Exploration of the P1 residue in 3CL protease inhibitors leading to the discovery of a 2-tetrahydrofuran P1 replacement.

The virally encoded 3C-like protease (3CLpro) is a well-validated drug target for the inhibition of coronaviruses including Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2). Most inhibitors of 3CLpro are peptidomimetic, with a γ-lactam in place of Gln at the P1 position of the pseudopeptide chain. An effort was pursued to identify a viable alternative to the γ-lactam P1 mimetic which would improve physicochemical properties while retaining affinity for the target. Discovery of a 2-tetrahydrofuran as a suitable P1 replacement that is a potent enzymatic inhibitor of 3CLpro in SARS-CoV-2 virus is described herein.

Parasite manipulation of host phenotypes inferred from transcriptional analyses in a trematode-amphipod system.

Manipulation of host phenotypes by parasites is hypothesized to be an adaptive strategy enhancing parasite transmission across hosts and generations. Characterizing the molecular mechanisms of manipulation is important to advance our understanding of host-parasite coevolution. The trematode (Levinseniella byrdi) is known to alter the colour and behaviour of its amphipod host (Orchestia grillus) presumably increasing predation of amphipods which enhances trematode transmission through its life cycle. We sampled 24 infected and 24 uninfected amphipods from a salt marsh in Massachusetts to perform differential gene expression analysis. In addition, we constructed novel genomic tools for O. grillus including a de novo genome and transcriptome. We discovered that trematode infection results in upregulation of amphipod transcripts associated with pigmentation and detection of external stimuli, and downregulation of multiple amphipod transcripts implicated in invertebrate immune responses, such as vacuolar ATPase genes. We hypothesize that suppression of immune genes and the altered expression of genes associated with coloration and behaviour may allow the trematode to persist in the amphipod and engage in further biochemical manipulation that promotes transmission. The genomic tools and transcriptomic analyses reported provide new opportunities to discover how parasites alter diverse pathways underlying host phenotypic changes in natural populations.

Novel Bent Conformation of CD4 Induced by HIV-1 Inhibitor Indirectly Prevents Productive Viral Attachment.

GSK3732394 is a multi-specific biologic inhibitor of HIV entry currently under clinical evaluation. A key component of this molecule is an adnectin (6940_B01) that binds to CD4 and inhibits downstream actions of gp160. Studies were performed to determine the binding site of the adnectin on CD4 and to understand the mechanism of inhibition. Using hydrogen-deuterium exchange with mass spectrometry (HDX), CD4 peptides showed differential rates of deuteration (either enhanced or slowed) in the presence of the adnectin that mapped predominantly to the interface of domains 2 and 3 (D2-D3). In addition, an X-ray crystal structure of an ibalizumab Fab/CD4(D1-D4)/adnectin complex revealed an extensive interface between the adnectin and residues on CD4 domains D2-D4 that stabilize a novel T-shaped CD4 conformation. A cryo-EM map of the gp140/CD4/GSK3732394 complex clearly shows the bent conformation for CD4 while bound to gp140. Mutagenic analyses on CD4 confirmed that amino acid F202 forms a key interaction with the adnectin. In addition, amino acid L151 was shown to be a critical indirect determinant of the specificity for binding to the human CD4 protein over related primate CD4 molecules, as it appears to modulate CD4's flexibility to adopt the adnectin-bound conformation. The significant conformational change of CD4 upon adnectin binding brings the D1 domain of CD4 in proximity to the host cell membrane surface, thereby re-orienting the gp120 binding site in a direction that is inaccessible to incoming virus due to a steric clash between gp160 trimers on the virus surface and the target cell membrane.

Overcoming the Pregnane X Receptor Liability: Rational Design to Eliminate PXR-Mediated CYP Induction.

