Of over 7000 patients referred to a diagnostic laboratory, 28% had diagnoses based on DNA sequencing, 5% of whom had two or more diagnoses. Their phenotypes could be better understood by considering ...whether the implicated genes affect independent biologic processes or organ systems.
Medical genetics focuses on the relationship between observed phenotypes and their underlying genotypes, modes of transmission, and risks of recurrence. Expected patterns of mendelian inheritance are often used to confirm the identification of disease genes, and deviations from mendelian expectations have led to the discovery of more complicated genetic underpinnings of disease (Fig. S1 in the Supplementary Appendix, available with the full text of this article at NEJM.org).
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Multiple (or dual) molecular diagnoses involve more than one clinical diagnosis and more than one genetic locus (Figure 1), each segregating independently.
Diagnostic whole-exome sequencing affords opportunities for providing insights into relationships . . .
Mechanisms by which attainment of specific body sizes trigger developmental transitions to adulthood (e.g. puberty or metamorphosis) are incompletely understood. In Drosophila, metamorphosis is ...triggered by ecdysone synthesis from the prothoracic gland (PG), whereas growth rate is increased by insulin/insulin growth factor signalling (IIS). Transgene-induced activation of PI3K, the major effector of IIS, within the PG advances the onset of metamorphosis via precocious ecdysone synthesis, raising the possibility that IIS triggers metamorphosis via PI3K activation in the PG. Here we show that blocking the protein kinase A (PKA) pathway in the insulin producing cells (IPCs) increases IIS. This increased IIS increases larval growth rate and also advances the onset of metamorphosis, which is accompanied by precocious ecdysone synthesis and increased transcription of at least one ecdysone biosynthetic gene. Our observations suggest that IIS is regulated by PKA pathway activity in the IPCs. In addition, taken together with previous findings, our observations are consistent with the possibility that, in Drosophila, attainment of a specific body size triggers metamorphosis via the IIS-mediated activation of PI3K and hence ecdysone synthesis in the PG.
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Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
IMPORTANCE: Clinical whole-exome sequencing is increasingly used for diagnostic evaluation of patients with suspected genetic disorders. OBJECTIVE: To perform clinical whole-exome sequencing and ...report (1) the rate of molecular diagnosis among phenotypic groups, (2) the spectrum of genetic alterations contributing to disease, and (3) the prevalence of medically actionable incidental findings such as FBN1 mutations causing Marfan syndrome. DESIGN, SETTING, AND PATIENTS: Observational study of 2000 consecutive patients with clinical whole-exome sequencing analyzed between June 2012 and August 2014. Whole-exome sequencing tests were performed at a clinical genetics laboratory in the United States. Results were reported by clinical molecular geneticists certified by the American Board of Medical Genetics and Genomics. Tests were ordered by the patient’s physician. The patients were primarily pediatric (1756 88%; mean age, 6 years; 888 females 44%, 1101 males 55%, and 11 fetuses 1% gender unknown), demonstrating diverse clinical manifestations most often including nervous system dysfunction such as developmental delay. MAIN OUTCOMES AND MEASURES: Whole-exome sequencing diagnosis rate overall and by phenotypic category, mode of inheritance, spectrum of genetic events, and reporting of incidental findings. RESULTS: A molecular diagnosis was reported for 504 patients (25.2%) with 58% of the diagnostic mutations not previously reported. Molecular diagnosis rates for each phenotypic category were 143/526 (27.2%; 95% CI, 23.5%-31.2%) for the neurological group, 282/1147 (24.6%; 95% CI, 22.1%-27.2%) for the neurological plus other organ systems group, 30/83 (36.1%; 95% CI, 26.1%-47.5%) for the specific neurological group, and 49/244 (20.1%; 95% CI, 15.6%-25.8%) for the nonneurological group. The Mendelian disease patterns of the 527 molecular diagnoses included 280 (53.1%) autosomal dominant, 181 (34.3%) autosomal recessive (including 5 with uniparental disomy), 65 (12.3%) X-linked, and 1 (0.2%) mitochondrial. Of 504 patients with a molecular diagnosis, 23 (4.6%) had blended phenotypes resulting from 2 single gene defects. About 30% of the positive cases harbored mutations in disease genes reported since 2011. There were 95 medically actionable incidental findings in genes unrelated to the phenotype but with immediate implications for management in 92 patients (4.6%), including 59 patients (3%) with mutations in genes recommended for reporting by the American College of Medical Genetics and Genomics. CONCLUSIONS AND RELEVANCE: Whole-exome sequencing provided a potential molecular diagnosis for 25% of a large cohort of patients referred for evaluation of suspected genetic conditions, including detection of rare genetic events and new mutations contributing to disease. The yield of whole-exome sequencing may offer advantages over traditional molecular diagnostic approaches in certain patients.
