Identification of genetic overlap and novel risk loci for attention-deficit/hyperactivity disorder and bipolar disorder.

Differential diagnosis between childhood onset attention-deficit/hyperactivity disorder (ADHD) and bipolar disorder (BD) remains a challenge, mainly due to overlapping symptoms and high rates of comorbidity. Despite this, genetic correlation reported for these disorders is low and non-significant. Here we aimed to better characterize the genetic architecture of these disorders utilizing recent large genome-wide association studies (GWAS). We analyzed independent GWAS summary statistics for ADHD (19,099 cases and 34,194 controls) and BD (20,352 cases and 31,358 controls) applying the conditional/conjunctional false discovery rate (condFDR/conjFDR) statistical framework that increases the power to detect novel phenotype-specific and shared loci by leveraging the combined power of two GWAS. We observed cross-trait polygenic enrichment for ADHD conditioned on associations with BD, and vice versa. Leveraging this enrichment, we identified 19 novel ADHD risk loci and 40 novel BD risk loci at condFDR <0.05. Further, we identified five loci jointly associated with ADHD and BD (conjFDR < 0.05). Interestingly, these five loci show concordant directions of effect for ADHD and BD. These results highlight a shared underlying genetic risk for ADHD and BD which may help to explain the high comorbidity rates and difficulties in differentiating between ADHD and BD in the clinic. Improving our understanding of the underlying genetic architecture of these disorders may aid in the development of novel stratification tools to help reduce these diagnostic difficulties.

Rapid fingerprinting of a highly glycosylated fusion protein by microfluidic chip-based capillary electrophoresis-mass spectrometry.

Protein glycosylation can impact the efficacy, safety, and pharmacokinetics of therapeutic proteins. Achieving uniform and consistent protein glycosylation is an important requirement for product quality control at all stages of therapeutic protein drug discovery and development. The development of a new microfluidic CE device compatible with MS offers a fast and sensitive orthogonal mode of high-resolution separation with MS characterization. Here, we describe a fast and robust chip-based CE-MS method for intact glycosylation fingerprinting of a therapeutic fusion protein with complex sialylated N and O-linked glycoforms. The method effectively separates multiple sialylated glycoforms and offers a rapid detection of changes in glycosylation profile in 6 min.

[Diagnosis, rehabilitation and development of disability 2000-2019 in Iceland].

INTRODUCTION: The disability assessment standard based on medically recognized illnesses or disabilities was introduced in Iceland 1999. The aim of this study is to examine the development of Social Insurance Administration (Tryggingastofnun ríkisins, TR) rulings regarding rehabilitation and disability pensions over a twenty-year period, since its introduction. MATERIAL AND METHODS: All registered diagnoses in the medical certificates of TR due to the approved rehabilitation or disability pension were examined in the period 2000-2019. The gender distribution and age distribution of these applicants and the number development during the period are described. At the same time, costs as a percentage of government expenditure are examined. RESULTS: The number of younger rehabilitation pensioners has increased rapidly in recent years, at the same time as the relative increase in disability pensioners has slowed slightly. Mental and musculoskeletal disorders are by far the most common types of illness leading to disability. Mental illnesses differ in terms of age distribution and increase over time. The proportion of individuals aged 18-66 with a 75% disability assessment has increased by a third during the period, from about 6% to 8%. The gender distribution of disability pensioners remains similar, with women accounting for 62% in total. Women are much more likely to receive disability pension due to musculoskeletal disorders than men and men are somewhat more likely to suffer from mental illness. The relative development of central government expenditure on total payments to rehabilitation and pensioners continues to grow as a proportion of central government expenditure. CONCLUSION: The number of rehabilitation pensioners has increased significantly since 2018, at the same time as the number of disability pensioners has decreased and there are indications that rehabilitation results in a lower number of new disability pensioners. Mental and musculoskeletal disorders are by far the most common types of illness leading to disability. A slightly lower proportion of disabled people have psychiatric diagnosis as a first diagnosis in the period 2000-2019 compared to those with a valid disability assessment in 2005, but the proportion of musculoskeletal disorders is slightly higher. Nevertheless, mental illnesses differ in age distribution and increase over time.

