Four-dimensional Lipidomics Profiling in X-linked Adrenoleukodystrophy using Trapped Ion Mobility Mass Spectrometry.

Lipids play pivotal roles in an extensive range of metabolic and physiological processes. In recent years, the convergence of trapped ion mobility (TIMS) and mass spectrometry (MS) has enabled 4D-lipidomics, a highly promising technology for comprehensive lipid analysis. 4D-lipidomics assesses lipid annotations across four distinct dimensions-retention time, collisional cross section, m/z (mass-to-charge ratio), and MS/MS spectra-providing a heightened level of confidence in lipid annotation. These advantages prove particularly valuable when investigating complex disorders involving lipid metabolism, such as adrenoleukodystrophy (ALD). ALD is characterized by the accumulation of very-long-chain fatty acids (VLCFA) due to pathogenic variants in the ABCD1 gene. A comprehensive 4D-lipidomics strategy of ALD fibroblasts demonstrated significant elevations of various lipids from multiple classes. This indicates that the changes observed in ALD are not confined to a single lipid class and likely impacts a broad spectrum of lipid-mediated physiological processes. Our findings highlight the incorporation of mainly saturated and monounsaturated VLCFA variants into a range of lipid classes, encompassing phosphatidylcholines, triacylglycerols, and cholesterol esters. These include ultra-long-chain fatty acids with a length of up to thirty carbon atoms. Lipid species containing C26:0, C26:1 were the most frequently detected VLCFA lipids in our study. Furthermore, we report a panel of 121 new candidate biomarkers in fibroblasts, exhibiting significant differentiation between controls and individuals with ALD. In summary, this study demonstrates the capabilities of a 4D-lipid profiling workflow in unraveling novel insights into the intricate lipid modifications associated with metabolic disorders like ALD.

Disruption of polyunsaturated fatty acid biosynthesis drives STING-dependent acute myeloid leukemia cell maturation and death.

The role of polyunsaturated fatty acid (PUFA) biosynthesis in acute myeloid leukemia (AML) remains largely undefined. A comparative expression analysis of 35 genes encoding fatty acid biosynthesis enzymes showed that fatty acid desaturase 1 (FADS1) was highly expressed across multiple AML subtypes relative to healthy controls and that elevated FADS1 expression correlates with worse overall AML patient survival. Functionally, shRNA-mediated inhibition of FADS1 reduced AML cell growth in vitro and significantly delayed leukemia onset in an AML mouse model. AML cell lines depleted of FADS1 arrested in the G1/S-phase of the cell cycle, acquired characteristics of myeloid maturation and subsequently died. To understand the molecular consequences of FADS1 inhibition, a combination of mass spectrometry-based analysis of complex lipids and gene expression analysis (RNA-seq) was performed. FADS1 inhibition caused AML cells to exhibit significant lipidomic remodeling, including depletion of PUFAs from the phospholipids, phosphatidylserine, and phosphatidylethanolamine. These lipidomic alterations were accompanied by an increase induction of inflammatory and stimulator of interferon genes (STING)-mediated type-1 interferon signaling. Remarkably, genetic deletion of STING largely prevented the AML cell maturation and death phenotypes mediated by FADS1 inhibition. Highlighting the therapeutic implications of these findings, pharmacological blockade of PUFA biosynthesis reduced patient-derived AML cell numbers ex vivo but not that of healthy donor cells. Similarly, STING agonism attenuated patient-derived-AML survival; however, STING activation also reduced healthy granulocyte numbers. Collectively, these data unveil a previously unrecognized importance of PUFA biosynthesis in leukemogenesis and that imbalances in PUFA metabolism can drive STING-mediated AML maturation and death.

The impact of bed rest on human skeletal muscle metabolism.

