Screening For Congenital Adrenal Hyperplasia Research Articles

Can incorporating molecular testing improve the accuracy of newborn screening for congenital adrenal hyperplasia?

Single-tier newborn screening (NBS) for CAH using 17-hydroxyprogesterone (17OHP) measured by fluoroimmunoassay (FIA) in samples collected at 24-48 hours produces a high false-positive rate (FPR). 2nd tier steroid testing can reduce the FPR and has been widely implemented. We investigated the accuracy of an alternative multi-tier CAH NBS protocol that incorporates molecular testing of the CYP21A2 gene and reduces the 1st tier 17OHP cutoff to minimize missed cases. Created a Minnesota-specific CYP21A2 pathogenic variants panel; develop a rapid, high-throughput multiplex, allele-specific-primer-extension assay; perform 1-year retrospective analysis of Minnesota NBS results comparing metrics between a conventional steroid-based two-tier protocol and a molecular-based multi-tier NBS protocol, applied post-hoc. CYP21A2 gene sequencing of 103 Minnesota families resulted in a Minnesota-specific panel of 21 pathogenic variants. Centers for Disease Control and Prevention (CDC) created a molecular assay with 100% accuracy and reproducibility. Two-tier steroid-based screening of 68,659 live births during 2015 resulted in 2 false negatives (FNs), 91 FPs, and 1 true positive (TP). A three-tier protocol with a lower 1st-tier steroid cutoff, 2nd-tier 21-variant CYP21A2 panel and 3rd-tier CYP21A2 sequencing would have resulted in 0 FNs, 52 FPs and 3 TPs. Incorporation of molecular testing could improve the accuracy of CAH NBS, although some distinct challenges of molecular testing may need to be considered before implementation by NBS programs.

Inclusion of 21-deoxycortisol in mass spectrometry testing improves newborn screening for congenital adrenal hyperplasia

Inclusion of 21-deoxycortisol in mass spectrometry testing improves newborn screening for congenital adrenal hyperplasia

Service evaluation suggests variation in clinical care provision in adults with congenital adrenal hyperplasia in the UK and Ireland.

Congenital adrenal hyperplasia (CAH) encompasses a rare group of autosomal recessive disorders, characterised by enzymatic defects in steroidogenesis. Heterogeneity in management practices has been observed internationally. The International Congenital Adrenal Hyperplasia registry (I-CAH, https://sdmregistries.org/) was established to enable insights into CAH management and outcomes, yet its global adoption by endocrine centres remains unclear. We sought (1) to assess current practices amongst clinicians managing patients with CAH in the United Kingdom and Ireland, with a focus on choice of glucocorticoid, monitoring practices and screening for associated co-morbidities, and (2) to assess use of the I-CAH registry. We designed and distributed an anonymised online survey disseminated to members of the Society for Endocrinology and Irish Endocrine Society to capture management practices in the care of patients with CAH. Marked variability was found in CAH management, with differences between general endocrinology and subspecialist settings, particularly in glucocorticoid use, biochemical monitoring and comorbidity screening, with significant disparities in reproductive health monitoring, notably in testicular adrenal rest tumours (TARTs) screening (p = .002), sperm banking (p = .0004) and partner testing for CAH (p < .0001). Adoption of the I-CAH registry was universally low. Differences in current management of CAH continue to exist. It appears crucial to objectify if different approaches result in different long-term outcomes. New studies such as CaHASE2, incorporating standardised minimum datasets including replacement therapies and monitoring strategies as well as longitudinal data collection, are now needed to define best-practice and standardise care.

Association of newborn screening of congenital adrenal hyperplasia with outcomes in the first 90 days of life: a multi-centre I-CAH registry analysis of contemporary practice

Association of newborn screening of congenital adrenal hyperplasia with outcomes in the first 90 days of life: a multi-centre I-CAH registry analysis of contemporary practice

The use of liquid chromatography-tandem mass spectrometry in newborn screening for congenital adrenal hyperplasia: improvements and future perspectives.

