Mastigoneme structure reveals insights into the O-linked glycosylation code of native hydroxyproline-rich helices.

Hydroxyproline-rich glycoproteins (HRGPs) are a ubiquitous class of protein in the extracellular matrices and cell walls of plants and algae, yet little is known of their native structures or interactions. Here, we used electron cryomicroscopy (cryo-EM) to determine the structure of the hydroxyproline-rich mastigoneme, an extracellular filament isolated from the cilia of the alga Chlamydomonas reinhardtii. The structure demonstrates that mastigonemes are formed from two HRGPs (a filament of MST1 wrapped around a single copy of MST3) that both have hyperglycosylated poly(hydroxyproline) helices. Within the helices, O-linked glycosylation of the hydroxyproline residues and O-galactosylation of interspersed serine residues create a carbohydrate casing. Analysis of the associated glycans reveals how the pattern of hydroxyproline repetition determines the type and extent of glycosylation. MST3 possesses a PKD2-like transmembrane domain that forms a heteromeric polycystin-like cation channel with PKD2 and SIP, explaining how mastigonemes are tethered to ciliary membranes.

Structural determination and modeling of ciliary microtubules.

The axoneme, a microtubule-based array at the center of every cilium, has been the subject of structural investigations for decades, but only recent advances in cryo-EM and cryo-ET have allowed a molecular-level interpretation of the entire complex to be achieved. The unique properties of the nine doublet microtubules and central pair of singlet microtubules that form the axoneme, including the highly decorated tubulin lattice and the docking of massive axonemal complexes, provide opportunities and challenges for sample preparation, 3D reconstruction and atomic modeling. Here, the approaches used for cryo-EM and cryo-ET of axonemes are reviewed, while highlighting the unique opportunities provided by the latest generation of AI-guided tools that are transforming structural biology.

Automated model building and protein identification in cryo-EM maps.

Interpreting electron cryo-microscopy (cryo-EM) maps with atomic models requires high levels of expertise and labour-intensive manual intervention in three-dimensional computer graphics programs1,2. Here we present ModelAngelo, a machine-learning approach for automated atomic model building in cryo-EM maps. By combining information from the cryo-EM map with information from protein sequence and structure in a single graph neural network, ModelAngelo builds atomic models for proteins that are of similar quality to those generated by human experts. For nucleotides, ModelAngelo builds backbones with similar accuracy to those built by humans. By using its predicted amino acid probabilities for each residue in hidden Markov model sequence searches, ModelAngelo outperforms human experts in the identification of proteins with unknown sequences. ModelAngelo will therefore remove bottlenecks and increase objectivity in cryo-EM structure determination.

Vitamin D levels of pregnant immigrant women and developmental disorders of language, learning and coordination in offspring.

BACKGROUND: Prenatal vitamin D deficiency is a common health concern among immigrants. No previous studies have examined the associations between prenatal vitamin D levels and developmental disorders of language, scholastic skills, and coordination in an immigrant sample. METHODS: The sample included 542 immigrant mothers of cases with language, scholastic, coordination or mixed developmental disorders, 443 immigrant mothers of controls without these disorders and 542 Finnish mothers of controls. Maternal vitamin D was measured in serum samples collected during early pregnancy and stored in a national biobank. RESULTS: The mean vitamin D levels during pregnancy were 25.0 (SD 14.4) nmol/L for immigrant mothers of cases, 25.4 (SD 15.5) for immigrant mothers of controls and 42.3 (SD 19.1) for Finnish mothers of controls. Low maternal vitamin D levels during pregnancy were not associated with the selected developmental disorders in offspring when immigrant mothers of cases were compared to immigrant mothers of controls (adjusted OR for continuous log-transformed vitamin D: 1.01, 95% CI 0.75-1.36, p = 0.96). When immigrant mothers of cases were compared to Finnish mothers of controls, the adjusted OR for continuous vitamin D was 18.94 (95% CI 11.47-31.25), p <0.001). The results were similar when vitamin D was examined as a categorical variable or divided into quintiles. CONCLUSIONS: Prenatal vitamin D levels were low, and similar, among immigrant mothers of cases with selected developmental disorders and unaffected controls. This indicates that vitamin D unlikely mediates previously observed associations between maternal immigrant status and the selected developmental disorders in offspring. The proportion of immigrant mothers with severe vitamin D deficiency was very high, which underlines the importance of prenatal counselling and overall public health efforts to improve immigrant health.

