The impact of treatment for childhood classical Hodgkin lymphoma according to the EuroNet-PHL-C2 protocol on serum anti-Müllerian Hormone.

STUDY QUESTION: What is the impact of the EuroNet-PHL-C2 treatment protocol for children with classical Hodgkin lymphoma (cHL) on gonadal function in girls, based on assessment of serum anti-Müllerian hormone (AMH)? SUMMARY ANSWER: Serum AMH levels decreased after induction chemotherapy and increased during subsequent treatment and 2 years of follow-up, with lowest levels in patients treated for advanced stage cHL. WHAT IS KNOWN ALREADY: Treatment for cHL, particularly alkylating agents and pelvic irradiation, can be gonadotoxic and result in premature reduction of primordial follicles in females. The current EuroNet-PHL-C2 trial aims to reduce the use of radiotherapy in standard childhood cHL treatment, by intensifying chemotherapy. This study aims to assess the gonadotoxic effect of the EuroNet-PHL-C2 protocol. STUDY DESIGN, SIZE, DURATION: This international, prospective, multicenter cohort study is embedded in the EuroNet-PHL-C2 trial, an European phase-3 treatment study evaluating the efficacy of standard cHL treatment with OEPA-COPDAC-28 (OEPA: vincristine, etoposide, prednisone, and doxorubicin; COPDAC-28: cyclophosphamide, vincristine, prednisone, and dacarbazine) versus intensified OEPA-DECOPDAC-21 (DECOPDAC-21: COPDAC with additional doxorubicin and etoposide and 25% more cyclophosphamide) in a randomized setting. Participants were recruited between January 2017 and September 2021. PARTICIPANTS/MATERIALS, SETTING, METHODS: Female patients aged ≤18 years, treated according to the EuroNet-PHL-C2 protocol for cHL were recruited across 18 sites in the Netherlands, Belgium, Germany, Austria, and Czech Republic. All parents and patients (aged ≥12 years old) provided written informed consent. Serum AMH levels and menstrual cycle characteristics were evaluated over time (at diagnosis, one to three times during treatment and 2 up to 5 years post-diagnosis) and compared between treatment-levels (TL1, TL2, and TL3) and treatment-arms (OEPA-COPDAC-28 and OEPA-DECOPDAC-21). Serum samples obtained from patients after receiving pelvic radiotherapy were excluded from the main analyses. MAIN RESULTS AND THE ROLE OF CHANCE: A total of 104 females, with median age at diagnosis of 15.6 years (IQR 13.7; 17.0), were included in the analysis. Ninety-nine were (post)pubertal. Eighteen girls were diagnosed with an early stage of cHL (TL1) and 86 with intermediate or advanced stage disease (50 TL2 and 36 TL3, 66% received COPDAC-28 and 34% DECOPDAC-21). Five patients received pelvic radiotherapy. Median AMH level at diagnosis was 1.7 µg/l (IQR 0.9; 2.7). After two courses of OEPA chemotherapy, AMH levels decreased substantially in all patients (98% <0.5 µg/l), followed by a significant increase during the consolidation treatment and follow-up. After 2 years, 68% of patients reached their baseline AMH value, with overall median recovery of 129% (IQR 75.0; 208.9) compared to baseline measurement. Five patients (7%) had AMH <0.5 µg/l. In patients treated for advanced stage disease, AMH levels remained significantly lower compared to early- or intermediate stage disease, with median serum AMH of 1.3 µg/l (IQR 0.8; 2.1) after 2 years. Patients who received DECOPDAC-21 consolidation had lower AMH levels during treatment than patients receiving COPDAC-28, but the difference was no longer statistically significant at 2 years post-diagnosis. Of the 35 postmenarchal girls who did not receive hormonal co-treatment, 19 (54%) experienced treatment-induced amenorrhea, two girls had persisting amenorrhea after 2 years. LIMITATIONS, REASONS FOR CAUTION: The studied population comprises young girls with diagnosis of cHL often concurring with pubertal transition, during which AMH levels naturally rise. There was no control population, while the interpretation of AMH as a biomarker during childhood is complex. The state of cHL disease may affect AMH levels at diagnosis, potentially complicating assessment of AMH recovery as a comparison with baseline AMH. The current analysis included data up to 2-5 years post-diagnosis. WIDER IMPLICATIONS OF THE FINDINGS: The current PANCARE guideline advises to use the cyclophosphamide-equivalent dose score (CED-score, as an estimation of cumulative alkylating agent exposure) with a cut-off of 6000 mg/m2 to identify females aged <25 years at high risk of infertility. All treatment-arms of the EuroNet-PHL-C2 protocol remain below this cut-off, and based on this guideline, girls treated for cHL should therefore be considered low-risk of infertility. However, although we observed an increase in AMH after chemotherapy, it should be noted that not all girls recovered to pre-treatment AMH levels, particularly those treated for advanced stages of cHL. It remains unclear how our measurements relate to age-specific expected AMH levels and patterns. Additional (long-term) data are needed to explore clinical reproductive outcomes of survivors treated according to the EuroNet-PHL-C2 protocol. STUDY FUNDING/COMPETING INTEREST(S): The fertility add-on study was funded by the Dutch charity foundation KiKa (project 257) that funds research on all forms of childhood cancer. C.M-K., D.K., W.H.W., D.H., M.C., A.U., and A.B. were involved in the development of the EuroNet-PHL-C2 regimen. The other authors indicated no potential conflicts of interest. TRIAL REGISTRATION NUMBER: N/A.