Newborn screening for congenital adrenal hyperplasia using 17-hydroxyprogesterone by immunoassay remains controversial despite screening been available for almost 40 years. Screening is confounded by poor immunoassay specificity, fetal adrenal physiology, stress, and illness which can result in a large number of false positive screening tests. Screening programmes apply higher screening thresholds based on co-variates such as birthweight or gestational age but the false positive rate using immunoassay remains high. Mass spectrometry was first applied to newborn screening for congenital adrenal hyperplasia over 15 years ago. Elevated 17-hydroxprogesterone by immunoassay can be retested with a specific liquid chromatography tandem mass spectrometry assay that may include additional steroid markers. Laboratories register with quality assurance programme providers to ensure accurate steroid measurements. This has led to improvements in screening but there are additional costs and added laboratory workload. The search for novel steroid markers may inform further improvements to screening. Studies have shown that 11-oxygenated androgens are elevated in untreated patients and that the adrenal steroidogenesis backdoor pathway is more active in babies with congenital adrenal hyperplasia. There is continual interest in 21-deoxycortisol, a specific marker of 21-hydroxylase deficiency. The measurement of androgenic steroids and their precursors by liquid chromatography tandem mass spectrometry in bloodspots may inform improvements for screening, diagnosis, and treatment monitoring. In this review, we describe how liquid chromatography tandem mass spectrometry has improved newborn screening for congenital adrenal hyperplasia and explore how future developments may inform further improvements to screening and diagnosis.

Differences in Hyperandrogenism Related to Early Detection of Non-Classical Congenital Adrenal Hyperplasia on Second Newborn Screen.

Screening for congenital adrenal hyperplasia (CAH) remains heterogenous across geographies-we sought to determine the proportion of non-classical CAH (NCAH) detection by one vs. two newborn screens (NBS) in two U.S. regions. Data were collected at tertiary centers in Houston (HOU) and Los Angeles (LA) on 35 patients with NCAH, comparing patients identified via the NBS vs. during childhood, 17-hydroxyprogesterone (17-OHP) levels, genotype, and phenotype. The NBS filter-paper 17-OHP levels and daily cutoffs were recorded on initial and second screens. In all, 53% of patients with NCAH in the HOU cohort were identified as infants via the second NBS. Patients identified clinically later in childhood presented at a similar age (HOU: n = 9, 5.5 ± 3.1 years; LA: n = 18, 7.9 ± 4 years) with premature pubarche in almost all. Patients in LA had more virilized phenotypes involving clitoromegaly and precocious puberty and were older at treatment onset compared with those identified in HOU by the second NBS (HOU: 3.2 ± 3.9 years; LA: 7.9 ± 4.0 years, p = 0.02). We conclude that the early detection of NCAH could prevent hyperandrogenism and its adverse consequences, with half of the cases in HOU detected via a second NBS. Further studies of genotyping and costs are merited.

Long-read sequencing: An effective method for genetic analysis of CYP21A2 variation in congenital adrenal hyperplasia

BackgroundThe sequence similarity between CYP21A2 gene and its inactive pseudogene CYP21A1P, and copy number variation (CNV) caused by unequal crossover, make it challenging to characterize the CYP21A2 gene through traditional methods. This study aimed to evaluate the clinical utility of the long-read sequencing (LRS) method in carrier screening and genetic diagnosis of congenital adrenal hyperplasia (CAH) by comparing the efficiency of the LRS method with the conventional multiplex ligation-dependent probe amplification (MLPA) plus Sanger sequencing approaches in CYP21A2 analysis. MethodsIn a retrospective study, full sequence analysis of the CYP21A2 and CYP21A1P was performed for three pedigrees through long-range locus-specific PCR followed by LRS based on the Pacific Biosciences (PacBio, California, USA) single-molecule real-time (SMRT) platform, and the results were compared with those obtained from next-generation sequencing (NGS)-based whole exome sequencing (WES) and the traditional methods of MLPA plus Sanger sequencing. ResultsThe LRS method successfully identified seven CYP21A2 variants, including three single nucleotide variants (NM_000500.9:c.1451G > C p.(Arg484Pro), c.293-13A/C > G (IVS2-13A/C > G), c.518 T > A p.(Ile173Asn)), one 111-bp polynucleotide insertion, one set of 3′URT variants (NM_000500.9:c.*368 T > C, c.*390A > G, c.*440C > T, c.*443 T > C) and two types of chimeric genes and straightforwardly depicted the inheritance patterns of these variants within families. Moreover, the LRS method enabled us to determine the cis–trans configuration of multiple variants in one assay, without the need to analyze additional family samples. Compared with traditional methods, this LRS method can achieve a precise, comprehensive and intuitive result in the genetic diagnosis of 21-hydroxylase deficiency (21-OHD). ConclusionThe LRS method is comprehensive in CYP21A2 analysis and intuitive in result presentation, which holds substantial promise in clinical application as a crucial tool for carrier screening and genetic diagnosis of CAH.