Immunocompromised patients, including those with inborn errors of immunity (IEI), may be at increased risk for severe or prolonged infections with SARS-CoV-2 (Zhu et al. N Engl J Med. 382:727–33,
...2020
; Guan et al.
2020
; Minotti et al. J Infect. 81:e61–6,
2020
). While antibody and T cell responses to SARS-CoV-2 structural proteins are well described in healthy convalescent donors, adaptive humoral and cellular immunity has not yet been characterized in patients with antibody deficiency (Grifoni et al. Cell. 181:1489–1501 e1415,
2020
; Burbelo et al.
2020
; Long et al. Nat Med. 26:845–8,
2020
; Braun et al.
2020
). Herein, we describe the clinical course, antibody, and T cell responses to SARS-CoV-2 structural proteins in a cohort of adult and pediatric patients with antibody deficiencies (
n
= 5) and controls (related and unrelated) infected with SARS-CoV-2. Five patients within the same family (3 with antibody deficiency, 2 immunocompetent controls) showed antibody responses to nucleocapsid and spike proteins, as well as SARS-CoV-2 specific T cell immunity at days 65–84 from onset of symptoms. No significant difference was identified between immunocompromised patients and controls. Two additional unrelated, adult patients with common variable immune deficiency were assessed. One did not show antibody response, but both demonstrated SARS-CoV-2-specific T cell immunity when evaluated 33 and 76 days, respectively, following SARS-CoV-2 diagnosis. This report is the first to show robust T cell activity and humoral immunity against SARS-CoV-2 structural proteins in some patients with antibody deficiency. Given the reliance on spike protein in most candidate vaccines (Folegatti et al. Lancet. 396:467–78,
2020
; Jackson et al. N Engl J Med. 383:1920–31,
2020
), the responses are encouraging. Additional studies will be needed to further define the timing of onset of immunity, longevity of the immune response, and variability of response in immunocompromised patients.
Idiopathic CD4 lymphocytopenia was initially described approximately 30 years ago as HIV infections were emerging. In this report, longer-term characteristics of this syndrome are described.
Truncating mutations in the maternally imprinted, paternally expressed gene MAGEL2, which is located in the Prader-Willi critical region 15q11–13, have recently been reported to cause Schaaf-Yang ...syndrome, a Prader-Willi-like disease that manifests as developmental delay/intellectual disability, hypotonia, feeding difficulties, and autism spectrum disorder. The causality of the reported variants in the context of the patients’ phenotypes was questioned, as MAGEL2 whole-gene deletions seem to cause little or no clinical phenotype.
Here we report a total of 18 newly identified individuals with Schaaf-Yang syndrome from 14 families, including 1 family with 3 individuals found to be affected with a truncating variant of MAGEL2, 11 individuals who are clinically affected but were not tested molecularly, and a presymptomatic fetal sibling carrying the pathogenic MAGEL2 variant.
All cases harbor truncating mutations of MAGEL2, and nucleotides c.1990–1996 arise as a mutational hotspot, with 10 individuals and 1 fetus harboring a c.1996dupC (p.Q666fs) mutation and 2 fetuses harboring a c.1996delC (p.Q666fs) mutation. The phenotypic spectrum of Schaaf-Yang syndrome ranges from fetal akinesia to neurobehavioral disease and contractures of the small finger joints.
This study provides strong evidence for the pathogenicity of truncating mutations of the paternal allele of MAGEL2, refines the associated clinical phenotypes, and highlights implications for genetic counseling for affected families.
Inborn errors of immunity (IEI) are a genetically heterogeneous group of disorders with a broad clinical spectrum. Identification of molecular and functional bases of these disorders is important for ...diagnosis, treatment, and an understanding of the human immune response. We identified 6 unrelated males with neutropenia, infections, lymphoproliferation, humoral immune defects, and in some cases bone marrow failure associated with 3 different variants in the X-linked gene TLR8, encoding the endosomal Toll-like receptor 8 (TLR8). Interestingly, 5 patients had somatic variants in TLR8 with <30% mosaicism, suggesting a dominant mechanism responsible for the clinical phenotype. Mosaicism was also detected in skin-derived fibroblasts in 3 patients, demonstrating that mutations were not limited to the hematopoietic compartment. All patients had refractory chronic neutropenia, and 3 patients underwent allogeneic hematopoietic cell transplantation. All variants conferred gain of function to TLR8 protein, and immune phenotyping demonstrated a proinflammatory phenotype with activated T cells and elevated serum cytokines associated with impaired B-cell maturation. Differentiation of myeloid cells from patient-derived induced pluripotent stem cells demonstrated increased responsiveness to TLR8. Together, these findings demonstrate that gain-of-function variants in TLR8 lead to a novel childhood-onset IEI with lymphoproliferation, neutropenia, infectious susceptibility, B- and T-cell defects, and in some cases, bone marrow failure. Somatic mosaicism is a prominent molecular mechanism of this new disease.
