Non-classical Phenotype Research Articles

Resolution of the Expert Council on the problem of early diagnosis of Alzheimer's disease

Alzheimer's disease (AD) is the most common neurodegenerative disease and the most common cause of dementia. In daily practice, AD is often diagnosed late, while the early stages of the disease are overlooked or mistaken for cerebrovascular pathology. However, the efficacy of existing and newly developed (disease-modifying) AD therapies is the greatest in the early stages of the disease. An accurate diagnosis of AD is possible when biological markers of the main pathological process (cerebral amyloidosis, tauopathy) are detected using positron emission tomography or neurochemical examination of cerebrospinal fluid, which are gradually being introduced into practice in Russia. The experts discussed the clinical aspects of the use of biological markers, obtained in the leading specialized centers of our country for the diagnosis and treatment of cognitive impairment (CI). First and foremost, biomarker testing is indicated in patients with mild CI and mild dementia possibly associated with AD, so that disease-modifying (pathogenetic) therapy can be initiated as early as possible upon on its availability (currently, drugs for anti-amyloid disease-modifying therapy are not registered in the Russian Federation). Patients with a non-classical (non-amnestic) or atypical AD phenotype are another group of patients in whom it is also advisable to analyze biomarkers for differential diagnostic purposes.

Celiac Disease: A Forty-Year Analysis in an Italian Referral Center.

Celiac disease (CD) is an autoimmune disorder triggered by gluten ingestion. Herein, we assessed clinical, serological and histopathological findings of a single-center, large cohort of CD patients diagnosed and followed-up over forty years. From January 1980 to December 2020, 1547 CD patients (1170 females; age range: 8-81 years; F:M ratio = 3.1:1) were diagnosed in an Italian tertiary referral center. Comorbidities and complications were recorded at diagnosis and during follow-up. CD diagnoses quadrupled after 2000. The most frequent phenotype was the non-classical CD (63.3%), and the most prevalent histotype was Marsh 3C (44.7%). Gastrointestinal manifestations, detectable in 51% of patients, were diarrhea (24.3%), bloating (28%) and aphthous stomatitis (19.7%). The most common CD-associated disorder was osteopenia (59.9%), predominant in females (64.3%); extraintestinal manifestations included anemia (35.8% iron-deficiency; 87% folic acid malabsorption), cryptogenic hypertransaminasemia (27.9%), and recurrent miscarriages (11.5%). Thyroiditis (26.9%), type 1 diabetes mellitus (2.9%), and dermatitis herpetiformis (1.4%) were the most common CD-related autoimmune disorders. Six patients had inflammatory bowel disease. Complications and mortality rate occurred in 1.8% and 1.9%, respectively. This single-center, large cohort analysis confirmed that CD presentation changed over the years, with an increase of non-classical and subclinical clinical phenotypes.

Cornelia de Lange Spectrum

Cornelia de Lange syndrome (CdLS) is a rare congenital developmental disorder with multisystemic involvement. The clinical presentation is highly variable, but the classic phenotype, characterized by distinctive craniofacial features, pre- and postnatal growth retardation, extremity reduction defects, hirsutism and intellectual disability can be distinguished from the nonclassic phenotype, which is generally milder and more difficult to diagnose. In addition, the clinical features overlap with those of other neurodevelopmental disorders, so the use of consensus clinical criteria and artificial intelligence tools may be helpful in confirming the diagnosis.Pathogenic variants in NIPBL, which encodes a protein related to the cohesin complex, have been identified in more than 60% of patients, and pathogenic variants in other genes related to this complex in another 15%: SMC1A, SMC3, RAD21, and HDAC8. Technical advances in large-scale sequencing have allowed the description of additional genes (BRD4, ANKRD11, MAU2), but the lack of molecular diagnosis in 15% of individuals and the substantial clinical heterogeneity of the syndrome suggest that other genes and mechanisms may be involved.Although there is no curative treatment, there are symptomatic/palliative treatments that paediatricians should be aware of. The main medical complication in classic SCdL is gastro-esophageal reflux (GER), which should be treated early.