Impact of Newborn Screening on Adult Height in Patients with Congenital Adrenal Hyperplasia (CAH).

Treatment of children with classical congenital adrenal hyperplasia (CAH) due to 21-hydroxylase deficiency is challenging. Linear growth and adult height are compromised according to recent publications. However, most of these data were obtained in the era before CAH newborn screening. Body height of patients with classical CAH diagnosed before and after the establishment of newborn screening were analyzed retrospectively. We identified 600 patients with classical CAH (227 male) with data on near-adult height (NAH), target height (TH), and information on newborn screening from the electronic German CAH registry (DGPAED society). Newborn screening was performed in 101 (16.8%) patients. All patients received hydrocortisone with or without fludrocortisone.To assess the effects of newborn screening, a linear regression model adjusted/stratified for sex and phenotype was used (SAS 9.4). TH corrected NAH (mean; 95% confidence interval) was closer to zero in patients with CAH and newborn screening (-0.25 SDS; -0.44 - -0.06) than in patients without newborn screening (-0.44 SDS; -0.52 - -0.36) (p=0.069). Screening had no effect on NAH in female patients. In male patients, NAH was significantly better (p=0.033) with screening than without screening. After stratifying for CAH phenotype, screening did not affect the NAH of patients with Salt-wasting-CAH. Patients with Simple-virilizing-CAH had a significantly better cNAH (p=0.034) with screening (0.15 SDS; -0.28 - 0.59) than without screening (-0.35 SDS; -0.52 - -0.18). Our data suggest that newborn screening might be associated with improved near-adult height in male CAH patients and in patients with SV-CAH.

International Newborn Screening Practices for the Early Detection of Congenital Adrenal Hyperplasia

Introduction: Newborn screening (NBS) programmes vary internationally in their approach to screening. Guidelines for congenital adrenal hyperplasia (CAH) screening recommend the use of two-tier testing and gestational age cutoffs to minimise false-positive results. The aims of this study were to describe (1) the approaches; (2) protocols used; and (3) available outcomes for CAH screening internationally. Methods: All members of the International Society for Neonatal Screening were asked to describe their CAH NBS protocols, with an emphasis on the use of second-tier testing, 17-hydroxyprogesterone (17OHP) cutoffs, and gestational age and birth weight adjustments. If available, screening outcomes were requested. Results: Representatives from 23 screening programmes provided data. Most (n = 14; 61%) recommend sampling at 48–72 h of life. Fourteen (61%) use single-tier testing and 9 have a two-tier testing protocol. Gestational age cutoffs are used in 10 programmes, birth weight cutoffs in 3, and a combination of both in 9. One programme does not use either method of adjusting 17OHP cutoffs. Case definition of a positive test and the response to a positive test differed between programmes. Conclusions: We have demonstrated significant variation across all aspects of NBS for CAH, including timing, the use of single versus two-tier testing and cutoff interpretation. Collaboration between international screening programmes and implementation of new techniques to improve screen efficacy will facilitate ongoing expansion and quality improvement in CAH NBS.

