The Synthesis and Reactivity of Naphthoquinonynes.

The first systematic exploration of the synthesis and reactivity of naphthoquinonynes is described. Routes to two regioisomeric Kobayashi-type naphthoquinonyne precursors have been developed, and the reactivity of the ensuing 6,7- and 5,6-aryne intermediates has been investigated. Remarkably, these studies have revealed that a broad range of cycloadditions, nucleophile additions and difunctionalizations can be achieved while maintaining the integrity of the highly sensitive quinone unit. The methodologies offer a powerful diversity oriented approach to C6 and C7 functionalized naphthoquinones, which are typically challenging to access. From a reactivity viewpoint, the study is significant because it demonstrates that aryne-based functionalizations can be utilized strategically in the presence of highly reactive and directly competing functionality.

Integrative Molecular Structure Elucidation and Construction of an Extended Metabolic Pathway Associated with an Ancient Innate Immune Response in COVID-19 Patients.

We present compelling evidence for the existence of an extended innate viperin-dependent pathway, which provides crucial evidence for an adaptive response to viral agents, such as SARS-CoV-2. We show the in vivo biosynthesis of a family of novel endogenous cytosine metabolites with potential antiviral activities. Two-dimensional nuclear magnetic resonance (NMR) spectroscopy revealed a characteristic spin-system motif, indicating the presence of an extended panel of urinary metabolites during the acute viral replication phase. Mass spectrometry additionally enabled the characterization and quantification of the most abundant serum metabolites, showing the potential diagnostic value of the compounds for viral infections. In total, we unveiled ten nucleoside (cytosine- and uracil-based) analogue structures, eight of which were previously unknown in humans allowing us to propose a new extended viperin pathway for the innate production of antiviral compounds. The molecular structures of the nucleoside analogues and their correlation with an array of serum cytokines, including IFN-α2, IFN-γ, and IL-10, suggest an association with the viperin enzyme contributing to an ancient endogenous innate immune defense mechanism against viral infection.

Chemoenzymatic Synthesis of 3'-Deoxy-3',4'-didehydro-cytidine triphosphate (ddhCTP).

3'-Deoxy-3',4'-didehydro-cytidine triphosphate (ddhCTP) is a novel antiviral molecule produced by the enzyme viperin during the early stages of the innate immune response. ddhCTP has been shown to act as a chain terminator of flavivirus RNA-dependent RNA polymerases. To date, synthesis of ddhCTP requires complicated synthetic protocols or isolation of the enzyme viperin to catalyze the production of ddhCTP from CTP. Recombinant viperin approaches preclude the production of highly pure ddhCTP (free of contaminants such as CTP), whereas the chemical synthesis involves techniques or equipment not readily available to most laboratories. Herein, we describe the chemoenzymatic synthesis of ddhCTP, starting from commercially available ddhC. We utilize these methods to produce milligram quantities of ddhCTP, ddhCDP, and ddhCMP. Using purified semisynthetic ddhCTP and fully synthetic ddhCTP, we also show ddhCTP does not inhibit NAD+-dependent enzymes such as glyceraldehyde 3-phosphate dehydrogenase, malate dehydrogenase, or lactate dehydrogenase, contrary to a recent report.

Galidesivir Triphosphate Promotes Stalling of Dengue-2 Virus Polymerase Immediately Prior to Incorporation.

