Publications by authors named "Claes Wahlestedt"

174 Publications

Emerging Technologies for Genome-Wide Profiling of DNA Breakage.

Front Genet 2020 27;11:610386. Epub 2021 Jan 27.

Department of Psychiatry and Behavioral Sciences, University of Miami Miller School of Medicine, Miami, FL, United States.

Genome instability is associated with myriad human diseases and is a well-known feature of both cancer and neurodegenerative disease. Until recently, the ability to assess DNA damage-the principal driver of genome instability-was limited to relatively imprecise methods or restricted to studying predefined genomic regions. Recently, new techniques for detecting DNA double strand breaks (DSBs) and single strand breaks (SSBs) with next-generation sequencing on a genome-wide scale with single nucleotide resolution have emerged. With these new tools, efforts are underway to define the "breakome" in normal aging and disease. Here, we compare the relative strengths and weaknesses of these technologies and their potential application to studying neurodegenerative diseases.
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http://dx.doi.org/10.3389/fgene.2020.610386DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7873462PMC
January 2021

Sexual Dimorphism in the 3xTg-AD Mouse Model and Its Impact on Pre-Clinical Research.

J Alzheimers Dis 2021 ;80(1):41-52

Department of Psychiatry & Behavioral Sciences, University of Miami Miller School of Medicine, Miami, FL, USA.

Female sex is a leading risk factor for developing Alzheimer's disease (AD). Sexual dimorphism in AD is gaining attention as clinical data show that women are not only more likely to develop AD but also to experience worse pathology and faster cognitive decline. Pre-clinical AD research in animal models often neglects to address sexual dimorphism in evaluation of behavioral or molecular characteristics and outcomes. This can compromise its translation to a clinical setting. The triple-transgenic AD mouse model (3xTg-AD) is a commonly used but unique AD model because it exhibits both amyloid and tau pathology, essential features of the human AD phenotype. Mounting evidence has revealed important sexually dimorphic characteristics of this animal model that have yet to be reviewed and thus, are often overlooked in studies using the 3xTg-AD model. In this review we conduct a thorough analysis of reports of sexual dimorphism in the 3xTg-AD model including findings of molecular, behavioral, and longevity-related sex differences in original research articles through August 2020. Importantly, we find results to be inconsistent, and that strain source and differing methodologies are major contributors to lack of consensus regarding traits of each sex. We first touch on the nature of sexual dimorphism in clinical AD, followed by a brief summary of sexual dimorphism in other major AD murine models before discussing the 3xTg-AD model in depth. We conclude by offering four suggestions to help unify pre-clinical mouse model AD research inspired by the NIH expectations for considering sex as a biological variable.
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http://dx.doi.org/10.3233/JAD-201014DOI Listing
January 2021

Alcohol use disorder causes global changes in splicing in the human brain.

Transl Psychiatry 2021 01 5;11(1). Epub 2021 Jan 5.

Center for Therapeutic Innovation, University of Miami Miller School of Medicine, Miami, FL, 33136, USA.

Alcohol use disorder (AUD) is a widespread disease leading to the deterioration of cognitive and other functions. Mechanisms by which alcohol affects the brain are not fully elucidated. Splicing constitutes a nuclear process of RNA maturation, which results in the formation of the transcriptome. We tested the hypothesis as to whether AUD impairs splicing in the superior frontal cortex (SFC), nucleus accumbens (NA), basolateral amygdala (BLA), and central nucleus of the amygdala (CNA). To evaluate splicing, bam files from STAR alignments were indexed with samtools for use by rMATS software. Computational analysis of affected pathways was performed using Gene Ontology Consortium, Gene Set Enrichment Analysis, and LncRNA Ontology databases. Surprisingly, AUD was associated with limited changes in the transcriptome: expression of 23 genes was altered in SFC, 14 in NA, 102 in BLA, and 57 in CNA. However, strikingly, mis-splicing in AUD was profound: 1421 mis-splicing events were detected in SFC, 394 in NA, 1317 in BLA, and 469 in CNA. To determine the mechanism of mis-splicing, we analyzed the elements of the spliceosome: small nuclear RNAs (snRNAs) and splicing factors. While snRNAs were not affected by alcohol, expression of splicing factor heat shock protein family A (Hsp70) member 6 (HSPA6) was drastically increased in SFC, BLA, and CNA. Also, AUD was accompanied by aberrant expression of long noncoding RNAs (lncRNAs) related to splicing. In summary, alcohol is associated with genome-wide changes in splicing in multiple human brain regions, likely due to dysregulation of splicing factor(s) and/or altered expression of splicing-related lncRNAs.
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http://dx.doi.org/10.1038/s41398-020-01163-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7790816PMC
January 2021

Molecular mechanisms of psychiatric diseases.

Neurobiol Dis 2020 12 17;146:105136. Epub 2020 Oct 17.

Center for Therapeutic Innovation, University of Miami, Miami, FL, United States of America; Department of Psychiatry and Behavioral Sciences, University of Miami, Miami, FL, United States of America. Electronic address:

For most psychiatric diseases, pathogenetic concepts as well as paradigms underlying neuropsychopharmacologic approaches currently revolve around neurotransmitters such as dopamine, serotonin, and norepinephrine. However, despite the fact that several generations of neurotransmitter-based psychotropics including atypical antipsychotics, selective serotonin reuptake inhibitors, and serotonin-norepinephrine reuptake inhibitors are available, the effectiveness of these medications is limited, and relapse rates in psychiatric diseases are relatively high, indicating potential involvement of other pathogenetic pathways. Indeed, recent high-throughput studies in genetics and molecular biology have shown that pathogenesis of major psychiatric illnesses involves hundreds of genes and numerous pathways via such fundamental processes as DNA methylation, transcription, and splicing. Current review summarizes these and other molecular mechanisms of such psychiatric illnesses as schizophrenia, major depressive disorder, and alcohol use disorder and suggests a conceptual framework for future studies.
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http://dx.doi.org/10.1016/j.nbd.2020.105136DOI Listing
December 2020

Dual Screen for Efficacy and Toxicity Identifies HDAC Inhibitor with Distinctive Activity Spectrum for BAP1-Mutant Uveal Melanoma.

Mol Cancer Res 2021 02 19;19(2):215-222. Epub 2020 Oct 19.

Bascom Palmer Eye Institute, University of Miami Miller School of Medicine, Miami, Florida.

