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High resolution melting analysis of KRAS, BRAF and PIK3CA in KRASexon 2 wild-type metastatic colorectal cancer

2013; BioMed Central; Volume: 13; Issue: 1 Linguagem: Inglês

10.1186/1471-2407-13-169

1471-2407

Joana G. Guedes, Isabel Veiga, Patrícia Rocha, Pedro Pinto, Carla Pinto, Manuela Pinheiro, Ana Peixoto, Maria Fragoso, A. Raimundo, P. Ferreira, Manuela Machado, Nuno Sousa, Paula Lopes, António Araújo, Joana Martins‐Macedo, Jorge Lino Alves, C. Coutinho, Rui Henrique, Lúcio Lara Santos, Manuel R. Teixeira,

Cancer Genomics and Diagnostics

Abstract Background KRAS is an EGFR effector in the RAS/RAF/ERK cascade that is mutated in about 40% of metastatic colorectal cancer (mCRC). Activating mutations in codons 12 and 13 of the KRAS gene are the only established negative predictors of response to anti-EGFR therapy and patients whose tumors harbor such mutations are not candidates for therapy. However, 40 to 60% of wild-type cases do not respond to anti-EGFR therapy, suggesting the involvement of other genes that act downstream of EGFR in the RAS-RAF-MAPK and PI3K-AKT pathways or activating KRAS mutations at other locations of the gene. Methods DNA was obtained from a consecutive series of 201 mCRC cases (FFPE tissue), wild-type for KRAS exon 2 (codons 12 and 13). Mutational analysis of KRAS (exons 3 and 4), BRAF (exons 11 and 15), and PIK3CA (exons 9 and 20) was performed by high resolution melting (HRM) and positive cases were then sequenced. Results One mutation was present in 23.4% (47/201) of the cases and 3.0% additional cases (6/201) had two concomitant mutations. A total of 53 cases showed 59 mutations, with the following distribution: 44.1% (26/59) in KRAS (13 in exon 3 and 13 in exon 4), 18.6% (11/59) in BRAF (two in exon 11 and nine in exon 15) and 37.3% (22/59) in PIK3CA (16 in exon 9 and six in exon 20). In total, 26.4% (53/201) of the cases had at least one mutation and the remaining 73.6% (148/201) were wild-type for all regions studied. Five of the mutations we report, four in KRAS and one in BRAF , have not previously been described in CRC. BRAF and PIK3CA mutations were more frequent in the colon than in the sigmoid or rectum: 20.8% vs. 1.6% vs. 0.0% ( P= 0.000) for BRAF and 23.4% vs. 12.1% vs. 5.4% ( P =0.011) for PIK3CA mutations. Conclusions About one fourth of mCRC cases wild-type for KRAS codons 12 and 13 present other mutations either in KRAS , BRAF , or PIK3CA , many of which may explain the lack of response to anti-EGFR therapy observed in a significant proportion of these patients.