COVID-19 Vaccines-All You Want to Know.

The severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) pandemic has led to an unprecedented public health crisis. The collective global response has led to production of multiple safe and effective vaccines utilizing novel platforms to combat the virus that have propelled the field of vaccinology forward. Significant challenges to universal vaccine effectiveness remain, including immune evasion by SARS-CoV-2 variants, waning of immune response, inadequate knowledge of correlates of protection, and dosing in special populations. This review serves as a detailed evaluation of the development of the current SARS-CoV-2 vaccines, their effectiveness, and challenges to their deployment as a preventive tool.

Impulse oscillometry measurement of distal airways obstruction in depleted uranium-exposed Gulf War veterans.

INTRODUCTION: A cohort of Gulf War I veterans who sustained exposure to depleted uranium undergoes biennial surveillance for potential uranium-related health effects. We performed impulse oscillometry and hypothesized that veterans with higher uranium body burdens would have more obstructive abnormalities than those with lower burdens. METHODS: We compared pulmonary function of veterans in high versus low urine uranium groups by evaluating spirometry and oscillometry values. RESULTS: Overall mean spirometry and oscillometry resistance values fell within the normal ranges. There were no significant differences between the high and low uranium groups for any parameters. However, more veterans were classified as having obstruction by oscillometry (42%) than spirometry (8%). CONCLUSIONS: While oscillometry identified more veterans as obstructed, obstruction was not uranium-related. However, the added sensitivity of this method implies a benefit in wider surveillance of exposed cohorts and holds promise in identifying abnormalities in areas of the lung historically described as silent.

Characterization of the membrane-inserted C-terminus of cytoprotective BCL-XL.

BCL-XL is a dominant inhibitor of apoptosis and a significant anti-cancer drug target. Endogenous BCL-XL is integral to the mitochondrial outer membrane (MOM). BCL-XL reconstituted in detergent-free lipid bilayer nanodiscs is anchored to the nanodisc lipid bilayer membrane by tight association of its C-terminal tail, while the N-terminal head retains the canonical structure determined for water-soluble, tail-truncated BCL-XL, with the surface groove solvent-exposed and available for BH3 ligand binding. To better understand the conformation and dynamics of this key region of BCL-XL we have developed methods for isolating the membrane-embedded C-terminal tail from its N-terminal head and for preparing protein suitable for structural and biochemical studies. Here, we outline the methods for sample preparation and characterization and describe previously unreported structural and dynamics features. We show that the C-terminal tail of BCL-XL forms a transmembrane α-helix that retains a significant degree of conformational dynamics. We also show that the presence of the intact C-terminus destabilizes the soluble state of the protein, and that the small fraction of soluble recombinant protein produced in Escherichia coli is susceptible to proteolytic degradation of C-terminal residues beyond M218. This finding impacts the numerous previous studies where recombinant soluble BCL-XL was presumed to be full-length. Nevertheless, the majority of recombinant BCL-XL produced in E. coli is insoluble and protected from proteolysis. This protein retains the complete C-terminal tail and can be reconstituted in lipid bilayers in a folded and active state.

Pharmacologic inhibition of RORγt regulates Th17 signature gene expression and suppresses cutaneous inflammation in vivo.

IL-17-producing CD4(+)Th17 cells, CD8(+)Tc17 cells, and γδ T cells play critical roles in the pathogenesis of autoimmune psoriasis. RORγt is required for the differentiation of Th17 cells and expression of IL-17. In this article, we describe a novel, potent, and selective RORγt inverse agonist (TMP778), and its inactive diastereomer (TMP776). This chemistry, for the first time to our knowledge, provides a unique and powerful set of tools to probe RORγt-dependent functions. TMP778, but not TMP776, blocked human Th17 and Tc17 cell differentiation and also acutely modulated IL-17A production and inflammatory Th17-signature gene expression (Il17a, Il17f, Il22, Il26, Ccr6, and Il23) in mature human Th17 effector/memory T cells. In addition, TMP778, but not TMP776, inhibited IL-17A production in both human and mouse γδ T cells. IL-23-induced IL-17A production was also blocked by TMP778 treatment. In vivo targeting of RORγt in mice via TMP778 administration reduced imiquimod-induced psoriasis-like cutaneous inflammation. Further, TMP778 selectively regulated Th17-signature gene expression in mononuclear cells isolated from both the blood and affected skin of psoriasis patients. In summary, to our knowledge, we are the first to demonstrate that RORγt inverse agonists: 1) inhibit Tc17 cell differentiation, as well as IL-17 production by γδ T cells and CD8(+) Tc17 cells; 2) block imiquimod-induced cutaneous inflammation; 3) inhibit Th17 signature gene expression by cells isolated from psoriatic patient samples; and 4) block IL-23-induced IL-17A expression. Thus, RORγt is a tractable drug target for the treatment of cutaneous inflammatory disorders, which may afford additional therapeutic benefit over existing modalities that target only IL-17A.