Drug screens leading to successful targeted therapies in cancer have been mainly based on cell viability assays identifying inhibitors of dominantly acting oncogenes. In contrast, there has been little success in discovering targeted therapies that reverse the effects of inactivating mutations in tumor-suppressor genes. BAP1 is one such tumor suppressor that is frequently inactivated in a variety of cancers, including uveal melanoma, renal cell carcinoma, and mesothelioma. Because BAP1 is an epigenetic transcriptional regulator of developmental genes, we designed a two-phase drug screen involving a cell-based rescue screen of transcriptional repression caused by BAP1 loss, followed by an screen of lead compounds for rescue of a BAP1-deficient phenotype with minimal toxicity in embryos. The first screen identified 9 compounds, 8 of which were HDAC inhibitors. The second screen eliminated all except one compound due to inefficacy or toxicity. The resulting lead compound, quisinostat, has a distinctive activity spectrum, including high potency against HDAC4, which was recently shown to be a key target of BAP1. Quisinostat was further validated in a mouse model and found to prevent the growth of BAP1-mutant uveal melanomas. This innovative strategy demonstrates the potential for identifying therapeutic compounds that target tumor-suppressor mutations in cancer. IMPLICATIONS: Few drugs have been identified that target mutations in tumor suppressors. Using a novel 2-step screening approach, strategy, we identified quisinostat as a candidate for therapy in BAP1-mutant uveal melanoma. HDAC4 is implicated as a key target in uveal melanoma and perhaps other BAP1-mutant cancers.
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http://dx.doi.org/10.1158/1541-7786.MCR-20-0434DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7864865PMC
February 2021

The bromodomain inhibitor IBET-151 attenuates vismodegib-resistant esophageal adenocarcinoma growth through reduction of GLI signaling.

Oncotarget 2020 Aug 18;11(33):3174-3187. Epub 2020 Aug 18.

Molecular Oncology Program, Division of Surgical Oncology, Dewitt Daughtry Family Department of Surgery, University of Miami, Miami, FL, USA.

The Hedgehog/GLI (HH/GLI) signaling pathway plays a critical role in human oncogenesis. Unfortunately, the clinical use of HH inhibitor(s) has been associated with serious adverse effects and mutation-related drug resistance. Since the efficacy of SMO (Smoothened) and GLI inhibitors is limited in clinical trials, there remains a critical need for the HH/GLI pathway inhibitors with different mechanisms of action. Here, we show that esophageal adenocarcinoma (EAC) cell lines are insensitive to vismodegib (SMO inhibitor) but respond to GANT61 (GLI1 inhibitor). Furthermore, we examine the role of GLI1 in tumorigenicity of EAC and how a selective bromodomain inhibitor IBET-151 downregulates transcriptional activity of the GLI1 transcription factor in EAC. Our study demonstrates that GLI1 plays an important role in tumorigenicity of EAC and that elevated GLI1 expression in patients' ultrasound-assisted endoscopic biopsy may predict the response to neoadjuvant chemotherapy (NAC) FOLFOX. Importantly, IBET-151 abrogates the growth of vismodegib-resistant EAC cells and downregulates HH/GLI by reducing the occupancy of BRD4 at the GLI1 locus. IBET-151 also attenuates tumor growth of EAC-PDXs and does so in an on-target manner as it reduces the expression of GLI1. We identify HH/GLI signaling as a novel druggable pathway in EAC as well as validate an ability of clinically relevant GLI inhibitor to attenuate the viability of vismodegib-resistant EAC cells. Therefore, we propose that selective bromodomain inhibitors, such as IBET-151, could be used as novel therapeutic agents for EAC patients harboring GLI-dependent tumors.
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http://dx.doi.org/10.18632/oncotarget.27699DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7443367PMC
August 2020

Antitumor activity of the dual BET and CBP/EP300 inhibitor NEO2734.

Blood Adv 2020 09;4(17):4124-4135

Institute of Oncology Research, Faculty of Biomedical Sciences, USI, Bellinzona, Switzerland.

Bromodomain and extra-terminal domain (BET) proteins, cyclic adenosine monophosphate response element-binding protein (CBP), and the E1A-binding protein of p300 (EP300) are important players in histone acetylation. Preclinical evidence supports the notion that small molecules targeting these proteins individually or in combination can elicit antitumor activity. Here, we characterize the antitumor activity of the pan BET/CBP/EP300 inhibitor NEO2734 and provide insights into its mechanism of action through bromodomain-binding assays, in vitro and in vivo treatments of cancer cell lines, immunoblotting, and transcriptome analyses. In a panel of 60 models derived from different tumor types, NEO2734 exhibited antiproliferative activity in multiple cell lines, with the most potent activity observed in hematologic and prostate cancers. Focusing on lymphoma cell lines, NEO2374 exhibited a pattern of response and transcriptional changes similar to lymphoma cells exposed to either BET or CBP/EP300 inhibitors alone. However, NEO2734 was more potent than single-agent BET or CBP/EP300 inhibitors alone. In conclusion, NEO2734 is a novel antitumor compound that shows preferential activity in lymphomas, leukemias, and prostate cancers.
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http://dx.doi.org/10.1182/bloodadvances.2020001879DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7479962PMC
September 2020

A Neurite Outgrowth Assay and Neurotoxicity Assessment with Human Neural Progenitor Cell-Derived Neurons.

J Vis Exp 2020 08 6(162). Epub 2020 Aug 6.

Center for Therapeutic Innovation and Department of Psychiatry & Behavioral Sciences, University of Miami Miller School of Medicine; Persian BayanGene Research and Training Center;

Neurite outgrowth assay and neurotoxicity assessment are two major studies that can be performed using the presented method herein. This protocol provides reliable analysis of neuronal morphology together with quantitative measurements of modifications on neurite length and synaptic protein localization and abundance upon treatment with small molecule compounds. In addition to the application of the presented method in neurite outgrowth studies, neurotoxicity assessment can be performed to assess, distinguish and rank commercial chemical compounds based on their potential developmental neurotoxicity effect. Even though cell lines are nowadays widely used in compound screening assays in neuroscience, they often differ genetically and phenotypically from their tissue origin. Primary cells, on the other hand, maintain important markers and functions observed in vivo. Therefore, due to the translation potential and physiological relevance that these cells could offer neurite outgrowth assay and neurotoxicity assessment can considerably benefit from using human neural progenitor cells (hNPCs) as the primary human cell model. The presented method herein can be utilized to screen for the ability of compounds to induce neurite outgrowth and neurotoxicity by taking advantage of the human neural progenitor cell-derived neurons, a cell model closely representing human biology."
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http://dx.doi.org/10.3791/60955DOI Listing
August 2020

Molecular Transducers of Human Skeletal Muscle Remodeling under Different Loading States.

