Biallelic Heterozygous Mutations Research Articles

Bi-Allelic Heterozygous Mutations in LPR2, Presenting as Donnai-Barrow Syndrome in Two Brothers

Bi-Allelic Heterozygous Mutations in LPR2, Presenting as Donnai-Barrow Syndrome in Two Brothers

WES Reveals Novel Heterozygous NBAS Gene Mutations Associated with Fanconi Syndrome in a Patient with SOPH Syndrome: Case Report

Variations in the NBAS gene is known to cause a spectrum of phenotypes ranging from isolated recurrent acute liver failure (RALF) to a multisystemic presentation known as SOPH syndrome. Patients with SOPH present with optic atrophy, acute liver failure, short stature, and Pelger-Huet anomaly. We report the presence of a novel pair of biallelic heterozygous mutations c.5139-5T>G and c.2203-2A>G in the NBAS gene of a patient with SOPH syndrome. A 9-year-old patient was clinically diagnosed with SOPH following clinical laboratory analyses. Current interventions for managing the disease encompass IVIG, methylprednisolone, calcium, and vitamin D administration. Whole-exome sequencing (WES) results showed two mutations: c.2203-2A>G and c.5139-5T>G, in the NBAS gene, which had not been previously reported. Notably, we hypothesize that NBAS mutations could potentially contribute to the development of Fanconi syndrome, a clinical diagnosis reported in our patient. Our study also supports the renaming of SOPH to SOPHIA to allow early detection and effective treatment.

Une maladie veino-occlusive pulmonaire familiale avec mutation biallélique hétérozygote composite d’EIF2AK4

Pulmonary veno-occlusive disease (PVOD) is a rare cause of pulmonary hypertension. Heritable and sporadic forms have been distinguished. Hypoxemia, profound reduction in the diffusion of carbon monoxide and haemodynamic confirmation of pre-capillary pulmonary hypertension are the major diagnostic criteria. Thoracic CT scanning and a response to pharmaceutical therapy provide additional information to confirm the diagnosis. A 52-year-old patient, three of whose siblings had pulmonary hypertension, was admitted with dyspnoea, malaise and palpitations. Right heart catheterisation confirmed pre-capillary pulmonary hypertension. A search for an EIF2AK4 mutation was carried out, and this showed a composite biallelic heterozygous mutation compatible with the diagnosis of familial PVOD, identical to that showed in one of his brothers. Given the signs of severity of the disease and the diagnosis of PVOD, whose response to pharmaceutical therapy is often poor, the patient was placed on a waiting list for lung transplantation. Despite a similar diagnosis in 3 brothers and follow-up proposed 11 years before the diagnosis, pulmonary hypertension appeared within a few weeks and led immediately to a severe clinical situation. Annual clinical and echocardiographic monitoring had been strongly advised to the patient, but had not allowed diagnosis at a mild or moderate stage of the disease. This clinical case shows that the identification of factors predicting the development of heritable PVOD at a pre-symptomatic stage is an important issue for clinical research.

Novel biallelic missense mutations in CTC1 gene identified in a Chinese family with Coats plus syndrome

BackgroundCoats plus syndrome is a recently described, very rare multisystem disorder. The clinical phenotype is wide and variable, which making the diagnosis more difficulty. The genetic study of Coats plus syndrome has been reported recently. The biallelic heterozygous mutations in CTC1 gene, encoding conserved telomere maintenance component 1, were identified in families with Coats plus from different ancestry (European, American, and African). To data, there has not been a report about genetically confirmed Coats plus syndrome from China. ResultsWe firstly identify a novel biallelic heterozygous missense variants (c.775G>A p.V259M and c.2066A>G p.Y689C) of CTC1 gene in a Chinese family with Coats plus. The c.2066A>G mutation (p.Y689C) in CTC1 is a novel variant. Such variant was not found in any of the 85 healthy individuals in the same community. ConclusionThis is the first report of a genetically confirmed case of Coats plus from China. Targeted sequencing of CTC1 gene is useful for genetic diagnosis in Coats plus and differential diagnosis for other patients with similar disease manifestations.