Improving the mental health of adolescents with epilepsy through a group cognitive behavioral therapy program.

The threat of unpredictable seizures makes epilepsy unique among childhood chronic illnesses. One consequence is that people who have childhood-onset epilepsy often have poor social adjustment and competence in adulthood. Better emotional and social functioning could improve long-term outcomes. Thirty-four adolescents with epilepsy participated in a group cognitive behavioral therapy program designed to enhance their level of psychosocial functioning. Baseline Strength and Difficulty Questionnaire scores suggested that many participants had difficulties with emotions, concentration, and social functioning, with parent-reported Impact scores significantly worse than adolescent-reported scores (p=0.005). Four months after the intervention, adolescent-reported Prosocial Behavior scores significantly improved (p=0.03). Parent-reported scores improved significantly at follow-up, compared with baseline, in Peer Problems (p=0.04), Impact (p=0.001), and Prosocial Behavior (p=0.004) scores. Adolescents with lower socioeconomic status reported the greatest improvements (p=0.01). A brief CBT intervention was effective and resulted in improved mental health indices and social functioning for adolescents with epilepsy.

Quantification of Thiopurine/UVA-Induced Singlet Oxygen Production.

Thiopurines were examined for their ability to produce singlet oxygen ((1)O(2)) with UVA light. The target compounds were three thiopurine prodrugs, azathioprine (Aza), 6-mercaptopurine (6-MP) and 6-thioguanine (6-TG), and their S-methylated derivatives of 6-methylmercaptopurine (me6-MP) and 6-methylthioguanine (me6-TG). Our results showed that these thiopurines were efficient (1)O(2) sensitizers under UVA irradiation but rapidly lost their photoactivities for (1)O(2) production over time by a self-sensitized photooxidation of sulfur atoms in the presence of oxygen and UVA light. The initial quantum yields of (1)O(2) production were determined to be in the range of 0.30-0.6 in aqueous solutions. Substitution of a hydrogen atom with a nitroimidazole or methyl group at S decreased the efficacy of photosensitized (1)O(2) production as found for Aza, me6-MP and me6-TG. (1)O(2)-induced formation of 8-oxo-7,8-dihydro-2'-dexyguanosine (8-oxodGuo) was assessed by incubation of 6-methylthiopurine/UVA-treated calf thymus DNA with human repair enzyme 8-oxodGuo DNA glycosylase (hOGG1), followed by apurinic (AP) site determination. Because more 8-oxodGuo was formed in Tris D(2)O than in Tris H(2)O, (1)O(2) is implicated as a key species in the reaction. These findings provided quantitative information on the photosensitization efficacy of thiopurines and to some extent revealed the correlations between photoactivity and phototoxicity.

The identification of a series of novel, soluble non-peptidic neuropeptide Y Y2 receptor antagonists.

The identification and subsequent optimisation of a selective non-peptidic NPY Y2 antagonist series is described. This led to the development of amine 2, a selective, soluble NPY Y2 receptor antagonist with enhanced CNS exposure.

Author Correction: G Protein-Coupling of Adhesion GPCRs ADGRE2/EMR2 and ADGRE5/CD97, and Activation of G Protein Signalling by an Anti-EMR2 Antibody.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Atypical Reactivation of Varicella Zoster Virus Associated with Pancreatitis in a Heart Transplant Patient.

BACKGROUND Acute pancreatitis is rare following solid organ transplantation but is associated with high mortality. It has been most commonly reported following renal transplant but can occur with other solid organ transplantations. CASE REPORT A 46-year-old male who had an orthotopic heart transplant 6 months ago presented with a 3-week history of abdominal pain. The patient described it as intermittent, sharp, and stabbing, originating in the periumbilical area and radiating to the back. His lipase was elevated at 232 U/L. Given that the patient's symptoms and lipase were elevated to greater than three times the upper limit of normal, he patient was diagnosed with acute pancreatitis. The patient also mentioned a diffuse itchy rash that started a few days prior to admission. Dermatology was consulted, and given the man's clinical presentation, there was concern for atypical reactivation of varicella zoster virus (VZV). VZV polymerase chain reaction of the vesicles returned positive. The patient was started on acyclovir and his symptoms improved. CONCLUSIONS This is the first reported case of VZV-associated pancreatitis in a heart transplant patient. Our patient presented with acute pancreatitis and was treated supportively. However, he did not receive antiviral treatment until his rash was discovered. Timely treatment of VZV resulted in resolution of both the rash and pancreatitis. Timely diagnosis of pancreatitis and VZV is important to prevent development of multiorgan failure and death.