Non-classical congenital adrenal hyperplasia. Clinical case

Congenital adrenal hyperplasia (CAH) is an autosomal recessive disease emerging from mutations of genes for enzymes that lead to the biochemical shifts in the production of glucocorticoids, mineralocorticoids, or sex steroids from cholesterol by the adrenal glands. Universal newborn screening for CAH is recommended for early diagnosis and initiation of therapy. The development of CAH is due to a defect in the CYP21 gene, which encodes 21-hydroxylase enzyme involved in the synthesis of cortisol. This leads to an increase in the secretion of adrenocorticotropic hormone and the accumulation of cortisol precursors, which are converted into adrenal androgens— the classical form of the disease develops. With a point mutation of the CYP21 gene, an incomplete defect occurs in 21-hydroxylase, which leads to an unpronounced disorder of adrenal steroidogenesis— a non-classical form of congenital adrenal hyperplasia, which happens more often. In this form, the clinical symptoms are erased with moderate hirsutism, acne vulgaris, infertility. In comparison to the classical form of the disease, which is diagnosed at birth or during the neonatal period because of ambiguous genitalia and/or salt-wasting symptoms or through screening programs used in some countries, most cases of non-classical CAH are not easy to detect. Additionally, many individuals remain asymptomatic during childhood and adolescence, have normal reproductive function, and only become aware of non-classical CAH due to the diagnosis of another family member and consequent testing. However, most women with non-classical CAH seek medical assistance when they experience symptoms of androgen excess and, when clinical suspicion prompts testing, elevated basal 17-OH progesterone levels may primarily point to the diagnosis of non-classical CAH. A case of a non-classical form of the disease which manifested itself in infertility is given. Pregnancy occurred after 4 months treatment with prednisolone (5 mg/day).

Establishing 17-Hydroxyprogesterone Cutoff Values for Congenital Adrenal Hyperplasia in Preterm, Low Birth Weight, and Sick Newborns.

Newborn screening for congenital adrenal hyperplasia (CAH) has benefits with a high adoption rate worldwide. It also has problems of high false positives, which can cause stress to the patient's family with economic losses and unnecessary visits of newborns to hospitals. Therefore, we investigated the influence of birth weight (BW), gestational age (GA), and GA with sampling time on 17-hydroxyprogesterone (17-OHP) concentration and attempted to establish the 17-OHP cutoff values in preterm, low birth weight (LBW), and sick newborns. Newborns (n=1,071) born between October 2020 and January 2022 were screened for CAH. Samples from neonates were collected on filter paper with the heel prick method. 17-OHP concentration was measured by time-resolved immunofluorescence with an AutoDELFIA Neonatal 17-hydroxyprogesteron kit and grouped in relation to BW, GA, and GA with sampling time. The median age of newborns at neonatal sample collection was 6 days. 17-OHP concentration showed a statistically significant negative correlation with BW (r=-0.488, p<0.001) and GA (r=-0.560, p<0.001). Full-term and preterm subgroups had a similar decreasing tendency of 17-OHP concentration with increasing sampling time. Application of newly establishing cutoff criteria significantly reduced recall rates to 1.16%, 0.9%, and 1.75% according to each criterion of BW, GA, and GA with sampling time, respectively. This study presents new 17-OHP cutoff values for preterm, LBW, and sick newborns. These data in our laboratory can be used as a reference by other laboratories for establishing new cutoff criteria to help lower the high recall rate and reduce unnecessary follow-up tests.

Neonatal Screening for Congenital Adrenal Hyperplasia in Indian Newborns with Reflex Genetic Analysis of 21-Hydroxylase Deficiency