Integrated Approach for Characterizing Bispecific Antibody/Antigens Complexes and Mapping Binding Epitopes with SEC/MALS, Native Mass Spectrometry, and Protein Footprinting.

Bispecific antibodies (BsAbs), with a unique mechanism of recognizing two different epitopes or antigens, have shown potential in various therapeutic areas. Molecular characterization of BsAbs' epitopes not only allows for detailed understanding of their mechanism of actions but also guides the design and selection of drug candidate molecules. In this study, we illustrate the practical utility of an integrated approach, including size exclusion chromatography with multiangle light scattering and native mass spectrometry (MS) for the biophysical characterization of complex formation of a BsAb with two target antigens, cluster of differentiation 3 (CD3) and B-cell maturation antigen (BCMA). MS-based protein footprinting strategies, including hydrogen/deuterium exchange MS, fast photochemical oxidation of proteins, and carboxyl group footprinting with glycine ethyl ester, were further applied to determine BsAb's binding epitopes. This combination approach provides molecular details on the binding mechanisms of BsAb to the two distinct antigens with rapid output and high resolution.

Enhancement of carotenoid biosynthesis in the green microalga Dunaliella salina with light-emitting diodes and adaptive laboratory evolution.

There is a particularly high interest to derive carotenoids such as ß-carotene and lutein from higher plants and algae for the global market. It is well known that ß-carotene can be overproduced in the green microalga Dunaliella salina in response to stressful light conditions. However, little is known about the effects of light quality on carotenoid metabolism, e.g., narrow spectrum red light. In this study, we present UPLC-UV-MS data from D. salina consistent with the pathway proposed for carotenoid metabolism in the green microalga Chlamydomonas reinhardtii. We have studied the effect of red light-emitting diode (LED) lighting on growth rate and biomass yield and identified the optimal photon flux for D. salina growth. We found that the major carotenoids changed in parallel to the chlorophyll b content and that red light photon stress alone at high level was not capable of upregulating carotenoid accumulation presumably due to serious photodamage. We have found that combining red LED (75 %) with blue LED (25 %) allowed growth at a higher total photon flux. Additional blue light instead of red light led to increased ß-carotene and lutein accumulation, and the application of long-term iterative stress (adaptive laboratory evolution) yielded strains of D. salina with increased accumulation of carotenoids under combined blue and red light.

Psychiatric disorders in an urban sample of preschool children.

BACKGROUND: To increase limited epidemiological knowledge of early childhood psychopathology, a study of prevalence estimates and demographic correlates of psychiatric disorders was conducted in a sample of preschool children. METHODS: In a two-stage study, parents of 339 children aged 4-6 years who came for a medical check-up at three primary care centres in Reykjavik were invited to participate. First, the participants were screened with Brigance Screens and the Strengths and Difficulties Questionnaire (SDQ) for parents and teachers. Subsequently, the children were tested with a short version of Wechsler Preschool and Primary Scales of Intelligence - Revised and their parents were interviewed with the Schedule for Affective Disorders and Schizophrenia for School Aged Children Present and Lifetime Version. Weighted prevalence estimates were calculated and logistic regression was used to analyse the association between risk factors and psychiatric disorders. RESULTS: Of those invited to participate, 317 (93.5%) were included in the screening and of those, 131 received a full diagnostic assessment. The final study sample included 151 girls (47.6%) and 166 boys (52.4%) who represented 11.6% of the total birth cohort in Reykjavik. Weighted prevalence of DSM-IV psychiatric disorders was 10.1% (95% CI 6.7-13.5%) and 57/317 or 18.0% (95% CI 13.8-22.2%), including elimination disorders. Anxiety disorders (5.7%) and attention deficit hyperactivity disorder (3.8%) were the most common disorders in this preschool sample. Poor physical health of parents and higher education was associated with DSM-IV psychiatric disorders of the children. SDQ Total Difficulties score was associated with male gender and poor physical health of parents. CONCLUSIONS: This study indicates that psychiatric disorders in preschool children are common and may be correlated with parental health factors.

Genome-wide analyses of ADHD identify 27 risk loci, refine the genetic architecture and implicate several cognitive domains.

