HER2Testing, Clinical Pathological Analysis And Epidemiologic Study In Chinese Breast Cancer Patients

Molecular techniques play an increasingly important role in breast cancer detection and help in the prediction of prognosis and treatment response. According to the receptors status to choose the endocrine therapy and molecular targeted therapy for breast cancer patients is the model of the individual treatment. Especially, epidermal growth factor receptor2(HER2) is an important therapeutic target for breast cancer treatment. Trastuzumab (Herceptin(?), Roche) is a recombinant humanised monoclonal antibody directed against the extracellular domain of the HER2protein that significantly increases the survival of HER2positive patients, both as monotherapy or in combination with chemotherapy. Furthermore, HER2positive tumors are more sensi tive to anthracyclin chemotherapy and seem to respond better to aromatase inhibitors than to tamoxifen. Given its prognostic, predictive and therapeutic impli-cations, an accurate assessment of HER2status for breast cancer patients is crucial.HER2, a185KDa transmemb rane receptor, is a proto-oncogene located on the long arm of chromosome17that encodes a transmembrane receptor protein of the epidermal growth factor receptor family. The tyrosine kinase domains are activated by both homodimerization and heterodimerization. Transcription factors activated by the pathway regulate many genes involved in cell proliferation, survival, differentiation, angiogenesis, and invasion and metastasis. However, with the development of HER2testing and clinical application of trastuzumab in China, a number of problems have cropped up in this field as followed.1. There are many techniques to evaluate HER2status, but the standardization of techniques, the accuracy of detection results, the concordance of different methods, and the expensive reagent from aboard are also controversial.2. The epidermiology of HER2expression of breast cancer patients in China is not clear.3. The incidence of chromosome17polysomy and the association with HER2overexpression is also not clear.4. Whether the serum is a surrogate for tissues to response HER2expression.According to the questions above, this study focused on four parts as followed,Part1. Study on comparison of GLP and PathVysion FISH assays on measuring HER2gene status in breast cancer. This study detected HER2gene status from108cases with invasive ductal breast cancer using GLP and PathVysion HER2probe kits by FISH, compared HER2gene expression differentiation measured by GLP and PathVysion HER2probe, and assessed the sensitivity, the specificity and the accuracy of GLP HER2probe kit. HER2gene amplification cases were25.0%(27/108) and26.9%(29/108) detected by GLP probe kit and PathVysion probe kit, respectively. As assessed with PathVysion HER2probe as the reference, the sensitivity, the specificity and the accuracy of the GLP HER2kit were calculated to be89.7%(26/29),98.7%(78/79) and96.3%(104/108), respectively, whereas the positive predictive value (PPV) and negative predictive value (NPV). GLP probe kit can be considered to be high sensitivity and specificity, and it has the widespread clinical application value in assessing HER2gene status for breast cancer patients.Part2. The epidermiological characteristics of HER2expression in Chinese breast cancer. We performed a multicenter study on the HER2status in3149breast cancer specimens from different ethnic populations and areas in China by immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH) assays. The positive rates for HER2over-expression and HER2amplification were23.3%and27.5%in this population, respectively. The concordance between IHC and FISH was71.2%(Kappa=0.494, p<0.001). The HER2amplification was associated significantly with advanced tumor stage (Ⅲ or Ⅳ)(p=0.002), large tumor size (>5cm)(p=0.002), moderately and poorly histological grades (p<0.001), postmenopause (p<0.001), ER/PR-(p=0.002), and having≥4lymph nodes affected (p<0.001) in this population. The positive rates of HER2amplification in specimens from Man and Hui Chinese were significantly higher than that in other Chinese populations. There are slightly higher positive rates of HER2expression and amplification in Chinese patients with breast cancer. These findings may provide new insights into understanding the epidemiological features of HER2expression and amplification, and may be valuable for clinical practice.Part3. Impact of polysomy17on HER2testing of breast cancer. We collected384cases of breast cancer collected from Cancer Institute/Hospital of Chinese Academy of Medical Sciences&Peking Union Medical College. Chromosome17copies and HER2gene status were identified by fluorescent in situ hybridization, and the corresponding HER2immunohistochemistry was obtained. The average CEP17copy number