Cornelia de Lange syndrome type 4 with a novel heterozygous missense variant in RAD 21 gene

Cornelia de Lange syndrome (CdLS) is a congenital multisystemic disorder characterized by genetic heterogeneity. It presents with features such as growth and cognitive retardation, upper limb deformities, cardiac, ophthalmologic, and genitourinary anomalies, alongside distinctive facial characteristics. The CdLS phenotype represents a spectrum that includes both classic and non-classic phenotypes resulting from pathogenic variants in genes associated with cohesin functioning, including NIBPL, SMC1A, SMC3, RAD21, BRD4, HDAC8, and ANKRD11. Mutations in these genes manifest diverse clinical features, with RAD21 variants accounting for a small percentage of cohesinopathies in humans. RAD21-related cohesinopathy typically exhibits growth retardation, minor skeletal anomalies, and facial features overlapping with CdLS. However, cognitive involvement tends to be milder. Despite this, due to the limited number of reported cases with RAD21 mutations, establishing genotype-phenotype correlations remain challenging. We present the case of an 18-month-old boy exhibiting developmental delay and distinct morphological features including micro-brachycephaly, depressed nasal bridge, upturned nose, long philtrum, low-set ears, mesomelic limb dwarfism, and a complete endocardial cushion defect. Exome sequencing revealed a novel RAD21 variant in this individual.

Multiparametric arterial spin-labeling and diffusion-weighted imaging of phenotyping of polycystic ovaries

BackgroundPolycystic ovarian syndrome [PCO] is a worldwide endocrine disorder affecting women of reproductive age. Diagnosis and differentiation of PCO phenotypes are crucial for disease prognosis, fertility outcome, and treatment planning. This study aims to assess arterial spin labeling perfusion (ASL) and diffusion-weighted imaging (DWI) derived metrics in the diagnosis of PCO, differentiation of its phenotypes, and correlation of these metrics with laboratory measurements.ResultsASL and DWI of the pelvis were performed on 72 PCO patients and another 20 age-matched control group. Two observers measured the blood flow (BF) and ADC in the ovarian stroma. Serum levels of testosterone, dehydroepiandrosterone sulfate (DHEAS), and body mass index (BMI) were calculated. BF values were significantly higher in PCO patients than in control cases (P = 0.001), with area under the curve (AUC) of (0.94 and 0.89) and accuracy of (96% and 92%) for both observers, respectively. Also, BF values were significantly higher in classic than in non-classic PCO cases (P = 0.001), with AUC of (0.92 and 0.90) and accuracy of (91%) for both observers, respectively. ADC values were significantly lower in PCO patients than in control cases (P = 0.001), with AUC of (0.85 and 0.84) for the first observer and second observer, respectively. ADC values were significantly lower in classic PCO patients than in non-classic patients (P = 0.001), with AUC of (0.85 and 0.84) and accuracy of (77% and 81%) for both observers, respectively. Combined values of BF and ADC showed an accuracy of 91% and 86% for differentiating patient from control cases for both observers, respectively, and an accuracy of 92% for differentiating classic from non-classic PCO phenotypes. A significant correlation was found between ADC, BF metrics, and both serum testosterone and DHEAS levels (P < 0.05).ConclusionsCombination of ASL and ADC can be used in PCO diagnosis and can help in the differentiation of its phenotypes. Serum levels of testosterone and DHEAS have a significant correlation with ADC and BF metrics.

Anderson-Fabry disease management: role of the cardiologist.

Anderson-Fabry disease (AFD) is a lysosomal storage disorder characterized by glycolipid accumulation in cardiac cells, associated with a peculiar form of hypertrophic cardiomyopathy (HCM). Up to 1% of patients with a diagnosis of HCM indeed have AFD. With the availability of targeted therapies for sarcomeric HCM and its genocopies, a timely differential diagnosis is essential. Specifically, the therapeutic landscape for AFD is rapidly evolving and offers increasingly effective, disease-modifying treatment options. However, diagnosing AFD may be difficult, particularly in the non-classic phenotype with prominent or isolated cardiac involvement and no systemic red flags. For many AFD patients, the clinical journey from initial clinical manifestations to diagnosis and appropriate treatment remains challenging, due to late recognition or utter neglect. Consequently, late initiation of treatment results in an exacerbation of cardiac involvement, representing the main cause of morbidity and mortality, irrespective of gender. Optimal management of AFD patients requires a dedicated multidisciplinary team, in which the cardiologist plays a decisive role, ranging from the differential diagnosis to the prevention of complications and the evaluation of timing for disease-specific therapies. The present review aims to redefine the role of cardiologists across the main decision nodes in contemporary AFD clinical care and drug discovery.

