Testing and validation of pre-analytical parameters to make cell-free tumor DNA detection a reliable minimal-invasive diagnostic tool

Introduction:

Neuroblastoma (NB) is the most common extra-cranial solid tumor of infancy and responsible for 15% of cancer-related pediatric deaths. This tumor type is characterized by a remarkable biological and clinical heterogeneity, making diagnostic approaches and therapeutic decisions challenging. Circulating tumor DNA (ctDNA) can possibly serve as an additional, minimal-invasive source for tumor DNA diagnostics and disease monitoring.

Aim:

Harmonize pre-analytical and analytical procedures and validate different technique(s) of sample handling and DNA-isolation of cell-free DNA (cfDNA) to provide clear guidelines and standards (generate SOPs) to implement liquid biopsy in the clinical routine.

Methods/Results:

The effect of sample conditions, such as selection of the optimal blood collection tube, sample stability over time, and influence of storage-temperature on the quality of the isolated cfDNA, was investigated. Moreover, cfDNA from blood and BM plasma from NB patients was analyzed for the proportion of ctDNA in the different size-fractions of the sample (166/332/498 bp fraction vs. high molecular weight cfDNA). These data allow drawing conclusions concerning the most reliable pre-analytical steps.