The role of magnetic resonance imaging in the diagnosis of central nervous system involvement in children with acute lymphoblastic leukemia.

PURPOSE: Acute lymphoblastic leukemia (ALL) is the most frequent malignancy in childhood. As central nervous system (CNS) involvement requires an intensified CNS-targeted therapy, timely diagnosis is essential. The aim of this retrospective analysis was to evaluate whether cranial magnetic resonance imaging (MRI) examinations findings correlate with cerebrospinal fluid (CSF) analysis on CNS involvement and whether MRI examinations reveal incidental findings with a clinical consequence. METHODS: All pediatric patients with ALL at our institution between 1998 and 2016 were identified. Patients were divided into two groups: de novo and relapsed ALL. Both groups were analyzed separately for the presence of CNS involvement. Incidental findings were also evaluated. RESULTS: Two hundred fifteen patients with de novo ALL and 31 with relapsed ALL were identified. In the de novo group, no patient was diagnosed CNS positive based on MRI results alone. In relapsed patients, only one patient had a positive MRI with negative CSF results and no neurological symptoms, thus was classified CNS positive solely on the basis of the MRI. In both groups, no patient showed an incidental finding that required therapy. CONCLUSION: In our study, MRI examinations do not improve the detection of CNS involvement compared with CSF analysis alone. In addition, the analysis of incidental findings does not add value to the performance of an MRI examination performed prior to treatment. Overall, MRI prior to treatment in pediatric patients with ALL is not necessary.

Frequency and epitope specificity of anti-factor VIII C1 domain antibodies in acquired and congenital hemophilia A.

Several studies showed that neutralizing anti-factor VIII (anti-fVIII) antibodies (inhibitors) in patients with acquired hemophilia A (AHA) and congenital hemophilia A (HA) are primarily directed to the A2 and C2 domains. In this study, the frequency and epitope specificity of anti-C1 antibodies were analyzed in acquired and congenital hemophilia inhibitor patients (n = 178). The domain specificity of antibodies was studied by homolog-scanning mutagenesis (HSM) with single human domain human/porcine fVIII proteins and antibody binding to human A2, C1, and C2 domains presented as human serum albumin (HSA) fusion proteins. The analysis with HSA-fVIII domain proteins confirmed the results of the HSM approach but resulted in higher detection levels. The higher detection levels with HSA-fVIII domain proteins are a result of antibody cross-reactivity with human and porcine fVIII leading to false-negative HSM results. Overall, A2-, C1-, and C2-specific antibodies were detected in 23%, 78%, and 68% of patients with AHA (n = 115) and in 52%, 57%, and 81% of HA inhibitor patients (n = 63). Competitive binding of the human monoclonal antibody (mAb) LE2E9 revealed overlapping epitopes with murine C1-specific group A mAbs including 2A9. Mutational analyses identified distinct crucial binding residues for LE2E9 (E2066) and 2A9 (F2068) that are also recognized by anti-C1 antibodies present in patients with hemophilia. A strong contribution of LE2E9- and 2A9-like antibodies was particularly observed in patients with AHA. Overall, our study demonstrates that the C1 domain, in addition to the A2 and C2 domains, contributes significantly to the humoral anti-fVIII immune response in acquired and congenital hemophilia inhibitor patients.

Crizotinib in ALK+ inflammatory myofibroblastic tumors-Current experience and future perspectives.

Inflammatory myofibroblastic tumor (IMT) and its subtype epithelioid inflammatory myofibroblastic sarcoma (EIMS) are rare soft-tissue tumors. As about 50% of IMT and 100% of EIMS contain activating rearrangements of the anaplastic lymphoma kinase (ALK) gene, targeted kinase inhibition of ALK by compounds such as crizotinib is a potential treatment option. We performed a literature review and analyzed a total of 30 patients with IMT/EIMS treated with crizotinib. A total of 12 patients achieved complete or partial remission. As preliminary data are promising, a prospective study evaluating crizotinib treatment in patients with unresectable/multifocal ALK+ IMT/EIMS is warranted.

Treatment of EBV-associated nodular sclerosing Hodgkin lymphoma in a patient with ataxia telangiectasia with brentuximab vedotin and reduced COPP plus rituximab.

