Identification of the molecular etiology in rare congenital hemolytic anemias using next-generation sequencing with exome-based copy number variant analysis.

OBJECTIVES: In congenital hemolytic anemias (CHA), it is not always possible to determine the specific diagnosis by evaluating clinical findings and conventional laboratory tests. The aim of this study is to evaluate the utility of next-generation sequencing (NGS) and clinical-exome-based copy number variant (CNV) analysis in patients with CHA. METHODS: One hundred and forty-three CHA cases from 115 unrelated families referred for molecular analysis were enrolled in the study. Molecular analysis was performed using two different clinical exome panels in 130 patients, and whole-exome sequencing in nine patients. Exome-based CNV calling was incorporated into the traditional single-nucleotide variant and small insertion/deletion analysis pipeline for NGS data in 92 cases. In four patients from the same family, the PK Gypsy variant was investigated using long-range polymerase chain reaction. RESULTS: Molecular diagnosis was established in 86% of the study group. The most frequently mutated genes were SPTB (31.7%) and PKLR (28.5%). CNV analysis of 92 cases revealed that three patients had different sizes of large deletions in the SPTB and six patients had a deletion in the PKLR. CONCLUSIONS: In this study, NGS provided a high molecular diagnostic rate in cases with rare CHA. Analysis of the CNVs contributed to the diagnostic success.

A rare cause of intestinal obstruction in children: signet-ring cell adenocarcinoma of the colon.

BACKGROUND: Signet-ring cell adenocarcinoma of the colon is well-recognized in adult patients who are extremely rare and not well-documented in children. Our study aims to raise awareness about this rare disease and its long-term outcomes. METHODS: We retrospectively evaluated patients with signet-ring cell colon adenocarcinoma. RESULTS: Six patients, three boys and three girls, with a mean age of 14.83 (range, 13-17 years), presented with signs of intesti-nal obstruction and were diagnosed with signet-ring cell colon adenocarcinoma. All patients had air-fluid levels on abdominal X-ray. Abdominal ultrasonography of all patients revealed subileus. Abdominal computed tomography was performed in five patients, and pre-operative colonoscopy was conducted in two patients before the emergency intervention. All of the patients underwent emergent exploratory laparotomy with the preliminary diagnosis of acute abdomen. In two patients, debulking surgery followed by a stoma was performed. The remaining four patients were treated with anastomosis following intestinal resection. All girls had metastases on the ovary. One of the patients died due to the burden of multiple metastases in the early period, and three died in the sixth post-operative year. We have been following the remaining two patients since then. CONCLUSION: Although signet-ring cell carcinomas (SRCCs) are rare, they should be considered in the differential diagnosis of acute abdomen and intestinal obstruction in pediatric patients. Despite early diagnosis and treatment, SRCC has a poor prognosis in the pediatric population.

Prognostic Factors and Long-term Outcomes in 41 Children With Primary Hemophagocytic Lymphohistiocytosis: Report of a Single-center Experience and Review of the Literature.

Hemophagocytic lymphohistiocytosis (HLH) is a rare life-threatening hyperinflammatory syndrome with diverse clinical manifestations leading to major diagnostic and therapeutic difficulties. This study aimed to evaluate clinical manifestations, prognostic factors, and long-term outcomes in children with primary HLH. Forty-one patients diagnosed with primary HLH were retrospectively evaluated for patient characteristics, HLH gene mutations, clinical and laboratory manifestations, prognostic factors, and long-term outcomes. The median age of the patients at the time of diagnosis was 3 months (minimum to maximum: 1 to 144 mo). There were 23 patients who had HLH mutation analysis performed, 10 patients with PRF1 mutation, 6 with STX11 mutation, and 7 with UNC13D mutation. Thirteen patients (31.7%) had central nervous system involvement. No correlation was found between overall survival and central nervous system involvement. The estimated 5-year overall survival for the patient who had hematopoietic stem cell transplantation was 9.4 times better than the patients who did not receive hematopoietic stem cell transplantation (81.3% vs 16.7%; P = 0.001). Median serum sodium and blood urea nitrogen levels were significantly higher in deceased HLH patients compared with surviving HLH patients ( P = 0.043, and P = 0.017, respectively). Primary HLH has a poor outcome with high mortality, which necessitates well-designed and international clinical trials to improve diagnosis, therapy, and long-term outcomes.

Vaccine Adherence and Postvaccination Serological Status of Pediatric Allogeneic Hematopoietic Stem Cell Transplant Recipients: A Single-center Experience.