Electrophysiological Changes in the Rabbit Ventricular Wedge and Human-Induced Pluripotent Stem-Cell Derived (IPSC) Cardiomyocytes Translate to Severe Arrhythmia Observed in a Canine Toxicology Study, Not Predicted by Standard In Vitro Ion Channel Assays.

During drug discovery, small molecules are typically assayed in vitro for secondary pharmacology effects, which include ion channels relevant to cardiac electrophysiology. Compound A was an irreversible inhibitor of myeloperoxidase investigated for the treatment of peripheral artery disease. Oral doses in dogs at ≥5 mg/kg resulted in cardiac arrhythmias in a dose-dependent manner (at Cmax, free ≥1.53 µM) that progressed in severity with time. Nevertheless, a panel of 13 different cardiac ion channel (K, Na, and Ca) assays, including hERG, failed to identify pharmacologic risks of the molecule. Compound A and a related Compound B were evaluated for electrophysiological effects in the isolated rabbit ventricular wedge assay. Compounds A and B prolonged QT and Tp-e intervals at ≥1 and ≥.3 µM, respectively, and both prolonged QRS at ≥5 µM. Compound A produced early after depolarizations and premature ventricular complexes at ≥5 µM. These data indicate both compounds may be modulating hERG (Ikr) and Nav1.5 ion channels. In human IPSC cardiomyocytes, Compounds A and B prolonged field potential duration at ≥3 µM and induced cellular dysrhythmia at ≥10 and ≥3 µM, respectively. In a rat toxicology study, heart tissue: plasma concentration ratios for Compound A were ≥19X at 24 hours post-dose, indicating significant tissue distribution. In conclusion, in vitro ion channel assays may not always identify cardiovascular electrophysiological risks observed in vivo, which can be affected by tissue drug distribution. Risk for arrhythmia may increase with a "trappable" ion channel inhibitor, particularly if cardiac tissue drug levels achieve a critical threshold for pharmacologic effects.

Species-specific Bioactivation of Morpholines as a Causative of Drug Induced Liver Injury Observed in Monkeys