Comparison of Th1/Th2 cytokine profiles between primary and secondary haemophagocytic lymphohistiocytosis.

BackgroundHaemophagocytic lymphohistiocytosis (HLH) is a life-threatening disorder of immune regulation, and HLH patients with mutations in genes including PRF1, UNC13D, STX11, STXBP2, SH2D1A, XIAP, and ITK were reported to be primary HLH. Due to the different treatment options, the differentiation between primary and secondary HLH is critical. Our previous studies have showed that a Th1/Th2 cytokine profile is diagnostic for HLH, yet the cytokine profiles between primary and secondary HLH have not been compared. The aim of the study was to test whether the Th1/Th2 cytokine profile could be used as a tool to differentiate between primary and secondary HLH.MethodsA total of 45 hospitalized Chinese children with HLH during the period of February 2010 through September 2012 were enrolled in the study. Fifty healthy children were enrolled as controls. Primary HLH related genes were sequenced using genomic DNA samples. The Th1/Th2 cytokine levels including interferon-γ (IFN-γ), tumor necrosis factor-alpha (TNF-α), interleukin (IL)-10, IL-6, IL-4 and IL-2 were quantitatively determined by cytometric bead assay techniques.ResultsPrimary HLH group (n = 4) included one patient with biallelic heterozygous mutations in PRF1 gene, and three patients with hemizygous mutation in SH2D1A gene. Based on the available genetic data, the other 41 patients were classified into the secondary HLH group. When compared the cytokine levels between the two groups, IL-4 level in primary-HLH was significantly lower than that in secondary HLH (P = 0.025), while IFN-γ level in primary HLH had a tendency of statistically lower than that in secondary HLH (P = 0.051). Area under receiver operating characteristic (ROC) curves of IL-4 and IFN-γ, IL-10, TNF-α, IL-2, and IL-6 levels were 0.841, 0.799, 0.506, 0.494, 0.457, and 0.250, respectively. ROC curves showed that 1.7 pg/ml of IL-4 had sensitivity and specificity for differentiation between primary and secondary HLH as 70.7 and 100.0 %, while 433.9 pg/ml of IFN-γ had sensitivity and specificity as 51.2 and 100.0 %, respectively.ConclusionsHLH patients with lower IL-4 and IFN-γ levels have higher possibility to be primary HLH. The cytokine profile may be used as an additional tool for the quick differential diagnosis between primary and secondary HLH.Electronic supplementary materialThe online version of this article (doi:10.1186/s13052-016-0262-7) contains supplementary material, which is available to authorized users.

At the crossroads of bench and bedside: how gonadotropins do their thing!

