Ben Readhead

Gene-regulatory networks (GRNs) offer a mechanistic framework for dissecting the complex molecular and genetic architecture of coronary artery disease and for identifying new therapeutic opportunities. Here, we present a GRN-driven drug-repurposing strategy that integrates transcriptional signatures induced by silencing key drivers of the human arterial wall foam cell regulator GRN 42 with drug-induced gene expression profiles from the NIH LINCS program. In vitro screening of top candidate compounds validates the computational predictions, identifying candidate foam cell modulators and showing that auranofin, an FDA-approved gold salt used to treat rheumatoid arthritis, effectively reduces foam cell formation. In vivo, auranofin attenuates atherosclerosis and inflammation in both male mice and rabbits. Clinically, auranofin was associated with reduced cardiovascular risk in a retrospective cohort of patients with rheumatoid arthritis. Together, these findings show that a GRN-based drug-repurposing framework, coupled with preclinical and clinical validation, can uncover new therapeutic applications for existing drugs, including auranofin, in coronary artery disease.

INTRODUCTION: While there may be microbial contributions to Alzheimer's disease (AD), findings have been inconclusive. We recently reported an AD‐associated CD83(+) microglia subtype associated with increased immunoglobulin G4 (IgG4) in the transverse colon (TC). METHODS: We used immunohistochemistry (IHC), IgG4 repertoire profiling, and brain organoid experiments to explore this association. RESULTS: CD83(+) microglia in the superior frontal gyrus (SFG) are associated with elevated IgG4 and human cytomegalovirus (HCMV) in the TC, anti‐HCMV IgG4 in cerebrospinal fluid, and both HCMV and IgG4 in the SFG and vagal nerve. This association was replicated in an independent AD cohort. HCMV‐infected cerebral organoids showed accelerated AD pathophysiological features (Aβ42 and pTau‐212) and neuronal death. DISCUSSION: Findings indicate complex, cross‐tissue interactions between HCMV and the adaptive immune response associated with CD83(+) microglia in persons with AD. This may indicate an opportunity for antiviral therapy in persons with AD and biomarker evidence of HCMV, IgG4, or CD83(+) microglia. Highlights: Cross‐tissue interaction between HCMV and the adaptive immune response in a subset of persons with AD. Presence of CD83(+) microglial associated with IgG4 and HCMV in the gut. CD83(+) microglia are also associated presence of HCMV and IgG4 in the cortex and vagal nerve. Replication of key association in an independent cohort of AD subjects. HCMV infection of cerebral organoids accelerates the production of AD neuropathological features.

Viral infections leading to inflammation have been implicated in several common diseases, such as Alzheimer’s disease (AD) and type 1 diabetes (T1D). Of note, herpes simplex virus 1 (HSV-1) has been reported to be associated with AD. We sought to identify the transcriptomic changes due to HSV-1 infection and anti-viral drug (acyclovir, ACV) treatment of HSV-1 infection in dissociated cells from human cerebral organoids (dcOrgs) versus stem cell-derived pancreatic islets (sc-islets) to gain potential biological insights into the relevance of HSV-1-induced inflammation in AD and T1D. We observed that differentially expressed genes (DEGs) in HSV-1-infected sc-islets were enriched for genes associated with several autoimmune diseases, most significantly, T1D, but also rheumatoid arthritis, psoriasis, Crohn’s disease, and multiple sclerosis, whereas DEGs in HSV-1-infected dcOrgs were exclusively enriched for genes associated with AD. The ACV treatment of sc-islets was not as effective in rescuing transcript perturbations of autoimmune disease-associated genes. Finally, we identified gene ontology categories that were enriched for DEGs that were in common across, or unique to, viral treatment of dcOrgs and sc-islets, such as categories involved in the transferase complex, mitochondrial, and autophagy function. In addition, we compared transcriptomic signatures from HSV-1-infected sc-islets with sc-islets that were infected with the coxsackie B virus (CVB) that had been associated with T1D pathogenesis. Collectively, this study provides tissue-specific insights into the molecular effects of inflammation in AD and T1D.