Insulin sensitivity and metabolic flexibility decrease in response to bed rest, but the temporal and causal adaptations in human skeletal muscle metabolism are not fully defined. Here, we use an integrative approach to assess human skeletal muscle metabolism during bed rest and provide a multi-system analysis of how skeletal muscle and the circulatory system adapt to short- and long-term bed rest (German Clinical Trials: DRKS00015677). We uncover that intracellular glycogen accumulation after short-term bed rest accompanies a rapid reduction in systemic insulin sensitivity and less GLUT4 localization at the muscle cell membrane, preventing further intracellular glycogen deposition after long-term bed rest. We provide evidence of a temporal link between the accumulation of intracellular triglycerides, lipotoxic ceramides, and sphingomyelins and an altered skeletal muscle mitochondrial structure and function after long-term bed rest. An intracellular nutrient overload therefore represents a crucial determinant for rapid skeletal muscle insulin insensitivity and mitochondrial alterations after prolonged bed rest.

The Concise Guide to PHARMACOLOGY 2023/24: Transporters.

The Concise Guide to PHARMACOLOGY 2023/24 is the sixth in this series of biennial publications. The Concise Guide provides concise overviews, mostly in tabular format, of the key properties of approximately 1800 drug targets, and over 6000 interactions with about 3900 ligands. There is an emphasis on selective pharmacology (where available), plus links to the open access knowledgebase source of drug targets and their ligands (https://www.guidetopharmacology.org/), which provides more detailed views of target and ligand properties. Although the Concise Guide constitutes almost 500 pages, the material presented is substantially reduced compared to information and links presented on the website. It provides a permanent, citable, point-in-time record that will survive database updates. The full contents of this section can be found at http://onlinelibrary.wiley.com/doi/10.1111/bph.16182. Transporters are one of the six major pharmacological targets into which the Guide is divided, with the others being: G protein-coupled receptors, ion channels, nuclear hormone receptors, catalytic receptors and enzymes. These are presented with nomenclature guidance and summary information on the best available pharmacological tools, alongside key references and suggestions for further reading. The landscape format of the Concise Guide is designed to facilitate comparison of related targets from material contemporary to mid-2023, and supersedes data presented in the 2021/22, 2019/20, 2017/18, 2015/16 and 2013/14 Concise Guides and previous Guides to Receptors and Channels. It is produced in close conjunction with the Nomenclature and Standards Committee of the International Union of Basic and Clinical Pharmacology (NC-IUPHAR), therefore, providing official IUPHAR classification and nomenclature for human drug targets, where appropriate.

ABCD1 Transporter Deficiency Results in Altered Cholesterol Homeostasis.

X-linked adrenoleukodystrophy (X-ALD), the most common peroxisomal disorder, is caused by mutations in the peroxisomal transporter ABCD1, resulting in the accumulation of very long-chain fatty acids (VLCFA). Strongly affected cell types, such as oligodendrocytes, adrenocortical cells and macrophages, exhibit high cholesterol turnover. Here, we investigated how ABCD1 deficiency affects cholesterol metabolism in human X-ALD patient-derived fibroblasts and CNS tissues of Abcd1-deficient mice. Lipidome analyses revealed increased levels of cholesterol esters (CE), containing both saturated VLCFA and mono/polyunsaturated (V)LCFA. The elevated CE(26:0) and CE(26:1) levels remained unchanged in LXR agonist-treated Abcd1 KO mice despite reduced total C26:0. Under high-cholesterol loading, gene expression of SOAT1, converting cholesterol to CE and lipid droplet formation were increased in human X-ALD fibroblasts versus healthy control fibroblasts. However, the expression of NCEH1, catalysing CE hydrolysis and the cholesterol transporter ABCA1 and cholesterol efflux were also upregulated. Elevated Soat1 and Abca1 expression and lipid droplet content were confirmed in the spinal cord of X-ALD mice, where expression of the CNS cholesterol transporter Apoe was also elevated. The extent of peroxisome-lipid droplet co-localisation appeared low and was not impaired by ABCD1-deficiency in cholesterol-loaded primary fibroblasts. Finally, addressing steroidogenesis, progesterone-induced cortisol release was amplified in X-ALD fibroblasts. These results link VLCFA to cholesterol homeostasis and justify further consideration of therapeutic approaches towards reducing VLCFA and cholesterol levels in X-ALD.

Attitudes of Patients with Adrenoleukodystrophy towards Sex-Specific Newborn Screening.

