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Inflammation impacts androgen receptor signaling in basal prostate stem cells through interleukin 1 receptor antagonist.
Get PDFChronic prostate inflammation in patients with benign prostate hyperplasia (BPH) correlates with the severity of symptoms. How inflammation contributes to prostate enlargement and/or BPH symptoms and the underlying mechanisms remain unclear. In this study, we utilize a unique transgenic mouse model that mimics chronic non-bacterial prostatitis in men and investigate the impact of inflammation on androgen receptor (AR) in basal prostate stem cells (bPSC) and their differentiation in vivo. We find that inflammation significantly enhances AR levels and activity in bPSC. More importantly, we identify interleukin 1 receptor antagonist (IL-1RA) as a crucial regulator of AR in bPSC during inflammation. IL-1RA is one of the top molecules upregulated by inflammation, and inhibiting IL-1RA reverses the enhanced AR activity in organoids derived from inflamed bPSC. Additionally, IL-1RA appears to activate AR by counteracting IL-1α\u27s inhibitory effect. Furthermore, using a lineage tracing model, we observe that inflammation induces bPSC proliferation and differentiation into luminal cells even under castrate conditions, indicating that AR activation driven by inflammation is sufficient to promote bPSC proliferation and differentiation. Taken together, our study uncovers mechanisms through which inflammation modulates AR signaling in bPSC and induces bPSC luminal differentiation that may contribute to prostate hyperplasia
Biallelic EPB41L3 variants underlie a developmental disorder with seizures and myelination defects.
No full textErythrocyte membrane protein band 4.1 like 3 (EPB41L3: NM_012307.5), also known as DAL1, encodes the ubiquitously expressed, neuronally enriched 4.1B protein, part of the 4.1 superfamily of membrane-cytoskeleton adaptors. The 4.1B protein plays key roles in cell spreading, migration and cytoskeletal scaffolding that support oligodendrocyte axon adhesions essential for proper myelination. We herein describe six individuals from five unrelated families with global developmental delay, intellectual disability, seizures, hypotonia, neuroregression and delayed myelination. Exome sequencing identified biallelic variants in EPB41L3 in all affected individuals: two nonsense [c.466C\u3eT, p.(R156*); c.2776C\u3eT, p.(R926*)] and three frameshift [c.666delT, p.(F222Lfs*46); c.2289dupC, p.(V764Rfs*19); c.948_949delTG, p.(A317Kfs*33)]. Quantitative-real time PCR and western blot analyses of human fibroblasts harbouring EPB41L3:c.666delT, p.(F222Lfs*46) indicated ablation of EPB41L3 mRNA and 4.1B protein expression. Inhibition of the nonsense mediated decay (NMD) pathway led to an upregulation of EPB41L3:c.666delT transcripts, supporting NMD as a pathogenic mechanism. Epb41l3-deficient mouse oligodendroglia cells showed significant reduction in mRNA expression of key myelin genes, reduced branching and increased apoptosis. Our report provides the first clinical description of an autosomal recessive disorder associated with variants in EPB41L3, which we refer to as EPB41L3-associated developmental disorder (EADD). Moreover, our functional studies substantiate the pathogenicity of EPB41L3 hypothesized loss-of-function variants
Hematopoietic stem cell gene therapy improves outcomes in a clinically relevant mouse model of multiple sulfatase deficiency.
No full textMultiple sulfatase deficiency (MSD) is a severe, lysosomal storage disorder caused by pathogenic variants in the gene SUMF1, encoding the sulfatase modifying factor formylglycine-generating enzyme. Patients with MSD exhibit functional deficiencies in all cellular sulfatases. The inability of sulfatases to break down their substrates leads to progressive and multi-systemic complications in patients, similar to those seen in single-sulfatase disorders such as metachromatic leukodystrophy and mucopolysaccharidoses IIIA. Here, we aimed to determine if hematopoietic stem cell transplantation with ex vivo SUMF1 lentiviral gene therapy could improve outcomes in a clinically relevant mouse model of MSD. We first tested our approach in MSD patient-derived cells and found that our SUMF1 lentiviral vector improved protein expression, sulfatase activities, and glycosaminoglycan accumulation. In vivo, we found that our gene therapy approach rescued biochemical deficits, including sulfatase activity and glycosaminoglycan accumulation, in affected organs of MSD mice treated post-symptom onset. In addition, treated mice demonstrated improved neuroinflammation and neurocognitive function. Together, these findings suggest that SUMF1 HSCT-GT can improve both biochemical and functional disease markers in the MSD mouse
Mutation in Prkra results in cerebellar abnormality and reduced eIF2α phosphorylation in a model of DYT-PRKRA.
