Abstract
BackgroundLiquid biopsies do not reflect the complete mutation profile of the tumor but have the potential to identify actionable mutations when tumor biopsies are not available as well as variants with low allele frequency. Most retrospective studies conducted in small cohorts of pediatric cancers have illustrated that the technology yield substantial potential in neuroblastoma.AimThe molecular landscape of neuroblastoma harbors potentially actionable genomic alterations. We aimed to study the utility of liquid biopsy to characterize the mutational landscape of primary neuroblastoma using a custom gene panel for ctDNA targeted sequencing.MethodsTargeted next-generation sequencing (NGS) was performed on ctDNA of 11 patients with primary neuroblastoma stage 4. To avoid the detection of false variants, we used UMIs (unique molecular identifiers) for the library construction, increased the sequencing depth and developed ad hoc bioinformatic analyses including the hard filtering of the variant calls.ResultsWe identified 9/11 (81.8%) patients who carry at least one pathogenic variation. The most frequently mutated genes were KMT2C (five cases), NOTCH1/2 (four cases), CREBBP (three cases), ARID1A/B (three cases), ALK (two cases), FGFR1 (two cases), FAT4 (two cases) and CARD11 (two cases).ConclusionsWe developed a targeted NGS approach to identify tumor-specific alterations in ctDNA of neuroblastoma patients. Our results show the reliability of our approach to generate genomic information which can be integrated with clinical and pathological data at diagnosis.
Highlights
Circulating tumor DNA, a subfraction of cell-free DNA, is fragmented genomic DNA poured in the blood flow and other biological fluids as the result of apoptosis and necrosis of tumor cells [1, 2]
We report the development of a targeted Generation Sequencing (NGS) gene panel for Circulating tumor DNA (ctDNA) sequencing that is tailored to the genetic landscape of neuroblastoma
Nine patients with Stage 4 and two with Stage 2 neuroblastoma were recruited as described in Methods (Supplementary Table 1)
Summary
Circulating tumor DNA (ctDNA), a subfraction of cell-free DNA (cfDNA), is fragmented genomic DNA poured in the blood flow and other biological fluids as the result of apoptosis and necrosis of tumor cells [1, 2]. The isolation and sequencing of ctDNA from biological fluids is called Liquid Biopsy (LB). LB is a lowcost and safe non-surgical procedure to access tumor’s genetic information. It is a valid alternative for tumors that are not easy to tissue biopsy and represents a complementary tool for more accurate diagnoses. Liquid biopsies do not reflect the complete mutation profile of the tumor but have the potential to identify actionable mutations when tumor biopsies are not available as well as variants with low allele frequency. Most retrospective studies conducted in small cohorts of pediatric cancers have illustrated that the technology yield substantial potential in neuroblastoma
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