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This report documents the clinical activity of caspofungin in a 13-year-old girl with acute myeloid leukemia (AML) and chronic disseminated candidiasis (CDC), refractory to amphotericin B and fluconazole. Caspofungin was started at 50 mg/d resulting in a temporary response. With no further clinical improvement and radiological progress after 2 months of therapy, the dose of caspofungin was increased to 100 mg/d, leading to a sustained clinical response. Therapy was given for a total of 12 months and had no attributable adverse effects. This approach resulted in successful treatment of refractory CDC with caspofungin., (Copyright (c) 2006 Wiley-Liss, Inc.)

Ewing's sarcoma cells are highly susceptible to apoptosis via tumor necrosis factor apoptosis-inducing ligand (TRAIL). Resistance to TRAIL has been linked to deficient expression of caspase-8 in vitro. Here, we report on the status of caspase-8 expression in tumors from patients with Ewing's sarcoma, the effect of interferon-gamma on caspase-8 expression and apoptosis, and the role of caspase-8 for TRAIL- and chemotherapy-mediated apoptosis in Ewing's sarcoma. Using immunohistochemistry, we show that low expression of caspase-8 is seen in about 24% of tumors. Interferon-gamma induces expression of caspase-8 at concentrations achievable in humans and sensitizes cells to TRAIL. Transfection of wild type but not mutant caspase-8 into caspase-8-deficient Ewing's sarcoma cells restored sensitivity to TRAIL, indicating that up-regulation of caspase-8 is sufficient to restore TRAIL sensitivity. In contrast, no role for caspase-8 in chemotherapy-induced apoptosis was identified, because 1) transfection of caspase-8 or treatment with interferon-gamma did not alter the sensitivity of caspase-8-deficient cells to chemotherapeutics, 2) application of chemotherapy did not select for caspase-8-negative tumor cells in vivo, and 3) the caspase-8 status of tumors did not influence survival after chemotherapy-based protocols. In conclusion, our data provide a rationale for the inclusion of interferon-gamma in upcoming clinical trials with TRAIL.

Background: During successful chemotherapy of osteosarcomas tumour size does not diminish significantly because the therapy has limited impact on the mineralized matrix of the tumour. Treatment response is considered successful if, histologically, more than 90% of tumour cells show necrosis., Objective: To determine if osteosarcomas change their water diffusion during preoperative chemotherapy in relation to the amount of tumour necrosis., Materials and Methods: Eight patients (age 11-19 years) with histologically proven limb osteosarcoma underwent T1-weighted, fat-suppressed T2-weighted and contrast-enhanced T1-weighted spin-echo imaging together with diffusion-weighted EPI sequences (b = 700) at 1.5 T before and after five cycles of standard chemotherapy. Tumour volume and apparent diffusion coefficient (ADC) maps were calculated before and after chemotherapy. The degree of tumour necrosis after chemotherapy was assessed using the histological Salzer-Kuntschik classification (grades 1-6)., Results: During chemotherapy, the ADC values of osteosarcomas changed significantly. The ADC of untreated tumour was 2.1 +/- 0.4 x 10(-3) mm(2)/s (mean +/- SD) (95% CI 1.6-2.0). The ADC of chemotherapy-treated sarcomas was 2.5 +/- 0.4 x 10(-3) mm(2)/s (95% CI 1.8-2.2). Necrotic areas, which were confirmed by macroscopic examination, showed ADC values up to 2.7 x 10(-3) mm(2)/s. Four patients with little viable tumour tissue within the neoplasm (Salzer-Kuntschik grades 1-2) had an increase in ADC of 0.4 up to 0.7 x 10(-3) mm(2)/s. Four patients with larger areas of viable tumour (Salzer-Kuntschik grade 4) showed a lesser increase in ADC of 0.0 up to 0.3 x 10(-3) mm(2)/s. The differences in ADC values in tumour tissue before and after chemotherapy were highly significant (P = 0.01)., Conclusion: During chemotherapy of osteosarcomas, tumour ADC changes are related to the degree of tumour necrosis.