Cell Rep 2020 08;32(5):107980

Department of Kinesiology, McMaster University, Hamilton, ON, Canada. Electronic address:

Loading of skeletal muscle changes the tissue phenotype reflecting altered metabolic and functional demands. In humans, heterogeneous adaptation to loading complicates the identification of the underpinning molecular regulators. A within-person differential loading and analysis strategy reduces heterogeneity for changes in muscle mass by ∼40% and uses a genome-wide transcriptome method that models each mRNA from coding exons and 3' and 5' untranslated regions (UTRs). Our strategy detects ∼3-4 times more regulated genes than similarly sized studies, including substantial UTR-selective regulation undetected by other methods. We discover a core of 141 genes correlated to muscle growth, which we validate from newly analyzed independent samples (n = 100). Further validating these identified genes via RNAi in primary muscle cells, we demonstrate that members of the core genes were regulators of protein synthesis. Using proteome-constrained networks and pathway analysis reveals notable relationships with the molecular characteristics of human muscle aging and insulin sensitivity, as well as potential drug therapies.
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http://dx.doi.org/10.1016/j.celrep.2020.107980DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7408494PMC
August 2020

Direct Administration and Gene Modulation Using Antisense Oligonucleotides Within the CNS.

Cell Mol Neurobiol 2020 Jul 12. Epub 2020 Jul 12.

Opko Health, Miramar, FL, USA.

Besides his vast contribution to the opioid receptor studies, Dr. G. W. Pasternak was among the early pioneers in the antisense oligonucleotide (ASO) field at the time when the crucial in vivo studies using ASO-mediated gene knockdown in the CNS were still impeded by the ASO's inability to cross the blood-brain barrier. This changed at the start of 1990s, when administration of oligonucleotides through intracerebroventricular or, later, intrathecal injection was undertaken at Cornell University Medical College and further developed in close collaboration with Pasternak lab. These early studies eventually led to the practical realization of the significant therapeutic potential of ASO-based drugs we see today.
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http://dx.doi.org/10.1007/s10571-020-00919-xDOI Listing
July 2020

Dipeptide repeat proteins inhibit homology-directed DNA double strand break repair in C9ORF72 ALS/FTD.

Mol Neurodegener 2020 02 24;15(1):13. Epub 2020 Feb 24.

Department of Psychiatry & Behavioral Sciences, Center for Therapeutic Innovation, University of Miami Miller School of Medicine, 1501 NW 10th Ave, Biomedical Research Building Room 413, Florida, Miami, 33136, USA.

Background: The C9ORF72 hexanucleotide repeat expansion is the most common known genetic cause of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD), two fatal age-related neurodegenerative diseases. The C9ORF72 expansion encodes five dipeptide repeat proteins (DPRs) that are produced through a non-canonical translation mechanism. Among the DPRs, proline-arginine (PR), glycine-arginine (GR), and glycine-alanine (GA) are the most neurotoxic and increase the frequency of DNA double strand breaks (DSBs). While the accumulation of these genotoxic lesions is increasingly recognized as a feature of disease, the mechanism(s) of DPR-mediated DNA damage are ill-defined and the effect of DPRs on the efficiency of each DNA DSB repair pathways has not been previously evaluated.

Methods And Results: Using DNA DSB repair assays, we evaluated the efficiency of specific repair pathways, and found that PR, GR and GA decrease the efficiency of non-homologous end joining (NHEJ), single strand annealing (SSA), and microhomology-mediated end joining (MMEJ), but not homologous recombination (HR). We found that PR inhibits DNA DSB repair, in part, by binding to the nucleolar protein nucleophosmin (NPM1). Depletion of NPM1 inhibited NHEJ and SSA, suggesting that NPM1 loss-of-function in PR expressing cells leads to impediments of both non-homologous and homology-directed DNA DSB repair pathways. By deleting NPM1 sub-cellular localization signals, we found that PR binds NPM1 regardless of the cellular compartment to which NPM1 was directed. Deletion of the NPM1 acidic loop motif, known to engage other arginine-rich proteins, abrogated PR and NPM1 binding. Using confocal and super-resolution immunofluorescence microscopy, we found that levels of RAD52, a component of the SSA repair machinery, were significantly increased iPSC neurons relative to isogenic controls in which the C9ORF72 expansion had been deleted using CRISPR/Cas9 genome editing. Western analysis of post-mortem brain tissues confirmed that RAD52 immunoreactivity is significantly increased in C9ALS/FTD samples as compared to controls.

Conclusions: Collectively, we characterized the inhibitory effects of DPRs on key DNA DSB repair pathways, identified NPM1 as a facilitator of DNA repair that is inhibited by PR, and revealed deficits in homology-directed DNA DSB repair pathways as a novel feature of C9ORF72-related disease.
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http://dx.doi.org/10.1186/s13024-020-00365-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7041170PMC
February 2020

Novel approach reveals genomic landscapes of single-strand DNA breaks with nucleotide resolution in human cells.

Nat Commun 2019 12 20;10(1):5799. Epub 2019 Dec 20.

Institute of Genomics, School of Biomedical Sciences, Huaqiao University, 668 Jimei Road, Xiamen, 361021, China.

Single-strand breaks (SSBs) represent the major form of DNA damage, yet techniques to map these lesions genome-wide with nucleotide-level precision are limited. Here, we present a method, termed SSiNGLe, and demonstrate its utility to explore the distribution and dynamic changes in genome-wide SSBs in response to different biological and environmental stimuli. We validate SSiNGLe using two very distinct sequencing techniques and apply it to derive global profiles of SSBs in different biological states. Strikingly, we show that patterns of SSBs in the genome are non-random, specific to different biological states, enriched in regulatory elements, exons, introns, specific types of repeats and exhibit differential preference for the template strand between exons and introns. Furthermore, we show that breaks likely contribute to naturally occurring sequence variants. Finally, we demonstrate strong links between SSB patterns and age. Overall, SSiNGLe provides access to unexplored realms of cellular biology, not obtainable with current approaches.
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http://dx.doi.org/10.1038/s41467-019-13602-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6925131PMC
December 2019

The novel BET-CBP/p300 dual inhibitor NEO2734 is active in SPOP mutant and wild-type prostate cancer.

