Accepting or declining preconception expanded carrier screening: An exploratory study with 407 couples.

Rapidly evolving genomic technologies have made genetic expanded carrier screening (ECS) possible for couples considering a pregnancy. The aim of ECS is to identify couples at risk of having a child affected with a severe disorder and to facilitate their reproductive decision-making process. The ECS test we offer at our center, called BeGECS (Belgian Genetic ECS), consists of 1268 autosomal recessive (AR) and X-linked pathogenic genes, including severe childhood-onset disorders. However, thus far data are scarce regarding the actual uptake of preconception ECS in a clinical setting. Therefore, our aim was to describe the characteristics of 407 couples to whom ECS was offered at the Center for Medical Genetics of the University Hospital Ghent (CMGG). In addition, we aimed to identify their reasons for accepting or declining BeGECS. Between October 2019 and January 2023, 407 preconception couples were offered BeGECS and were asked to fill in a questionnaire after their decision. Of the 407 couples participating in the survey, 270 (66%) decided to take the test and 137 (34%) declined. We observed that age, highest education level as well as indication for consultation were statistically different between the group that accepted to take the test and the group that declined (p = 0.037). In particular, age and education level were substantially higher in the group that accepted the test. Major reasons for taking BeGECS include prevention, wishing to obtain all information possible, helping preparing their future reproductive decision and increasing their sense of control by being informed. However, couples that do not chose to take BeGECS stated that too much information would make them anxious, that the result would not change their decision to have children, that they do not want to spend money on something that will not happen and that they do not worry about their family history. These findings show that the majority of preconception couples that were offered ECS, accepted the test.

Association of antenatal evaluations with postmortem and genetic findings in the series of fetal osteogenesis imperfecta.

INTRODUCTION: Counseling Osteogenesis Imperfecta (OI) pregnancies is challenging due to the wide range of onsets and clinical severities, from perinatal lethality to milder forms detected later in life. METHODS: Thirty-eight individuals from 36 families were diagnosed with OI through prenatal ultrasonography and/or postmortem clinical and radiographic findings. Genetic analysis was conducted on 26 genes associated with OI in these subjects that emerged over the past 20 years, while some genes were examined progressively, all 26 genes were examined in the group where no pathogenic variations were detected. RESULTS: Prenatal and postnatal observations both consistently showed short limbs in 97%, followed by bowing of the long bones in 89%. Among 32 evaluated cases, all exhibited cranial hypomineralization. Fractures were found in 29 (76%) cases, with multiple bones involved in 18 of them. Genetic associations were disclosed in 27 families with 22 (81%) autosomal dominant and five (19%) autosomal recessive forms, revealing 25 variants in six genes (COL1A1, COL1A2, CREB3L1, P3H1, FKBP10, and IFITM5), including nine novels. Postmortem radiological examination showed variability in intra-family expression of CREBL3 and P3H1-related OI. CONCLUSION: Prenatal diagnosis for distinguishing OI and its subtypes relies on factors such as family history, timing, ultrasound, genetic and postmortem evaluation.

Syntaxin 18 Defects in Human and Zebrafish Unravel Key Roles in Early Cartilage and Bone Development.

SNARE proteins comprise a conserved protein family responsible for catalyzing membrane fusion during vesicle traffic. Syntaxin18 (STX18) is a poorly characterized endoplasmic reticulum (ER)-resident t-SNARE. Recently, together with TANGO1 and SLY1, its involvement was shown in ER to Golgi transport of collagen II during chondrogenesis. We report a fetus with a severe osteochondrodysplasia in whom we identified a homozygous substitution of the highly conserved p.Arg10 to Pro of STX18. CRISPR/Cas9-mediated Stx18 deficiency in zebrafish reveals a crucial role for Stx18 in cartilage and bone development. Furthermore, increased expression of multiple components of the Stx18 SNARE complex and of COPI and COPII proteins suggests that Stx18 deficiency impairs antero- and retrograde vesicular transport in the crispant stx18 zebrafish. Taken together, our studies highlight a new candidate gene for a recessive form of osteochondrodysplasia, thereby possibly broadening the SNAREopathy phenotypic spectrum and opening new doors toward future research avenues. © 2023 American Society for Bone and Mineral Research (ASBMR).