Purpose: We sought to evaluate diffusion and perfusion weighted 1.5 T magnetic resonance imaging (MRI) in detecting tumor necrosis with histologic correlation after preoperative chemotherapy., Materials and Methods: Eight patients (ages 11-19 years) with histologic proven osteosarcoma of the limbs underwent T1- and fat-suppressed T2-weighted spin echo and diffusion-weighted EPI sequences (b value = 700) after 5 cycles of standard chemotherapy. Tumor volume and apparent diffusion coefficients (ADC) were calculated. Tumor signal intensities were measured in dynamic contrast enhanced T1-weighted fast gradient echo-sequences obtained every 3 seconds after an intravenous injection of gadolinium-DTPA. Perfusion parameters of first-pass tracing of contrast medium (time-to-peak, slope of contrast enhancement curve) were calculated, and perfusion maps were established. After MRI, all patients underwent limb resection, and the specimens were investigated macroscopically and histologically. The degree of tumor necrosis was assessed using the histologic Salzer-Kuntschik classification (grades 1-6) after chemotherapy., Results: Necrotic areas, which were confirmed by macroscopic/histologic examination, showed ADC values up to 2.7 (mean, 2.3 +/- 0.2). Viable tumor areas revealed lower apparent diffusion coefficients (mean, 0.8 +/- 0.3). The differences in ADC between viable and necrotic tumor were highly significant (paired t test; P = 0.01). Slopes of necrotic areas ranged from 0.1 up to 5.2%/min (mean, 1.5%/min) and those of viable tumor areas from 2.8 to 31.5%/min (mean, 16.1%/min). The time-to-peak-values (TTPs) ranged from 40 to 210 seconds (mean, 131 seconds, SD 60 seconds) in necrotic tumors and from 30 to 96 seconds (mean, 55 seconds, SD 21) in viable areas of sarcomas. The differences in slope and TTP between viable and necrotic tumor were highly significant. In necrotic areas, the linear correlation between slope (%/min) and ADC (mm/s) and between TTP (s) and ADC were weak, respectively., Conclusion: Both dynamic contrast-enhanced MRI and diffusion-weighted MRI permit recognition of tumor necrosis induced by chemotherapy in osteosarcomas. We hypothesized that diffusion-weighted imaging is correlated directly with tumor necrosis. Perfusion-weighted imaging is correlated with microvessel density, vascular permeability, local blood volume, and flow. Therefore, perfusion weighed MRI depicts areas of tumor cell necrosis indirectly.

Griscelli syndrome (GS) was diagnosed in a 2-year-old patient with oculocutaneous albinism and immunodeficiency, but sequencing of RAB27a revealed only a heterozygous mutation. Due to impaired natural killer (NK) and T-cell cytotoxicity implying a high risk of developing hemophagocytic lymphohistiocytosis (HLH), he was prepared for hematopoietic stem cell transplantation (HSCT). Unexpectedly, a severe bleeding episode occurred that led to the demonstration of disturbed platelet aggregation, reduced plateletdense granules, and impaired platelet degranulation. In combination with neutropenia, this suggested the diagnosis of Hermansky-Pudlak syndrome type II (HPSII) and a novel homozygous mutation in AP3B1 was detected. None of the 3 reported HPSII patients had developed HLH, and our patient seroconverted to Epstein-Barr virus (EBV) without clinical symptoms. HSCT was therefore withheld, and granulocyte-colony-stimulating factor (G-CSF) therapy was initiated and prevented further bacterial infections. At 3 years of age, however, the patient developed, without an obvious trigger, fulminant HLH that was resistant to therapy. This patient shows that careful clinical and molecular diagnosis is essential to differentiate the complex disorders of lysosomal trafficking. HPSII belongs to the group of familial hemophagocytic syndromes and may represent an indication for HSCT.