EMBO Mol Med 2019 11 26;11(11):e10659. Epub 2019 Sep 26.

Department of Biochemistry and Molecular Biology, Mayo Clinic College of Medicine and Science, Rochester, MN, USA.

CULLIN3-based E3 ubiquitin ligase substrate-binding adaptor gene SPOP is frequently mutated in prostate cancer (PCa). PCa harboring SPOP hotspot mutants (e.g., F133V) are resistant to BET inhibitors because of aberrant elevation of BET proteins. Here, we identified a previously unrecognized mutation Q165P at the edge of SPOP MATH domain in primary and metastatic PCa of a patient. The Q165P mutation causes structural changes in the MATH domain and impairs SPOP dimerization and substrate degradation. Different from F133V hotspot mutant tumors, Q165P mutant patient-derived xenografts (PDXs) and organoids were modestly sensitive to the BET inhibitor JQ1. Accordingly, protein levels of AR, BRD4 and downstream effectors such as RAC1 and phosphorylated AKT were not robustly elevated in Q165P mutant cells as in F133V mutant cells. However, NEO2734, a novel dual inhibitor of BET and CBP/p300, is active in both hotspot mutant (F133V) and non-hotspot mutant (Q165P) PCa cells in vitro and in vivo. These data provide a strong rationale to clinically investigate the anti-cancer efficacy of NEO2734 in SPOP-mutated PCa patients.
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http://dx.doi.org/10.15252/emmm.201910659DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6835201PMC
November 2019

Dysregulation of the histone demethylase KDM6B in alcohol dependence is associated with epigenetic regulation of inflammatory signaling pathways.

Addict Biol 2021 01 1;26(1):e12816. Epub 2019 Aug 1.

Center for Therapeutic Innovation, University of Miami Miller School of Medicine, Miami, Florida, USA.

Epigenetic enzymes oversee long-term changes in gene expression by integrating genetic and environmental cues. While there are hundreds of enzymes that control histone and DNA modifications, their potential roles in substance abuse and alcohol dependence remain underexplored. A few recent studies have suggested that epigenetic processes could underlie transcriptomic and behavioral hallmarks of alcohol addiction. In the present study, we sought to identify epigenetic enzymes in the brain that are dysregulated during protracted abstinence as a consequence of chronic and intermittent alcohol exposure. Through quantitative mRNA expression analysis of over 100 epigenetic enzymes, we identified 11 that are significantly altered in alcohol-dependent rats compared with controls. Follow-up studies of one of these enzymes, the histone demethylase KDM6B, showed that this enzyme exhibits region-specific dysregulation in the prefrontal cortex and nucleus accumbens of alcohol-dependent rats. KDM6B was also upregulated in the human alcoholic brain. Upregulation of KDM6B protein in alcohol-dependent rats was accompanied by a decrease of trimethylation levels at histone H3, lysine 27 (H3K27me3), consistent with the known demethylase specificity of KDM6B. Subsequent epigenetic (chromatin immunoprecipitation [ChIP]-sequencing) analysis showed that alcohol-induced changes in H3K27me3 were significantly enriched at genes in the IL-6 signaling pathway, consistent with the well-characterized role of KDM6B in modulation of inflammatory responses. Knockdown of KDM6B in cultured microglial cells diminished IL-6 induction in response to an inflammatory stimulus. Our findings implicate a novel KDM6B-mediated epigenetic signaling pathway integrated with inflammatory signaling pathways that are known to underlie the development of alcohol addiction.
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http://dx.doi.org/10.1111/adb.12816DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7757263PMC
January 2021

Ovarian Cancer Treatment Stratification Using Drug Sensitivity Testing.

Anticancer Res 2019 Aug;39(8):4023-4030

Center for Therapeutic Innovation, Miller School of Medicine, University of Miami, Miami, FL, U.S.A.

Background: Treatment options for patients with platinum-resistant ovarian cancer are generally palliative in nature and rarely have realistic potential to be curative. Because many patients with recurrent ovarian cancer receive aggressive chemotherapy for prolonged periods, sometimes continuously, therapy-related toxicities are a major factor in treatment decisions. The use of ex vivo drug sensitivity screens has the potential to improve the treatment of patients with platinum-resistant ovarian cancer by providing personalized treatment plans and thus reducing toxicity from unproductive therapy attempts.

Materials And Methods: We evaluated the treatment responses of a set of six early-passage patient-derived ovarian cancer cell lines towards a set of 30 Food and Drug Administration-approved chemotherapy drugs using drug-sensitivity testing.

Results: We observed a wide range of treatment responses of the cell lines. While most compounds displayed vastly different treatment responses between cell lines, we found that some compounds such as docetaxel and cephalomannine reduced cell survival of all cell lines.

Conclusion: We propose that ex vivo drug-sensitivity screening holds the potential to greatly improve patient outcomes, especially in a population where multiple continuous treatments are not an option due to advanced disease, rapid disease progression, age or poor overall health. This approach may also be useful to identify potential novel therapeutics for patients with ovarian cancer.
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http://dx.doi.org/10.21873/anticanres.13558DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7323502PMC
August 2019

Defective HNF4alpha-dependent gene expression as a driver of hepatocellular failure in alcoholic hepatitis.

Nat Commun 2019 07 16;10(1):3126. Epub 2019 Jul 16.

UCL Institute for Liver and Digestive Health, Division of Medicine, Royal Free Campus, University College London, London, WC1E 6BT, UK.

Alcoholic hepatitis (AH) is a life-threatening condition characterized by profound hepatocellular dysfunction for which targeted treatments are urgently needed. Identification of molecular drivers is hampered by the lack of suitable animal models. By performing RNA sequencing in livers from patients with different phenotypes of alcohol-related liver disease (ALD), we show that development of AH is characterized by defective activity of liver-enriched transcription factors (LETFs). TGFβ1 is a key upstream transcriptome regulator in AH and induces the use of HNF4α P2 promoter in hepatocytes, which results in defective metabolic and synthetic functions. Gene polymorphisms in LETFs including HNF4α are not associated with the development of AH. In contrast, epigenetic studies show that AH livers have profound changes in DNA methylation state and chromatin remodeling, affecting HNF4α-dependent gene expression. We conclude that targeting TGFβ1 and epigenetic drivers that modulate HNF4α-dependent gene expression could be beneficial to improve hepatocellular function in patients with AH.
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http://dx.doi.org/10.1038/s41467-019-11004-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6635373PMC
July 2019

Longevity-related molecular pathways are subject to midlife "switch" in humans.