Comprehensive review and expanding the genetic landscape of Cornelia-de-Lange spectrum: insights from novel mutations and skin biopsy in exome-negative cases

BackgroundCornelia de Lange Syndrome (CdLS) is a rare genetic disorder characterized by a range of physical, cognitive, and behavioral abnormalities. This study aimed to perform a comprehensive review of the literature on CdLS and investigate two cases of CdLS with distinct phenotypes that underwent WES to aid in their diagnosis.MethodsWe conducted a comprehensive review of the literature on CdLS along with performing whole-exome sequencing on two CdLS patients with distinct phenotypes, followed by Sanger sequencing validation and in-silico analysis.ResultsThe first case exhibited a classic CdLS phenotype, but the initial WES analysis of blood-derived DNA failed to identify any mutations in CdLS-related genes. However, a subsequent WES analysis of skin-derived DNA revealed a novel heterozygous mutation in the NIPBL gene (NM_133433.4:c.6534_6535del, p.Met2178Ilefs*8). The second case was presented with a non-classic CdLS phenotype, and WES analysis of blood-derived DNA identified a heterozygous missense variant in the SMC1A gene (NM_006306.4:c.2320G>A, p.Asp774Asn).ConclusionsThe study shows the importance of considering mosaicism in classic CdLS cases and the value of WES for identifying genetic defects. These findings contribute to our understanding of CdLS genetics and underscore the need for comprehensive genetic testing to enhance the diagnosis and management of CdLS patients.

Immunophenotypic variations in syphilis: insights from Mendelian randomization analysis.

Infection with Treponema pallidum instigates complex immune responses. Prior research has suggested that persistent Treponema pallidum infection can manipulate host immune responses and circumvent host defenses. However, the precise role of immune cells in Treponema pallidum infection across different stages remains a contentious issue. Utilizing summary data from genome-wide association studies, we employed a two-sample Mendelian randomization method to investigate the association between 731 immunophenotypes and syphilis. Syphilis was categorized into early and late stages in this study to establish a more robust correlation and minimize bias in database sources. Our findings revealed that 33, 36, and 27 immunophenotypes of peripheral blood were associated with syphilis (regardless of disease stage), early syphilis and late syphilis, respectively. Subsequent analysis demonstrated significant variations between early and late syphilis in terms of immunophenotypes. Specifically, early syphilis showcased activated, secreting, and resting regulatory T cells, whereas late syphilis was characterized by resting Treg cells. More B cells subtypes emerged in late syphilis. Monocytes in early syphilis exhibited an intermediate and non-classical phenotype, transitioning to classical in late syphilis. Early syphilis featured naive T cells, effector memory T cells, and terminally differentiated T cells, while late syphilis predominantly presented terminally differentiated T cells. Immature myeloid-derived suppressor cells were evident in early syphilis, whereas the dendritic cell immunophenotype was exclusive to late syphilis. Multiple immunophenotypes demonstrated associations with syphilis, showcasing substantial disparities between the early and late stages of the disease. These findings hold promise for informing immunologically oriented treatment strategies, paving the way for more effective and efficient syphilis interventions.

#313 : The Correlation Between Ultrasound Morphological Parameters and Phenotypes of Polycystic Ovary Syndrome in Indonesian Women

Background and Aims: Polycystic ovary syndrome (PCOS) is a complex endocrine disorder that affects women of reproductive age. Ultrasound morphological parameters of ovarian morphology, ovarian volume, follicle number, endometrial plate thickness, and doppler analysis are necessary for the diagnosis of PCOS. The aim of this study is to evaluate the correlation between ultrasound morphological parameters and classic and nonclassic phenotypes of PCOS in Indonesian Women. Method: A single-center, observational analytic study from May 2022 until October 2022 was performed upon 50 women with PCOS at Halim Fertility Center (HFC), Indonesia. Fifty PCOS women were classified into two groups: 20 PCOS women with classic phenotype and 30 PCOS women with non-classic phenotype. Data on patient’s age, body mass index (BMI), LH level, FSH level, LH: FSH ratio, Testosterone, Free testosterone index, SHBG, HOMA-IR, Ultrasound parameters (antral follicle count, follicle size, ovarian volume, stromal volume, Stromal/total area (S/A) ratio, Pulsatility index (PI), Peak systolic velocity (PS) and follicle distribution) were assessed and compared between the two groups. Results: The antral follicle count was higher in classic phenotype of PCOS than in nonclassic phenotype and there was a significant difference between the groups (p&lt;0.05). The stromal volume and PI were higher in non-classic phenotype PCOS than in classic phenotype and there was significantly difference between the groups (p&lt;0.05). Most of the follicle distribution in the classic phenotype of PCOS was central and the follicle distribution in the non-classic phenotype was peripheral and there was significantly difference between the groups (p&lt;0.05). The number of antral follicles, stromal volume, PI, and follicle distribution were positively correlated with the PCOS phenotypes (r=0.734; r=0.594; r=0.577; r=0.196), respectively. Conclusion: Classic phenotype of PCOS in Indonesian women has high antral follicle count and central follicle distribution and nonclassic phenotype has high stromal volume, PI and peripheral follicle distribution.