Retained chromosomal integrity following CRISPR-Cas9-based mutational correction in human embryos.

Human germline gene correction by targeted nucleases holds great promise for reducing mutation transmission. However, recent studies have reported concerning observations in CRISPR-Cas9-targeted human embryos, including mosaicism and loss of heterozygosity (LOH). The latter has been associated with either gene conversion or (partial) chromosome loss events. In this study, we aimed to correct a heterozygous basepair substitution in PLCZ1, related to infertility. In 36% of the targeted embryos that originated from mutant sperm, only wild-type alleles were observed. By performing genome-wide double-digest restriction site-associated DNA sequencing, integrity of the targeted chromosome (i.e., no deletions larger than 3 Mb or chromosome loss) was confirmed in all seven targeted GENType-analyzed embryos (mutant editing and absence of mutation), while short-range LOH events (shorter than 10 Mb) were clearly observed by single-nucleotide polymorphism assessment in two of these embryos. These results fuel the currently ongoing discussion on double-strand break repair in early human embryos, making a case for the occurrence of gene conversion events or partial template-based homology-directed repair.

Cell differentiation and matrix organization are differentially affected during bone formation in osteogenesis imperfecta zebrafish models with different genetic defects impacting collagen type I structure.

Osteogenesis imperfecta (OI) is a family of rare heritable skeletal disorders associated with dominant mutations in the collagen type I encoding genes and recessive defects in proteins involved in collagen type I synthesis and processing and in osteoblast differentiation and activity. Historically, it was believed that the OI bone phenotype was only caused by abnormal collagen type I fibrils in the extracellular matrix, but more recently it became clear that the altered bone cell homeostasis, due to mutant collagen retention, plays a relevant role in modulating disease severity in most of the OI forms and it is correlated to impaired bone cell differentiation. Despite in vitro evidence, in vivo data are missing. To better understand the physiopathology of OI, we used two zebrafish models: Chihuahua (Chi/+), carrying a dominant p.G736D substitution in the α1 chain of collagen type I, and the recessive p3h1-/-, lacking prolyl 3-hydroxylase (P3h1) enzyme. Both models share the delay of collagen type I folding, resulting in its overmodification and partial intracellular retention. The regeneration of the bony caudal fin of Chi/+ and p3h1-/- was employed to investigate the impact of abnormal collagen synthesis on bone cell differentiation. Reduced regenerative ability was evident in both models, but it was associated to impaired osteoblast differentiation and osteoblastogenesis/adipogenesis switch only in Chi/+. On the contrary, reduced osteoclast number and activity were found in both models during regeneration. The dominant OI model showed a more detrimental effect in the extracellular matrix organization. Interestingly, the chemical chaperone 4-phenylbutyrate (4-PBA), known to reduce cellular stress and increase collagen secretion, improved bone formation only in p3h1-/- by favoring caudal fin growth without affecting bone cell markers expression. Taken together, our in vivo data proved the negative impact of structurally abnormal collagen type I on bone formation but revealed a gene mutation-specific effect on bone cell differentiation and matrix organization in OI. These, together with the distinct ability to respond to the chaperone treatment, underline the need for precision medicine approaches to properly treat the disease.

Inorganic Pyrophosphate Plasma Levels Are Decreased in Pseudoxanthoma Elasticum Patients and Heterozygous Carriers but Do Not Correlate with the Genotype or Phenotype.