Congenital adrenal hyperplasia (CAH), screened for in neonates, is the second most common endocrinopathy after congenital hypothyroidism.Newborn screening for CAH due to CYP21A2 deficiency is performed by immunologic assay for 17-hydroxyprogesterone (17-OHP). The second-tier test for confirmation of diagnosis is carried out on recall venous blood sample from screen positives measuring 17-OHP, or other metabolites of steroid metabolism by liquid chromatography-tandem mass spectroscopy. However, as steroid metabolism is dynamic, it can affect these parameters even in the recall sample of a stressed neonate. Moreover, there is some time delay in recalling the neonate for repeat testing. Reflex genetic analysis of blood spot from the initial Guthrie cards of screen positive neonates, if used for confirmatory testing, can avoid this time delay as well as the effect of stress on steroid metabolism. In this study, we used Sanger sequencing and MLPA in a reflex manner for molecular genetic analysis to confirm CYP21A2-mediated CAH. Out of 220,000 newborns screened, 97 were positive on the initial biochemical screen, of which 54 were confirmed true positives with genetic reflex testing, giving incidence of CAH as 1:4074. Point mutations were more common than deletions, indicating that Sanger sequencing should be used ahead of MLPA for molecular diagnosis in India. Amongst the variants detected, the most common was I2G-Splice variant (44.5%), followed by c.955C>T (p.Gln319Ter) (21.2%); Del 8 bp and c.-113G>A were detected with frequencies of 20.3% and 20%, respectively. In conclusion, reflex genetic testing is an effective strategy for identifying true positives in CAH screening in neonates. This will obviate need for recall samples and also aid effective counselling and timely prenatal diagnosis in the future. In Indian newborns, as point mutations are more common than large deletions, Sanger sequencing should be the initial method of choice for genotyping, ahead of MLPA.

Newborn screening for congenital adrenal hyperplasia

This presentation discusses newborn screening for congenital adrenal hyperplasia (CAH) which is available in most developed countries. Early detection and hormone replacement prevents the progression of salt-wasting adrenal crises that can become life-threatening in the first two weeks of life. Screening by 17-hydroxyprogesterone immunoassay is sensitive for salt-wasting disease but confounded by poor specificity in premature and ill neonates. The use of liquid chromatography tandem mass spectrometry as a second-tier test offers improvements to screening through improved analytical specificity which reduces the number of false positive tests.

Evaluation of a New Laboratory Protocol for Newborn Screening for Congenital Adrenal Hyperplasia in New Zealand.

Between 2005 and 2021, 49 cases of classical congenital adrenal hyperplasia were diagnosed in New Zealand, 39 were detected in newborns and 10 were not detected by screening. For every case of CAH detected by screening, 10 false-positive tests are encountered. Second-tier liquid chromatography-tandem mass spectrometry (LCMSMS) has the potential to improve screening sensitivity and specificity. A new laboratory protocol for newborn screening for CAH was evaluated. Birthweight-adjusted thresholds for first- and second-tier 17-hydroxyprogesterone, second-tier 21-deoxyicortisol and a steroid ratio were applied to 4 years of newborn screening data. The study was enriched with 35 newborn screening specimens from confirmed CAH cases. Newborn screening was conducted on 232,542 babies, and 11 cases of classical CAH were detected between 2018 and 2021. There were 98 false-positive tests (specificity 99.96%, PPV = 10.1%) using the existing protocol. Applying the new protocol, the same 11 cases were detected, and there were 13 false-positive tests (sensitivity > 99.99%, PPV = 45.8%, (X2 test p < 0.0001). Incorporating the retrospective specimens screening sensitivity for classical CAH was 78% (existing protocol), compared to 87% for the new protocol (X2 test p = 0.1338). Implementation of LCMSMS as a second-tier test will improve newborn screening for classical CAH in New Zealand.

Diagnosis and Management of Adrenal Crisis in 46XX Congenital Adrenal Hyperplasia Infant

Highlight: The diagnosis and therapy of Congenital Adrenal Hyperplasia (CAH) children with Adrenal crisis (AC) case report. Adrenal crisis (AC) is a life-threatening emergency that contributes to the high death rate of children with adrenal insufficiency. The early detection and prompt treatment can improve the outcomes of patients with CAH and AC. Abstract: Adrenal crisis is the acute complication of the patient with congenital adrenal hyperplasia. Congenital adrenal hyperplasia (CAH) is a rare condition. Children with CAH commonly come to the emergency room due to acute complications. The condition has high mortality and thus needs early recognition. Newborn screening for CAH in Indonesia is not routinely performed and has not been suggested yet. The purpose of this case report was to report a case of adrenal crisis in a congenital adrenal hyperplasia patient focused on diagnosis and therapy. A female, 10 months old infant, was admitted to the emergency department with a chief complaint of a decrease of consciousness for 3 hours before admission and frequent vomiting since born. On physical examination, there was clitoromegaly. Laboratory showed 17-OH progesterone: 173 ng/dL (7-77 ng/dL) and karyotyping: 46 XX. Management of adrenal crisis is a stress dose of hydrocortisone and rehydration. Education is the key to optimal outcomes and normal growth and development.

