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Aim: The European Network of Excellence for Paediatric Clinical Research, known as the TEDDY Network, carried out a survey to determine the capacity and competence of paediatric centres to perform research studies. Methods: A cross-sectional, web-based pilot survey was conducted from October 2016 to April 2017 with paediatric clinical research centres in 11 countries: Albania, Austria, Belgium, Denmark, Iceland, Ireland, Italy, Norway, Spain, Switzerland and the United Kingdom. All were registered with the TEDDY Network database. Results: We approached 107 centres and 63 provided data on their experiences and expertise in paediatric clinical trials. Four groups of performance indicators were identified, referring to scientific experience, trial readiness, trial competence, regulatory issues, ethics and patients. Most centres were actively involved in paediatric clinical research: 53 centres (84.1%) had received funds for more than five paediatric studies in the last 5 years, and 42 (66.7%) had a specific clinical trial unit and dedicated study coordinators. We concluded that the European centres we studied had the capability and capacity to conduct paediatric trials, but there was still room for improvement, including enhanced collaboration. Conclusion: This pilot survey demonstrated that there is potential for performing paediatric trials across Europe, but improvements are possible.

Lymphoproliferative diseases (LPDs) associated with Epstein-Barr virus (EBV) infection cause significant morbidity and mortality in bone marrow and solid organ transplant recipients. To gain insight into LPD pathogenesis and to identify potential effective therapeutic approaches, we investigated early molecular events leading to B-cell transformation by gene expression profiling of EBV-infected B-cells from tonsils by Affymetrix microarray 72 hr postinfection when the B-cells hyperproliferation phase starts. Cell cycle and apoptosis were the most significantly affected pathways and enriched gene sets. In particular, we found significantly increased expression of cyclin-dependent kinase (CDK)1 and CCNB1 (cyclin B1) and of one of their downstream targets BIRC5 (survivin). Importantly, the strong upregulation of the antiapoptotic protein survivin was confirmed in lymphoblastoid cell lines (LCLs) and 71% of EBV-positive post-transplant EBV-LPD lesions scored positive for survivin. The validity of early transforming events for the identification of therapeutic targets for EBV-LPD was confirmed by the marked antiproliferative effect of the CDK inhibitor flavopiridol on LCLs and by the strong induction of apoptosis by survivin inhibition with YM155 or terameprocol. Our results suggest that targeting of CDKs and/or survivin in post-transplant EBV-LPD by specific inhibitors might be an important approach to control and eliminate EBV-transformed B-cells that should be further considered.

Fever and chemotherapy-induced neutropenia (FN) is the most frequent potentially lethal complication of therapy in children with cancer. This study aimed to describe serious medical complications (SMC) in children with FN regarding incidence, clinical spectrum, and associated characteristics.

HIV infection is characterized by sustained immune activation, which is reflected by activated T cells and, in particular, by increased levels of phosphorylated STAT proteins. Here, we hypothesized that T-cell activation in HIV infection is partially due to the inability of SOCS-1 and SOCS-3 to control the JAK/STAT pathway. We found higher levels of SOCS-1/3 mRNA levels in CD4(+) T cells of HIV-infected patients than in healthy controls. However, SOCS protein levels were lower, explaining the lack of attenuation of the JAK/STAT pathway. Infection of CD4(+) T cells alone did not activate STATs, while ex vivo infection of PBMC did, indicating that non-T cells critical for shaping the immune response, e.g. DC were responsible for the STAT-1 activation. Supernatants from ex vivo-infected PBMC transferred to CD4(+) T cells induced JAK/STAT activation, pointing to a central role of soluble factors. Notably, over-expression of SOCS-1/3 in CD4(+) T cells prevented JAK/STAT activation. Thus, HIV infection interferes with SOCS-1/3 expression driving immune activation. Sustained immune activation disrupts the lymphoid system and favors HIV replication since HIV preferentially infects activated cells. We speculate that regulating SOCS may be a potential way to counteract immune activation in HIV disease.

Quantitative real-time polymerase chain reaction (PCR) was utilized to measure serum EBV DNA levels of BamHI W fragment and latent membrane protein 1 (LMP1) in 20 nasal natural killer (NK)/T-cell lymphoma patients. Both serum EBV DNAs were detected at high levels in all patients, but the levels were below the limit of detection in all healthy controls. The BamHI Z fragment, Epstein-Barr-replication activator (ZEBRA) expression was detected in a small proportion (0.1-3%) of lymphoma cells from 10 (50%) of the patients. Patients with ZEBRA expression showed significantly higher DNA levels of BamHI W and LMP1 (P = 0.0081, P = 0.004), suggesting that EBV DNA may be caused by EBV replication from lymphoma cells. Kaplan-Meier and univariate analyses revealed that high DNA levels of BamHI W and LMP1 at pre-treatment and high BamHI W DNA level at post-treatment were associated with short disease-free survival and overall survival (P < 0.05, each). Although the DNA levels of BamHI W and LMP1 correlated significantly, their dynamics were not always parallel. Patients with low pre-treatment level of both EBV DNAs showed a favorable course, in contrast to patients with high pre-treatment level of both EBV DNAs who showed an aggressive course (P = 0.0085). More importantly, the high pre-treatment level of both EBV DNAs was determined as the only independent prognostic factor among various prognostic factors. These data suggest that simultaneous measurement of serum levels of both BamHI W and LMP1 DNAs may be useful for diagnosis, disease monitoring, and prediction of prognosis for nasal NK/T-cell lymphoma patients.

