Loss of a FYN-regulated differentiation and growth arrest pathway in advanced stage neuroblastoma.

Tumor stage, age of patient, and amplification of MYCN predict disease outcome in neuroblastoma. To gain insight into the underlying molecular pathways, we have obtained expression profiles from 94 primary neuroblastoma specimens. Advanced tumor stages show a characteristic expression profile that includes downregulation of multiple genes involved in signal transduction through Fyn and the actin cytoskeleton. High expression of Fyn and high Fyn kinase activity are restricted to low-stage tumors. In culture, expression of active Fyn kinase induces differentiation and growth arrest of neuroblastoma cells. Expression of Fyn predicts long-term survival independently of MYCN amplification. Amplification of MYCN correlates with deregulation of a distinct set of genes, many of which are target genes of Myc. Our data demonstrate a causal role for Fyn kinase in the genesis of neuroblastoma.

The coamplification pattern of the MYCN amplicon is an invariable attribute of most MYCN-amplified human neuroblastomas.

PURPOSE: Fifteen percent to 20% of human neuroblastomas show amplification of the MYCN oncogene physiologically located at chromosome 2p24-25, indicating an aggressive subtype of human neuroblastoma with a poor clinical outcome. Recent findings revealed that the structure of the amplicon differs interindividually and that coamplification of genes in telomeric proximity to MYCN might play a relevant role in neuroblastoma development and response to treatment, respectively. We now asked if the amplicon structure is an invariable attribute of an individual tumor or if the coamplification pattern could change during progress or in case of recurrent disease. EXPERIMENTAL DESIGN: We used a previously described multiplex PCR approach to analyze the coamplification status of MYCN-amplified human neuroblastomas (n = 33) in tumor tissue at the time of initial diagnosis and in consecutive tissue specimens at later time points after initial treatment or from relapsing disease. The MYCN copy number per haploid genome (Mcn/hg) in these specimens was determined in a separate duplex PCR. RESULTS: In 32 of the 33 investigated tumors, the amplicon structure showed no changes after initial chemotherapy and in recurrent disease. Mcn/hg showed a decrease after initial treatment (n = 23), whereas we found a significant increase in recurrent disease (n = 10). CONCLUSION: Our data indicate that the initial determined structure of the 2p24-25 amplicon is a consistent attribute in the great majority of the individual MYCN-amplified neuroblastomas and shows no plasticity during or after chemotherapy. Observed changes in the Mcn/hg over the course of disease are in line with preexisting cell culture findings.

Coamplification of DDX1 correlates with an improved survival probability in children with MYCN-amplified human neuroblastoma.

PURPOSE: Amplification of the MYCN oncogene at chromosome 2p24-25 identifies an aggressive subtype of human neuroblastoma with a poor clinical outcome. Differences in amplicon structure and coamplification of genes telomeric and centromeric to the MYCN oncogene have previously been described. A relevant role of gene coamplification for neuroblastoma pathogenesis remains elusive. PATIENTS AND METHODS: We analyzed 98 primary neuroblastoma tumors with MYCN amplification for coamplification of seven additional genes at chromosome 2p24-25 (DDX1, NAG, NSE1, LPIN1, EST-AA581763, SMC6, and SDC1). Two semiquantitative multiplex polymerase chain reactions were used to obtain the amplification status of the target genes in relation to a reference gene on chromosome 2q (Inhibin-beta-b). Furthermore, mRNA expression pattern of coamplified genes in a subset of tumors was analyzed. RESULTS: Our results show that the frequency of gene coamplification on 2p24-25 in neuroblastoma is correlated directly to the physical distance to MYCN. Coamplification is correlated to an upregulated gene expression for DDX1 and NAG. Coamplification of the DDX1 gene within 400kb telomeric to MYCN identifies a subgroup of advanced stage neuroblastoma tumors with a more favorable outcome (P =.027, log-rank test). A high expression level of DDX1 is associated with a trend towards a better survival probability (P =.058, log-rank test). CONCLUSION: Our results indicate that DDX1 coamplification correlates with a better prognosis and improved patient survival in MYCN-amplified neurobastoma.