Neuroblastoma (NB) expresses the tyrosine kinase receptors c-Kit, PDGFR-alpha and -beta-targets for STI-571. We investigated a possible combination therapy of STI-571 with retinoic acid (RA) and gamma-irradiation on NB cell viability in vitro. Expression of tyrosine kinase receptors and their ligands was examined in 6 NB cell lines by RT-PCR and FACS. The effect on cell viability was determined by MTT assay. Cell viability of all 6 NB cell lines was significantly inhibited after treatment with 20 microM STI-571 for 72h, two cell lines responding already to 10 microM. Cell lines responded irrespective of their mRNA status or cell surface expression of c-Kit, PDGFR-alpha and -beta. Co-incubation with 9-cis RA sensitized cells to the inhibitory effects of STI-571. However, pre-treatment with 9-cis RA resulted in resistance of NB cell lines to STI-571 and gamma-irradiation. Treatment of NB with STI-571 in combination with 9-cis RA might be a therapeutic strategy for patients in consolidation therapy who have completed gamma-irradiation therapy.

Purpose: In the Non-Hodgkin's Lymphoma-Berlin-Frankfurt-Munster (NHL-BFM) 95 trial, we tested, against the historical control of the combined trials NHL-BFM90 and NHL-BFM86, whether prophylactic cranial radiotherapy (PCRT) can be omitted for CNS-negative patients with stage III or IV lymphoblastic lymphoma (LBL) with sufficient early response., Patients and Methods: Apart from the removal of PCRT in NHL-BFM95, the chemotherapy of the three trials was identical except for the amount of l-asparaginase and daunorubicin during induction. The therapy in NHL-BFM95 was accepted to be noninferior when compared with trials NHL-BFM90/86 if the lower limit of the one-sided 95% CI for the difference in the 2-year probability of event-free-survival (pEFS) between target patients of NHL-BFM95 and the historical controls of NHL-BFM90/86 did not exceed -14%. The target patient group consisted of stage III and IV patients who were CNS negative and responded well to induction therapy., Results: The number of target patients was 156 in NHL-BFM95 (median age, 8.6 years; range, 0.2 to 19.5 years) and 163 in NHL-BFM90/86 (median age, 8.4 years; range, 0.6 to 16.6 years). For the target group, the pEFS rates at 2 and 5 years were 86% +/- 3% and 82% +/- 3%, respectively, in NHL-BFM95 (median follow-up time, 5.1 years; range, 2.1 to 9.1 years) compared with 91% +/- 2% and 88% +/- 3%, respectively in NHL-BFM90/86 (median follow-up time, 10.7 years; range, 5 to 15.4 years). The lower limit of the one-sided 95% CI for the difference in pEFS was -11% at 2 years and -13% at 5 years. In NHL-BFM95, one isolated and two combined CNS relapses occurred compared with one combined CNS relapse in NHL-BFM90/86. Five-year disease-free-survival rate was 88% +/- 3% in NHL-BFM95 compared with 91% +/- 2% in NHL-BFM90/86., Conclusion: For CNS-negative patients with stage III or IV LBL and sufficient response to induction therapy, treatment without PCRT may be noninferior to treatment including PCRT.

Background: Paravertebral masses of the fetus are often detected during routine prenatal ultrasonography. The most common differential diagnoses of these tumors are neuroblastoma, adrenal hemorrhage, schwannoma and germ cell tumors., Case Report: We report on a mature male newborn, who was diagnosed antenatally at 23 + 3 weeks of gestation with a tumor in the left paravertebral region. After birth the child was transferred to a neonatal unit. Tumor markers like urinary catecholamines were within normal limits, neuron-specific enolase was slightly elevated. MRI as well as ultrasonography confirmed a 3.2 x 2.2 x 1.6 cm large smoothed edged tumor in the left paravertebral region at the level of T10-T12. An open biopsy was performed, and the tumor which was located below the diaphragma was subtotally resected. Histopathology showed an extralobar pulmonary sequestration. Surgery as well as postoperative course was uneventful., Conclusions: Extralobar pulmonary sequestrations represent rare congenital anomalies, which are usually asymptomatic. Clear differentiation between tumor and pulmonary sequestration is seldom possible despite high resolution imaging studies. Hence, a biopsy procedure should be done for diagnosis of paravertebral masses.

