Ismail Beshlawi

Optimizing Hydroxyurea use in children with sickle cell disease: Low dose regimen is effective

Hydroxyurea (HU) is the standard treatment for severely affected children with sickle cell disease (SCD). Starting dose is 15–20 mg/kg/day that can be escalated up to 35 mg/kg/day. Ethnic neutropenia is common in this area of the world that requires judicious usage of myelosuppressive drugs. Aim was to assess the efficacy of a lower initial dose of HU and cautious dose escalation regimen in patients with SCD.

dRTA and hemolytic anemia: first detailed description of SLC4A1 A858D mutation in homozygous state

Mutations in the anion exchanger 1 (AE1) gene encoding the erythroid and kidney anion (chloride–bicarbonate) exchanger 1 may result in familial distal renal tubular acidosis (dRTA) in association with membrane defect hemolytic anemia. Seven children presenting with hyperchloremic normal anion gap metabolic acidosis, failure to thrive, and compensated hemolytic anemia were studied. Analysis of red cell AE1/Band 3 surface expression by Eosin 5′‐maleimide (E5M) was performed in patients and their family members using flow cytometry. Genetic studies showed that all patients carried a common SLC4A1 mutation, c.2573C>A; p.Ala858Asp in exon 19, found as homozygous (A858D/A858D) mutation in the patients and heterozygous (A858D/N) in the parents. Analysis by flowcytometry revealed a single uniform fluorescence peak, with the mean channel fluorescence (MCF) markedly reduced in cases with homozygous mutation, along with a left shift of fluorescence signal but was only mildly reduced in the heterozygous state. Red cell morphology showed striking acanthocytosis in the homozygous state [patients] and only a mild acanthocytosis in heterozygous state [parents]. In conclusion, this is the first description of a series of homozygous cases with the A858D mutation. The E5M flowcytometry test is specific for reduction in the Band 3 membrane protein and was useful in conjunction with a careful morphological examination of peripheral blood smears in our patient cohort.

Thiamine responsive megaloblastic anemia: The puzzling phenotype

Thiamine responsive megaloblastic anemia (TRMA) is characterized by a triad of megaloblastic anemia, non‐type 1 diabetes mellitus and sensorineural deafness. Other clinical findings have been described in few cases. The SLC19A2 gene on chromosome 1q 23.3 is implicated in all cases with TRMA. Our aim is to discuss the clinical manifestations of all Omani children diagnosed with TRMA and determine genotype–phenotype relationship.

Brain is not always the last fortress; osteosarcoma with large brain metastasis.

Osteosarcomas are the most common malignant primary bone tumors in children and adolescents. Brain metastases of osteosarcoma are very rare and carry a dismal prognosis. We report a case of chondroblastic osteosarcoma of right humerus presented with right frontal lobe metastasis in a 10-year-old girl with small pulmonary lesions.

Severity ranking of non-deletional alpha thalassemic alleles: insights from an Omani family study

In an Omani family, four different alpha thalassemic alleles, one single‐gene deletional (−α3.7) and three non‐deletional forms (αTSaudi, αΔ5nt, and αΔG), interact in various combinations and result in two distinct hematological phenotypes, with and without HbH inclusions. After excluding the presence of potential genetic modifiers, viz associated β‐thalassemic alleles or functional alpha hemoglobin stabilizing protein (AHSP) polymorphisms, we observed that only the genetic combinations involving αTSaudi mutation are associated with HbH inclusions (a marker of degree of α/β‐chain imbalance) and high reticulocyte count (a marker of ongoing hemolysis). Overall, the αTSaudi mutation is associated with a more severe α‐globin deficiency than the other two (αΔ5nt and αΔG) non‐deletional α0 thalassemic mutations. The likely molecular explanation is that the compensatory increase in the linked α1 globin gene expression is much more compromised in cases with αTSaudi mutation.

Echocardiography in PICU: when the heart sees what is invisible to the eye

OBJECTIVE Echocardiography has become an indispensable bedside diagnostic tool in the realm of pediatric intensive care units (PICU). It has proven to be an influential factor in the formula of clinical decision-making. This study aimed to delineate the impact of echocardiography on the management of critically ill pediatric patients in the PICU at Sultan Qaboos University Hospital, Oman. METHOD This was a retrospective cohort study conducted in a five-bed PICU. Patients admitted to the PICU from January of 2011 to December of 2012 were reviewed. Those who have undergone bedside echocardiography during their ICU stay were recruited. Electronic patient record was used as data source. RESULTS Over a-24-month period, 424 patients were admitted in this PICU. One hundred and one clinically indicated transthoracic echocardiograms were performed. 81.8% of these presented new findings (n=82) that significantly impacted the clinical decision of patient management, namely, alteration in drug therapy and procedure, whereas no difference in the management was yielded in the remaining 17.8% of the studied cases. CONCLUSIONS Echocardiography had a significant impact on the management of PICU patients. Such salutary effect was consequently reflected on the outcome. Pediatric intensivists are encouraged to acquire such bedside skill.