OP58 Challenges In Maintaining Up To Date Health Technology Assessments in Rare Diseases: Lessons From Fabry Disease In Australia

IntroductionFabry disease is a rare, inherited X-linked lysosomal storage disease characterized by a wide spectrum of heterogeneously progressive clinical phenotypes, and which results in progressive kidney disease, cardiomyopathy, cerebrovascular disease, and reduced life expectancy. Disease-specific therapy aims to improve symptoms, stabilize current disease and delay progression. In Australia treatment access requires that patients meet pre-specified criteria, which have been in place for more than 15 years. Patient questions prompted the patient organization, Fabry Australia, to investigate why these criteria had remained unchanged despite significant progress in the understanding and management of Fabry disease.MethodsA panel comprising two members of Fabry Australia and its Medical Advisory Committee conducted a review of the literature. The aim of this was to inform the clinical quality of the Australian treatment access criteria with reference to international guidelines and contemporary data. The findings from the literature were applied to develop consensus recommendations for classification and Fabry-specific treatment initiation criteria in diagnosed patients.ResultsEvidence supports earlier treatment with reduced barriers to access in some circumstances. Australian access criteria are misaligned with this. They do not distinguish between classical and non-classical Fabry phenotypes, neglect the impact of quality of life and gastrointestinal symptoms, and impose symptom-severity related criteria, which may lead to unnecessary treatment initiation delay. An updated framework is presented. It differentiates phenotypes, facilitates more timely access to Fabry-specific treatment for classical males, and supports relevant organ involvement criteria in classical females and patients with non-classical disease.ConclusionsA well-performing health technology assessment system facilitates patient access to cost-effective treatments that improve health outcomes. Timely treatment initiation is important to avoid irreversible organ damage in Fabry patients. Patients’ questions about out-dated access criteria has prompted research and uncovered barriers that are no longer clinically valid. The perspectives of the patient as a stakeholder in their disease management should not be overlooked when assessing the value of health technologies in the rare disease setting.

Oncogenic RAS Signaling Promotes an Immunosuppressive Niche in Acute Lymphoblastic Leukemia

Emerging evidence suggests that genomic aberrations in cancer cells may impact the composition of the tumor microenvironment leading to a tumor-specific supporting niche. Previous genomic profiling studies have identified somatic alterations in RAS pathway genes that drive treatment resistance in B-cell acute lymphoblastic leukemia (B-ALL): yet it is unknown whether such genetic events may specifically hijack the leukemic niche. Here, we analyzed bulk RNA-sequencing data of bone marrow mononuclear cells from 185 pediatric B-ALL patients entering our center and performed mutation calling of RAS pathway genes, namely KRAS, NRAS, BRAF and PTPN11. Transcriptomic analysis revealed attenuated immune responses and decreased T cell activation in samples harboring RAS mutations (n= 62) compared with wild type cases (n= 123). To further explore this finding at single-cell resolution, we sorted bone marrow mononuclear cells from 8 primary B-ALL pediatric patient samples (4 RAS mutant and 4 wild type), enriching on the hematopoietic and non-hematopoietic compartments, followed by single-cell RNA-sequencing using the 10x Genomics platform. Twelve cell populations were identified by the expression of bona fide markers, mapping different immune and stromal cells in the ALL landscape. Specifically, the RAS-mutant microenvironment was defined by dysfunctional CD8+ T-cells, characterized by high expression of TIM-3, TIGIT and LAG-3, as well as regulatory T cells, displaying expression of CTLA-4, ICOS and FOXP3, suggesting a RAS-mediated immunosuppressive mechanism. In light of these changes, we examined flow cytometry data of bone marrow mononuclear cells collected at diagnosis from our cohort of 185 patient samples. Whereas the overall proportion of lymphoid subsets (T, B and NK cells) was not significantly changed among RAS mutant and wild type groups, we detected a decrease in myeloid subsets (neutrophils, eosinophils, and monocytes) in RAS mutant ALL, which was more prominent in monocytes. Cross comparison of the transcriptional state of monocytes in our single-cell data unravel that monocytes in the RAS mutant niche displayed a non-classical phenotype, characterized with high expression of the immune checkpoint TIM-3, which is a major driver of T cell exhaustion. Taken together, the bone marrow microenvironment of ALL patients harbouring oncogenic RAS lymphoblasts is defined by dysfunctional immune cell populations. Further studies on the cellular dynamics in the niche will provide the basis for the development of therapeutic opportunities to target the immune micro-environment in RAS mutant ALL.