Patients with ataxia telangiectasia (AT) with malignancies face poor prognosis due to increased treatment-related toxicity. Here, we report a 14-year-old male with AT and Hodgkin lymphoma (HL) who received brentuximab vedotin and reduced COPP plus rituximab courses. This treatment resulted in complete remission and showed no severe toxicity.

Interleukin-2-stimulated natural killer cells are less susceptible to mycophenolate mofetil than non-activated NK cells: possible consequences for immunotherapy.

In a clinical phase I/II trial, pediatric patients with high-risk malignancies were treated with ex vivo IL-2-stimulated donor natural killer (NK) cells after transplantation with haploidentical stem cells. To evaluate the potential negative effects of the immunosuppressive drug mycophenolate mofetil (MMF) used for immunotherapy, the functionality and signaling of ex vivo NK cells was investigated. Our results show that during NK cell expansion, long-term (9 days) incubation with mycophenolic acid (MPA), the active metabolite of MMF, in therapeutically relevant concentrations led to the severe inhibition of NK cell proliferation. This correlated with a significantly reduced cytokine/chemokine secretion and the inhibited acquisition of surface receptors regarding cytotoxicity (e.g., NKp30, NKp44, NKp46, NKG2D), adhesion/migration (e.g., ICAM-1/CD54, LFA-1/CD11a, CD62L, CXCR3) and activation (e.g., CD25). Moreover, MPA prevented phosphorylation of the central signaling molecules STAT-3/-4/-5, AKT and ERK1/2. In contrast, short-term (24 h) MPA incubation of IL-2-stimulated NK cells had no or only marginal effects on the activated NK cell phenotype, including receptor expression, cytokine/chemokine secretion and intracellular signaling. Further, short-term MPA incubation only moderately affected the highly cytotoxic activity of previously IL-2-stimulated NK cells. In conclusion, while long-term MPA incubation significantly compromised ex vivo NK cell functionality, previously IL-2-activated NK cells seemed to be rather resistant to short-term MPA treatment. This finding supports the use of IL-2-activated NK cells as immunotherapy, especially for patients treated with MMF after haploidentical stem cell transplantation.

Treatment of patients with advanced cancer with the natural killer cell line NK-92.

BACKGROUND AIMS: Natural killer (NK) cells, either naive or genetically engineered, are increasingly considered for cellular therapy of patients with malignancies. When using NK cells from peripheral blood, the number of expanded NK cells can be highly variable and the need for NK cell enrichment can make the process expensive. The NK-92 cell line (CD56+/CD3-) that was isolated from a patient with lymphoma has predictable high cytotoxic activity and can be expanded under good manufacturing practice conditions in recombinant interleukin-2. METHODS: Fifteen patients (age, 9-71 years) with advanced, treatment-resistant malignancies, either solid tumors/sarcomas (n = 13) or leukemia/lymphoma (n = 2), received two infusions of NK-92 cells, given 48 h apart. Three cohorts of patients were treated with escalating doses of NK-92 cells (n = 7 at 1 × 10(9), n = 6 at 3 × 10(9) and n = 2 at 1 × 10(10) cells/m(2)). RESULTS: No infusion-related or long-term side effects were observed. The dose of 10(10) cells/m(2) was considered the maximum expandable cell dose with the use of an established culture bag system. Three fourths of patients with lung cancer had some anti-tumor response. Only one patient of seven had development of human leukocyte antigen antibodies. The persistence of NK-92 cells (male origin) in the circulation was confirmed by Y chromosome-specific polymerase chain reaction in two female patients. CONCLUSIONS: Infusions of NK-92 cells up to 10(10) cells/m(2) were well tolerated. Despite the allogeneic nature of NK-92, development of human leukocyte antigen antibodies in these patients with cancer appears to be rare. The cells can persist in the recipient's circulation for at least 48 h. Some encouraging responses were seen in patients with advanced lung cancer.

Safety and Feasibility of Long-Distance Aeromedical Transport of Neonates and Children in Fixed-Wing Air Ambulance.