BCG lymphadenitis in neonates with familial hemophagocytic lymphohistiocytosis.

To the Editors: Familial hemophagocytic lymphohistiocytosis (FHLH) is an autosomal recessive immune disorder characterized by fever, hepatosplenomegaly, pancytopenia, hypertriglyceridemia, hypofibrinogenemia, markedly elevated inflammatory cytokines, and impaired cytotoxic activity of lymphocytes. This disease is invariably fatal in infancy if the treatment is not initiated immediately with specific chemotherapy. In Oman, we have previously reported a number of families (10) with known FHLH mutation, mainly in the perforin gene, and our patients commonly present in the first 2 months of life. Due to the high rate of consanguinity and interfamilial marriage in Oman, almost all the new cases of hemophagocytic lymphohistiocytosis can be traced to these families. BCG is a live attenuated bacterial vaccine that is included in the expanded program on immunization in Oman, which is administered on the first day of life. We recently treated 2 cases (male, 1 week old and female, 9 weeks old) coming from known families, fulfilling all the diagnostic criteria of FHLH, and both of them have no perforin by FACS analysis and perforin gene mutations (674G3C(Arg225Pro)). After initiating chemotherapy as per HLH 2004 protocol, they developed severe BCG adenitis confirmed by excisional biopsy. They needed triple antituberculous therapy for 3 months and continued to receive isoniazid and rifampin for 1 year. This has delayed the definitive treatment of these patients with bone marrow transplant until the adenitis resolves. As these patients came from known high risk families of FHLH, this complication could have been prevented by proper counseling, highlighting the need to screen the newborns of such families for FHLH known mutation before administering BCG vaccine. We recommend delaying the BCG vaccine for these patients until recovery of their immune system.

Coexistence of sickle cell disease and severe congenital neutropenia: first impressions can be deceiving

We report an Omani family in whom the propositus had a rare coexistence of sickle cell disease and severe congenital neutropenia associated with a mutation in ELANE. In contrast to his siblings with sickle cell disease, the severity of HbSS‐associated complications such as painful crises and acute chest syndrome was significantly reduced. His course of the disease had markedly worsened after initiating G‐CSF therapy. These clinical observations suggest that neutropenia may ameliorate inflammatory responses and thus display a modulating factor with respect to the clinical course of sickle cell disease.

Wolman's Disease with Secondary Hemophagocytic Lymphohistiocytosis

We describe a 2-month-old male presenting with one week history of fever, irritability and pallor. He had history of progressive abdominal distension since the age of 2 weeks. The child was born by spontaneous vaginal delivery at full term following a normal pregnancy. His birth weight was 2.9 kg. He was the first child of first-degree consanguineous parents with no significant family history. On examination coarse facial features, proptosis, jaundice, tachypnea, and tachycardia were noted. The child had grossly distended abdomen with umbilical hernia and dilated veins. Liver was palpable 10 cm below the costal margin while the spleen was felt12 cm below the left costal margin. Broad-spectrum antibiotic therapy was initiated empirically for sepsis. Initial investigations revealed the following: hemoglobin 6.9 g/dL, WBC 11.9 × 109/L, ANC 3.7 × 109/L, platelet count 73 × 109/L, alanine transaminase 38 iu/L, total bilirubin 19 umol/L, conjugated bilirubin 13 umol/L, LDH 3290 u/L, ferritin 5600 ug/L, PT >120 seconds, APTT 63.2 seconds, cross-linked D-dimer > 10 mg/L (N 0.2–0.7 mg/L), fibrinogen 1.24 g/L, triglycerides 18 mmol/L (N 0.0–2.3 mmol/L), and albumin 18 g/L. Blood and urine cultures, TORCH and EBV serology studies were negative. The abdominal radiograph showed bilateral adrenal calcification (Figure 1A). MRI brain unveiled mild brain atrophy. With the constellation of splenomegaly, fever, bicytopenia, hyperferritinemia, hypertriglyceridemia and hypofibrinogenemia, and hemophagocytic lymphohistiocytosis (HLH) was suspected. Therefore, the patient was posted for bone marrow aspirate that revealed active marrow with numerous macrophages occasionally showing hemophagocytosis, while the majority of cells have soap bubble like cytoplasm (Figure 1B). The leukocytic cholesteryl esterase assay made the diagnosis of Wolman Disease (WD); 0–0.1 nmol/17 hours/mg Protein (Normal Range: 15–20). WD is a rare autosomal recessive lysosomal disorder characterized by generalized lipid storage with no reported ethnic association [1, 2]. WD has been previously reported among Saudis, Bahrainis, Palestinians, and Iraqi Jews [3, 4]. Sequencing LIPA gene did not yield a mutation. However, it does not rule out Wolman disease as the patient has severe enzyme deficiency with adrenal calcification. The gap in

High incidence of intracranial bleeding in factor V-deficient patients with homozygous F5 splicing mutations.