G Protein-Coupling of Adhesion GPCRs ADGRE2/EMR2 and ADGRE5/CD97, and Activation of G Protein Signalling by an Anti-EMR2 Antibody.

The experimental evidence that Adhesion G Protein-Coupled Receptors (aGPCRs) functionally couple to heterotrimeric G proteins has been emerging in incremental steps, but attributing biological significance to their G protein signalling function still presents a major challenge. Here, utilising activated truncated forms of the receptors, we show that ADGRE2/EMR2 and ADGRE5/CD97 are G protein-coupled in a variety of recombinant systems. In a yeast-based assay, where heterologous GPCRs are coupled to chimeric G proteins, EMR2 showed broad G protein-coupling, whereas CD97 coupled more specifically to Gα12, Gα13, Gα14 and Gαz chimeras. Both receptors induced pertussis-toxin (PTX) insensitive inhibition of cyclic AMP (cAMP) levels in mammalian cells, suggesting coupling to Gαz. EMR2 was shown to signal via Gα16, and via a Gα16/Gαz chimera, to stimulate IP1 accumulation. Finally, using an NFAT reporter assay, we identified a polyclonal antibody that activates EMR2 G protein signalling in vitro. Our results highlight the potential for the development of soluble agonists to understand further the biological effects and therapeutic opportunities for ADGRE receptor-mediated G protein signalling.

The identification and optimisation of novel and selective diamide neuropeptide Y Y2 receptor antagonists.

A novel small molecule NPY Y2 antagonist (3) identified from high throughput screening is described. A subsequent SAR study and optimisation programme based around this molecule is also described, leading to the identification of potent and soluble pyridyl analogue 36.

High Throughput Screening Assay to Identify Modulators of IL-17 Expression.

OBJECTIVE: Herein we demonstrate the successful development of a new RORγt-enhanced IL-17F promoter-luciferase reporter assay and its use in a parallel high throughput screening approach, alongside a RORγt TR-FRET assay, to rapidly identify new small molecule RORγt/IL-17 inhibitors and evaluate their mode of action. MATERIAL & METHODS: We sought to identify cell-permeable small-molecule inhibitors of RORγt for rapid progression into hit-to-lead chemistry. As such, we developed the IL-17F promoter luciferase reporter assay in a stable human T-cell (Jurkat) line expressing the RORγt receptor and miniaturised it to a final volume of 8 µL in 1536 well plates for HTS use in screening a library of > 350k compounds. In parallel, a RORγt TR-FRET binding assay was employed to cross-screen the same set of compounds. This enabled the rapid identification of a small number of cell permeable RORγt antagonists showing promising activity in both assays and also highlighted a larger group of potentially very interesting hits which inhibited IL-17 reporter activity, but did not appear to modulate RORγt directly. RESULT: A rigorous triaging process of the novel non-RORγt IL-17 antagonists was followed, making use of in-silico filtering, historical screening data, selectivity screening using an IL-2 reporter assay with an identical cellular background, and final profiling in a phenotypic PBMC IL-17A production assay. This resulted in the identification of a set of promising small molecule compounds which show IL-17 inhibition via potentially novel pathways. CONCLUSION: This technique for the fast identification of cell-permeable IL-17 modulators acting through different mechanisms, highlights the benefits of adopting a parallel approach combining high throughput profiling of hits in multiple assay formats, with robust in-silico triaging.

9alpha,11beta-PGF2 and its stereoisomer PGF2alpha are novel agonists of the chemoattractant receptor, CRTH2.

CRTH2 is a recently described chemoattractant receptor for the prostaglandin, PGD(2), expressed by Th2 cells, eosinophils and basophils, and believed to play a role in allergic inflammation. Here we describe the potency of several PGD(2) metabolites at the receptor to induce cell migration and activation. We report for the first time that the PGD(2) metabolite, 9alpha,11beta-PGF(2), and its stereoisomer, PGF(2alpha), are CRTH2 agonists. 9alpha,11beta-PGF(2) is a major metabolite produced in vivo following allergen challenge, whilst PGF(2alpha) is generated independently of PGD synthetase, with implications for CRTH2 signalling in the presence or absence of PGD(2) production.

