Measurement of 3-O-methyldopa from dried plasma microsamples by high-resolution mass spectrometry: A tool for the diagnosis of patients with high-risk neuroblastoma

Background: Risk assessment at diagnosis is crucial for neuroblastoma (NB) in order to address patients at highrisk to the most timely and appropriate treatments. 3-O-methyldopa (3-OMD), a direct metabolite of L-Dopa, is a promising biomarker of NB at diagnosis able to stratify high-risk patients. Methods: We show the development and validation of a method for measuring 3-OMD from dried plasma samples (DPS) and plasma using liquid chromatography coupled with high resolution mass spectrometry (LC-HRMS) on a Thermo Fisher Scientific Orbitrap Exploris 120. Results: The method was accurate and reproducible in the range 7.8-4000 ng/mL, from small amounts (50 mL) of plasma and DPS (obtained starting from 30 mL plasma). 3-OMD concentrations measured in plasma and DPS were highly correlated (R = 0.99 95 %CI 0.993-0.996). Differences of 3-OMD levels across stages L1 and M and L1 and L2 (p-value < 0.05) were statistically significant. Receiving Operator Curve (ROC) analysis showed that 3OMD was able to discriminate patients at high-risk with high sensitivity and specificity both from plasma or DPS (AUC = 0.8295 %CI 0.71-0.94, P < 0.0001). Conclusions: 3-OMD is confirmed as an interesting biomarker of high-risk NB. The described method is an added value for further prospective studies involving multiple sites. The stability of 3-OMD in DPS allows for easy shipment and storage at room temperature.