The injured monocyte: The link to chronic critical illness and mortality following injury.

This study aimed to understand the altered innate immune response in severely injured patients leading to chronic critical illness (CCI). Specifically, it focused on characterizing the monocyte populations and their correlation with CCI development and long-term complications. Over a 3-year period, we monitored patients with severe injuries for up to 1-year postinjury. Chronic critical illness was defined as an ICU stay exceeding 14 days with persistent organ failure. Blood samples were collected on Days 1 and 5 for monocyte phenotypic expression analysis using cytometry by time flight. The monocyte subpopulations studied were classical (CL), intermediate (INT), and nonclassical (NC), along with cell surface receptor expression and activation. Out of 80 enrolled patients, 26 (32.5%) developed CCI. Patients with CCI had more severe injuries (Injury Severity Score, 32.4 + 5.2 vs. 29.6 + 4.1, p = 0.01) and received a higher number of red blood cells (8.9 + 4.1 vs. 4.7 + 3.8 units, p < 0.01) compared with those without CCI. In patients with CCI, the NC monocytes were significantly reduced by over twofold early, and significantly increased later, compared with those without CCI. Moreover, significant changes in intracellular cytokine expression and cell receptors were observed within each monocyte subpopulation in patients with CCI, indicating an increased proinflammatory phenotype but decreased phagocytic capacity and antigen presentation. The development of CCI and the presence of this unique monocyte phenotype were associated with a significantly increased risk of infection, discharge to a long-term care facility, and 1-year mortality of 27%. Development of CCI following severe injury is associated with significant long-term morbidity and unacceptably high mortality. The altered NC phenotype with reduced phagocytic capacity and antigen presentation in patients developing CCI after severe injury is appears partially responsible. Early identification of this unique phenotype may help predict and treat patients at risk for CCI, leading to improved outcomes. Prognostic and Epidemiological; Level III.

Increased Prevalence of Accidents and Injuries in Congenital Adrenal Hyperplasia: A Population-based Cohort Study.

It has been suggested that injuries and accidents are increased in females with congenital adrenal hyperplasia (CAH), but the prevalence is unclear. To study the prevalence of injuries and accidents in females and males with CAH. Patients with CAH (n = 714, all 21-hydroxylase deficiency) were compared with matched controls (n = 71 400). Data were derived by linking National Population-Based Registers. Prevalence of injuries and accidents. Mean age was 29.8 ± 18.4 years. Injuries were more prevalent in patients with CAH than in controls (relative risk, 1.34; 95% CI, 1.24-1.44), and this was found in both sexes (females: 1.43; 1.29-1.58; males: 1.25; 1.12-1.38). In the classical phenotype, the prevalence of injuries was higher, especially in females but not in the nonclassic phenotype. In the genotype groups, injuries were mainly increased in females. Head injuries were increased in all patients with CAH and in the different phenotypes and were mainly driven by females. More patients with CAH born before the introduction of neonatal screening had had an injury compared with controls (1.48; 1.35-1.62); this was seen in both sexes. In patients with CAH born after the introduction of screening, the prevalence of injuries was overall increased (1.20; 1.07-1.35), and in females with CAH but not in males. Accidents showed a similar pattern to injuries in all comparisons. Patients with CAH had an increased prevalence of both injuries and accidents, especially in females and in those born before the neonatal screening program. Patients with nonclassic phenotype were hardly affected.

