A comprehensive procedural approach to genotyping KRAS and BRAF from paraffin embedded tissues for diagnostic purposes

Raffaele Palmirotta, Giorgia Ludovici, Maria Laura De Marchis, Barbara Leone, Vincenzo Formica, Giuseppe Maria Ettorre, Francesco Cavaliere, David Della-Morte, Patrizia Ferroni, Mario Roselli, Fiorella Guadagni

Research output: Contribution to journalArticle

6 Citations (Scopus)

Abstract

Background: Mutations in the Kirsten Ras 1 (KRAS) and V-Raf Murine Sarcoma Viral Oncogene Homolog B1 (BRAF) genes may be predictive of response to drugs directly linked to the Epidermal Growth Factor Receptor (EGFR) signaling pathway. Materials and Methods: A total of 230 samples from patients with metastatic colorectal cancer were analyzed for KRAS exon 1 and 2 and for BRAF exon 15 mutations. DNA from paraffinembedded tumor sections was analyzed using microdissection, direct sequencing analysis and allelic separation by cloning. Results: KRAS mutations were present in 44.3% of the tumor samples. The mutation frequency at hot-spot codons of exon 1 was 84.2%, whereas noncanonical variants had a frequency of 11.8%. Approximately 4% of the cases exhibited concomitant variations. BRAF mutations were present in 3.9% of the tumor samples. Conclusion: Our experience suggests that sequential microdissection, direct sequencing and allelic separation by cloning may improve the approach to mutational analysis of KRAS and BRAF in patients with colorectal cancer.

Original languageEnglish
Pages (from-to)537-548
Number of pages12
JournalIn Vivo
Volume26
Issue number4
Publication statusPublished - Jul 2012

Fingerprint

Microdissection
Paraffin
Tumors
Exons
Cloning
Tissue
Mutation
Organism Cloning
Colorectal Neoplasms
Epidermal Growth Factor Receptor
Neoplasms
Mutation Rate
Genes
Oncogenes
Codon
Sarcoma
DNA
Pharmaceutical Preparations

Keywords

  • BRAF
  • Colorectal cancer
  • Concurrent mutation
  • Genotyping
  • KRAS
  • Mutational analysis

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)
  • Pharmacology

Cite this

Palmirotta, R., Ludovici, G., De Marchis, M. L., Leone, B., Formica, V., Ettorre, G. M., ... Guadagni, F. (2012). A comprehensive procedural approach to genotyping KRAS and BRAF from paraffin embedded tissues for diagnostic purposes. In Vivo, 26(4), 537-548.

A comprehensive procedural approach to genotyping KRAS and BRAF from paraffin embedded tissues for diagnostic purposes. / Palmirotta, Raffaele; Ludovici, Giorgia; De Marchis, Maria Laura; Leone, Barbara; Formica, Vincenzo; Ettorre, Giuseppe Maria; Cavaliere, Francesco; Della-Morte, David; Ferroni, Patrizia; Roselli, Mario; Guadagni, Fiorella.

In: In Vivo, Vol. 26, No. 4, 07.2012, p. 537-548.

Research output: Contribution to journalArticle

Palmirotta, R, Ludovici, G, De Marchis, ML, Leone, B, Formica, V, Ettorre, GM, Cavaliere, F, Della-Morte, D, Ferroni, P, Roselli, M & Guadagni, F 2012, 'A comprehensive procedural approach to genotyping KRAS and BRAF from paraffin embedded tissues for diagnostic purposes', In Vivo, vol. 26, no. 4, pp. 537-548.
Palmirotta R, Ludovici G, De Marchis ML, Leone B, Formica V, Ettorre GM et al. A comprehensive procedural approach to genotyping KRAS and BRAF from paraffin embedded tissues for diagnostic purposes. In Vivo. 2012 Jul;26(4):537-548.
Palmirotta, Raffaele ; Ludovici, Giorgia ; De Marchis, Maria Laura ; Leone, Barbara ; Formica, Vincenzo ; Ettorre, Giuseppe Maria ; Cavaliere, Francesco ; Della-Morte, David ; Ferroni, Patrizia ; Roselli, Mario ; Guadagni, Fiorella. / A comprehensive procedural approach to genotyping KRAS and BRAF from paraffin embedded tissues for diagnostic purposes. In: In Vivo. 2012 ; Vol. 26, No. 4. pp. 537-548.
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