A real-time polymerase chain reaction was employed to detect and quantitate Epstein-Barr virus (EBV) DNA in tonsils and autologous sera from EBV-seropositive children. EBV DNA was found in 95% of tonsils from 21 children and in 50% from 18 children with serum IgG titers to the virus capsid antigen (VCA) of ≥1:160 and 1:10 to 1:80, respectively (P = 0.002). Tonsils from children with titers ≥1:160 harbored more EBV DNA copies per mg tissue (mean, 1,237; range

A real-time polymerase chain reaction assay for quantitation of Epstein-Barr virus (EBV) DNA in serum was developed. This assay detected EBV DNA in 24 (89%) of 27 sera from patients with infectious mononucleosis, but only in 9 (18%) of 51 sera from EBV carriers (P < 0.001) and in none of the sera from 32 EBV-seronegative individuals. EBV DNA levels were higher in sera from infectious mononucleosis (median 8,000, range 1833-150,069 copies/ml) than from carriers (median < 2, range < 2-2980; P < 0.001). In sera of 36 children with infectious mononucleosis followed prospectively, EBV DNA levels correlated inversely with the duration of symptoms. Among 18 children with tumors including Hodgkin's disease (n = 7), non-Hodgkin's lymphoma (n = 6), Burkitt's lymphoma (n = 1), lymphoproliferative disorder (n = 4), and osteosarcoma (n = 1), EBV DNA was detected in serum from those 9 (100%) expressing EBV in the tumor (Hodgkin's disease, 3; non-Hodgkin's lymphoma, 2; lymphoproliferative disorder, 4), the levels peaking at diagnosis and correlating with disease activity. Quantitation of EBV DNA in serum may offer a simple means of monitoring patients at risk of EBV-associated lymphoproliferation.

The latent membrane proteins LMP1 and LMP2A are co-expressed in most malignancies associated with Epstein-Barr virus (EBV). In contrast with the transforming LMP1 oncoprotein, LMP2A is expressed in lymphocytes of healthy EBV carriers and considered to maintain viral latency. Critical for these LMP2A functions are a transmembranous epitope recognized by specific cytotoxic T lymphocytes (CTLs) and the N-terminal immunoreceptor tyrosine-based activation motif (ITAM), blocking B-cell receptor signaling. To characterize ITAM and CTL motifs of LMP2A and to correlate them with C-terminal variants of LMP1 including the 30-bp deletion variant (LMP1Δ), comparative sequence analysis was performed on 76 samples from patients with reactive and malignant lympho-proliferation (infectious mononucleosis, n = 21; tonsillar hyperplasia, n = 16, chronic lympho-proliferation, n = 9; Hodgkin's disease, n = 8; Non-Hodgkin's lymphoma, n = 5; AIDS-related large-cell lymphoma, n = 17). The CTL motif was conserved in all but 2 cases (C426S). The ITAM motif was characterized by strictly conserved YXXL sequences in all cases, with a sequence polymorphism in between. The B95.8 prototype was found in 17% (13/76) of cases, while in 72% a variant with 3 point mutations (166796 CA, 166805 CA, 166810 CT) was detected; 11% had 1 or 2 of these mutations in addition to GA at 166793. In the C terminus of LMP1, a hypervariable region including LMP1Δ was described in 61% of cases. There was no significant association of a particular LMP2A variant with either malignant phenotype or LMP1Δ, demonstrating that the functional domains of LMP2A are conserved and that the sequence polymorphisms in LMP1 and LMP2A are independent. Int. J. Cancer 81:371–375, 1999. © 1999 Wiley-Liss, Inc.

Background. Reports on lymphoid malignancy and its treatment in children infected with human immunodeficiency virus (HIV) are limited. Methods. Antibodies to Epstein-Barr virus (EBV) were detected by indirect immunofluorescence. DNA was extracted from peripheral blood lymphocytes or biopsy specimens. Polymerase chain reaction was run using primers for EBV. Reaction products underwent Southern blot analysis to confirm EBV specificity. Tumor clonality was assessed by immunohistochemistry and by Southern blot analysis of immunoglobulin heavy-chain and T-cell receptor β-gene rearrangement. Results. Within 1 year, non-Hodgkin's lymphoma (NHL) was diagnosed in four children infected with HIV. All four were EBV-seropositive and had detectable EBV DNA in peripheral blood lymphocytes. The EBV-linked disorders lymphoid interstitial pneumonia and recurrent parotid enlargement preceded NHL in three and two of the children, respectively. In all four patients, NHL involved at one time the central nervous system (CNS). All three tested NHL tissues were positive for EBV DNA. A 12-week course of chemotherapy given to two children resulted in rapid tumor regression. One of these children experienced meningeal relapse and died 16 months after diagnosis. The other child, who in addition received local irradiation of the affected eye and who underwent surgical removal of the involved ovaries, has been in continuous remission for 20 months. Conclusions. EBV-associated NHL may be seen more frequently in pediatric patients with HIV. Treatment protocols taking into account NHL propensity for the CNS in this age group need to be developed.

HIV-infected children have impaired antibody responses after exposure to certain antigens. Our aim was to determine whether HIV-infected children had lower varicella zoster virus (VZV) antibody levels compared with HIV-infected adults or healthy children and, if so, whether this was attributable to an impaired primary response, accelerated antibody loss, or failure to reactivate the memory VZV response.

ss Open Acce Oral presentation O222 Treatment interruption in children with chronic HIV-infection: the results of the paediatric European network for treatment of AIDS (PENTA) 11 trial DM Gibb*1, A Compagnucci2, H Green3, M Lallemant4, Y Saidi2, N NgoGiang-Huong4, C Taylor3, L Mofenson5, F Monpoux6, MIG Tome7, M Marczynska8, D Nadal9, U Wintergerst10, S Kanjavanit11, H Lyall12, C Giaquinto13 and J Moye4