Coexpression of insulin receptor-related receptor and insulin-like growth factor 1 receptor correlates with enhanced apoptosis and dedifferentiation in human neuroblastomas.

PURPOSE: We compared the expression of the insulin receptor-related receptor (IRR) in primary human neuroblastomas with other biological and clinical parameters and the impact of its expression on prognostic outcome. EXPERIMENTAL DESIGN: We studied 49 neuroblastomas of different clinical stages and histological subtypes for (a) IRR, insulin-like growth factor 1 receptor (IGF-1R), TrkA, p75 neurotrophin receptor, and MYCN mRNA expression by reverse transcription-PCR; (b) MYCN gene amplification by Southern blot analyses; (c) cyclin A protein expression by Western blot analyses indicating proliferation rate; and (d) apoptotic index (AI) by terminal deoxynucleotidyl transferase (Tdt)-mediated dUTP nick end-labeling assay. RESULTS: IRR mRNA expression was found in 25 (51%) neuroblastomas and correlated with stages 1, 2, 3, and 4S disease and with age

Detection of human tumor cells by amplicon fusion site polymerase chain reaction (AFS-PCR).

Reliable diagnostic strategies for individuals with cancer demand practical methods for highly sensitive and specific detection of tumor cells. Amplification of genomic regions that include putative oncogenes is common in tumor cells of various types. Genomic array platforms offer the opportunity to identify and precisely map amplified genomic regions (ampGRs). The stable existence of these tumor cell­specific genomic aberrations during and after therapy, in theory, make ampGRs optimal targets for cancer diagnostics. In this study, we mapped ampGRs around the proto-oncogene MYCN of human neuroblastomas using a high-resolution tiling array (HR-TA). Based on the HR-TA data, we were able to precisely describe the telomeric and centromeric borders of the ampGRs and deduce virtual fusion sites of the joined ampGRs (amplicon fusion sites [AFSs]). These AFSs served as blueprints for the subsequent design of AFS bridging PCR assays (AFS-PCRs). Strikingly, these assays were absolutely tumor cell specific and capable of detecting 1 tumor cell in 1 × 10(6) to 8 × 10(6) control cells. We successfully proved the in vivo practicability of AFS-PCR by detecting and quantifying the specific AFS DNA of human MYCN-amplified neuroblastomas in the patients' corresponding peripheral blood and bone marrow samples. Thus, we believe AFS-PCR could become a powerful and nevertheless feasible personalized diagnostic tool applicable to a large number of cancer patients, including children with MYCN-amplified neuroblastomas.

OECD Health Care Quality Indicator Project. The expert panel on primary care prevention and health promotion.

PURPOSE: This article describes a project undertaken as part of the Organization for Economic Co-operation and Development (OECD)'s Healthcare Quality Indicator (HCQI) Project, which aimed to develop a set of quality indicators representing the domains of primary care, prevention and health promotion, and which could be used to assess the performance of primary care systems. METHODS: Existing quality indicators from around the world were mapped to an organizing framework which related primary care, prevention, and health promotion. The indicators were judged against the US Institute of Medicine's assessment criteria of importance and scientific soundness, and only those which met these criteria and were likely to be feasible were included. An initial large set of indicators was reduced by the primary care expert panel using a modified Delphi process. RESULTS: A set of 27 indicators was produced. Six of them were related to health promotion, covering health-related behaviours that are typically targeted by health education and outreach campaigns, 13 to preventive care with a focus on prenatal care and immunizations and eight to primary clinical care mainly addressing activities related to risk reduction. The indicators selected placed a strong emphasis on the public health aspects of primary care. CONCLUSIONS: This project represents an important but preliminary step towards a set of measures to evaluate and compare primary care quality. Further work is required to assess the operational feasibility of the indicators and the validity of any benchmarking data drawn from international comparisons. A conceptual framework needs to be developed that comprehensively captures the complex construct of primary care as a basis for the selection of additional indicators.