Pseudoxanthoma elasticum (PXE) is a rare ectopic calcification disorder affecting soft connective tissues that is caused by biallelic ABCC6 mutations. While the underlying pathomechanisms are incompletely understood, reduced circulatory levels of inorganic pyrophosphate (PPi)-a potent mineralization inhibitor-have been reported in PXE patients and were suggested to be useful as a disease biomarker. In this study, we explored the relation between PPi, the ABCC6 genotype and the PXE phenotype. For this, we optimized and validated a PPi measurement protocol with internal calibration that can be used in a clinical setting. An analysis of 78 PXE patients, 69 heterozygous carriers and 14 control samples revealed significant differences in the measured PPi levels between all three cohorts, although there was overlap between all groups. PXE patients had a ±50% reduction in PPi levels compared to controls. Similarly, we found a ±28% reduction in carriers. PPi levels were found to correlate with age in PXE patients and carriers, independent of the ABCC6 genotype. No correlations were found between PPi levels and the Phenodex scores. Our results suggest that other factors besides PPi are at play in ectopic mineralization, which limits the use of PPi as a predictive biomarker for severity and disease progression.

Generation of human induced pluripotent stem cell line UGENTi001-A from a patient with Marfan syndrome carrying a heterozygous c.7754 T > C variant in FBN1 and the isogenic control UGENT001-A-1 using CRISPR/Cas9 editing.

Marfan syndrome is an autosomal dominant genetic disorder resulting from pathogenic variants in FBN1 gene. FBN1 encodes for fibrillin-1, an important extracellular matrix protein. Impaired fibrillin-1 affects multiple organ systems, including the cardiovascular system. We generated an iPSC line carrying a heterozygous variant c.7754 T > C (p.Ile2585Thr, missense) in FBN1 from a patient with Marfan syndrome. Also, an isogenic control is generated, where the pathogenic variant is repaired using CRISPR-Cas9. This isogenic pair provides a valuable resource for in vitro disease modelling.

Zebrafish Tric-b is required for skeletal development and bone cells differentiation.

Introduction: Trimeric intracellular potassium channels TRIC-A and -B are endoplasmic reticulum (ER) integral membrane proteins, involved in the regulation of calcium release mediated by ryanodine (RyRs) and inositol 1,4,5-trisphosphate (IP3Rs) receptors, respectively. While TRIC-A is mainly expressed in excitable cells, TRIC-B is ubiquitously distributed at moderate level. TRIC-B deficiency causes a dysregulation of calcium flux from the ER, which impacts on multiple collagen specific chaperones and modifying enzymatic activity, leading to a rare form of osteogenesis imperfecta (OI Type XIV). The relevance of TRIC-B on cell homeostasis and the molecular mechanism behind the disease are still unknown. Results: In this study, we exploited zebrafish to elucidate the role of TRIC-B in skeletal tissue. We demonstrated, for the first time, that tmem38a and tmem38b genes encoding Tric-a and -b, respectively are expressed at early developmental stages in zebrafish, but only the latter has a maternal expression. Two zebrafish mutants for tmem38b were generated by CRISPR/Cas9, one carrying an out of frame mutation introducing a premature stop codon (tmem38b-/- ) and one with an in frame deletion that removes the highly conserved KEV domain (tmem38bΔ120-7/Δ120-7 ). In both models collagen type I is under-modified and partially intracellularly retained in the endoplasmic reticulum, as described in individuals affected by OI type XIV. Tmem38b-/- showed a mild skeletal phenotype at the late larval and juvenile stages of development whereas tmem38bΔ120-7/Δ120-7 bone outcome was limited to a reduced vertebral length at 21 dpf. A caudal fin regeneration study pointed towards impaired activity of osteoblasts and osteoclasts associated with mineralization impairment. Discussion: Our data support the requirement of Tric-b during early development and for bone cell differentiation.

A tapt1 knock-out zebrafish line with aberrant lens development and impaired vision models human early-onset cataract.

