Third line systemic treatment for patients with RAS wild-type metastatic colorectal cancer (mCRC) includes anti-EGFR monoclonal antibodies such as cetuximab. Here, we examined cell-free DNA (cfDNA) to gain insight into mechanisms underlying primary and acquired resistance in patients with mCRC receiving cetuximab.
34 patients with RAS wild-type mCRC (KRAS and NRAS exon 2-4) received biweekly cetuximab monotherapy (500mg/m2). cfDNA was isolated from plasma obtained at baseline, after 2 weeks of treatment and at disease progression (PD). ± 20 ng DNA was used for targeted next generation sequencing using the Oncomine™ Colon cfDNA Assay (14 genes, 242 hotspots). Mutation analysis of tumor tissue was performed according standard of care, at least including KRAS and NRAS. Outcome was defined as clinical benefit (CB; PD > 8 weeks, n = 21) versus no CB (NCB; PD ≤ 8 weeks, n = 13).
Baseline cfDNA concentration correlated with the sum of diameters on CT (p = 0.043) and metabolically active tumor volume on [18F]FDG PET (p < 0.001). In 6/13 (46%) of patients with NCB, mutations in KRAS (n = 3) and BRAF (n = 3) were detected in baseline cfDNA. Two KRAS mutations were detected cfDNA, but not in tissue. All BRAF mutations in cfDNA were present in tissue, one BRAF mutation in tissue was not detected in cfDNA. In one patient (5%) with CB a polyclonal KRAS mutation was detected in cfDNA, which was not found in tumor tissue. In 9 patients with CB, cfDNA concentrations were measured and decreased from a median of 45 ng/mL plasma (range 13 – 784 ng/mL) at baseline to 19 ng/mL after 2 weeks of treatment (range 9 – 42 ng/mL) (p = 0.008). In patients with CB an enrichment of mutations in genes associated with resistance (KRAS, NRAS and BRAF) was found in 12/17 (70%) at PD compared to baseline. Moreover, in 8/17 (47%) of these patients EGFR mutations in codons coding for the epitope binding site of cetuximab emerged and in 9/17 patients (53%) multiple mutations in the same gene occurred suggesting the presence of multiple subclones.
A subset of mCRC patients with NCB could be identified based on baseline cfDNA mutations in genes associated with cetuximab resistance. By using cfDNA we can optimize patient selection for cetuximab therapy and elucidate mechanisms of resistance.
Clinical trial identification
Legal entity responsible for the study
Erasmus University Medical Center.
Dutch Cancer Society.
L. Angus: Advisory board: Merck B.V. All other authors have declared no conflicts of interest.