BACKGROUND: Everolimus, an allosteric mechanistic target of rapamycin (mTOR) inhibitor, recently demonstrated the therapeutic value of mTOR inhibitors for Central Nervous System (CNS) indications driven by hyperactivation of mTOR. A newer, potent brain-penetrant analog of everolimus, referred to as (1) in this manuscript [(S)-3-methyl-4-(7-((R)-3-methylmorpholino)-2-(thiazol-4-yl)-3H-imidazo[4,5-b]pyridin-5-yl)morpholine,(1)] catalytically inhibits mTOR function in the brain and increases the lifespan of mice with neuronal mTOR hyperactivation. INTRODUCTION: Early evaluation of the safety of 1 was conducted in cynomolgus monkeys in which oral doses were administered to three animals in a rising-dose fashion (from 2 to 30 mg/kg/day). 1 produced severe toxicity including the evidence of hepatic toxicity, along with non-dose proportional increases in drug exposure. Investigations of cross-species hepatic bioactivation of 1 were conducted to assess whether the formation of reactive drug metabolites was associated with the mechanism of liver toxicity. METHOD: 1 contained two morpholine rings known as structural alerts and can potentially form reactive intermediates through oxidative metabolism. Bioactivation of 1 was investigated in rat, human and monkey liver microsomes fortified with trapping agents such as methoxylamine or potassium cyanide. RESULTS: Our results suggest that bioactivation of the morpholine moieties to reactive intermediates may have been involved in the mechanism of liver toxicity observed with 1. Aldehyde intermediates trappable by methoxylamine were identified in rat and monkey liver microsomal studies. In addition, a total of four cyano conjugates arising from the formation of iminium ion intermediates were observed and identified. These findings may potentially explain the observed monkey toxicity. Interestingly, methoxylamine or cyano adducts of 1 were not observed in human liver microsomes. CONCLUSION: The bioactivation of 1 appears to be species-specific. Circumstantial evidence for the toxicity derived from 1 point to the formation of iminium ion intermediates trappable by cyanide in monkey liver microsomes. The cyano conjugates were only observed in monkey liver microsomes, potentially pointing to cause at least the hepatotoxicity observed in monkeys. In contrast, methoxylamine conjugates were detected in both rat and monkey liver microsomes, with only a trace amount in human liver microsomes. Cyano conjugates were not observed in human liver microsomes, challenging the team on the drugability and progressivity of 1 through drug development. The mechanisms for drug-induced liver toxicity are multifactorial. These results are highly suggestive that the iminium ion may be an important component in the mechanism of liver toxicity 1 observed in the monkey.

Recent synthetic strategies of spiro-azetidin-2-one, -pyrrolidine, -indol(one) and -pyran derivatives-a review.

Spiro-heterocycles have received special attention in medicinal chemistry because of their promising biological activity. Over the years, many synthetic methodologies have been established for the construction of spirocyclic compounds. Spiro heterocycles such as spiro-azetidin-2-one, -pyrrolidine, -indol(one) and -pyran derivatives have been found to exhibit diversified biological and pharmacological activity in addition to their therapeutic properties. In view of these facts, we decided in this review to present representative synthetic approaches of the aforementioned spiro heterocycles, especially in the past 20 years.

The Role of Emotion Regulation, Affect, and Sleep in Individuals With Sleep Bruxism and Those Without: Protocol for a Remote Longitudinal Observational Study.

BACKGROUND: Sleep bruxism (SB) is an oral behavior characterized by high levels of repetitive jaw muscle activity during sleep, leading to teeth grinding and clenching, and may develop into a disorder. Despite its prevalence and negative outcomes on oral health and quality of life, there is currently no cure for SB. The etiology of SB remains poorly understood, but recent research suggests a potential role of negative emotions and maladaptive emotion regulation (ER). OBJECTIVE: This study's primary aim investigates whether ER is impaired in individuals with SB, while controlling for affective and sleep disturbances. The secondary aim tests for the presence of cross-sectional and longitudinal mediation pathways in the bidirectional relationships among SB, ER, affect, and sleep. METHODS: The study used a nonrandomized repeated-measures observational design and was conducted remotely. Participants aged 18-49 years underwent a 14-day ambulatory assessment. Data collection was carried out using electronic platforms. We assessed trait and state SB and ER alongside affect and sleep variables. We measured SB using self-reported trait questionnaires, ecological momentary assessment (EMA) for real-time reports of SB behavior, and portable electromyography for multinight assessment of rhythmic masticatory muscle activity. We assessed ER through self-reported trait questionnaires, EMA for real-time reports of ER strategies, and heart rate variability derived from an electrocardiography wireless physiological sensor as an objective physiological measure. Participants' trait affect and real-time emotional experiences were obtained using self-reported trait questionnaires and EMA. Sleep patterns and quality were evaluated using self-reported trait questionnaires and sleep diaries, as well as actigraphy as a physiological measure. For the primary objective, analyses will test for maladaptive ER in terms of strategy use frequency and effectiveness as a function of SB using targeted contrasts in the general linear model. Control analyses will be conducted to examine the persistence of the SB-ER relationship after adjusting for affective and sleep measures, as well as demographic variables. For the secondary objective, cross-sectional and longitudinal mediation analyses will test various competing models of directional effects among self-reported and physiological measures of SB, ER, affect, and sleep. RESULTS: This research received funding in April 2017. Data collection took place from August 2020 to March 2022. In all, 237 participants were eligible and completed the study. Data analysis has not yet started. CONCLUSIONS: We hope that the effort to thoroughly measure SB and ER using gold standard methods and cutting-edge technology will advance the knowledge of SB. The findings of this study may contribute to a better understanding of the relationship among SB, ER, affect, and sleep disturbances. By identifying the role of ER in SB, the results may pave the way for the development of targeted interventions for SB management to alleviate the pain and distress of those affected. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID): DERR1-10.2196/41719.

