Noninvasive Detection of ctDNA Reveals Intratumor Heterogeneity and Is Associated with Tumor Burden in Gastrointestinal Stromal Tumor

Abstract

Molecular analysis of circulating tumor DNA (ctDNA) has a large potential for clinical application by capturing tumor-specific aberrations through noninvasive sampling. In gastrointestinal stromal tumor (GIST), analysis of and mutations is important for therapeutic decisions, but the invasiveness of traditional biopsies limits the possibilities for repeated sampling. Using targeted next-generation sequencing, we have analyzed circulating cell-free DNA from 50 GIST patients. Tumor-specific mutations were detected in 16 of 44 plasma samples (36%) from treatment-naïve patients and in three of six (50%) patients treated with tyrosine kinase inhibitors. A significant association between detection of ctDNA and the modified National Institutes of Health risk classification was found. All patients with metastatic disease had detectable ctDNA, and tumor burden was the most important detection determinant. Median tumor size was 13.4 cm for patients with detectable mutation in plasma compared with 4.4 cm in patients without detectable mutation ( = 0.006). ctDNA analysis of a patient with disease progression on imatinib revealed that multiple resistance mutations were synchronously present, and detailed analysis of tumor tissue showed that these were spatially distributed in the primary tumor. Plasma samples taken throughout the course of treatment demonstrated that clonal evolution can be monitored over time. In conclusion, we have shown that detection of GIST-specific mutations in plasma is particularly feasible for patients with high tumor burden. In such cases, we have demonstrated that mutational analysis by use of liquid biopsies can capture the molecular heterogeneity of the whole tumor, and may guide treatment decisions during progression. .

Circulating cKIT and PDGFRA DNA indicates disease activity in Gastrointestinal Stromal Tumor (GIST)

“…Recently, Namlos et al . showed that detection of ctDNA in plasma of GIST patients using NGS correlated with high tumor burden but at a relevantly low ctDNA detection rate (19 of 50 patients, i.e., 38%) . Similarly, Xu et al .…”

“…22,24,26,27 It has been shown that in GIST the clonal genetic composition changes over time and secondary mutations causing treatment resistance can be detected in tissue 28,29 and plasma samples of patients with progression. 17,30 Accordingly, in Patient #5, L-PCR detected a cKIT exon 17 D820G mutation known to mediate imatinib resistance in ctDNA, 28 in addition to the preexistent cKIT exon 11 mutation, reflecting selection of a treatment-resistant disease clone contributing to subsequent progression. Targeted NGS of ctDNA offers the advantage of covering hundreds to thousands of amplicons in one assay read in an unbiased fashion.…”

“…Analysis of ctDNA has detected cancer mutations that were missed in corresponding tissue specimen and thus can provide information about clonal and subclonal mutations . It has been shown that in GIST the clonal genetic composition changes over time and secondary mutations causing treatment resistance can be detected in tissue and plasma samples of patients with progression . Accordingly, in Patient #5, L‐PCR detected a cKIT exon 17 D820G mutation known to mediate imatinib resistance in ctDNA, in addition to the preexistent cKIT exon 11 mutation, reflecting selection of a treatment‐resistant disease clone contributing to subsequent progression.…”

“…Targeted NGS of ctDNA offers the advantage of covering hundreds to thousands of amplicons in one assay read in an unbiased fashion . NGS was shown to successfully detect ctDNA in plasma, and in single cases to capture clonal heterogeneity in GIST patients . However, GIST liquid biopsy studies that used NGS had either low patient numbers or were limited to localized disease, and data showing a long‐term follow up of individual GIST patients are lacking.…”

“…31,32 NGS was shown to successfully detect ctDNA in plasma, 30,[33][34][35][36] and in single cases to capture clonal heterogeneity in GIST patients. 30,34,35 However, GIST liquid biopsy studies that used NGS had either low patient numbers 34,35 or were limited to localized disease, 36 and data showing a long-term follow up of individual GIST patients are lacking. Recently, Namlos et al showed that detection of ctDNA in plasma of GIST patients using NGS correlated with high tumor burden but at a relevantly low ctDNA detection rate (19 of 50 patients, i.e., 38%).…”