Get PDFVariants in the PRKRA gene, which encodes PACT, cause the early-onset primary dystonia DYT-PRKRA, a movement disorder associated with disruption of coordinated muscle movements. PACT and its murine homolog RAX activate protein kinase R (PKR; also known as EIF2AK2) by a direct interaction in response to cellular stressors to mediate phosphorylation of the α subunit of eukaryotic translation initiation factor 2 (eIF2α). Mice homozygous for a naturally arisen, recessively inherited frameshift mutation, Prkralear-5J, exhibit progressive dystonia. In the present study, we investigated the biochemical and developmental consequences of the Prkralear-5J mutation. Our results indicated that the truncated PACT/RAX protein retains its ability to interact with PKR but inhibits PKR activation. Mice homozygous for the mutation showed abnormalities in cerebellar development as well as a severe lack of dendritic arborization of Purkinje neurons. Additionally, reduced eIF2α phosphorylation was noted in the cerebellum and Purkinje neurons of the homozygous Prkralear-5J mice. These findings indicate that PACT/RAX-mediated regulation of PKR activity and eIF2α phosphorylation plays a role in cerebellar development and contributes to the dystonia phenotype resulting from the Prkralear-5J mutation
Leveraging clinical intuition to improve accuracy of phenotype-driven prioritization.
Get PDFPURPOSE: Clinical intuition is commonly incorporated into the differential diagnosis as an assessment of the likelihood of candidate diagnoses based either on the patient population being seen in a specific clinic or on the signs and symptoms of the initial presentation. Algorithms to support diagnostic sequencing in individuals with a suspected rare genetic disease do not yet incorporate intuition and instead assume that each Mendelian disease has an equal pretest probability.
METHODS: The LIkelihood Ratio Interpretation of Clinical AbnormaLities (LIRICAL) algorithm calculates the likelihood ratio of clinical manifestations represented by Human Phenotype Ontology terms to rank candidate diagnoses. The initial version of LIRICAL assumed an equal pretest probability for each disease in its calculation of the posttest probability (where the test is diagnostic exome or genome sequencing). We introduce Clinical Intuition for Likelihood Ratios (ClintLR), an extension of the LIRICAL algorithm that boosts the pretest probability of groups of related diseases deemed to be more likely.
RESULTS: The average rank of the correct diagnosis in simulations using ClintLR showed a statistically significant improvement over a range of adjustment factors.
CONCLUSION: ClintLR successfully encodes clinical intuition to improve ranking of rare diseases in diagnostic sequencing. ClintLR is freely available at https://github.com/TheJacksonLaboratory/ClintLR
Steering research on mRNA splicing in cancer towards clinical translation.
No full textSplicing factors are affected by recurrent somatic mutations and copy number variations in several types of haematologic and solid malignancies, which is often seen as prima facie evidence that splicing aberrations can drive cancer initiation and progression. However, numerous spliceosome components also \u27moonlight\u27 in DNA repair and other cellular processes, making their precise role in cancer difficult to pinpoint. Still, few would deny that dysregulated mRNA splicing is a pervasive feature of most cancers. Correctly interpreting these molecular fingerprints can reveal novel tumour vulnerabilities and untapped therapeutic opportunities. Yet multiple technological challenges, lingering misconceptions, and outstanding questions hinder clinical translation. To start with, the general landscape of splicing aberrations in cancer is not well defined, due to limitations of short-read RNA sequencing not adept at resolving complete mRNA isoforms, as well as the shallow read depth inherent in long-read RNA-sequencing, especially at single-cell level. Although individual cancer-associated isoforms are known to contribute to cancer progression, widespread splicing alterations could be an equally important and, perhaps, more readily actionable feature of human cancers. This is to say that in addition to \u27repairing\u27 mis-spliced transcripts, possible therapeutic avenues include exacerbating splicing aberration with small-molecule spliceosome inhibitors, targeting recurrent splicing aberrations with synthetic lethal approaches, and training the immune system to recognize splicing-derived neoantigens
Targeting immune-fibroblast cell communication in heart failure.