Millions of people are infected by the dengue and Zika viruses each year, resulting in significant morbidity and mortality. Galidesivir is an adenosine nucleoside analog that can attenuate flavivirus replication in cell-based assays and animal models of infection. Galidesivir is converted to the triphosphorylated form by host kinases and subsequently incorporated into viral RNA by viral RNA polymerases. This has been proposed to lead to the delayed termination of RNA synthesis. Here, we report direct in vitro testing of the effects of Galidesivir triphosphate on dengue-2 and Zika virus polymerase activity. Galidesivir triphosphate was chemically synthesized, and inhibition of RNA synthesis followed using a dinucleotide-primed assay with a homopolymeric poly(U) template. Galidesivir triphosphate was equipotent against dengue-2 and Zika polymerases, with IC50 values of 42 ± 12 µM and 47 ± 5 µM, respectively, at an ATP concentration of 20 µM. RNA primer extension assays show that the dengue-2 polymerase stalls while attempting to add a Galidesivir nucleotide to the nascent RNA chain, evidenced by the accumulation of RNA products truncated immediately upstream of Galidesivir incorporation sites. Nevertheless, Galidesivir is incorporated at isolated sites with low efficiency, leading to the subsequent synthesis of full-length RNA with no evidence of delayed chain termination. The incorporation of Galidesivir at consecutive sites is strongly disfavored, highlighting the potential for modulation of inhibitory effects of nucleoside analogs by the template sequence. Our results suggest that attenuation of dengue replication by Galidesivir may not derive from the early termination of RNA synthesis following Galidesivir incorporation.

An Isomer of Galidesivir That Potently Inhibits Influenza Viruses and Members of the Bunyavirales Order.

We report for the first time the antiviral activities of two iminovirs (antiviral imino-C-nucleosides) 1 and 2, structurally related to galidesivir (Immucillin A, BCX4430). An iminovir containing the 4-aminopyrrolo[2,1-f][1,2,4-triazine] nucleobase found in remdesivir exhibited submicromolar inhibition of multiple strains of influenza A and B viruses, as well as members of the Bunyavirales order. We also report the first syntheses of ProTide prodrugs of iminovir monophosphates, which unexpectedly displayed poorer viral inhibition than their parent nucleosides in vitro. An efficient synthesis of the 4-aminopyrrolo[2,1-f][1,2,4-triazine]-containing iminovir 2 was developed to enable preliminary in vivo studies, wherein it displayed significant toxicity in BALB/c mice and limited protection against influenza. Further modification of this anti-influenza iminovir will therefore be required to improve its therapeutic value.

Geochemical evidence for the internal migration of gas condensate in a major unconventional tight petroleum system.

Unconventional petroleum systems go through multiple episodes of internal hydrocarbon migration in response to evolving temperature and pressure conditions during burial and uplift. Migrated fluid signatures can be recognized using stable carbon isotope and PVT compositional data from produced samples representative of in-situ petroleum fluids. Such samples, however, are seldom collected due to operational complexity and high cost. Here, we use carbon isotope and PVT data from co-produced hydrocarbon gas and liquid to provide evidence for widespread migration of gas-condensate in the Montney unconventional petroleum system of western Canada. Extended C1-C33 isotopic profiles exhibit convex upward signatures with C4-C5 maxima at low molecular weight, and increasing or nearly uniform signatures at high molecular weight. Additionally, recombination PVT compositional data show C6-C15 condensate concentrations are higher than expected for unmodified oils. The combined convex upward and increasing or uniform isotopic signatures are interpreted as mixing profiles formed by the introduction of high-maturity gas-condensate (C1-C15) to shallower zones with in-situ hydrocarbon fluids of lower thermal maturity. The recognition of widespread gas-condensate migration adds to the complex history of internal hydrocarbon migration within the Montney tight-petroleum system including previously identified migration episodes of early oil and late-stage methane-rich gas.

Viperin triggers ribosome collision-dependent translation inhibition to restrict viral replication.

Innate immune responses induce hundreds of interferon-stimulated genes (ISGs). Viperin, a member of the radical S-adenosyl methionine (SAM) superfamily of enzymes, is the product of one such ISG that restricts the replication of a broad spectrum of viruses. Here, we report a previously unknown antiviral mechanism in which viperin activates a ribosome collision-dependent pathway that inhibits both cellular and viral RNA translation. We found that the radical SAM activity of viperin is required for translation inhibition and that this is mediated by viperin's enzymatic product, 3'-deoxy-3',4'-didehydro-CTP (ddhCTP). Viperin triggers ribosome collisions and activates the MAPKKK ZAK pathway that in turn activates the GCN2 arm of the integrated stress response pathway to inhibit translation. The study illustrates the importance of translational repression in the antiviral response and identifies viperin as a translation regulator in innate immunity.

