Detection of HER2 status in breast cancer: comparison of current methods with MLPA and real-time RT-PCR.
Human epidermal growth factor receptor (HER) status is an important prognostic factor in breast cancer. There is no globally accepted method for determining its status, and which method is most precise is still a matter of debate. We here analyzed HER2 mRNA expression by quantitative reverse transcription-PCR (qRT-PCR) and HER2 DNA amplification using multiplex ligation-dependent probe amplification (MLPA). In parallel, we performed a routine evaluation of HER2 protein by immunohistochemistry (IHC). To assess the accuracy of the RT-PCR and MLPA techniques, a combination of IHC and fluorescence in situ hybridization (FISH) was used, substituting FISH when the results of IHC were ambiguous (2+) and for those IHC results that disagreed with MLPA and qRT-PCR, this approach being termed IHC-FISH. The IHC results for four samples were not compatible with the MLPA and qRT-PCR results; the MLPA and qRT-PCR results for these samples were confirmed by FISH. The correlations between IHC-FISH and qRT-PCR or MLPA were 0.945 and 0.973, respectively. The ASCO/CAP guideline IHC/FISH correlation with MLPA was (0.827) and with RT-PCR was (0.854). The correlations between the IHC results (0, 1+ as negative, and 3+ as positive) and qRT-PCR and MLPA techniques were 0.743 and 0.831, respectively. Given the shortcomings of IHC analysis and greater correlations between MLPA, qRT-PCR, and FISH methods than IHC analysis alone with each of these three methods, we propose that MLPA and real-time PCR are good alternatives to IHC. However a suitable cut-off point for qRT- PCR is a prerequisite for determining the exact status of HER2.
Genetics Research Center, University of Social Welfare and Rehabilitation Sciences, Tehran, Iran E-mail : fbehjati@gmail.com.
Reza
R
Pazhoomand
Elahe
E
Keyhani
Mehdi
M
Banan
Hossein
H
Najmabadi
Faranak
F
Khodadadi
Alireza
A
Iraniparast
Farnaz
F
Feiz
Keivan
K
Majidzadeh
Ideh
I
Bahman
Fatemeh Aghakhani
FA
Moghadam
Atoosa Madadkar
AM
Sobhani
Ahad
A
Muhammadnejad
Seyedeh Sedigheh
SS
Abedini
Farkhondeh
F
Behjati
D001943:Breast Neoplasms / Q000175:diagnosis / Q000235:genetics* / Q000378:metabolism*; D005260:Female; D005784:Gene Amplification; D006801:Humans; D007124:Immunoenzyme Techniques; D017404:In Situ Hybridization, Fluorescence; D009361:Neoplasm Invasiveness; D011379:Prognosis; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000235:genetics* / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*
D018719:Erb-b2 Receptor Tyrosine Kinases; C508053:ERBB2 protein, human