Milos Kuzmanovic

Fever of unknown origin in 185 paediatric patients: A single‐centre experience

AIM We conducted a prospective study to evaluate the causes and outcome in children with fever of unknown origin (FUO). METHODS From 1990 to 1999, 185 children with FUO were evaluated. Initial evaluation included routine haematological analysis, Epstein-Barr virus (EBV) serology, urine, stool or blood cultures, chest X-ray and tuberculin probe. RESULTS In 131 (70%) patients diagnosis was established, and 70 (37.8%) had infectious disease. EBV infection was the most common infection followed by visceral leishmaniasis (VL), urinary tract infection (UTI) and tuberculosis. Autoimmune disorders were diagnosed in 24 (12.9%), Kawasaki disease in 12 (6.4%), malignant diseases in 12 (6.4%) and miscellaneous conditions in 15 (8.1%) patients. In the remaining 54 (30%) patients, diagnosis was not established and most of them had self-limited disease. During the investigation, 26 (14%) patients developed serious organ dysfunction and five patients (two with virus-associated haemophagocytic syndrome, one with VL and two unknown) died. CONCLUSION The most important infectious causes of FUO in our study were EBV infection and VL. Kawasaki disease represented a significant cause of FUO at the beginning of our study because it was not recognized by primary-care physicians. We report myelodysplastic syndrome as another emerging cause of paediatric FUO. Repeated clinical examination and careful use of specific laboratory examinations, invasive diagnostic procedures or imaging are crucial in approaching paediatric FUO.

Epstein-Barr virus-associated haemophagocytic lymphohistiocytosis in Wiskott-Aldrich syndrome.

A patient with Wiskott‐Aldrich syndrome who developed Epstein‐Barr virus‐associated haemophagocytic lymphohistiocytosis (EBV‐HLH) is described in this study. At 4 mo of age the patient developed fever associated with bicytopenia and splenomegaly. Analysis of a bone marrow specimen revealed extensive haemophagocytosis, and in situ hybridization for EBV of the bone marrow specimen using an EBV‐encoded RNA probe was positive. Diagnosis of EBV‐HLH was established and immunotherapy with HLH‐94 protocol was started. HLH has been described in patients with other well‐defined primary immunodeficiencies such as X‐linked lymphoproliferative syndrome, Chediak‐Higashi syndrome and Griscelli disease. Also, HLH was reported recently in severe combined immunodeficiency and DiGeorge syndrome.

Incidence of FLT3 and nucleophosmin gene mutations in childhood acute myeloid leukemia: Serbian experience and the review of the literature.

AbstractMutations in the fms-like tyrosine kinase 3 (FLT3) gene (internal tandem duplication (ITD) and point mutation in the tyrosine kinase domain, FLT3/D835) as well as the nucleophosmin (NPM1) gene are the most common abnormalities in adult acute myeloid leukemia (AML). Their significance in pediatric AML is still unclear. In this study we evaluated the frequency of FLT3 and NPM1 mutations in childhood AML. We also examined clinical features and outcome of these patients. FLT3 and NPM1 mutations were analysed in 42 and 37 childhood AML patients, respectively, using polymerase chain reaction (PCR) and direct sequencing. FLT3 mutations were detected in 4/42 patients (9.5%). The frequencies of FLT3/ITD and FLT3/D835 were the same, 2/42 (4.7%). NMP1 mutations were found in 1/37 patients (2.7%). FLT3 gene mutations were correlated with induction failure. Here we report the results of the study of FLT3 and NPM1 gene mutations in childhood AML patients in Serbia. Low frequencies of these molecular markers point out that these abnormalities are rare in this cohort of patients. Comparative study of data on NPM1 mutations in childhood AML revealed that various NPM1 gene mutation types are associated with childhood AML. Our findings as well as previously reported data, contributes to a hypothesis of different biology and etiology of adult and childhood AML. More extensive studies of NPM1 and FLT3 mutations in childhood AML are needed to determine their biological and clinical importance.

