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Abstract CRISPR-Cas12a systems are becoming an attractive genome editing tool for cell engineering due to their broader editing capabilities compared to CRISPR-Cas9 counterparts. As opposed to Cas9, the Cas12a endonucleases are characterized by a lack of trans-activating crRNA (tracrRNA), which reduces the complexity of the editing system and simultaneously makes CRISPR RNA (crRNA) engineering a promising approach toward further improving and modulating editing activity of the CRISPR-Cas12a systems. Here, we design and validate sixteen types of structurally engineered Cas12a crRNAs targeting various immunologically relevant loci in-vitro and in-cellulo. We show that all our structural modifications in the loop region, ranging from engineered breaks (STAR-crRNAs) to large gaps (Gap-crRNAs), as well as nucleotide substitutions, enable gene-cutting in the presence of various Cas12a nucleases. Moreover, we observe similar insertion rates of short HDR templates using the engineered crRNAs compared to the wild-type crRNAs, further demonstrating that the introduced modifications in the loop region led to comparable genome editing efficiencies. In conclusion, we show that Cas12a nucleases can broadly utilize structurally engineered crRNAs with breaks or gaps in the otherwise highly-conserved loop region, which could further facilitate a wide range of genome editing applications.

Introduction Children represent a large and vulnerable patient group. However, the evidence base for most paediatric diagnostic and therapeutic procedures remains limited or is often inferred from adults. There is an urgency to improve paediatric healthcare provision based on real-world evidence generation. Digital transformation is a unique opportunity to shape a data-driven, agile, learning healthcare system and deliver more efficient and personalised care to children and their families. The goal of Paediatric Personalized Research Network Switzerland (SwissPedHealth) is to build a sustainable and scalable infrastructure to make routine clinical data from paediatric hospitals in Switzerland interoperable, standardised, quality-controlled, and ready for observational research, quality assurance, trials and health-policy creation. This study describes the design, aims and current achievements of SwissPedHealth.Methods and analysis SwissPedHealth was started in September 2022 as one of four national data streams co-funded by the Swiss Personalized Health Network (SPHN) and the Personalized Health and Related Technologies (PHRT). SwissPedHealth develops modular governance and regulatory strategies and harnesses SPHN automatisation procedures in collaboration with clinical data warehouses, the Data Coordination Center, Biomedical Information Technology Network, and other SPHN institutions and funded projects. The SwissPedHealth consortium is led by a multisite, multidisciplinary Steering Committee, incorporating patient and family representatives. The data stream contains work packages focusing on (1) governance and implementation of standardised data collection, (2) nested projects to test the feasibility of the data stream, (3) a lighthouse project that enriches the data stream by integrating multi-omics data, aiming to improve diagnoses of rare diseases and 4) engagement with families through patient and public involvement activities and bioethics interviews.Ethics and dissemination The health database regulation of SwissPedHealth was approved by the ethics committee (AO_2022-00018). Research findings will be disseminated through national and international conferences and publications in peer-reviewed journals, and in lay language via online media and podcasts.

Abstract Background Law enforcement traffic stops are one of the most common entryways to the US justice system. Conventional frameworks suggest traffic stops promote public safety by reducing dangerous driving practices and non-vehicular crime with little to no collateral damage to individuals and communities. Critical frameworks interrogate these assumptions, identifying significant individual and community harms that disparately impact Black, Indigenous, and People of Color (BIPOC) and low-income communities. Methods The Public Health Critical Race Praxis (PHCRP) and multi-level frameworks from community anti-racist training were combined into a structured diagram to guide intervention and research teams in contrasting conventional and critical perspectives on traffic stops. The diagram divides law enforcement and drivers/residents as two separate agent types that interact during traffic stops. These two agent types have different conventional and critical histories, priorities, and perspectives at multiple levels, including individual, interpersonal, institutional, and cultural levels. Conventional solutions (identifying explicitly racist officers, “meet-a-cop” programs, police interaction training for drivers) are born from conventional frameworks (rewarding crime prevention regardless of cost, the war on drugs saves lives, driver behavior perfectionism). While conventional perspectives focus on individual and interpersonal levels, critical perspectives more deeply acknowledge dynamics at institutional and cultural levels. Critical solutions may be hard to discover without critical frameworks, including that law enforcement creates measurable collateral damage and disparate social control effects; neighborhood patrol priorities can be set without community self-determination or accountability and may trump individual and interpersonal dynamics; and the war on drugs is highly racialized and disproportionally enforced through traffic stop programs. Conclusions Traffic stop enforcement and crash prevention programs that do not deeply and critically consider these dynamics at multiple levels, not just law enforcement-driver interactions at the individual and interpersonal levels, may be at increased risk of propagating histories of BIPOC discrimination. In contrast, public health and transportation researchers and practitioners engaged in crash and injury prevention strategies that employ law enforcement should critically consider disparate history and impacts of law enforcement in BIPOC communities. PHCRP, anti-racism frameworks, and the included diagram may assist them in organizing critical thinking about research studies, interventions, and impacts.

