Conversion of AML-blasts to leukemia-derived dendritic cells (DCleu) in 'DC-culture-media' shifts correlations of released chemokines with antileukemic T-cell reactions.

Dendritic cells (DC) and T-cells are mediators of CTL-responses. Autologous (from patients with acute myeloid leukaemia (AML) or myelodysplasia (MDS)) or allogeneic (donor)-T-cells stimulated by DCleu, gain an efficient lysis of naive blasts, although not in every case. CXCL8, -9, -10, CCL2, -5 and Interleukin (IL-12) were quantified by Cytometric Bead Array (CBA) in supernatants from 5 DC-generating methods and correlated with AML-/MDS-patients' serum-values, DC-/T-cell-interactions/antileukemic T-cell-reactions after mixed lymphocyte culture (MLC) and patients' clinical course. The blast-lytic activity of T-cells stimulated with DC or mononuclear cells (MNC) was quantified in a cytotoxicity assay. Despite great variations of chemokine-levels, correlations with post-stimulation (after stimulating T-cells with DC in MLC) improved antileukemic T-cell activity were seen: higher released chemokine-values correlated with improved T-cells' antileukemic activity (compared to stimulation with blast-containing MNC) - whereas with respect to the corresponding serum values higher CXCL8-, -9-, and -10- but lower CCL5- and -2-release correlated with improved antileukemic activity of DC-stimulated (vs. blast-stimulated) T-cells. In DC-culture supernatants higher chemokine-values correlated with post-stimulation improved antileukemic T-cell reactivity, whereas higher serum-values of CXCL8, -9, and -10 but lower serum-values of CCL5 and -2 correlated with post-stimulation improved antileukemic T-cell-reactivity. In a context of 'DC'-stimulation (vs serum) this might point to a change of (CCL5 and -2-associated) functionality from a more 'inflammatory' or 'tumor-promoting' to a more 'antitumor'-reactive functionality. This knowledge could contribute to develop immune-modifying strategies that promote antileukemic (adaptive) immune-responses.

Serum Chemokine-release Profiles in AML-patients Might Contribute to Predict the Clinical Course of the Disease.

In cancer or hematologic disorders, chemokines act as growth- or survival factors, regulating hematopoiesis and angiogenesis, determining metastatic spread and controlling leukocyte infiltration into tumors to inhibit antitumor immune responses. The aim was to quantify the release of CXCL8, -9, -10, CCL2, -5, and IL-12 in AML/MDS-pts' serum by cytometric bead array and to correlate data with clinical subtypes and courses. Minimal differences in serum-levels subdivided into various groups (e.g. age groups, FAB-types, blast-proportions, cytogenetic-risk-groups) were seen, but higher release of CXCL8, -9, -10 and lower release of CCL2 and -5 tendentially correlated with more favorable subtypes (<50 years of age, <80% blasts in PB). Comparing different stages of the disease higher CCL5-release in persisting disease and a significantly higher CCL2-release at relapse were found compared to first diagnosis - pointing to a change of 'disease activity' on a chemokine level. Correlations with later on achieved response to immunotherapy and occurrence of GVHD were seen: Higher values of CXCL8, -9, -10 and CCL2 and lower CCL5-values correlated with achieved response to immunotherapy. Predictive cut-off-values were evaluated separating the groups in 'responders' and 'non-responders'. Higher levels of CCL2 and -5 but lower levels of CXCL8, -9, -10 correlated with occurrence of GVHD. We conclude, that in AML-pts' serum higher values of CXCL8, -9, -10 and lower values of CCL5 and in part of CCL2 correlate with more favorable subtypes and improved antitumor'-reactive function. This knowledge can contribute to develop immune-modifying strategies that promote antileukemic adaptive immune responses.

[Abortive course of malignant hyperthermia following preclinical induction of general anesthesia using succinylcholine]. / Abortiver Verlauf einer malignen Hyperthermie nach präklinischer Narkoseinduktion mit Succinylcholin.