Inhibition of SARS-CoV-2 polymerase by nucleotide analogs from a single-molecule perspective.

The absence of 'shovel-ready' anti-coronavirus drugs during vaccine development has exceedingly worsened the SARS-CoV-2 pandemic. Furthermore, new vaccine-resistant variants and coronavirus outbreaks may occur in the near future, and we must be ready to face this possibility. However, efficient antiviral drugs are still lacking to this day, due to our poor understanding of the mode of incorporation and mechanism of action of nucleotides analogs that target the coronavirus polymerase to impair its essential activity. Here, we characterize the impact of remdesivir (RDV, the only FDA-approved anti-coronavirus drug) and other nucleotide analogs (NAs) on RNA synthesis by the coronavirus polymerase using a high-throughput, single-molecule, magnetic-tweezers platform. We reveal that the location of the modification in the ribose or in the base dictates the catalytic pathway(s) used for its incorporation. We show that RDV incorporation does not terminate viral RNA synthesis, but leads the polymerase into backtrack as far as 30 nt, which may appear as termination in traditional ensemble assays. SARS-CoV-2 is able to evade the endogenously synthesized product of the viperin antiviral protein, ddhCTP, though the polymerase incorporates this NA well. This experimental paradigm is essential to the discovery and development of therapeutics targeting viral polymerases.

To multiply and spread from cell to cell, the virus responsible for COVID-19 (also known as SARS-CoV-2) must first replicate its genetic information. This process involves a 'polymerase' protein complex making a faithful copy by assembling a precise sequence of building blocks, or nucleotides. The only drug approved against SARS-CoV-2 by the US Food and Drug Administration (FDA), remdesivir, consists of a nucleotide analog, a molecule whose structure is similar to the actual building blocks needed for replication. If the polymerase recognizes and integrates these analogs into the growing genetic sequence, the replication mechanism is disrupted, and the virus cannot multiply. Most approaches to study this process seem to indicate that remdesivir works by stopping the polymerase and terminating replication altogether. Yet, exactly how remdesivir and other analogs impair the synthesis of new copies of the virus remains uncertain. To explore this question, Seifert, Bera et al. employed an approach called magnetic tweezers which uses a magnetic field to manipulate micro-particles with great precision. Unlike other methods, this technique allows analogs to be integrated under conditions similar to those found in cells, and to be examined at the level of a single molecule. The results show that contrary to previous assumptions, remdesivir does not terminate replication; instead, it causes the polymerase to pause and backtrack (which may appear as termination in other techniques). The same approach was then applied to other nucleotide analogs, some of which were also found to target the SARS-CoV-2 polymerase. However, these analogs are incorporated differently to remdesivir and with less efficiency. They also obstruct the polymerase in distinct ways. Taken together, the results by Seifert, Bera et al. suggest that magnetic tweezers can be a powerful approach to reveal how analogs interfere with replication. This information could be used to improve currently available analogs as well as develop new antiviral drugs that are more effective against SARS-CoV-2. This knowledge will be key at a time when treatments against COVID-19 are still lacking, and may be needed to protect against new variants and future outbreaks.

Chemical Synthesis of the Antiviral Nucleotide Analogue ddhCTP.

3'-Deoxy-3',4'-didehydro-cytidine triphosphate (ddhCTP) is a novel antiviral molecule produced by the enzyme viperin as part of the innate immune response. ddhCTP has been shown to act as an obligate chain terminator of flavivirus and SARS-CoV-2 RNA-dependent RNA polymerases; however, further biophysical studies have been precluded by limited access to this promising antiviral. Herein, we report a robust and scalable synthesis of ddhCTP as well as the mono- and diphosphates ddhCMP and ddhCDP, respectively. Identification of a 2'-silyl ether protection strategy enabled selective synthesis and facile purification of the 5'-triphosphate, culminating in the preparation of ddhCTP on a gram scale.

