title: Challenges in EGFRvIII detection in head and neck squamous cell carcinoma.
abstract: OBJECTIVE
  Head and neck squamous cell carcinoma (HNSCC) accounts for more than 5% of all cancers worldwide. The mortality rate of HNSCC has remained unchanged (approximately 50%) over the last few decades. Ubiquitous overexpression of wild type EGFR in many solid tumors has led to the development of EGFR targeted therapies. EGFR can be constitutively activated via several mechanisms including the truncated, EGFR variant III isoform (EGFRvIII). EGFRvIII lacks exons 2-7 and has been reported to be present in up to 20-40% of HNSCC. EGFRvIII has been shown to contribute to cetuximab resistance. The mechanisms leading to EGFRvIII expression in HNSCC are unknown. The present investigation was undertaken to determine the etiology of EGFRvIII in HNSCC.
  
  
  METHODS
  Fixed HNSCC and glioma tissues were analyzed by fluorescence in situ hybridization for EGFR amplification. DNA and RNA from fresh frozen specimens were used to determine the presence of EGFRvIII transcripts and the mechanisms of expression via PCR, RT-PCR and RNA sequencing.
  
  
  RESULTS
  Unlike glioma, EGFRvIII expression in HNSCC did not correlate with EGFR amplification. We found evidence of genomic deletion of the exon 2-7 in 6 of 7 HNSCC cases examined, however, the presence of genomic deletion did not always result in mRNA expression of EGFRvIII. RNA sequencing with automated alignment did not identify EGFRvIII due to microhomology between intron 1 and exon 8. RNA sequencing analyzed by manual alignment methods did not correlate well with RT-PCR and PCR findings.
  
  
  CONCLUSIONS
  These findings suggest that genomic deletion as well as additional regulatory mechanisms may contribute to EGFRvIII expression in HNSCC. Further, large scale automated alignment of sequencing are unlikely to identify EGFRvIII and an assay specifically designed to detect EGFRvIII may be necessary to detect this altered form of EGFR in HNSCC tumors.
authors: Department of Pathology, University of Pittsburgh, Pittsburgh, PA, United States of America.
authors: Sarah E
authors: SE
authors: Wheeler
authors: Department of Otolaryngology, University of Pittsburgh and University of Pittsburgh Cancer Institute, Pittsburgh, PA, United States of America.
authors: Ann Marie
authors: AM
authors: Egloff
authors: Department of Pathology, University of Pittsburgh, Pittsburgh, PA, United States of America.
authors: Lin
authors: L
authors: Wang
authors: Department of Neurological Surgery, Northwestern University, Chicago, IL, United States of America.
authors: C David
authors: CD
authors: James
authors: Dana-Farber Cancer Institute, Harvard Medical School, Boston, MA, United States of America.
authors: Peter S
authors: PS
authors: Hammerman
authors: Department of Pathology, University of Pittsburgh, Pittsburgh, PA, United States of America; Department of Otolaryngology, University of Pittsburgh and University of Pittsburgh Cancer Institute, Pittsburgh, PA, United States of America; Dana-Farber Cancer Institute, Harvard Medical School, Boston, MA, United States of America; Department of Pharmacology & Chemical Biology, University of Pittsburgh, Pittsburgh, PA, United States of America.
authors: Jennifer R
authors: JR
authors: Grandis
mesh_terms: D061067:Antibodies, Monoclonal, Humanized / Q000008:administration & dosage; D000970:Antineoplastic Agents / Q000008:administration & dosage; D001483:Base Sequence; D002294:Carcinoma, Squamous Cell / Q000188:drug therapy / Q000235:genetics* / Q000401:mortality; D000068818:Cetuximab; D015331:Cohort Studies; D019008:Drug Resistance, Neoplasm / Q000187:drug effects / Q000235:genetics; D066246:ErbB Receptors / Q000235:genetics*; D005091:Exons; D005260:Female; D015972:Gene Expression Regulation, Neoplastic / Q000235:genetics; D006258:Head and Neck Neoplasms / Q000188:drug therapy / Q000235:genetics* / Q000401:mortality; D006801:Humans; D008297:Male; D017384:Sequence Deletion
chemical_list: D061067:Antibodies, Monoclonal, Humanized; D000970:Antineoplastic Agents; C512478:EGFR protein, human; D066246:ErbB Receptors; D000068818:Cetuximab