Prevalence of antibodies against influenza A and B viruses in children in Germany, 2008 to 2010.

The prevalence of influenza A and B virus-specific IgG was determined in sera taken between 2008 and 2010 from 1,665 children aged 0-17 years and 400 blood donors in Germany. ELISA on the basis of whole virus antigens was applied. Nearly all children aged nine years and older had antibodies against influenza A. In contrast, 40% of children aged 0-4 years did not have any influenza A virus-specific IgG antibodies. Eightysix percent of 0-6 year-olds, 47% of 7-12 year-olds and 20% of 13-17 year-olds were serologically naïve to influenza B viruses. By the age of 18 years, influenza B seroprevalence reached approximately 90%. There were obvious regional differences in the seroprevalence of influenza B in Germany. In conclusion, seroprevalences of influenza A and influenza B increase gradually during childhood. The majority of children older than eight years have basal immunity to influenza A, while comparable immunity against influenza B is only acquired at the age of 18 years. Children aged 0-6 years, showing an overall seroprevalence of 67% for influenza A and of 14% for influenza B, are especially at risk for primary infections during influenza B seasons.

Response of children with stage IV soft tissue sarcoma to topotecan and carboplatin: a phase II window trial of the cooperative soft tissue sarcoma group.

To investigate antitumor activity and toxicity associated with combined topotecan and carboplatin treatment in children and adolescents with metastasized, untreated soft tissue sarcoma (STS).Patients (n=34) less than 21 years old and untreated, stage IV STS. Patients were treated with topotecan (1 mg/m²/d for 4 days) and carboplatin (150 mg/m²/d for 4 days) (TC course) during week 1 and 4 of a chemotherapy window trial, which was followed by chemotherapy and local therapy from week 6 on. We evaluated the side effects, toxicity and tumor response (using RECIST criteria) 6 weeks after starting the 2 TC chemotherapy courses.The objective response rate (ORR) was 38% (n=13 patients with a partial response (PR)), and a stable disease (SD) was reached in 11 cases. No patient showed a complete response (CR) of all metastatic lesions, although 1 patient showed a CR of the target lesion. 2 patients died of progress of disease (PD). Toxicity was mainly hematological (grade III/IV toxicity 79%), and nonhematological toxicities mainly included infection, fever, nausea,and vomiting. Regarding adverse events, 4 probable and 8 possible events related to study medication occurred among the 66 courses of TC.In conclusion, TC was potent against high-risk STS, but results and toxicity data were not superior to former published monotherapeutic topotecan therapies.

Prevalence of antibodies against hepatitis A virus among children and adolescents in Germany.

Since hepatitis A virus (HAV) infection during childhood is mostly asymptomatic, only seroprevalence studies can provide reliable information on incidence of HAV infection in children. The prevalence of anti-HAV antibodies was determined in sera taken in 2008 to 2010 from 1,645 children aged 0-17 years and in sera taken in 2010-2011 from 400 adult blood donors in Germany. For examination of trend over time, 715 sera collected between 1999 and 2006 from children at the age of 0-17 years within the federal state Thuringia were included. Antibody testing was carried out using the test kits ETI-AB-HAVK PLUS and ETI-HA-IGMK PLUS from DiaSorin. In children, the overall prevalence of antibodies was 10.8 %. After the seroprevalence declined from 8.8 % among the 0-2 year-olds to 2.4 % among the 3-4 year-olds, there was a significant increase to 20.5 % in the group of the 15-17 year-olds. Boys had with 12.7 % a significantly higher seroprevalence of anti-HAV antibodies compared to 8.8 % among girls. In adult blood donors, there was a HAV seroprevalence of 19.3 %. The likelihood of past infection or immunization within the age groups of children from 0 to 12 years differed significantly from that of adults. In conclusion, in Germany, only a small number of HAV infections occur in children, especially up to the age of 12 years. The proportion of susceptible children is greater than the proportion of susceptible adults. Thus, during outbreaks, the rate of infection among children would usually be higher than the rate among adults.

[GATA1-mutation associated leukemia in children with trisomy 21 mosaic]. / GATA1-Mutations-assoziierte Leukämien bei Kindern mit Trisomie 21-Mosaik.

