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Serum HER2 extracellular domain as a potential alternative for tissue HER2 status in metastatic gastric cancer patients
Aim: We investigated whether serum concentrations of the HER2 extracellular domain (ECD) can be used as an alternative to test tissue HER2 status in metastatic gastric cancer. Materials & methods: A total of 133 cases of metastatic gastric cancer were included in present study. Serum HER2 ECD was measured by chemiluminescence immunoassay (CLIA). Receiver operating characteristic curve analysis was used to determine optimal serum HER2 ECD concentrations for differentiation between positive and negative HER2 status. Results: The median level of serum HER2 ECD was 9.6 ng/ml in metastatic gastric cancer patients. There was a significant relationship between serum and tissue levels of HER2 protein (p < 0.001). Area under the curve for serum HER2 ECD was 0.771 (95% CI: 0.682–0.860). Conclusion: Levels of serum HER2 ECD are highly correlated with tissue HER2 status in metastatic gastric cancer. Serum HER2 ECD assay can be considered as a potential alternative for tissue HER2 status. Keywords: ECD • extracellular domain • HER2 • metastatic gastric cancer • serum
The human epidermal growth factor receptor-2 proto-oncogene, also known as HER2 and c-erbB-2, encoding a growth factor receptor, has been found to play an important role in gastric cancer [1] . HER2 has been shown to be overexpressed and amplified in 6–35% of gastric tumors [2] . The Phase III ToGA trial showed that adding the HER2targeted humanized monoclonal antibody trastuzumab to chemotherapy significantly improved survival of patients with advanced gastric cancer. Moreover, the study indicated that patients with immunohistochemistry (IHC) 2+/FISH+ and IHC 3+ (HER2 positive) had an increased benefit from trastuzumab treatment [3] . HER2-positive patients are eligible for HER2-targeted (trastuzumab) treatment, although the methods of assessment and prognostic value of HER2 have been subject to debate [4] . However, both the methods IHC and FISH for detecting HER2 status have limitations. One such limitation is the lack of ‘real-time’ follow-up due to the dependency of both assays on tumor biopsies. In many
10.2217/BMM.14.10 © 2014 Future Medicine Ltd
Zhi Peng1, Yi Liu2, Yanyan Li1, Xiaotian Zhang1, Jun Zhou1, Ming Lu1, Qingqing Li1 & Lin Shen*,1 Department of Gastrointestinal Oncology, Key Laboratory of Carcinogenesis & Translational Research (Ministry of Education), Peking University Cancer Hospital & Institute, Beijing, China 2 Translational Medicine Center, Laboratory of Oncology, Affiliated Hospital of Academy of Military Medical Sciences, Beijing, China *Author for correspondence Tel.: +86 10 8819 6561 Fax: +86 10 8819 6561 lin100@ medmail.com.cn 1
patients with metastatic gastric cancer, the re-evaluation of HER2 status by a tissue biopsy of the metastatic lesion is not feasible due to location of the metastatic site. Another important issue in HER2-targeted therapy is HER2 heterogeneity [5] . The heterogeneity may result in the inaccurate assessment of the HER2 status and the consequent rendering patients unsuitable for treatment of trastuzumab [6] . In addition, HER2 status may vary between different metastatic sites and also change during treatment [7] . New tests have been continuing development. To find more convenient, reproducible and accurate detection to identify patients with gastric cancer for anti-HER2 therapy is particularly urgent [8] . Serum extracellular domain (ECD) fragment of HER2 represents a noninvasive, relatively fast, reproducible and quantifiable biomarker that could supplement existing HER2 testing [9] . This extracellular fragment of the receptor is released from the surface of tumor cells into the circulation by the ADAM proteases [10,11] . It retains its sig-
Biomarkers Med. (2014) 8(5), 663–670
part of
ISSN 1752-0363
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Research Article Peng, Liu, Li et al. naling activity and, upon phosphorylation, can contribute to disease progression [12] . Studies have found elevated levels of HER2 ECD in the serum of many breast cancer patients, and in most cases, high levels of serum HER2 ECD are associated with higher relapse rates and worse prognosis [13] . The association between serum HER2 ECD and tissue HER2 status in gastric cancer patients has been investigated in a number of studies, too [14–17] . However, most of these studies included different groups of patients and used different methods. In the present study, we evaluated the correlation of HER2 tissue expression with serum levels of soluble HER2 ECD in patients with metastatic gastric cancer. Furthermore, we investigated whether serum HER2 ECD could be an alternative method to choose patients who are suitable for anti-HER2 therapy. Materials & methods Subjects