Aging Cell 2019 08 6;18(4):e12970. Epub 2019 Jun 6.

Department of Psychiatry and Behavioral Sciences, Center for Therapeutic Innovation, University of Miami Miller School of Medicine, Miami, Florida.

Emerging evidence indicates that molecular aging may follow nonlinear or discontinuous trajectories. Whether this occurs in human neuromuscular tissue, particularly for the noncoding transcriptome, and independent of metabolic and aerobic capacities, is unknown. Applying our novel RNA method to quantify tissue coding and long noncoding RNA (lncRNA), we identified ~800 transcripts tracking with age up to ~60 years in human muscle and brain. In silico analysis demonstrated that this temporary linear "signature" was regulated by drugs, which reduce mortality or extend life span in model organisms, including 24 inhibitors of the IGF-1/PI3K/mTOR pathway that mimicked, and 5 activators that opposed, the signature. We profiled Rapamycin in nondividing primary human myotubes (n = 32 HTA 2.0 arrays) and determined the transcript signature for reactive oxygen species in neurons, confirming that our age signature was largely regulated in the "pro-longevity" direction. Quantitative network modeling demonstrated that age-regulated ncRNA equaled the contribution of protein-coding RNA within structures, but tended to have a lower heritability, implying lncRNA may better reflect environmental influences. Genes ECSIT, UNC13, and SKAP2 contributed to a network that did not respond to Rapamycin, and was associated with "neuron apoptotic processes" in protein-protein interaction analysis (FDR = 2.4%). ECSIT links inflammation with the continued age-related downwards trajectory of mitochondrial complex I gene expression (FDR < 0.01%), implying that sustained inhibition of ECSIT may be maladaptive. The present observations link, for the first time, model organism longevity programs with the endogenous but temporary genome-wide responses to aging in humans, revealing a pattern that may ultimately underpin personalized rates of health span.
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http://dx.doi.org/10.1111/acel.12970DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6612641PMC
August 2019

Vitamin C supplementation expands the therapeutic window of BETi for triple negative breast cancer.

EBioMedicine 2019 May 8;43:201-210. Epub 2019 Apr 8.

John P. Hussman Institute for Human Genomics, Dr. John T. Macdonald Foundation Department of Human Genetics, University of Miami Miller School of Medicine, Miami, FL 33136, USA; Sylvester Comprehensive Cancer Center, University of Miami Miller School of Medicine, Miami, FL 33136, USA. Electronic address:

Background: Bromodomain and extra-terminal inhibitors (BETi) have shown efficacy for the treatment of aggressive triple negative breast cancer (TNBC). However, BETi are plagued by a narrow therapeutic window as manifested by severe toxicities at effective doses. Therefore, it is a limitation to their clinical implementation in patient care.

Methods: The impact of vitamin C on the efficacy of small compounds including BETi was assessed by high-throughput screening. Co-treatment of TNBC by BETi especially JQ1 and vitamin C was evaluated in vitro and in vivo.

Findings: High-throughput screening revealed that vitamin C improves the efficacy of a number of structurally-unrelated BETi including JQ1, I-BET762, I-BET151, and CPI-203 in treating TNBC cells. The synergy between BETi and vitamin C is due to suppressed histone acetylation (H3ac and H4ac), which is in turn caused by upregulated histone deacetylase 1 (HDAC1) expression upon vitamin C addition. Treatment with JQ1 at lower doses together with vitamin C induces apoptosis and inhibits the clonogenic ability of cultured TNBC cells. Oral vitamin C supplementation renders a sub-therapeutic dose of JQ1 able to inhibit human TNBC xenograft growth and metastasis in mice.

Interpretation: Vitamin C expands the therapeutic window of BETi by sensitizing TNBC to BETi. Using vitamin C as a co-treatment, lower doses of BETi could be used to achieve an increased therapeutic index in patients, which will translate to a reduced side effect profile. FUND: University of Miami Sylvester Comprehensive Cancer Center, Bankhead Coley Cancer Research program (7BC10), Flight Attendant Medical Research Institute, and NIH R21CA191668 (to GW) and 1R56AG061911 (to CW and CHV).
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http://dx.doi.org/10.1016/j.ebiom.2019.04.006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6557781PMC
May 2019

HDAC Inhibitors Induce Expression and Promote Neurite Outgrowth in Human Neural Progenitor Cells-Derived Neurons.

Int J Mol Sci 2019 Mar 5;20(5). Epub 2019 Mar 5.

Center for Therapeutic Innovation and Department of Psychiatry & Behavioral Sciences, University of Miami Miller School of Medicine, Miami, FL 33136, USA.

Besides its key role in neural development, brain-derived neurotrophic factor (BDNF) is important for long-term potentiation and neurogenesis, which makes it a critical factor in learning and memory. Due to the important role of BDNF in synaptic function and plasticity, an in-house epigenetic library was screened against human neural progenitor cells (HNPCs) and WS1 human skin fibroblast cells using Cell-to-Ct assay kit to identify the small compounds capable of modulating the expression. In addition to two well-known hydroxamic acid-based histone deacetylase inhibitors (hb-HDACis), SAHA and TSA, several structurally similar HDAC inhibitors including SB-939, PCI-24781 and JNJ-26481585 with even higher impact on expression, were discovered in this study. Furthermore, by using well-developed immunohistochemistry assays, the selected compounds were also proved to have neurogenic potential improving the neurite outgrowth in HNPCs-derived neurons. In conclusion, we proved the neurogenic potential of several hb-HDACis, alongside their ability to enhance expression, which by modulating the neurogenesis and/or compensating for neuronal loss, could be propitious for treatment of neurological disorders.
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http://dx.doi.org/10.3390/ijms20051109DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6429164PMC
March 2019

BET Bromodomain Inhibitors Which Permit Treg Function Enable a Combinatorial Strategy to Suppress GVHD in Pre-clinical Allogeneic HSCT.