Despite developing consensus guidelines addressing immunization after hematopoietic stem cell transplantation (HSCT), studies showed deviations from recommended immunization practices commonly occur. Difference between the ideal scenario presented in guidelines and real-life scenarios is one of the most recognized barriers to implementing recommended practices. Therefore, this study aimed to evaluate pediatric allogeneic HSCT recipients' adherence to revaccination schedule and evaluate the serological status after immunization. Transplant and vaccination records of children who were followed up at least 2 years after HSCT, postvaccination antibody results of vaccine-preventable diseases were evaluated retrospectively. Total of 173 patients have enrolled in this study. Median revaccination onset time was post-transplant 15 months. Adherence to revaccination program was 30% for inactive and 11.4% for live vaccines. Oral polio vaccine was given to 22 patients, and Bacille-Calmette-Guerin vaccine was applied to 3. Seropositivity after revaccination was >90% for Hepatitis B, Hepatitis A, pertussis, and measles, and it was 88.5% for rubella, 80% for mumps and varicella. Measles seropositivity was low in children with hemoglobinopathy. In subgroup assessments of pertussis, patients vaccinated with low antigen-containing pertussis vaccine (Tdap) had higher seropositivity of adenylate cyclase toxin. Our findings revealed the importance of careful monitoring of current practices in pediatric HSCT recipients.

Central Nervous System Fungal Infections in Children With Leukemia and Undergoing Hematopoietic Stem Cell Transplantation: A Retrospective Multicenter Study.

BACKGROUND: Central nervous system fungal infections (CNSFI) are seen in patients with hematologic malignancies and have high morbidity and mortality. Because of their rarity, there is limited data on CNSFI in children with no established treatment protocols or guidelines. MATERIALS AND METHODS: In this multicenter retrospective study, 51 pediatric patients with leukemia, 6 of whom had undergone bone marrow transplantation, with proven or probable CNSFI were evaluated. Fungal infections were defined as proven or probable based on European Organisation for Research and Treatment of Cancer criteria. Proven CNSFI was diagnosed by appropriate central nervous system (CNS) imaging or tissue sample findings in combination with positive microbiological results of cerebrospinal fluid. A positive culture, microscopic evidence of hyphae, a positive result of the galactomannan assays are defined as positive microbiological evidence. Probable CNSFI was defined as appropriate CNS imaging findings together with proven or probable invasive fungal infections at another focus without CNS when there is no other explanatory condition. Data was collected by using the questionnaire form (Supplemental Digital Content 1, http://links.lww.com/JPHO/A541 ). RESULTS: Seventeen patients had proven, 34 patients had probable CNSFI. Headaches and seizures were the most common clinical findings. The median time between the onset of fever and diagnosis was 5 days. The most common fungal agent identified was Aspergillus . Sixteen patients received single-agent, 35 received combination antifungal therapy. Surgery was performed in 23 patients. Twenty-two patients (43%) died, 29 of the CNSFI episodes recovered with a 20% neurological sequelae. CONCLUSION: CNSFIs should be considered in the differential diagnosis in patients with leukemia and refractory/recurrent fever, headache, neurologicalocular symptoms, and a radiologic-serological evaluation should be performed immediately. Early diagnosis and prompt management, both medical and surgical, are essential for improving clinical outcomes.

Bladder granulocytic sarcoma in a child: case report and literature review.

BACKGROUND: Granulocytic sarcoma (GS) is an extramedullary solid tumor composed of immature myeloid cells. GS has been associated with acute myeloid leukemia (AML), myelodysplastic syndromes or myeloproliferative diseases. Although GS can affect various tissues of the human body, it has rarely been reported in other soft tissues such as the breast, gastrointestinal, respiratory and genitourinary tracts. We report a pediatric case diagnosed with granulocytic sarcoma of the bladder and concomitant AML. CASE: A twelve-year-old previously healthy girl was admitted to the pediatric urology clinic with a ten-day history of hematuria and pollakiuria. Laboratory examinations revealed anemia, thrombocytopenia and neutrophilic leukocytosis. Bone marrow aspiration results were consistent with acute myeloid leukemia -FAB subtype M2-. Abdominal magnetic resonance imaging (MRI) showed an irregularly bounded 12 cm mass on the right side of the bladder. Transurethral resection (TUR) pathology was consistent with granulocytic sarcoma. After a multimodal treatment approach, complete remission was achieved. CONCLUSIONS: Malignant bladder masses are rare causes of macroscopic hematuria in childhood. The diagnostic spectrum is wide, ranging from rhabdomyosarcoma to leukemia involvement. The bladder is a rare site of extramedullary involvement in pediatric patients with AML. Multimodal treatment should be considered on a per-patient basis.