Bi-allelic mutations in the gene coding for human trans-membrane anterior-posterior transformation protein 1 (TAPT1) result in a broad phenotypic spectrum, ranging from syndromic disease with severe skeletal and congenital abnormalities to isolated early-onset cataract. We present here the first patient with a frameshift mutation in the TAPT1 gene, resulting in both bilateral early-onset cataract and skeletal abnormalities, in addition to several dysmorphic features, in this way further expanding the phenotypic spectrum associated with TAPT1 mutations. A tapt1a/tapt1b double knock-out (KO) zebrafish model generated by CRISPR/Cas9 gene editing revealed an early larval phenotype with eye malformations, loss of vision, increased photokinetics and hyperpigmentation, without visible skeletal involvement. Ultrastructural analysis of the eyes showed a smaller condensed lens, loss of integrity of the lens capsule with formation of a secondary lens and hyperplasia of the cells in the ganglion and inner plexiform layers of the retina. Transcriptomic analysis pointed to an impaired lens development with aberrant expression of many of the crystallin and other lens-specific genes. Furthermore, the phototransduction and visual perception pathways were found to be significantly disturbed. Differences in light perception are likely the cause of the increased dark photokinetics and generalized hyperpigmentation observed in this zebrafish model. In conclusion, this study validates TAPT1 as a new gene for early-onset cataract and sheds light on its ultrastructural and molecular characteristics.

A generated induced pluripotent stem cell (iPSC) line (CMGANTi005-A) of a Marfan syndrome patient with an FBN1 c.7754T > C (p.Ile2585Thr) variation.

Marfan syndrome (MFS) is a connective tissue disorder with pleiotropic manifestations in the ocular, skeletal and cardiovascular system; and is typically cause by pathogenic variants in the fibrillin-1 (FBN1) gene. We report a generated induced pluripotent cell (iPSC) line of a MFS patient with an FBN1 c.7754T > C (p.Ile2585Thr) variant. The cell line was generated from peripheral blood mononuclear cells (PBMCs) and after reprogramming the line showed a no relevant copy number alterations, expression of pluripotency markers and was able to differentiate into three germ layers while carrying the original genotype.

G protein-coupled receptor kinase 6 (GRK6) regulates insulin processing and secretion via effects on proinsulin conversion to insulin.

Recent studies identified a missense mutation in the gene coding for G protein-coupled receptor kinase 6 (GRK6) that segregates with type 2 diabetes (T2D). To better understand how GRK6 might be involved in T2D, we used pharmacological inhibition and genetic knockdown in the mouse ß-cell line, MIN6, to determine whether GRK6 regulates insulin dynamics. We show inhibition of GRK5 and GRK6 increased insulin secretion but reduced insulin processing while GRK6 knockdown revealed these same processing defects with reduced levels of cellular insulin. GRK6 knockdown cells also had attenuated insulin secretion but enhanced proinsulin secretion consistent with decreased processing. In support of these findings, we demonstrate GRK6 rescue experiments in knockdown cells restored insulin secretion after glucose treatment. The altered insulin profile appears to be caused by changes in the proprotein convertases, the enzymes responsible for proinsulin to insulin conversion, as GRK6 knockdown resulted in significantly reduced convertase expression and activity. To identify how the GRK6-P384S mutation found in T2D patients might affect insulin processing, we performed biochemical and cell biological assays to study the properties of the mutant. We found that while GRK6-P384S was more active than WT GRK6, it displayed a cytosolic distribution in cells compared to the normal plasma membrane localization of GRK6. Additionally, GRK6 overexpression in MIN6 cells enhanced proinsulin processing, while GRK6-P384S expression had little effect. Taken together, our data show that GRK6 regulates insulin processing and secretion in a glucose-dependent manner and provide a foundation for understanding the contribution of GRK6 to T2D.

Exploring the Mutational Landscape of Isolated Congenital Heart Defects: An Exome Sequencing Study Using Cardiac DNA.