FRI402 “An Unusual Presentation Of Polycystic Ovary Syndrome” - Isolated DHEA-S Elevation In A Patient With Difficulty Conceiving And Recent Onset of Hirsutism

Abstract Disclosure: C. Musurakis: None. J.L. Gilden: None. Background: Isolated DHEA-S levels and worsening hirsutism often prompt the clinician to investigate more serious causes for the elevation, such as an adrenal mass. Clinical Case: A 29-year-old female G0P0 with Norwegian and English heritage and familial hypertriglyceridemia, was referred for elevated DHEA-S levels, and difficulty conceiving for past three years. She had hirsutism of face, chest, and abdomen, worsening over the last two years. Acne had also progressively worsened to cystic type. Menarche was at age 12, and menses were regular. She had never used contraception. Medications: docosahexanoic acid, eicosapentanoic acid, topical azelaic acid, benzoyl peroxide, clindamycin, pantoprazole, prenatal vitamins, and cholecalciferol. There was a family history of acne, but not for hirsutism or menstrual irregularities. Vitals: BP 137/73, HR 81, Ht 165.10 cm, Wt 86.18 Kg, BMI 31.68. Ferriman-Gallwey score was 10 with normal genitalia. Labs (luteal phase): DHEA-S 756 mcg/dL with repeat of 596 mcg/dl(n=35-430), total testosterone 25 ng/dL(n=2-45), free testosterone 4.3 pg/mL(n=0.1-6.4), prolactin 6.6 ng/mL(n=2.2-30.3), LH 5.1 mIU/mL(n=0.6-16.6), FSH 4.7 mIU/mL(n=1.7-12.9), 17-OH progesterone 74 ng/dL(n=139-431), androstenedione 163 ng/dL(n=73-184), estradiol 59.79 pg/mL(n=55.8-214.2), AMH 1.38 ng/mL(n=0.69-13.39), midnight salivary cortisol &amp;lt;0.03 and 0.05 mcg/dL(n&amp;lt;5), IGF-1 258 ng/mL(n=63-373), ACTH 31 pg/mL(n=0-47), AM cortisol 12.58 ug/dL(n=5.3-22.5), TSH 1.27 uIU/mL(n=0.358-3.74), Free T4 0.88 ng/dL (n=0.76-1.46), peroxidase antibodies 49.8 U/m (n=0-60), thyroglobulin antibodies &amp;lt;15(n=0-60), HbA1c 5.5% (n&amp;lt;=5.6%), 25-OH vitamin D 42.7 ng/mL(n=30-100) and negative pregnancy urine test. US pelvis: small follicles in ovaries and normal size. MRI abdomen w/wo contrast: normal bilateral adrenal glands. Cosyntropin stimulation test: baseline cortisol 15.03 ug/dL,17-OH progesterone 168 ng/dL. 30 mins: Cortisol 30.39ug/dL(n&amp;gt;18),17-OH progesterone 130ng/dL(n&amp;lt;200). 60 mins: cortisol 38.95ug/dL(n&amp;gt;18), 17-OH progesterone 153ng/dL(n&amp;lt;200). Non-Classical Congenital Adrenal Hyperplasia was excluded. Polycystic Ovary Syndrome (PCOS) was diagnosed based on clinical and biochemical hyperandrogenism, lack of adrenal mass, and clinical signs of anovulation. Conclusion: As reported by Carmina et al, few patients with PCOS exhibit isolated increase of serum DHEA-S. It is important to recognize that non-classic forms of PCOS (normal ovaries or normal ovulatory cycles) can present with a higher prevalence of adrenal hyperandrogenism than the common classical forms of PCOS. Reference: Carmina E, Longo RA. Increased Prevalence of Elevated DHEAS in PCOS Women with Non-Classic (B or C) Phenotypes: A Retrospective Analysis in Patients Aged 20 to 29 Years. Cells. 2022 Oct 17;11(20):3255. doi: 10.3390/cells11203255. PMID: 36291122; PMCID: PMC9601254. Presentation: Friday, June 16, 2023

Genotyping in patients with congenital adrenal hyperplasia by sequencing of newborn bloodspot samples.