In a 59-year-old woman with a perforating eyeball injury to the right eye, the emergency physician induced a preclinical general anesthesia with propofol, fentanyl and the depolarizing muscle relaxant succinylcholine. Anesthesia was maintained using propofol and remifentanil infusion throughout the preoperative period and the subsequent surgical procedure. Postoperatively, isolated rhabdomyolysis with an increase in serum creatine kinase to >20,000 U/l was observed. The diagnosis of malignant hyperthermia (MH) susceptibility could be confirmed in the patient 4 months after the acute event by the in vitro contracture test and detection of the MH causative mutation p.Val4849Ile in exon 101 of the ryanodine receptor gene. Due to the variable expression, for a long time MH often remained unrecognized. Warning symptoms, such as unspecific tachycardia or masseter spasm following succinylcholine injection, should alert emergency physicians to include MH susceptibility in the differential diagnostics. With an estimated genetic MH prevalence of 1:2000-3000, individuals with known or so far unrecognized MH disposition are likely to be among patients treated in the preclinical setting. If a person develops MH symptoms after exposure to triggering agents, immediate hospital admission is essential in order to initiate guideline-conform treatment without further delay because preclinically the life-saving causal measures are not possible due to the lack of supply of dantrolene.

Follicular variant of papillary thyroid cancer in Alström syndrome.

Alström syndrome (AS) is an autosomal recessive disorder, characterized by cone-rod dystrophy, sensorineural hearing loss, obesity, hyperinsulinemia with insulin resistance, type 2 diabetes mellitus and progressive pulmonary, hepatic and renal dysfunction. AS is caused by mutations in the ALMS1 gene, located on the short arm of chromosome 2. We report a 35-year-old woman with known history of AS, who developed a follicular variant of papillary thyroid carcinoma. To our knowledge this is the first association of AS with thyroid malignancy, among the approximately 450 cases reported since the first description of the syndrome. We conclude that papillary thyroid carcinoma should be considered in the differential diagnosis of thyroid nodules in patients with AS.

Dynamics of fertility impairment and recovery after allogeneic haematopoietic stem cell transplantation in childhood and adolescence: results from a longitudinal study.

PURPOSE: Fertility impairment and recovery after haematopoietic stem cell transplantation (HSCT) have been reported in both sexes, but little is known about how they develop over time. Our aim was to describe the dynamics of fertility impairment and recovery after HSCT. METHODS: We retrieved treatment and fertility data for up to 12 years of 361 paediatric patients with malignant and non-malignant diseases from seven European centres. The patients had been treated with allogeneic HSCT between 2000 and 2005. RESULTS: Development of fertility impairment was observed in males (123/217, 56%) after a median time of 2.6 years (range 0.1-11.4) and in females (82/144, 57%) after 2.3 years (range 0.1-12.0) after HSCT. Different busulfan dosages had only a slight impact on the onset of fertility impairment (busulfan ≥ 16 mg/kg with a median time to fertility impairment of 2.9 vs. 3.9 years after busulfan <14 mg/kg). Recovery from fertility impairment was observed in 17 participants after a median time of 4.1 years (range 1-10.6) in females (10/144, 7%) and 2.0 years (range 1-6.3) in males (7/217, 3 %) after fertility impairment first appeared. CONCLUSIONS: In the light of the dynamics of fertility impairment and recovery in the HSCT patients reviewed, these patients should be counselled comprehensively regarding fertility preservation measures.

[Malignant hyperthermia]. / Maligne Hyperthermie.

Malignant hyperthermia (MH) is a rare hereditary, mostly subclinical myopathy. Trigger substances, such as volatile anesthetic agents and the depolarizing muscle relaxant succinylcholine can induce a potentially fatal metabolic increase in predisposed patients caused by a dysregulation of the myoplasmic calcium (Ca) concentration. Mutations in the dihydropyridine ryanodine receptor complex in combination with the trigger substances are responsible for an uncontrolled release of Ca from the sarcoplasmic reticulum. This leads to activation of the contractile apparatus and a massive increase in cellular energy production. Exhaustion of the cellular energy reserves ultimately results in local muscle cell destruction and subsequent cardiovascular failure. The clinical picture of MH episodes is very variable. Early symptoms are hypoxia, hypercapnia and cardiac arrhythmia whereas the body temperature rise, after which MH is named, often occurs later. Decisive for the course of MH episodes is a timely targeted therapy. Following introduction of the hydantoin derivative dantrolene, the previously high mortality of fulminant MH episodes could be reduced to well under 10 %. An MH predisposition can be detected using the invasive in vitro contracture test (IVCT) or mutation analysis. Few elaborate diagnostic procedures are in the developmental stage.

Sevoflurane is less sensitive than halothane for in vitro detection of malignant hyperthermia susceptibility.

