Roctavian Gene Therapy for Hemophilia A.

Following successful efforts in adeno-associated virus (AAV) gene addition for hemophilia B gene therapy, the development of valoctocogene roxaparvovec (Roctavian; Biomarin) over the past decade represents a potential new hemophilia A (HA) treatment paradigm. Roctavian is the first licensed HA gene therapy and was conditionally approved in Europe in August of 2022 and approved in the U.S. in June of 2023. Beyond Roctavian, there are ongoing pivotal trials of additional AAV vectors for HA and others that are progressing through pre-clinical development or early-phase clinical trial as well as non-AAV approaches also in clinical development. This review focuses on the clinical development of Roctavian for which the collective clinical trials represent the largest body of work thus far available for any licensed AAV product. From this pioneering clinical development, several outstanding questions have emerged for which the answers will undoubtedly be important to the clinical adaptation of Roctavian and future efforts in HA gene therapy. Most notably, unexplained year-over-year declines in factor VIII (FVIII) expression after Roctavian treatment contrast with observed stable FVIII expression in AAV HA gene therapy clinical trials with more modest initial FVIII expression. This observation has been qualitatively replicated in animal models. The development and approval of Roctavian is a landmark in HA therapeutics, though next-generation approaches are needed before HA gene therapy fulfills its promise of stable FVIII expression that normalizes hemostasis.

LarvaTagger: Manual and automatic tagging of drosophila larval behaviour.

MOTIVATION: As more behavioural assays are carried out in large-scale experiments on Drosophila larvae, the definitions of the archetypal actions of a larva are regularly refined. In addition, video recording and tracking technologies constantly evolve. Consequently, automatic tagging tools for Drosophila larval behaviour must be retrained to learn new representations from new data. However, existing tools cannot transfer knowledge from large amounts of previously accumulated data.We introduce LarvaTagger, a piece of software that combines a pre-trained deep neural network, providing a continuous latent representation of larva actions for stereotypical behaviour identification, with a graphical user interface to manually tag the behaviour and train new automatic taggers with the updated ground truth. RESULTS: We reproduced results from an automatic tagger with high accuracy, and we demonstrated that pre-training on large databases accelerates the training of a new tagger, achieving similar prediction accuracy using less data. AVAILABILITY: All the code is free and open source. Docker images are also available. See gitlab.pasteur.fr/nyx/LarvaTagger.jl. SUPPLEMENTARY INFORMATION: Supplementary material is available at Bioinformatics online.

Enhanced Procoagulant Activity of Select Hemophilia B Causing Factor IX Variants with Emicizumab.

Emicizumab improves the procoagulant activity of select loss-of-function FIX variants with likely dysfunctional assembly of the intrinsic Xase complex that cause hemophilia B. FVIII-mimetics may represent an alternative non-factor therapy for select hemophilia B patients.

Chimpanzees (Pan troglodytes) recognize that their guesses could be wrong and can pass a two-cup disjunctive syllogism task.

When chimpanzees search for hidden food, do they realize that their guesses may not be correct? We applied a post-decision wagering paradigm to a simple two-cup search task, varying whether we gave participants visual access to the baiting and then asking after they had chosen one of the cups whether they would prefer a smaller but certain reward instead of their original choice (experiment 1). Results showed that chimpanzees were more likely to accept the smaller reward in occluded than visible conditions. Experiment 2 found the same effect when we blocked visual access but manipulated the number of hiding locations for the food piece, showing that the effect is not owing to representation type. Experiments 3 and 4 showed that when given information about the contents of the unchosen cup, chimpanzees were able to flexibly update their choice behaviour accordingly. These results suggest that language is not a pre-requisite to solving the disjunctive syllogism and provides a valuable contribution to the debate on logical reasoning in non-human animals.

Challenges in clinical diagnosis of Clade I Mpox: Highlighting the need for enhanced diagnostic approaches.