Mutations of the hematopoietic transcription factor GATA1 (GATA1s) are pathognomonic in newborn with transient leukemia and children with Down syndrome and myeloid leukemia (ML-DS). Both TL and ML-DS can also occur in children with trisomy 21 mosaic.Between 2002 and 2011, 15 newborns and infants were diagnosed with DS mosaic. 9 of them presented with TL and 8 children suffered from ML-DS; 2 of them with a history of TL. In children without stigmata the special morphology and immunophenotype of blasts triggered the screening for GATA1 mutation and trisomy 21 mosaic.All newborns with TL achieved complete remission (CR). Due to clinical symptoms caused by the leukemic blasts, in 3 children low-dose cytarabine was applied. 1 patient died due to cardiac defect. In all patients GATA 1 s was confirmed. 6 children with ML-DS were initially treated according the AML-BFM protocol. After ML-DS was confirmed, therapy was continued with the intensity reduced schedule according to the ML-DS 2006 protocol. All children are still in CR (follow-up 1.8-7 years, median 2.7 yrs). 2 children with unknown trisomy 21 mosaic were diagnosed as acute megakaryoblastic leukemia (AMKL) and treated according the high risk arm of the AML-BFM 2004 including allogeneic stem cell transplantation in one child). GATA1 mutation was identified retrospectively. Both children are alive in CR.GATA1s associated leukemia has to be excluded in all young children with AMKL (<5 years old) to prevent overtreatment. Treatment with reduced intensity seems sufficient in children trisomy 21 mosaic and ML-DS.

Redirected behavior in learning tasks: the commercial laying hen (Gallus gallus domesticus) as model.

Redirected behaviors occur when some course of action is thwarted or inhibited (frustration). They also occur as adjunctive behaviors in operant conditioning tasks, where they might reflect frustration about unrewarded responses. Because frustration is associated with stress, which could interfere with learning and memory, we studied whether the occurrence of redirected behavior is correlated with learning success in a series of visual-cue discrimination tasks. Eleven hens, aged 34 wk, were tested on acquisition, reversal, extinction, and relearning of a simple visual discrimination task. The experimenters randomly assigned red and blue cardboard discs as discriminative stimuli. A correct response was recorded when a hen pecked at the correct disc. The learning criterion was 90% correct responses in 20 trials in 2 consecutive task sessions. The following data were documented: number of pecks needed to achieve the learning criterion, latency in choosing, pecks at the experimenter, and pecks at the surroundings. The behavioral responses were analyzed using linear mixed model ANOVA. Redirected pecking at the surroundings was a significant indicator of learning failure in that the more the hens performed this behavior, the more trials they needed to complete the discrimination tasks (P = 0.012). The number of pecks at the experimenter during the tasks significantly influenced learning success (P = 0.020), with hens directing more pecks at the experimenter during reversal, reaching the learning criterion in fewer trials (P = 0.027). The more the hens pecked at the experimenter during acquisition and extinction, however, the more trials they needed to meet the learning criteria (acquisition: P = 0.048; extinction: P = 0.003). Thus, laying hens are susceptible to the effects of frustration as measured in terms of redirected pecking elicited by operant procedures in visual discrimination tasks. In general, any situation in which a desirable goal is obstructed or an expected reward is omitted may lead to frustration-related activities, such as redirected behavior, which could in turn lead to abnormal behavior and welfare issues for the animals.

[Desmopressin testing in children with von Willebrand syndrome in haemostaseologic centers of Saxonia, Saxonia-Anhalt and Thuringia]. / Desmopressin-Tests bei Kindern mit von-Willebrand-Syndrom in den Gerinnungszentren Sachsens, Sachsen-Anhalts und Thüringens.