Hepatitis B Vaccination in Children With Ongoing Cancer Treatment: A Safety and Efficacy Study of Super-Accelerated Vaccination Scheme.

OBJECTIVE: Children with cancer have an increased risk for hepatitis B virus (HBV) infections due to chemotherapy-induced secondary immunodeficiency and frequent blood transfusions. The aim of this study is to evaluate the efficacy and safety of hepatitis B vaccination during the intensive induction chemotherapy in children with cancer found to be seronegative for hepatitis B on admission. MATERIALS AND METHODS: Children newly diagnosed with cancer were evaluated for the presence of hepatitis B surface antigen (HBsAg) and antibody on admission. The children negative for both were included in the study. A super-accelerated vaccination scheme (3 booster doses at days 1-5, 8-12, and 28-33) was administered to these seronegative children concurrently with induction chemotherapy. Antibody response was checked 4-8 weeks after the last vaccination and 6 months after the end of the treatment. RESULTS: Eleven out of 122 children were seronegative for hepatitis B on admission (9%). Acute lymphoblastic leukemia, lymphoma, and solid tumors were diagnosed in 5, 4, and 2 children, respectively. Complete seroconversion was achieved in 4-8 weeks after the last vaccination with high titers of anti-HBs antibody, and all patients remained antibody-positive until 6 months after the completion of chemotherapy. CONCLUSION: The risk of transfusion-related infections increases with a number of transfused products and donor exposures, and it is more significant for immunosuppressed children with hematologic and oncologic malignancies. Hepatitis B vaccination could safely be applied with brisk and sustained responses in this vulnerable population, based on the local epidemiological data.

Congenital Factor XIII Deficiency With the Presence of Inhibitor: A Case Study.

Coagulation factor XIII (FXIII) is a fibrin-stabilizing factor with additional roles in wound healing and interactions between the decidua and fetus. Congenital FXIII deficiency is rare bleeding disorder. Inhibitor development against FXIII in inherited FXIII deficency is also uncommon, but may cause severe, life-threatening bleeding. FXIII is the last step in the coagulation cascade with normal coagulation paramaters (PT, aPTT), the detection of inhibitor to FXIII is quite difficult. The treatment of inhibitor-positive congenital FXIII deficiency is challenging due to the lack of a role of by-pass agents such as FVII. The best known ways of treatment in these cases are the use of high-dose FXIII concentrates and immunosuppression. Herein, we report the management of postoperative bleeding diathesis in a patient with FXIII deficiency who developed inhibitors, and to follow the clinical course of the disease with FXIII concentrate and immunosuppression.

Intrapericardial pneumonectomy for unicentric hilar castleman disease.

INTRODUCTION: The intrapulmonary involvement of Castleman disease is extremely rare with very few cases reported in the literature. CASE REPORT: We present a case of a 16-year-old male who underwent pulmonary resection for Castleman disease. The tumor was located at the right hilum and measured 10 cm in diameter. The patient underwent an intrapericardial pneumonectomy with a mediastinal lymph node dissection because the tumor showed infiltration into the lung parenchyma, the major pulmonary vessels and the main bronchus. CONCLUSION: We emphasize that some cases with intrapulmonary Castleman disease may require a pneumonectomy if the tumor is centrally located.

Mixed Langerhans Cell Histiocytosis and Erdheim-Chester Disease in a Girl: A Rare and Puzzling Diagnosis.

OBJECTIVE: The objective of this study was to report the case of a girl diagnosed as suffering from multisystem, BRAF V600E-positive refractory Langerhans cell histiocytosis (LCH) and coexistent Erdheim-Chester disease (ECD) with perirenal, intracranial involvement and the dramatic response to clofarabine treatment. OBSERVATIONS: Histiocytoses are rare diseases with a broad clinical spectrum. Recent evidence supports a molecular and clinical overlap between LCH and ECD, and mixed LCH/ECD is now a separate entity. However, only a few pediatric cases of mixed disease have been reported in the literature. CONCLUSIONS: In a child with refractory, multisystem histiocytosis and atypical presentations, mixed LCH/ECD should be suspected in the differential diagnosis.

The Role of Anti-Neutrophil Antibodies in the Etiologic Classification of Childhood Neutropenia: A Cross-Sectional Study in a Tertiary Center.