Attention-deficit hyperactivity disorder (ADHD) is a prevalent neurodevelopmental disorder with a major genetic component. Here, we present a genome-wide association study meta-analysis of ADHD comprising 38,691 individuals with ADHD and 186,843 controls. We identified 27 genome-wide significant loci, highlighting 76 potential risk genes enriched among genes expressed particularly in early brain development. Overall, ADHD genetic risk was associated with several brain-specific neuronal subtypes and midbrain dopaminergic neurons. In exome-sequencing data from 17,896 individuals, we identified an increased load of rare protein-truncating variants in ADHD for a set of risk genes enriched with probable causal common variants, potentially implicating SORCS3 in ADHD by both common and rare variants. Bivariate Gaussian mixture modeling estimated that 84-98% of ADHD-influencing variants are shared with other psychiatric disorders. In addition, common-variant ADHD risk was associated with impaired complex cognition such as verbal reasoning and a range of executive functions, including attention.

Improved therapeutic index of an acidic pH-selective antibody.

Although therapeutically efficacious, ipilimumab can exhibit dose-limiting toxicity that prevents maximal efficacious clinical outcomes and can lead to discontinuation of treatment. We hypothesized that an acidic pH-selective ipilimumab (pH Ipi), which preferentially and reversibly targets the acidic tumor microenvironment over the neutral periphery, may have a more favorable therapeutic index. While ipilimumab has pH-independent CTLA-4 affinity, pH Ipi variants have been engineered to have up to 50-fold enhanced affinity to CTLA-4 at pH 6.0 compared to pH 7.4. In hCTLA-4 knock-in mice, these variants have maintained anti-tumor activity and reduced peripheral activation, a surrogate marker for toxicity. pH-sensitive therapeutic antibodies may be a differentiating paradigm and a novel modality for enhanced tumor targeting and improved safety profiles.

Rare chromosomal deletions and duplications in attention-deficit hyperactivity disorder: a genome-wide analysis.

BACKGROUND: Large, rare chromosomal deletions and duplications known as copy number variants (CNVs) have been implicated in neurodevelopmental disorders similar to attention-deficit hyperactivity disorder (ADHD). We aimed to establish whether burden of CNVs was increased in ADHD, and to investigate whether identified CNVs were enriched for loci previously identified in autism and schizophrenia. METHODS: We undertook a genome-wide analysis of CNVs in 410 children with ADHD and 1156 unrelated ethnically matched controls from the 1958 British Birth Cohort. Children of white UK origin, aged 5-17 years, who met diagnostic criteria for ADHD or hyperkinetic disorder, but not schizophrenia and autism, were recruited from community child psychiatry and paediatric outpatient clinics. Single nucleotide polymorphisms (SNPs) were genotyped in the ADHD and control groups with two arrays; CNV analysis was limited to SNPs common to both arrays and included only samples with high-quality data. CNVs in the ADHD group were validated with comparative genomic hybridisation. We assessed the genome-wide burden of large (>500 kb), rare (<1% population frequency) CNVs according to the average number of CNVs per sample, with significance assessed via permutation. Locus-specific tests of association were undertaken for test regions defined for all identified CNVs and for 20 loci implicated in autism or schizophrenia. Findings were replicated in 825 Icelandic patients with ADHD and 35,243 Icelandic controls. FINDINGS: Data for full analyses were available for 366 children with ADHD and 1047 controls. 57 large, rare CNVs were identified in children with ADHD and 78 in controls, showing a significantly increased rate of CNVs in ADHD (0·156 vs 0·075; p=8·9×10(-5)). This increased rate of CNVs was particularly high in those with intellectual disability (0·424; p=2·0×10(-6)), although there was also a significant excess in cases with no such disability (0·125, p=0·0077). An excess of chromosome 16p13.11 duplications was noted in the ADHD group (p=0·0008 after correction for multiple testing), a finding that was replicated in the Icelandic sample (p=0·031). CNVs identified in our ADHD cohort were significantly enriched for loci previously reported in both autism (p=0·0095) and schizophrenia (p=0·010). INTERPRETATION: Our findings provide genetic evidence of an increased rate of large CNVs in individuals with ADHD and suggest that ADHD is not purely a social construct. FUNDING: Action Research; Baily Thomas Charitable Trust; Wellcome Trust; UK Medical Research Council; European Union.