No full textInflammation and tissue fibrosis co-exist and are causally linked to organ dysfunction1,2 . However, the molecular mechanisms driving immune–fibroblast cell communication in human cardiac disease remain unexplored and there are at present no approved treatments that directly target cardiac fibrosis3,4 . Here we performed multiomic single-cell gene expression, epitope mapping and chromatin accessibility profiling in 45 healthy donor, acutely infarcted and chronically failing human hearts. We identified a disease-associated fibroblast trajectory that diverged into distinct populations reminiscent of myofibroblasts and matrifibrocytes, the latter expressing fibroblast activator protein (FAP) and periostin (POSTN). Genetic lineage tracing of FAP+ fibroblasts in vivo showed that they contribute to the POSTN lineage but not the myofibroblast lineage. We assessed the applicability of experimental systems to model cardiac fibroblasts and demonstrated that three different in vivo mouse models of cardiac injury were superior compared with cultured human heart and dermal fibroblasts in recapitulating the human disease phenotype. Ligand–receptor analysis and spatial transcriptomics predicted that interactions between C-C chemokine receptor type 2 (CCR2) macrophages and fibroblasts mediated by interleukin-1β (IL-1β) signalling drove the emergence of FAP/POSTN fibroblasts within spatially defined niches. In vivo, we deleted the IL-1 receptor on fibroblasts and the IL-1β ligand in CCR2 + monocytes and macrophages, and inhibited IL-1β signalling using a monoclonal antibody, and showed reduced FAP/POSTN fibroblasts, diminished myocardial fibrosis and improved cardiac function. These findings highlight the broader therapeutic potential of targeting inflammation to treat tissue fibrosis and preserve organ function
Ontology-based modeling, integration, and analysis of heterogeneous clinical, pathological, and molecular kidney data for precision medicine.
No full textMany data resources generate, process, store, or provide kidney related molecular, pathological, and clinical data. Reference ontologies offer an opportunity to support knowledge and data integration. The Kidney Precision Medicine Project (KPMP) team contributed to the representation and addition of 329 kidney phenotype terms to the Human Phenotype Ontology (HPO), and identified many subcategories of acute kidney injury (AKI) or chronic kidney disease (CKD). The Kidney Tissue Atlas Ontology (KTAO) imports and integrates kidney-related terms from existing ontologies (e.g., HPO, CL, and Uberon) and represents 259 kidney-related biomarkers. We have also developed a precision medicine metadata ontology (PMMO) to integrate 50 variables from KPMP and CZ CellxGene data resources and applied PMMO for integrative kidney data analysis. The gene expression profiles of kidney gene biomarkers were specifically analyzed under healthy control or AKI/CKD disease states. This work demonstrates how ontology-based approaches support multi-domain data and knowledge integration in precision medicine
Computing Minimal Boolean Models of Gene Regulatory Networks.
No full textModels of gene regulatory networks (GRNs) capture the dynamics of the regulatory processes that occur within the cell as a means to understanding the variability observed in gene expression between different conditions. Arguably the simplest mathematical construct used for modeling is the Boolean network, which dictates a set of logical rules for transition between states described as Boolean vectors. Due to the complexity of gene regulation and the limitations of experimental technologies, in most cases knowledge about regulatory interactions and Boolean states is partial. In addition, the logical rules themselves are not known a priori. Our goal in this work is to create an algorithm that finds the network that fits the data optimally, and identify the network states that correspond to the noise-free data. We present a novel methodology for integrating experimental data and performing a search for the optimal consistent structure via optimization of a linear objective function under a set of linear constraints. In addition, we extend our methodology into a heuristic that alleviates the computational complexity of the problem for datasets that are generated by single-cell RNA-Sequencing (scRNA-Seq). We demonstrate the effectiveness of these tools using simulated data, and in addition a publicly available scRNA-Seq dataset and the GRN that is associated with it. Our methodology will enable researchers to obtain a better understanding of the dynamics of GRNs and their biological role
Effects of CD47 Expression on Macrophage-Mediated Phagocytosis of Acute Myeloid Leukemia
No full textAcute Myeloid Leukemia (AML) is a hematological disease caused by the proliferation of immature myeloid precursors in the bone marrow. While chemotherapy remains the first-line treatment approach and exerts anti-tumor effects, this treatment is often less tolerated by older patients and can significantly increase the risk of patient death. This finding has encouraged investigation into the use of immunotherapeutic targets to overcome the obstacles arising from chemotherapies. The cell surface receptor CD47 presents a favorable target due to its role in suppressing phagocytosis and allowing leukemia cells to evade immune detection. CD4 7 has been shown to be upregulated in many cancers, including AML. As therapeutics directly targeting CD47 caused adverse reactions in clinical trials, we investigated upstream regulators and regulatory sites on CD4 7 and assessed the functional consequences of modulations in surface expression. This study was successful in identifying DYRK1A and CBL as positive and negative regulators of CD47, respectively. We found that genetic ablation of these molecules modulated CD47 surface expression, as did elimination of regulatory sites directly on the CD47 protein. These manipulations significantly impacted the progression of macrophage-mediated phagocytosis, providing novel evidence that targeting CD4 7 regulation may present a powerful mechanism by which the CD47-SIRPa axis can be attenuated