early detection of Aß accumulation is crucial. We previously reported the applicability of the QPLEXTM Alz plus assay kit for the prescreening of Aß accumulation. Here, we tested the specific application of the kit in a large cohort of cognitively normal (CN) individuals of varying ages for the early detection of Aß accumulation. We included a total of 221 CN participants with or without brain Aß. The QPLEXTM biomarkers were characterized based on age groups (1st-3rd tertile) and across various brain regions with cerebral amyloid deposition. The 3rd tertile group (>65 years) was found to be the most suitable age group for the application of our assay kit. Receiver operating characteristic curve analysis showed that the area under the curve (AUC, discrimination power) was 0.878 with 69.7% sensitivity and 98.4% specificity in the 3rd tertile group. Additionally, specific correlations between biomarkers and cerebral amyloid deposition in four different brain regions revealed an overall correlation with general amyloid deposition, consistent with previous findings. Furthermore, the combinational panel with plasma Aß1-42 levels maximized the discrimination efficiency and achieved an AUC of 0.921 with 95.7% sensitivity and 67.3% specificity. Thus, we suggest that the QPLEXTM Alz plus assay is useful for prescreening brain Aß levels in CN individuals, especially those aged >65 years, to prevent disease progression via the early detection of disease initiation.

Intracellular Ca2+ accumulation triggered by caffeine provokes resistance against a broad range of biotic stress in rice.

Chemical pesticides are still frequently overused to diminish such crop loss caused by biotic stress despite the threat to humans and the environment. Thus, it is urgent to find safer and more effective defense strategies. In this study, we report that caffeine, implanted through a transgenic approach, enhances resistance against variable biotic stresses in rice without fitness cost. Caffeine-producing rice (CPR) was generated by introducing three N-methyltransferase genes involved in the biosynthesis of caffeine in coffee plants. The CPR plants have no differences in morphology and growth compared to their wild-type counterparts, but they show strongly enhanced resistance to both bacterial leaf blight, rice blast, and attack of white-backed planthoppers. Caffeine acts as a repellent agent against rice pathogens. Moreover, caffeine triggers a series of Ca2+ signalling-like processes to synthesize salicylic acid (SA), a hormone associated with plant resistance. In CPR, phosphodiesterase was inhibited by caffeine, cAMP and cGMP increased, intracellular Ca2+ increased, phenylalanine lyase (PAL) was activated by OsCPK1, and SA synthesis was activated. This finding is a novel strategy to improve resistance against the biotic stresses of crops with a special type of defense inducer.

Natural Sulfurs Inhibit LPS-Induced Inflammatory Responses through NF-κB Signaling in CCD-986Sk Skin Fibroblasts.

Lipopolysaccharide (LPS)-induced inflammatory response leads to serious damage, up to and including tumorigenesis. Natural mineral sulfur, non-toxic sulfur (NTS), and methylsulfonylmethane (MSM) have anti-inflammatory activity that may inhibit LPS-induced inflammation. We hypothesized that sulfur compounds could inhibit LPS-induced inflammatory responses in CCD-986Sk skin fibroblasts. We used Western blotting and real-time PCR to analyze molecular signaling in treated and untreated cultures. We also used flow cytometry for cell surface receptor analysis, comet assays to evaluate DNA damage, and ELISA-based cytokine detection. LPS induced TLR4 activation and NF-κB signaling via canonical and protein kinase C (PKC)-dependent pathways, while NTS and MSM downregulated that response. NTS and MSM also inhibited LPS-induced nuclear accumulation and binding of NF-κB to proinflammatory cytokines COX-2, IL-1ß, and IL-6. Finally, the sulfur compounds suppressed LPS-induced ROS accumulation and DNA damage in CCD-986Sk cells. These results suggest that natural sulfur compounds could be used to treat inflammation and may be useful in the development of cosmetics.

The clinical use of blood-test factors for Alzheimer's disease: improving the prediction of cerebral amyloid deposition by the QPLEXTM Alz plus assay kit.