This retrospective study included unselected consecutive 133 patients who were diagnosed with metastatic gastric cancer at the Department of Gastroenterology Oncology in Peking University Cancer Hospital in China from February 2010 to May 2013. All patients were collected as following criteria: patients had histologically confirmed metastatic gastric cancer with serum and primary gastric tumor specimens available; patients were >18 years old. Their peripheral bloods before chemotherapy were drawn, after signed a written informed consents for their blood to be used for research. Demographic and clinicopathologic features such as age, gender, disease site, metastatic site and number of metastases were retrospectively obtained from the patient records. This study was approved by the medical ethics committee of Peking University Cancer Hospital and was performed according to the Declaration of Helsinki Principles. Tissue expression of HER2: IHC & FISH
Tumor biopsies of primary lesion were fixed in 10% neutral-buffered formalin overnight and embedded in paraffin blocks. Paraffin-embedded tissues were cut into 4-µm thick slices, and IHC was performed using HercepTest II™ (Dako, Glostrup, Denmark) according to manufacturer instructions. All IHC specimens were stained at the Department of Pathology, Peking University Cancer Hospital. The scoring criteria used in the ToGA trial were applied to give IHC scores for HER2 expression. Furthermore, FISH was performed with Abbott-Vysis PathVysion™ (Abbott Laboratories, IL, USA) according to manufacturer
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instructions if HER2 protein expression in IHC 2+ cases. Tumor specimens with a HER2:CEP17 signal ratio ≥2.0 were considered HER2 FISH positive. Serum HER2 levels using CLIA
Venous blood specimens were collected into separator tubes before treatment. Serum specimens were recovered by centrifuging blood specimens at 3000 rpm for 10 min and stored at -70°C. All samples stored in a standardized fashion and were detected at the same time. Serum HER2 levels were measured using ADVIA Centaur CP Immunoassay System (Siemens Healthcare Diagnostics, Erlangen, Germany) according to manufacturer instructions. Quality control was ensured by assaying the two levels of control sera supplied with the kit in each series. The intra-assay coefficient of variation (CV) was less than 6% and the interassay CV was 10%. Serum HER2 ECD measurement was carried out in a blinded manner without knowledge about the results of HER2 IHC and FISH. Statistical analysis
We used SAS software (version 8.0, SAS Institute Inc., NC, USA) for statistical analysis. The χ 2 test was used to assess the association between clinical and pathological features and the different levels of expression of HER2 ECD. Wilcoxon rank sum test or Kruskal–Wallis H test was used to compare the serum HER2 concentration in the patients with different tissue HER2 status. p-values less than 0.05 were considered statistically significant. To estimate sample size, we carried out a preliminary experiment first. We calculated that group sample sizes of 37 and 74 achieve 80% power to detect a difference of 0.4 between the null hypothesis that both group (HER2 positive and HER2 negative) means are 2.5 (log conversion) and the alternative hypothesis that the mean of HER2 negative group is 2.1 with estimated standard deviations of 0.8 and 0.4 and with a significance level (alpha) of 0.05 using a two-sided t wo-sample t-test. Based on the above calculation, over 111 samples are required to be involved in this study. Therefore, we choose the latest consecutive periods including over 111 patients to screen subjects. The receiver operating characteristic (ROC) curve was analyzed to evaluate the discriminating ability of serum HER2 and to arrive at a serum HER2 cutoff for predicting tissue HER2 positivity. To obtain appropriate serum level cutoff values, we calculated the total sensitivity and specificity for each cutoff value and then chose the cutoff values that maximized the sum of s ensitivity plus 1-specificity.