Congenital heart defects (CHD) are the most common congenital anomalies in liveborn children. In contrast to syndromic CHD (SCHD), the genetic basis of isolated CHD (ICHD) is complex, and the underlying pathogenic mechanisms appear intricate and are incompletely understood. Next to rare Mendelian conditions, somatic mosaicism or a complex multifactorial genetic architecture are assumed for most ICHD. We performed exome sequencing (ES) in 73 parent-offspring ICHD trios using proband DNA extracted from cardiac tissue. We identified six germline de novo variants and 625 germline rare inherited variants with 'damaging' in silico predictions in cardiac-relevant genes expressed in the developing human heart. There were no CHD-relevant somatic variants. Transmission disequilibrium testing (TDT) and association testing (AT) yielded no statistically significant results, except for the AT of missense variants in cilia genes. Somatic mutations are not a common cause of ICHD. Rare de novo and inherited protein-damaging variants may contribute to ICHD, possibly as part of an oligogenic or polygenic disease model. TDT and AT failed to provide informative results, likely due to the lack of power, but provided a framework for future studies in larger cohorts. Overall, the diagnostic value of ES on cardiac tissue is limited in individual ICHD cases.

Serum Calcification Propensity T50 Associates with Disease Severity in Patients with Pseudoxanthoma Elasticum.

Pseudoxanthoma elasticum (PXE) is a currently intractable genetic disorder characterized by progressive ectopic calcification in the skin, eyes and arteries. Therapeutic trials in PXE are severely hampered by the lack of reliable biomarkers. Serum calcification propensity T50 is a blood test measuring the functional anticalcifying buffer capacity of serum. Here, we evaluated T50 in PXE patients aiming to investigate its determinants and suitability as a potential biomarker for disease severity. Fifty-seven PXE patients were included in this cross-sectional study, and demographic, clinical, imaging and biochemical data were collected from medical health records. PXE severity was assessed using Phenodex scores. T50 was measured using a validated, nephelometry-based assay. Multivariate models were then created to investigate T50 determinants and associations with disease severity. In short, the mean age of patients was 45.2 years, 68.4% was female and mean serum T50 was 347 min. Multivariate regression analysis identified serum fetuin-A (p < 0.001), phosphorus (p = 0.007) and magnesium levels (p = 0.034) as significant determinants of T50, while no correlations were identified with serum calcium, eGFR, plasma PPi levels or the ABCC6 genotype. After correction for covariates, T50 was found to be an independent determinant of ocular (p = 0.013), vascular (p = 0.013) and overall disease severity (p = 0.016) in PXE. To conclude, shorter serum T50­indicative of a higher calcification propensity­was associated with a more severe phenotype in PXE patients. This study indicates, for the first time, that serum T50 might be a clinically relevant biomarker in PXE and may thus be of importance to future therapeutic trials.

Hypergastrinemia, a clue leading to the identification of an atypical form of diabetes mellitus type 2.

BACKGROUND: A hitherto undescribed form of diabetes mellitus type 2 is reported in a Flemish family. In these patients, markedly elevated gastrin levels were observed, which could not be linked to gastrointestinal symptoms. MATERIALS AND METHODS: Gel permeation chromatography was performed for gastrin, insulin, and proinsulin. Proprotein convertase subtilisin/kexin type (PCSK1 and PCSK2)] were sequenced. Whole-exome sequencing was performed on the genomic DNA extracted from leukocytes of the proband of the family. RESULTS: Gel permeation chromatography revealed that the apparent hypergastrinemia was caused by the accumulation of biologically inactive progastrin. Besides, high serum concentrations of proinsulin and intact fibroblast growth factor 23 (FGF23) were also detected. Sequencing of PCSK1 and PCSK2 genes did not reveal any mutations in these genes. Whole exome sequencing revealed a c.1150C > T (p.Pro384Ser) mutation in G protein-coupled receptor kinase 6 (GRK6), which cosegregated with the disease. Expression of the mutant enzyme in mammalian cells revealed that it was mislocalized compared to the wild-type GRK6. CONCLUSIONS: In the affected patients, prohormone processing is impaired likely due to the altered function of mutant GRK6. Delayed pro-insulin processing causes hypoglycaemia episodes a couple of hours following meals. In addition, increased plasma concentrations of progastrin and intact FGF23 in the affected individuals can be explained by incomplete processing of the precursor hormones.

Shortcutting the diagnostic odyssey: the multidisciplinary Program for Undiagnosed Rare Diseases in adults (UD-PrOZA).