Genotype-phenotype correlation in congenital adrenal hyperplasia (CAH) caused by 21-hydroxylase deficiency ranges from 45 to 97 %. We performed massively parallel sequencing of CYP21A2 on stored newborn bloodspot samples to catalogue the genotypes present in our patients with CAH and enable genotype-phenotype comparison. Participants≤15 years old with clinically diagnosed CAH were recruited from The Sydney Children's Hospitals Network. Phenotype was classified from clinical and biochemical details in the medical record as salt wasting (SW), simple virilising (SV), non-classic (NC) or an intermediate phenotype (SW/SV; SV/NC). Amplicon-based sequencing for CYP21A2 was performed on stored newborn bloodspot samples by the New South Wales Newborn Bloodspot Screening Laboratory on MiSeq™Dx (Illumina, California). Available genetic test results were also obtained from the medical records. Samples from 67 participants (43 % female, age 0.3-15 years) were sequenced, including 9 sibships. SW phenotype was present in 33/67 participants (49 %), SV in 9 (13 %) and NC in 16 (24 %). Intermediate phenotypes included SW/SV in seven participants (10 %) and SV/NC in two (3 %). Variants were identified in 90/116 alleles (78 %). A complete genotype was available in 47/67 participants (70 %). The most common genotype was homozygous c.293-13A/C>G (I2G) in 7/47 participants (15 %). Genotype correlated with the mostcommonly reported phenotype in 36/44 cases (82 %). Correlation was higher in SW and NC phenotypes. This study uses genetic testing of newborn bloodspots to identify and characterise the genotypes present in an ethnically diverse Australian population with CAH. It further strengthens our knowledge of genotype-phenotype correlations in CAH.

Myocardial extracellular volume fraction is positively associated with activated monocyte subsets among cART-treated persons living with HIV in South Africa

BackgroundDespite treatment with combination antiretroviral therapy (cART), persons living with HIV (PLWH) are at higher risk of cardiac structural abnormalities that may presage clinical heart failure, including myocardial fibrosis. This study assessed whether circulating cellular and soluble protein markers of immune activation cross-sectionally associate with myocardial fibrosis among cART-treated PLWH in South Africa. MethodsParticipants were enrolled in Khayelitsha township near Cape Town, SA. Cardiac magnetic resonance imaging was performed. Plasma protein biomarkers were measured using enzyme-linked immunoassays and monocyte phenotypes were evaluated using flow cytometry. Associations were assessed using multivariable linear and logistic regression. ResultsAmong 69 cART-treated PLWH, mean (SD) age was 48 (10) years, 71% were female, and time since HIV diagnosis was 9 (6) years. Evidence of left ventricular fibrosis by late gadolinium enhancement was present in 74% of participants and mean (SD) extracellular volume fraction (ECV) was 30.9 (5.9)%. Degree of myocardial fibrosis/inflammation measured by ECV was positively associated with percentages of circulating non-classical and intermediate monocyte phenotypes reflecting inflammation and tissue injury. ConclusionThese data generate hypotheses on possible immune mechanisms of HIV-associated non-ischemic myocardial disease, specifically among cART-treated PLWH in sub-Saharan Africa, where the majority of the HIV burden exists globally.

Genetic Characterization of a Cohort of Italian Patients with Congenital Adrenal Hyperplasia Due to 21-Hydroxylase Deficiency.

Defects in the steroid 21-hydroxylase gene (CYP21A2) cause 21-hydroxylase deficiency (21OHD), the main cause of congenital adrenal hyperplasia (CAH). The disease shows a broad spectrum of clinical forms, ranging from severe or classical (salt wasting, SW, and simple virilizing, SV), to mild late onset or nonclassical (NC). 21OHD affects 1 in 15,000 in its severe classic form and 1 in 200-1000 in its mild NC form. There are many studies reporting the frequency of CYP21A2 pathogenic variants in different populations; however, few of them provide comprehensive information about Italian patients. Here, we present genetic results from a cohort of 245 unrelated Italian individuals with clinical diagnosis of CAH due to 21OHD. A specific polymerase chain reaction (PCR) protocol combined with Sanger sequencing was used for CYP21A2 analysis. The multiplex ligation-dependent probe amplification (MLPA) assay was employed for copy number variation (CNV) determination. One hundred fourteen (46.5%) of the index cases had the NC form, 57 (23.3%) had the SV form, and 74 (30.2%) presented the SW form of the disease. The most prevalent variant found in NC patients was the p.Val282Leu (51.3%), while the most frequent variants in the classical form were p.Ile173Asn (8.6%) and c.293-13C>G (26.0%). In our study, the frequency of large rearrangements was 15.3%, with CAH-X alleles representing 40% of all DEL/CONV. In addition, 12 alleles carried rare variants, and 1 had a novel variant p.(Arg342Gln). We observed phenotype-genotype correlation in 94.7% of cases. A complete concordance was observed in Groups 0 (enzyme activity completely impaired) where all patients had the SW form as expected. In Group A (0-1% residual enzyme activity), 78.4% of patients had the anticipated SW form while 21.6% were diagnosed with the SV form. Within Group B (~2% residual enzyme activity), 93.4% of patients exhibited SV form and 6.5% SW disease. Finally, 92.6% and 7.4% of patients belonging to Group C (enzyme partially impaired to ~20-60% residual activity) exhibited NC and SV phenotypes, respectively. This work, representing a comprehensive genetic study, expanded the CYP21A2 variants spectrum of Italian patients with 21OHD and could be helpful in prenatal diagnosis and genetic counseling.