Higher-Order Structure Characterization of NKG2A/CD94 Protein Complex and Anti-NKG2A Antibody Binding Epitopes by Mass Spectrometry-Based Protein Footprinting Strategies.

NK group 2 member A (NKG2A), an immune checkpoint inhibitor, is an emerging therapeutic target in immuno-oncology. NKG2A forms a heterodimer with CD94 on the cell surface of NK and a subset of T cells and recognizes the nonclassical human leukocyte antigen (HLA-E) in humans. Therapeutic blocking antibodies that block the ligation between HLA-E and NKG2A/CD94 have been shown to enhance antitumor immunity in mice and humans. In this study, we illustrate the practical utilities of mass spectrometry (MS)-based protein footprinting in areas from reagent characterization to antibody epitope mapping. Hydrogen/deuterium exchange mass spectrometry (HDX-MS) in the higher-order structure characterization of NKG2A in complex with CD94 provides novel insights into the conformational dynamics of NKG2A/CD94 heterodimer. To fully understand antibody/target interactions, we employed complementary protein footprinting methods, including HDX-MS and fast photochemical oxidation of proteins (FPOP)-MS, to determine the binding epitopes of therapeutic monoclonal antibodies targeting NKG2A. Such a combination approach provides molecular insights into the binding mechanisms of antibodies to NKG2A with high specificity, demonstrating the blockade of NKG2A/HLA-E interaction.

Identification of Genetic Loci Shared Between Attention-Deficit/Hyperactivity Disorder, Intelligence, and Educational Attainment.

BACKGROUND: Attention-deficit/hyperactivity disorder (ADHD) is a neurodevelopmental disorder that is consistently associated with lower levels of educational attainment. A recent large genome-wide association study identified common gene variants associated with ADHD, but most of the genetic architecture remains unknown. METHODS: We analyzed independent genome-wide association study summary statistics for ADHD (19,099 cases and 34,194 controls), educational attainment (N = 842,499), and general intelligence (N = 269,867) using a conditional/conjunctional false discovery rate (FDR) statistical framework that increases power of discovery by conditioning the FDR on overlapping associations. The genetic variants identified were characterized in terms of function, expression, and biological processes. RESULTS: We identified 58 linkage disequilibrium-independent ADHD-associated loci (conditional FDR < 0.01), of which 30 were shared between ADHD and educational attainment or general intelligence (conjunctional FDR < 0.01) and 46 were novel risk loci for ADHD. CONCLUSIONS: These results expand on previous genetic and epidemiological studies and support the hypothesis of a shared genetic basis between these phenotypes. Although the clinical utility of the identified loci remains to be determined, they can be used as resources to guide future studies aiming to disentangle the complex etiologies of ADHD, educational attainment, and general intelligence.

Attention-deficit hyperactivity disorder shares copy number variant risk with schizophrenia and autism spectrum disorder.

Attention-deficit/hyperactivity disorder (ADHD) is a highly heritable common childhood-onset neurodevelopmental disorder. Some rare copy number variations (CNVs) affect multiple neurodevelopmental disorders such as intellectual disability, autism spectrum disorders (ASD), schizophrenia and ADHD. The aim of this study is to determine to what extent ADHD shares high risk CNV alleles with schizophrenia and ASD. We compiled 19 neuropsychiatric CNVs and test 14, with sufficient power, for association with ADHD in Icelandic and Norwegian samples. Eight associate with ADHD; deletions at 2p16.3 (NRXN1), 15q11.2, 15q13.3 (BP4 & BP4.5-BP5) and 22q11.21, and duplications at 1q21.1 distal, 16p11.2 proximal, 16p13.11 and 22q11.21. Six of the CNVs have not been associated with ADHD before. As a group, the 19 CNVs associate with ADHD (OR = 2.43, P = 1.6 × 10-21), even when comorbid ASD and schizophrenia are excluded from the sample. These results highlight the pleiotropic effect of the neuropsychiatric CNVs and add evidence for ADHD, ASD and schizophrenia being related neurodevelopmental disorders rather than distinct entities.

Discovery of the first genome-wide significant risk loci for attention deficit/hyperactivity disorder.

