title: Alterations in epidermal growth factor receptors 1 and 2 in esophageal squamous cell carcinomas.
abstract: BACKGROUND
  Esophageal squamous cell carcinoma (ESCC) shows a 5-year survival rate below 10%, demonstrating the urgency in improving its treatment. Alterations in epidermal growth factor receptors are closely related to malignancy transformation in a number of tumors and recent successful targeted therapies have been directed to these molecules. Therefore, in this study, we analyzed the expression of EGFR and HER2 and evaluated EGFR mutation profile as well as the presence of mutations in hotspots of KRAS and BRAF in ESCC patients.
  
  
  METHODS
  We performed RT-qPCR, immunohistochemistry and Fluorescent in situ hybridization to determine EGFR and HER2 expression in ESCC patients, and direct sequencing and PCR-RFLP for mutations and polymorphism analysis.
  
  
  RESULTS
  Our results showed an increased EGFR mRNA expression in tumors compared to surrounding tissue (p <0.05), with 11% of the cases presenting at least a four-fold difference between tumor and paired adjacent mucosa. EGFR protein overexpression was present only in 4% of the cases. The median expression of HER2 mRNA was not different between tumors and adjacent mucosa. Still, 7% of the tumors presented at least a 25-fold higher expression of this gene when compared to its paired counterpart. Immunohistochemical analysis revealed that 21% of the tumors were positive for HER2 (scores 2+ and 3+), although only 3+ tumors presented amplification of this gene. Mutation analysis for EGFR (exons 18-21), KRAS (codons 12 and 13) and BRAF (V600E) showed no mutations in any of the hotspots of these genes in almost 100 patients analyzed. EGFR presented synonymous polymorphisms at codon 836 (C>T) in 2.1% of the patients, and at codon 787 (G>A) in 79.2% of the cases. This last polymorphism was also evaluated in 304 healthy controls, which presented a similar frequency (73.7%) in comparison with ESCC patients. The absence of mutations of EGFR, KRAS and BRAF as well as the overexpression of EGFR and HER2 in less than 10% of the patients suggest that this signaling pathway is altered in only a small proportion of patients with ESCC.
  
  
  CONCLUSIONS
  HER receptors target therapies may have the potential to be effective in only a minor fraction of patients with ESCC.
authors.affiliation: Programa de Carcinogênese Molecular, Instituto Nacional de Câncer, Coordenação de Pesquisa, Rua André Cavalcanti, 37 - 6º andar, Bairro de Fátima, Rio de Janeiro, Rio de Janeiro, CEP: 20231-050, Brazil.
authors.forename: Isabela Martins
authors.initials: IM
authors.lastname: Gonzaga
authors.forename: Sheila Coelho
authors.initials: SC
authors.lastname: Soares-Lima
authors.forename: Paulo Thiago Souza
authors.initials: PT
authors.lastname: de Santos
authors.forename: Tania Cristina Moita
authors.initials: TC
authors.lastname: Blanco
authors.forename: Bruno Souza Bianchi
authors.initials: BS
authors.lastname: de Reis
authors.forename: Danielle Carvalho
authors.initials: DC
authors.lastname: Quintella
authors.forename: Ivanir Martins
authors.initials: IM
authors.lastname: de Oliveira
authors.forename: Paulo Antonio Silvestre
authors.initials: PA
authors.lastname: de Faria
authors.forename: Cleber Dario Pinto
authors.initials: CD
authors.lastname: Kruel
authors.forename: Nelson Adami
authors.initials: NA
authors.lastname: Andreollo
authors.forename: Tatiana Almeida
authors.initials: TA
authors.lastname: de Simão
authors.forename: Luis Felipe Ribeiro
authors.initials: LF
authors.lastname: Pinto
mesh_terms: D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D002294:Carcinoma, Squamous Cell / Q000235:genetics* / Q000378:metabolism; D004252:DNA Mutational Analysis; D066246:ErbB Receptors / Q000235:genetics* / Q000378:metabolism; D004938:Esophageal Neoplasms / Q000235:genetics* / Q000378:metabolism; D005260:Female; D018773:Genes, erbB-1; D006801:Humans; D007150:Immunohistochemistry; D017404:In Situ Hybridization, Fluorescence; D008297:Male; D008875:Middle Aged; D012150:Polymorphism, Restriction Fragment Length; D020641:Polymorphism, Single Nucleotide; D011518:Proto-Oncogene Proteins / Q000235:genetics; D048493:Proto-Oncogene Proteins B-raf / Q000235:genetics; D016283:Proto-Oncogene Proteins p21(ras); D012333:RNA, Messenger / Q000032:analysis; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000096:biosynthesis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D018631:ras Proteins / Q000235:genetics
chemical_list: D066246:ErbB Receptors; D011518:Proto-Oncogene Proteins; D048493:Proto-Oncogene Proteins B-raf; D016283:Proto-Oncogene Proteins p21(ras); D012333:RNA, Messenger; D018719:Erb-b2 Receptor Tyrosine Kinases; D018631:ras Proteins; C117307:KRAS protein, human; C512478:EGFR protein, human; C508053:ERBB2 protein, human; C482119:BRAF protein, human