Post-transplantation lymphoproliferative disorder is an infrequent complication after hematopoietic stem cell transplantation. It is hypothesized that lack of T-cell surveillance following transplantation permits reactivation of latent EBV leading to polyclonal B-cell expansion and finally outgrowth of a predominant clone. Most cases are of donor origin. Here, we describe an 8-year old boy with early onset post-transplantation lymphoproliferative disorder following matched-unrelated stem cell transplantation for high-risk T-cell leukemia whose disease was unusual for two reasons. First, his B-cell clone was of host origin and, in contrast to the few PTLD of host origin described so far, not associated with autologous reconstitution. Secondly, using clonal analysis, we could retrospectively show that the B-cell clone emerged during consolidation chemotherapy for T-cell leukemia, 3 months before stem cell transplantation.

There is a lack of valid epidemiological data on malignancy-associated pain in modern pediatric oncology. Pediatric oncology patients (self-assessment) and their parents from 28 hospitals were questioned using age-adapted, structured interviews and validated pain assessment tools. Pain intensity was measured by the NRS and Bieri faces scale. We conducted 363 interviews with patients and their parents, and 46 with the parents alone (if patients <2.5 years). Pain was reported at the time of the interview or within the last 24 h, 7 d, or 4 weeks in 15%, 28%, 50% and 58% of cases, respectively. The proportion of patients suffering severe to maximal pain (NRS>3; Bieri>2) increased significantly (p=0.001, chi2 test). The median pain intensity for the most severe pain episode within the last 4 weeks was 6.7 (NRS 0-10). Adverse effects of anti-tumor therapy were the most frequent cause of pain. Multivariate analyses depicted general physical condition either "severely reduced" (ASA status 3) (OR 4.0, 95% CI 1.1-14.7, p=0.037) or "moderately reduced" (ASA status 2) (OR 1.8, 95% CI 1.1-2.9, p=0.018), "in-patient status" (OR 1.8, 95% CI 1.2-2.9, p=0.010), and "co-morbidity present" (OR 3.5, 95% CI 1.1-10.7, p=0.030) as risk factors for severe to maximal pain. General anesthesia was the only factor significantly (OR 0.14, 95% CI 0.05-0.39, p<0.01) associated with a reduction in the proportion of patients suffering severe to maximal pain during bone marrow aspiration. Our data emphasize both the importance of in-house acute pain control and the need for general anesthesia during painful procedures in pediatric oncology.

Chimaerism analysis was performed by polymerase chain reaction amplification of short-tandem repeat markers in 30 children following haematopoietic stem cell transplantation for juvenile myelomonocytic leukaemia (JMML). Fourteen patients always had complete chimaerism (CC); one of them relapsed after the discontinuation of the study and 13 continued in complete remission (CR). Mixed chimaerism (MC) was noted in 16 patients. Of those 12 patients demonstrated increasing MC (i-MC); 10 relapsed and two achieved CC following discontinuation of immunosuppressive therapy (IST). Four other patients demonstrating only transient MC are alive in CR. MC with up to 20% of autologous cells could be successfully eradicated without induction of severe graft-versus-host disease when IST was reduced or discontinued directly after the first demonstration of MC. At the same time, MC with up to 10% of autologous cells could disappear without intervention. The interval between MC and relapse ranged from 0-320 d (median 38 d). Donor leucocyte infusion was given to six patients with i-MC, but only one patient responded. Peripheral blood seems as valuable as bone marrow for chimaerism studies. In conclusion, serial quantitative chimaerism studies can identify patients with i-MC who are at high risk for relapse of JMML. Immediate withdrawal of IST is advised in these patients.

Unlabelled: Neonates with Down syndrome can present with a haematological disorder called transient abnormal myelopoiesis (TAM). While TAM is usually a self-limiting disease, patients with severe complications such as hydrops fetalis, cardiorespiratory failure and liver fibrosis have been described. Here, we present five consecutive neonates with trisomy 21 and TAM, four of whom were critically ill and were therefore treated with cytosine-arabinoside. All five patients survived., Conclusion: severely affected neonates with Down syndrome and transient abnormal myelopoiesis might benefit from early cytostatic treatment with cytosine-arabinoside., (Copyright 2004 Springer-Verlag)