Infections, drugs, malignancies, immunodeficiency, and autoimmunity may cause neutropenia. In primary autoimmune neutropenia, anti-neutrophil antibodies (ANeuA) bind to membrane antigens of neutrophils, which give rise to peripheral destruction of neutrophils. However, it is not always easy to detect these antibodies. This study aims to investigate the etiology of neutropenia, and at the same time to evaluate the immune mechanisms by ANeuA testing using granulocyte indirect immunofluorescence test. In our study, 310 neutropenic patients who were between 3 months and 18 years of age were evaluated. ANeuA screening tests were performed in 108 neutropenic patients (group 1), and these patients were divided into 2 subgroups as persistent neutropenia (group 1P, n=12) and recovered neutropenia (group 1R, n=96). Besides, a control group in the same age range was formed, consisting of 39 non-neutropenic children (group 2). ANeuA serum levels were also checked in these groups, and no statistically significant difference could be found between groups 1 and 2, or between groups 1P and 1R, regarding ANeuA levels. As a conclusion, our study was the first comprehensive research in Turkey investigating the large-scale etiology of neutropenia. Moreover, while ANeuA screening tests did not provide sufficient insight for immune neutropenia, we argue that it is not necessary for routine use and that further research in the etiology of neutropenia is required.

Hepatosplenic Fungal Infections in Children With Leukemia-Risk Factors and Outcome: A Multicentric Study.

BACKGROUND: Invasive fungal infections, including hepatosplenic fungal infections (HSFI), cause significant morbidity and mortality in children with leukemia. There are not enough data to support for the best approach to diagnosis of HSFI in children, nor for the best treatment. PROCEDURE: In this multicentric study, we assessed the demographic data, clinical and radiologic features, treatment, and outcome of 40 children with leukemia and HSFI from 12 centers. RESULTS: All cases were radiologically diagnosed with abdominal ultrasound, which was performed at a median of 7 days, of the febrile neutropenic episode. Mucor was identified by histopathology in 1, and Candida was identified in blood cultures in 8 patients. Twenty-two had fungal infection in additional sites, mostly lungs. Nine patients died. Four received a single agent, and 36 a combination of antifungals. CONCLUSIONS: Early diagnosis of HSFI is challenging because signs and symptoms are usually nonspecific. In neutropenic children, persistent fever, back pain extending to the shoulder, widespread muscle pain, and increased serum galactomannan levels should alert clinicians. Abdominal imaging, particularly an abdominal ultrasound, which is easy to perform and available even in most resource-limited countries, should be recommended in children with prolonged neutropenic fever, even in the absence of localizing signs and symptoms.

Diagnostic performances of superb microvascular imaging, shear wave elastography and shape index in pediatric lymph nodes categorization: a comparative study.

OBJECTIVE: To determine the diagnostic utility of a vascularity index via superb microvascular imaging in lymph nodes of children with malignant lymphoma and acute lymphadenitis compared to normal lymph nodes. METHODS: We performed a retrospective study for multiparametric lymph node (LN) evaluation. Malignant lymphoma diagnosed via histopathological examination and lymph nodes receiving an acute lymphadenitis diagnosis based on clinical and laboratory findings constituted the study subgroups. We calculated a shape index [SI (percent of shortest to longest diameter)] using grayscale ultrasonography and elasticity and velocity values via shear wave elastography (SWE) as well as a vascularity index (VI) using superb microvascular imaging (SMI) for comparison with normal lymph nodes. RESULTS: 45 lymph nodes diagnosed with malignant lymphoma, 72 lymph nodes diagnosed with acute lymphadenitis and 146 normal lymph nodes were evaluated. For differentiating lymphoma from normal lymph nodes, vascularity index cut-off values higher than 15% represented a diagnostic accuracy of 95%; cut-off elasticity values higher than 17 kPa exhibited a diagnostic accuracy of 99%. Optimal VI, elasticity, velocity and SI cut-off values in differentiating lymphoma from lymphadenitis were 15%, 17 kPa, 2.45 m sn-1 (p < 0.001) and 65% (p < 0.002) with calculated diagnostic accuracies of 83, 87, 88 and 68%, respectively. CONCLUSION: Vascularity index values obtained via superb microvascular imaging and SWE would be reasonably useful in differentiating malignant lymphoma and acute lymphadenitis from normal LNs. SWE would be more efficient in distinguishing malignant lymph nodes from acute lymphadenitis compared with superb microvascular imaging. Advances in knowledge: Vascularity index by superb microvascular imaging would be a novel Doppler parameter in differentiating both lymphoma from lymphadenitis and also lymphadenitis from normal lymph nodes.