In cases of critical injury or illness abroad, fixed-wing air ambulance aircraft is employed to repatriate children to their home country. Air ambulance aircraft also transport children to foreign countries for treatment not locally available and newborns back home that have been born prematurely abroad. In this retrospective observational study, we investigated demographics, feasibility, and safety and outcomes of long-distance and international aeromedical transport of neonates and children. The study included 167 pediatric patients, 56 of those preterm neonates. A total of 41 patients were ventilated, 45 requiring oxygen prior to the transport, 57 transferred from an intensive care unit (ICU), and 48 to an ICU. Patients were transported by using Learjet 31A, Learjet 45, Learjet 55, and Bombardier Challenger 604, with a median transport distance of 1,008 nautical miles (NM), median transport time of 04:45 hours (median flight time = 03:00 hours), flight time ≥8 hours in 15 flights, and transport time ≥8 hours in 29 missions. All transports were accompanied by a pediatric physician/nurse team. An increase in FiO 2 during the transport was documented in 47/167 patients (28%). Therapy escalation (other than increased oxygen) was reported in 18 patients, and technical adverse events in 3 patients. No patient required CPR or died during the transport. Clinical transport outcome was rated by the accompanying physician as unchanged in 163 transports, improved in 4, and deteriorated in none. In summary, international, long-distance transport of neonatal and pediatric patients performed by experienced and well-equipped transport teams is feasible. Neither major adverse events nor physician-rated clinical deteriorations were observed in this group of patients.

IL-2-activated haploidentical NK cells restore NKG2D-mediated NK-cell cytotoxicity in neuroblastoma patients by scavenging of plasma MICA.

NK group 2D (NKG2D)-expressing NK cells exhibit cytolytic activity against various tumors after recognition of the cellular ligand MHC class I chain-related gene A (MICA). However, release of soluble MICA (sMICA) compromises NKG2D-dependent NK-cell cytotoxicity leading to tumor escape from immunosurveillance. Although some molecular details of the NKG2D-MICA interaction have been elucidated, its impact for donor NK (dNK) cell-based therapy of solid tumors has not been studied. Within an ongoing phase I/II trial, we used allogeneic IL-2 activated dNK cells after haploidentical stem cell transplantation for immunotherapy of patients with high-risk stage IV neuroblastoma. NKG2D levels on activated dNK cells increased strongly when compared with freshly isolated dNK cells and correlated with enhanced NK-cell cytotoxicity. Most importantly, elevated sMICA levels in patients plasma correlated significantly with impaired dNK-cell-mediated cytotoxicity. This effect could be reversed by high-dose infusion of activated dNK cells, which display high levels of surface NKG2D. Our data suggest that the provided excess of NKG2D leads to clearance of sMICA and preserves cytotoxicity of dNK cells via non-occupied NKG2D. In conclusion, our results identify this tumor immune escape mechanism as a target to improve immunotherapy of neuroblastoma and presumably other tumors.

Leptomeningeal metastases in pediatrics: magnetic resonance image manifestations and correlation with cerebral spinal fluid cytology.

BACKGROUND: Detection of leptomeningeal metastases is fundamental to a complete evaluation of central nervous system (CNS) or non-CNS tumor with suspected involvement of the neuroaxis. Our purpose was to assess the appearances of different magnetic resonance (MR) sequences in the diagnosis of leptomeningeal metastases and correlate those positive findings with the cerebral spinal fluid (CSF) cytology results. METHODS: The authors reviewed the medical records and MR image manifestations of leptomeningeal metastases from 18 children who had positive MR findings and retrospectively correlated them with CSF cytologic results. There was a uniform MR protocol and the patients were examined with the same sequences. RESULTS: The abnormalities included pial-arachnoid disease (n = 16), disease coating the nerves (n = 12), hydrocephalus (n = 3) and subependymal metastases (n = 2). Enhanced T1 images were better than unenhanced fluid attenuated inversion recovery (FLAIR) and T2 to delineate cranial and spinal leptomeningeal metastases. In our sample, seven out of 18 cases were cytologically negative on a single lumbar puncture. CONCLUSIONS: Contrast-enhanced MR imaging can be invaluable, detecting the false-negative lumbar punctures. FLAIR and diffusion images can be helpful in diagnosing leptomeningeal metastases of non-enhancing primary tumors. Prognosis was more related to the primary tumor type than to the leptomeningeal enhancement MR pattern.