No single sentinel better captures the reproductive fitness of organisms than the gonadotropic hormones. Whether alluding to the sustaining forces of spermatogenesis or the cascade of events ushering in ovulation, this remarkable duo of glycoproteins, FSH and LH, purposely and heroically serve as the centerpiece of the reproductive axis for those multicellular organisms whose survival and propagation lie in the waiting. So it is after a protracted period of neglect that we draw attention in this issue of JARG to tales of FSH and LH of immediate clinical and experimental import. First to the bedside! In one of those rare case reports, suffused with implications rather than speculations, the group at the Toronto Centre for Advanced Reproductive Technology of the University of Toronto and the Mount Sinai Hospital have discovered a mutation in the LH/HCG receptor that is certain to raise more questions than it answers regarding that age-old problem of how do we get from the LH surge to ovulation of a developmentally competent oocyte. The black box that is the Graafian follicle before and after the LH surge retains a mystical quality thus continuing the saga of bemusement and wonder that so much of human ART is centered around. Readers are encouraged to follow the case history in detail (Mitri et al., A novel compound heterozygous mutation of the luteinizing hormone receptor: implications for fertility, DO10.1007/s10815-014-0249-5). Those whose interest has been piqued by the genetic underpinnings of this case should see the accompanying commentary by our Senior Editor Professor T. Raj Kumar (“Been Hit Twice”: a novel bi-allelic heterozygous mutation LHCGR). As is often the case when managing difficult cases, Mitri and colleagues were presented with a confounding initial result when oocyte retrieval following conventional COH yielded mostly immature oocytes. Not be stonewalled, the group next extended their strategy for treating this patient within the purview of recent advances in ovarian follicle physiology. Accordingly, they reasoned that deficiencies in the signaling pathway for LH likely to contribute to the phenotype observed could be based in either a failure to propagate sufficient prostaglandins or a defect in what had become a central tenet of ovulation biology--the cascade of events mediated by EGF-like molecules as exemplified by a number of studies on mammalian oocytes and follicles. Would the application of conventional wisdom derived from studies on animals provide the long-awaited coup de grâce for the bench to bedside paradigm? As reported, things are never as straightforward as one would like when it comes to solving problems of this kind. Prostaglandin treatments did little to encourage or ameliorate the suspected defect in LH signaling. They next attempted in vitro maturation using amphiregulin, one of those EGF look-alikes that has worked so well on animal cumulus complexes; this strategy did result in maturation of a fraction of the retrieved GV oocytes to metaphase-2, but following ICSI, the resultant embryos failed to develop to the blastocyst stage. Clearly, patients of this ilk will continue to seek treatment for their infertility, and the case at hand is illustrative of how challenging resolutions will be in genetically complex cases. Never fear, however, as the human ART repertoire was brought to bear on this problem as oocyte donation has resulted in a live birth for this couple, interestingly validating the initiation and sustainment of a pregnancy while lacking functional LH receptor! Our readers are also encouraged to view another case, this time affecting males, where the need for treatment for Leydig cell hypoplasia (LCH) could have its origins in a similar genetic disorder (see Successful testicular sperm recovery and IVF treatment in a man with Leydig cell hypoplasia, DOI 10.1007/s10815-014-0241-0). No matter the gender, so many cases of unexplained infertility continue to escape detection and/or explanation with our present armament of technology. Our continuing reliance on the evolution of deep-sequencing approaches stands as an example of what will come to pass in the coming years, bringing patients and physicians a step closer to achieving the level of satisfaction and success they so deserve. JARG will continue to bring advances in molecular genetics to our readership as we serve your needs as educators aimed at understanding and applying the new technologies taking residence in the field of human ARTs.

Survival Outcomes in Patients with FLT3-Itd Positive Acute Myeloid Leukaemia Relative to Biological Stratification: Smaller FLT3-Itd Clone Size Predict Better Overall Survival