Prevention of activated brown adipose tissue on 18F-FDG-PET scans of young lymphoma patients: results of an ancillary study within the EuroNet-PHL-C2 trial.

Activated brown fat (aBAT) is known to affect the evaluation of 18F-FDG PET scans, especially in young patients. The aim of this study was to determine factors influencing the occurrence of aBAT, and to investigate the effectiveness of the two preventive measures, warming and beta-blocker (propranolol) administration. Five-hundred-twenty-eight 18F-FDG-PET scans of 241 EuroNet-PHL-C2 trial patients from 41 nuclear medicine departments in Germany and Czech Republic were screened for aBAT. The occurrence of aBAT was analyzed with patient characteristics (age, sex, body mass index, predisposition to aBAT), weather data at the day of 18F-FDG PET scanning as well as the preventive measures taken. Potentially important factors from univariate analyses were included into a logistic regression model. Warming as a preventive measure was used in 243 18F-FDG-PET scans, propranolol was administered in 36, warming and propranolol were combined in 84, and no preventive measures were taken in 165 scans. Whereas age, sex and body mass index had no clear impact, there was an individual predisposition to aBAT. Logistic regression model revealed that the frequency of aBAT mainly depends on the outside temperature (p = 0.005) and can be effectively reduced by warming (p = 0.004), the administration of unselective beta-blocker or the combination of both. Warming is a simple, cheap and non-invasive method to reduce the frequency of aBAT. However, the effect of warming decreases with increasing outside temperatures. Administration of propranolol seems to be equally effective and provides advantages whenever the positive effect of warming is compromised. The combination of both preventive measures could have an additive effect.

Advantage of proton-radiotherapy for pediatric patients and adolescents with Hodgkin's disease.

Radiotherapy is frequently used in the therapy of lymphoma. Since lymphoma, for example Hodgkin's disease, frequently affect rather young patients, the induction of secondary cancer or other long-term adverse effects after irradiation are important issues to deal with. Especially for mediastinal manifestations numerous organs and substructures at risk play a role. The heart, its coronary vessels and cardiac valves, the lungs, the thyroid and, for female patients, the breast tissue are only the most important organs at risk. In this study we investigated if proton-radiotherapy might reduce the dose delivered to the organs at risk and thus minimize the therapy-associated toxicity. METHODS: In this work we compared the dose delivered to the heart, its coronary vessels and valves, the lungs, the thyroid gland and the breast tissue by different volumetric photon plans and a proton plan, all calculated for a dose of 28.8 Gy (EURO-NET-PHL-C2). Target Volumes have been defined by F18-FDG PET-positive areas, following a modified involved node approach. Data from ten young female patients with mediastinal lymphoma have been evaluated. Three different modern volumetric IMRT (VMAT) photon plans have been benchmarked against each other and against proton-irradiation concepts. For plan-evaluation conformity- and homogeneity-indices have been calculated as suggested in ICRU 83. The target volume coverage as well as the dose to important organs at risk as the heart with its substructures, the lungs, the breast tissue, the thyroid and the spinal cord were calculated and compared. For statistical evaluation mean doses to organs at risk were evaluated by non- parametric Kruskal-Wallis calculations with pairwise comparisons. RESULTS: Proton-plans and three different volumetric photon-plans have been calculated. Proton irradiation results in significant lower doses delivered to organ at risk. The median doses and the mean doses could be decreased while PTV coverage is comparable. As well conformity as homogeneity are slightly better for proton plans. For several organs a risk reduction for secondary malignancies has been calculated using literature data as reference. According to the used data derived from literature especially the secondary breast cancer risk, the secondary lung cancer risk and the risk for ischemic cardiac insults can be reduced significantly by using protons for radiotherapy of mediastinal lymphomas. CONCLUSION: Irradiation with protons for mediastinal Hodgkin-lymphoma results in significant lower doses for almost all organs at risk and is suitable to reduce long term side effects for pediatric and adolescent patients.