Ethnic and National Differences in Congenital Adrenal Hyperplasia Incidence: A Systematic Review and Meta-Analysis

Background: Congenital adrenal hyperplasia (CAH) is an autosomal recessive genetic disorder that causes defects in the adrenal cortex enzymes that impair the biosynthesis of cortisol, aldosterone, or both. The most common type is the 21-hydroxylase enzyme deficiency in approximately 95% of cases resulting from CYP21A2 gene mutations or deletions. Objectives: This study aimed to systematically review the national differences in CAH incidence and analyze the pooled results to determine disparities and whether ethnicity can predispose people to develop CAH. Methods: PubMed, Scopus, and LILACS were used to achieve results until June 22, 2018. Study eligibility criteria included availability of full-text; English, Spanish, or Portuguese languages; incidence or number of new cases; and number of live births or sample population. Only the classic CAH type (salt-wasting and simple-virilizing) was considered, and no distinction was made between the enzyme deficiency types. Results: This study summarizes the findings of 58 studies and 31 countries (from 1969 to 2017), in which the overall CAH incidence was 1:9,498 (95% confidence interval: 1:9,089, 1:9,945). Countries from the Eastern Mediterranean and Southeast Asia revealed the highest CAH incidence. The lowest incidence was reported in countries of the Western Pacific of Asia. No remarkable difference was observed in the Hispanics/Latino and White groups. However, they manifested a higher incidence of CAH than people identified as Black or of African descent. Published studies on CAH incidence in the sub-Saharan African region and parts of Europe were insufficient. Conclusions: This study highlights the at-risk population for CAH and regions that need monitoring for CAH. The highest CAH incidence could be attributed to higher consanguinity, less genetic diversity, or other genetic causes since CAH is an inherited genetic disorder. Cultural practices in some places regarding consanguineous unions or geographic isolation may directly affect the incidence. Newborn screening for CAH may be unavailable in many developing countries, thereby affecting the actual CAH incidence. Therefore, healthcare workers should be trained to recognize CAH at an early stage to reduce its complications and mortality.

Simultaneous quantitation of 17 endogenous adrenal corticosteroid hormones in human plasma by UHPLC-MS/MS and their application in congenital adrenal hyperplasia screening.

Accurate investigation of adrenal hormone levels plays a vital role in pediatric endocrinology for the detection of steroid-related disorders. This study aims to develop a straightforward, sensitive UHPLC-MS/MS method to quantify 17 endogenous adrenal corticosteroid hormones in human plasma. These hormones are the main ingredients in the synthetic and metabolic pathways of adrenal corticosteroid hormones. Chromatographic separation was achieved on a C18 column before electrospray ionization triple-quadrupole mass spectrometry in multiple reaction monitoring mode with a run time of 7 min. The samples were extracted by liquid-liquid extraction and required no derivatization. Analytical performance was evaluated, including linearity, analytical sensitivity, accuracy, precision, and specificity. Plasma specimens from 32 congenital adrenal hyperplasia (CAH) patients and 30 healthy volunteers were analyzed to further reveal the diagnostic value of multiple steroid hormones in the synthetic and metabolic pathways of adrenal corticosteroid in CAH diagnosis. All hormones were effectively extracted and separated using our method. The method was essentially free from potential interference of isomers or structural analogues. The imprecisions were <10%. The lower limits of quantification varied from 0.05 to 15.0 ng/ml. Good linearity coefficients (r 2 > 0.998) were also obtained for most hormones in the required concentration range, except for 21-deoxycortisol (r 2 = 0.9967) and androstenediol (r 2 = 0.9952). The recoveries for the steroid hormones ranged from 91.7 to 109.8%. We developed the UHPLC-MS/MS method for the simultaneous measurement of steroid hormones. The results showed that measurement of steroid hormones simultaneously could improve the diagnostic efficiency of CAH.