Recent studies suggest that heparan sulfate proteoglycans (HSPG) contribute to the predisposition to, protection from, and potential treatment and prevention of Alzheimer’s disease (AD). Here, we used electronic health records (EHR) from two different health systems to examine whether heparin therapy was associated with a delayed diagnosis of AD dementia. Longitudinal EHR data from 15,183 patients from the Mount Sinai Health System (MSHS) and 6207 patients from Columbia University Medical Center (CUMC) were used in separate survival analyses to compare those who did or did not receive heparin therapy, had a least 5 years of observation, were at least 65 years old by their last visit, and had subsequent diagnostic code or drug treatment evidence of possible AD dementia. Analyses controlled for age, sex, comorbidities, follow-up duration and number of inpatient visits. Heparin therapy was associated with significant delays in age of clinical diagnosis of AD dementia, including +1.0 years in the MSMS cohort (P < 0.001) and +1.0 years in the CUMC cohort (P < 0.001). While additional studies are needed, this study supports the potential roles of heparin-like drugs and HSPGs in the protection from and prevention of AD dementia.

Viral infections leading to inflammation have been implicated in several common diseases such as Alzheimer’s disease (AD) and type 1 diabetes (T1D). Of note, herpes simplex virus 1 (HSV-1) has been reported to be associated with AD. We sought to identify the transcriptomic changes due to HSV-1 infection and anti-viral drug (acyclovir, ACV) treatment of HSV-1 infection in dissoci-ated cells from human cerebral organoids (dcOrgs) versus stem cell-derived pancreatic islets (sc-islets) to gain potential biological insights into the relevance of HSV-1-induced inflammation in AD and T1D. We observed that differentially expressed genes (DEGs) in HSV-1-infected sc-islets were enriched for genes associated with several autoimmune diseases, most significantly T1D but also rheumatoid arthritis, psoriasis, Crohn’s disease, and multiple sclerosis, whereas DEGs in HSV-1-infected dcOrgs were exclusively enriched for genes associated with AD. ACV treatment of sc-islets did not rescue transcript expression of autoimmune disease-associated genes. Finally, we identified gene ontology categories that were enriched for DEGs that were in common across, or unique to, viral treatment of dcOrgs and sc-islets, such as categories involved in the transferase complex, mitochondrial and autophagy function. Collectively, this studies pro-vide insight to the molecular effects of inflammation in AD and T1D.

The emergence of single nucleus RNA sequencing (snRNA-seq) offers to revolutionize the study of Alzheimer’s disease (AD). Integration with complementary multiomics data such as genetics, proteomics and clinical data provides powerful opportunities to link cell subpopulations and molecular networks with a broader disease-relevant context. We report snRNA-seq profiles from superior frontal gyrus samples from 101 well characterized subjects from the Banner Brain and Body Donation Program in combination with whole genome sequences. We report findings that link common AD risk variants with CR1 expression in oligodendrocytes as well as alterations in hematological parameters. We observed an AD-associated CD83(+) microglial subtype with unique molecular networks and which is associated with immunoglobulin IgG4 production in the transverse colon. Our major observations were replicated in two additional, independent snRNA-seq data sets. These findings illustrate the power of multi-tissue molecular profiling to contextualize snRNA-seq brain transcriptomics and reveal disease biology.

The emergence of technologies that can support high-throughput profiling of single cell transcriptomes offers to revolutionize the study of brain tissue from persons with and without Alzheimer’s disease (AD). Integration of these data with additional complementary multiomics data such as genetics, proteomics and clinical data provides powerful opportunities to link observed cell subpopulations and molecular network features within a broader disease-relevant context. We report here single nucleus RNA sequencing (snRNA-seq) profiles generated from superior frontal gyrus cortical tissue samples from 101 exceptionally well characterized, aged subjects from the Banner Brain and Body Donation Program in combination with whole genome sequences. We report findings that link common AD risk variants with CR1 expression in oligodendrocytes as well as alterations in peripheral hematological lab parameters, with these observations replicated in an independent, prospective cohort study of ageing and dementia. We also observed an AD-associated CD83(+) microglial subtype with unique molecular networks that encompass many known regulators of AD-relevant microglial biology, and which are associated with immunoglobulin IgG4 production in the transverse colon. These findings illustrate the power of multi-tissue molecular profiling to contextualize snRNA-seq brain transcriptomics and reveal novel disease biology. The transcriptomic, genetic, phenotypic, and network data resources described within this study are available for access and utilization by the scientific community.