The pregnane X receptor (PXR) regulates expression of proteins responsible for all three phases required for the detoxification mechanism, which include CYP450 enzymes, phase II enzymes, and multidrug efflux pumps. Therefore, PXR is a prominent receptor that is responsible for xenobiotic excretion and drug-drug interactions. Pyrimidinone 1 is an antagonist of the calcium sensing receptor (CaSR) and a strong activator of PXR. Repeat oral administration revealed diminished exposures over time, which prohibited further progression. A medicinal chemistry campaign was initiated to understand and abolish activation of PXR in order to increase systemic exposures. Rational structure-activity relationship investigations utilizing cocrystal structures and a de novo pharmacophore model resulted in compounds devoid of PXR activation. These studies culminated in the first orally active CaSR antagonist 8 suitable for progression. Cocrystallography, the pharmacophore model employed, and additional observations reported herein supported rational elimination of PXR activation and have applicability across diverse chemical classes to help erase PXR-driven drug-drug interactions.

Design of N-Benzoxaborole Benzofuran GSK8175-Optimization of Human Pharmacokinetics Inspired by Metabolites of a Failed Clinical HCV Inhibitor.

We previously described the discovery of GSK5852 (1), a non-nucleoside polymerase (NS5B) inhibitor of hepatitis C virus (HCV), in which an N-benzyl boronic acid was essential for potent antiviral activity. Unfortunately, facile benzylic oxidation resulted in a short plasma half-life (5 h) in human volunteers, and a backup program was initiated to remove metabolic liabilities associated with 1. Herein, we describe second-generation NS5B inhibitors including GSK8175 (49), a sulfonamide- N-benzoxaborole analog with low in vivo clearance across preclinical species and broad-spectrum activity against HCV replicons. An X-ray structure of NS5B protein cocrystallized with 49 revealed unique protein-inhibitor interactions mediated by an extensive network of ordered water molecules and the first evidence of boronate complex formation within the binding pocket. In clinical studies, 49 displayed a 60-63 h half-life and a robust decrease in viral RNA levels in HCV-infected patients, thereby validating our hypothesis that reducing benzylic oxidation would improve human pharmacokinetics and lower efficacious doses relative to 1.

Classification of crystallization outcomes using deep convolutional neural networks.

The Machine Recognition of Crystallization Outcomes (MARCO) initiative has assembled roughly half a million annotated images of macromolecular crystallization experiments from various sources and setups. Here, state-of-the-art machine learning algorithms are trained and tested on different parts of this data set. We find that more than 94% of the test images can be correctly labeled, irrespective of their experimental origin. Because crystal recognition is key to high-density screening and the systematic analysis of crystallization experiments, this approach opens the door to both industrial and fundamental research applications.

An orally available, brain-penetrant CAMKK2 inhibitor reduces food intake in rodent model.

Hypothalamic CAMKK2 represents a potential mechanism for chemically affecting satiety and promoting weight loss in clinically obese patients. Single-digit nanomolar inhibitors of CAMKK2 were identified in three related ATP-competitive series. Limited optimization of kinase selectivity, solubility, and pharmacokinetic properties were undertaken on all three series, as SAR was often transferrable. Ultimately, a 2,4-diaryl 7-azaindole was optimized to afford a tool molecule that potently inhibits AMPK phosphorylation in a hypothalamus-derived cell line, is orally bioavailable, and crosses the blood-brain barrier. When dosed orally in rodents, compound 4 t limited ghrelin-induced food intake.

Plant Translation Initiation Complex eIFiso4F Directs Pokeweed Antiviral Protein to Selectively Depurinate Uncapped Tobacco Etch Virus RNA.

Pokeweed antiviral protein (PAP) is a ribosome inactivating protein (RIP) that depurinates the sarcin/ricin loop (SRL) of rRNA, inhibiting protein synthesis. PAP depurinates viral RNA, and in doing so, lowers the infectivity of many plant viruses. The mechanism by which PAP accesses uncapped viral RNA is not known, impeding scientists from developing effective antiviral agents for the prevention of the diseases caused by uncapped RNA viruses. Kinetic rates of PAP interacting with tobacco etch virus (TEV) RNA, in the presence and absence of eIFiso4F, were examined, addressing how the eIF affects selective PAP targeting and depurination of the uncapped viral RNA. PAP-eIFs copurification assay and fluorescence resonance energy transfer demonstrate that PAP forms a ternary complex with the eIFiso4G and eIFiso4E, directing the depurination of uncapped viral RNA. eIFiso4F selectively targets PAP to depurinate TEV RNA by increasing PAP's specificity constant for uncapped viral RNA 12-fold, when compared to the depurination of an oligonucleotide RNA that mimics the SRL of large rRNA, and cellular capped luciferase mRNA. This explains how PAP is able to lower infectivity of pokeweed viruses, while preserving its own ribosomes and cellular RNA from depurination: PAP utilizes cellular eIFiso4F in a novel strategy to target uncapped viral RNA. It may be possible to modulate and utilize these PAP-eIFs interactions for their public health benefit; by repurposing them to selectively target PAP to depurinate uncapped viral RNA, many plant and animal diseases caused by these viruses could be alleviated.

