Whole genome-based circulating tumor DNA analysis as an ultrasensitive biomarker in pediatric sarcomas.

Abstract

10053 Background: Disease monitoring in pediatric sarcomas relies on invasive surgical biopsies and radiologic imaging, which often trail disease activity. Circulating tumor DNA (ctDNA) is an attractive potential biomarker, but studies in pediatric sarcomas have reported low detection rates by tracking copy number alterations or recurrent genetic alterations. We aim to establish a personalized ctDNA pipeline in pediatric sarcomas based on identification of tumor-specific single nucleotide variants (SNVs) from whole genome sequencing (WGS) to enhance ctDNA detection. Methods: Patients < 25 years old with new or relapsed sarcoma treated at our institution are eligible to enroll in our study, which is ongoing. Plasma is collected at imaging response evaluations. Matched tumor and germline DNA are collected at baseline and undergo paired WGS to identify tumor-specific SNVs, and personalized hybrid capture panels are designed to track up to 5000 mutations per patient. Cell-free DNA from each time point is analyzed with CAncer Personalized Profiling by deep Sequencing (CAPP-Seq) using duplex sequencing to minimize the background error rate and maximize sensitivity. Results: This approach was piloted in 8 pediatric patients with osteosarcoma (5), Ewings sarcoma (2), and rhabdomyosarcoma (1). A median of 619 tumor-derived SNVs (range 160-4937) were tracked per patient. ctDNA was detected in all patients at baseline with mean allele fractions ranging from 0.000937-27.1. Table 1 summarizes characteristics, number of SNVs tracked, and mean allele fraction for each patient. Seven patients had longitudinal ctDNA samples available for analysis. In all patients, ctDNA broadly correlated with radiologic response with ctDNA becoming undetectable in patients with an imaging response and ctDNA remaining detectable in patients who experienced progression. One patient achieved initial remission with undetectable ctDNA and relapsed with ctDNA re-emergence. Conclusions: Personalized whole genome-based ctDNA analysis may improve ctDNA detection in pediatric sarcomas. In this pilot, ctDNA was detected at baseline in 8/8 patients, and ctDNA levels correlated with treatment response on imaging. A personalized approach for ctDNA detection in pediatric sarcomas has the potential to aid in disease monitoring and treatment selection. Patient characteristics and baseline ctDNA allele fraction. Disease New (N) v Relapsed/ Refractory (R) Localized (L) v Metastatic (M) Sex (M, F) Age (years) SNVs (n) Baseline ctDNA Mean Allele fraction Osteosarcoma N L M 17 692 0.25 Osteosarcoma N L M 14 301 4.29 Osteosarcoma R M M 20 4937 0.00094 Osteosarcoma R M F 11 1684 27.10 Osteosarcoma R M M 12 546 2.70 Ewings sarcoma N L M 5 160 14.11 Ewings sarcoma N L M 6 198 0.78 Rhabdomyosarcoma R M F 19 935 2.79 Each row represents 1 patient with disease and ctDNA detection characteristics displayed.