BACKGROUND: Sevoflurane is a known triggering agent of malignant hyperthermia (MH). The present study analyzed different effects of sevoflurane on skeletal muscle of MH susceptible and nonsusceptible individuals in vitro and compared the results to the standardized test protocol with halothane and caffeine. A potential influence of a present ryanodine receptor type 1 (RyR1) mutation was investigated. METHODS: Muscle bundles of 24 MH-susceptible patients with or without an RyR1 mutation, 35 MH-nonsusceptible and 10 MH-equivocal patients were exposed either to sevoflurane 8 vol% bolus or increasing doses of 2, 4, 6, and 8 vol%. In MH-positive patients, a screening for mutations in the RyR1 gene was performed. RESULTS: The in vitro parameters initial length, weight, predrug resting tension, and predrug twitch height did not differ between the groups. Sevoflurane caused significant contractures in MH-susceptible but not in MH-nonsusceptible muscle after increasing doses [1.4 (0.3-6.0) vs. 0 (0-0) mN] and after bolus application [6.9 (2.4-21.4) vs. 0 (0-0) mN]. However, only 50% of the susceptible patients developed contractures ≥ 2 mN after increasing concentrations while 83% did so after rapid bolus administration. Presence of an RyR1 mutation was detected in 36% of the examined MH-positive patients but had no influence on developing contractures. CONCLUSION: Sevoflurane-induced contractures do not reliably detect MH susceptibility on an individual level. Therefore, sevoflurane is no suitable alternative for diagnostic use. Mutation-specific effects regarding contracture sizes after incubation with sevoflurane, halothane, or caffeine were not found.

Allo-SCT using BU, CY and melphalan for children with AML in second CR.

Based on the results from the AML-BFM 98 trial, hematopoietic SCT (HSCT) is recommended for children with AML in second CR only. Here, we retrospectively analyze interphase data of children who underwent HSCT after myeloablative conditioning with BU, CY, and melphalan (BuCyMel) for AML in second remission (CR2) between 1998 and 2009. Out of 152 children, transplant data were available on 109 individuals. Sixty out of 109 children (55%) received BuCyMel. Median age at HSCT was 12.2 years (range 3.0; 18.3). GVHD prophylaxis mostly consisted of CsA and short term MTX with or without antithymocyte globulin. Matched-sibling donors were used for 6/60 analyzed recipients, the remainder either received grafts from matched unrelated (30/60) or mismatched donors. OS after 5 years was 62% (s.e. 6%), relapse incidence 35% (18/60 children) and treatment-related mortality accounted for 12% (7/60) of fatal events. In conclusion, even taking into account possible selection bias in this retrospective analysis, HSCT in CR2 using BuCyMel resulted in a respectable OS. Based on this data the prospective, controlled and centrally monitored AML SCT-BFM 2007 trial has started to recruit patients in January 2010 aiming to generate valid outcome data for further strategy decisions.

Efficient control of pandemic 2009 H1N1 virus infection with intravenous zanamivir despite the lack of immune function.

A teenager who acquired 2009 H1N1 influenza A lower respiratory tract infection during total bone marrow and lymphoid aplasia, in the setting of human leukocyte antigen-haploidentical hematopoietic stem cell transplantation, was successfully treated with intravenous zanamivir. This case demonstrates efficient control of pandemic influenza infection by intravenous zanamivir in the absence of any functional immune system, thus suggesting profound antiviral activity.

Epidemiology and clinical characteristics of pandemic (H1N1) 2009 influenza infection in pediatric hemato-oncology and hematopoietic stem cell transplantation patients.

BACKGROUND: For children with hemato-oncologic diseases, especially after hematopoietic stem cell transplantation (HSCT), the risk for developing complications related to pandemic influenza A (H1N1) 2009 (pH1N1) infection is largely unknown. METHODS: A retrospective chart study was performed of pH1N1 cases diagnosed between October 2009 to January 2010 in the hemato-oncologic unit of the University Children's Hospital of Düsseldorf, Germany. FINDINGS: In total, 21 children were diagnosed with laboratory-confirmed pH1N1; in 16 patients with malignancies (acute leukemia 7, lymphoma 4, solid tumors 2, others 3) and in 5 with benign hematologic disorders. Five patients had undergone prior HSCT, although 1 patient was diagnosed during conditioning therapy with high-dose chemotherapy in preparation for haploidentical HSCT. Most frequent symptoms were fever (>38.5°C) and cough (in 100%), and rhinorrhea (57%). The 2 patients acquiring pH1N1 infection under high-dose or intensive chemotherapy did not require intensive care or mechanical ventilation, and both recovered under antiviral therapy. Oseltamivir was administered to 11 patients; in 1 patient, therapy was switched, on a compassionate-use basis, to intravenous zanamivir because of lack of clinical improvement after oseltamivir therapy. Complications were hospitalization (19%), demand of oxygen supplementation, delay/interruption of antineoplastic therapy, and prolonged administration of antibiotics and antipyretics. CONCLUSION: In the investigated patient population, pH1N1 was mild in most cases, but was associated with substantial morbidity in a proportion of patients and led to interruption and delay in anticancer treatment.