The role of magnetic resonance imaging in children with hematogenous brain metastases from primary solid tumors.

Within a 10-year period, 4 out of 429 children with solid tumors treated at the pediatric oncology department developed brain metastases. Lesions secondary to direct extension from the skull or dura were excluded. The tumors causing brain metastases were non-small cell lung carcinoma, Wilms' tumor, osteosarcoma, und hepatoblastoma. All patients had single brain metastasis. All tumors were subcortical/cortical based and isointense on T1-images and, in 2 cases, mildly hyperintense on T2-images. Two patients showed diffusion abnormalities. Three showed enhancement. In the patient with osteosarcoma, metastasis was calcified. Central nervous system (CNS) metastasis may not in itself be a terminal event; metastasis in patients with Wilms' tumor might behave differently. Neuroimaging should be considered in children with pediatric solid tumors with neurological symptoms on follow up.

Long-Distance Aeromedical Transport of Patients with COVID-19 in Fixed-Wing Air Ambulance Using a Portable Isolation Unit: Opportunities, Limitations and Mitigation Strategies.

INTRODUCTION: Aeromedical transport of patients with highly-infectious diseases, particularly over long distances with extended transport times, is a logistical, medical and organizational challenge. Following the 2014-2016 Ebola Crisis, sophisticated transport solutions have been developed, mostly utilizing large civilian and military airframes and the patient treated in a large isolation chamber. In the present COVID-19 pandemic, however, many services offer aeromedical transport of patients with highly-infectious diseases in much smaller portable medical isolation units (PMIU), with the medical team on the outside, delivering care through portholes. METHODS: We conducted a retrospective review of all transports of patients with proven or suspected COVID-19 disease, transported by Jetcall, Idstein, Germany, between April 1 and August 1, 2020, using a PMIU (EpiShuttle, EpiGuard AS, Oslo, Norway). Demographics and medical data were analyzed using the services' standardized transport protocols. Transport-associated challenges and optimization strategies were identified by interviewing and debriefing all transport teams after each transport. RESULTS: Thirteen patients with COVID-19 have been transported in a PMIU over distances up to 7,400 kilometers (km), with flight times ranging from 02:15 hours to 11:10 hours. We identified the main limitations of PMIU transports as limited access to the patient and reduced manual dexterity when delivering care through the porthole gloves and disconnection of lines and tubes during loading and unloading procedures. Technical solutions such as bluetooth-enabled stethoscopes, cordless ultrasound scanners and communication devices, meticulous preparation of the PMIU and the patient following standardized protocols and scenario-based training of crew members can reduce some of the risks. DISCUSSION: Transporting a patient with COVID-19 or any other highly infectious disease in a PMIU is a feasible option even over long distances, but adding a significant layer of additional risk, thus requiring a careful and individualized risk-benefit analysis for each patient prior to transport.

Treatment with sirolimus results in complete responses in patients with autoimmune lymphoproliferative syndrome.

We hypothesized that sirolimus, an mTOR inhibitor, may be effective in patients with autoimmune lymphoproliferative syndrome (ALPS) and treated patients who were intolerant to or failed other therapies. Four patients were treated for autoimmune cytopenias; all had a rapid complete or near complete response. Two patients were treated for autoimmune arthritis and colitis, demonstrating marked improvement. Three patients had complete resolution of lymphadenopathy and splenomegaly and all patients had a reduction in double negative T cells, a population hallmark of the disease. Based on these significant responses, we recommend that sirolimus be considered as second-line therapy for patients with steroid-refractory disease.

Position of nonmuscle myosin heavy chain IIA (NMMHC-IIA) mutations predicts the natural history of MYH9-related disease.