Glomerular involvement in myelodysplastic syndromes

Abstract. Several reports have documented various forms of glomerular diseases in adults with myelodysplastic syndromes (MDS), but similar reports in children are lacking. We describe two children with MDS-associated steroid-responsive nephrotic syndrome (NS). Patient 1, who had MDS with myelofibrosis, presented with hepatosplenomegaly, pancytopenia, chronic hepatitis, moderate proteinuria, hypocomplementemia and elevated ANA titer. During initial prednisone treatment proteinuria markedly diminished and partial but transient hematological improvement occurred. Relapse subsequently occurred that manifested by overt NS and pancytopenia. High doses of prednisolone led to remission of the renal disease, but hematological remission did not occur. Persisting pancytopenia and repeated infections terminated in sepsis, 2 years after the onset of the MDS. Patient 2, who had refractory anemia with clonal monosomy 19, presented with bowel disease, hepatosplenomegaly, anemia and non-organ-specific autoantibodies. Prednisone led to both clinical and hematological remission. The hematologic disease relapsed 12 months later, when nephrotic-range proteinuria, hematuria and mild azotemia were also found. Corticosteroid treatment led to long-lasting renal and hematologic remission, maintained by a small dosage of prednisone. In both patients, renal biopsy findings were consistent with those seen in idiopathic NS. A Medline search disclosed 16 cases of glomerulopathy in the course of MDS in adult patients. Clinical features included NS, usually accompanied by renal insufficiency with acute, chronic, or rapidly progressive glomerulonephritis. On biopsy, membranous nephropathy, crescentic or mesangial proliferative glomerulonephritis, and AL amyloidosis were found. We conclude: (1) that glomerular disease may be present and should be searched for in patients with MDS and (2) that MDS can be added to the list of rare conditions associated with corticosteroid-responsive NS in children.

Epstein-Barr virus associated hemophagocytic lymphohystiocytosis during maintenance treatment of acute lymphoblastic leukemia.

To the Editor: Hemophagocytic lymphohystiocytosis (HLH) is a hyperinflammatory syndrome caused by ineffective immune response to diverse stimuli, such as infections, metabolic products or tissue damage [1,2]. HLH occurs in patients with genetic background for this condition, in the setting of immunodeficiency, as well as in healthy individuals. Diagnostic criteria includes clinical, laboratory and histological data [3,4]. Our patient was a 9 years old boy who presented with high fever and pancytopenia during maintenance treatment of acute lymphoblastic leukemia (ALL). He is the second child of healthy, unrelated parents, with uneventful personal and family history. Nine months before this admission we made a diagnosis of preB ALL. Treatment according to BFM protocol induced complete hematological remission after a month, and further intensive chemotherapy took an uneventful course. In the second month of maintenance treatment the boy suffered high fever, accompanied with loose stools and pharyngitis. Physical finding, except sore throat, was completely normal. He had severe pancytopenia, and serum biochemistry revealed LDH 1747 U/L, AST 261 U/L, ALT 223 U/L, feritin of 1987 mcg/ L, triglycerides 3.05mmol/L.Also, he had slightly prolonged prothrombin time, but normal activated partial thromboplastin time and elevated level of fibrinogen of 5.45 g/L. Bone marrow aspirate was hypocelullar, without lymphoblasts, and phenomenon of hemophagocytosis was observed. Etiology of HLH was confirmed with the results of serological testing—concentration IgM on EBV was 170 U/ml (upper limit in our laboratory is 13 U/ml). NK cells in peripheral blood was almost absent (1.6%, normal value is 11%), and there was very low cytotoxic NK activity. We introduced treatment with cyclosporin A, dexametason and VP 16, according to HLH 2004 protocol. After starting this regimen fever resolved after 2 days and there was an improvement in complete blood counts and in general condition, as well. After the fourth dose of VP 16 fever reappearead, followed with the finding of distended and painful abdomen. Despite wide spectrum antimicrobials and G-CSF, the boy succumbed due to typhlitis and septicemia caused bymultiresistant Pseudomonas aeruginosa. HLH is a rare cause of fever in children during the treatment of malignant diseases. Although our patient did not fulfill all criteria for HLH (he had no splenomegaly and coagulopathy), persistent fever with unexplained pancyto-peniashould alert for diagnostic work up for HLH [5].