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Serum HER2 extracellular domain in metastatic gastric cancer patients
Research Article
Table 1. Clinicopathologic characteristics and tissue HER2 status of 133 patients with metastatic gastric cancer. Characteristic
All patients, n (%)
HER2 positive, n (%)
HER2 negative, n (%)
Total
133 (100)
43 (32.3)
90 (67.7)
Age (years): – >60 – ≥60
73 (54.9) 60 (45.1)
28 (21.15) 15 (11.3)
45 (33.8) 45 (33.8)
Gender: – Male – Female
102 (76.7) 31 (23.3)
36 (27.1) 7 (5.3)
66 (49.6) 24 (18.0)
Primary site: – GEJ – Non-GEJ
48 (36.1) 85 (63.9)
13 (9.8) 30 (22.6)
35 (26.3) 55 (41.3)
Tumor type (Laurén classification): – Diffuse type 34 (25.6) – Intestinal type 67 (45.9) – Mixed type 32 (24.0)
10 (7.5) 23 (17.3) 10 (7.5)
24 (18.0) 44 (33.1) 22 (16.5)
Metastatic site: – Liver – Lung – Lymph node
62 (47.4) 23 (20.3) 96 (72.2)
17 (12.8) 12 (12.0) 27 (20.3)
45 (34.6) 11 (8.3) 69 (51.9)
Metastasis (n): –1 –2 – ≥3
15 (11.3) 52 (35.3) 66 (53.4)
6 (4.5) 16 (12.0) 21 (15.8)
9 (6.8) 36 (27.1) 45 (33.8)
GEJ: Gastroesophageal junction.
Results Patient characteristics
A total of 133 unselected consecutive patients (102 males and 31 females) with metastatic gastric cancer were enrolled in this study. Patient characteristics are summarized in Table 1. All patients were diagnosed as stage IV gastric cancer according to the American Joint Committee on Cancer (AJCC) 7th edition at the time of initial diagnosis. The median age was 59 years (range: 24–79 years). For the selection bias, we further analyzed the demographic and clinicopathologic features, no statistically differences were found in this study (data not shown). Therefore, it was considered that these subjects could represent the patients treated in our department. There were 66 IHC 1+/0 cases, 31 IHC 2+ cases and 36 IHC 3+ cases. Seven cases were HER2 gene amplification in IHC 2+ cases (Figure 1). According to ToGA trial criteria for HER2 positivity (IHC 3+ or IHC 2+ plus FISH+), 43 cases were positive of tissue HER2 expression and 90 cases were negative of tissue HER2 expression. Correlation between serum HER2 ECD levels & tissue HER2 status
The median serum HER2 level was 9.6 ng/ml in all metastatic gastric cancer patients (interquartile range:
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8.3–13.3 ng/ml; range: 4.4–350.0 ng/ml). Significant differences was found in serum HER2 ECD levels between patients with 3+ and those with 2+ and 0/1+ (χ2: 26.505; p < 0.0001) (Figure 2). Serum HER2 ECD levels were significantly higher in patients with 3+ (median: 13.3 ng/ml; interquartile range: 9.5–22.6 ng/ml; range: 6.9–350.0 ng/ml) than those in patients with 2+ (median: 10.1 ng/ml; interquartile range: 8.5–13.0 ng/ml; range: 4.8–23.7 ng/ml) and with 0/1+ (median: 8.9 ng/ml; interquartile: 7.5–10.3 ng/ml; range: 4.4–18.6 ng/ml). There was a significant relationship between serum and tissue levels of HER2 protein (p < 0.001). We defined IHC 3+ and IHC 2+ plus FISH positive as HER2 positive according to the ToGA trial. The HER2 ECD levels of HER2 positive (median: 13.0 ng/ml; interquartile range: 9.5–21.3 ng/ml; range: 5.5–350.0 ng/ml) were significantly higher than HER2 negative (median: 9.0 ng/ml; interquartile range: 7.7–10.3 ng/ml; range: 4.4–23.7 ng/ml; p < 0.0001) (Figure 3). The ROC curves plotted for serum HER2 ECD in patients with metastatic gastric cancer versus tissue status are shown in Figure 4. The area under the curve generated from the serum HER2 ECD was 0.771 (95% CI: 0.682–0.860) for all 133 patients with metastatic gastric cancer. The set point of serum HER2 ECD at 10.65 ng/ml to identify tissue HER2 status has
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A
B
Figure 1. HER2 gene amplification evaluated by FISH. (A) FISH-positive case; (B) FISH-negative case (magnification ×1000). Please see color figures online at www.futuremedicine.com/doi/pdf/10.2217/bmm.14.10.
