Circulating tumour DNA sequencing to determine therapeutic response and identify tumour heterogeneity in patients with paediatric solid tumours

Abstract

Objective: Clinical diagnostic sequencing of circulating tumour DNA (ctDNA) is well advanced for adult patients, but application to paediatric cancer patients lags behind. Methods: To address this, we have developed a clinically relevant (67 gene) NGS capture panel and accompanying workflow that enables sensitive and reliable detection of lowfrequency genetic variants in cell-free DNA (cfDNA) from children with solid tumours. We combined gene panel sequencing with low pass whole-genome sequencing of the same library to inform on genome-wide copy number changes in the blood. Results: Analytical validity was evaluated using control materials, and the method was found to be highly sensitive (0.96 for SNVs and 0.97 for INDEL), specific (0.82 for SNVs and 0.978 for INDEL), repeatable (>0.93 [95% CI: 0.89e0.95]) and reproducible (>0.87 [95% CI: 0.87 e0.95]). Potential for clinical application was demonstrated in 39 childhood cancer patients with a spectrum of solid tumours in which the single nucleotide variants expected from tumour sequencing were detected in cfDNA in 94.4% (17/18) of cases with active extracranial disease. In 13 patients, where serial samples were available, we show a close correlation between events detected in cfDNA and treatment response, demonstrate that cfDNA analysis could be a useful tool to monitor disease progression, and show cfDNA sequencing has the potential to identify targetable variants that were not detected in tumour samples. Conclusions: This is the first pan-cancer DNA sequencing panel that we know to be optimised for cfDNA in children for blood-based molecular diagnostics in paediatric solid tumours.

Blood-Derived Liquid Biopsies Using Foundation One® Liquid CDx for Children and Adolescents with High-Risk Malignancies: A Monocentric Experience

“…There is currently very little pediatric data reporting the use of liquid biopsy for a wide panel of genetic alterations in pediatric oncology. Stankunaite et al reported a series of 39 children with solid tumors, for whom they were able to perform analysis of 67 genes [ 17 ]. Interestingly, molecular abnormalities were found in all patients with extra-cranial malignancies and not under treatment using LB.…”

“…Although these results are based a small subset of patients among the series we report here, and although LB and tumor sampling was not conducted during the course of the disease, this might illustrate the absence of pediatric enhancement of the Foundation One ® Liquid CDx test. As previously described, some genomic events enriched in pediatric cancer may not be captured by adult panels [ 17 ]. This represents a risk of underestimating and missing an opportunity to treat an actionable target, although according to OncoKB classification, Class 1, 2, and 3 alterations were missed using the LB.…”

“…We therefore advocate for LB to be performed in the context of high-risk malignancy and further explored, as it may represent a genuine alternative to tissue biopsy. Additionally, as suggested by Stankunaite et al and previous authors, Ct DNA in LB could also be used to monitor response to treatment [ 16 , 17 , 23 ].…”

“…Experience and knowledge in LB are increasing rapidly in the adult setting, with evidence of its clinical impact [ 12 , 13 , 14 ]. In children, feasibility of LB has been described in research settings with successful molecular profiling [ 15 , 16 , 17 , 18 , 19 , 20 ]. However, data is still quite limited, as well as reports on the real-world feasibility and usefulness of this tool.…”

Circulating Tumor DNA in Pediatric Cancer

“…There is strong evidence for the importance of profiling tumors at relapse, including RAS-MAPK pathway mutations known to occur in relapsed/refractory disease ( Eleveld et al, 2015 ). Several pediatric studies have achieved this without prior knowledge of tumor-derived biomarkers in neuroblastoma ( Cimmino et al, 2020 ) and more recently, in a panel of pediatric solid tumors ( Stankunaite et al, 2021 ). These studies have shown the value of ctDNA for identifying potential mutations associated with drug resistance.…”