The rockefeller university

In this paper, the hepatitis C virus inhibitors Pibrentasvir and Ombitasvir are found to inhibit the SARS-CoV-2 exonuclease and are shown to have therapeutic potential when combined with SARS-CoV-2 polymerase inhibitors in viral cell cultures. SARS-CoV-2 has an exonuclease-based proofreader, which removes nucleotide inhibitors such as Remdesivir that are incorporated into the viral RNA during replication, reducing the efficacy of these drugs for treating COVID-19. Combinations of inhibitors of both the viral RNA-dependent RNA polymerase and the exonuclease could overcome this deficiency. Here we report the identification of hepatitis C virus NS5A inhibitors Pibrentasvir and Ombitasvir as SARS-CoV-2 exonuclease inhibitors. In the presence of Pibrentasvir, RNAs terminated with the active forms of the prodrugs Sofosbuvir, Remdesivir, Favipiravir, Molnupiravir and AT-527 were largely protected from excision by the exonuclease, while in the absence of Pibrentasvir, there was rapid excision. Due to its unique structure, Tenofovir-terminated RNA was highly resistant to exonuclease excision even in the absence of Pibrentasvir. Viral cell culture studies also demonstrate significant synergy using this combination strategy. This study supports the use of combination drugs that inhibit both the SARS-CoV-2 polymerase and exonuclease for effective COVID-19 treatment.

Public Health Significance Persons with dual severe opioid and cocaine use disorders are at particular risk of morbidity and mortality. This study shows that in persons with such dual diagnoses, there is a shared vulnerability to escalation of both heroin and cocaine. Persons with dual severe opioid and cocaine use disorders are at risk of considerable morbidity, and the bidirectional relationship of escalation of mu-opioid agonists and cocaine use is not well understood. The aim of this study was to examine the bidirectional relationship between escalation of heroin and cocaine use in volunteers dually diagnosed with opioid and cocaine dependence (OD + CD). Volunteers from New York with OD + CD (total n = 295; male = 182, female = 113; age >= 18 years) were interviewed with the Structured Clinical Interview for the DSM-IV Axis I Disorders and Kreek-McHugh-Schluger-Kellogg scales for dimensional measures of drug exposure, which also collect ages of 1st use and onset of heaviest use. Time of escalation was defined as age of onset of heaviest use minus age of 1st use in whole years. Times of escalation of heroin and cocaine were positively correlated in both men (Spearman r = .34, 95% confidence interval [CI: .17, .48], p < .0001) and women (Spearman r = .51, [.27, .50], p < .0001) volunteers. After we adjusted for demographic variables, a Cox regression showed that time of cocaine escalation was a predictor of time of heroin escalation (hazard ratio [HR] = 0.97, 95% CI [0.95, 0.99], p = .003). Another Cox regression showed that this relationship is bidirectional, because time of heroin escalation was also a predictor of time of cocaine escalation (HR = 0.98, [0.96-0.99], p = .016). In these adjusted models, gender was not a significant predictor of time of escalation of either heroin or cocaine. Therefore, escalation did not differ robustly by gender when adjusting for demographics and other major variables. Overall, rapid escalation of cocaine use was a predictor of rapid escalation of heroin use, and vice versa, in persons with dual severe opioid and cocaine use disorders. These findings suggest a shared vulnerability to rapid escalation of these 2 drugs in persons with dual severe opioid and cocaine use disorders.

Catecholaminergic polymorphic ventricular tachycardia (CPVT) is an arrhythmia syndrome caused by gene mutations that render RYR2 Ca release channels hyperactive, provoking spontaneous Ca release and delayed afterdepolarizations (DADs). What remains unknown is the cellular source of ventricular arrhythmia triggered by DADs: Purkinje cells in the conduction system or ventricular cardiomyocytes in the working myocardium. To answer this question, we used a genetic approach in mice to knock out cardiac calsequestrin either in Purkinje cells or in ventricular cardiomyocytes. Total loss of calsequestrin in the heart causes a severe CPVT phenotype in mice and humans. We found that loss of calsequestrin only in ventricular myocytes produced a full-blown CPVT mice. Subendocardial chemical ablation or restoration of calsequestrin expression in subendocardial cardiomyocytes neighboring Purkinje cells was sufficient to protect against catecholamine-induced arrhythmias. In silico modeling demonstrated that DADs in ventricular myocardium can trigger full generated at the Purkinje-myocardial junction via a heretofore unrecognized tissue mechanism, whereby DADs in the ventricular myocardium trigger full action potentials in adjacent Purkinje cells.

A sarcomere is the contractile unit of the myofibril in striated muscles such as cardiac and skeletal muscles. The assembly of sarcomeres depends on multiple molecules that serve as raw materials and participate in the assembly process. However, the mechanism of this critical assembly process remains largely unknown. Here, we found that the cell fate determinant Numb and its homolog Numblike regulated sarcomere assembly and maintenance in striated muscles. We discovered that Numb and Numblike are sarcomeric molecules that were gradually confined to the Z-disc during striated muscle development. Conditional knockout of Numb and Numblike severely compromised sarcomere assembly and its integrity and thus caused organelle dysfunction. Notably, we identified that Numb and Numblike served as sarcomeric alpha-Actin-binding proteins (ABPs) and shared a conserved domain that can bind to the barbed end of sarcomeric alpha-Actin. In vitro fluorometric alpha-Actin polymerization assay showed that Numb and Numblike also played a role in the sarcomeric alpha-Actin polymerization process. Last, we demonstrate that Numb and Numblike regulate sarcomeric alpha-Actinin-dependent (ACTN-dependent) Z-disc consolidation in the sarcomere assembly and maintenance. In summary, our studies show that Numb and its homolog Numblike regulate sarcomere assembly and maintenance in striated muscles, and demonstrate a molecular mechanism by which Numb/Numblike, sarcomeric alpha-Actin, and ACTN cooperate to control thin filament formation and Z-disc consolidation.