Abstract Background Acute intoxication-type inborn errors of metabolism (IT-IEM) such as urea cycle disorders and non-acute IT-IEM such as phenylketonuria (PKU) and their treatment have a major impact on the life of affected children and families. Yet patients’ and parents’ perspectives on the burdens of IT-IEM and its effects on everyday functioning and well-being have rarely been addressed. Patient- and observer-reported outcomes (PROs/ObsROs) are critically important to evaluate and target health care and treatment efficacy. Therefore, it is mandatory to define PROs/ObsROs relevant to patients with IT-IEM, their families, and health care professionals and to provide valid, standardised and reliable measuring instruments. To achieve consensus we performed a two-round, electronic-based modification of a Delphi survey including 27 parents of affected children, nine teenage patients and 35 health professionals (physicians, nutritionists, psychologists). The final set of PROs/ObsROs was discussed and defined in an online consensus meeting with a subsample of three health professionals, three parents and two patients. For this final set, appropriate measures (PROMs/ObsROMs) were assembled. Results Seventeen PROs/ObsROs constitute the final core set for paediatric IT-IEM. They cover social (e.g. social participation), emotional (e.g. positive affect), and disease-related aspects (e.g. attitude towards treatment) of patients’ lives as well as the experience of parents (e.g. parental stress). Conclusion To promote a holistic treatment approach, this consensus-driven set of relevant PROs/ObsROs should be incorporated into daily IT-IEM care and considered as the key psychological outcomes in clinical trials. We have identified existing—psychometrically and contextual—appropriate PROMs/ObsROMs with open access to facilitate this process.

Abstract Background The number of market approvals of orphan medicinal products (OMPs) has been increasing steadily in the last 3 decades. While OMPs can offer a unique chance for patients suffering from rare diseases, they are usually very expensive. The growing number of approved OMPs increases their budget impact despite their low prevalence, making it pressing to find solutions to ethical challenges on how to fairly allocate scarce healthcare resources under this context. One potential solution could be to grant OMPs special status when considering them for reimbursement, meaning that they are subject to different, and less stringent criteria than other drugs. This study aims to provide a systematic analysis of moral reasons for and against such a special status for the reimbursement of OMPs in publicly funded healthcare systems from a multidisciplinary perspective. Results With a systematic review of reasons, we identified 39 reasons represented in 243 articles (scientific and grey literature) for and against special status for the reimbursement of OMPs, then categorized them into nine topics. Taking a multidisciplinary perspective, we found that most articles came from health policy (n = 103) and health economics (n = 49). More articles took the position for a special status of OMPs (n = 97) than those against it (n = 31) and there was a larger number of reasons identified in favour (29 reasons) than against (10 reasons) this special status. Conclusion Results suggest that OMP reimbursement issues should be assessed and analysed from a multidisciplinary perspective. Despite the higher occurrence of reasons and articles in favour of a special status, there is no clear-cut solution for this ethical challenge. The binary perspective of whether or not OMPs should be granted special status oversimplifies the issue: both OMPs and rare diseases are too heterogeneous in their characteristics for such a binary perspective. Thus, the scientific debate should focus less on the question of disease prevalence but rather on how the important variability of different OMPs concerning e.g. target population, cost-effectiveness, level of evidence or mechanism of action could be meaningfully addressed and implemented in Health Technology Assessments.