Newborn screening (NBS) for X-linked adrenoleukodystrophy (ALD) can identify affected individuals before the onset of life-threatening manifestations. Some countries have decided to only screen boys (sex-specific screening). This study investigates the attitudes of individuals with ALD towards sex-specific NBS for ALD. A questionnaire was sent to all patients in the Dutch ALD cohort. Invitees were asked who they thought should be screened for ALD: only boys, both boys and girls or neither. The motives and background characteristics of respondents were compared between screening preferences. Out of 108 invitees, 66 participants (61%), 38 men and 28 women, participated in this study. The majority (n = 53, 80%) favored screening both newborn boys and girls for ALD, while 20% preferred boys only. None of the respondents felt that newborns should not be screened for ALD. There were no differences in the background characteristics of the respondents between screening preferences. Our study revealed a diverse range of motivations underlying respondents' screening preferences. This study is one of the first to investigate the attitudes of patients towards sex-specific screening for ALD. The outcomes of this study can offer insights to stakeholders engaged in the implementation of NBS programs. ALD patients are important stakeholders who can provide valuable input in this process.

VUS: Variant of uncertain significance or very unclear situation?

The advancements in population screening, including newborn screening, enables the identification of disease-causing variants and timely initiation of treatment. However, screening may also identify mild variants, non-disease variants, and variants of uncertain significance (VUS). The identification of a VUS poses a challenge in terms of diagnostic uncertainty and confusion. X-linked adrenoleukodystrophy (ALD) serves as an illustrative example of this complex issue. ALD is a monogenic neurometabolic disease with a complex clinical presentation and a lack of predictive tests for clinical severity. Despite the success of ALD newborn screening, a significant proportion (62%) of missense variants identified through newborn screening exhibit uncertainty regarding their pathogenicity. Resolving this issue requires ongoing efforts to accurately classify variants and refine screening protocols. While it is undisputable that ALD newborn screening greatly benefits boys with the disease, the identification of VUS underscores the need for continuous research and collaboration in improving screening practices.

Sex-specific newborn screening for X-linked adrenoleukodystrophy.

Males with X-linked adrenoleukodystrophy (ALD) are at high risk for developing adrenal insufficiency and/or progressive leukodystrophy (cerebral ALD) at an early age. Pathogenic variants in ABCD1 result in elevated levels of very long-chain fatty acids (VLCFA), including C26:0-lysophosphatidylcholine (C26:0-LPC). Newborn screening for ALD enables prospective monitoring and timely therapeutic intervention, thereby preventing irreversible damage and saving lives. The Dutch Health Council recommended to screen only male newborns for ALD without identifying untreatable conditions associated with elevated C26:0-LPC, like Zellweger spectrum disorders and single peroxisomal enzyme defects. Here, we present the results of the SCAN (Screening for ALD in the Netherlands) study which is the first sex-specific newborn screening program worldwide. Males with ALD are identified based on elevated C26:0-LPC levels, the presence of one X-chromosome and a variant in ABCD1, in heel prick dried bloodspots. Screening of 71 208 newborns resulted in the identification of four boys with ALD who, following referral to the pediatric neurologist and confirmation of the diagnosis, enrolled in a long-term follow-up program. The results of this pilot show the feasibility of employing a boys-only screening algorithm that identifies males with ALD without identifying untreatable conditions. This approach will be of interest to countries that are considering ALD newborn screening but are reluctant to identify girls with ALD because for girls there is no direct health benefit. We also analyzed whether gestational age, sex, birth weight and age at heel prick blood sampling affect C26:0-LPC concentrations and demonstrate that these covariates have a minimal effect.

Saturated very long-chain fatty acids regulate macrophage plasticity and invasiveness.