a sensitivity of 67.4% and a specificity of 78.9%. ROC curve analysis showed 94% specificity and 40% sensitivity for tissue HER2 positivity at 15.0 ng/ml serum HER2 ECD. Relationships between serum HER2 ECD levels and clinicopathological variables are summarized in Table 2. Gender, Laurén classification, live metastases and lung metastases were significantly associated with serum HER2 levels. There was no association between serum HER2 ECD levels and age, primary tumor site and number of metastases. Discussion This study indicated a strong correlation between serum HER2 ECD concentration and tissue IHC
Log HER2 ECD
3.0 2.5
1+/0 2+ 3+
2.0 1.5 1.0 0.5 1+/0
2+ HER2 IHC
3+
Figure 2. Serum HER2 extracellular domain concentration according to tissue HER2 status in 133 patients with metastatic gastric cancer. IHC staining intensity and serum HER2 ECD level. Please see color figures online at www.futuremedicine. com/doi/pdf/10.2217/bmm.14.10. ECD: Extracellular domain; IHC: Immunohistochemistry.
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HER2 test results in metastatic gastric cancer, which suggested detection of serum HER2 ECD concentration can be a potential alternative way to choose patients for trastuzumab treatment. Serum HER2 ECD detection in breast cancer patients has been discovered in many studies. The elevated serum HER2 ECD levels are more frequently observed in metastatic breast cancers. The concentration of serum HER2 ECD is dynamic during disease progression and treatment. However, detection of serum HER2 ECD and establishment of its potential clinical usefulness is still under debate. Monitoring of HER2 ECD will not replace FISH/IHC assays right now but definitely will complement the tissue assay to offer a real-time picture of HER2/neu status in breast cancer patients [8,12] . There are two methods for detecting serum HER2 ECD approved in breast cancer for monitoring patients with metastatic disease: enzyme immunoassay (EIA; Bayer Immuno 1, Siemens Healthcare Diagnostics, Erlangen, Germany) and chemiluminescence immunoassay (CLIA, ADVIA Centaur System, Siemens Healthcare Diagnostics) [18] . Previously, serum HER2 ECD levels were usually determined by EIA. However, trastuzumab therapy influences EIA results since the therapeutic monoclonal antibody acts as a competitive inhibitor. CLIA was a more sensitive method than EIA for measuring serum HER2 ECD levels in patients who received trastuzumab [19] . Bayer Immuno 1 and ADVIA Centaur assays (used in automated systems) are approved for monitoring patients with metastatic disease. To date, only one study tested serum HER2 ECD levels in gastric cancer patients by CLIA [17] . In this study, CLIA was used to detect the serum HER2 ECD. To our knowledge, four articles have reported the serum HER2 ECD in gastric cancer patients
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Serum HER2 extracellular domain in metastatic gastric cancer patients
up to 1 May 2013. However, the relationship between serum and tissue levels of HER2 remains controversial. Three of them detect serum HER2 levels in gastric cancer patients by EIA [14–16] , one by CLIA [17] . Three studies are not consistent with our result, which showed no correlation with tissue HER2 status [15–17] . One explanation might be the different characteristics of included patients. All of the four studies used different stages (I–IV) of gastric cancer. This is the first study to compare the consistency of serum HER2 ECD and tissue HER2 status in patients with metastatic gastric cancer. The cutoff value (10.65 ng/ml) of serum HER2 ECD in present study was lower than in other studies (15 ng/ml) [20,21] . However, in Witzel’s study, the cutoff value of HER2 ECD equal to 10 ng/ml led to better sensitivity and specificity instead of 15 ng/ml [20] . Moreover, the calculated cutoff value of the serum HER2 assay used in our study might be appropriate for Chinese patients with metastatic gastric cancer in general. Different cutoff values need to be established for each population for specific disease from the prospective randomized trial. Because of higher specificity and sensitivity of CLIA test relative to tissue HER2 status, it is recommended to use serum HER2 ECD level measurement as complementary of histological methods. The technique is rapid, cost effective and fully automated, thereby offering a high degree of standardization and reproducibility. Due to the correlation between serum HER2 ECD levels and HER2 expression in tumor tissue, patients could be considered for trastuzumab therapy even without tissue, just on the basis of high HER2 ECD levels. It has been indicated that a certain percentage of primary breast cancers with negative tissue HER2 status presented abnormal HER2 ECD levels in serum [22] . In our experiment, some cases with negative tissue HER2 also showed high serum HER2 ECD concentrations. Ardavanis et al. found that the HER2 tissue negative but serum positive patients can be benefited from trastuzumab therapy [23] . Thus, a potential utility of serum HER2 ECD is to identify patients who missed the opportunity of being treated with HER2-targeted therapies due to a lack of overexpression of HER2 in tumor tissues. Trastuzumab can bind to the HER2 ECD on the surface of tumor cells, eliciting an immune response via antibody-dependent cellular cytotoxicity, inhibiting receptor downstream signaling, inhibiting of tumor induced angiogenesis tumor cell repair after chemotherapy-induced and radiation-induced DNA damage [24–26] . A number of studies have monitored the serum HER2 ECD in response to HER2-tar[14–17]
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Research Article
3.0
Log HER2 ECD
2.5
HER2 negative HER2 positive
2.0 1.5 1.0 0.5 HER2 positive
HER2 negative
Figure 3. Serum HER2 extracellular domain concentration in metastatic gastric cancer patients tissue HER2 negative and positive. Tissue HER2 status and serum HER2 ECD level. ECD: Extracellular domain.