Frequency of and risk factors associated with lingual lesions in dogs: 1,196 cases (1995-2004).

OBJECTIVE: To categorize histologic lesions affecting the tongue, determine the frequency with which they develop, and identify risk factors associated with their development in dogs. DESIGN: Retrospective case series. ANIMALS: 1,196 dogs. PROCEDURES: Diagnostic reports of lingual biopsy specimens from dogs evaluated from January 1995 to October 2004 were reviewed. RESULTS: Neoplasia comprised 54% of lingual lesions. Malignant tumors accounted for 64% of lingual neoplasms and included melanoma, squamous cell carcinoma, hemangiosarcoma, and fibrosarcoma. Large-breed dogs, especially Chow Chows and Chinese Shar-Peis, were at increased risk for melanoma. Females of all breeds and Poodles, Labrador Retrievers, and Samoyeds were more likely to have squamous cell carcinomas. Hemangiosarcomas and fibrosarcomas were commonly diagnosed in Border Collies and Golden Retrievers, respectively. Benign neoplasms included squamous papilloma, plasma cell tumor, and granular cell tumor. Small-breed dogs, especially Cocker Spaniels, were at increased risk for plasma cell tumors. Glossitis accounted for 33% of diagnoses; in most cases, the inciting cause was not apparent. Whereas large-breed dogs were more likely to have lingual neoplasia, small-breed dogs were more likely to have glossitis. Calcinosis circumscripta accounted for 4% of lingual lesions and predominately affected young large-breed dogs. The remaining submissions consisted mostly of various degenerative or wound-associated lesions. CONCLUSIONS AND CLINICAL RELEVANCE: The frequency of lingual lesions was not evenly distributed across breeds, sexes, or size classes of dogs. Veterinarians should be aware of the commonly reported lingual lesions in dogs so that prompt diagnosis and appropriate management can be initiated.

Changes in male partner abuser attachment styles in group treatment.

This study examined attachment pattern change of partner violent men over the course of 17 weeks of integrated cognitive-behavioral/psychodynamic group treatment. Results indicated a significant increase in the number of men reporting a secure attachment from pre- to posttreatment. Subsequent analysis indicated that secure-changed men reported significant increases in comfort with closeness and depending on others from pre- to posttreatment. From pre- to posttreatment, insecure and secure-unchanged men reported an increase in avoidance of closeness. Additionally, secure-changed men reported lower anxiety and depression than insecure men at posttreatment. Finally, the total sample reported significant reductions in partner violence from pre to posttreatment. Based on these results, it would be important for clinicians and researchers to distinguish between men who changed to a secure attachment pattern over the course of treatment and securely attached men who remained unchanged. (PsycINFO Database Record (c) 2010 APA, all rights reserved).

RFX transcription factors are essential for hearing in mice.

Sensorineural hearing loss is a common and currently irreversible disorder, because mammalian hair cells (HCs) do not regenerate and current stem cell and gene delivery protocols result only in immature HC-like cells. Importantly, although the transcriptional regulators of embryonic HC development have been described, little is known about the postnatal regulators of maturating HCs. Here we apply a cell type-specific functional genomic analysis to the transcriptomes of auditory and vestibular sensory epithelia from early postnatal mice. We identify RFX transcription factors as essential and evolutionarily conserved regulators of the HC-specific transcriptomes, and detect Rfx1,2,3,5 and 7 in the developing HCs. To understand the role of RFX in hearing, we generate Rfx1/3 conditional knockout mice. We show that these mice are deaf secondary to rapid loss of initially well-formed outer HCs. These data identify an essential role for RFX in hearing and survival of the terminally differentiating outer HCs.

A coding polymorphism in the CYSLT2 receptor with reduced affinity to LTD4 is associated with asthma.

BACKGROUND: Cysteinyl leukotrienes (CYSLTR) are potent biological mediators in the pathophysiology of asthma for which two receptors have been characterized, CYSLTR1 and CYSLTR2. The leukotriene modifying agents currently used to control bronchoconstriction and inflammation in asthmatic patients are CYSLTR1-specific leukotriene receptor antagonists. In this report, we investigated a possible role for therapeutic modulation of CYSLTR2 in asthma by investigating genetic association with asthma and further characterization of the pharmacology of a coding polymorphism. METHODS: The association of CYSLTR2 polymorphisms with asthma was assessed by transmission disequilibrium test in two family-based collections (359 families from Denmark and Minnesota, USA and 384 families from the Genetics of Asthma International Network). RESULTS: A significant association of the coding polymorphism, 601A>G, with asthma was observed (P = 0.003). We replicated these findings in a collection of 384 families from the Genetics of Asthma International Network (P = 0.04). The G allele is significantly under-transmitted to asthmatics, indicating a possible role for this receptor in resistance to asthma. The potency of cysteinyl leukotrienes at the wild-type CYSLTR2 and the coding polymorphism 601A>G were assessed using a calcium mobilization assay. The potency of LTC4 and LTE4 was similar for both forms of the receptor and LTB4 was inactive, however, LTD4 was approximately five-fold less potent on 601A>G compared to wild-type CYSLTR2. CONCLUSIONS: Since 601A>G alters the potency of LTD4 and this variant allele may be associated with resistance to asthma, it is possible that modulation of the CYSLTR2 may be useful in asthma pharmacotherapy.