The influence of desmopressin on hemostasis is mediated by the release of von Willebrand factor and of coagulation factor VIII from vascular endothelium. The necessity of testing desmopressin effectiveness on hemostasis is a matter of controversy and the performance of the test is not yet standardized. For this reason the desmopressin tests in 114 children with von Willebrand syndrome (type 1, n=98; type 2A, n=12; type 2M, n=2; type 2N, n=2) carried out in 7 paediatric haemostaseologic centers were retrospectively analyzed. The effectiveness of desmopressin was assessed using defined response criteria. As expected, the test performance showed a wide variation among the centers. In 99 children desmopressin was given intravenously as a short infusion at a dosage ranging from 0.25 to 0.41 microg/kg and in 15 intranasally at an absolute dose of 40 to 300 microg. The points of time for blood taking after desmopressin application ranged from 0.5 to 12 h. The absent desmopressin response in 7 patients (6%) and the partial response in 15 indicate the necessity of testing desmopressin effectiveness before the first therapeutic use. The application of desmopressin was well tolerated by the patients.

Protein expression profile of newly diagnosed acute lymphoblastic leukemia in children developing relapses.

The purpose of this investigation was to evaluate the expression profile of proteins involved in children with newly diagnosed acute lymphoblastic leukemia (ALL) children who are developing relapses. For this reason, the expressions of 10 proteins including proto-oncogene and tumor suppressor gene products, proliferative factors and resistance parameters in 104 initial cases of childhood ALL were analyzed and the proteins correlated with ALL patients who experienced relapses. Applying immunocytochemical assays, we found that 4 out of the 10 parameters revealed a relationship to developing relapses (Fisher's exact tests). These were the oncogene product Fos (p=0.002), the drug resistance proteins glutathione S-transferase (p=0.008) and P-glycoprotein (P-pg/MDR1) (p=0.07) and protein kinase C (p=0.01). By means of hierarchical cluster analysis, we were able to show that the patients could be separated according to their protein expression profile into clusters consisting of patients whose ALL relapsed later and of patients who did not show relapses in the future.

BCRP gene expression is associated with a poor response to remission induction therapy in childhood acute myeloid leukemia.

Breast cancer resistance protein (BCRP), also known as mitoxantrone resistance protein (MRX) or placenta ABC protein (ABC-P), is the second member of the ABCG subfamily of ABC transport proteins (gene symbol ABCG2). Transfection and enforced expression of BCRP in drug-sensitive cells confers resistance to mitoxantrone, doxorubicin, daunorubicin and topotecan. In this study the expression of BCRP gene was measured using TaqMan real-time PCR in 59 children with newly diagnosed AML. Nine patients were also analyzed in relapse. The median of BCRP gene expression was more than 10 times higher in patients who did not achieve remission after the first phase of chemotherapy (n = 24) as compared to patients who did achieve remission at this stage (n = 21; P = 0.012). In first relapse the expression of the BCRP gene was higher than at diagnosis (P = 0.038). Although high levels of BCRP gene expression were more frequent in subtypes of AML with a favorable prognosis, we found that within both risk groups (high and low risk), patients who expressed high levels of BCRP had a worse prognosis (P = 0.023). Our results strongly suggest that the expression of the BCRP gene reduces the response to chemotherapy in AML and that BCRP expression is higher at the time of relapse.

Vascular endothelial growth factor in newly diagnosed and recurrent childhood acute lymphoblastic leukemia as measured by real-time quantitative polymerase chain reaction.

PURPOSE: Overexpression of vascular endothelial growth factor (VEGF) is associated with increased angiogenesis, growth, and metastasis in solid tumors, but to date the significance of VEGF in leukemia has received only limited attention. Therefore, this study examined the cellular VEGF levels in 31 newly diagnosed and 22 recurrent cases of childhood acute lymphoblastic leukemia (ALL). EXPERIMENTAL DESIGN: VEGF was determined with real-time quantitative PCR methods. Kaplan-Meier statistical analyses were conducted for the relapse-free intervals and the overall survival times. The groups were compared by log-rank and rank-sum tests. RESULTS: The VEGF levels were significantly higher in recurrent ALL compared with newly diagnosed ALL (28.0 versus 3.1 units; P = 0.001). Kaplan-Meier estimates were conducted to analyze the prognostic value of VEGF levels in newly diagnosed ALL with regard to the relapse-free intervals and the overall survival times. In this analysis, the median relapse-free interval of patients with low VEGF levels was more than 10 years, whereas the relapse-free interval of patients with high VEGF expression was only 1.2 years. The median overall survival time for the collective with low VEGF levels was >10 years, whereas the survival of the group of patients with high VEGF levels was 3.9 years. This difference was not statistically significant. This may be attributable to the small number of patients involved. CONCLUSION: Our data suggest that VEGF may play an important role in the pathophysiology of ALL. The expression of VEGF raises the possibility of using angiogenesis inhibitors as a novel therapeutic strategy in childhood ALL.