Front Immunol 2018 24;9:3104. Epub 2019 Jan 24.

Department of Microbiology and Immunology, Miller School of Medicine, University of Miami, Miami, FL, United States.

A recent approach for limiting production of pro-inflammatory cytokines has been to target bromodomain and extra-terminal (BET) proteins. These epigenetic readers of histone acetylation regulate transcription of genes involved in inflammation, cardiovascular disease, and cancer. Development of BET inhibitors (BETi) has generated enormous interest for their therapeutic potential. Because inflammatory signals and donor T cells promote graft-versus-host disease (GVHD), regulating both pathways could be effective to abrogate this disorder. The objective of the present study was to identify a BETi which did not interfere with CD4FoxP3 regulatory T cell (Treg) expansion and function to utilize together with Tregs following allogeneic hematopoietic stem cell transplantation (aHSCT) to ameliorate GVHD. We have reported that Tregs can be markedly expanded and selectively activated with increased functional capacity by targeting TNFRSF25 and CD25 with TL1A-Ig and low dose IL-2, respectively. Here, mice were treated over 7 days (TL1A-Ig + IL-2) together with BETi. We found that the BETi EP11313 did not decrease frequency/numbers or phenotype of expanded Tregs as well as effector molecules, such as IL-10 and TGF-β. However, BETi JQ1 interfered with Treg expansion and altered subset distribution and phenotype. Notably, in Treg expanded mice, EP11313 diminished tnfa and ifng but not il-2 RNA levels. Remarkably, Treg pSTAT5 expression was not affected by EP11313 supporting the notion that Treg IL-2 signaling remained intact. MHC-mismatched aHSCT (B6 → BALB/c) was performed using expanded donor Tregs with or without EP11313 short-term treatment in the recipient. Early post-transplant, improvement in the splenic and LN CD4/CD8 ratio along with fewer effector cells and high Treg levels in aHSCT recipients treated with expanded Tregs + EP11313 was detected. Interestingly, this group exhibited a significant diminution of GVHD clinical score with less skin and ocular involvement. Finally, using low numbers of highly purified expanded Tregs, improved clinical GVHD scores were observed in EP11313 treated recipients. In total, we conclude that use of this novel combinatorial strategy can suppress pre-clinical GVHD and posit, EP11313 treatment might be useful combined with Treg expansion therapy for treatment of diseases involving inflammatory responses.
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http://dx.doi.org/10.3389/fimmu.2018.03104DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6353853PMC
October 2019

Precision medicine in the treatment stratification of AML patients: challenges and progress.

Oncotarget 2018 Dec 28;9(102):37790-37797. Epub 2018 Dec 28.

Center for Therapeutic Innovation, Miller School of Medicine, University of Miami, Miami, FL, USA.

Recent advances in high throughput technologies have led to the generation of vast amounts of clinical data and the development of personalized medicine approaches in acute myeloid leukemia (AML). The ability to treat cancer patients based upon their individual molecular characteristics or drug sensitivity profiles is expected to significantly advance cancer treatment and improve the long-term survival of patients with refractory AML, for whom current treatment options are restricted to palliative approaches. The clinical development of omics-based and phenotypic screens, however, is limited by a number of bottlenecks including the generation of cost-effective high-throughput data, data interpretation and integration of multiple approaches, sample availability, clinically relevant timelines, and the development and education of multidisciplinary teams. Recently, a number of small clinical trials have shown survival benefits in patients treated based on personalized medicine approaches. While these preliminary studies are encouraging, larger trials are needed to evaluate the utility of these technologies in routine clinical settings.
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http://dx.doi.org/10.18632/oncotarget.26492DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6340870PMC
December 2018

Purification of H3 and H4 Histone Proteins and the Quantification of Acetylated Histone Marks in Cells and Brain Tissue.

J Vis Exp 2018 11 30(141). Epub 2018 Nov 30.

Department of Psychiatry & Behavioral Sciences, University of Miami Miller School of Medicine; Center for Therapeutic Innovation, University of Miami Miller School of Medicine;

In all eukaryotic organisms, chromatin, the physiological template of all genetic information, is essential for heredity. Chromatin is subject to an array of diverse posttranslational modifications (PTMs) that mostly occur in the amino termini of histone proteins (i.e., histone tail) and regulate the accessibility and functional state of the underlying DNA. Histone tails extend from the core of the nucleosome and are subject to the addition of acetyl groups by histone acetyltransferases (HATs) and the removal of acetyl groups by histone deacetylases (HDACs) during cellular growth and differentiation. Specific acetylation patterns on lysine (K) residues on histone tails determine a dynamic homeostasis between transcriptionally active or transcriptionally repressed chromatin by (1) influencing the core histone assembly and (2) recruiting synergistic or antagonistic chromatin-associated proteins to the transcription site. The fundamental regulatory mechanism of the complex nature of histone tail PTMs influences the majority of chromatin-templated processes and results in changes in cell maturation and differentiation in both normal and pathological development. The goal of the current report is to provide novices with an efficient method to purify core histone proteins from cells and brain tissue and to reliably quantify acetylation marks on histones H3 and H4.
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http://dx.doi.org/10.3791/58648DOI Listing
November 2018

Drug and disease signature integration identifies synergistic combinations in glioblastoma.

Nat Commun 2018 12 14;9(1):5315. Epub 2018 Dec 14.

Center for Therapeutic Innovation, Department of Psychiatry and Behavioral Sciences, University of Miami Miller School of Medicine, 1120 NW 14th St, Miami, FL, 33136, USA.

Glioblastoma (GBM) is the most common primary adult brain tumor. Despite extensive efforts, the median survival for GBM patients is approximately 14 months. GBM therapy could benefit greatly from patient-specific targeted therapies that maximize treatment efficacy. Here we report a platform termed SynergySeq to identify drug combinations for the treatment of GBM by integrating information from The Cancer Genome Atlas (TCGA) and the Library of Integrated Network-Based Cellular Signatures (LINCS). We identify differentially expressed genes in GBM samples and devise a consensus gene expression signature for each compound using LINCS L1000 transcriptional profiling data. The SynergySeq platform computes disease discordance and drug concordance to identify combinations of FDA-approved drugs that induce a synergistic response in GBM. Collectively, our studies demonstrate that combining disease-specific gene expression signatures with LINCS small molecule perturbagen-response signatures can identify preclinical combinations for GBM, which can potentially be tested in humans.
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http://dx.doi.org/10.1038/s41467-018-07659-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6294341PMC
December 2018

Enhancement of BDNF Expression and Memory by HDAC Inhibition Requires BET Bromodomain Reader Proteins.