Internalization of the chemokine receptor CCR4 can be evoked by orthosteric and allosteric receptor antagonists.

The chemokine receptor CCR4 has at least two natural agonist ligands, MDC (CCL22) and TARC (CCL17) which bind to the same orthosteric site with a similar affinity. Both ligands are known to evoke chemotaxis of CCR4-bearing T cells and also elicit CCR4 receptor internalization. A series of small molecule allosteric antagonists have been described which displace the agonist ligand, and inhibit chemotaxis. The aim of this study was to determine which cellular coupling pathways are involved in internalization, and if antagonists binding to the CCR4 receptor could themselves evoke receptor internalization. CCL22 binding coupled CCR4 efficiently to ß-arrestin and stimulated GTPγS binding however CCL17 did not couple to ß-arrestin and only partially stimulated GTPγS binding. CCL22 potently induced internalization of almost all cell surface CCR4, while CCL17 showed only weak effects. We describe four small molecule antagonists that were demonstrated to bind to two distinct allosteric sites on the CCR4 receptor, and while both classes inhibited agonist ligand binding and chemotaxis, one of the allosteric sites also evoked receptor internalization. Furthermore, we also characterize an N-terminally truncated version of CCL22 which acts as a competitive antagonist at the orthosteric site, and surprisingly also evokes receptor internalization without demonstrating any agonist activity. Collectively this study demonstrates that orthosteric and allosteric antagonists of the CCR4 receptor are capable of evoking receptor internalization, providing a novel strategy for drug discovery against this class of target.

Antagonism of human CC-chemokine receptor 4 can be achieved through three distinct binding sites on the receptor.

Chemokine receptor antagonists appear to access two distinct binding sites on different members of this receptor family. One class of CCR4 antagonists has been suggested to bind to a site accessible from the cytoplasm while a second class did not bind to this site. In this report, we demonstrate that antagonists representing a variety of structural classes bind to two distinct allosteric sites on CCR4. The effects of pairs of low-molecular weight and/or chemokine CCR4 antagonists were evaluated on CCL17- and CCL22-induced responses of human CCR4(+) T cells. This provided an initial grouping of the antagonists into sets which appeared to bind to distinct binding sites. Binding studies were then performed with radioligands from each set to confirm these groupings. Some novel receptor theory was developed to allow the interpretation of the effects of the antagonist combinations. The theory indicates that, generally, the concentration-ratio of a pair of competing allosteric modulators is maximally the sum of their individual effects while that of two modulators acting at different sites is likely to be greater than their sum. The low-molecular weight antagonists could be grouped into two sets on the basis of the functional and binding experiments. The antagonistic chemokines formed a third set whose behaviour was consistent with that of simple competitive antagonists. These studies indicate that there are two allosteric regulatory sites on CCR4.

Molecular identification of high and low affinity receptors for nicotinic acid.

Nicotinic acid has been used clinically for over 40 years in the treatment of dyslipidemia producing a desirable normalization of a range of cardiovascular risk factors, including a marked elevation of high density lipoprotein and a reduction in mortality. The precise mechanism of action of nicotinic acid is unknown, although it is believed that activation of a G(i)-G protein-coupled receptor may contribute. Utilizing available information on the tissue distribution of nicotinic acid receptors, we identified candidate orphan receptors. The selected orphan receptors were screened for responses to nicotinic acid, in an assay for activation of G(i)-G proteins. Here we describe the identification of the G protein-coupled receptor HM74 as a low affinity receptor for nicotinic acid. We then describe the subsequent identification of HM74A in follow-up bioinformatics searches and demonstrate that it acts as a high affinity receptor for nicotinic acid and other compounds with related pharmacology. The discovery of HM74A as a molecular target for nicotinic acid may facilitate the discovery of superior drug molecules to treat dyslipidemia.