High-dose chemotherapy (HDC) and autologous hematopoietic stem cell transplantation (ASCT) as salvage therapy for relapsed osteosarcoma.

In this report, we describe our experience with high-dose chemotherapy (HDC) and autologous stem cell transplantation (ASCT) in 15 children with relapsed osteosarcoma who were treated by members of the Cooperative Osteosarcoma Study Group. Eight patients received HDC after the first relapse, six patients after the second relapse and one after the sixth relapse. Thirteen patients underwent HDC and ASCT in complete remission and two patients had macroscopic tumor residues. Seven patients received HDC based on melphalan and etoposide. Four of these patients were treated with additional carboplatinum. Two patients received carboplatinum, etoposide, and thiotepa or cyclophosphamide. In six patients double HDC was performed. In all six of these, the first HDC consisted of thiotepa/ cyclophosphamide. The second regimens included melphalan/etposide (two patients), melphalan/etposide/ carboplatinum (one patient), and melphalan/busulfan (one patient). Three of the 15 patients died of toxic complications. Eight patients developed further relapses, two patients showed persistent disease, and two patients are presently in continuous complete remission. The probability of relapse-free survival was 0.20 +/- 0.12 within a median follow-up (MFU) of 8 months and the probability of overall survival was 0.29 +/- 0.12 after a MFU of 16 months. In conclusion, utilization of HDC and ASCT in this patient group did not significantly improve the treatment outcome compared to conventional relapse therapy.

The anti-malarial artesunate is also active against cancer.

Artesunate (ART) is a semi-synthetic derivative of artemisinin, the active principle of the Chinese herb Artemisia annua. ART reveals remarkable activity against otherwise multidrug-resistant Plasmodium falciparum and P. vivax malaria. ART has now been analyzed for its anti-cancer activity against 55 cell lines of the Developmental Therapeutics Program of the National Cancer Institute, USA. ART was most active against leukemia and colon cancer cell lines (mean GI50 values: 1.11+/-0.56 microM and 2.13+/-0.74 microM , respectively). Non-small cell lung cancer cell lines showed the highest mean GI50 value (25.62+/-14.95 microM) indicating the lowest sensitivity towards ART in this test panel. Intermediate GI50 values were obtained for melanomas, breast, ovarian, prostate, CNS, and renal cancer cell lines. Importantly, a comparison of ART's cytotoxicity with those of other standard cytostatic drugs showed that ART was active in molar ranges comparable to those of established anti-tumor drugs. Furthermore, we tested CEM leukemia sub-lines resistant to either doxorubicin, vincristine, methotrexate, or hydroxyurea which do not belong to the N.C.I. screening panel. None of these drug-resistant cell lines showed cross resistance to ART. To gain insight into the molecular mechanisms of ART's cytotoxicity, we used a panel of isogenic Saccaromyces cerevisiae strains with defined genetic mutations in DNA repair, DNA checkpoint and cell proliferation genes. A yeast strain with a defective mitosis regulating BUB3 gene showed increased ART sensitivity and another strain with a defective proliferation-regulating CLN2 gene showed increased ART resistance over the wild-type strain, wt644. None of the other DNA repair or DNA check-point deficient isogenic strains were different from the wild-type. These results and the known low toxicity of ART are clues that ART may be a promising novel candidate for cancer chemotherapy.

Non-Hodgkin's lymphoma in pediatric patients with chromosomal breakage syndromes (AT and NBS): experience from the BFM trials.