Abstract Methylmalonyl‐CoA mutase (MMUT) is part of the propionyl‐CoA catabolic pathway, responsible for the breakdown of branched‐chain amino acids, odd‐chain fatty acids and the side‐chain of cholesterol. Patients with deficient activity of MMUT suffer from isolated methylmalonic aciduria (MMAuria), frequently presenting in the newborn period with failure to thrive and metabolic crisis. Even well managed patients remain at risk for metabolic crises, of which one known trigger is acute illness, which may lead to poor feeding and vomiting, putting the patient in a catabolic state. This situation is believed to result in increased breakdown of propionyl‐CoA catabolic pathway precursors, producing massively elevated levels of disease related metabolites, including methylmalonic acid and propionylcarnitine. Here, we used fasting of a hemizygous mouse model (Mut‐ko/ki) of MMUT deficiency to study the role of induced catabolism on metabolite production. Although mice lost weight and displayed markers consistent with a catabolic state, contrary to expectation, we found strongly reduced levels of methylmalonic acid and propionylcarnitine in fasted conditions. Switching Mut‐ko/ki mice from a high‐protein diet to fasted conditions, or from a standard diet to a no‐protein diet, resulted in similar reductions of methylmalonic acid and propionylcarnitine levels. These results suggest, in our mouse model at least, induction of a catabolic state on its own may not be sufficient to trigger elevated metabolite levels.

Abstract The Comparative Agendas Project (CAP) collects, organizes, and makes freely available millions of bits of information concerning the objects of government attention over long periods of time (often back to the Second World War) for more than 25 political systems, worldwide. As researchers affiliated with the CAP expand their projects into Latin America, they confront some challenges similar to those from other regions, and some unique to their national political systems. In this introductory essay, we explore the background of the CAP and the opportunities posed by its expansion into Latin American political systems.

Methylmalonic acidemia is an inherited metabolic disease caused by loss or mutation of the enzyme MMUT. Here the authors use cell and animal models to show that MMUT mutations lead to defective mitophagy and stress in kidney cells, contributing to the pathogenesis in methylmalonic acidemia patients.

Abstract Background Inborn errors of metabolism (IEM) are a group of rare, heterogeneous and complex genetic conditions. Clinically, IEM often affect the central nervous system and other organs. Some carry the risk of progression and / or potentially life-threatening crises. Many patients have to adhere to lifelong dietary or drug treatment. The complexity of IEM makes it difficult for patients and caregivers to understand their pathophysiology, inheritance and therapy rationale. Especially patients reaching adolescence may have only limited knowledge of their condition since medical care has often entirely been handled by their parents. Knowledge about disease and treatment, however, constitute pillars of self-responsible disease management. Not many standardized patient education materials on IEM are available and their comprehensibility has not been systematically investigated. Methods We developed and tested patient education materials for school-aged children and adolescents with IEM. Informative texts and illustrations in paper form and as videos were developed by an international network of metabolic care professionals together with a graphic artist and experts for easy-to-read language. The materials were presented in standardized single or group training sessions to 111 individuals; first, to 74 healthy children and adolescents (recruited via public schools) and consecutively to 37 paediatric patients with IEM (phenylketonuria, galactosemia, urea cycle defects, lysosomal storage disorders) from six metabolic centres. Knowledge-gain was assessed by pre- and post-testing. Results Knowledge-gain was significant in healthy children and adolescents as well as in patients (p

Do democracies perform better than more autocratic political systems? Most existing literature focuses on single issues, such as maintaining the peace, avoiding famines, or promoting stable economic growth. The key to policy success for all these and other issues is adaptive policymaking in complex, dynamic environments. Relying on theory and empirical tests from policy process studies, we focus on extreme policy punctuations as indicators of maladaptive policymaking. We conceive of a continuum from the most open democracies to the most closed authoritarian systems, with intermediate forms of less open democracies, hybrid regimes, and less closed authoritarian systems. Based on a review of the existing literature, we extract four factors that seem to affect maladaptive and hence more punctuated policymaking: friction imposed by formal rules and informal norms on the policymaking process, the absence of incentives to address problems, centralization in policymaking, and lack of diversity in channels of information. Many of these factors cluster, so it is difficult to discern their specific effects, but our approach allows a start at doing so.

