title: Alterations in the human epidermal growth factor receptor 2-phosphatidylinositol 3-kinase-v-Akt pathway in gastric cancer.
abstract: OBJECTIVE
  To investigate human epidermal growth factor receptor 2 (HER2)-phosphatidylinositol 3-kinase (PI3K)-v-Akt murine thymoma viral oncogene homolog signaling pathway.
  
  
  METHODS
  We analyzed 231 formalin-fixed, paraffin-embedded gastric cancer tissue specimens from Japanese patients who had undergone surgical treatment. The patients' age, sex, tumor location, depth of invasion, pathological type, lymph node metastasis, and pathological stage were determined by a review of the medical records. Expression of HER2 was analyzed by immunohistochemistry (IHC) using the HercepTest(TM) kit. Standard criteria for HER2 positivity (0, 1+, 2+, and 3+) were used. Tumors that scored 3+ were considered HER2-positive. Expression of phospho Akt (pAkt) was also analyzed by IHC. Tumors were considered pAkt-positive when the percentage of positive tumor cells was 10% or more. PI3K, catalytic, alpha polypeptide (PIK3CA) mutations in exons 1, 9 and 20 were analyzed by pyrosequencing. Epstein-Barr virus (EBV) infection was analyzed by in situ hybridization targeting EBV-encoded small RNA (EBER) with an EBER-RNA probe. Microsatellite instability (MSI) was analyzed by polymerase chain reaction using the mononucleotide markers BAT25 and BAT26.
  
  
  RESULTS
  HER2 expression levels of 0, 1+, 2+ and 3+ were found in 167 (72%), 32 (14%), 12 (5%) and 20 (8.7%) samples, respectively. HER2 overexpression (IHC 3+) significantly correlated with intestinal histological type (15/20 vs 98 /205, P = 0.05). PIK3CA mutations were present in 20 cases (8.7%) and significantly correlated with MSI (10/20 vs 9/211, P < 0.01). The mutation frequency was high (21%) in T4 cancers and very low (6%) in T2 cancers. Mutations in exons 1, 9 and 20 were detected in 5 (2%), 9 (4%) and 7 (3%) cases, respectively. Two new types of PIK3CA mutation, R88Q and R108H, were found in exon1. All PIK3CA mutations were heterozygous missense single-base substitutions, the most common being H1047R (6/20, 30%) in exon20. Eighteen cancers (8%) were EBV-positive and this positivity significantly correlated with a diffuse histological type (13/18 vs 93/198, P = 0.04). There were 7 cases of lymphoepithelioma-like carcinomas (LELC) and 6 of those cases were EBV-positive (percent/EBV: 6/18, 33%; percent/all LELC: 6/7, 86%). pAkt expression was positive in 119 (53%) cases but showed no correlation with clinicopathological characteristics. pAkt expression was significantly correlated with HER2 overexpression (16/20 vs 103/211, P < 0.01) but not with PIK3CA mutations (12/20 vs 107/211, P = 0.37) or EBV infection (8/18 vs 103/211, P = 0.69). The frequency of pAkt expression was higher in cancers with exon20 mutations (100%) than in those with exon1 (40%) or exon9 (56%) mutations. One case showed both HER2 overexpression and EBV infection and 3 cases showed both PIK3CA mutations and EBV infection. However, no cases showed both PIK3CA mutations and HER2 overexpression. One EBV-positive cancer with PIK3CA mutation (H1047R) was MSI-positive. Three of these 4 cases were positive for pAkt expression. In survival analysis, pAkt expression significantly correlated with a poor prognosis (hazard ratio 1.75; 95%CI: 1.12-2.80, P = 0.02).
  
  
  CONCLUSIONS
  HER2 expression, PIK3CA mutations and EBV infection in gastric cancer were characterized. pAkt expression significantly correlates with HER2 expression and with a poor prognosis.
authors.affiliation: First Department of Internal Medicine, Sapporo Medical University School of Medicine, Sapporo 60-8543, Japan.
authors.forename: Yasutaka
authors.initials: Y
authors.lastname: Sukawa
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Yamamoto
authors.forename: Katsuhiko
authors.initials: K
authors.lastname: Nosho
authors.forename: Hiroaki
authors.initials: H
authors.lastname: Kunimoto
authors.forename: Hiromu
authors.initials: H
authors.lastname: Suzuki
authors.forename: Yasushi
authors.initials: Y
authors.lastname: Adachi
authors.forename: Mayumi
authors.initials: M
authors.lastname: Nakazawa
authors.forename: Takayuki
authors.initials: T
authors.lastname: Nobuoka
authors.forename: Mariko
authors.initials: M
authors.lastname: Kawayama
authors.forename: Masashi
authors.initials: M
authors.lastname: Mikami
authors.forename: Takashi
authors.initials: T
authors.lastname: Matsuno
authors.forename: Tadashi
authors.initials: T
authors.lastname: Hasegawa
authors.forename: Koichi
authors.initials: K
authors.lastname: Hirata
authors.forename: Kohzoh
authors.initials: K
authors.lastname: Imai
authors.forename: Yasuhisa
authors.initials: Y
authors.lastname: Shinomura
mesh_terms: D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000818:Animals; D058534:Class I Phosphatidylinositol 3-Kinases; D020031:Epstein-Barr Virus Infections / Q000175:diagnosis; D005091:Exons; D005260:Female; D004854:Herpesvirus 4, Human; D006801:Humans; D007150:Immunohistochemistry; D017403:In Situ Hybridization; D008207:Lymphatic Metastasis; D008297:Male; D051379:Mice; D018895:Microsatellite Repeats; D008875:Middle Aged; D009154:Mutation; D051058:Oncogene Protein v-akt / Q000235:genetics*; D019869:Phosphatidylinositol 3-Kinases / Q000235:genetics*; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000235:genetics*; D015398:Signal Transduction; D013274:Stomach Neoplasms / Q000235:genetics* / Q000821:virology
chemical_list: D058534:Class I Phosphatidylinositol 3-Kinases; D051058:Oncogene Protein v-akt; D019869:Phosphatidylinositol 3-Kinases; D018719:Erb-b2 Receptor Tyrosine Kinases; C484760:PIK3CA protein, human; C508053:ERBB2 protein, human