Chiropractors' perception of occupational stress and its influencing factors: a qualitative study using responses to open-ended questions.

BACKGROUND: Job stress and emotional exhaustion have been shown to have a negative impact on the helping professional. The development and causal relations of job stress and emotional exhaustion are rather unclear in the chiropractic profession. The objective of this study is to understand the main sources of occupational stress and emotional exhaustion among doctors of chiropractic. METHODS: Analysis of the written responses to web-based open-ended questionnaire was performed using an interpretive research methodology. Additionally, cross tabulation and Chi square statistical tests were conducted to match and couple the demographic data with the categorical themes. RESULTS: Fourteen professional stress categories emerged from the 970 completed surveys. "Managed Care Organization regulation", "Managed Care reimbursement" and "Scope of Practice Issues" were the most common stressors that negatively influenced chiropractors' professional and personal lives. The results of the categorical analysis suggests that age, marital status, number of years in practice and location of practice may have an influence on the category of stress reported by chiropractors. CONCLUSIONS: The qualitative approach revealed common, conventional and culture-specific job stressors in doctors of chiropractic. Notably, these findings suggest an association between third-party payer influences (increased regulation/decreased reimbursement) with that of increased job stress. Further research will be undertaken to refine the stress and satisfaction parameters and address stress interventions.

A human fatty acid synthase inhibitor binds ß-ketoacyl reductase in the keto-substrate site.

Human fatty acid synthase (hFAS) is a complex, multifunctional enzyme that is solely responsible for the de novo synthesis of long chain fatty acids. hFAS is highly expressed in a number of cancers, with low expression observed in most normal tissues. Although normal tissues tend to obtain fatty acids from the diet, tumor tissues rely on de novo fatty acid synthesis, making hFAS an attractive metabolic target for the treatment of cancer. We describe here the identification of GSK2194069, a potent and specific inhibitor of the ß-ketoacyl reductase (KR) activity of hFAS; the characterization of its enzymatic and cellular mechanism of action; and its inhibition of human tumor cell growth. We also present the design of a new protein construct suitable for crystallography, which resulted in what is to our knowledge the first co-crystal structure of the human KR domain and includes a bound inhibitor.

Prevalence and associated risk factors of burnout among US doctors of chiropractic.

OBJECTIVE: The purpose of this study was to establish the frequency of burnout among doctors of chiropractic in the United States. METHODS: Using a nonprobability convenience sampling methodology, we e-mailed the Maslach Burnout Inventory-Human Services Survey and a sociodemographic questionnaire to a randomized sample of licensed doctors of chiropractic (n = 8000). RESULTS: The survey return rate was 16.06%. Twenty-one percent of the participants had high emotional exhaustion (EE), 8% had low personal accomplishment, and 8% had high depersonalization. DISCUSSION: Significant differences (P < .001) were found in the level of EE, depersonalization, and personal accomplishment as a function of sex, time dedicated to clinical care and administrative duties, source of reimbursement, the type of practice setting, the nature of practitioners' therapeutic focus, the location of chiropractic college, self-perception of burnout, the effect of suffering from a work-related injury, the varying chiropractic philosophical perspectives, and the public's opinion of chiropractic. CONCLUSION: Although doctors of chiropractic in the United States who responded to the survey had a relatively low frequency of burnout, higher levels of EE remain workplace issues for this professional group.

Discovery of a potent boronic acid derived inhibitor of the HCV RNA-dependent RNA polymerase.