BACKGROUND: Due to limited diagnostic capacity and availability of point-of-care tests, diagnosis of Clade I mpox in the geographical regions most affected is usually on clinical grounds. This may be complicated due to the similarity between mpox and varicella (chickenpox) lesions. Visual assessment of lesions is also used for determining clinical progress and to assess patient outcomes in clinical trials. However, there has been no investigation into whether clinicians can (i) identify Clade I mpox compared to other viral lesions (ii) differentiate between Clade I mpox lesion stages. METHODOLOGY/PRINCIPLE FINDINGS: The objective of this study was to evaluate inter-rater reliability and agreement between clinicians assessing lesions in patients with Clade I mpox. We presented experienced clinicians with 17 images of Clade I mpox or varicella and asked them to independently indicate the most likely diagnosis-mpox or varicella-and to categorise the lesions according to their stage. When selecting the most likely diagnosis, accuracy varied across all images, the inter-rater reliability was poor (κ = 0.223; z = 10.1) and agreement was moderate (Po = 68%). When categorising lesions according to their type, if a single lesion type was present in the image, inter-rater reliability was moderate (κ = 0.671, z = 40.6) and agreement was good (Po = 78%), but when multiple lesion types were shown in an image, both inter-rater reliability (κ = 0.153, z = 10.5) and agreement (Po = 29%) decreased substantially. CONCLUSIONS: This study demonstrates that there are presently limitations in using visual assessment to diagnose Clade I mpox and evaluate lesion stage and treatment outcomes, which have an impact on clinical practice, public health and clinical trials. More robust indicators and tools are required to inform clinical, public-health, and research priorities, but these must be implementable in countries affected by mpox.

Biochemical and structural insights into a 5' to 3' RNA ligase reveal a potential role in tRNA ligation.

ATP-grasp superfamily enzymes contain a hand-like ATP-binding fold and catalyze a variety of reactions using a similar catalytic mechanism. More than 30 protein families are categorized in this superfamily, and they are involved in a plethora of cellular processes and human diseases. Here we identify C12orf29 as an atypical ATP-grasp enzyme that ligates RNA. Human C12orf29 and its homologs auto-adenylate on an active site Lys residue as part of a reaction intermediate that specifically ligates RNA halves containing a 5'-phosphate and a 3'-hydroxyl. C12orf29 binds tRNA in cells and can ligate tRNA within the anticodon loop in vitro. Genetic depletion of c12orf29 in female mice alters global tRNA levels in brain. Furthermore, crystal structures of a C12orf29 homolog from Yasminevirus bound to nucleotides reveal a minimal and atypical RNA ligase fold with a unique active site architecture that participates in catalysis. Collectively, our results identify C12orf29 as an RNA ligase and suggest its involvement in tRNA biology.

Post-fire stabilization of thaw-affected permafrost terrain in northern Alaska.

In 2007, the Anaktuvuk River fire burned more than 1000 km2 of arctic tundra in northern Alaska, ~ 50% of which occurred in an area with ice-rich syngenetic permafrost (Yedoma). By 2014, widespread degradation of ice wedges was apparent in the Yedoma region. In a 50 km2 area, thaw subsidence was detected across 15% of the land area in repeat airborne LiDAR data acquired in 2009 and 2014. Updating observations with a 2021 airborne LiDAR dataset show that additional thaw subsidence was detected in < 1% of the study area, indicating stabilization of the thaw-affected permafrost terrain. Ground temperature measurements between 2010 and 2015 indicated that the number of near-surface soil thawing-degree-days at the burn site were 3 × greater than at an unburned control site, but by 2022 the number was reduced to 1.3 × greater. Mean annual ground temperature of the near-surface permafrost increased by 0.33 °C/yr in the burn site up to 7-years post-fire, but then cooled by 0.15 °C/yr in the subsequent eight years, while temperatures at the control site remained relatively stable. Permafrost cores collected from ice-wedge troughs (n = 41) and polygon centers (n = 8) revealed the presence of a thaw unconformity, that in most cases was overlain by a recovered permafrost layer that averaged 14.2 cm and 18.3 cm, respectively. Taken together, our observations highlight that the initial degradation of ice-rich permafrost following the Anaktuvuk River tundra fire has been followed by a period of thaw cessation, permafrost aggradation, and terrain stabilization.

Withdrawal during outpatient low dose buprenorphine initiation in people who use fentanyl: a retrospective cohort study.