Invasive aspergillosis in pediatric oncology patients: a rare event with poor prognosis--case analysis to plan better targeted prophylactic or therapeutic measurement.

Despite the implementation of new antifungal drugs, invasive aspergillosis (IA) still remains a considerable challenge in pediatric oncology with a severe mortality. Prophylactic and therapeutic measurement have to be evaluated in these rare but poor prognostic patients. Therefore the entire group of patients at risk of developing IA has to be defined before cooperative prospective trials. In a retrospective analysis including all our patients with malignancies we looked for patients with proven/probable IA. Cases of the period from 2003 to 2008 were analyzed in detail.In the period between 2003 to 2008 24 of 755 patients were affected by proven/ probable IA. Compared to former studies incidence increased from 1.3%in 1980 to 3.4% in 2008. AML patients with or without allogeneic/haploidentical stem cell transplantation were at highest risk (24% and 25% respectively, in comparison to 1% in ALL-patients). Survival after 2 years was 50% for patients with AML and IA. In patients with high risk to develop IA the effect of intensified, intravenous antimycotic prophylaxis has to be proven prospectively in a cooperative and randomized setting.

An unusual manifestation of post-transplant lymphoproliferative disorder in the lip after pediatric heart transplantation.

PTLD is a serious and frequently observed complication after solid organ transplantation. We present a six-yr-old girl with a rapidly growing, solid tumor of the lip four yr after orthotopic heart transplantation, which was classified as monomorphic PTLD with the characteristics of a diffuse large B-cell lymphoma. Treatment with reduction in immunosuppression, ganciclovir, and anti B-cell monoclonal antibody (rituximab) resulted in full remission since 12 months. To the best of our knowledge, this report is the first description of PTLD in the lip in a pediatric patient after heart transplantation in the English literature.

Fertility after allogeneic haematopoietic stem cell transplantation in childhood and adolescence.

Infertility is a major late effect in patients receiving haematopoietic stem cell transplantation (HSCT). The aim of this study was to determine the proportion of patients having fertility impairment after allogeneic HSCT in childhood/adolescence and to identify the potential risk factors. Treatment and fertility data of paediatric patients with malignant and non-malignant diseases treated with allogeneic HSCT between 2000 and 2005 were collected from seven European centres. Data were obtained for 138 female and 206 male patients after a median follow-up of 6 years (range 3-12). The patients' median age was 13 years (range 4-28) at the time of HSCT and 19 (range 12-35) years at the time of the enquiry. Seven children were born to the overall group, all at term and healthy. Fertility impairment was suspected in 69% males and 83% females. Start of treatment at age 13 years was a risk factor in females (odds ratio (OR) 4.7; 95% confidence interval (CI), 1.5 to 14.9), whereas pre-pubertal therapy was a risk factor in males (OR 0.4; 95% CI, 0.2 to 0.8). The major treatment-related risk factors were BU in females (OR 47.4; 95% CI, 5.4 to 418.1) and TBI in males (OR 7.7; 95% CI, 2.3 to 25.4). In light of the significant proportion of HSCT patients reviewed with impaired fertility, fertility conservation procedures should be considered for all patients undergoing HSCT, particularly those receiving TBI or BU-based preparative regimens.

Sevoflurane as a potential replacement for halothane in diagnostic testing for malignant hyperthermia susceptibility: results of a preliminary study.