Saturated very long-chain fatty acids (VLCFA, ≥ C22), enriched in brain myelin and innate immune cells, accumulate in X-linked adrenoleukodystrophy (X-ALD) due to inherited dysfunction of the peroxisomal VLCFA transporter ABCD1. In its severest form, X-ALD causes cerebral myelin destruction with infiltration of pro-inflammatory skewed monocytes/macrophages. How VLCFA levels relate to macrophage activation is unclear. Here, whole transcriptome sequencing of X-ALD macrophages indicated that VLCFAs prime human macrophage membranes for inflammation and increased expression of factors involved in chemotaxis and invasion. When added externally to mimic lipid release in demyelinating X-ALD lesions, VLCFAs did not activate toll-like receptors in primary macrophages. In contrast, VLCFAs provoked pro-inflammatory responses through scavenger receptor CD36-mediated uptake, cumulating in JNK signalling and expression of matrix-degrading enzymes and chemokine release. Following pro-inflammatory LPS activation, VLCFA levels increased also in healthy macrophages. With the onset of the resolution, VLCFAs were rapidly cleared in control macrophages by increased peroxisomal VLCFA degradation through liver-X-receptor mediated upregulation of ABCD1. ABCD1 deficiency impaired VLCFA homeostasis and prolonged pro-inflammatory gene expression upon LPS treatment. Our study uncovers a pivotal role for ABCD1, a protein linked to neuroinflammation, and associated peroxisomal VLCFA degradation in regulating macrophage plasticity.

Peroxisomal very long-chain fatty acid transport is targeted by herpesviruses and the antiviral host response.

Very long-chain fatty acids (VLCFA) are critical for human cytomegalovirus replication and accumulate upon infection. Here, we used Epstein-Barr virus (EBV) infection of human B cells to elucidate how herpesviruses target VLCFA metabolism. Gene expression profiling revealed that, despite a general induction of peroxisome-related genes, EBV early infection decreased expression of the peroxisomal VLCFA transporters ABCD1 and ABCD2, thus impairing VLCFA degradation. The mechanism underlying ABCD1 and ABCD2 repression involved RNA interference by the EBV-induced microRNAs miR-9-5p and miR-155, respectively, causing significantly increased VLCFA levels. Treatment with 25-hydroxycholesterol, an antiviral innate immune modulator produced by macrophages, restored ABCD1 expression and reduced VLCFA accumulation in EBV-infected B-lymphocytes, and, upon lytic reactivation, reduced virus production in control but not ABCD1-deficient cells. Finally, also other herpesviruses and coronaviruses target ABCD1 expression. Because viral infection might trigger neuroinflammation in X-linked adrenoleukodystrophy (X-ALD, inherited ABCD1 deficiency), we explored a possible link between EBV infection and cerebral X-ALD. However, neither immunohistochemistry of post-mortem brains nor analysis of EBV seropositivity in 35 X-ALD children supported involvement of EBV in the onset of neuroinflammation. Collectively, our findings indicate a previously unrecognized, pivotal role of ABCD1 in viral infection and host defence, prompting consideration of other viral triggers in cerebral X-ALD.

Treatment of cerebral adrenoleukodystrophy: allogeneic transplantation and lentiviral gene therapy.

INTRODUCTION: Adrenoleukodystrophy (ALD) is an X-linked peroxisomal disorder with an incidence of 1 in 14-17,000 male births, caused by pathogenic variants within the ABCD1 gene. By adulthood, approximately 40% of the patients develop cerebral ALD, a severe, neuroinflammatory condition that is generally progressive and fatal without intervention. AREAS COVERED: Historically, only allogeneic hematopoietic stem cell transplantation (HSCT) has been shown to halt progression of cerebral ALD, with superior outcomes obtained when HSCT is performed early in the disease process. More recently, a lentiviral-based gene therapy approach has been investigated as therapy for cerebral ALD as an alternative to allogeneic transplantation. A focused literature review was performed using the terms 'hematopoietic stem cell transplantation,' 'gene therapy' and 'adrenoleukodystrophy' to include relevant literature, especially comparing the experience with gene therapy and HSCT outcomes. We review the history and experience with HSCT in cerebral ALD and its limitations, as well as the information currently available in association with the gene therapy trials for cerebral ALD. EXPERT OPINION: The data regarding this lentiviral-based gene therapy approach and its relative risks and benefits is still being evaluated. This information is explored in the context of the experience with allogeneic HSCT for cerebral ALD.

International Recommendations for the Diagnosis and Management of Patients With Adrenoleukodystrophy: A Consensus-Based Approach.