BACKGROUND: Buprenorphine is an effective treatment for opioid use disorder (OUD); however, buprenorphine initiation can be complicated by withdrawal symptoms including precipitated withdrawal. There has been increasing interest in using low dose initiation (LDI) strategies to reduce this withdrawal risk. As there are limited data on withdrawal symptoms during LDI, we characterize withdrawal symptoms in people with daily fentanyl use who underwent initiation using these strategies as outpatients. METHODS: We conducted a retrospective chart review of patients with OUD using daily fentanyl who were prescribed 7-day or 4-day LDI at 2 substance use disorder treatment clinics in San Francisco. Two addiction medicine experts assessed extracted chart documentation for withdrawal severity and precipitated withdrawal, defined as acute worsening of withdrawal symptoms immediately after taking buprenorphine. A third expert adjudicated disagreements. Data were analyzed using descriptive statistics. RESULTS: There were 175 initiations in 126 patients. The mean age was 37 (SD 10 years). 71% were men, 26% women, and 2% non-binary. 21% identified as Black, 16% Latine, and 52% white. 60% were unstably housed and 75% had Medicaid insurance. Substance co-use included 74% who used amphetamines, 29% cocaine, 22% benzodiazepines, and 19% alcohol. Follow up was available for 118 (67%) initiations. There was deviation from protocol instructions in 22% of these initiations with follow up. 31% had any withdrawal, including 21% with mild symptoms, 8% moderate and 2% severe. Precipitated withdrawal occurred in 10 cases, or 8% of initiations with follow up. Of these, 7 had deviation from protocol instructions; thus, there were 3 cases with follow up (3%) in which precipitated withdrawal occurred without protocol deviation. CONCLUSIONS: Withdrawal was relatively common in our cohort but was mostly mild, and precipitated withdrawal was rare. Deviation from instructions, structural barriers, and varying fentanyl use characteristics may contribute to withdrawal. Clinicians should counsel patients who use fentanyl that mild withdrawal symptoms are likely during LDI, and there is still a low risk for precipitated withdrawal. Future studies should compare withdrawal across initiation types, seek ways to support patients in initiating buprenorphine, and qualitatively elicit patients' withdrawal experiences.

Current and projected flood exposure for Alaska coastal communities.

Globally, coastal communities experience flood hazards that are projected to worsen from climate change and sea level rise. The 100-year floodplain or record flood are commonly used to identify risk areas for planning purposes. Remote communities often lack measured flood elevations and require innovative approaches to estimate flood elevations. This study employs observation-based methods to estimate the record flood elevation in Alaska communities and compares results to elevation models, infrastructure locations, and sea level rise projections. In 46 analyzed communities, 22% of structures are located within the record floodplain. With sea level rise projections, this estimate increases to 30-37% of structures by 2100 if structures remain in the same location. Flood exposure is highest in western Alaska. Sea level rise projections suggest northern Alaska will see similar flood exposure levels by 2100 as currently experienced in western Alaska. This evaluation of record flood height, category, and history can be incorporated into hazard planning documents, providing more context for coastal flood exposure than previously existed for Alaska. This basic flood exposure method is transferable to other areas with similar mapping challenges. Identifying current and projected hazardous zones is essential to avoid unintentional development in floodplains and improve long-term safety.

Successful preimplantation genetic testing for fibrodysplasia ossificans progressiva: a case report.

PURPOSE OF THE STUDY: Fibrodysplasia ossificans progressiva (FOP) is a rare autosomal dominant condition that leads to significant disability and morbidity, characterised by the formation of heterotopic hard tissues within connective tissues. The condition has an incidence of approximately one per two million people worldwide. There is no known single effective treatment available for FOP. We report the world's first case of a healthy infant born following in vitro fertilisation (IVF) and preimplantation genetic testing for monogenic disorder (PGT-M) using Karyomapping for FOP. CASE PRESENTATION: A 30-year-old Caucasian female with FOP presented with her partner seeking IVF with PGT-M to achieve a healthy pregnancy with an embryo unaffected by FOP. METHODS: The couple underwent IVF and PGT-M using Karyomapping as the testing method. A multi-disciplinary team approach was utilised in planning this case, considering the additional risks of oocyte retrieval, pregnancy and childbirth in women with FOP. MAIN FINDINGS: The oocyte retrieval was covered with a 5-day course of prednisolone to reduce the risk of a localised inflammatory reaction, which could result in subsequent heterotopic ossification. This was subsequently weaned down with reducing doses every two days. The patient underwent uncomplicated oocyte retrieval, yielding 12 mature oocytes. Following intracytoplasmic sperm injection (ICSI), ten zygotes having two pro-nuclei were cultured, and six underwent trophoectoderm biopsy and vitrification 5-6 days after retrieval. PGT-M via Karyomapping revealed four out of six (66.7%) of blastocysts were not carriers of the maternal high-risk FOP allele. In total, the patient had three separate embryo transfers. Pregnancy was achieved following the third frozen embryo transfer, which went to 37 weeks' gestation, and delivered by Caesarean section. The baby was born in excellent condition and is unaffected by FOP. CONCLUSION: IVF/ICSI and PGT-M using Karyomapping was successfully implemented to identify embryos carrying the high-risk FOP allele resulting in a healthy livebirth.

