Abstract 1502: A rapid and fully automated multiplex assay for KRAS-BRAF mutations with high mutation sensitivity using novel selective amplification and detection technologies

• DOI: 10.1158/1538-7445.am2014-1502
• Publication Date: 2015-05-01T21:52:08Z
• Authors: Ina Vandenbroucke; Katrien Vermeiren; Elisa Mokany; Lit Yeen Tan; Nicole Lima; Samantha Walker; Geneviève Vanderc...; Bart Claes; Inky D. Baere; Pascale Holemans; Evelien Rondelez; Alison Todd; Geert Maertens; Erwin Sablon
• Keywords: Multiplex; Primer (cosmetics); Molecular diagnostics

Abstract Introduction The MAPK/ERK pathway is a complex signaling cascade involved in many cancer types. KRAS and BRAF gene mutations are present number of cancers, including colon, lung pancreas, identification these genes great importance clinical diagnostics. Moreover, there growing demand for performing multiple tests simultaneously on single sample an increased need to provide answers oncologists short timeframe. Methods IdyllaTM fully integrated automated molecular diagnostics platform (1) that combines speed ease use with high sensitivity multiplexing capabilities. it overcomes the current time-consuming step processing formalin-fixed paraffin-embedded tissue (FFPE) samples. After insertion FFPE slice into cartridge, complete process preparation, real-time PCR reporting takes less than 2 hours. We here KRAS-BRAF prototype assay allows sensitive detection 13 5 one assay. discriminates individual at codons 12, 61 codon 600 using novel “Primer Assisted Sequence Switching” (PASS) primers along “Multi-component Nucleic Acid enzyme” (MNAzyme) detection. These technologies confer advantages multiplex mutation analysis; PASS selectively amplify target sequences interest resulting enhanced specificity between wild type (WT) mutant, mutants, MNAzymes allow efficient discrimination simultaneously. Results Several performance characteristics were examined: specificity, cross-reactivity, Specificity mutant versus WT as well cross-reactivity each was evaluated. demonstrated excellent all targets, delta Cq values >7 mutants >12 WT. Sensitivity assessed cell lines embedded containing defined ratios dilutions background. results indicated sensitivities <1% allele. evaluated set colon melanoma samples characterized by Sequenom MassARRAY, Illumina MiSeq, or Biocartis BRAF-only which >96 % concordance. Conclusion new extended capabilities sensitivity, use, turnaround time analysis For research only Citation Format: Ina Vandenbroucke, Katrien Vermeiren, Elisa Mokany, Lit Yeen Tan, Nicole Lima, Samantha Walker, Geneviève Vandercruyssen, Bart Claes, Inky De Baere, Pascale Holemans, Evelien Rondelez, Alison Todd, Geert Maertens, Erwin Sablon. A rapid selective amplification technologies. [abstract]. In: Proceedings 105th Annual Meeting American Association Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Res 2014;74(19 Suppl):Abstract nr 1502. doi:10.1158/1538-7445.AM2014-1502