Only strongly enhanced residual FDG uptake in early response PET (Deauville 5 or qPET ≥ 2) is prognostic in pediatric Hodgkin lymphoma: Results of the GPOH-HD2002 trial.

PURPOSE: In 2014, we published the qPET method to quantify fluorodeoxyglucose positron emission tomography (FDG-PET) responses. Analysis of the distribution of the quantified signals suggested that a clearly abnormal FDG-PET response corresponds to a visual Deauville score (vDS) of 5 and high qPET values ≥ 2. Evaluation in long-term outcome data is still pending. Therefore, we analyzed progression-free survival (PFS) by early FDG-PET response in a subset of the GPOH-HD2002 trial for pediatric Hodgkin lymphoma (PHL). PATIENTS/METHODS: Pairwise FDG-PET scans for initial staging and early response assessment after two cycles of chemotherapy were available in 93 PHL patients. vDS and qPET measurement were performed and related to PFS. RESULTS: Patients with a qPET value ≥ 2.0 or vDS of 5 had 5-year PFS rates of 44%, respectively 50%. Those with qPET values < 2.0 or vDS 1 to 4 had 5-year PFS rates of 90%, respectively 80%. The positive predictive value of FDG-PET response assessment increased from 18% (9%; 33%) using a qPET threshold of 0.95 (vDS ≤ 3) to 30% (13%; 54%) for a qPET threshold of 1.3 (vDS ≤ 4) and to 56% (23%; 85%) when the qPET threshold was ≥ 2.0 (vDS 5). The negative predictive values remained stable at ≥92% (CI: 82%; 98%). CONCLUSION: Only strongly enhanced residual FDG uptake in early response PET (vDS 5 or qPET ≥ 2, respectively) seems to be markedly prognostic in PHL when treatment according to the GPOH-HD-2002 protocol is given.

Pediatric Nodular Lymphocyte-predominant Hodgkin Lymphoma: Treatment Recommendations of the GPOH-HD Study Group.

Nodular lymphocyte-predominant Hodgkin lymphoma (nLPHL) is a very rare disease in childhood and adolescence. In Germany, about 15 newly diagnosed patients present with this disease annually; this number comprises less than 10% of all pediatric Hodgkin lymphoma cases. Since the EuroNet-PHL-LP1 trial for early stage nLPHL patients stopped recruiting in Germany in October 2014, the GPOH-HD writing committee reviewed the literature and decided to deliver treatment recommendations for childhood and adolescent nLPHL patients. These guidelines shall be applicable to young nLPHL patients in European countries that will no longer be able to participate in nLPHL trials for young patients. Therefore, the EuroNet-PHL-nLPHL-registry will be installed to provide quality assured central review of staging and response assessment for registered patients by the Central Review Board of EuroNet-PHL in Halle/Leipzig, Germany.

[Early childhood intervention - access to risk families and support through actors from the health-care sector]. / Frühe Hilfe für Familien und Frühgeborene - Zugangswege und Unterstützungen aus dem Gesundheitsbereich.

Interdisciplinary cooperation and networking determine the success of activities for supporting families at risk for early childhood abuse. The integration of the healthcare sector might be important.The medical standard of perinatal care at the University hospital includes information exchange about family risk factors which may contribute to an increased risk of child abuse within the first year of life. As a result, the -pediatrician offered supporting services for the families at the time of the second examination during the official childhood health screening program (U2). A team of family-sponsorship was established and evaluated.In 281 of 1238 risk-factor questionnaires at least one stress factor was detected and 97 families had high-impact family stress. Families under the supervision of a family midwife or youth services had a significantly higher number of risk factors. The family-sponsorship program was institutionalized and positively evaluated by the families.The time of a hospital delivery is an excellent opportunity for the evaluation of familial risk factors and for the provision of supporting services. To increase the acceptance of such services by the families at risk repeated assessment of risk factors and support offers are required.