Background: Although hidradenitis suppurativa (HS) shares some transcriptomic and cellular infiltrate features with psoriasis, their skin proteome remains unknown. Objective: To define and compare inflammatory protein biomarkers of HS and psoriasis skin. Methods: We assessed 92 inflammatory biomarkers in HS (n = 13), psoriasis (n = 11), and control skin (n = 11) using Olink high-throughput proteomics. We also correlated HS skin and blood biomarkers using proteomics and RNA sequencing. Results: We identified 57 differentially expressed proteins (DEPs) in lesional psoriasis and 64 DEPs in lesional HS skin, compared to healthy controls. Both HS and psoriasis lesional skin demonstrated a significant upregulation of T helper 1 and T helper 17 proteins. Healthy-appearing perilesional HS skin had 63 DEPs compared to healthy controls. Nonlesional HS and psoriasis skin had 24 and 7 DEPs, respectively, compared to healthy controls. Tumor necrosis factor and 8 other proteins were significantly correlated with clinical severity in perilesional HS skin (2 cm from a nodule). Limitations: Inclusion of only moderate-to-severe patients and the cohort size. Conclusion: HS has a greater inflammatory profile and is more diffusely distributed compared with psoriasis. Proteins correlated with disease severity are potential disease mediators. Perilesional skin is comparably inflamed to lesional skin, suggesting the need to treat beyond skin nodules. ( J Am Acad Dermatol 2022;86:322-30.)

Chemical synthesis of secondary metabolites isolated from nature, and derivatives thereof, is still a paradigm of significance to drug development. Here the authors instead use bioinformatics to analyze a biosynthetic gene cluster found in the soil metagenome, and chemical synthesis of its predict product to produce lapcin, a dual topoisomerase I/II inhibitor with promising activity against cancer cell lines. In natural product discovery programs, the power of synthetic chemistry is often leveraged for the total synthesis and diversification of characterized metabolites. The synthesis of structures that are bioinformatically predicted to arise from uncharacterized biosynthetic gene clusters (BGCs) provides a means for synthetic chemistry to enter this process at an early stage. The recent identification of non-ribosomal peptides (NRPs) containing multiple rho-aminobenzoic acids (PABAs) led us to search soil metagenomes for BGCs that polymerize PABA. Here, we use PABA-specific adenylation-domain sequences to guide the cloning of the lap BGC directly from soil. This BGC was predicted to encode a unique N-acylated PABA and thiazole containing structure. Chemical synthesis of this structure gave lapcin, a dual topoisomerase I/II inhibitor with nM to pM IC50s against diverse cancer cell lines. The discovery of lapcin highlights the power of coupling metagenomics, bioinformatics and total chemical synthesis to unlock the biosynthetic potential contained in even complex uncharacterized BGCs.

Retinal degenerative diseases such as retinitis pigmentosa (RP) are of the major causes of vision loss in developed countries. Despite the unclear pathophysiology, treatment methods have been investigated vastly in the past decades. This review article mainly discusses the advances in application of stem cell and progenitor transplantation for retinitis pigmentosa. Stem cell sources such as mesenchymal stem cells, embryonic stem cells, induced pluripotent stem cells, neural stem cells, retinal progenitor cells, and olfactory ensheathing cells are discussed separately in addition to a brief description of two approaches for treatment of early-stage RP, including gene therapy and nutritional therapy.

Flaviviruses are zoonotic pathogens transmitted by the bite of infected mosquitos and ticks and represent a constant burden to human health. Here we review recent literature aimed at uncovering how flaviviruses interact with the cells that they infect. A better understanding of these interactions may ultimately lead to novel therapeutic targets. We highlight several studies that employed low-biased methods to discover new protein-protein, protein-RNA, and genetic interactions, and spotlight recent work characterizing the host protein, TMEM41B, which has been shown to be critical for infection by diverse flaviviruses and coronaviruses.

The molecular mechanisms underlying the clinical manifestations of coronavirus disease 2019 (COVID-19), and what distinguishes them from common seasonal influenza virus and other lung injury states such as acute respiratory distress syndrome, remain poorly understood. To address these challenges, we combine transcriptional profiling of 646 clinical nasopharyngeal swabs and 39 patient autopsy tissues to define body wide transcriptome changes in response to COVID-19. We then match these data with spatial protein and expression profiling across 357 tissue sections from 16 representative patient lung samples and identify tissue-compartment-specific damage wrought by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, evident as a function of varying viral loads during the clinical course of infection and tissue-type-specific expression states. Overall, our findings reveal a systemic disruption of canonical cellular and transcriptional pathways across all tissues, which can inform subsequent studies to combat the mortality of COVID-19 and to better understand the molecular dynamics of lethal SARS-CoV-2 and other respiratory infections.