J Neurosci 2019 01 30;39(4):612-626. Epub 2018 Nov 30.

Center for Therapeutic Innovation and Department of Psychiatry and Behavioral Sciences, Miller School of Medicine, University of Miami, Miami, Florida 33136,

Histone deacetylase (HDAC) inhibitors may have therapeutic utility in multiple neurological and psychiatric disorders, but the underlying mechanisms remain unclear. Here, we identify BRD4, a BET bromodomain reader of acetyl-lysine histones, as an essential component involved in potentiated expression of brain-derived neurotrophic factor (BDNF) and memory following HDAC inhibition. In studies, we reveal that pharmacological inhibition of BRD4 reversed the increase in mRNA induced by the class I/IIb HDAC inhibitor suberoylanilide hydroxamic acid (SAHA). Knock-down of HDAC2 and HDAC3, but not other HDACs, increased mRNA expression, whereas knock-down of BRD4 blocked these effects. Using dCas9-BRD4, locus-specific targeting of BRD4 to the promoter increased mRNA. In additional studies, RGFP966, a pharmacological inhibitor of HDAC3, elevated expression and BRD4 binding to the BDNF promoter, effects that were abrogated by JQ1 (an inhibitor of BRD4). Examining known epigenetic targets of BRD4 and HDAC3, we show that H4K5ac and H4K8ac modifications and H4K5ac enrichment at the promoter were elevated following RGFP966 treatment. In electrophysiological studies, JQ1 reversed RGFP966-induced enhancement of LTP in hippocampal slice preparations. Last, in behavioral studies, RGFP966 increased subthreshold novel object recognition memory and cocaine place preference in male C57BL/6 mice, effects that were reversed by cotreatment with JQ1. Together, these data reveal that BRD4 plays a key role in HDAC3 inhibitor-induced potentiation of expression, neuroplasticity, and memory. Some histone deacetylase (HDAC) inhibitors are known to have neuroprotective and cognition-enhancing properties, but the underlying mechanisms have yet to be fully elucidated. In the current study, we reveal that BRD4, an epigenetic reader of histone acetylation marks, is necessary for enhancing brain-derived neurotrophic factor (BDNF) expression and improved memory following HDAC inhibition. Therefore, by identifying novel epigenetic regulators of BDNF expression, these data may lead to new therapeutic targets for the treatment of neuropsychiatric disorders.
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http://dx.doi.org/10.1523/JNEUROSCI.1604-18.2018DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6343644PMC
January 2019

Inhibition of HDAC3 reverses Alzheimer's disease-related pathologies in vitro and in the 3xTg-AD mouse model.

Proc Natl Acad Sci U S A 2018 11 5;115(47):E11148-E11157. Epub 2018 Nov 5.

Department of Psychiatry & Behavioral Sciences, University of Miami Miller School of Medicine, Miami, FL 33136;

Alzheimer's disease (AD) is the leading cause of age-related dementia. Neuropathological hallmarks of AD include brain deposition of β-amyloid (Aβ) plaques and accumulation of both hyperphosphorylated and acetylated tau. RGFP-966, a brain-penetrant and selective HDAC3 inhibitor, or HDAC3 silencing, increases BDNF expression, increases histone H3 and H4 acetylation, decreases tau phosphorylation and tau acetylation at disease-associated sites, reduces β-secretase cleavage of the amyloid precursor protein (APP), and decreases Aβ accumulation in HEK-293 cells overexpressing APP with the double Swedish mutation (HEK/APP). In the triple transgenic AD mouse model (3xTg-AD), repeated administration of 3 and 10 mg/kg of RGFP-966 reverses pathological tau phosphorylation at Thr, Ser, and Ser, increases levels of the Aβ degrading enzyme Neprilysin in plasma, decreases Aβ protein levels in the brain and periphery, and improves spatial learning and memory. Finally, we show that RGFP-966 decreases Aβ accumulation and both tau acetylation and phosphorylation at disease residues in neurons derived from induced pluripotent stem cells obtained from APOEε4-carrying AD patients. These data indicate that HDAC3 plays an important regulatory role in the expression and regulation of proteins associated with AD pathophysiology, supporting the notion that HDAC3 may be a disease-modifying therapeutic target.
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http://dx.doi.org/10.1073/pnas.1805436115DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6255210PMC
November 2018

Ex vivo drug sensitivity testing as a means for drug repurposing in esophageal adenocarcinoma.

PLoS One 2018 13;13(9):e0203173. Epub 2018 Sep 13.

Center for Therapeutic Innovation, Miller School of Medicine, University of Miami, Miami, Florida, United States of America.

Background: Esophageal cancer remains one of the hardest cancers to treat with rising incidence rates, low overall survival and high levels of treatment resistance. The lack of clinically available biomarkers hinder diagnosis and treatment stratification. While large scale sequencing approaches have uncovered a number of molecular makers, little has translated in the routine treatment of esophageal cancer patients.

Material And Methods: We evaluate the treatment response towards a panel of 215 FDA-approved and 163 epigenetic compounds of 4 established and 2 patient-derived esophageal cancer cell lines. Cell viability was evaluated after 72h of treatment using cell titer glow. The drug sensitivity testing results for gemcitabine and cisplatin were validated using clonogenic assays.

Results: The tested cell lines display different drug sensitivity profiles, although we found compounds that display efficacy in all of the tested established or patient-derived cell lines. Clonogenic assays confirmed the validity of the drug sensitivity testing results. Using the epigenetic library, we observed high sensitivity towards a number of epigenetic modifiers.

Discussion: Ex vivo drug sensitivity testing may present a viable option for the treatment stratification of esophageal cancer patients and holds the potential to greatly improve patient outcome while reducing treatment toxicity.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0203173PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6136712PMC
February 2019

EZH1 is an antipsychotic-sensitive epigenetic modulator of social and motivational behavior that is dysregulated in schizophrenia.

Neurobiol Dis 2018 11 9;119:149-158. Epub 2018 Aug 9.