Quantitative versus qualitative emotion regulation goals: Differential effects on emotional responses.

Emotion regulation (ER) involves both a goal (e.g., to feel less emotion) and a strategy (e.g., reappraisal). To clarify the impact of ER goals on emotional responding, we conducted a within-participant study (N = 156) in which we held the strategy constant (reappraisal) to isolate the impact of regulation goals. We compared the impact of a quantitative goal (changing emotion quantity/intensity) with that of a qualitative goal (changing emotion quality/type) on emotional responses to negative and positive pictures. We manipulated ER goals by cuing participants to continue viewing the picture (unregulated/no ER goal) or to reappraise it to decrease its predominant affective impact (quantitative goal) or increase its opposite-valence impact (qualitative goal). We assessed emotional responses through self-reported feelings and facial expressions (corrugator supercilii and zygomaticus major electromyography). Our findings suggest that the type of regulation goal has a differential effect on emotional responses, with qualitative goals being more effective in modulating both negative and positive emotions. For negative stimuli, attempts to use a quantitative goal decreased negative but not positive emotional responses (uncoupled negative deactivation). Conversely, attempts to use a qualitative goal decreased negative and increased positive emotional responses (reciprocal positive activation). For positive stimuli, the quantitative goal generated uncoupled positive deactivation, while the qualitative goal produced reciprocal negative activation. Results highlight the importance of considering specific regulation goals in shaping emotional responses. Future research in the field of ER may benefit from identifying and manipulating different goals and strategies to understand how to effectively regulate emotions.

Automated model building and protein identification in cryo-EM maps.

Interpreting electron cryo-microscopy (cryo-EM) maps with atomic models requires high levels of expertise and labour-intensive manual intervention. We present ModelAngelo, a machine-learning approach for automated atomic model building in cryo-EM maps. By combining information from the cryo-EM map with information from protein sequence and structure in a single graph neural network, ModelAngelo builds atomic models for proteins that are of similar quality as those generated by human experts. For nucleotides, ModelAngelo builds backbones with similar accuracy as humans. By using its predicted amino acid probabilities for each residue in hidden Markov model sequence searches, ModelAngelo outperforms human experts in the identification of proteins with unknown sequences. ModelAngelo will thus remove bottlenecks and increase objectivity in cryo-EM structure determination.

A Saliva-Based Serological and Behavioral Analysis of SARS-CoV-2 Antibody Prevalence in Howard County, Maryland.

