Identification of patients with transitional cell carcinoma of the bladder overexpressing ErbB2, ErbB3, or specific ErbB4 isoforms: real-time reverse transcription-PCR analysis in estimation of ErbB receptor status from cancer patients.
OBJECTIVE
The purpose of this research was to quantitatively analyze tumor-specific overexpression of all ErbB receptors and ErbB4 isoforms in transitional cell carcinoma (TCC) of the bladder.


METHODS
A real-time reverse transcription-PCR protocol was set up to simultaneously quantitate the mRNA levels of all four of the ErbB receptors and ErbB4 isoforms. Exon-intron structure of the ErbB4 gene was determined for ErbB4 isoform analysis. The assay was validated by analyzing: (a) defined ErbB cDNAs; (b) cell lines transfected with defined ErbB cDNAs; and (c) cancer cell lines with ErbB status controlled by Western blotting. ErbB mRNA expression was quantitated from 29 clinical samples representing TCC, interstitial cystitis, or histologically normal bladder. Cutoff expression levels predicting neoplasia at 95% probability were determined. ErbB expression and amplification was analyzed by immunohistochemistry and chromogenic in situ hybridization.


RESULTS
Experiments with control cDNAs and cell lines demonstrated that the assay was both specific and sensitive, and that ErbB mRNA levels closely correlated with protein levels in cancer cell lines. Determination of cutoff expression levels indicated tumor-specific overexpression of ErbB2, ErbB3, and specific ErbB4 isoforms in a subset of TCC patients. Significant overexpression of ErbB mRNAs was also detected in cases without amplification of the respective gene or when the protein product was not localized at the cell membrane.


CONCLUSIONS
Bladder cancer patients with tumor-specific overexpression of ErbB receptors or their isoforms were identified. Real-time reverse transcription-PCR could be used for ErbB receptor status quantitation to produce prognostic and predictive information for cancer therapy.
Junttila
Teemu T
TT
Medicity Research Laboratories, and Department of Medical Biochemistry and Molecular Biology, Turku Graduate School of Biomedical Sciences, University of Turku, FIN-20520 Turku.
Laato
Matti
M
Vahlberg
Tero
T
Söderström
Karl-Ove
KO
Visakorpi
Tapio
T
Isola
Jorma
J
Elenius
Klaus
K
D000818:Animals; D015153:Blotting, Western; D002295:Carcinoma, Transitional Cell / Q000175:diagnosis / Q000235:genetics / Q000378:metabolism*; D018856:Cystitis, Interstitial / Q000175:diagnosis / Q000235:genetics / Q000378:metabolism; D066246:ErbB Receptors / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D051379:Mice; D041681:NIH 3T3 Cells; D011379:Prognosis; D020033:Protein Isoforms; D012333:RNA, Messenger / Q000378:metabolism; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000235:genetics* / Q000378:metabolism; D020893:Receptor, ErbB-3 / Q000235:genetics* / Q000378:metabolism; D066247:Receptor, ErbB-4; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection; D001749:Urinary Bladder Neoplasms / Q000175:diagnosis / Q000235:genetics / Q000378:metabolism*
D066246:ErbB Receptors; D020033:Protein Isoforms; D012333:RNA, Messenger; D018719:Erb-b2 Receptor Tyrosine Kinases; D020893:Receptor, ErbB-3; D066247:Receptor, ErbB-4; C587689:ERBB4 protein, human; C587690:Erbb4 protein, mouse