The resistance of neuroblastoma (NB) cells to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-induced apoptosis has been attributed to a lack of caspase 8 expression. Here we demonstrate a clinically applicable molecular targeting strategy that not only increases caspase 8 expression ex vivo in NB cell lines but also in the tumor tissues of NB patients receiving IFN-gamma treatment. We identify the functional caspase 8 promoter, which is different from the methylated region reported previously, and show promoter activity is up-regulated by IFN-gamma through a IFN-gamma activation site-containing region. IFN-gamma also induces TRAIL expression in NB cell lines. However, the IFN-gamma restoration of caspase 8 in some NB cells revealed persistent TRAIL resistance in most NB cell lines examined. This additional lesion in the TRAIL path is because of a loss of cell membrane TRAIL receptors (TR1/TR2) not only in cell lines but in most of the NB tumor tissues evaluated. Restoration of TR2 expression by transfection enhances IFN-gamma-induced TRAIL sensitivity. Furthermore, we have found that we can improve TRAIL sensitivity in NB by reconstituting caspase 8 with IFN-gamma and TR2 with chemotherapeutic agents.

The purpose of this study was to evaluate diffusion-weighted MR imaging in neuroblastomas. We prospectively examined seven children (age range 1-3 years) with seven solid body neuroblastomas. Diagnosis was established histologically. Diffusion-weighted echo-planar imaging (EPI) sequence was performed in all patients, with a repetition time of 5400 ms and an echo time of 103 ms, and with a b-value of 1000 s/mm(2). The contrast of tumour tissue depicted with T2-weighted images and diffusion-weighted images were evaluated by means of region-of-interest measurements and a calculation of the apparent diffusion coefficient (ADC) was done. The ADC calculation showed a mean ADC of 1.1x10(-3) (SD 0.14x10(-3), range 0.9-1.2x10(-3)) mm(2)/s of all tumours. Diffusion-weighted images showed an increased tumour signal. Water proton diffusion within the tumour matrix of neuroblastomas is especially restricted by the molecular and macromolecular barriers due to the very dense structure of this tumour tissue. We hypothesize that high nuclear-to-cytoplasm ratio of neuroblastoma cells limits intracellular motion. Furthermore, the very densely packed tumour cells inhibit effective motion of extracellular water protons. Restricted proton motion leads to a reduction in the rate of apparent diffusion and to a marked increase in signal on diffusion-weighted EPI MR images.

Paroxysmal nocturnal haemoglobinuria (PNH) is a clonal haematopoietic disorder characterized by chronic or intermittent intravascular haemolysis, variable cytopenia and an increased risk of thrombosis. Stem cell transplantation (SCT) is a curative therapeutic option, but its risks must be carefully weighed against the natural course of PNH. World-wide experience with SCT for PNH in the paediatric age group is scarce. We report on two adolescents suffering from PNH with life-threatening complications who were successfully transplanted from unrelated donors. Indications and techniques of SCT in childhood PNH are discussed and an overview of the literature is given.

In the cellular response to genotoxic stress, cell cycle checkpoint and apoptosis are considered to be two of the major biological events in maintaining genomic stability. The tumor suppressor p53 has been shown to play critical roles in these stress-induced cellular responses at least in part through the activation of its down-stream genes, such as p21CIP1/WAF1, GADD45 and BAX. In addition, p53 has been found to down-regulate the expression of BCL-2, which is able to block apoptosis induced by both p53-dependent and independent signaling events. In this report, we have found that increased expression of Bcl-2 protein in the human Burkitt's lymphoma WMN cell line suppressed apoptosis induced by different DNA-damaging agents. The induction of p53-regulated genes including GADD45, p21CIP1/WAF1 and BAX by genotoxic stress was substantially reduced in cells expressing high levels of Bcl-2 protein. Furthermore, Bcl-2 protein was shown to specifically suppress the p53-mediated transactivation of p21CIP1/WAF1 and PG13-CAT, which is a typical p53-binding-site reporter construct. Similarly, the inhibitory effect of Bcl-2 protein was seen in a GADD45 promoter reporter construct after treatment with methylmethane sulfonate or UV-radiation. These results indicate that in addition to its apoptosis-suppressing activity, Bcl-2 protein is able to inhibit transactivation of p53-regulated genes, which function in multiple important cellular responses to genotoxic stress, including the control of cell cycle checkpoints, cell growth suppression and DNA repair.