BACKGROUND: In order to facilitate the diagnostic process for adult patients suffering from a rare disease, the Undiagnosed Disease Program (UD-PrOZA) was founded in 2015 at the Ghent University Hospital in Belgium. In this study we report the five-year results of our multidisciplinary approach in rare disease diagnostics. METHODS: Patients referred by a healthcare provider, in which an underlying rare disease is likely, qualify for a UD-PrOZA evaluation. UD-PrOZA uses a multidisciplinary clinical approach combined with state-of-the-art genomic technologies in close collaboration with research facilities to diagnose patients. RESULTS: Between 2015 and 2020, 692 patients (94% adults) were referred of which 329 (48%) were accepted for evaluation. In 18% (60 of 329) of the cases a definite diagnosis was made. 88% (53 of 60) of the established diagnoses had a genetic origin. 65% (39 of 60) of the genetic diagnoses were made through whole exome sequencing (WES). The mean time interval between symptom-onset and diagnosis was 19 years. Key observations included novel genotype-phenotype correlations, new variants in known disease genes and the identification of three new disease genes. In 13% (7 of 53), identifying the molecular cause was associated with therapeutic recommendations and in 88% (53 of 60), gene specific genetic counseling was made possible. Actionable secondary findings were reported in 7% (12 of 177) of the patients in which WES was performed. CONCLUSION: UD-PrOZA offers an innovative interdisciplinary platform to diagnose rare diseases in adults with previously unexplained medical problems and to facilitate translational research.

Human germline nuclear transfer to overcome mitochondrial disease and failed fertilization after ICSI.

PURPOSE: Providing additional insights on the efficacy of human nuclear transfer (NT). Here, and earlier, NT has been applied to minimize transmission risk of mitochondrial DNA (mtDNA) diseases. NT has also been proposed for treating infertility, but it is still unclear which infertility indications would benefit. In this work, we therefore additionally assess the applicability of NT to overcome failed fertilization. METHODS: Patient 1 carries a homoplasmic mtDNA mutation (m.11778G > A). Seventeen metaphase II (MII) oocytes underwent pre-implantation genetic testing (PGT), while five MII oocytes were used for spindle transfer (ST), and one in vitro matured (IVM) metaphase I oocyte underwent early pronuclear transfer (ePNT). Patients 2-3 experienced multiple failed intracytoplasmic sperm injection (ICSI) and ICSI-assisted oocyte activation (AOA) cycles. For these patients, the obtained MII oocytes underwent an additional ICSI-AOA cycle, while the IVM oocytes were subjected to ST. RESULTS: For patient 1, PGT-M confirmed mutation loads close to 100%. All ST-reconstructed oocytes fertilized and cleaved, of which one progressed to the blastocyst stage. The reconstructed ePNT-zygote reached the morula stage. These samples showed an average mtDNA carry-over rate of 2.9% ± 0.8%, confirming the feasibility of NT to reduce mtDNA transmission. For patient 2-3 displaying fertilization failure, ST resulted in, respectively, 4/5 and 6/6 fertilized oocytes, providing evidence, for the first time, that NT can enable successful fertilization in this patient population. CONCLUSION: Our study showcases the repertoire of disorders for which NT can be beneficial, to overcome either mitochondrial disease transmission or failed fertilization after ICSI-AOA.

Negative Molecular Diagnostics in Non-Syndromic Hearing Loss: What Next?

Congenital hearing loss has an impact on almost every facet of life. In more than 50% of cases, a genetic cause can be identified. Currently, extensive genetic testing is available, although the etiology of some patients with obvious familial hearing loss remains unknown. We selected a cohort of mutation-negative patients to optimize the diagnostic yield for genetic hearing impairment. In this retrospective study, 21 patients (17 families) with negative molecular diagnostics for non-syndromic hearing loss (gene panel analysis) were included based on a positive family history with a similar type of hearing loss. Additional genetic testing was performed using a whole exome sequencing panel (WESHL panel v2.0) in four families with the strongest likelihood of genetic hearing impairment. In this cohort (n = 21), the severity of hearing loss was most commonly moderate (52%). Additional genetic testing revealed pathogenic copy number variants in the STRC gene in two families. In summary, regular re-evaluation of hearing loss patients with presumably genetic etiology after negative molecular diagnostics is recommended, as we might miss newly discovered deafness genes. The switch from gene panel analysis to whole exome sequencing or whole genome sequencing for the testing of congenital hearing loss seems promising.