Inhibition of SARS-CoV-2 polymerase by nucleotide analogs: a single molecule perspective.

The nucleotide analog Remdesivir (RDV) is the only FDA-approved antiviral therapy to treat infection by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The physical basis for efficient utilization of RDV by SARS-CoV-2 polymerase is unknown. Here, we characterize the impact of RDV and other nucleotide analogs on RNA synthesis by the polymerase using a high-throughput, single-molecule, magnetic-tweezers platform. The location of the modification in the ribose or in the base dictates the catalytic pathway(s) used for its incorporation. We reveal that RDV incorporation does not terminate viral RNA synthesis, but leads the polymerase into deep backtrack, which may appear as termination in traditional ensemble assays. SARS-CoV-2 is able to evade the endogenously synthesized product of the viperin antiviral protein, ddhCTP, though the polymerase incorporates this nucleotide analog well. This experimental paradigm is essential to the discovery and development of therapeutics targeting viral polymerases. TEASER: We revise Remdesivir's mechanism of action and reveal SARS-CoV-2 ability to evade interferon-induced antiviral ddhCTP.

The Different Facets of Metal-Catalyzed C-H Functionalization Involving Quinone Compounds.

Metal-catalysed C-H functionalization has emerged as a powerful platform for the derivatization of quinones, a class of compounds with wide-ranging applications. This review organises and discusses the evolution of this chemistry from early Fujiwara-Moritani reactions, through to modern directing-group assisted C-H functionalization processes, including C-H functionalization reactions directed by the quinone ring itself. Mechanistic details of these reactions are provided to afford insight into how the unique reactivity of quinoidal compounds has been leveraged in each example.

Uncoupling Molecular Testing for SARS-CoV-2 From International Supply Chains.

The rapid global rise of COVID-19 from late 2019 caught major manufacturers of RT-qPCR reagents by surprise and threw into sharp focus the heavy reliance of molecular diagnostic providers on a handful of reagent suppliers. In addition, lockdown and transport bans, necessarily imposed to contain disease spread, put pressure on global supply lines with freight volumes severely restricted. These issues were acutely felt in New Zealand, an island nation located at the end of most supply lines. This led New Zealand scientists to pose the hypothetical question: in a doomsday scenario where access to COVID-19 RT-qPCR reagents became unavailable, would New Zealand possess the expertise and infrastructure to make its own reagents onshore? In this work we describe a review of New Zealand's COVID-19 test requirements, bring together local experts and resources to make all reagents for the RT-qPCR process, and create a COVID-19 diagnostic assay referred to as HomeBrew (HB) RT-qPCR from onshore synthesized components. This one-step RT-qPCR assay was evaluated using clinical samples and shown to be comparable to a commercial COVID-19 assay. Through this work we show New Zealand has both the expertise and, with sufficient lead time and forward planning, infrastructure capacity to meet reagent supply challenges if they were ever to emerge.

Strategies towards potent trypanocidal drugs: Application of Rh-catalyzed [2 + 2 + 2] cycloadditions, sulfonyl phthalide annulation and nitroalkene reactions for the synthesis of substituted quinones and their evaluation against Trypanosoma cruzi.

Rhodium-catalyzed [2 + 2 + 2] cycloadditions, sulfonyl phthalide annulations and nitroalkene reactions have been employed for the synthesis of 56 quinone-based compounds. These were evaluated against Trypanosoma cruzi, the parasite that causes Chagas disease. The reactions described here are part of a program that aims to utilize modern, versatile and efficient synthetic methods for the one or two step preparation of trypanocidal compounds. We have identified 9 compounds with potent activity against the parasite; 3 of these were 30-fold more potent than benznidazole (Bz), a drug used for the treatment of Chagas disease. This article provides a comprehensive outline of reactions involving over 120 compounds aimed at the discovery of new quinone-based frameworks with activity against T. cruzi.