The EuroNet paediatric hodgkin network - modern imaging data management for real time central review in multicentre trials.

Since 2007, children and adolescents with Hodgkin lymphomas are treated in the Europe-wide EuroNet-PHL trials. A real time central review process for stratification of the patients enhances quality control and efficient therapy management. This process includes reading of all cross-sectional-images. Since reference evaluation is time critical, a fast, easy to handle and safe data transfer is important. In addition, immediate and constant access to all the data has to be guaranteed in case of queries and for regulatory reasons. To meet the mentioned requirements the EuroNet Paediatric Hodgkin Data Network (funded by the European Union - Project Number: 2007108) was established between 2008 and 2011. A respective tailored data protection plan was formulated. The aim of this article is to describe the networks' mode of operation and the advantages for multi-centre trials that include centralized image review.

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.

Evaluation of the prognostic meaning of C-reactive protein (CRP) in children and adolescents with classical Hodgkin's lymphoma (HL).

BACKGROUND: In adult cancer patients the negative predictive value of elevated CRP levels has been described for several malignancies. Only few studies have analyzed the prognostic role of CRP in children and adolescents with classical HL. In these studies elevated CRP levels correlate with the presence of classical risk factors and adverse outcome. PATIENTS AND METHODS: The prognostic role of CRP for patients with classical HL admitted to the GPOH-HD-2002 study was analyzed retrospectively. RESULTS: CRP levels were documented for 369 of 573 patients. Significant (p<0.05) increased median CRP levels were found in the presence of B-Symptoms (25.7 vs. 5.1 mg/l), extranodal involvement (21.5 vs. 7.5 mg/l), elevated erythrocyte sedimentation rate (ESR, 13.0 vs. 1.0 mg/l) and stage III/IV disease (15.5 vs. 5.3 mg/l). 83.9% of patients with elevated and 45.8% of patients with normal CRP had an ESR >30 mm/h. CONCLUSION: Elevated CRP levels were associated with classical risk factors of HL. CRP and ESR may reflect different biological processes. CRP was prognostic within early stage TG-1 patients treated with reduced treatment, but not within advanced stage TG-2+3.

Human leukocyte antigen distribution in German Caucasians with advanced Ewing's sarcoma.

BACKGROUND: Risk stratification criteria for patients with Ewing's sarcoma family of tumors (ESFT) are still limited. We hypothesized divergent human leukocyte antigen (HLA) patterns in ESFT patients and compared HLA-A, -B and -DR phenotype frequencies of patients with advanced ESFT with those of healthy controls. PATIENTS: HLA types of all German Caucasian patients with advanced ESFT and available HLA-A, -B and -DR data registered in the European Group for Blood and Marrow Transplantation, Paediatric Registry for Stem Cell Transplantation and the MetaEICESS data bases (study group, n=30) were retrospectively compared with HLA types of healthy German stem cell donors (control group, n=8 862 for single HLA frequencies and n=8 839 for allele combinations). Study group patients had been immuno-typed due to eligibility for allogeneic stem cell transplantation for high risk of treatment failure, and thus constituted a selected subgroup of ESFT patients. RESULTS: After Bonferroni correction for multiple testing (PC), phenotype frequencies of HLA-A24 remained significantly higher in the study group compared to controls (PC<0.05). Furthermore, several HLA combinations were significantly more frequent in the study group compared to controls (all PC<0.05). CONCLUSION: We report an increased incidence of circumscribed HLA patterns in German Caucasians with advanced ESFT. The possible clinical significance of this observation has to be re-assessed in prospective trials comprising larger ESFT patient numbers of all risk groups.

High dose methotrexate treatment in childhood ALL: pilot study on the impact of the MTHFR 677C>T and 1298A>C polymorphisms on MTX-related toxicity.

Methotrexate (MTX) is commonly administered in high doses for treatment of childhood acute lymphoblastic leukemia (ALL). The aim of this analysis was to study the influence of 2 common MTHFR polymorphisms (MTHFR 677C>T and 1298 A>C) on MTX toxicity in children with ALL.Retrospective analysis of 129 MTX courses in 34 pediatric patients with ALL.677C>T variants (CT or TT) were found in 19 (14 heterozygous, 5 homozygous) and 1298A>C variants (AC or CC) in 20 (16 heterozygous, 4 homozygous) patients. The MTHFR 677C>T wild type was associated with an increased frequency of grade III and IV leukopenia (60% vs. 31%, p<0.05) compared to the variants. The rate of severe infections (21% vs. 0%, p<0.05) and grade III-IV anemia (26% vs. 5%, p<0.05) was increased in carriers of the MTHFR 677C>T wild type compared to patients with the TT variant. Grade III-IV anemia was more frequent in patients with the MTHFR 1298A>C CC variant compared to the wild type (56% vs. 21%, p<0.05). The differences were not significant in a patient-based analysis.MTX related toxicity might be influenced by the MTHFR 677C>T or the MTHFR 1298A>C polymorphisms. Differences in MTX toxicity are only partially explainable by these 2 polymorphisms.

