Tumor fraction-guided cell-free DNA profiling in metastatic solid tumor patients

Abstract

Background Cell-free DNA (cfDNA) profiling is increasingly used to guide cancer care, yet mutations are not always identified. The ability to detect somatic mutations in plasma depends on both assay sensitivity and the fraction of circulating DNA in plasma that is tumor-derived (i.e., cfDNA tumor fraction). We hypothesized that cfDNA tumor fraction could inform the interpretation of negative cfDNA results and guide the choice of subsequent assays of greater genomic breadth or depth. Methods Plasma samples collected from 118 metastatic cancer patients were analyzed with cf-IMPACT, a modified version of the FDA-authorized MSK-IMPACT tumor test that can detect genomic alterations in 410 cancer-associated genes. Shallow whole genome sequencing (sWGS) was also performed in the same samples to estimate cfDNA tumor fraction based on genome-wide copy number alterations using z-score statistics. Plasma samples with no somatic alterations detected by cf-IMPACT were triaged based on sWGS-estimated tumor fraction for analysis with either a less comprehensive but more sensitive assay (MSK-ACCESS) or broader whole exome sequencing (WES). Results cfDNA profiling using cf-IMPACT identified somatic mutations in 55/76 (72%) patients for whom MSK-IMPACT tumor profiling data were available. A significantly higher concordance of mutational profiles and tumor mutational burden (TMB) was observed between plasma and tumor profiling for plasma samples with a high tumor fraction (z-score≥5). In the 42 patients from whom tumor data was not available, cf-IMPACT identified mutations in 16/42 (38%). In total, cf-IMPACT analysis of plasma revealed mutations in 71/118 (60%) patients, with clinically actionable alterations identified in 30 (25%), including therapeutic targets of FDA-approved drugs. Of the 47 samples without alterations detected and low tumor fraction (z-score<5), 29 had sufficient material to be re-analyzed using a less comprehensive but more sensitive assay, MSK-ACCESS, which revealed somatic mutations in 14/29 (48%). Conversely, 5 patients without alterations detected by cf-IMPACT and with high tumor fraction (z-score≥5) were analyzed by WES, which identified mutational signatures and alterations in potential oncogenic drivers not covered by the cf-IMPACT panel. Overall, we identified mutations in 90/118 (76%) patients in the entire cohort using the three complementary plasma profiling approaches. Conclusions cfDNA tumor fraction can inform the interpretation of negative cfDNA results and guide the selection of subsequent sequencing platforms that are most likely to identify clinically-relevant genomic alterations.

From APC to the genetics of hereditary and familial colon cancer syndromes

“…Unfortunately, availability of tumor DNA is often a limiting factor for NGS-based studies. Liquid biopsies could be a reliable source of tumor DNA for such purposes, as significant numbers of clinically actionable variants have been present in cell-free DNA samples when tumor samples were either unavailable or had not yielded any results ( 192 ). Similar conclusions can be drawn from studies utilizing mutation-specific PCR ( 193 ) and methylation analysis ( 194 , 195 ) of cell-free DNA from patients with metastatic disease.…”

The utility of cfDNA in TGCT patient management: a systematic review

“…The study investigating mutations in cfDNA had 10 recruited patients with TGCT and 10 healthy controls. 32 Of the 10, NGS analysis of cfDNA and tumor genomic DNA has managed to identify mutations in six patients, while the remaining four have not had any mutations detected. Of the six, four patients had mutations present in the tumor, which were also detected in the cfDNA of three patients, and undetected in one.…”

“… 59 , 60 This would make most commercial cfDNA assays, which are highly focused, not well suited for TGCT, and more broader sequencing assays should be employed which remain cost-prohibitive for near-term clinical implementation. 32 , 49 This is only additionally complicated by ultra-deep sequencing picking up inconsequential mutations in other high turnover components (such as bone marrow) when needing to interpret rare alleles. 19 As research has shown, 32 while 6 of the 10 TGCTs have been detected by NGS, only 3 of them have had the same mutations identified in cfDNA and DNA from the solid tumor, 2 have had mutations detected only in cfDNA (which could be the before-mentioned inconsequential mutations), and 1 had tumor mutations not detected in cfDNA.…”

Dynamics of RAS/BRAF Mutations in cfDNA from Metastatic Colorectal Carcinoma Patients Treated with Polychemotherapy and Anti-EGFR Monoclonal Antibodies

“…However, we must acknowledge that the true positive fraction of cases could be underestimated because of the limited sensitivity of the tests that we used. In this respect, the estimation of tumor fraction in plasma samples might in the future allow for a better definition of this phenomenon [ 44 , 45 , 46 ]. Studies on larger patients’ cohorts will allow us to better define the prognostic value of acquired, likely sub-clonal RAS and BRAF variants, which could be different as compared with de novo clonal mutations of these genes.…”