geted therapies such as trastuzumab and lapatinib in patients with breast cancer [20,27–28] . It can be anticipated that the change of serum HER2 ECD might be a new prognostic biomarker and predict response to HER2-targeted therapy in metastatic gastric cancer. Conclusion & future perspective The present study documented an association between serum HER2 ECD and tissue HER2 status in patients with metastatic gastric cancer. Determination of serum HER2 ECD concentration in metastatic gastric cancer may provide a useful alternative of tissue HER2 status, especially in light of its relaROC curve
1.0
Sensitivity
0.8 0.6 0.4 0.2 0.0 0.0
0.2
0.4
0.6
0.8
1.0
1-specificity Figure 4. ROC curve of serum HER2 extracellular domain concentration for tissue HER2-negative and -positive patients. The area under the curve for serum HER2 extracellular domain was 0.771 (95% CI: 0.682–0.860). Specificities and sensitivity for tissue HER2 positivity were 78.9 and 67.4%, respectively, at 10.65 ng/ml serum HER2 extracellular domain. ROC: Receiver operating characteristic.
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Table 2. Relationship between clinicopathological factors and serum HER2 extracellular domain levels. Characteristic
Patients, n (%)
Median serum HER2 (interquartile range), ng/ml
p-value
Age (years): – >60 – ≥60
73 (54.9) 60 (45.1)
9.60 (8.3–12.0) 9.65 (8.3–13.1)
0.643
Gender: – Male – Female
102 (76.7) 31 (23.3)
9.9 (8.5–13.1) 8.6 (7.6–10.2)
0.005
Primary site: – GEJ – Non-GEJ
8 (36.1) 85 (63.9)
9.85 (8.2–13) 9.6 (8.4–11.4)
0.568
Tumor type (Laurén classification): – Intestinal type – Diffuse or mixed
67 (50.4) 66 (49.6)
9.7 (8.8–14.2) 8.85 (7.7–10.8)
0.0048
Live metastases: – Yes – No
62 (46.6) 71 (53.4)
10.45 (9.0–14.2) 8.7 (7.7–11.0)
0.0008
Lung metastases: – Yes – No
23 (17.3) 110 (82.7)
13 (9.0–23.4) 9.3 (8.1–11.0)
0.0033
Metastases (n): – 1–2 – ≥3
67 (50.4) 66 (49.6)
9.6 (8.4–11.4) 9.7 (8.1–13.5)
0.6805
GEJ: Gastroesophageal junction.
tively easy, inexpensive and reproductive methodology and the simplicity of sample collection. Future study, especially prospective clinical trials to explore serum HER2 ECD in clinical applications such as determining the precise cutoff value, choosing suitable patients for anti-HER2 therapy and monitoring the treatment response, are needed.
dation (2013M530494). The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript apart from those disclosed. No writing assistance was utilized in the production of this manuscript.
Financial & competing interests disclosure
Ethical conduct of research
This work was supported by National Natural Science Foundation of China (No. 81172110), National High Technology Research and Development Program (No. 2012AA 02A 504), Beijing Municipal Science & Technology Commission Program (No. Z11110706730000), Beijing Natural Science Foundation (7142034) and China Postdoctoral Science Foun-
The authors state that they have obtained appropriate institutional review board approval or have followed the principles outlined in the Declaration of Helsinki for all human or animal experimental investigations. In addition, for investigations involving human subjects, informed consent has been obtained from the participants involved.