Magnetic resonance imaging during management of patients with transfusion-dependent thalassemia: a single-center experience.

BACKGROUND: Cardiac and hepatic magnetic resonance imaging evaluation during treatment can tailor physicians' chelation therapy titrations. AIM: The aim of the study was to assess the relationship of cardiac and hepatic T2* values with chelation therapy in patients with transfusion-dependent thalassemia (TDT). METHODS: A total of 106 patients with TDT who were followed up in Istanbul Medical Faculty Thalassemia Center were evaluated for the study. Forty-eight (45%) patients with TDT had more than one consecutive MRI examination. The patients were divided into three subgroups according to the cardiac T2* values as the high-risk group (T2* MRI < 10 ms), medium-risk group (T2* MRI 10-20 ms), and the low-risk group (T2* MRI > 20 ms). RESULTS: The majority of patients used DFX (deferasirox) (79%) and deferiprone (DFP) (17%). Approximately 80% of patients according to cardiac T2* value and 40% of patients according to hepatic T2* value were initially in the low-risk group. Patients with follow-up MRI examinations exhibited significant improvement in liver iron concentration, which correlated with an increase in hepatic T2* values. The decrease of liver iron concentration was prominent in the DFX group (p < 0.01). The serum ferritin level was significantly correlated with liver iron concentrations (rs = 0.65, p < 0.001), hepatic T2* value (rs = - 0.62, p < 0.001), but not with cardiac T2* value (rs = - 0.20, p = 0.07). CONCLUSION: Cardiovascular and hepatic MRI is a useful follow-up tool during the assessment of risk groups and chelation therapy of patients with TDT. Consecutive MRI tests showed good monitoring of cardiac and liver iron overload.

Congenital Factor Deficiencies in Children: A Report of a Single-Center Experience.

Congenital factor deficiencies (CFDs) refer to inherited deficiency of coagulation factors in the blood. A total of 481 patients with CFDs, who were diagnosed and followed at our Pediatric Hematology and Oncology Clinic between 1990 and 2015, were retrospectively evaluated. Of the 481 cases, 134 (27.8%) were hemophilia A, 38 (7.9%) were hemophilia B, 57 (11.8%) were von Willebrand disease (vWD), and 252 (52.3%) were rare bleeding disorders (RBDs). The median age of the patients at the time of diagnosis and at the time of the study was 4.1 years (range: 2 months to 20.4 years) and 13.4 years (range: 7 months to 31.3 years), respectively. The median duration of the follow-up time was 6.8 years (range: 2.5 months to 24.8 years). One hundred nineteen (47.2%) of 252 patients with RBDs were asymptomatic, 49 (41.1%) of whom diagnosed by family histories, 65 (54.6%) through preoperative laboratory studies, and 5 (4.2%) after prolonged bleeding during surgeries. Consanguinity rate for the RBDs was 47.2%. Prophylactic treatment was initiated in 80 patients, 58 of whom were hemophilia A, 7 were hemophilia B, 13 were RBDs, and 2 were vWD. Significant advances have been achieved during the past 2 decades in the treatment of patients with CFDs, particularly in patients with hemophilias. The rarity and clinical heterogeneity of RBDs lead to significant diagnostic challenges and improper management. In this regard, multinational collaborative efforts are needed with the hope that can improve the management of patients with RBDs.

Olfactory dysfunction and quality of life in patients with transfusion-dependent thalassemia.

Transfusion-dependent thalassemia (TDT) is a group of thalassemia syndromes that require regular blood transfusions for survival. It is unknown whether the sense of smell of patients with TDT is affected, and if so, whether smell loss has an adverse effect on quality of life (QOL). Olfactory thresholds were measured using Snap & Sniff® wands. QOL was assessed via the Short Form-36 (SF-36) questionnaire. Forty-two TDT patients from the Thalassemia Center in Istanbul Medical Faculty were tested (mean age 26.6 years), along with 42 healthy controls (mean age 28.0 years). Mean olfactory sensitivity was lower in the TDT patients than the controls (p < 0.0001). TDT was associated with lower mean QOL scores on the domains of physical function (p < 0.0001), physical role limitation (p = 0.011), and general health (p < 0.0001). Within the TDT group, significant correlations were present between the threshold scores and physical function, physical role limitation, emotional role limitation, and general health. Patients with TDT are less sensitive to odors than healthy controls and the lower olfactory test scores are related to lower quality of life, suggesting that decreased smell function is an additional complication of this disease.