The California Current System (CCS) has experienced large fluctuations in environmental conditions in recent years that have dramatically affected the biological community. Here we synthesize remotely sensed, hydrographic, and biological survey data from throughout the CCS in 2019–2020 to evaluate how recent changes in environmental conditions have affected community dynamics at multiple trophic levels. A marine heatwave formed in the north Pacific in 2019 and reached the second greatest area ever recorded by the end of summer 2020. However, high atmospheric pressure in early 2020 drove relatively strong Ekman-driven coastal upwelling in the northern portion of the CCS and warm temperature anomalies remained far offshore. Upwelling and cooler temperatures in the northern CCS created relatively productive conditions in which the biomass of lipid-rich copepod species increased, adult krill size increased, and several seabird species experienced positive reproductive success. Despite these conditions, the composition of the fish community in the northern CCS remained a mixture of both warm- and cool-water-associated species. In the southern CCS, ocean temperatures remained above average for the seventh consecutive year. Abundances of juvenile fish species associated with productive conditions were relatively low, and the ichthyoplankton community was dominated by a mixture of oceanic warm-water and cosmopolitan species. Seabird species associated with warm water also occurred at greater densities than cool-water species in the southern CCS. The population of northern anchovy, which has been resurgent since 2017, continued to provide an important forage base for piscivorous fishes, offshore colonies of seabirds, and marine mammals throughout the CCS. Coastal upwelling in the north, and a longer-term trend in warming in the south, appeared to be controlling the community to a much greater extent than the marine heatwave itself.

The human enzyme MTHFR links the folate and methionine cycles, which are essential for the biosynthesis of nucleotides and proteins. Here, the authors present the crystal structure and biochemical analysis of human MTHFR, providing molecular insights into its function and regulation in higher eukaryotes.

We demonstrate strong self-referential effects in county-level data concerning use of the death penalty. We first show event-dependency using a repeated-event model. Higher numbers of previous events reduce the expected time delay before the next event. Second, we use a cross-sectional time-series approach to model the number of death sentences imposed in a given county in a given year. This model shows that the cumulative number of death sentences previously imposed in the same county is a strong predictor of the number imposed in a given year. Results raise troubling substantive implications: The number of death sentences in a given county in a given year is better predicted by that county's previous experience in imposing death than by the number of homicides. This explains the previously observed fact that a large share of death sentences come from a small number of counties and documents the self-referential aspects of use the death penalty. A death sentencing system based on racial dynamics and then amplified by self-referential dynamics is inconsistent with equal protection of the law, but this describes the United States system well.

Chronic cocaine use has been associated with impairments in social cognition, self-serving and antisocial behavior, and socially relevant personality disorders (PD). Despite the apparent relationship between Machiavellianism and stimulant use, no study has explicitly examined this personality concept in cocaine users so far. In the frame of the longitudinal Zurich Cocaine Cognition Study, the Machiavellianism Questionnaire (MACH-IV) was assessed in 68 recreational and 30 dependent cocaine users as well as in 68 psychostimulant-naïve controls at baseline. Additionally, three closely related personality dimensions from the Temperament and Character Inventory (TCI)-cooperativeness, (social) reward dependence, and self-directedness-and the screening questionnaire of the Structured Clinical Interview for DSM-IV Axis II Personality Disorders (SCID-II) were acquired. At the one-year follow-up, 57 cocaine users and 48 controls were reassessed with the MACH-IV. Finally, MACH-IV scores were correlated with measures of social cognition and interaction (cognitive/emotional empathy, Theory-of-Mind, prosocial behavior) and with SCID-II PD scores assessed at baseline. Both recreational and dependent cocaine users showed significantly higher Machiavellianism than controls, while dependent cocaine users additionally displayed significantly lower levels of TCI cooperativeness and self-directedness. During the one-year interval, MACH-IV scores showed high test-retest reliability and also the significant gap between cocaine users and controls remained. Moreover, in cocaine users, higher Machiavellianism correlated significantly with lower levels of cooperativeness and self-directedness, with less prosocial behavior, and with higher cluster B PD scores. However, Machiavellianism was not correlated with measures of cocaine use severity (r