Core versus cuttings samples for geochemical and petrophysical analysis of unconventional reservoir rocks.

Core samples from petroleum wells are costly to obtain, hence drill cuttings are commonly used as an alternative source of rock measurements for reservoir, basin modelling, and sedimentology studies. However, serious issues such as contamination from drilling mud, geological representativeness, and physical alteration can cast uncertainty on the results of studies based on cuttings samples. This paper provides a unique comparative study of core and cuttings samples obtained from both vertical and horizontal sections of a petroleum well drilled in the Canadian Montney tight gas siltstone reservoir to investigate the suitability of cuttings for a wide range of geochemical and petrophysical analyses. The results show that, on average, the bulk quantity of kerogen or solid bitumen measured in cuttings is comparable to that of the core samples. However, total organic carbon (TOC) measurements are influenced by oil-based drilling mud (OBM) contamination. Solvent-cleaning of cuttings has been shown to effectively remove OBM contamination in light, medium, and heavy range hydrocarbons and to produce similar kerogen/solid bitumen measurements to that of core samples. Similarly, pyrolysis methods provide an alternative to the solvent-cleaning procedure for analysis of kerogen/solid bitumen in as-received cuttings. Microscopic study substantiates the presence of significant contamination by OBM and caved organic and inorganic matter in the cuttings, which potentially influence the bulk geochemistry of the samples. Furthermore, minerals in the cuttings display induced micro-fractures due to physical impacts of the drilling process. These drilling-induced micro-fractures affect petrophysical properties by artificially enhancing the measured porosity and permeability.

Rh-Catalyzed [2 + 2 + 2] Cycloadditions with Benzoquinones: De Novo Access to Naphthoquinones for Lignan and Type II Polyketide Synthesis.

The first examples of Rh-catalyzed [2 + 2 + 2] cycloadditions between diynes and benzoquinones are described. The method enables de novo and step-economical access to challenging naphthoquinones that are of relevance to lignan and type II polyketide synthesis. The value of the chemistry is demonstrated by a short total synthesis of justicidone.

Methodological advances in statistical prediction.

Thirty years ago, Dawes, Faust, and Meehl (1989) argued that mental health professionals should routinely use statistical prediction rules to describe and diagnose clients, predict behaviors, and formulate treatment plans. Subsequent research has supported their claim that statistical prediction performs well when compared to clinical judgment. However, many of the things we thought we knew about statistical prediction have changed. The purpose of this literature review is to describe methodological advances in statistical prediction. Three broad areas are covered. First, while statistical prediction rules are valuable for criterion-referenced assessment (e.g., predicting violence, recidivism, treatment outcomes), they are valuable only for some norm-referenced assessment tasks (e.g., diagnosis but not describing personality and psychopathology). Second, statistical prediction is particularly prominent for the prediction of violence and criminal recidivism. Results from this area will be used to describe the validity of traditional clinical judgment, structured professional judgment, and statistical prediction. The results support the use of both structured professional judgment and statistical prediction. The effect of allowing professionals to override statistical predictions consistently led to lower validity. Third, issues in building statistical prediction rules are described, including the assignment of weights to predictors, the emergence of new statistical analyses (e.g., machine learning), and the role of theory. As research has progressed, statistical prediction has become one of the most exciting areas of psychological assessment. (PsycINFO Database Record (c) 2019 APA, all rights reserved).

2-Formylpyrrole natural products: origin, structural diversity, bioactivity and synthesis.

Covering: up to April 2018 2-Formylpyrroles are ubiquitous in nature, arising from the non-enzymatic Maillard reactions of amines and sugars. Often confused for secondary metabolites, these Maillard products display interesting biological activities including hepatoprotective, immunostimulatory, antiproliferative and antioxidant effects. This review presents all 2-formylpyrrole natural products reported to date and identifies structural sub-classes for their categorisation. The origin, biological activity and chemical syntheses of these natural products are discussed herein.