Pathogenic variants in the ABCD1 gene cause adrenoleukodystrophy (ALD), a progressive metabolic disorder characterized by 3 core clinical syndromes: a slowly progressive myeloneuropathy, a rapidly progressive inflammatory leukodystrophy (cerebral ALD), and primary adrenal insufficiency. These syndromes are not present in all individuals and are not related to genotype. Cerebral ALD and adrenal insufficiency require early detection and intervention and warrant clinical surveillance because of variable penetrance and age at onset. Newborn screening has increased the number of presymptomatic individuals under observation, but clinical surveillance protocols vary. We used a consensus-based modified Delphi approach among 28 international ALD experts to develop best-practice recommendations for diagnosis, clinical surveillance, and treatment of patients with ALD. We identified 39 discrete areas of consensus. Regular monitoring to detect the onset of adrenal failure and conversion to cerebral ALD is recommended in all male patients. Hematopoietic cell transplant (HCT) is the treatment of choice for cerebral ALD. This guideline addresses a clinical need in the ALD community worldwide as the number of overall diagnoses and presymptomatic individuals is increasing because of newborn screening and greater availability of next-generation sequencing. The poor ability to predict the disease course informs current monitoring intervals but remains subject to change as more data emerge. This knowledge gap should direct future research and illustrates once again that international collaboration among physicians, researchers, and patients is essential to improving care.

Parents' views on accepting, declining, and expanding newborn bloodspot screening.

INTRODUCTION: The goal of newborn bloodspot screening (NBS) is the early detection of treatable disorders in newborns to offer early intervention. Worldwide, the number of conditions screened for is expanding, which might affect public acceptance. In the Netherlands, participation is high (>99%), but little is known about how parents perceive NBS. This study assessed parents' views on accepting, declining and expanding NBS. METHODS: A total of 804 of 6051 (13%) invited parents who participated in NBS in the Netherlands during the last two weeks of December 2019, and 48 of 1162 (4%) invited parents who declined participation in NBS in 2019 and 2020, completed a questionnaire. RESULTS: The most important reason for parents to participate in NBS was to prevent health complaints, whereas the most important reason to decline NBS was parents' viewpoint on life and the belief that the heel prick would be painful for the child. Compared to NBS participants, respondents who declined NBS were more actively religious, considered alternative medicine or lifestyle more important, were less inclined to vaccinate their child for infectious diseases, and reported more doubt about NBS participation (all differences p < .001). Informed choice was lower among respondents who declined NBS (44%) compared to participants in NBS (83%, p < .001), mostly due to insufficient knowledge. Of the NBS participants, 95% were positive about NBS expansion. Most NBS participants agreed to include conditions that could unintentionally reveal a diagnosis in the mother instead of the child (86%) or a condition that may not cause symptoms until later in the child's life (84%). CONCLUSION: Most participants made an informed decision to participate in NBS and are positive about screening for more conditions. Insights into parents' views on (non-)participation and expansion of NBS can help to ensure that NBS suits the population needs while safeguarding ethical principles for screening.

Peroxisome Metabolism Contributes to PIEZO2-Mediated Mechanical Allodynia.

Mutations in the peroxisomal half-transporter ABCD1 cause X-linked adrenoleukodystrophy, resulting in elevated very long-chain fatty acids (VLCFA), progressive neurodegeneration and an associated pain syndrome that is poorly understood. In the nervous system of mice, we found ABCD1 expression to be highest in dorsal root ganglia (DRG), with satellite glial cells (SGCs) displaying higher expression than neurons. We subsequently examined sensory behavior and DRG pathophysiology in mice deficient in ABCD1 compared to wild-type mice. Beginning at 8 months of age, Abcd1-/y mice developed persistent mechanical allodynia. DRG had a greater number of IB4-positive nociceptive neurons expressing PIEZO2, the mechanosensitive ion channel. Blocking PIEZO2 partially rescued the mechanical allodynia. Beyond affecting neurons, ABCD1 deficiency impacted SGCs, as demonstrated by high levels of VLCFA, increased glial fibrillary acidic protein (GFAP), as well as genes disrupting neuron-SGC connectivity. These findings suggest that lack of the peroxisomal half-transporter ABCD1 leads to PIEZO2-mediated mechanical allodynia as well as SGC dysfunction. Given the known supportive role of SGCs to neurons, this elucidates a novel mechanism underlying pain in X-linked adrenoleukodystrophy.

Structure and Function of the ABCD1 Variant Database: 20 Years, 940 Pathogenic Variants, and 3400 Cases of Adrenoleukodystrophy.