A 10-month-old girl with acute monocytic leukemia is presented. Ultrasound of the abdomen on admission showed an unusual distribution of leukemia infiltrates in the stomach and liver. The leukemic infiltrates in the wall of the stomach presented as intraluminal polyps, while the leukemic infiltration of the liver was strictly confined to the portal vessels.

Using flow cytometric analysis we investigated the distribution of major lymphocyte surface antigens in newborn infants. A total of 221 newborns entered the study, of whom 53 fulfilled our criteria of healthy mature neonates. Percentages of immunofluorescent-positive cells were as follows (median and range from 25th to 75th percentiles given): for CD1 3.8%; 2.3%-5.8%. CD2 60.9%; 52.4%-66.8%. CD3 57.5%; 50.5%-63.3%. TcRass 57.7%; 48.1%-60.0%. CD4 36.3%; 28.0%-42.6%. CD8 23.0%; 20.0%-27.4%. CD11a 56.3%; 46.3%-68.5%. CD19 12.1%; 8.6%-14.8%. CD20 10.9%; 8.4%-12.9%. CD25 2.6%; 2.1%-4.5%. CDw52 61.0%; 51.2%-76.1%. CD71 5.2%; 3.1%-9.3%. While the ranges for the percentage of immunofluorescent-positive cells were rather small, there was a wide variation in the absolute numbers of marker immunofluorescent-positive cells.

The incidence of shunt infections and possible risk factors was investigated by chart analysis. From 1986 to 1989 350 shunt procedures were performed including 273 ventriculoperitoneal shunts and 75 ventriculoatrial shunts. Twenty-eight infectious episodes (8%) occurred in 25 patients during a median follow-up time of 20 months. For 204 patients the follow-up time could be prolonged until September 1992. In these patients no infectious episodes occurred in the extended observation period. In 24 cases (85.7%) a causative organism could be isolated. The infecting organisms were gram-positive cocci in 22 cases (78.6%) and gram-negative bacilli in two cases. The main signs and symptoms were fever, shunt malfunction and meningeal irritation, and with VP-shunts only, abdominal pain. Twenty-four infectious episodes were treated with antibiotics and immediate removal of the shunt. The remaining were managed with antibiotics only. The risk for shunt infection did not correlate with age or sex of patients, nor with the etiology of hydrocephalus, type of shunt implanted or perioperative antibiotic prophylaxis. However, a trend showing a higher risk for shunt infections with prolonged operation time was noticed. The infection rate was 13.6% for an operation lasting more than 90 minutes versus 5.2% for procedures of less than 30 minutes duration.

Twenty three human mononuclear cell lines including ten myelomonocytic cell lines, eight B cell lines and five T cell lines, were examined to determine whether they could be infected with dengue-2 virus. All the cell lines were infected with dengue-2 virus as determined by immunofluorescent staining and by virus titration of culture supernatant fluids. K562, Jiyoye and Jurkat, respectively, showed the highest percentage of infected cells of these myelomonocytic, B and T cell lines. Antibody to dengue-2 virus at subneutralizing concentrations augmented dengue-2 virus infection of myelomonocytic cell lines, but not of B cell lines or of T cell lines. Persistent dengue-2 virus infection was established using a myelomonocytic cell line (K562), a B cell line (Raji), and a T cell line (HSB-2). These cell lines maintained a high percentage (more than 70%) of dengue-2 virus antigen-positive cells for at least 25 weeks. Very low titers of infectious dengue-2 virus were detected in the culture supernatant fluids of the persistently infected cells. Dengue-2 virus antigen-positive Raji cell clones were established from persistently-infected Raji cells using limiting dilutions and all of the cells in these clones were dengue-2 virus antigen-positive. These findings demonstrate that a variety of human mononuclear cell lines can be infected with dengue-2 virus and may be useful as models for the analysis of dengue virus-human cell interactions in dengue virus infections.