AbstractAbstract 1460 Background:FLT3 internal tandem duplication (itd) mutations are found in 25% of adult patients with acute myeloid leukaemia (AML) and are associated with an adverse prognosis. This mutation results in constitutive activation of downstream pathways.Distinct biological subgroups can be identified based on FLT3-itd mutation type: clones with heterozygous FLT3-itd mutated/wildtype; homozygous mutated FLT3 allele; FLT3 heterozygous biallelic mutant and clones with evidence of overexpression of FLT3-itd. There is evidence to suggest that these mechanisms are important in the clonal evolution of AML cells. We sought to investigate their clinical impact. Method:Itd mutation analysis within exon 14 and/or exon 15 of the FLT3 gene was carried out at diagnosis by PCR of genomic DNA (gDNA) and cDNA for all new AML referrals in the region over 8 years. PCR products were identified and sized using fluorescent based fragment analysis. Allelic ratio (AR) and expression ratio (ER) (mutation: wild type ratio in gDNA and cDNA respectively) was determined from the relative peak heights. High relative expression was defined as 10 fold difference of ER/AR. Copy neutral loss of heterozygosity for the wild type allele resulting in homozygosity of the FLT3-itd mutation (acquired isodisomy (AID)) was determined by analysis of microsatellite markers along chromosome 13. Overall survival (OS) (time of diagnosis to death), event free survival (EFS) (time from diagnosis to induction failure, relapse or death) was calculated. Survival rates were estimated by the Kaplan-Meier method. Differences between the survival distributions were compared with the log-rank test. Results:177 patients positive for the FLT3-itd mutation were identified. Median follow-up for patients alive was 3.4 years. A separate group of 49 patients tested negative for this mutation during this period had better OS and EFS (p=0.02) compared to the patients who were FLT3-itd positive (median survival 1715 and 307 days respectively).Patients who were FLT3-itd positive had statistically significant (p<0.05) differences in outcomes based on age, presenting white cell count, treatment intensity and cytogenetic risk. The characteristics of this group of patients are described below.Patients with lower AR (less than/equal to 0.3) as compared to higher AR (greater than 0.3) had an improved OS (p=0.018) and EFS (p=0.02). The impact of AR on OS had borderline significance (p=0.05) when only patients treated with intensive chemotherapy were considered.AID provides true evidence of FLT3 mutant homozygosity and was detected in 15 (142 tested) patients. In 38 patients who relapsed and had samples at these stages, 5 had developed AID, but were heterozygous (mutant/wildtype) at diagnosis. 6 patients with multiple FLT3-itd products may comprise patients who have multiple different mutant/wildtype clones but may include patients with a second, independent FLT3-itd mutation resulting in biallelic heterozygous mutations, although this cannot be confirmed. A high relative expression level of FLT3-itd was seen in 12 patients. The clinical significance of these findings is uncertain due to the small numbers. Conclusion:The impact of FLT3-itd mutation and other known prognostic factors has been confirmed in a heterogeneous, real life cohort of patients.An AR over 1 provides firm evidence of loss of the wild type allele (9 patients). AID also occurs at an AR of less than 1 because of the presence of normal cells or due to preferential amplification of the wildtype allele. Direct testing for AID is a more sensitive measure of FLT3 mutant homozygosity (detected in 14% of tested patients). The development of AID in patients who relapse may be an important mechanism by which an AML clone gains a further advantage.Lower AR was associated with improved survival. In the context of a high blast count (median bone marrow blast 80%), this implies having a small sub-clone of FLT3-itd positive cells is advantageous to having a larger FLT3-itd clone in their population of AML cells.FLT3-itd positive patientsTotal (n)177Age (years)Median, range61, 17-89Sex (n)M:F87:88Presenting white cell count (×109/l)Median, range36, 0-388NPM1 mutation (n)Positive90Negative79Unknown7Treatment (n)Intensive133Non-intensive37Unknown6MRC Cytogenetic classification (n)Good26Intermediate143Adverse7 Disclosures:No relevant conflicts of interest to declare.

RUNX1 mutations are frequent in chronic myelomonocytic leukemia and mutations at the C-terminal region might predict acute myeloid leukemia transformation

Runt-related transcription factor 1 (RUNX1) is essential for normal hematopoiesis. RUNX1 mutations have rarely been reported in chronic myelomonocytic leukemia (CMML). We examined RUNX1 mutations in 81 patients with CMML at initial diagnosis. Mutational analysis was performed on bone marrow samples by direct sequencing of all reverse transcription PCR products amplified with three primer pairs that cover the entire coding sequences of RUNX1b. Thirty-two RUNX1 mutations were detected in 30 patients (37%); 23 mutants were located in the N-terminal part and 9 in the C-terminal region. The mutations consisted of 9 missense, 1 silent, 7 nonsense and 15 frameshift mutations. Two patients had biallelic heterozygous mutations. There was no difference in overall survival between patients with and without RUNX1 mutations, but a trend of higher risk of acute myeloid leukemia (AML) progression was observed in mutation-positive patients (16/30 vs 17/51, P=0.102), especially in patients with C-terminal mutations (P=0.023). The median time to AML progression was 6.8 months in patients with C-terminal mutations compared with 28.3 months in those without mutations (P=0.022). This study showed for the first time a high frequency of RUNX1 mutations in CMML. C-terminal mutations might be associated with a more frequent and rapid AML transformation.