Addressing air quality challenges: Comparative analysis of Barcelona, Venezuela, and Guayaquil, Ecuador.

This study presents a willingness-to-pay (WtP) questionnaire that was designed, validated, and applied to assess perceptions of air quality and self-reported health in two middle-income South American cities: Barcelona and its neighboring cities (Venezuela) and Guayaquil (Ecuador). These cities lack air quality monitoring and control measures. The questionnaire is a reliable tool to assess air quality based on citizens' perceptions, and the results reveal that both populations perceive low air quality and accurately identify emission sources and air pollutants (industrial emissions and particulate matter in Barcelona and vehicular emissions and carbon monoxide in Guayaquil). The study also evaluated the efforts made by both cities to improve air quality using the United Nations Environment Programme to strengthen air quality in South America. Based on this evaluation, strengths were identified for enhancing air quality in both cities. The study finds that in Barcelona and its surroundings, investment is needed to improve urban transport, waste management, and update the environmental legislation regarding air quality at the national level. In contrast, Guayaquil has already taken some measures to improve air quality, but more investment in public transport and measures to lower vehicle emissions are needed.

Discovery of Pyridopyrimidinones that Selectively Inhibit the H1047R PI3Kα Mutant Protein.

The H1047R mutation of PIK3CA is highly prevalent in breast cancers and other solid tumors. Selectively targeting PI3KαH1047R over PI3KαWT is crucial due to the role that PI3KαWT plays in normal cellular processes, including glucose homeostasis. Currently, only one PI3KαH1047R-selective inhibitor has progressed into clinical trials, while three pan mutant (H1047R, H1047L, H1047Y, E542K, and E545K) selective PI3Kα inhibitors have also reached the clinical stage. Herein, we report the design and discovery of a series of pyridopyrimidinones that inhibit PI3KαH1047R with high selectivity over PI3KαWT, resulting in the discovery of compound 17. When dosed in the HCC1954 tumor model in mice, 17 provided tumor regressions and a clear pharmacodynamic response. X-ray cocrystal structures from several PI3Kα inhibitors were obtained, revealing three distinct binding modes within PI3KαH1047R including a previously reported cryptic pocket in the C-terminus of the kinase domain wherein we observe a ligand-induced interaction with Arg1047.

Menstrual cycles and the impact upon performance in elite British track and field athletes: a longitudinal study.

Objective: To assess the prevalence of menstrual disorders and the perceived effect of menstrual cycles upon performance in elite athletes. Methodology: A longitudinal survey in the form of a questionnaire was sent to female track and field athletes at British Athletics every 6 months, over a five-year period between 1st October 2014 and 1st October 2019 in the United Kingdom (UK). Results: 128 athletes completed an average of 4.2 ± 2.9 questionnaires across the study period. The mean age of menarche was 14.2 ± 1.4 years, 13.4 ± 1.3 years and 12.8 ± 1.4 years in endurance, power, and thrower athletes respectively (p < 0.05). Two-thirds (66%; n = 82) reported consistently regular cycles, 30% (n = 37) irregular at some point during the period of observation and 4% (n = 5) were amenorrhoeic. 87 athletes (68%) reported dysmenorrhoea and 40 (31%) menorrhagia. More than three quarters (76.8%; n = 96) described their cycle negatively affected performance. Amongst those who reported when the negative impact occurred (n = 91), 40% (n = 36) reported this in the late luteal phase and 35% (n = 32) during the early follicular phase. 79% (n = 100) of athletes reported at least one cyclical symptom, of which bloating, lower back and pelvic pain were most frequently experienced. Conclusion: This data highlights the complex interrelationship between women's health and elite athletic performance. Athletes perceive a negative impact from their menstrual cycles upon performance with a desire to manage these more effectively, particularly during competition. Female reproductive health expertise in the multi-disciplinary management of elite athletes is required.

Lewis Acid-Driven Inverse Hydride Shuttle Catalysis.