Attention deficit/hyperactivity disorder (ADHD) is a highly heritable childhood behavioral disorder affecting 5% of children and 2.5% of adults. Common genetic variants contribute substantially to ADHD susceptibility, but no variants have been robustly associated with ADHD. We report a genome-wide association meta-analysis of 20,183 individuals diagnosed with ADHD and 35,191 controls that identifies variants surpassing genome-wide significance in 12 independent loci, finding important new information about the underlying biology of ADHD. Associations are enriched in evolutionarily constrained genomic regions and loss-of-function intolerant genes and around brain-expressed regulatory marks. Analyses of three replication studies: a cohort of individuals diagnosed with ADHD, a self-reported ADHD sample and a meta-analysis of quantitative measures of ADHD symptoms in the population, support these findings while highlighting study-specific differences on genetic overlap with educational attainment. Strong concordance with GWAS of quantitative population measures of ADHD symptoms supports that clinical diagnosis of ADHD is an extreme expression of continuous heritable traits.

Moisture-Induced Amorphous Phase Separation of Amorphous Solid Dispersions: Molecular Mechanism, Microstructure, and Its Impact on Dissolution Performance.

Amorphous phase separation (APS) is commonly observed in amorphous solid dispersions (ASD) when exposed to moisture. The objective of this study was to investigate: (1) the phase behavior of amorphous solid dispersions composed of a poorly water-soluble drug with extremely low crystallization propensity, BMS-817399, and PVP, following exposure to different relative humidity (RH), and (2) the impact of phase separation on the intrinsic dissolution rate of amorphous solid dispersion. Drug-polymer interaction was confirmed in ASDs at different drug loading using infrared (IR) spectroscopy and water vapor sorption analysis. It was found that the drug-polymer interaction could persist at low RH (≤75% RH) but was disrupted after exposure to high RH, with the advent of phase separation. Surface morphology and composition of 40/60 ASD at micro-/nano-scale before and after exposure to 95% RH were also compared. It was found that hydrophobic drug enriched on the surface of ASD after APS. However, for the 40/60 ASD system, the intrinsic dissolution rate of amorphous drug was hardly affected by the phase behavior of ASD, which may be partially attributed to the low crystallization tendency of amorphous BMS-817399 and enriched drug amount on the surface of ASD. Intrinsic dissolution rate of PVP decreased resulting from APS, leading to a lower concentration in the dissolution medium, but supersaturation maintenance was not anticipated to be altered after phase separation due to the limited ability of PVP to inhibit drug precipitation and prolong the supersaturation of drug in solution. This study indicated that for compounds with low crystallization propensity and high hydrophobicity, the risk of moisture-induced APS is high but such phase separation may not have profound impact on the drug dissolution performance of ASDs. Therefore, application of ASD technology on slow crystallizers could incur low risks not only in physical stability but also in dissolution performance.

Oral Delivery of Highly Lipophilic, Poorly Water-Soluble Drugs: Self-Emulsifying Drug Delivery Systems to Improve Oral Absorption and Enable High-Dose Toxicology Studies of a Cholesteryl Ester Transfer Protein Inhibitor in Preclinical Species.

BMS-A is an inhibitor of cholesteryl ester transfer protein and is a highly lipophilic compound (clogP 10.5) with poor aqueous solubility (<0.0001 mg/mL at pH 6.5). The compound exhibits low oral exposure when dosed as cosolvent solution formulations. The purpose of this study was to evaluate lipid-based formulations for enabling high-dose toxicology studies and enhancing toxicology margins of BMS-A in preclinical studies in nonrodent species. The solubility of BMS-A was screened in lipid and cosolvent/surfactant excipients, and prototype formulations were developed. In vitro tests showed that fine/microemulsions were formed after aqueous dilution of lipid formulations, and BMS-A was transferred from oil phase to aqueous phase with enhanced solubility following lipid digestion. When dosed in dogs at 200 mg/kg, a Gelucire-based formulation exhibited more than 10-fold higher exposure compared to the solution formulation and was thus selected for toxicology studies in dogs. For monkeys, an olive oil formulation was developed, and the exposure was about 7-fold higher than that from the solution. In summary, lipid-based drug delivery could be applied in early stages of drug discovery to enhance oral exposure and enable preclinical toxicology studies of highly lipophilic compounds, while facilitating the candidate selection of a molecule which is more specifically designed for bioperformance in a lipid-based drug delivery strategy.