•Mosaic and germline gain-of-function variants in TLR8 cause neutropenia, antibody deficiency, lymphoproliferation, and bone marrow failure.•TLR8 gain of function leads to an inflammatory environment with activated T cells and abnormalities in B-cell differentiation.
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Though copy number variants (CNVs) have been suggested to play a significant role in inborn errors of immunity (IEI), the precise nature of this role remains largely unexplored. We sought to ...determine the diagnostic contribution of CNVs using genome-wide chromosomal microarray analysis (CMA) in children with IEI.
We performed exome sequencing (ES) and CMA for 332 unrelated pediatric probands referred for evaluation of IEI. The analysis included primary, secondary, and incidental findings.
Of the 332 probands, 134 (40.4%) received molecular diagnoses. Of these, 116/134 (86.6%) were diagnosed by ES alone. An additional 15/134 (11.2%) were diagnosed by CMA alone, including two likely
changes. Three (2.2%) participants had diagnostic molecular findings from both ES and CMA, including two compound heterozygotes and one participant with two distinct diagnoses. Half of the participants with CMA contribution to diagnosis had CNVs in at least one non-immune gene, highlighting the clinical complexity of these cases. Overall, CMA contributed to 18/134 diagnoses (13.4%), increasing the overall diagnostic yield by 15.5% beyond ES alone.
Pairing ES and CMA can provide a comprehensive evaluation to clarify the complex factors that contribute to both immune and non-immune phenotypes. Such a combined approach to genetic testing helps untangle complex phenotypes, not only by clarifying the differential diagnosis, but in some cases by identifying multiple diagnoses contributing to the overall clinical presentation.
Newborn screening can identify neonatal T-cell lymphopenia through detection of a low number of copies of T-cell receptor excision circles in dried blood spots collected at birth. After a positive ...screening result, further diagnostic testing is required to determine whether the subject has severe combined immunodeficiency or other causes of T-cell lymphopenia. Even after thorough evaluation, approximately 15% of children with a positive result of newborn screening for T-cell receptor excision circles remain genetically undiagnosed. Identifying the underlying genetic etiology is necessary to guide subsequent clinical management and family planning.
We sought to elucidate the genetic basis of patients with T-cell lymphopenia without an apparent genetic diagnosis.
We used clinical genomic testing as well as functional and immunologic assays to identify and elucidate the genetic and mechanistic basis of T-cell lymphopenia.
We report 2 unrelated individuals with nonsevere T-cell lymphopenia and abnormal T-cell receptor excision circles who harbor heterozygous loss-of-function variants in forkhead box I3 transcription factor (FOXI3).
Our findings support the notion that haploinsufficiency of FOXI3 results in T-cell lymphopenia with variable expressivity and that FOXI3 may be a key modulator of thymus development.
Background: In Drosophila, each of the three larval instars ends with a molt, triggered by release of steroid molting hormone ecdysone from the prothoracic gland (PG). Because all growth occurs ...during the larval stages, final body size depends on both the larval growth rate and the duration of each larval stage, which in turn might be regulated by the timing of ecdysone release.
Results: Here, we show that the expression of activated Ras, PI3 kinase (PI3K), or Raf specifically in the PG reduces body size, whereas activated Ras or PI3K, but not Raf, increases PG cell size. In contrast, expression of either dominant-negative (dn) Ras, Raf, or PI3K increases body size and prolongs the larval stages, leading to delayed pupariation, whereas expression of dn-PI3K, but not of dn-Raf or dn-Ras, reduces PG cell size. To test the possibility that altered ecdysone release is responsible for these phenotypes, we measured larval ecdysone levels indirectly, via the transcriptional activation of two ecdysone targets, E74A and E74B. We found that the activation of Ras within the PG induces precocious ecdysone release, whereas expression of either dn-PI3K or dn-Raf in the PG greatly attenuates the ecdysone increase that causes growth cessation and pupariation onset.
Conclusions: We conclude that Ras activity in the PG regulates body size and the duration of each larval stage by regulating ecdysone release. We also suggest that ecdysone release is regulated in two ways: a PI3K-dependent growth-promoting effect on PG cells, and a Raf-dependent step that may involve the transcriptional regulation of ecdysone biosynthetic genes.
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