Severe central nervous system thrombotic events in hemoglobin Sabine patient

Hemoglobin (Hb) Sabine is a rare, unstable Hb variant resulting from the point mutation in codon 91 (CTG → CCG) of β‐globin gene. We report a case of Hb Sabine patient with mild hemolytic anemia, unusually high Hb F level and severe central nervous system thrombotic disturbances. We have tried to elucidate possible genetic background of this unusual Hb Sabine phenotype. Extremely high level of Hb F and rather mild anemia in our patient could be partially explained by the presence of Gγ Xmn I polymorphism. This case of Hb Sabine, unlike all other reported to date, shows extremely severe thromboembolic complications. It is our opinion that the hypercoagulable state described in thalassemia is not the only factor responsible for this specific clinical state. The presence of MTHFR C677T mutation in heterozygous state found in our patient and unstable Hb Sabine molecules could contribute to development of thromboembolic phenomena. However, it remains unclear whether other factors participate in pathogenesis of the disease. In this paper we emphasize different genetic background of father and son both affected with Hb Sabine, but with markedly different severity of the disease.

Leukapheresis in management hyperleucocytosis induced complications in two pediatric patients with chronic myelogenous leukemia.

Complications caused by elevated white blood cell count in pediatric patients with CML could be a presenting feature of the disease. Here, we present two adolescents, aged 16 and 17years, who were admitted for investigation of extremely elevated leukocytes and complications of leucostasis. Initial manifestations were priapism and blurred vision, respectively. Diagnosis of chronic phase of chronic myeloid leukemia is established, and conventional measures for leucoreduction began. However, since there were no improvements, a leukapheresis procedure was initiated. After undergoing 3 daily procedures the leukocyte count declined for each patient, with resolution of pripaism and ophtalmological disturbances. Leukapheresis is safe and effective therapeutic option for patients with complications of hyperleucocytosis. If started in a timely manner, permanent organ damage or death could be avoided.

Prognostic Impact of NPM1 Mutations in Serbian Adult Patients with Acute Myeloid Leukemia

Based on current findings, the presence of NPM1 mutations in acute myeloid leukemia (AML) patients is associated with an increased probability of complete remission (CR) and better overall survival (OS). We determined the incidence and prognostic relevance of NPM1 mutations, their association with FLT3 and IDH mutations, and other clinical characteristics in Serbian adult AML patients. Samples from 111 adult de novo AML patients, including 73 AML cases with a normal karyotype (NK-AML), were studied. NPM1, FLT3, and IDH mutations were detected by PCR and direct sequencing. NPM1 mutations were detected in 22.5% of patients. The presence of NPM1 mutations predicted a low CR rate and shorter OS. NPM1 mutations showed an association with both FLT3 and IDH mutations. Survival analysis based on NPM1/FLT3 mutational status revealed a lower OS for NPM1+/FLT3– compared to the NPM1–/FLT3– group in NK-AML patients. The lack of impact or unfavorable prognostic effect of NPM1 mutations found in this study can be assigned to a small cohort of analyzed AML patients, as can the presence of FLT3 and IDH mutations or other genetic lesions that cooperate with NPM1 mutations influencing prognosis.

Arterial Ischemic Stroke in a Child with β-Thalassemia Trait and Methylentetrahydrofolate Reductase Mutation

Genetic and acquired disorders that foster a procoagulable state represent risk factors for stroke in childhood. Although an increased incidence of thromboembolic complications has been reported in patients with thalassemia, severe cerebral thromboembolism has rarely been observed in patients with β-thalassemia minor. This article describes a case study of a 1-year-old boy who presented with left-sided hemiparesis, seizures, microcytic anemia, and recent infection with reactive thrombocytosis. Ischemic infarction in the territory of the right middle cerebral artery was confirmed by magnetic resonance imaging and magnetic resonance angiography. Genetic tests showed that the patient was heterozygous for the β°-thalassemia IVS-I-1 mutation and homozygous for the methylentetrahydrofolate reductase C677T mutation. Based on these findings, it was concluded that the synergistic effects of multiple, genetic, and acquired prothrombotic risk factors brought about the hypercoagulable state that resulted in overt stroke in a thalassemic patient in early childhood.

Bone marrow processing for transplantation using Cobe Spectra cell separator

Concentration of bone marrow aspirates is an important prerequisite prior to infusion of ABO incompatible allogeneic marrow and prior to cryopreservation and storage of autologous marrow. In this paper we present our experience in processing 15 harvested bone marrow for ABO incompatible allogeneic and autologous bone marrow (BM) transplantation using Cobe Spectra® cell separator. BM processing resulted in the median recovery of 91.5% CD34+ cells, erythrocyte depletion of 91% and volume reduction of 81%. BM processing using cell separator is safe and effective technique providing high rate of erythrocyte depletion and volume reduction, and acceptable recovery of the CD34+ cells.