The Center for Therapeutic Innovation and Department of Psychiatry & Behavioral Sciences, University of Miami Miller School of Medicine, Miami, FL, USA. Electronic address:

Background: With the capacity to modulate gene networks in an environmentally-sensitive manner, the role of epigenetic systems in mental disorders has come under intense investigation. Dysregulation of epigenetic effectors, including microRNAs and histone-modifying enzymes, may better explain the role of environmental risk factors and the observed heritability rate that cannot be fully attributed to known genetic risk alleles. Here, we aimed to identify novel epigenetic targets of the schizophrenia-associated microRNA 132 (miR-132).

Methods: Histone modifications were quantified by immunodetection in response to viral-mediated overexpression of miR-132 while a luminescent reporter system was used to validate targets of miR-132 in vitro. Genome-wide profiling, quantitative PCR and NanoSting were used to quantify gene expression in post-mortem human brains, neuronal cultures and prefrontal cortex (PFC) of mice chronically exposed to antipsychotics. Following viral-mediated depletion of Enhancer of Zeste 1 (EZH1) in the murine PFC, behaviors including sociability and motivation were assessed using a 3-chambered apparatus and forced-swim test, respectively.

Results: Overexpression of miR-132 decreased global histone 3 lysine 27 tri-methylation (H3K27me3), a repressive epigenetic mark. Moreover, the polycomb-associated H3K27 methyltransferase, EZH1, is regulated by miR-132 and upregulated in the PFC of schizophrenics. Unlike its homolog EZH2, expression of EZH1 in the murine PFC decreased following chronic exposure to antipsychotics. Viral-mediated depletion of EZH1 in the mouse PFC attenuated sociability, enhanced motivational behaviors, and affected gene expression pathways related to neurotransmission and behavioral phenotypes.

Conclusions: EZH1 is dysregulated in schizophrenia, sensitive to antipsychotic medications, and a brain-enriched miR-132 target that controls neurobehavioral phenotypes.
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http://dx.doi.org/10.1016/j.nbd.2018.08.005DOI Listing
November 2018

Developments in lncRNA drug discovery: where are we heading?

Expert Opin Drug Discov 2018 09 6;13(9):837-849. Epub 2018 Aug 6.

a Center for Therapeutic Innovation and Department of Psychiatry and Behavioral Sciences , University of Miami Miller School of Medicine , Miami , FL , USA.

Introduction: The central dogma of molecular biology, which states that the only role of long RNA transcripts is to convey information from gene to protein, was brought into question in recent years due to discovery of the extensive presence and complex roles of long noncoding RNAs (lncRNAs). Furthermore, lncRNAs were found to be involved in pathogenesis of multiple diseases and thus represent a new class of therapeutic targets. Translational efforts in the lncRNA field have been augmented by progress in optimizing the chemistry and delivery platforms of lncRNA-targeting modalities, including oligonucleotide-based drugs and CRISPR-Cas9. Areas covered: This review covers the current advances in characterizing diversity and biological functions of lncRNA focusing on their therapeutic potential in selected therapeutic areas. Expert opinion: Due to accelerating parallel progress in lncRNA biology and lncRNA-compatible therapeutic modalities, it is likely that lncRNA-dependent mechanisms of pathogenesis will soon be targeted in various disorders, including neurological, psychiatric, cardiovascular, infectious diseases, and cancer. Significant efforts, however, are still required to better understand the biology of both lncRNAs and lncRNA-targeting drugs. Further work is needed in the areas of lncRNA nomenclature, database representation, intra/interfield communication, and education of the community at large.
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http://dx.doi.org/10.1080/17460441.2018.1501024DOI Listing
September 2018

Strategies to Annotate and Characterize Long Noncoding RNAs: Advantages and Pitfalls.

Trends Genet 2018 09 17;34(9):704-721. Epub 2018 Jul 17.

Institute of Genomics, School of Biomedical Sciences, Huaqiao University, 201 Pan-Chinese S & T Building, 668 Jimei Road, Xiamen 361021, China. Electronic address:

The past decade has seen an explosion of interest in long noncoding RNAs (lncRNAs). However, despite the massive volume of scientific data implicating these transcripts in a plethora of molecular and cellular processes, a great deal of controversy surrounds these RNAs. One of the main reasons for this lies in the multiple unique features of lncRNAs which limit the available methods used to characterize them. Combined with their vast numbers and inadequate classification, comprehensive annotation of these transcripts becomes a daunting task. The solution to this complex challenge likely lies in deep understanding of the strengths and weaknesses of each computational and empirical approach, and integration of multiple strategies to reduce noise, authenticate the results, and classify lncRNAs. We review here both the advantages and caveats of strategies commonly used for functional characterization and annotation of lncRNAs in the context of emerging conceptual guidelines for their application.
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http://dx.doi.org/10.1016/j.tig.2018.06.002DOI Listing
September 2018

A coding and non-coding transcriptomic perspective on the genomics of human metabolic disease.

Nucleic Acids Res 2018 09;46(15):7772-7792

Duke University School of Medicine, Durham, USA.

Genome-wide association studies (GWAS), relying on hundreds of thousands of individuals, have revealed >200 genomic loci linked to metabolic disease (MD). Loss of insulin sensitivity (IS) is a key component of MD and we hypothesized that discovery of a robust IS transcriptome would help reveal the underlying genomic structure of MD. Using 1,012 human skeletal muscle samples, detailed physiology and a tissue-optimized approach for the quantification of coding (>18,000) and non-coding (>15,000) RNA (ncRNA), we identified 332 fasting IS-related genes (CORE-IS). Over 200 had a proven role in the biochemistry of insulin and/or metabolism or were located at GWAS MD loci. Over 50% of the CORE-IS genes responded to clinical treatment; 16 quantitatively tracking changes in IS across four independent studies (P = 0.0000053: negatively: AGL, G0S2, KPNA2, PGM2, RND3 and TSPAN9 and positively: ALDH6A1, DHTKD1, ECHDC3, MCCC1, OARD1, PCYT2, PRRX1, SGCG, SLC43A1 and SMIM8). A network of ncRNA positively related to IS and interacted with RNA coding for viral response proteins (P < 1 × 10-48), while reduced amino acid catabolic gene expression occurred without a change in expression of oxidative-phosphorylation genes. We illustrate that combining in-depth physiological phenotyping with robust RNA profiling methods, identifies molecular networks which are highly consistent with the genetics and biochemistry of human metabolic disease.
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http://dx.doi.org/10.1093/nar/gky570DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6125682PMC
September 2018