Inverse hydride shuttle catalysis provides a multicomponent platform for the highly efficient synthesis of alkaloid frameworks with exquisite diastereoselectivity. However, a number of limitations hinder this method, primarily the strict requirement for highly electron-deficient acceptors. Herein, we present a general Lewis acid-driven approach to address this constraint, and have developed two broad strategies enabling the modular synthesis of complex azabicycles that were entirely unattainable using the previous method. The enhanced synthetic flexibility facilitates a streamlined asymmetric cyclization, leading to a concise total synthesis of the alkaloid (-)-tashiromine.

Systematic Review of Scales for Measuring Infectious Disease-Related Stigma.

Infectious disease outbreaks are associated with substantial stigma, which can have negative effects on affected persons and communities and on outbreak control. Thus, measuring stigma in a standardized and validated manner early in an outbreak is critical to disease control. We reviewed existing scales used to assess stigma during outbreaks. Our findings show that many different scales have been developed, but few have been used more than once, have been adequately validated, or have been tested in different disease and geographic contexts. We found that scales were usually developed too slowly to be informative early during an outbreak and were published a median of 2 years after the first case of an outbreak. A rigorously developed, transferable stigma scale is needed to assess and direct responses to stigma during infectious disease outbreaks.

AAV gene therapy in companion dogs with severe hemophilia: Real-world long-term data on immunogenicity, efficacy, and quality of life.

The hemophilias are the most common severe inherited bleeding disorders and are caused by deficiency of clotting factor (F) VIII (hemophilia A) or FIX (hemophilia B). The resultant bleeding predisposition significantly increases morbidity and mortality. The ability to improve the bleeding phenotype with modest increases in clotting factor levels has enabled the development and regulatory approval of adeno-associated viral (AAV) vector gene therapies for people with hemophilia A and B. The canine hemophilia model has proven to be one of the best predictors of therapeutic response in humans. Here, we report long-term follow-up of 12 companion dogs with severe hemophilia that were treated in a real-world setting with AAV gene therapy. Despite more baseline bleeding than in research dogs, companion dogs demonstrated a 94% decrease in bleeding rates and 61% improvement in quality of life over a median of 4.1 years (range 2.6-8.9). No new anti-transgene immune responses were detected; one dog with a pre-existing anti-FVIII inhibitor achieved immune tolerance with gene therapy. Two dogs expressing 1%-5% FVIII post gene therapy experienced fatal bleeding events. These data suggest AAV liver-directed gene therapy is efficacious in a real-world setting but should target expression >5% and closely monitor those with levels in the 1%-5% range.

Stereoselective Amine Synthesis Mediated by a Zirconocene Hydride to Accelerate a Drug Discovery Program.

Chiral amine synthesis remains a significant challenge in accelerating the design cycle of drug discovery programs. A zirconium hydride, due to its high oxophilicity and lower reactivity, gave highly chemo- and stereoselective reductions of sulfinyl ketimines. The development of this zirconocene-mediated reduction helped to accelerate our drug discovery efforts and is applicable to several motifs commonly used in medicinal chemistry. Computational investigation supported a cyclic half-chair transition state to rationalize the high selectivity in benzyl systems.

Molecular and Structural Basis for Cγ-C Bond Formation by PLP-Dependent Enzyme Fub7.

Pyridoxal 5'-phosphate (PLP)-dependent enzymes that catalyze γ-replacement reactions are prevalent, yet their utilization of carbon nucleophile substrates is rare. The recent discovery of two PLP-dependent enzymes, CndF and Fub7, has unveiled unique C-C bond forming capabilities, enabling the biocatalytic synthesis of alkyl- substituted pipecolic acids from O-acetyl-L-homoserine and ß-keto acid or aldehyde derived enolates. This breakthrough presents fresh avenues for the biosynthesis of pipecolic acid derivatives. However, the catalytic mechanisms of these enzymes remain elusive, and a dearth of structural information hampers their extensive application. Here, we have broadened the catalytic scope of Fub7 by employing ketone-derived enolates as carbon nucleophiles, revealing Fub7's capacity for substrate-dependent regioselective α-alkylation of unsymmetrical ketones. Through an integrated approach combining X-ray crystallography, spectroscopy, mutagenesis, and computational docking studies, we offer a detailed mechanistic insight into Fub7 catalysis. Our findings elucidate the structural basis for its substrate specificity, stereoselectivity, and regioselectivity. Our work sets the stage ready for subsequent protein engineering effort aimed at expanding the synthetic utility of Fub7, potentially unlocking novel methods to access a broader array of noncanonical amino acids.