BACKGROUND: The purpose of this study was to evaluate whether levels of neuroblastoma mRNAs in bone marrow and peripheral blood from stage M infants (≤12 months of age at diagnosis, MYCN amplified) and toddlers (between 12 and 18 months, any MYCN status) predict event-free survival (EFS). METHODS: Bone marrow aspirates and peripheral blood samples from 97 infants/toddlers enrolled in the European High-Risk Neuroblastoma trial were collected at diagnosis in PAXgene™ blood RNA tubes. Samples were analyzed by reverse transcription quantitative polymerase chain reaction according to standardized procedures. RESULTS: Bone marrow tyrosine hydroxylase (TH) or paired-like homeobox 2b (PHOX2B) levels in the highest tertile were associated with worse EFS; hazard ratios, adjusted for age and MYCN status, were 1.5 and 1.8 respectively. Expression of both TH and PHOX2B in the highest tertile predicted worse outcome (p = 0.015), and identified 20 (23%) infants/toddlers with 5-year EFS of 20% (95%CI: 4%-44%). Prognostic significance was maintained after adjusting for over-fitting bias (p = 0.038), age and MYCN status. In peripheral blood, PHOX2B levels in the highest tertile predicted a two-fold increased risk of an event (p = 0.032), and identified 23 (34%) infants/toddlers with 5-year EFS of 29% (95%CI: 12%-48%). Time-dependent receiver operating characteristic analysis confirmed the prognostic value of combined TH and PHOX2B in bone marrow and of PHOX2B in peripheral blood during the first year of follow-up. CONCLUSIONS: High levels of bone marrow TH and PHOX2B and of peripheral blood PHOX2B at diagnosis allow early identification of a group of high-risk infant and toddlers with neuroblastoma who may be candidates for alternative treatments. Integration with additional biomarkers, as well as validation in additional international trials is warranted.
- MeSH
- doba přežití bez progrese choroby MeSH
- homeodoménové proteiny analýza biosyntéza MeSH
- Kaplanův-Meierův odhad MeSH
- kojenec MeSH
- lidé MeSH
- messenger RNA analýza MeSH
- nádorové biomarkery analýza MeSH
- neuroblastom metabolismus mortalita MeSH
- novorozenec MeSH
- plocha pod křivkou MeSH
- prognóza MeSH
- proporcionální rizikové modely MeSH
- ROC křivka MeSH
- senzitivita a specificita MeSH
- transkripční faktory analýza biosyntéza MeSH
- tyrosin-3-monooxygenasa analýza biosyntéza MeSH
- Check Tag
- kojenec MeSH
- lidé MeSH
- mužské pohlaví MeSH
- novorozenec MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- klinické zkoušky MeSH
- práce podpořená grantem MeSH
An assay of L-tyrosine (Tyr) hydroxylating activity operating in lincomycin biosynthesis is described. The assay development consisted of HPLC procedure development, assessing the effect of reaction mixture components on non-enzymatic Dopa and Tyr oxidation, and sample stability evaluation. The HPLC procedure with isocratic elution and fluorescence detection was developed and validated. The method showed a wide linear range of Dopa determination of 0.125-25 micromol/L with lower limit of quantification (LLOQ) of 0.125 micromol/L, RSD of 7.2% and accuracy of 101.7%. The studied linear range of Tyr was 15.625 mmol/L to 500 mmol/L with LLOQ of 15.625 mmol/L, RSD of 1.1%, and accuracy of 98.1%. Recoveries for Dopa and Tyr were 100.66 +/- 0.89% and 94.76 +/- 0.94%, respectively. The inter- and intra-day accuracies and precisions were all within 10%. Samples of the reaction mixture were stable for at least 24 h at room temperature (RT) and 28 days at -20 degrees C. The method was tested for the enzyme activity monitoring in purified as well as crude preparations and enabled micro preparation of the enzyme product during confirmation of its identity. The influence of pH and ascorbic acid content in reaction mixture was studied with respect to non-enzymatic Tyr oxidation.
- MeSH
- aktivace enzymů MeSH
- biotest MeSH
- časové faktory MeSH
- dopaminové látky analýza chemie MeSH
- financování organizované MeSH
- koncentrace vodíkových iontů MeSH
- levodopa analýza chemie MeSH
- oxidace-redukce MeSH
- reprodukovatelnost výsledků MeSH
- senzitivita a specificita MeSH
- stabilita léku MeSH
- teplota MeSH
- tyrosin-3-monooxygenasa analýza metabolismus MeSH
- tyrosin analýza MeSH
- vysokoúčinná kapalinová chromatografie metody MeSH
- MeSH
- dopamin-beta-hydroxylasa analýza metabolismus MeSH
- finanční podpora výzkumu jako téma MeSH
- hypoxie buňky MeSH
- hypoxie patologie MeSH
- krysa rodu rattus MeSH
- messenger RNA analýza MeSH
- mezencefalon MeSH
- polymerázová řetězová reakce s reverzní transkripcí MeSH
- tyrosin-3-monooxygenasa analýza metabolismus MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- zvířata MeSH
- Publikační typ
- techniky in vitro MeSH
- MeSH
- hypertenze patofyziologie MeSH
- krysa rodu rattus MeSH
- noradrenalin analýza MeSH
- synthasa oxidu dusnatého analýza MeSH
- tyrosin-3-monooxygenasa analýza MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- zvířata MeSH
- Publikační typ
- kongresy MeSH
- MeSH
- deoxyglukosa farmakologie MeSH
- dřeň nadledvin enzymologie MeSH
- hypoglykemie chemicky indukované metabolismus MeSH
- inzulin farmakologie MeSH
- krysa rodu rattus MeSH
- locus coeruleus enzymologie MeSH
- messenger RNA analýza fyziologie MeSH
- tyrosin-3-monooxygenasa analýza biosyntéza genetika MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- zvířata MeSH
- Publikační typ
- srovnávací studie MeSH
- MeSH
- cholin-O-acetyltransferasa MeSH
- krysa rodu rattus MeSH
- mezencefalon anatomie a histologie cytologie enzymologie MeSH
- pons anatomie a histologie cytologie enzymologie MeSH
- substance P analýza MeSH
- tegmentum mesencephali anatomie a histologie cytologie enzymologie MeSH
- tyrosin-3-monooxygenasa analýza MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- zvířata MeSH