MYH9-related disease (MYH9-RD) is a rare autosomal-dominant disorder caused by mutations in MYH9, the gene for the heavy chain of nonmuscle myosin IIA (NMMHC-IIA). All patients present from birth with macrothrombocytopenia, but in infancy or adult life, some of them develop sensorineural deafness, presenile cataracts, and/or progressive nephritis leading to end-stage renal failure. No consistent correlations have been identified between the 27 different MYH9 mutations identified so far and the variable clinical evolution of the disease. We have evaluated 108 consecutive MYH9-RD patients belonging to 50 unrelated pedigrees. The risk of noncongenital manifestations associated with different genotypes was estimated over time by event-free survival analysis. We demonstrated that all subjects with mutations in the motor domain of NMMHC-IIA present with severe thrombocytopenia and develop nephritis and deafness before the age of 40 years, while those with mutations in the tail domain have a much lower risk of noncongenital complications and significantly higher platelet counts. We also evaluated the clinical course of patients with mutations in the four most frequently affected residues of NMMHC-IIA (responsible for 70% of MYH9-RD cases). We concluded that mutations at residue 1933 do not induce kidney damage or cataracts and cause deafness only in the elderly, those in position 702 result in severe thrombocytopenia and produce nephritis and deafness at a juvenile age, while alterations at residue 1424 or 1841 result in intermediate clinical pictures. These findings are relevant not only to patients' clinical management but also to the elucidation of the pathogenesis of the disease.

Enumeration of functionally active anti-Aspergillus T-cells in human peripheral blood.

Invasive aspergillosis remains a life-threatening complication in patients undergoing allogeneic stem cell transplantation (SCT). Since CD4(+) T-cells provide a critical secondary defense against Aspergillus spp., the quantification of "functional" anti-Aspergillus T-cells might be important in the clinical care of allogeneic transplant patients. We present a rapid, simple and reproducible method to enumerate functionally active, cytokine-producing anti-Aspergillus T-cells in peripheral blood by means of flow cytometry, by which these cells were also phenotypically characterized as memory CD4(+) T-cells. When using 100,000 PBMCs and requiring a minimum of 50 events, at least one anti-Aspergillus T-cell among 1000 CD4(+) T-cells can be detected. Compared to healthy individuals, the number of anti-Aspergillus T-cells in patients up to one year after SCT was significantly lower. The presented method might help to define hematopoietic transplant recipients who will benefit from adoptive transfer of anti-Aspergillus T cells.

Cytotoxic effects of treosulfan and busulfan against leukemic cells of pediatric patients.

PURPOSE: The alkylating agent treosulfan exerts a high cytotoxic activity against various malignant cells. Due to limited non-hematological toxicity, treosulfan might be a promising compound in myeloablative therapy for hematopoietic transplantation in children. Since in vitro data regarding the activity of treosulfan against childhood leukemic cells are limited, we compared the effect of treosulfan and busulfan against pediatric leukemic and non-malignant cells. EXPERIMENTAL DESIGN: Both agents were tested alone and in combination with fludarabine by means of the MTT and/or a five color-flow cytometric assay. Moreover, the induction of apoptosis by treosulfan was investigated via regulation of the proteinase caspase 3. RESULTS: Treosulfan was more active against leukemic cells of 20 children as well as against 3 leukemia-derived cell lines than busulfan, with increasing IC50 values from initial diagnosis to relapse. Overall purified stem cells were most sensitive, followed by CD56+CD3- NK and CD3+ T cells. The combination of treosulfan with fludarabine resulted in a synergistic effect against leukemic cells. In malignant cells, treosulfan induced rapid cell apoptosis measured by the activation of the centrally proteinase caspase 3. CONCLUSION: Our results indicate that treosulfan has activity against pediatric leukemic cells, myeloablative potential and immunosuppressive properties suitable for conditioning regimen in childhood malignancies.

MECOM-associated syndrome: a heterogeneous inherited bone marrow failure syndrome with amegakaryocytic thrombocytopenia.