BACKGROUND: Lymphoma and leukemia are the commonest malignant diseases in patients with chromosomal breakage syndromes and immunodeficiency (Ataxia teleangiectasia (AT) and Nijmegen breakage syndrome (NBS)). With improved management of infections, malignant disease is more frequently diagnosed and has become one of the commonest causes of death in pediatric AT and NBS. PATIENTS AND METHODS: In three consecutive multicenter therapy trials for pediatric non-Hodgkin's lymphoma (NHL) (NHL-BFM), 1569 patients with newly diagnosed NHL have been registered between 1986 and 1997. Nine patients with AT (n = 5) and NBS (n = 4) were identified and analysed. RESULTS: Median age of patients with AT and NBS at diagnosis of NHL was nine years. NHL-entities differed from non-AT/NBS-patients: diffuse large B-cell lymphomas, n = 7 (78%); ALCL, n = 1; lymphoblastic T-cell lymphoma, n = 1. Cervical nodes, paranasal sinuses and epipharynx were the sites most frequently involved. Stages were: I and II in three patients, III in five and IV in one patient. All patients received polychemotherapy according to tumor-entity and stage, none received radiation. Dose reductions according to individual tolerance concerned mainly ethotrexate, alkylating agents and epipodophyllotoxines. One patient died of toxic complications, two patients relapsed and died, one patient suffered from second malignancy. Five of nine patients are in 1. CCR after a median follow-up of five years. CONCLUSIONS: Patients with AT and NBS suffer from rare entities of pediatric NHL. Curative treatment is possible and should be attempted. Intensity of therapy should be adjusted to individual risk factors and tolerance. Alkylating agents, epipodophyllotoxines should be omitted, dose of MTX should be limited to 1 g/m2. Further cooperative trials using standardized approaches are required.

A Meckel's diverticulum containing pancreatic tissue and nesidioblastosis in a patient with Beckwith-Wiedemann syndrome.

A 30-month-old twin with Beckwith-Wiedemann syndrome underwent exstirpation of a nephroblastoma from the right kidney. A Meckel's diverticulum (MD), an incidental finding intraoperatively, was resected because it was indurated and enlarged. Histologically, the MD contained ectopic gastric and pancreatic mucosa. Nesidioblastosis was identified within the pancreatic tissue. This is probably the first description of ectopic nesidioblastosis within a MD.

Therapy for non-Hodgkin lymphoma in children with primary immunodeficiency: analysis of 19 patients from the BFM trials.

BACKGROUND: Non-Hodgkin lymphomas (NHL) represent an important complication of primary immunodeficiency (ID), posing new therapeutic challenges in this patient population. This study analyzes clinical data and therapy results of pediatric patients with primary ID and NHL in three consecutive NHL-BFM trials. PROCEDURE: Retrospective analysis of children with primary ID and NHL, treated according to protocol NHL-BFM, was performed regarding clinical presentation, diagnostic features, therapy, and outcome. RESULTS: From October, 1986, to April, 1997, 19 of 1,413 newly diagnosed patients with NHL were registered as suffering from primary ID. Age at diagnosis of NHL was lower in patients with ID. Six patients suffered from humoral ID, 13 patients from combined ID (ataxia teleangiectasia n = 3; Nijmegen breakage syndrome n = 4; PNP deficiency n = 1; IL2 receptor defect n = 1, other combined ID n = 4). Thirteen lymphomas were of B-cell and six of T-cell-lineage. Four of thirteen patients with combined ID were diagnosed with T-NHL, nine with B-NHL. Two of six patients with humoral ID presented with T-NHL and four with B-NHL. NHL entities differed significantly between ID and non-ID patients (P < or = 0.01): centroblastic and immunoblastic lymphomas (31.6% vs. 8.1%), anaplastic large cell lymphoma (26.3% vs. 10.7%), Burkitt lymphoma and B-ALL (21% vs. 47. 8%). Seventeen patients received polychemotherapy. Therapy-related toxicity was increased in ID- compared to non-ID-patients. Three patients died of sepsis; three died of tumor progression; one patient relapsed; one died of BMT-related toxicity; one died of second malignancy. Ten patients are in first continuous remission after a median follow-up of 4 years. CONCLUSIONS: Curative treatment of NHL in the presence of primary ID is possible and should be attempted.

Expression of a novel double-mutant dihydrofolate reductase-cytidine deaminase fusion gene confers resistance to both methotrexate and cytosine arabinoside.