Minocycline Attenuates Excessive DNA Damage Response and Reduces Ectopic Calcification in Pseudoxanthoma Elasticum.

Pseudoxanthoma elasticum (PXE) is a hereditary ectopic calcification disorder affecting the skin, eyes, and blood vessels. Recently, the DNA damage response (DDR), in particular PARP1, was shown to be involved in aberrant mineralization, raising the hypothesis that excessive DDR/PARP1 signaling also contributes to PXE pathogenesis. Using dermal fibroblasts of patients with PXE and healthy controls, (lesional) skin tissue, and abcc6a‒/‒ zebrafish, we performed expression analysis of DDR/PARP1 targets with QRT-PCR, western blot, immunohistochemistry, and enzyme activity assays before and after treatment with the PARP1 inhibitor minocycline. PARP1 and the ATM‒p21‒p53 axis was found to be significantly increased in PXE. In addition, PARP1 downstream targets IL-6, signal transducer and activator of transcription 1/3, TET1, and RUNX2 were upregulated, whereas the RUNX2 antagonist microRNA-204 was decreased. In PXE fibroblasts, DDR/PARP1 signaling increased with advancing ectopic calcification. Minocycline treatment attenuated DDR/PARP1 overexpression and reduced aberrant mineralization in PXE fibroblasts and abcc6a‒/‒ zebrafish. In summary, we showed the involvement of excessive DDR/PARP1 signaling in PXE pathophysiology, identifying a signal transducer and activator of transcription‒driven cascade resulting in increased expression of the epigenetic modifier TET1 and procalcifying transcription factor RUNX2. Minocycline attenuated this deleterious molecular mechanism and reduced ectopic calcification both in vitro and in vivo, fueling the exciting prospect of a therapeutic compound for PXE.

Clinical and subclinical findings in heterozygous ABCC6 carriers: results from a Belgian cohort and clinical practice guidelines.

BACKGROUND: Biallelic pathogenic variants in the ATP-binding cassette subfamily C member 6 (ABCC6) gene cause pseudoxanthoma elasticum, a multisystemic ectopic calcification disorder, while heterozygous ABCC6 variants are associated with an increased risk of cardiovascular and cerebrovascular disease. As the prevalence of pathogenic ABCC6 variants in the general population is estimated at ~1%, identifying additional ABCC6-related (sub)clinical manifestations in heterozygous carriers is of the utmost importance to reduce this burden of disease. Here, we present a large Belgian cohort of heterozygous ABCC6 carriers with comprehensive clinical, biochemical and imaging data. Based on these results, we formulate clinical practice guidelines regarding screening, preventive measures and follow-up of ABCC6 carriers. METHODS: The phenotype of 56 individuals carrying heterozygous pathogenic ABCC6 variants was assessed using clinical (eg, detailed ophthalmological examinations), biochemical, imaging (eg, cardiovascular and abdominal ultrasound) and genetic data. Clinical practice guidelines were then drawn up. RESULTS: We found that ABCC6 heterozygosity is associated with distinct retinal alterations ('comet-like') (24%), high prevalence of hypercholesterolaemia (>75%) and diastolic dysfunction (33%), accelerated lower limb atherosclerosis and medial vascular disease, abdominal organ calcification (26%) and testicular microlithiasis (28%), though with highly variable expression. CONCLUSION: In this study, we delineated the multisystemic ABCC6 heterozygosity phenotype characterised by retinal alterations, aberrant lipid metabolism, diastolic dysfunction and increased vascular, abdominal and testicular calcifications. Our clinical practice guidelines aimed to improve early diagnosis, treatment and follow-up of ABCC6-related health problems.