The effectiveness of army field manual interrogation approaches for educing information and building rapport.

In 2016, the U.S. Congress mandated that federal intelligence interrogators adhere to the methods of the U.S. Army Field Manual FM 2-22.3 (AFM) and that the manual be revised based upon empirically based evaluations of the interrogation methods' effectiveness with interviewees motivated to withhold information. In the present study, 120 participants took part in a testing situation in which half were induced to cheat. All participants were then accused of cheating and interrogated with either (a) a combination of AFM interrogation approaches that focused on the potential benefits of cooperation with the interviewer (cooperation-focused condition), or (b) a combination of AFM approaches that focused on the potential risks of withholding information (withholding-focused condition). Participants who cheated on the test were significantly more likely to admit their wrongdoing and to provide additional relevant information when interrogated with the withholding-focused approaches than when questioned with the cooperation-focused approaches. The "we know all" AFM approach was especially effective for eliciting truthful admission-related details. Participants reported high rapport with the interrogator in both the cooperation-focused and withholding-focused conditions. These findings indicate that the we-know-all approach can be effective for maintaining rapport and eliciting accurate information in brief interrogations. (PsycINFO Database Record (c) 2018 APA, all rights reserved).

The Lackland Behavioral Questionnaire: The use of biographical data and statistical prediction rules for public safety screening.

Screening for public safety positions (e.g., police officers, fire fighters, military service members) is a difficult and challenging task. Notably, the military has been widely criticized because of the general lack of an empirically based system or program for mental health screening. The purpose of the present study is to describe the use of statistical prediction rules for this task. Prediction rules were derived and validated using U.S. Air Force (USAF) recruits in basic military training (N = 50,322). Items from the Lackland Behavioral Questionnaire were used as predictors. General attrition (discharge for any reason before completing term of service) and disciplinary offenses (including criminal charges) were used as outcomes. For trainees in the top 2% or 1% of the general attrition prediction rule, 63% and 68% were discharged before they completed their first 4 years. The base rate was 25%. Similarly, for trainees in the top 2% or 1% of the disciplinary offenses prediction rule, 35% and 39% had a significant disciplinary offense over the following 4 years. The base rate was 15.5%. The results suggest that we may be able to use biographical data inventories and statistical prediction rules to identify a small percentage of trainees in public safety fields with significant mental health or behavioral histories who are at elevated risk for general attrition and disciplinary offenses. (PsycINFO Database Record

Prevalence of internalizing, externalizing, and psychotic disorders among low-risk juvenile offenders.

To effectively allocate mental health services, agencies must be able to predict what proportion of youth will have a mental disorder. Prevalence estimates are available for juvenile offenders at intake, detained youth, and incarcerated youth, but there is limited research on prevalence of mental disorders for juvenile offenders who are low-risk to reoffend, many of whom are first time offenders (i.e., low-risk youth). To complicate matters, ethnic minorities are disproportionately represented in the justice system, and specifically, little is known about culturally sensitive clinical interviewing. To aid service providers and administrators in allocating mental health resources for low-risk offenders and to contribute to knowledge on culturally sensitive clinical assessment techniques, the present study reports the prevalence of mental disorders for a mostly Mexican American sample of 503 low-risk youth in diversion programming. We found that approximately 1 of every 6 (17.1%) low-risk juvenile offenders had a current affective, anxiety, or psychotic disorder, and 24.9% of low-risk juvenile offenders met criteria for a current substance/alcohol abuse disorder. These results suggest that allocating a portion of specialty mental health services and substance abuse treatment for low-risk juvenile offenders may help agencies combat the issue of repeat offending by offering public health interventions proactively to indirectly prevent recidivism rather than reacting afterward. Lastly, recommendations are given to help service providers incorporate culturally sensitive techniques into clinical assessment in order to better identify Mexican American juvenile offenders with mental health needs. (PsycINFO Database Record