Coagulation factor V (FV) deficiency (Huang & Koerper, 2008) is a rare autosomal recessive bleeding disorder caused by mutations in the F5 gene. Symptoms range from mucosal and post-traumatic bleeding, which are present in virtually all patients, to life-threatening intracranial and gastro-intestinal haemorrhages, which are rare and confined to the most severe cases (Lak et al, 1998; Huang & Koerper, 2008). As no FV concentrate is available, standard treatment relies on the administration of fresh frozen plasma (FFP). Currently, 150 different F5 mutations have been described, most of which are missense or nonsense (Human Genome Mutation Database, www.hgmd.cf.ac.uk). Splicing mutations represent only ~10% of the F5 mutational spectrum, but this is probably an underestimate, as splicing mutations outside the canonical splice sites are not readily recognized (Castoldi et al, 2011; Nuzzo et al, 2015). Here we report a novel F5 splicing mutation responsible for a particularly severe case of FV deficiency and we present evidence, based on a review of the literature, that F5 splicing mutations tend to confer severe bleeding tendencies. An Omani baby girl, born to consanguineous parents (Fig 1A), came to clinical attention because of multiple episodes of intracranial bleeding in the first days of life, which led to the diagnosis of severe FV deficiency (FV:C = FV FV coagulant activity). Despite immediate prophylaxis with FFP twice weekly, she continued to experience breakthrough intracranial haemorrhages, prompting an upgrade of her prophylactic regimen to three times weekly. After three years on this regimen, treatment returned to two plasma transfusions per week with no further bleeding for the past 21⁄2 years. Following written informed consent, blood was collected from the proband and her family members at Sultan Qaboos University Hospital (Muscat, Sultanate of Oman) in compliance with the Helsinki Declaration. Citrated plasma and genomic DNA were shipped to Maastricht (The Netherlands) for analysis. F5 gene sequencing in the proband revealed a novel homozygous mutation (F5 IVS3+2T>C, c.373+2T>C), affecting the second nucleotide of intron 3 (Fig 1B). Both parents and the proband’s eldest sister were heterozygous for this mutation, whereas the proband’s other sister had a normal genotype. The observed F5 IVS3+2T>C genotypes were consistent with the plasma FV antigen (FV:Ag) and activity levels determined by enzyme-linked immunosorbent assay (Zymutest Factor V, NODIA, Amsterdam, The Netherlands) and a prothrombinase-based assay, respectively (Fig 1A). In particular, FV activity was undetectable in the proband’s (platelet-poor) plasma by a prothrombinase-based assay and by measuring thrombin generation at 10 pM tissue factor (not shown). The F5 IVS3+2T>C mutation affects positon +2 of the canonical donor splice site of intron 3 (Fig 1C). In silico analysis indicated that the mutation reduces the donor splice site consensus score from 0 99 to undetectable (according to the NNsplice software; http://www.fruitfly.org/seq_tools/splice.html) or from 0 96 to 0 69 (according to the Human Splicing Finder software; http://www.umd.be/HSF3/), predicting an effect on pre-mRNA splicing. Unfortunately, the splicing defect associated with the F5 IVS3+2T>C mutation could not be studied experimentally due to the lack of patient material suitable for RNA isolation and to the technical challenge of cloning a F5 minigene containing intron 3 (>11 kb). However, the most likely consequence of this mutation is exon 3 skipping, an aberrant splicing event that also occurs at a low rate in the wild-type F5 transcript (Dall’Osso et al, 2008). The resulting mRNA would maintain the correct reading frame, but it would encode a putative FV mutant lacking 41 amino acids in the A1 domain, which is unlikely to be correctly folded and secreted. The unusually severe bleeding phenotype of the proband of this study prompted us to review the literature for the relationship between splicing mutations and bleeding symptoms in FV deficiency. A clinically well-characterized cohort of 35 genetically unselected FV-deficient patients (Lak et al, 1998) served as a reference group. A literature search retrieved seven cases of FV deficiency attributable to homozygous splicing mutations, including the present report (Table I). Plasma FV activity levels were undetectable (<1%) or barely detectable (1 6%) in 6/7 patients, the remaining patient having moderate FV deficiency (5% FV). All patients with homozygous splicing mutations, except the one with 5% FV, showed remarkably severe clinical histories in terms of early onset and severity of bleeding symptoms. In particular, 3/7 patients with homozygous splicing mutations (42 9%) had been diagnosed at birth as opposed to only 1/35 unselected patients (2 9%). Moreover, 5/7 patients with homozygous splicing mutations (71 4%) had experienced one or more episodes of intracranial haemorrhage. Although numbers are small and do not allow proper statistics, this figure is quite striking considering that intracranial bleeding is rather uncommon in unselected Correspondence