for the group was2.1(range,1.0-12.4). Fourty-eight cases (13.8%,48/348) were idenfied as chromosome17polosomy with CEP17copy number≥3. Ninety-two (26.4%) cases had>6copies of HER2per nucleus, and95cases (25.3%) qualified as HER2gene amplified using the HER2/CEP17ratio (>2.2) guideline. All these cases had>6HER2signals, presented38.0%cases with>3CEP17copy number. HER2protein expression showed significant positive correlations with both HER2gene copy number and HER2/CEP17ratio (p<0.01, r=0.56and0.64, respectively). In conclusion, increased CEP17signals detected in invasive breast carcinomas may lead to discordant interpretation of HER2gene amplification in a significant proportion of the cases, depending on which criterion (ratio vs absolute number) is used for interpretation. However, increased gene dosage (>6HER2genes or HER2/CEP17ratio>2.2), regardless of the evaluation method, is positively correlated with HER2protein expression. The polysomy of chromosome17can not be a independent predictive factor for HER2gene amplification.Part4. Relationship of serum HER2level and tissue HER2expression in early stage of breast cancer. The measurement of the HER2protein in the serum of metastatic breast cancer patients has now been reported, but there are no consistent data to support the clinical utility of serum HER2extracellular domain (ECD) for patients with early breast cancer. We aimed to evaluate the correlation between serum HER2ECD levels and tumor HER2status, and analyze their relationship with clinicopathological parameters in patients with early stage disease. This study was conducted on232breast cancer patients with stage Ⅰ-ⅢA diseases before treatment. Preoperative serum samples were measured by enzyme-linked immunosorbent assay. Tissue HER2status was analyzed by immunohistochemistry and fluorescence in situ hybridization assays. The median serum HER2ECD concentration was6.8ng/ml. The best diagnostic cut-off value was7.4ng/ml, with62.9%sensitivity and85.3%specificity. High serum HER2ECD levels were reported in89patients (38.3%) and HER2tissue positive expression was observed in77patients (33.2%) with a moderate concordance of76.7%. Elevated serum HER2ECD correlated with postmenopausal (p<0.001), high tumor grade (p<0.001), negativity of both estrogen (p=0.007) and progesterone receptors (p<0.001),high level of CA153(p=0.039) and TPS (p=0.018). HER2ECD may provide more additional information compared with HER2tissue alone. We supported that it is necessary to decrease the cut-off value in evaluating serum HER2ECD level for early stage of breast cancer.Taken together, we cleared the wide application value of GLP HER2DNA probe kit, got the epidemiological data of HER2expression in Chinese breast cancer patients, found the influence of chromosome17polysomy on HER2status, and explicated the feasibility of serum instead of tissue to detect HER2expression in new diagnosis breast cancer, which may provide the basis for accurately interpreting HER2status in the future clinical practice, and promot individualized targeted therapy in clinical application. Objective To evaluate clinical application of Jin Pujia GLP HER2probe kit in testing HER2gene status of breast cancer through comparing with Vysis PathVysion HER2probe kit. Methods This study detected HER2gene status from108cases with invasive ductal breast cancer using GLP and PathVysion HER2probe kits by FISH, compared HER2gene expression differentiation measured by GLP and PathVysion HER2probe, and assessed the sensitivity, the specificity and the accuracy of GLP HER2probe kit. Results HER2gene amplification cases were25.0%(27/108) and26.9%(29/108) detected by GLP probe kit and PathVysion probe kit, respectively. As assessed with PathVysion HER2probe as the reference, the sensitivity, the specificity and the accuracy of the GLP HER2kit were calculated to be89.7%(26/29),98.7%(78/79) and96.3%(104/108), respectively, whereas the positive predictive value (PPV) and negative predictive value (NPV) were96.3%(26/27) and96.3%(78/81), respectively. With regard to the ability to presume polysomy17, the GLP HER2kit had a sensitivity of93.3%(14/15), a specificity of100%(93/93) and an accuracy of99.1%(107/108). Each lot was run on five non-consecutive days over one month. The GLP and PathVysion HER2probe kit all had high reproducibility with concordance rate of100%. Conclusion GLP probe kit can be considered to be high sensitivity and specificity, and it has the widespread clinical application value in assessing HER2gene status for breast cancer patients. We performed a multicenter study on the HER2status in3149breast cancer specimens from different ethnic populations and areas in China by immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH) assays. The potential association of HER2status with demographic and clinical characteristics