BACKGROUND: To diagnose malignant hyperthermia (MH) susceptibility, muscle bundles are exposed to halothane and caffeine. We investigated whether sevoflurane, which is more clinically relevant but less potent of an anesthetic, could replace halothane in diagnostic MH testing. METHODS: With prior written consent, muscle bundles from 6 malignant hyperthermia susceptible (MHS) and 5 non-susceptible (MHN) individuals were exposed to increasing concentrations of sevoflurane (1.3; 2.6; 5.2 vol%). In addition, muscles from 9 MHS and 8 MHN were tested with a rapid exposure to 8 vol% of sevoflurane. Maximal contractures were measured and statistically analyzed (Mann-Whitney-U-test; P<0.05). RESULTS: There were no differences in weight, length or pre-drug tension of the muscle bundles. Incremental sevoflurane concentrations induced no differences in contracture between susceptible and non-susceptible muscles. The rapid application of sevoflurane induced significant contractures in all malignant hyperthermia susceptible compared with non-susceptible individuals. CONCLUSION: The rapid application of a high sevoflurane concentration but not an increasing stepwise application allowed for the diagnostic discrimination of susceptible individuals.

Magnetization transfer imaging provides no evidence of demyelination in methotrexate-induced encephalopathy.

Subacute MTX-induced encephalopathy is characterized by an abrupt onset of focal neurologic deficits within days after intrathecal or systemic therapy. Demyelination is one proposed mechanism. We describe the neuroimaging features of 2 patients with clinical symptoms of subacute encephalopathy after intrathecal and systemic MTX therapy. DWI showed restricted diffusion, indicating cytotoxic edema. MTI yielded no evidence of demyelination in either patient because there was no loss of MTR in areas of restricted diffusion.

Highly skewed T-cell receptor V-beta chain repertoire in the bone marrow is associated with response to immunosuppressive drug therapy in children with very severe aplastic anemia.

One of the major obstacles of immunosuppressive therapy (IST) in children with severe aplastic anemia (SAA) comes from the often months-long unpredictability of bone-marrow (BM) recovery. In this prospective study in children with newly diagnosed very severe AA (n=10), who were enrolled in the therapy study SAA-BFM 94, we found a dramatically reduced diversity of both CD4+ and CD8+ BM cells, as scored by comprehensive V-beta chain T-cell receptor (TCR) analysis. Strongly skewed TCR V-beta pattern was highly predictive for good or at least partial treatment response (n=6, CD8+ complexity scoring median 35.5, range 24-73). In contrast, IST in patients with rather moderate reduction of TCR V-beta diversity (n=4, CD8+ complexity scoring median 109.5, range 82-124) always failed (P=0.0095). If confirmed in a larger series of patients, TCR V-beta repertoire in BM may help to assign children with SAA up-front either to IST or to allogeneic stem-cell transplantation.

Statins alter intracellular calcium homeostasis in malignant hyperthermia susceptible individuals.

AIMS: Statines, HMG-CoA reductase inhibitors, are widely used to treat hypercholesterinemia. These substances are well tolerated, but myotoxic effects have been reported. The exact mechanisms of the induced myotoxicity are unknown but an involvement of intracellular calcium handling is suspected. Individuals susceptible to malignant hyperthermia (MH) have an impaired calcium homeostasis. An in vitro test measuring contracture responses of isolated muscle bundles is used to investigate cellular processes of MH. Aim of this study was to investigate if statins modify the contracture response of isolated muscle bundles from MH susceptible (MHS) and nonsusceptible (MHN) pigs. METHODOLOGY: With approval of the local ethics committee muscle biopsies of 18 MH susceptible and 12 nonsusceptible pigs were performed. Muscle bundles were mounted on an isometric force transducer, preloaded, and electrically stimulated. After establishment of a stable baseline, muscle bundles were exposed to simvastatin, atorvastatin, gemfibrocil, and the pure solvent. Baseline tension was measured and analyzed for changes with P < 0.05 considered to be significant. RESULTS: There were no differences in weight, length, and predrug baseline tension between the groups. Both simvastatin and atorvastatin induced significant contractures in muscle bundles from MHS pigs. Gemfibrocil and the solvent methanol showed no effect. In MHN muscle bundles, none of the tested substances induced a contracture. Statines induce contractures only in MHS muscle bundles. CONCLUSION: We therefore conclude that the underlying mechanism may be a pathologic influence on intracellular calcium handling that is absent in MHN. A preexisting impairment of the calcium homeostasis seems to be necessary for this behavior because muscle bundles of MHN pigs showed no pathologic reaction. A higher muscle cell vulnerability toward statins is assumed in MHS patients. Statins ought to be used with caution in these individuals. Analogous a diagnostic workup for MH should be considered for patients with statin-induced rhabdomyolyis.