General insufficiency of the classical CDC-based crossmatch to detect donor-specific anti-HLA antibodies leading to invalid results under recipients' medical treatment or underlying diseases.

Antibodies directed against HLA antigens of a given donor represent the most prominent cause for hyper-acute and acute rejections. In order to select recipients without donor-specific antibodies the complement-dependent cytotoxicity (CDC-) crossmatch as the standard procedure was established. As a functional assay it strongly depends on the availability of isolated donor lymphocytes and in particular on their vitality. However, due to several diseases or pharmacological treatment of a given recipient unexpected "false-positive" results of the CDC-crossmatch may arise. We here present three groups of patients which demonstrate the limits of the conventional crossmatch. 1) Kidney recipients before living donations exhibited positive CDC-reactions due to their conditioning using the therapeutical anti-CD20 mAb Rituximab (n=7), routinely used to deplete B-cells, or the anti-CD25 mAb basiliximab (n=2) to inhibit the proliferation of activated T-cells. 2) Recipients suffering from various leukaemias (n=5) exhibited "positive" CDC-crossmatches using PBL of the donors, although formerly these patients had never shown anti-HLA antibodies. Instead of donor-specific allo-antibodies, cytostatic agents such as 6-mercaptopurine led to an unspecific cell death. 3) Patients projected for post mortem or living kidney donations (n=44) exhibited "positive" CDC-crossmatch results which were not in accordance with their former antibody status and, partially, with high degrees of HLA-matching. These implausible results were due to underlying auto-immune diseases, mainly of the systemic immune complex type III such as lupus erythematosus, mainly leading to false-positive B-cell crossmatches by immune complexes binding to Fcγ-receptors. In all these 58 cases the alternatively performed ELISA-based "Antibody Monitoring System" (AMS-) crossmatch assay was not artifically affected, suggesting that this assay may be comprehensively established at least for the cases described.

Acute psychosis in an adolescent girl during treatment for advanced nodular lymphocyte-predominant Hodgkin's lymphom.

A 16,5 year old female adolescent was diagnosed with nodular lymphocyte-predominant Hodgkin lymphoma (nLPHL), presenting bilateral inguinal and right iliac lymphadenopathy accompanied by B-symptoms. The patient was due to treatment according to the German Interim Guidelines of HD 2002-Pilot Study with 2x OPPA (vincristine, adriamycine, prednisone, procarbacine) and 2x COPP (cyclophosphamide, vincristine, prednisone, procarbazine) and radiotherapy. After 2x OPPA the patient presented a severe episode of a presumably prednisone-induced acute psychosis with need for psychiatric treatment and change of therapy regimen. She was successfully treated with a chimeric monoclonal anti-CD20 antibody (rituximab) and subsequent radiotherapy.

Management of oncology patients admitted to the paediatric intensive care unit of a general children's hospital - a single center analysis.

BACKGROUND: The improving prognosis of children with cancer has partially been attributed to the increasing importance of pediatric intensive care units (PICU). We analyze whether outcome of these patients on a PICU improved during the last decade and which factors may influence the outcome in our hospital. PATIENTS AND METHODS: The charts of all oncology patients admitted to the PICU between 1998 and 2009 have been reviewed retrospectively. The survival of patients admitted for life threatening complications has been correlated with basic data, organ failure and the PRISM score. The results of 2 consecutive treatment periods (1998-2003 and 2004-2009) were compared. RESULTS: 644 admissions of 226 patients were recorded. 79 admissions were performed because of potentially life threatening complications (Group A), 236 for monitoring (B) and 329 admissions for interventions (C). 62% of Group A patients and all Group B and C patients were discharged alive. Poor outcome was associated with admission >28 days after initial diagnosis, PRISM >10, organ failure >2 organs, sepsis, allogeneic stem cell transplantation, need for mechanical ventilation or for catecholamines. The PICU survival rate of Group A patients admitted between 2004 and 2009 (78%) was higher than in the period between 1998 and 2003 (48%). CONCLUSIONS: PICU provides essential services to support the pediatric oncology ward. Although children with cancer may have had benefit from advances in pediatric intensive care over the past decade, specific scoring systems for early identification of children with cancer needing PICU treatment are required. These systems might further improve PICU outcome in critical ill pediatric cancer patients.

