Background: According to current clinical guidelines mutational analysis for KRAS and NRAS is recommended prior to EGFR-directed therapy of colorectal cancer (CRC) in the metastatic setting. Therefore, reliable, fast, sensitive and cost-effective methods for routine tissue based molecular diagnostics are required that allow the assessment of the CRC mutational status in a high throughput fashion.
Methods: We have developed a custom designed assay for routine mass-spectrometric (MS) (MassARRAY, Agena Bioscience) analysis to test the presence/absence of 18 KRAS, 14 NRAS and 4 BRAF mutations. We have applied this assay to 93 samples from patients with CRC and have compared the results with Sanger sequencing and a chip hybridization assay (KRAS LCD-array Kit, Chipron). In cases with discordant results, next-generation sequencing (NGS) was performed.
Results: MS detected a KRAS mutation in 46/93 (49%), a NRAS mutation in 2/93 (2%) and a BRAF mutation in 1/93 (1%) of the cases. MS results were in agreement with results obtained by combination of the two other methods in 92 (99%) of 93 cases. In 1/93 (1%) of the cases a G12V mutation has been detected by Sanger sequencing and MS, but not by the chip assay. In this case, NGS has confirmed the G12V mutation in KRAS.
Conclusions: Mutational analysis by MS is a reliable method for routine diagnostic use, which can be easily extended for testing of additional mutations.