The objective of the study was to estimate severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) seroprevalence in the Howard County, Maryland, general population and demographic subpopulations attributable to natural infection or coronavirus disease 2019 (COVID-19) vaccination and to identify self-reported social behaviors that may affect the likelihood of recent or past SARS-CoV-2 infection. A cross-sectional, saliva-based serological study of 2,880 residents of Howard County, Maryland, was carried out from July through September 2021. Natural SARS-CoV-2 infection prevalence was estimated by inferring infections among individuals according to anti-nucleocapsid immunoglobin G levels and calculating averages weighted by sample proportions of various demographics. Antibody levels between BNT162b2 (Pfizer-BioNTech) and mRNA-1273 (Moderna) recipients were compared. Antibody decay rate was calculated by fitting exponential decay curves to cross-sectional indirect immunoassay data. Regression analysis was carried out to identify demographic factors, social behaviors, and attitudes that may be linked to an increased likelihood of natural infection. The estimated overall prevalence of natural infection in Howard County, Maryland, was 11.9% (95% confidence interval, 9.2% to 15.1%), compared with 7% reported COVID-19 cases. Antibody prevalence indicating natural infection was highest among Hispanic and non-Hispanic Black participants and lowest among non-Hispanic White and non-Hispanic Asian participants. Participants from census tracts with lower average household income also had higher natural infection rates. After accounting for multiple comparisons and correlations between participants, none of the behavior or attitude factors had significant effects on natural infection. At the same time, recipients of the mRNA-1273 vaccine had higher antibody levels than those of BNT162b2 vaccine recipients. Older study participants had overall lower antibody levels compared with younger study participants. The true prevalence of SARS-CoV-2 infection is higher than the number of reported COVID-19 cases in Howard County, Maryland. A disproportionate impact of infection-induced SARS-CoV-2 positivity was observed across different ethnic/racial subpopulations and incomes, and differences in antibody levels across different demographics were identified. Taken together, this information may inform public health policy to protect vulnerable populations. IMPORTANCE We employed a highly innovative noninvasive multiplex oral fluid SARS-CoV-2 IgG assay to ascertain our seroprevalence estimates. This laboratory-developed test has been applied in NCI's SeroNet consortium, possesses high sensitivity and specificity according to FDA Emergency Use Authorization guidelines, correlates strongly with SARS-CoV-2 neutralizing antibody responses, and is Clinical Laboratory Improvement Amendments-approved by the Johns Hopkins Hospital Department of Pathology. It represents a broadly scalable public health tool to improve understanding of recent and past SARS-CoV-2 exposure and infection without drawing any blood. To our knowledge, this is the first application of a high-performance salivary SARS-CoV-2 IgG assay to estimate population-level seroprevalence, including identifying COVID-19 disparities. We also are the first to report differences in SARS-CoV-2 IgG responses by COVID-19 vaccine manufacturers (BNT162b2 [Pfizer-BioNTech] and mRNA-1273 [Moderna]). Our findings demonstrate remarkable consistency with those of blood-based SARS-CoV-2 IgG assays in terms of differences in the magnitude of SARS-CoV-2 IgG responses between COVID-19 vaccines.

Structural specializations of the sperm tail.

Sperm motility is crucial to reproductive success in sexually reproducing organisms. Impaired sperm movement causes male infertility, which is increasing globally. Sperm are powered by a microtubule-based molecular machine-the axoneme-but it is unclear how axonemal microtubules are ornamented to support motility in diverse fertilization environments. Here, we present high-resolution structures of native axonemal doublet microtubules (DMTs) from sea urchin and bovine sperm, representing external and internal fertilizers. We identify >60 proteins decorating sperm DMTs; at least 15 are sperm associated and 16 are linked to infertility. By comparing DMTs across species and cell types, we define core microtubule inner proteins (MIPs) and analyze evolution of the tektin bundle. We identify conserved axonemal microtubule-associated proteins (MAPs) with unique tubulin-binding modes. Additionally, we identify a testis-specific serine/threonine kinase that links DMTs to outer dense fibers in mammalian sperm. Our study provides structural foundations for understanding sperm evolution, motility, and dysfunction at a molecular level.

Novel quinoline/thiazinan-4-one hybrids; design, synthesis, and molecular docking studies as potential anti-bacterial candidates against MRSA.

