Hemizygous IL2RG Variants Impair IL-2-Induced STAT5 Phosphorylation and Transcriptional Activity Causing X-Linked Severe Combined Immunodeficiency

We investigated the feasibility of interleukin-2 receptor gamma (IL2Rγ) gene based on gene mutation analysis and pre-natal diagnosis of X-linked severe combined immunodeficiency (X-SCID). Blood samples of patients and their parents of X-SCID (family 1) and X-SCID (family 2) were collected. IL2Rγ gene sequences of the 2 families were analyzed using bi-directional direct sequencing by polymerase chain reaction. DNA sequence changes in the IL2Rγ gene exon region and shear zone were also analyzed. We also sequenced the IL2Rγ gene in 100 healthy individuals. Prenatal genetic diagnoses for a high-risk fetus in family 1 were performed by chorionic villus sampling after determining each family's genotypes. The suspect fe-male in family 1 underwent carrier detection. Two novel mutations of IL2Rγ gene were identified, including c.361-363delGAG (p.E121del) in the patient and his mother in family 1, and c.510-511insGAACT (p.W173X) heterozygous mutation in the proband's mother in family 2. These mutations were absent in the 100 controls. Prenatal diagnosis of early pregnancy in the female fetus of family 1 was performed; the fetus was heterozygous, which was confirmed at postnatal follow-up. The suspect female in family 1 showed no mutation in carrier detection. The novel p.E121del and p.W173X mutations in IL2Rγ may have been the primary causes of disease in 2 families with X-SCID. In couples with an X-SCID reproductive history, prenatal gene mutation analysis of IL2Rγ can effectively prevent the birth of children with X-SCID and carrier detection for suspected females.

The use of fetal exome sequencing in prenatal diagnosis: a points to consider document of the American College of Medical Genetics and Genomics (ACMG)

New Insights and Unresolved Issues Regarding Insertional Mutagenesis in X-linked SCID Gene Therapy

To evaluate the diagnostic feasibility of mutation analysis and prenatal genetic diagnosis genetic analysis of IL2RG gene in two families with a birth history of X-linked severe combined immunodeficiency (X-SCID). Blood samples of a male infant patient of X-SCID and his mother in family 1 and the parents of another deceased child with X-SCID in family 2 from January 2012 to February 2013 were collected.Eight exons comprising IL2RG open reading frame and their exon/intron boundaries were analyzed by bi-directional direct sequencing of polymerase chain reaction (PCR) products. Prenatal genetic diagnoses were performed by chorionic villus sampling after the genotypes of maternal probands were identified in family 1. Two mutations of IL2RG gene were identified in these two families. The c.361-363delGAG (p.E121del) mutation was identified in family 1. The c.510-511insGAACT (p.W173X) mutation appeared in family 2. The two mutations of c.361-363delGAG (p.E121del) and c.510-511insGAACT (p.W173X) were novel. The two novel mutations were absent in 100 normal controls. The pregnancy in family 1 continued and the infant showed no symptom of X-SCID at 1 year after birth. The aunt (II-3) of proband in family 1 was not a carrier. The female fetus in family 1 had no mutation. Two novel mutations of c.361-363delGAG (p.E121del) and c.510-511insGAACT (p.W173X) in IL2RG gene may be a major cause of disease in two families with X-SCID. And direct sequencing of IL2RG gene provides genetic counseling, prenatal diagnosis and carrier screening for families with X-SCID.

To analyze the mutation of IL2RG gene in a Chinese family with a birth history of a dead child suspected of X-linked severe combined immunodeficiency (X-SCID), and to perform prenatal diagnosis with DNA sequencing. Blood samples of the parents of the dead child and chorionic villi at gestational age 11 weeks were collected. Eight exons comprising the open reading frame as well as their exon/intron boundaries of IL2RG gene were analyzed by PCR and bi-directional sequencing. A heterozygous nucleotide substitution c.690C > T (R226C) in exon 5 was detected in the mother, but not in the father. In the second pregnancy of the mother, the mutation of R226C was not detected in the male fetus by prenatal diagnosis, and the heterozygous mutation was detected in the female fetus of the third pregnancy. The reliability of the prenatal genetic diagnosis was confirmed by the one-year follow-up after the neonates were born. The mutation of c.690C>T in IL2RG gene may be the pathologic cause of the proband with X-SCID. DNA sequencing combining sex determination is a valid strategy for prenatal diagnosis of X-SCID.