The progressive neurometabolic disorder X-linked adrenoleukodystrophy (ALD) is caused by pathogenic variants in the ABCD1 gene, which encodes the peroxisomal ATP-binding transporter for very-long-chain fatty acids. The clinical spectrum of ALD includes adrenal insufficiency, myelopathy, and/or leukodystrophy. A complicating factor in disease management is the absence of a genotype-phenotype correlation in ALD. Since 1999, most ABCD1 (likely) pathogenic and benign variants have been reported in the ABCD1 Variant Database. In 2017, following the expansion of ALD newborn screening, the database was rebuilt. To add an additional level of confidence with respect to pathogenicity, for each variant, it now also reports the number of cases identified and, where available, experimental data supporting the pathogenicity of the variant. The website also provides information on a number of ALD-related topics in several languages. Here, we provide an updated analysis of the known variants in ABCD1. The order of pathogenic variant frequency, overall clustering of disease-causing variants in exons 1-2 (transmembrane domain spanning region) and 6-9 (ATP-binding domain), and the most commonly reported pathogenic variant p.Gln472Argfs*83 in exon 5 are consistent with the initial reports of the mutation database. Novel insights include nonrandom clustering of high-density missense variant hotspots within exons 1, 2, 6, 8, and 9. Perhaps more importantly, we illustrate the importance of collaboration and utility of the database as a scientific, clinical, and ALD-community-wide resource.

Molecular Biomarkers for Adrenoleukodystrophy: An Unmet Need.

X-linked adrenoleukodystrophy (ALD) is an inherited progressive neurometabolic disease caused by mutations in the ABCD1 gene and the accumulation of very long-chain fatty acids in plasma and tissues. Patients present with heterogeneous clinical manifestations which can include adrenal insufficiency, myelopathy, and/or cerebral demyelination. In the absence of a genotype-phenotype correlation, the clinical outcome of an individual cannot be predicted and currently there are no molecular markers available to quantify disease severity. Therefore, there is an unmet clinical need for sensitive biomarkers to monitor and/or predict disease progression and evaluate therapy efficacy. The increasing amount of biological sample repositories ('biobanking') as well as the introduction of newborn screening creates a unique opportunity for identification and evaluation of new or existing biomarkers. Here we summarize and review the many studies that have been performed to identify and improve knowledge surrounding candidate molecular biomarkers for ALD. We also highlight several shortcomings of ALD biomarker studies, which often include a limited sample size, no collection of longitudinal data, and no validation of findings in an external cohort. Nonetheless, these studies have generated a list of interesting biomarker candidates and this review aspires to direct future biomarker research.

Biochemical Studies in Fibroblasts to Interpret Variants of Unknown Significance in the ABCD1 Gene.

Due to newborn screening for X-linked adrenoleukodystrophy (ALD), and the use of exome sequencing in clinical practice, the detection of variants of unknown significance (VUS) in the ABCD1 gene is increasing. In these cases, functional tests in fibroblasts may help to classify a variant as (likely) benign or pathogenic. We sought to establish reference ranges for these tests in ALD patients and control subjects with the aim of helping to determine the pathogenicity of VUS in ABCD1. Fibroblasts from 36 male patients with confirmed ALD, 26 healthy control subjects and 17 individuals without a family history of ALD, all with an uncertain clinical diagnosis and a VUS identified in ABCD1, were included. We performed a combination of tests: (i) a test for very-long-chain fatty acids (VLCFA) levels, (ii) a D3-C22:0 loading test to study the VLCFA metabolism and (iii) immunoblotting for ALD protein. All ALD patient fibroblasts had elevated VLCFA levels and a reduced peroxisomal ß-oxidation capacity (as measured by the D3-C16:0/D3-C22:0 ratio in the D3-C22:0 loading test) compared to the control subjects. Of the VUS cases, the VLCFA metabolism was not significantly impaired (most test results were within the reference range) in 6/17, the VLCFA metabolism was significantly impaired (most test results were within/near the ALD range) in 9/17 and a definite conclusion could not be drawn in 2/17 of the cases. Biochemical studies in fibroblasts provided clearly defined reference and disease ranges for the VLCFA metabolism. In 15/17 (88%) VUS we were able to classify the variant as being likely benign or pathogenic. This is of great clinical importance as new variants will be detected.