Heterozygous mutations in MECOM (MDS1 and EVI1 complex locus) have been reported to be causative of a rare association of congenital amegakaryocytic thrombocytopenia and radioulnar synostosis. Here we report on 12 patients with congenital hypomegakaryocytic thrombocytopenia caused by MECOM mutations (including 10 novel mutations). The mutations affected different functional domains of the EVI1 protein. The spectrum of phenotypes was much broader than initially reported for the first 3 patients; we found familial as well as sporadic cases, and the clinical spectrum ranged from isolated radioulnar synostosis with no or mild hematological involvement to severe bone marrow failure without obvious skeletal abnormality. The clinical picture included radioulnar synostosis, bone marrow failure, clinodactyly, cardiac and renal malformations, B-cell deficiency, and presenile hearing loss. No single clinical manifestation was detected in all patients affected by MECOM mutations. Radioulnar synostosis and B-cell deficiency were observed only in patients with mutations affecting a short region in the C-terminal zinc finger domain of EVI1. We propose the term MECOM-associated syndrome for this heterogeneous hereditary disease and inclusion of MECOM sequencing in the diagnostic workup of congenital bone marrow failure.

An Exponential Regression Model Reveals the Continuous Development of B Cell Subpopulations Used as Reference Values in Children.

B lymphocytes are key players in humoral immunity, expressing diverse surface immunoglobulin receptors directed against specific antigenic epitopes. The development and profile of distinct subpopulations have gained awareness in the setting of primary immunodeficiency disorders, primary or secondary autoimmunity and as therapeutic targets of specific antibodies in various diseases. The major B cell subpopulations in peripheral blood include naïve (CD19+ or CD20+IgD+CD27-), non-switched memory (CD19+ or CD20+IgD+CD27+) and switched memory B cells (CD19+ or CD20+IgD-CD27+). Furthermore, less common B cell subpopulations have also been described as having a role in the suppressive capacity of B cells to maintain self-tolerance. Data on reference values for B cell subpopulations are limited and only available for older age groups, neglecting the continuous process of human B cell development in children and adolescents. This study was designed to establish an exponential regression model to produce continuous reference values for main B cell subpopulations to reflect the dynamic maturation of the human immune system in healthy children.

Vemurafenib in Langerhans cell histiocytosis: report of a pediatric patient and review of the literature.

Selective BRAF inhibitors such as vemurafenib have become a treatment option in patients with Langerhans cell Histiocytosis (LCH). To date, only 14 patients receiving vemurafenib for LCH have been reported. Although vemurafenib can stabilize the clinical condition of these patients, it does not seem to cure the patients, and it is unknown, when and how to stop vemurafenib treatment. We present a girl with severe multisystem LCH who responded only to vemurafenib. After 8 months of treatment, vemurafenib was tapered and replaced by prednisone and vinblastine, a strategy which has not been described to date. Despite chemotherapy, early relapse occurred, but remission was achieved by re-institution of vemurafenib. Further investigation needs to address the optimal duration of vemurafenib therapy in LCH and whether and which chemotherapeutic regimen may prevent disease relapse after cessation of vemurafenib.

Neurocognitive development in HIV-positive children is correlated with plasma viral loads in early childhood.

Because of neurocognitive impairments in perinatally human immunodeficiency virus (HIV)-infected children and adolescents, this study aimed to demonstrate the effect of plasma viral loads and early initiation of sufficient combined antiretroviral therapy (cART) on neurocognitive development.In total, 14 perinatally infected HIV-positive children (median age 8.24 years [range: 6.0-16.74]) receiving lopinavir/ritonavir (LPV/r)-based ART underwent neurocognitive testing using the Wechsler Intelligence Score for Children, 4th Edition (WISC-IV). All 14 patients participated in a pharmacokinetic study in which they were hospitalized for an entire day. As a child's ability to concentrate varies over the course of the day, all tests were performed in the morning.The patients' neurocognitive development did not significantly differ from the normative collective pattern for any of the following composite scores that were examined: full-scale intelligence quotient (IQ) (mean: 106.5, P = .1060), verbal comprehension index (mean: 106.0, P = .1356), perceptual reasoning index (mean: 106.0, P = .1357), working memory index (mean: 106.3, P = .1171), and processing speed index (mean: 98.1, P = .6313). The overall full-scale IQ scores were significantly higher in children who began ART within the first year of life (P = .0379), whereas low lopinavir/r plasma levels (P = .0070) and high viral load area under the curves (AUCs) in the first 3 years of life, but not later, significantly correlated with reduced neurocognitive performance (Spearman r = -0.64, P = .0278).In this cohort of cART treated HIV-positive children and adolescents, neurocognitive performance correlated with early and sufficient viral load suppression within the first 3 years of life.