In an attempt to develop effective and safe antibacterial agents, we synthesized novel thiazinanones by combining the quinolone scaffold and the 1,3-thiazinan-4-one group by reaction between ((4-hydroxy-2-oxo-1,2-dihydroquinolin-3-yl)methylene)hydrazinecarbothioamides and 2,3-diphenylcycloprop-2-enone in refluxing ethanol in the presence of triethyl amine as a catalyst. The structure of the synthesized compounds was characterized by spectral data and elemental analysis, IR, MS, 1H and 13C NMR spectroscopy which showed two doublet signals for CH-5 and CH-6 and four sharp singlets for the protons of thiazinane NH, CH[double bond, length as m-dash]N, quinolone NH and OH, respectively. Also, the 13C NMR spectrum clearly showed the presence of two quaternary carbon atoms which were assigned to thiazinanone-C-5 and C-6. All the 1,3-thiazinan-4-one/quinolone hybrids were screened for antibacterial activity. Compounds 7a, 7e and 7g showed broad spectrum antibacterial activity against most of the tested strains either G +ve or G -ve. Compound 7e is the most potent antibacterial agent against MRSA with the minimum inhibitory concentration against MRSA found to be 48 µg mL-1 compared to the drug ciprofloxacin (96 µg mL-1). Additionally, a molecular docking study was performed to understand the molecular interaction and binding mode of the compounds on the active site of S. aureus Murb protein. In silico docking assisted data strongly correlated with the experimental approach of antibacterial activity against MRSA.

Prenatal organochlorine pollutant exposure and risk of schizophrenia in a national birth cohort.

Non-genetic prenatal exposures have been associated with schizophrenia risk. However, the role of prenatal exposure to environmental neurotoxicants in offspring schizophrenia risk has been studied in only limited instances. Polychlorinated biphenyls (PCBs) and the pesticide metabolite p,p'-dichlorodiphenyl dichloroethylene (DDE) have been linked to neurodevelopmental outcomes, including impairments implicated in schizophrenia. To determine whether prenatal maternal levels of organochlorine pollutants including PCBs or DDE are associated with schizophrenia in the offspring, an investigation was conducted in the Finnish Prenatal Study of Schizophrenia (FIPS-S), a case-control study nested in a national birth cohort. Cases were born in 1987-1991 and had at least two diagnoses of schizophrenia (ICD-10 F20; ICD-9 295) or schizoaffective disorder (ICD-10 F25; ICD-9 295.7) recorded in the national Care Register for Health Care. Each case was individually matched to a control on sex, date of birth, and residence in Finland on the date of case diagnosis. In 500 case-control pairs, PCB congeners 74, 99, 118, 138, 153, 156, 170, 180, 183, 187, and some widespread organochlorine pesticides or their metabolites including DDE were measured in archived prenatal maternal sera using gas chromatography - high triple quadrupole mass spectrometry. Maternal total PCBs were quantified as the sum of concentrations of the measured congeners. Associations with schizophrenia were examined using conditional logistic regression. Maternal PCB or DDE levels greater than the 75th percentiles of the control distributions showed no evidence of association with offspring schizophrenia (PCBs: adjusted odds ratio (aOR) = 1.13, 95 % CI = 0.85-1.50), p = 0.41; DDE: aOR = 1.08, 95 % CI = 0.80-1.45; p = 0.63). Maternal levels of either pollutant dichotomized at the 90th percentile or considered as a continuous variable also did not show evidence for association with offspring schizophrenia. This study found a lack of evidence that prenatal maternal levels of the organochlorine pollutants DDE and PCBs are associated with offspring risk of schizophrenia.

Axonemal structures reveal mechanoregulatory and disease mechanisms.