was analyzed. The positive rates for HER2over-expression and HER2amplification were23.3%and27.5%in this population, respectively. The concordance between IHC and FISH was71.2%(Kappa=0.494, p<0.001). Furthermore,72.9%of specimens with IHC2+were negative to FISH. The discordance rates among laboratories were from5%to28%for IHC and1%to16%for FISH. The HER2amplification was associated significantly with advanced tumor stage (Ⅲ or Ⅳ)(p=0.002), large tumor size (>5cm)(p=0.002), moderately and poorly histological grades (p<0.001), postmenopause (p<0.001), ER-PR-(p=0.002), and having≥4lymph nodes affected (p<0.001) in this population. The positive rates of HER2amplification in specimens from Man and Hui Chinese were significantly higher than that in other Chinese populations. There are slightly higher positive rates of HER2expression and amplification in Chinese patients with breast cancer than that in Caucasian. These findings indicate the epidemiological features of HER2expression in Chinese breast cancer patients, state the difference between Chinese patients and Western Caucasian patients, which may provide new insights into understanding personalized targeted therapy for breast cancer, and may be valuable for early prevention in clinical practice. Backgroud:The current study was performed to determine the impact of polysomy17on the interpretation of HER2testing of invasive breast carcinomas using fluorescent in situ hybridization methods. Current American Societyof Clinical Oncology/College of American Pathologists guidelines define HER2positive tumors as those with>6HER2genes per nucleus or those with HER2/CEP17(chromosome17) ratio>2.2. These guidelines are potentially con-tradictory in tumors with polysomy of chromosome17. Methods:Chromosome17copies (≥3CEP17signals on average) and HER2gene status were identified by fluorescent in situ hybridization in the384cases of breast cancer collected from Cancer Institute/Hospital of Chinese Academy of Medical Sciences&Peking Union Medical College, and the corresponding HER2immunohistochemistry was obtained. Results:The average CEP17copy number for the group was2.1(range,1.0-12.4). Fourty-eight cases (13.8%,48/348) were idenfied as chromosome17polosomy with CEP17copy number≥3. Ninety-two (26.4%) cases had>6copies of HER2per nucleus, and95cases (25.3%) qualified as HER2gene amplified using the HER2/CEP17ratio (>2.2) guideline. All these cases had>6HER2signals, presented38.0%cases with>3CEP17copy number. HER2protein expression showed significant positive correlations with both HER2gene copy number and HER2/CEP17ratio (p<0.01, r=0.56and0.64, respectively). Conclusions:Increased CEP17signals detected in invasive breast carcinomas may lead to discordant interpretation of HER2gene amplification in a significant proportion of the cases, depending on which criterion (ratio vs absolute number) is used for interpretation. Increased gene dosage (>6HER2genes or HER2/CEP17ratio>2.2), regardless of the evaluation method, is positively correlated with HER2protein expression, but the polysomy of chromosome17can not be a independent predictive factor for HER2gene amplification. Background:The measurement of the human epidermal growth factor receptor2(HER2) protein in the serum of metastatic breast cancer patients has now been reported, but there are no consistent data to support the clinical utility of serum HER2extracellular domain (ECD) for patients with early breast cancer. We aimed to evaluate the correlation between serum HER2ECD levels and tumor HER2status, and analyze their relationship with clinicopathological parameters in patients with early stage disease. Methods:A prospective study was conducted on232breast cancer patients with stage Ⅰ-Ⅱ diseases before treatment. Preoperative serum samples were measured by enzyme-linked immunosorbent assay (ELISA). Tissue HER2status was analyzed by immunohistochemistry and fluorescence in situ hybridization assays. Results:The median serum HER2ECD concentration was6.8ng/ml (range1.3-42.1ng/ml). The best diagnostic cut-off value was7.4ng/ml, with72.9%sensitivity and85.3%specificity, which was lower than HER2ECD cut-off value with metastasis breast cancer. High serum HER2ECD levels were reported in89patients (38.3%) and HER2tissue positive expression was observed in77patients (33.2%) with a moderate concordance of76.7%. Elevated serum HER2ECD correlated with postmenopausal (p<0.001), high tumor grade (p<0.001), negativity of both estrogen (p=0.007) and progesterone receptors (p<0.001), high level of carbohydrate antigen153(CA153)(p=0.039) and tissue polypeptide specific antigen (TPS)(p=0.018). Conclusion: HER2ECD may provide more additional information compared with HER2tissue alone. We support that it is necessary to decrease the cut-off value in evaluating serum HER2ECD level for early stage of breast cancer.