Next Generation Sequencing of Tumor and Matched Plasma Samples: Identification of Somatic Variants in ctDNA From Ovarian Cancer Patients

Abstract

Genetic testing to detect somatic alterations is usually performed on formalin-fixed paraffin-embedded tumor samples. However, tumor molecular profiling through ctDNA analysis may be particularly interesting with the emergence of targeted therapies for ovarian cancer (OC), mainly when tumor is not available and biopsy is not viable, also allowing representation of multiple neoplastic subclones. Using a custom panel of 27 genes, next-generation sequencing (NGS) was performed on tumor and matched plasma samples from 96 OC patients, which were combined in two groups (treatment naive and post-treatment). Overall, at least one somatic variant present in the tumor sample was also detected in the matched plasma sample in 35.6% of the patients, a percentage that increased to 69.6% of the treatment naive patients and 83.3% of those with stage IV disease, showing the potential of ctDNA analysis as an alternative to identify somatic variants in these patients, namely those that have predictive value for targeted therapy. In fact, of the two treatment-naive patients with somatic BRCA1 variants identified in tumor samples, in one of them we detected in ctDNA a BRCA1 somatic variant that was present in the tumor with a VAF of 53%, but not in the one that had a VAF of 5.4%. We also showed that ctDNA analysis has a complementary role to molecular unraveling of inter- and intra-tumor heterogeneity, as exemplified by one patient diagnosed with bilateral OC in which different somatic variants from both tumors were detected in ctDNA. Interestingly, as these bilateral tumors shared a rare combination of two of the three variants identified in ctDNA, we could conclude that these morphologically different tumors were clonally related and not synchronous independent neoplasias. Moreover, in the post-treatment group of patients with plasma samples collected after surgery, those with detectable somatic variants had poor prognosis when compared with patients with no detectable somatic variants, highlighting the potential of ctDNA analysis to identify patients at higher risk of recurrence. Concluding, this study demonstrated that somatic variants can be detected in plasma samples of a significant proportion of OC patients, supporting the use of NGS-based ctDNA testing for noninvasive tumor molecular profiling and to stratify patients according to prognosis.

Whole-genome/exome analysis of circulating tumor DNA and comparison to tumor genomics from patients with heavily pre-treated ovarian cancer: subset analysis of the PERMED-01 trial

“…To our knowledge, we are the first to describe this technology in this setting. We identified at least one tumor mutation in 21/24 patients (88%), close to what we can expect with panel-based NGS in these tumors with low mutation burden (42), and to what has been described in pre-treated metastatic cancers (43). As already shown, plasma VAF was lower than tumor VAF, reflecting a lower tumor fraction in the bloodstream (44,45).…”

Liquid biopsy in ovarian cancer in China and the world: current status and future perspectives

“…A NGS analysis of tumor and plasma samples from 96 patients with OC showed that tumor somatic variants in at least one of 27 cancer-related genes were present in the serum of 83.3% of patients with stage IV OC; however, the sensitivity of this test was lower for early-stage disease ( 32 ). Mutations in TP53 and BRCA1 in ctDNA or cfDNA have also been shown to have diagnostic utility in OC ( 31 ), and analysis of Chinese patients has shown that mutation frequency in ctDNA using hybrid capture-based genomic profiling were generally similar between tissue biopsies and LB ( 50 ).…”

Potential clinical utility of liquid biopsies in ovarian cancer

“…More recently, Barbosa et. al used NGS and a custom panel of 27 genes to analyze tumour and matched plasma samples from 96 ovarian cancer patients and detected tumour somatic variants in 75% of patients with stage I disease [ 180 ]. Cohen et al analyzed circulating protein biomarkers and genetic alterations in cfDNA using a commercial blood test called CancerSEEK [ 177 ].…”