Alzheimer's disease (AD) is the leading cause of dementia, and many studies have focused on finding effective blood biomarkers for the accurate diagnosis of this disease. Predicting cerebral amyloid deposition is considered the key for AD diagnosis because a cerebral amyloid deposition is the hallmark of AD pathogenesis. Previously, blood biomarkers were discovered to predict cerebral amyloid deposition, and further efforts have been made to increase their sensitivity and specificity. In this study, we analyzed blood-test factors (BTFs) that can be commonly measured in medical health check-ups from 149 participants with cognitively normal, 87 patients with mild cognitive impairment, and 64 patients with clinically diagnosed AD dementia with brain amyloid imaging data available. We demonstrated that four factors among regular health check-up blood tests, cortisol, triglyceride/high-density lipoprotein cholesterol ratio, alanine aminotransferase, and free triiodothyronine, showed either a significant difference by or correlation with cerebral amyloid deposition. Furthermore, we made a prediction model for Pittsburgh compound B-positron emission tomography positivity, using BTFs and the previously discovered blood biomarkers, the QPLEXTM Alz plus assay kit biomarker panel, and the area under the curve was significantly increased up to 0.845% with 69.4% sensitivity and 90.6% specificity. These results show that BTFs could be used as co-biomarkers and that a highly advanced prediction model for amyloid plaque deposition could be achieved by the combinational use of diverse biomarkers.

Non-toxic sulfur inhibits LPS-induced inflammation by regulating TLR-4 and JAK2/STAT3 through IL-6 signaling.

Janus kinase 2 (JAK2) and STAT3 signaling is considered a major pathway in lipopolysaccharide (LPS)­induced inflammation. Toll­like receptor 4 (TLR­4) is an inflammatory response receptor that activates JAK2 during inflammation. STAT3 is a transcription factor for the pro­inflammatory cytokine IL­6 in inflammation. Sulfur is an essential element in the amino acids and is required for growth and development. Non­toxic sulfur (NTS) can be used in livestock feeds as it lacks toxicity. The present study aimed to inhibit LPS­induced inflammation in C2C12 myoblasts using NTS by regulating TLR­4 and JAK2/STAT3 signaling via the modulation of IL­6. The 3­(4,5­dimethylthiazol­2­yl)­2,5­diphenyltetrazolium bromide assay was conducted to analyze cell viability and reverse transcription polymerase chain reaction and western blotting performed to measure mRNA and protein expression levels. Chromatin immunoprecipitation and enzyme­linked immunosorbent assays were used to determine the binding activity of proteins. The results indicated that NTS demonstrated a protective effect against LPS­induced cell death and inhibited LPS­induced expression of TLR­4, JAK2, STAT3 and IL­6. In addition, NTS inhibited the expression of nuclear phosphorylated­STAT3 and its binding to the IL­6 promoter. Therefore, NTS may be a potential candidate drug for the treatment of inflammation.

A logical network-based drug-screening platform for Alzheimer's disease representing pathological features of human brain organoids.

Developing effective drugs for Alzheimer's disease (AD), the most common cause of dementia, has been difficult because of complicated pathogenesis. Here, we report an efficient, network-based drug-screening platform developed by integrating mathematical modeling and the pathological features of AD with human iPSC-derived cerebral organoids (iCOs), including CRISPR-Cas9-edited isogenic lines. We use 1300 organoids from 11 participants to build a high-content screening (HCS) system and test blood-brain barrier-permeable FDA-approved drugs. Our study provides a strategy for precision medicine through the convergence of mathematical modeling and a miniature pathological brain model using iCOs.

Performance of the QPLEX™ Alz plus assay, a novel multiplex kit for screening cerebral amyloid deposition.

BACKGROUND: Alzheimer's disease (AD) is an irreversible neurodegenerative disease characterized by the hallmark finding of cerebral amyloid deposition. Many researchers have tried to predict the existence of cerebral amyloid deposition by using easily accessible blood plasma samples, but the effectiveness of such strategies remains controversial. METHODS: We developed a new multiplex kit, the QPLEX™ Alz plus assay kit, which uses proteomics-based blood biomarkers to prescreen for cerebral amyloid deposition. A total of 300 participants who underwent Pittsburgh compound B (PiB)-positron emission tomography (PET) which allows imaging of cerebral amyloid deposition were included in this study. We compared the levels of QPLEX™ biomarkers between patients who were classified as PiB-negative or PiB-positive, regardless of their cognitive function. Logistic regression analysis followed by receiver operating characteristic (ROC) curve analysis was performed. The kit accuracy was tested using a randomized sample selection method. RESULTS: The results obtained using our assay kit reached 89.1% area under curve (AUC) with 80.0% sensitivity and 83.0% specificity. Further validation of the QPLEX™ Alz plus assay kit using a randomized sample selection method showed an average accuracy of 81.5%. CONCLUSIONS: Our QPLEX™ Alz plus assay kit provides preliminary evidence that it can be used as blood marker to predict cerebral amyloid deposition but independent validation is needed.