DNA-based screening and population health: a points to consider statement for programs and sponsoring organizations from the American College of Medical Genetics and Genomics (ACMG)

Purpose of the Study. Interleukin-2 (IL-2) and IL-2 receptors play a critical role in the regulation of the T cell immune response. IL-2 receptors are made of combinations of three different chains a, β, and γ. Different combinations of these chains result in the formation of three different classes of IL-2 receptors. The authors of this paper were interested in the interleukin-2 receptor γ chain (IL-2Rγ) gene and its relationship to X-linked severe combined immunodeficiency (XSCID). Findings. The IL-2Rγ gene was localized to the X chromosome in the same region as the XSCID locus. The IL-2Rγ gene from three patients with XSCID were found to have point mutations leading to premature stop codons and truncated IL-2Rγ chains. These shortened IL-2Rγ chains lack the cytoplasmic domain required for cell signal transduction. Reviewers' Comments. This is the first article showing the XSCID is associated with mutations in the IL-2Rγ gene. This suggests that IL-2Rγ plays a critical role in the early development of T-cells and their function in the immune system. These results also suggest that other genetic defects in components of the IL-2/IL-2R system could be responsible for other forms of SCID or other immunodeficiency syndromes. The fact that all three patients had different mutations suggests that there is no single dominant mutation. These findings have important implications for prenatal and postnatal diagnosis, carrier identification, and possibly gene therapy for XSCID.

Severe combined immunodeficiency (SCID) is a rare, fatal syndrome of diverse genetic etiology characterized by profound deficiencies of T and B cell function. In the U.S., it affects primarily male infants. Some are due to mutations in the genes encoding adenosine deaminase (ADA), the common gamma chain (Yc) of the IL-2, IL-4, IL-7, IL-9, and IL-15 receptors, or Janus kinase 3 (Jak3). However, for a significant proportion of SCID's, the molecular basis remains unknown. In an effort to identify unique phenotypic and/or functional features that could help identify fundamental causes, we analyzed the clinical, genetic and immunologic features of 95 SCID infants who presented consecutively to this institution between August of 1965 and December of 1995. Eighty (84%) were male and 15 (16%) female; 74 (78%) were white, 13 (14%) black, and 8 (8%) Hispanic. The 95 infants fit 7 categories: 16 (17%) were ADA deficient; 6 (6.3%) had Jak3 deficiency; 16 (17%) had autosomal recessive inheritance of unknown molecular cause; 1 (1.2%) had reticular dysgenesis; 1 (1.2%) had cartilage hair hypoplasia; 38 (40%) had X-linked SCID; and 17 (18%) (all males) had SCID of unknown type. Numbers of total lymphocytes and the various lymphocyte phenotypes distinguished the groups: ADA deficient SCID's had the lowest numbers of total lymphocytes; B cells were present in highest number and NK cells in lowest number in X-linked and Jak3-deficient SCIDs. Mean numbers of NK cells were highest in autosomal recessive and unknown types of SCID. All types were profoundly deficient in T cells. NK function was tested in 60 and found to be normal in 25. Although the family history was positive in only 34% of X-linked cases, it is likely that as many as two-thirds of the boys with SCID have mutations in Yc. The presence of NK function in ADA deficient, autosomal recessive, and some SCID's of unknown type, and abnormally low NK numbers and function in a majority of X-linked and Jak3-deficient SCID's implies that some molecular lesions affect T, B and NK cells (Yc and Jak3 mutations), but that others affect only T cells (ADA deficiency), and others T and B cells (possible recombinase abnormalities). The findings in this group of SCID's, the largest reported in the U.S., argue against a common lineage for T and NK cells and may provide clues as to the molecular bases of SCID in the 35 (37%) infants who did not have X-linked SCID, Jak3 deficiency or ADA deficiency.

Eligibility Criteria and Genetic Testing Results from a High-Risk Cohort for Hereditary Breast and Ovarian Cancer Syndrome in Southeastern Ontario

A new comprehensive paradigm for prenatal diagnosis: seeing the forest through the trees.