Lipophilic salts of poorly soluble compounds to enable high-dose lipidic SEDDS formulations in drug discovery.

Self-emulsifying drug delivery systems (SEDDS) have been used to solubilize poorly water-soluble drugs to improve exposure in high-dose pharmacokinetic (PK) and toxicokinetic (TK) studies. However, the absorbable dose is often limited by drug solubility in the lipidic SEDDS vehicle. This study focuses on increasing solubility and drug loading of ionizable drugs in SEDDS vehicles using lipophilic counterions to prepare lipophilic salts of drugs. SEDDS formulations of two lipophilic salts-atazanavir-2-naphthalene sulfonic acid (ATV-2-NSA) and atazanavir-dioctyl sulfosuccinic acid (ATV-Doc)-were characterized and their performance compared to atazanavir (ATV) free base formulated as an aqueous crystalline suspension, an organic solution, and a SEDDS suspension, using in vitro, in vivo, and in silico methods. ATV-2-NSA exhibited ∼6-fold increased solubility in a SEDDS vehicle, allowing emulsion dosing at 12mg/mL. In rat PK studies at 60mg/kg, the ATV-2-NSA SEDDS emulsion had comparable exposure to the free-base solution, but with less variability, and had better exposure at high dose than aqueous suspensions of ATV free base. Trends in dose-dependent exposure for various formulations were consistent with GastroPlus™ modeling. Results suggest use of lipophilic salts is a valuable approach for delivering poorly soluble compounds at high doses in Discovery.

Reproductive fitness and genetic risk of psychiatric disorders in the general population.

The persistence of common, heritable psychiatric disorders that reduce reproductive fitness is an evolutionary paradox. Here, we investigate the selection pressures on sequence variants that predispose to schizophrenia, autism, bipolar disorder, major depression and attention deficit hyperactivity disorder (ADHD) using genomic data from 150,656 Icelanders, excluding those diagnosed with these psychiatric diseases. Polygenic risk of autism and ADHD is associated with number of children. Higher polygenic risk of autism is associated with fewer children and older age at first child whereas higher polygenic risk of ADHD is associated with having more children. We find no evidence for a selective advantage of a high polygenic risk of schizophrenia or bipolar disorder. Rare copy-number variants conferring moderate to high risk of psychiatric illness are associated with having fewer children and are under stronger negative selection pressure than common sequence variants.

Discovery of a Parenteral Small Molecule Coagulation Factor XIa Inhibitor Clinical Candidate (BMS-962212).

Factor XIa (FXIa) is a blood coagulation enzyme that is involved in the amplification of thrombin generation. Mounting evidence suggests that direct inhibition of FXIa can block pathologic thrombus formation while preserving normal hemostasis. Preclinical studies using a variety of approaches to reduce FXIa activity, including direct inhibitors of FXIa, have demonstrated good antithrombotic efficacy without increasing bleeding. On the basis of this potential, we targeted our efforts at identifying potent inhibitors of FXIa with a focus on discovering an acute antithrombotic agent for use in a hospital setting. Herein we describe the discovery of a potent FXIa clinical candidate, 55 (FXIa Ki = 0.7 nM), with excellent preclinical efficacy in thrombosis models and aqueous solubility suitable for intravenous administration. BMS-962212 is a reversible, direct, and highly selective small molecule inhibitor of FXIa.

Validity of self-report and informant rating scales of adult ADHD symptoms in comparison with a semistructured diagnostic interview.

In a study of ADHD symptoms in the relatives of probands diagnosed with ADHD, the validity of self-reported and informant-reported symptoms in childhood and adulthood was investigated with a semistructured diagnostic interview, the Schedule for Affective Disorders and Schizophrenia for School-Age Children (K-SADS) adapted for adults, as a criterion. The participating relatives were 80 women and 46 men aged 17 to 77. Rating scales based on the Diagnostic and Statistical Manual of Mental Disorders (4th ed.) were completed by participants and informants. Internal consistency of the scales and interrater reliabilities of the diagnostic interview were satisfactory. Correlations between ratings across sources of information supported convergent and divergent validity. Self-report scales and informant scales predicted interview-based diagnoses in childhood and adulthood with adequate sensitivities and specificities. It was concluded that the rating scales have good psychometric properties, at least in at-risk populations.