Motile cilia and flagella beat rhythmically on the surface of cells to power the flow of fluid and to enable spermatozoa and unicellular eukaryotes to swim. In humans, defective ciliary motility can lead to male infertility and a congenital disorder called primary ciliary dyskinesia (PCD), in which impaired clearance of mucus by the cilia causes chronic respiratory infections1. Ciliary movement is generated by the axoneme, a molecular machine consisting of microtubules, ATP-powered dynein motors and regulatory complexes2. The size and complexity of the axoneme has so far prevented the development of an atomic model, hindering efforts to understand how it functions. Here we capitalize on recent developments in artificial intelligence-enabled structure prediction and cryo-electron microscopy (cryo-EM) to determine the structure of the 96-nm modular repeats of axonemes from the flagella of the alga Chlamydomonas reinhardtii and human respiratory cilia. Our atomic models provide insights into the conservation and specialization of axonemes, the interconnectivity between dyneins and their regulators, and the mechanisms that maintain axonemal periodicity. Correlated conformational changes in mechanoregulatory complexes with their associated axonemal dynein motors provide a mechanism for the long-hypothesized mechanotransduction pathway to regulate ciliary motility. Structures of respiratory-cilia doublet microtubules from four individuals with PCD reveal how the loss of individual docking factors can selectively eradicate periodically repeating structures.

The N-Acetylgalactosamine-conjugated small interfering RNA inclisiran can be coadministered safely with atorvastatin in cynomolgus monkeys resulting in additive low-density lipoprotein cholesterol reductions.

Inclisiran, a small interfering RNA, selectively inhibits proprotein convertase subtilisin/kexin type 9 (PCSK9) synthesis in the liver and has been shown to reduce low-density lipoprotein cholesterol (LDL-C) by ≥50% in patients with hypercholesterolemia receiving maximally tolerated statins. The toxicokinetic, pharmacodynamic, and safety profiles of inclisiran when coadministered with a statin were characterized in cynomolgus monkeys. Six cohorts of monkeys were administered either atorvastatin (40 mg/kg, reduced to 25 mg/kg during the study, daily, oral gavage), inclisiran (300 mg/kg every 28 days, subcutaneous administration), atorvastatin (40/25 mg/kg) and inclisiran combinations (30, 100, or 300 mg/kg), or control vehicles over 85 days followed by 90 days' recovery. Inclisiran and atorvastatin toxicokinetic parameters were similar in cohorts administered either agent alone or in combination. Inclisiran exposure increased in a dose-proportional manner. At Day 86, atorvastatin increased plasma PCSK9 levels four-fold from pretreatment levels but did not significantly lower serum LDL-C levels. Inclisiran (alone or in combination) reduced PCSK9 (mean decrease 66-85%) and LDL-C (mean decrease 65-92%) from pretreatment levels at Day 86; levels were significantly lower than the control group (p ≤ .05) and remained decreased during the 90-day recovery. Coadministration of inclisiran with atorvastatin resulted in greater reductions in LDL-C and total cholesterol compared with either drug alone. No toxicities or adverse effects were observed in any cohort receiving inclisiran, either alone or in combination. In summary, inclisiran significantly inhibited PCSK9 synthesis and decreased LDL-C in cynomolgus monkeys without increasing the risk of adverse effects when coadministered with atorvastatin.

Maternal Serum Vitamin B12 during Pregnancy and Offspring Autism Spectrum Disorder.

This study examined the association between maternal serum vitamin B12 levels during early pregnancy and offspring autism spectrum disorders (ASD) and subtypes. Based on a Finnish national birth cohort, case offspring (n = 1558) born in 1987-2007 and diagnosed with ASD by 2015 were matched with one control on date of birth, sex and place of birth. Maternal vitamin B12 levels were measured during first and early second trimesters of pregnancy. High maternal vitamin B12 levels (≥81th percentile) was associated with increased risk for offspring childhood autism, adjusted odds ratio, 1.59, 95% confidence interval 1.06-2.41 (p = 0.026). No significant associations were observed between maternal vitamin B12 levels and offspring Asperger's or pervasive developmental disorder/NOS.

Maternal SSRI use during pregnancy and offspring depression or anxiety disorders: A review of the literature and description of a study protocol for a register-based cohort study.