A boronic acid moiety was found to be a critical pharmacophore for enhanced in vitro potency against wild-type hepatitis C replicons and known clinical polymorphic and resistant HCV mutant replicons. The synthesis, optimization, and structure-activity relationships associated with inhibition of HCV replication in a subgenomic replication system for a series of non-nucleoside boron-containing HCV RNA-dependent RNA polymerase (NS5B) inhibitors are described. A summary of the discovery of 3 (GSK5852), a molecule which entered clinical trials in subjects infected with HCV in 2011, is included.

A 69-year-old presenting with musculoskeletal low back pain: a case of lumbosacral chordoma.

OBJECTIVE: The purpose of this case report is to describe the presentation of a patient with lumbosacral chordoma characterized by somatic chronic low back pain and intermittent sacral nerve impingement. CASE REPORT: A 69-year-old male presenting to an emergency department (ED) with low back pain was provided analgesics and muscle relaxants then referred for a series of chiropractic treatments. Chiropractic treatment included manipulation, physical therapy, and rehabilitation. After 3 times per week for a total of 4 weeks, re-examination showed little relief of his symptoms. His pain symptoms worsened and he presented to the ED for the second time. Magnetic resonance imaging was performed and revealed a high intensity mass. INTERVENTION AND OUTCOME: The soft tissue mass identified on magnetic resonance imaging was surgically removed. Shortly after the surgery, the patient developed post-operative bleeding and was returned to surgery. During the second procedure, he developed a post-operative hemorrhage related to the development of disseminated intravascular coagulation and subsequently died during the second procedure. A malignant lumbosacral chordoma was diagnosed on pathologic examination. CONCLUSION: This case report describes the presentation of a patient with lumbosacral chordoma presenting with musculoskeletal low back pain. Chordomas are rare with few prominent manifestations. An early diagnosis can potentially make a difference in morbidity and mortality. Due to its insidious nature, it is a difficult diagnosis and one that is often delayed.

Prevalence of burnout among doctors of chiropractic in the northeastern United States.

OBJECTIVE: The purpose of this study was to measure the prevalence of burnout among doctors of chiropractic (DCs) in the New York, New Jersey, and Pennsylvania geographical region and compare these results with burnout data from other health care professions. METHODS: This exploratory study applied cross-sectional data collection methods. Using nonprobability convenience sampling, a New York-New Jersey-Pennsylvania chiropractic governance body provided contact information of a randomized sample of licensed DCs from their membership directory. Participants included any DC licensed to practice chiropractic whose primary occupation encompassed the chiropractic profession. The Maslach Burnout Inventory-Human Services Survey (MBI-HSS) and a demographic questionnaire were e-mailed to a randomized sample of licensed DCs. RESULTS: Of the 772 surveys deployed, 90 returned the survey with usable data. Nearly 40% of the DCs reported a moderate (24%) or high (18%) level of emotional exhaustion, whereas the majority of respondents scored a high (72%) level of personal accomplishment. In total, only 2 participants (2%) met the criteria for high burnout, whereas 42 participants (47%) were low. Statistically significant relationships (P < .001) were found between burnout subscales and the effect of time dedicated to administrative duties, the type of practice setting, the varying chiropractic philosophical perspectives, the public's opinion of chiropractic, and the effect of suffering from a work-related injury. When compared with data from previously published studies using the MBI-HSS for other health professions (ie, medical, nursing, physical therapy, occupational therapy, and dentistry), the values for DCs were significantly lower. CONCLUSIONS: The sample of DCs in this study fared more favorably on all 3 dimensions of burnout. They reported lower emotional exhaustion and depersonalization scores and higher personal accomplishment scores than their medical, nursing, physical therapy, occupational therapy, and dentistry colleagues who have been evaluated using the MBI-HSS. However, the levels of emotional exhaustion remain a concern for this professional group.

Burnout among chiropractic practitioners: real or imagined an exploratory study protocol.