Objective To analyse the relationship between clinical manifestations and the mutation of interleukin-2 receptor common gamma chain (IL-2RG) mutation of X-linked severe combined immunodeficiency (SCID) caused by IL-2RG. Methods From May 2010 to September 2012, four boys who were clinically suspected as X-linked SCID were enrolled in this study. The study protocol was approved by the Ethical Review Board of Investigation in Human Being of West China Second University Hospital, Sichuan University.Informed consent was obtained from each participants' parents. IL-2RG mutation were detected and analyzed in four patients and their family members. Results Four boys were all suffered by severe pulmonary infection (severe pneumonia) and demonstrated to have absence of thymus or dysplastic thymus by chest X-ray film. Among them, three patients had tuberculosis infection after bacilli Calmette-Guerin (BCG) vaccination (75%), two patients had family history (50%), and all patients suffered lymphopenia (100%). All 4 male infants were demonstrated to be X-linked SCID due to the mutation of IL-2RG, including 2 patients with nonsense mutations (50%, c. 711.G>A, p. W237X and c. 578.G>A, p. W193X, respectively), one missense mutation (25%, c. 173C>A, p. P58Q), and 1 splice site mutation (25%, IVS5-2 G>T). The linear relatives of 3 patients and maternal relatives were detected the mutation of IL-2RG, and 3 patients were demonstrated to have family carriers (75%) and one patient was suffered by de novo mutation (25%). Conclusions The detection of IL-2RG gene mutation can confirm clinical diagnosis of X-linked SCID which is critical for patients to take hematopoietic stem cell transplantation as soon as possible to reconstruction of immunity before the fatal infection occurred. The early immune reconstruction can offer the best chance of long term survival. Key words: Severe combined immunodeficiency; Receptors, interleukin-2; Mutation

Interleukin-2 receptor γ chain mutation results in X-linked severe combined immunodeficiency in humans

Prenatal exome sequencing for fetuses with structural abnormalities: the next step.

Introduction:KMT2A (MLL) rearranged (KMT2A-r) infant acute lymphoblastic leukemia (ALL) is a distinct leukemia with a poor prognosis, especially when compared to childhood ALL. Previous studies on KMT2A-r leukemia have demonstrated that KMT2A rearrangement can occur in utero with a short latency after birth. Infant ALL with KMT2A-r also demonstrates a low frequency of other somatic mutations. Given that germline mutations in cancer predisposition genes are found in 5-15% of children with cancer, we hypothesized that there might be distinct germline mutations in infants with this type of leukemia that could inform about leukemogenesis and biologic behavior of this entity. Methods: We performed whole genome sequencing (WGS) and whole exome sequencing (WES) on DNA isolated from 43 peripheral blood samples at the time of remission, on 3 cohorts of infants. The cohorts were comprised of infants with KMT2A-r ALL who were known to relapse (n=14) (Cohort A), infants with KMT2A-r ALL who to date have remained in remission (n=14) (Cohort B), and infants with KMT2A germline ALL (n=15) (Cohort C). Sequencing was performed using an Illumina Hiseq 4000 or 2500 to a minimum depth of 90Gb. Alignment and variant calling were performed using the Dragon Bio-IT platform (v 3.2.8, Illumina). We then looked for variants in 347 genes associated with cancer predisposition and bone marrow failure syndromes. Variants present in remission samples at a variant allele frequency of ~50%, and rare (minor allele frequency <0.1%) in control population databases (gnomAD), were considered for analysis and interpreted using ACMG/AMP variant interpretation guidelines.Results:We found 64 germline variants in cohort A (63 variants of unknown significance (VUS) and 1 likely pathogenic (LP) variant), 39 of which are associated with cancer predisposition ( 15 autosomal dominant (AD), 11 autosomal recessive (AR) and 13 others) and 25 other variants associated with bone marrow failure (BMF). In cohort B, we found 67 germline variants (64 VUS and 3 LP), 42 of which are associated with cancer predisposition (12 AD, 14 AR, and 16 others) and 25 other variants associated with bone marrow failure (BMF). Cohort C had 67 germline variants (67 VUS), 50 of which are associated with cancer predisposition (21 AD, 15 AR, and 14 others) and 17 other variants associated with bone marrow failure (BMF).Conclusion:Overall, there were no significant differences across the 3 cohorts in terms of the number and types of variants identified. Although there were germline VUS present in cancer predisposition genes in KMT2A-r infant ALL, there were no distinct recurrent pathogenic variants seen to make a significant association with an underlying cancer predisposition syndrome. Citation Format: Azhar Saeed, Midhat Farooqi, Byunggil Yoo, Rumen Kostadinov, Emily Farrow, Neil Miller, Patrick Brown, Erin Guest. Genomic analysis of infant KMT2A-r acute lymphoblastic leukemia reveals weak association with underlying germline cancer predisposition [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 5417.

Germline Variants Associated with Cancer Predisposition and Bone Marrow Failure Are Common in KMT2A-r Infant Acute Lymphoblastic Leukemia Patients