Expanding Neonatal Bloodspot Screening: A Multi-Stakeholder Perspective.

Neonatal bloodspot screening (NBS) aims to detect treatable disorders in newborns. The number of conditions included in the screening is expanding through technological and therapeutic developments, which can result in health gain for more newborns. NBS expansion, however, also poses healthcare, ethical and societal challenges. This qualitative study explores a multi-stakeholders' perspective on current and future expansions of NBS. Semi-structured interviews were conducted with 22 Dutch professionals, including healthcare professionals, test developers and policy makers, and 17 parents of children with normal and abnormal NBS results. Addressed themes were (1) benefits and challenges of current expansion, (2) expectations regarding future developments, and (3) NBS acceptance and consent procedures. Overall, participants had a positive attitude toward NBS expansion, as long as it is aimed at detecting treatable disorders and achieving health gain. Concerns were raised regarding an increase in results of uncertain significance, diagnosing asymptomatic mothers, screening of subgroups ("males only"), finding untreatable disorders, along with increasingly complex consent procedures. Regarding the scope of future NBS expansions, two types of stakeholder perspectives emerged. Stakeholders with a "targeted-scope" perspective saw health gain for the neonate as the exclusive NBS aim. They thought pre-test information could be limited, and parents should be protected against too much options or information. Stakeholders with a "broad-scope" perspective thought the NBS aim should be formulated broader, for example, also taking (reproductive) life planning into account. They put more emphasis on individual preferences and parental autonomy. Policy-makers should engage with both perspectives when making further decisions about NBS.

THE CONCISE GUIDE TO PHARMACOLOGY 2021/22: Transporters.

The Concise Guide to PHARMACOLOGY 2021/22 is the fifth in this series of biennial publications. The Concise Guide provides concise overviews, mostly in tabular format, of the key properties of nearly 1900 human drug targets with an emphasis on selective pharmacology (where available), plus links to the open access knowledgebase source of drug targets and their ligands (www.guidetopharmacology.org), which provides more detailed views of target and ligand properties. Although the Concise Guide constitutes over 500 pages, the material presented is substantially reduced compared to information and links presented on the website. It provides a permanent, citable, point-in-time record that will survive database updates. The full contents of this section can be found at http://onlinelibrary.wiley.com/doi/bph.15543. Transporters are one of the six major pharmacological targets into which the Guide is divided, with the others being: G protein-coupled receptors, ion channels, nuclear hormone receptors, catalytic receptors and enzymes. These are presented with nomenclature guidance and summary information on the best available pharmacological tools, alongside key references and suggestions for further reading. The landscape format of the Concise Guide is designed to facilitate comparison of related targets from material contemporary to mid-2021, and supersedes data presented in the 2019/20, 2017/18, 2015/16 and 2013/14 Concise Guides and previous Guides to Receptors and Channels. It is produced in close conjunction with the Nomenclature and Standards Committee of the International Union of Basic and Clinical Pharmacology (NC-IUPHAR), therefore, providing official IUPHAR classification and nomenclature for human drug targets, where appropriate.

iBRET Screen of the ABCD1 Peroxisomal Network and Mutation-Induced Network Perturbations.

Mapping the network of proteins provides a powerful means to investigate the function of disease genes and to unravel the molecular basis of phenotypes. We present an automated informatics-aided and bioluminescence resonance energy transfer-based approach (iBRET) enabling high-confidence detection of protein-protein interactions in living mammalian cells. A screen of the ABCD1 protein, which is affected in X-linked adrenoleukodystrophy (X-ALD), against an organelle library of peroxisomal proteins demonstrated applicability of iBRET for large-scale experiments. We identified novel protein-protein interactions for ABCD1 (with ALDH3A2, DAO, ECI2, FAR1, PEX10, PEX13, PEX5, PXMP2, and PIPOX), mapped its position within the peroxisomal protein-protein interaction network, and determined that pathogenic missense variants in ABCD1 alter the interaction with selected binding partners. These findings provide mechanistic insights into pathophysiology of X-ALD and may foster the identification of new disease modifiers.