Burnout is a psychological syndrome of emotional exhaustion, depersonalization and reduced personal accomplishment that has been found to exist in a significant number of healthcare and helping professionals. It imposes a significant societal burden by shortened practitioner lifespan, decreased efficiency, negative health outcomes and poorer levels of patient care. Theoretical models suggest that it appears to be the result of a complex interaction between job resources and job demands. It may be reasonable to conclude that Chiropractic professionals experience similar vocational demands and thus experience significant levels of occupational stress and subsequent burnout. However the data on burnout within the chiropractic profession is limited. It is possible that this results in significant negative outcomes on chiropractors and their patients. Therefore, the objective of this paper is to demonstrate the need to explore burnout in chiropractic practice and offer a research protocol for a potential study.

Conformationally constrained farnesoid X receptor (FXR) agonists: alternative replacements of the stilbene.

To further explore the optimum placement of the acid moiety in conformationally constrained analogs of GW 4064 1a, a series of stilbene replacements were prepared. The benzothiophene 1f and the indole 1g display the optimal orientation of the carboxylate for enhanced FXR agonist potency.

Small molecule agonists of the orphan nuclear receptors steroidogenic factor-1 (SF-1, NR5A1) and liver receptor homologue-1 (LRH-1, NR5A2).

The crystal structure of LRH-1 ligand binding domain bound to our previously reported agonist 3-(E-oct-4-en-4-yl)-1-phenylamino-2-phenyl-cis-bicyclo[3.3.0]oct-2-ene 5 is described. Two new classes of agonists in which the bridgehead anilino group from our first series was replaced with an alkoxy or 1-ethenyl group were designed, synthesized, and tested for activity in a peptide recruitment assay. Both new classes gave very active compounds, particularly against SF-1. Structure-activity studies led to excellent dual-LRH-1/SF-1 agonists (e.g., RJW100) as well as compounds selective for LRH-1 (RJW101) and SF-1 (RJW102 and RJW103). The series based on 1-ethenyl substitution was acid stable, overcoming a significant drawback of our original bridgehead anilino-substituted series. Initial studies on the regulation of gene expression in human cell lines showed excellent, reproducible activity at endogenous target genes.

Conformationally constrained farnesoid X receptor (FXR) agonists: heteroaryl replacements of the naphthalene.

To improve on the drug properties of GSK8062 1b, a series of heteroaryl bicyclic naphthalene replacements were prepared. The quinoline 1c was an equipotent FXR agonist with improved drug developability parameters relative to 1b. In addition, analog 1c lowered body weight gain and serum glucose in a DIO mouse model of diabetes.

Potential unique causes of burnout for chiropractic professionals.

OBJECTIVE: The objective of this narrative review is to discuss the potential for burnout in chiropractic practitioners. This discussion is grounded in the job demands-resource model, the conservation of resources model, the unique profession-specific stressors experienced by chiropractors, and information from similar health care professions. METHODS: A search using both the indexed (PubMed and PsychLit) and nonindexed psychosocial literature was used. Other resources included the Cochrane Library, articles from governing bodies of the chiropractic profession, trade magazines, and research conferences and symposium proceedings. Articles were analyzed following the grounded theory principles: open coding and memos for conceptual labeling, axial coding and memos for category building, and selective coding for model building. RESULTS: Potential stressors unique to doctors of chiropractic include factors associated with physical workload, role stress, and mental and emotional demands. CONCLUSIONS: There are unique chiropractic-specific occupational characteristics that possibly contribute to burnout in the chiropractic professionals. These findings emphasize the need for assessing and measuring burnout and attrition within the chiropractic profession.

Polymer-fullerene bulk-heterojunction solar cells.

Solution-processed bulk heterojunction organic photovoltaic (OPV) devices have gained serious attention during the last few years and are established as one of the leading next generation photovoltaic technologies for low cost power production. This article reviews the OPV development highlights of the last two decades, and summarizes the key milestones that have brought the technology to today's efficiency performance of over 7%. An outlook is presented on what will be required to drive this young photovoltaic technology towards the next major milestone, a 10% power conversion efficiency, considered by many to represent the efficiency at which OPV can be adopted in wide-spread applications. With first products already entering the market, sufficient lifetime for the intended application becomes more and more critical, and the status of OPV stability as well as the current understanding of degradation mechanisms will be reviewed in the second part of this article.