Physician assessment of aortic stenosis severity, quantitative parameters, and long-term outcomes: Results from the KP-VALVE project

BackgroundContemporary outcomes for aortic stenosis (AS) and the association between physician-assessed AS severity and quantitative parameters is poorly understood. We aimed to evaluate AS natural history, compare outcomes for physicians’ AS assessment vs. quantitative parameters, and identify AS parameters with the most explanatory power. MethodsWe ascertained physician-assessed AS severity, echocardiographic parameters, and clinical data for 546,769 patients from 2008-2018, examined multivariable associations of physician-assessed AS severity and number of quantitative severe AS parameters with death, cardiovascular hospitalization, and aortic valve replacement, and estimated the relative contribution of different quantitative AS parameters on outcomes. ResultsAmong 49,604 AS patients (mean [SD] age 77 [11] years), 17.6% had moderate, 3.6% moderate-severe, and 9.4% severe AS. During median 3.7 [IQR 1.7-6.8] years, physician-assessed AS severity strongly correlated with outcomes, with moderate AS patients tracking closest to mild AS, and moderate-to-severe AS patients more comparable to severe AS. Although the number of quantitative severe AS parameters strongly predicted outcomes (adjusted HR [95% CI] for death 1.40 [1.34-1.46], 1.70 [1.56-1.85], and 1.78 [1.63-1.94] for 1, 2, and 3 parameters, respectively), aortic valve area <1.0 cm2 was the most frequent severe AS parameter, explained the largest relative contribution (67%), and was common in patients classified as moderate (21%) or moderate-severe (56%) AS. ConclusionsPhysician-assessed AS severity predicts outcomes, with cumulative effects for each severe AS parameter. Moderate AS includes a wide spectrum of patients, with discordant AVA <1.0 cm2 being both common and predictive. Better identification of non-classical severe AS phenotypes may improve outcomes.

Evidence of a non-classical phenotype of hypoparathyroidism in a cohort of adult patients with iron overload diseases

Searchable abstracts of presentations at key conferences in endocrinology ISSN 1470-3947 (print) | ISSN 1479-6848 (online)

Molecular and clinical features of papillary thyroid cancer in adult patients with a non-classical phenotype.

Genotyping is fundamental in papillary thyroid cancer (PTC) and helps to enhance diagnosis and prognosis and determine appropriate treatments. The phenotype-genotype association in PTC was previously studied, with BRAF V600E characterizing classic PTC and tall-cell PTC and RAS mutations characterizing follicular-variant PTC. In clinic, some non-classical histological subtypes of PTC were also identified, however, their genotype remains unclear. In this study, we collected samples of these non-classical PTC after the exclusion of classic phenotypes and examined their phenotypes, genotype and the relationship between phenotype and genotype. We screened out non-classical PTC by excluding classical PTC from 1,059 different thyroid samples, and a total of 24 cases was obtained and described from the morphological features, which is rare in differentiated PTC. DNA/RNA sequencing was performed using 18 available samples to describe the genetic features. PTC with the non-classical phenotype were characterized cuboidal to low columnar tumor cells with subtle nuclear features of PTC and without discernible nuclear elongation, concurrently with dense microfollicles, delicate papillae or solid nodules with delicate fibrovascular cores. They were associated with lymphatic vessel invasion (P<0.001) but not with a worse prognosis (P=0.791). Gene fusions were identified in 14 of 18 (77.8%) cases, including eight fusions of NTRK and six fusions of RET. The high percentage of fusions in this papillary thyroid cancer subgroup suggested a correlation of gene fusions with the phenotype that does not belong to the BRAF V600E-mutant or RAS-mutant group. Our study retrospectively screened a large cohort of different thyroid tissue samples, and presented the histopathological and genetic features of a non-classical phenotype of PTC from 24 patients. It may contribute to diagnose in PTC, and patients of these non-classical phenotype may benefit from targeted therapy, compared to a natural patient cohort without selection.