Women have a higher incidence of Alzheimer’s disease (AD), even after adjusting for increased longevity. Thus, there is an urgent need to identify the molecular networks that underpin the sex-associated risk of AD. Recent efforts have identified PIN1 as a key regulator of tau phosphorylation signaling pathway . Pin1 is the only gene, to date, that when deleted can cause both tau and Aβ-related pathologies in an age-dependent manner. We analyzed multiple brain transcriptomic datasets focusing on sex differences in PIN1 mRNA levels, in an aging and AD cohort, which revealed reduced PIN1 levels driven by females. Then, we validated this observation in an independent dataset (ROS/MAP) which also revealed that PIN1 is negatively correlated with multiregional neurofibrillary tangle density and global cognitive function, in females only. Additional analysis revealed a decrease in PIN1 in subjects with mild cognitive impairment (MCI) compared with aged individuals, again, driven predominantly by female subjects. Our results show that while both male and female AD patients show decreased PIN1 expression, changes occur before the onset of clinical symptoms of AD in females and correlate to early events associated with AD risk (e.g., synaptic dysfunction). These changes are specific to neurons, and may be a potential prognostic marker to assess AD risk in the aging population and even more so in AD females with increased risk of AD. Graphical

Glucocorticoids (GCs) are widely used for the treatment of inflammatory skin diseases despite significant adverse effects including skin atrophy. Effects of GCs are mediated by the glucocorticoid receptor (GR), a well‐known transcription factor. Previously, we discovered that one of the GR target genes, REDD1, is causatively involved in skin atrophy. Here, we investigated its role in GR function using HaCaT REDD1 knockout (KO) keratinocytes. We found large differences in transcriptome of REDD1 KO and control Cas9 cells in response to glucocorticoid fluocinolone acetonide (FA): both the scope and amplitude of response were significantly decreased in REDD1 KO. The status of REDD1 did not affect GR stability/degradation during self‐desensitization, and major steps in GR activation—its nuclear import and phosphorylation at activating Ser211. However, the amount of GR phosphorylated at Ser226 that may play negative role in GR signalling, was increased in the nuclei of REDD1 KO cells. GR nuclear import and transcriptional activity also depend on the composition of GR chaperone complex: exchange of chaperone FKBP51 (FK506‐binding protein 5) for FKBP52 (FK506‐binding protein 4) being a necessary step in GR activation. We found the increased expression and abnormal nuclear translocation of FKBP51 in both untreated and FA‐treated REDD1 KO cells. Overall, our results suggest the existence of a feed‐forward loop in GR signalling mediated by its target gene REDD1, which has translational potential for the development of safer GR‐targeted therapies.

Mitochondrial (MT) dysfunction has been associated with several neurodegenerative diseases including Alzheimer’s disease (AD). While MT-copy number differences have been implicated in AD, the effect of MT heteroplasmy on AD has not been well characterized. Here, we analyzed over 1800 whole genome sequencing data from four AD cohorts in seven different tissue types to determine the extent of MT heteroplasmy present. While MT heteroplasmy was present throughout the entire MT genome for blood samples, we detected MT heteroplasmy only within the MT control region for brain samples. We observed that an MT variant 10398A>G (rs2853826) was significantly associated with overall MT heteroplasmy in brain tissue while also being linked with the largest number of distinct disease phenotypes of all annotated MT variants in MitoMap. Using gene-expression data from our brain samples, our modeling discovered several gene networks involved in mitochondrial respiratory chain and Complex I function associated with 10398A>G. The variant was also found to be an expression quantitative trait loci (eQTL) for the gene MT-ND3. We further characterized the effect of 10398A>G by phenotyping a population of lymphoblastoid cell-lines (LCLs) with and without the variant allele. Examination of RNA sequence data from these LCLs reveal that 10398A>G was an eQTL for MT-ND4. We also observed in LCLs that 10398A>G was significantly associated with overall MT heteroplasmy within the MT control region, confirming the initial findings observed in post-mortem brain tissue. These results provide novel evidence linking MT SNPs with MT heteroplasmy and open novel avenues for the investigation of pathomechanisms that are driven by this pleiotropic disease associated loci.