Steroid profile in dried blood spots by liquid chromatography tandem mass spectrometry: Application to newborn screening for congenital adrenal hyperplasia in China

BackgroundNewborn screening for congenital adrenal hyperplasia (CAH) using 17-hydroxyprogesterone dissociation-enhanced, lanthanide fluorescence immunoassay (DELFIA) generates a large number of false-positive results. The present study aimed to improve the sensitivity of the CAH neonatal screening by including second-tier steroid profiling in dried blood spots (DBS) using liquid chromatography tandem mass spectrometry (LC-MS/MS). MethodsWe developed and validated a LC-MS/MS method for simultaneous determination of six steroids in DBS, including androstenedione, testosterone, 17-hydroxyprogesterone, 11-deoxycortisol, 21-deoxycortisol, and cortisol. Two 5-mm blood spots were eluted by internal standard working solution. We analyzed 1170 DBS samples from neonates to determine gestational age-specific reference intervals. In order to test the specificity of the second-tier method, we analyzed 707 cards with a positive screening by DELFIA. ResultsValues of intra- and inter-day precision coefficients of variance and accuracy were 2.0%–13.3% and 85.8%–114.5%, respectively. Recovery ranged from 85.0% to 106.9%. The lower limit of quantification was 0.5 ng/mL for 21-deoxycortisol, 0.25 ng/mL for 17-hydroxyprogesterone and cortisol, and 0.1 ng/mL for testosterone, androstenedione, and 11-deoxycortisol. In addition, the linearity range was 0.25–50 ng/mL (R2 > 0.99). According to the 17-hydroxyprogesterone levels and ratios of (androstenedione + 17-hydroxyprogesterone)/cortisol in the 707 positive screening samples, 77 neonates should receive recall visit. The number of false-positive results reduced by 89.1%. Totally, 18 newborns were diagnosed with 21-hydroxylase deficiency, one with P450 oxidoreductase deficiency and one with 11β-hydroxylase deficiency. With two-tier screening, the positive predictive value increased to 26.0%. ConclusionsThe second-tier steroid profiling by LC-MS/MS reduced the false-positive rate and improved the positive predictive value of CAH screening. We suggest applying this steroid profiling assay as a second-tier test for CAH screening in China.

Twenty years of newborn screening for congenital adrenal hyperplasia and congenital primary hypothyroidism – experiences from the DGKED/AQUAPE study group for quality improvement in Germany

Abstract Congenital primary hypothyroidism (CH) and congenital adrenal hyperplasia (CAH) are targeted by the German and Austrian newborn screening. For both diseases, there are registries for quality improvement, based on standardized observational data from long-term patient follow-up, under the auspices of the DGKED study group. By September 2021, the CH registry HypoDOK includes datasets from 23,348 visits of 1,840 patients, and the CAH registry contains datasets from 36,237 visits of 1,976 patients. Here, we report on the recruitment process, patient characteristics, and research contributions from the registries, and underline that the registries are an important tool to improve patient care and outcomes. Registries for rare conditions should thus be considered as an important public health measure and they should be adequately institutionalized and funded.

Analysis of a pitfall in congenital adrenal hyperplasia newborn screening: evidence of maternal use of corticoids detected on dried blood spot.

Neonatal screening for congenital adrenal hyperplasia (CAH) faces many specific challenges. It must be done using a performant analytical approach that combines sensitivity and specificity to capture the potential causes of mortality during the first week of life, such as salt wasting and glucocorticoid deficiency. Here, we confirm that maternal inhaled corticosteroid intake during pregnancy is a possible cause of missed CAH diagnosis. Thanks to liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) analysis, we were able to quantify endogenous steroid metabolites and also detect the presence of exogenous steroids in the dried blood spot of a newborn. Adding LC-MS/MS analysis as second-tier test, especially one that includes both 17-hydroxyprogesterone and 21-deoxycortisol measurements, would probably improve CAH diagnosis. In familial neonatal screening one could also look for maternal corticosteroid therapies that are hidden to prevent false-negative tests.