[Immune tolerance induction with high-dose FVIII and pulsed intravenous immunoglobulin]. / Immuntoleranzinduktion mit hoch dosiertem FVIII und intravenösen Immunglobulin-Pulsen. Ein Patient mit Hemmkörper-Hämophilie A.

UNLABELLED: The development of neutralizing allo-antibodies against factor VIII (FVIII) or FVIII inhibitors is a severe complication in the treatment of haemophilia A. About 25% of the children with severe haemophilia A develop FVIII inhibitors. Here we report on a boy with severe haemophilia A and intron 22 inversion of the FVIII gene who was diagnosed at ten months of age. After 16 exposure days to FVIII (81 days after initial exposure) he developed a FVIII inhibitor (maximum: 9.76 BU/ml). THERAPY: We started immune tolerance induction (ITI) according to the Bonn protocol with high dose plasma derived FVIII concentrate (100 IU per kg body weight) twice daily. For additional inhibitor elimination treatment the patient received intravenous immunoglobulin (ivIg) at a dose of 1-2 g/kg body weight every 4 to 6 weeks. After start of treatment a rapid decline of the inhibitor level was observed, nevertheless low FVIII inhibitor levels persisted (<5 BU/ml). Furthermore, the FVIII half-life was still accelerated. However, after every course of ivIg the inhibitor level declined and FVIII half-life was prolonged. Currently, the FVIII half-life is approaching normal values after more than seven months of ITI duration. CONCLUSION: Additional application of immunoglobulin is beneficial for immune tolerance induction.

[FDG-PET in Hodgkin lymphoma]. / FDG-PET beim Hodgkin-Lymphom.

The high negative predictive value of FDG-PET in therapy control of Hodgkin lymphoma is proven by the data acquired up to now. Thus, the analysis of the HD15 trial has shown that consolidation radiotherapy might be omitted in PET negative patients after effective chemotherapy. Further response adapted therapy guided by PET seems to be a promising approach in reducing the toxicity for patients undergoing chemotherapy. The criteria used for the PET interpretation have been standardized by the German study groups for Hodgkin lymphoma patients and will be reevaluated in the current studies.

Molecular mechanisms and gene regulation of melphalan- and hyperthermia-induced apoptosis in Ewing sarcoma cells.

The prognosis of high-risk Ewing tumours (HR-ET) remains poor. Melphalan-containing chemotherapy regimens are commonly applied for HR-ET patients. Moreover, melphalan (Mel) is a promising agent in thermochemotherapy. Therefore, we investigated the single effects, the synergism and the gene regulation of Mel and hyperthermia (HT) in an ET cell line (RD-ES). Dose-dependent cytotoxicity by Mel was demonstrated, which was enhanced by the concomitant application of HT (42 degrees C for 2 h). Mel, HT and their combination caused a significant activation of caspase-3. Using the pan-caspase inhibitor z-VAD-fmk, we demonstrated that both stimuli mediated predominantly caspase-dependent cytotoxicity. With cDNA array analysis, 20 out of 198 apoptosis-related genes were identified to be differentially expressed by Mel and/or HT. Although a significant enhancement of three selected genes could not be proven at the protein level in subsequent experiments, this study gives insight into the complex molecular and genetic response of tumour cells to cytotoxic stimulation.

Determination of terephthalic acid isopropylamide in urine with a liquid chromatography/mass spectrometry (LC/MS) method.

A sensitive and simple liquid chromatography/mass spectrometry (LC/MS) method was developed for the determination of terephthalic acid isopropylamide, the final metabolite of procarbazine in human urine. A solid-phase extraction with C(18) cartridges was used followed by LC/MS with a single mass spectrometer (SSQ 7000 from Finnigan). Terephthalic acid isobutylamide was the internal standard. The quantification limit was 30 ng/mL in urine (6 x noise). This assay was applied for drug monitoring of terephthalic acid isopropylamide in urine after oral administration of procarbazine in children and adolescents with Hodgkin lymphomas.