A novel fusion gene consisting of the open reading frame of a double-mutant (Phe22-Ser31) dihydrofolate reductase (dmDHFR) cDNA fused to the open reading frame of cytidine deaminase (CD) was constructed and characterized for the purpose of conferring simultaneous resistance to methotrexate (MTX) and cytosine arabinoside (ara-C). The kinetic properties of purified recombinant dmDHFR-CD fusion protein were compared with those of purified CD and dmDHFR. The fusion protein was found to retain enzymatic properties of both dmDHFR and CD, in that the Km and Kcat values of purified dmDHFR-CD protein were found to be virtually identical to those of CD and dmDHFR alone. Retrovirus-mediated expression of dmDHFR-CD in NIH 3T3 cells conferred significant resistance (10- to 12-fold) against MTX and ara-C, compared with mock- and single gene-infected cells and the level of resistance obtained was similar to that of cells expressing both CD and dmDHFR from a retroviral bicistronic vector. Infection of mouse bone marrow cells with the dmDHFR-CD construct also showed high levels of resistance to MTX and ara-C in a CFU-GM assay. This fusion protein confers resistance to two antineoplastic agents that differ in their mechanism of action, and may be useful in the design of gene transfer strategies for protection of target cells against multiple drugs. Since high-dose ara-C and MTX are used in the treatment of lymphomas, this vector may be of value in protecting human hematopoietic progenitor cells from the toxicity of these antimetabolites.

Analysis of cyclin D1 in de novo and relapsed childhood acute lymphoblastic leukemia.

In a retrospective analysis we investigated 59 children with de novo acute lymphoblastic leukemia (ALL) and 28 patients with relapsed ALL for the expression of cyclin D1 using RT-PCR. In addition, the relationships of cyclin D1 to the retinoblastoma tumor suppressor gene and the Ki-67-expression were analyzed. Cyclin D1-mRNA was detectable in 42 out of 58 patients with de novo ALL (72%) and in 26 out of 28 relapsed patients (93%). The blast cells of the relapsed patients contained significantly higher levels of cyclin D1-mRNA compared with the de novo group (p = 0.0007). The cyclin D1 expression was independent of the proliferative activity of the cells and inversely correlated to the expression of the retinoblastoma tumor suppressor gene (r = -0.27, p = 0.03). Prognostic considerations using Kaplan Meier estimates for the relapse-free interval showed that patients with high cyclin D1- mRNA levels had a poorer prognosis (p = 0.046).

Proliferation and apoptosis in newly diagnosed and relapsed childhood acute lymphoblastic leukemia.

Children with acute lymphoblastic leukemia (ALL) who are treated with chemotherapy have remissions in more than 95% of the cases. However, 25% of the patients relapse and show resistance to chemotherapy. In this investigation we compared 25 newly diagnosed and 25 relapsed cases of ALL with respect to proliferation and apoptosis. Using immunocytochemistry and Western blotting, we determined the expression of cyclin A protein as a measure of the proliferative activity and the pro-apoptotic and anti-apoptotic factors, Fas, Fas ligand, caspase-3 and Bcl-2. Cyclin A expression was observed in 32% of the newly diagnosed cases and in 52% of the relapsed cases. Expression of Fas was found in 58% of the newly diagnosed and in only 27% of the relapsed samples. Of the newly diagnosed ALL, 88% expressed the Fas ligand while such expression was observed in 54% of the relapsed ALL. Sixty-four percent of the newly diagnosed cases expressed caspase-3 while only 48% of the relapsed samples did so. The anti-apoptotic factor, Bcl-2, was more frequently expressed in relapsed than in newly diagnosed cases. These data indicate that relapsed ALL more frequently exhibits high proliferative activity and reduced apoptosis than does newly diagnosed ALL.

Expression of Fas ligand in newly diagnosed childhood acute lymphoblastic leukemia.

Immunocytochemistry was used to analyze the expression of the Fas ligand (FasL) in sixty-four children with newly diagnosed acute lymphoblastic leukemia (ALL). FasL expression was detected in 55 of 64 children (86%). Children with newly diagnosed ALL and FasL expression exhibited longer relapse-free and overall survival times under chemotherapy than did patients who did not express the Fas ligand. These data suggest that Fas/FasL mediated signaling may play a general role in the cytotoxicity of anticancer drugs.