Previous studies examining the relationship between in utero exposure to selective serotonin reuptake inhibitors (SSRI) and long-term offspring depressive or anxiety behaviors are inconclusive. We aimed to critically review the findings of previous studies and describe a new study protocol to investigate the association of prenatal SSRI exposure and offspring depression or anxiety using data from several Finnish national registers. The study includes 1,266,473 mothers and their live-born singleton offspring, born in 1996-2018. The study cohorts include the prenatally SSRI exposed group and three comparison groups: 1) depression exposed/antidepressants unexposed, 2) unexposed to antidepressants or antipsychotics and depression, and 3) discordant siblings. We aim to examine whether depression in prenatally SSRI exposed children is more common or severe than depression in the offspring of mothers with depression but without SSRI exposure. We aim to disambiguate the effects of maternal SSRI from the effects of maternal depression, severity of maternal depression and familial loading history of psychiatric disorders by including data from first-degree relatives of prenatally SSRI exposed and unexposed children. Associations between exposure and outcome are assessed by statistical modeling, accounting for within-family correlation. The study has potential public health significance and in guiding clinicians in considering treatment options for pregnant women.

Discovery of a novel, highly potent and orally bioavailable pyrrolidinone indole series of irreversible Myeloperoxidase (MPO) inhibitors.

Myeloperoxidase (MPO) is a heme-containing peroxidase from phagocytic cells, which plays an important role in the innate immune response. The primary anti-microbial function of MPO is achieved by catalyzing the oxidation of halides by hydrogen peroxide (H2O2). Upon activation of phagocytes, MPO activity is detectable in both phagosomes and extracellularly, where it can remain or transcytose into interstitial compartments. Activated MPO leads to oxidative stress and tissue damage in many inflammatory states, including cardiovascular disease. Starting from a low molecular weight (LMW) high throughput screening (HTS) hit, here we report the discovery of a novel pyrrolidinone indole (IN-4) as a highly potent MPO inhibitor. This compound displays similar in vitro potency across peroxidation, plasma and NETosis assays. In a dilution/dialysis study, <5% of the original MPO activity was detected post-incubation of MPO with IN-4, suggesting irreversible enzyme inhibition. A fast MPO inactivation rate (kinact/Ki) and low partition ratio (k3/k4) make IN-4 kinetic properties attractive for an MPO inhibitor. This compound also displays significant selectivity over the closely related thyroid peroxidase (TPO), and is selective for extracellular MPO over intracellular (neutrophil) MPO. Moreover, IN-4 shows good exposure, low clearance and high oral bioavailability in mice, rats and dogs. The high in vitro MPO activity and high oral exposure observed with IN-4 result in a dose-dependent inhibition of MPO activity in three mouse models of inflammation. In conclusion, IN-4 is a novel, potent, mechanism-based and selective MPO inhibitor, which may be used as superior therapeutic agent to treat multiple inflammatory conditions, including cardiovascular disease.

Mechanism of IFT-A polymerization into trains for ciliary transport.

Intraflagellar transport (IFT) is the highly conserved process by which proteins are transported along ciliary microtubules by a train-like polymeric assembly of IFT-A and IFT-B complexes. IFT-A is sandwiched between IFT-B and the ciliary membrane, consistent with its putative role in transporting transmembrane and membrane-associated cargoes. Here, we have used single-particle analysis electron cryomicroscopy (cryo-EM) to determine structures of native IFT-A complexes. We show that subcomplex rearrangements enable IFT-A to polymerize laterally on anterograde IFT trains, revealing a cooperative assembly mechanism. Surprisingly, we discover that binding of IFT-A to IFT-B shields the preferred lipid-binding interface from the ciliary membrane but orients an interconnected network of ß-propeller domains with the capacity to accommodate diverse cargoes toward the ciliary membrane. This work provides a mechanistic basis for understanding IFT-train assembly and cargo interactions.