Quantitative RT-PCR assay of HER2 mRNA expression in formalin-fixed and paraffin-embedded breast cancer tissues.
Detection of human epidermal growth factor receptor 2 gene (HER2, also known as erbB2) expression is a preparatory process to decide a treatment strategy for breast cancer patients. 20-30% of breast cancer patients have HER2 overexpression, and they usually show poor recovery rate. For detection of HER2 expression, immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH) methods are conventionally used. Although these methods are accurate and reliable, their time-consuming process and high cost need a concise method with high sensitivity and accuracy. As a complementary method to the current IHC/FISH standard techniques, PCR-based methods have been developed. Here we employed a quantitative PCR method to detect HER2 expression in one hundred ninety nine formalin-fixed and paraffin-embedded (FFPE) breast cancer tissue samples from the patients treated over two years at the Yonsei University Severance Hospital, Republic of Korea. Relative expression of HER2 mRNA in the FFPE samples was analyzed using a quantitative RT-PCR (RT-qPCR) method and the obtained HER2 expression levels were compared with those from IHC/FISH methods. Our results show that the RT-qPCR method was highly concordant with IHC/FISH methods for detecting HER2 expression. Overall sensitivity and specificity of the BrightGen HER2 RT-qDx assay kit (Syantra, Calgary, Canada), which is a kit we used for RT-qPCR analyses, were 93.0% and 89.8% (P < 0.0001), respectively. The diagnostic cut-off value of HER2 RT-qDx for the clinical samples was determined by likelihood ratio, among which the highest likelihood ratio of relative HER2 mRNA levels was over 105.5 (AUC = 0.9466) with the highest sensitivity and specificity. Our study indicates that quantification of HER2 mRNA expression with the RT-qPCR could be an alternative method of conventional IHC/FISH methods.
Department of Biomedical Laboratory Science, College of Health Sciences, Yonsei University Gangwon, Republic of Korea ; Department of Clinical Laboratory Science, College of Medical Science, Daegu Haany University Daegu, Republic of Korea.
Sangjung
S
Park
M&D, Inc., Wonju Eco Environmental Technology Center Gangwon, Republic of Korea.
Hye-Young
HY
Wang
Department of Biomedical Laboratory Science, College of Health Sciences, Yonsei University Gangwon, Republic of Korea ; Institute for Life Science and Biotechnology, Yonsei University Seoul, Republic of Korea.
Sunghyun
S
Kim
Department of Biomedical Laboratory Science, College of Health Sciences, Yonsei University Gangwon, Republic of Korea.
Sungwoo
S
Ahn
Department of Clinical Laboratory Science, Hyejeon College Chungnam, Republic of Korea.
Dongsup
D
Lee
Department of Biomedical Laboratory Science, College of Health Sciences, Yonsei University Gangwon, Republic of Korea.
Yoonjung
Y
Cho
Department of Pathology, Wonju College of Medicine, Yonsei University Gangwon, Republic of Korea.
Kwang Hwa
KH
Park
Department of Biomedical Laboratory Science, College of Natural Sciences, Hoseo University Chungnam, Republic of Korea.
Dongju
D
Jung
Department of Surgery, Yonsei University College of Medicine Seoul, Republic of Korea.
Seung Il
SI
Kim
Department of Biomedical Laboratory Science, College of Health Sciences, Yonsei University Gangwon, Republic of Korea.
Hyeyoung
H
Lee
D014408:Biomarkers, Tumor / Q000235:genetics*; D001706:Biopsy; D001943:Breast Neoplasms / Q000235:genetics* / Q000473:pathology; D045744:Cell Line, Tumor; D005260:Female; D005404:Fixatives; D005557:Formaldehyde; D006785:Hospitals, University; D006801:Humans; D007150:Immunohistochemistry; D017404:In Situ Hybridization, Fluorescence; D016612:Paraffin Embedding; D011237:Predictive Value of Tests; D012333:RNA, Messenger / Q000032:analysis*; D011933:Reagent Kits, Diagnostic; D060888:Real-Time Polymerase Chain Reaction / Q000295:instrumentation; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000235:genetics*; D056910:Republic of Korea; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000295:instrumentation; D016707:Tissue Fixation / Q000379:methods*
D014408:Biomarkers, Tumor; D005404:Fixatives; D005557:Formaldehyde; D012333:RNA, Messenger; D011933:Reagent Kits, Diagnostic; D018719:Erb-b2 Receptor Tyrosine Kinases; C508053:ERBB2 protein, human
Breast cancer; FFPE; HER2; RT-qPCR; molecular diagnosis