Executive summary Background • The correlation between serum HER2 extracellular domain (ECD) and tissue HER2 status in gastric cancer is still not well known.
Results • A significant difference was found in serum HER2 ECD levels between patients with HER2 IHC 3+ and those with 2+ and 0/1+. • There was a significant relationship between serum and tissue levels of HER2 protein.
Conclusion • This study suggests that the serum HER2 ECD assay can be considered as a potential alternative for tissue HER2 status in patients with metastatic gastric cancer.
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Serum HER2 extracellular domain in metastatic gastric cancer patients
combination predictor for poor prognosis in breast cancer patients. Clinical Cancer Res. 10(11), 3815–3824 (2004).
References Papers of special note have been highlighted as: • of interest; •• of considerable interest 1
Bang YJ. Advances in the management of HER2-positive advanced gastric and gastroesophageal junction cancer. J. Clin. Gastroenterol. 46(8), 637–648 (2012).
2
Ruschoff J, Hanna W, Bilous M et al. HER2 testing in gastric cancer: a practical approach. Modern Pathol. 25(5), 637–650 (2012).
•
Evaluation of HER2 testing in gastric cancer.
3
Bang YJ, Van Cutsem E, Feyereislova A et al. Trastuzumab in combination with chemotherapy versus chemotherapy alone for treatment of HER2-positive advanced gastric or gastro-oesophageal junction cancer (ToGA): a Phase III, open-label, randomised controlled trial. Lancet 376(9742), 687–697 (2010).
Research Article
13
Lam L, McAndrew N, Yee M, Fu T, Tchou JC, Zhang H. Challenges in the clinical utility of the serum test for HER2 ECD. Biochim. Biophys. Acta 1826(1), 199–208 (2012).
•
Major review of the relationship between serum HER2 and breast cancer.
14
Kono K, Naganuma H, Sekikawa T et al. Serum level of HER-2/neu in patients with gastric cancer: correlation with HER-2/neu overexpression in gastric carcinoma tissue. Tumour Biol. 21(3), 139–144 (2000).
15
Takehana T, Kunitomo K, Kono K et al. Status of c-erbB-2 in gastric adenocarcinoma: a comparative study of immunohistochemistry, fluorescence in situ hybridization and enzyme-linked immuno-sorbent assay. Int. J. Cancer 98(6), 833–837 (2002).
16
Tsigris C, Karayiannakis AJ, Syrigos KN et al. Clinical significance of soluble c-erbB-2 levels in the serum and urine of patients with gastric cancer. Anticancer Res. 22(5), 3061–3065 (2002).
•
Describes how entry into the trial was guided by centralized HER2 testing using both immunohistochemistry and FISH.
4
De Vita F, Giuliani F, Silvestris N et al. Current status of targeted therapies in advanced gastric cancer. Expert Opin. Ther. Targets 16(Suppl. 2), S29–S34 (2012).
17
5
Grabsch H, Sivakumar S, Gray S, Gabbert HE, Muller W. HER2 expression in gastric cancer: rare, heterogeneous and of no prognostic value – conclusions from 924 cases of two independent series. Cellular Oncol. 32(1–2), 57–65 (2010).
Narita T, Seshimo A, Suzuki M, Murata J, Kameoka S. Status of tissue expression and serum levels of HER2 in gastric cancer patients in Japan. Hepatogastroenterology 60(125), 1083–1088 (2013).
18
6
Warneke VS, Behrens HM, Boger C et al. Her2/neu testing in gastric cancer: evaluating the risk of sampling errors. Ann. Oncol. 24(3), 725–733 (2013).
Carlson RW, Moench SJ, Hammond ME et al. HER2 testing in breast cancer: NCCN Task Force report and recommendations. J. Natl Compr. Canc. Netw. 4(Suppl. 3), S1–S22; quiz S23–S24 (2006).
19
7
Fusco N, Rocco EG, Del Conte C et al. HER2 in gastric cancer: a digital image analysis in pre-neoplastic, primary and metastatic lesions. Modern Pathol. 26(6), 816–824 (2013).
Hayashi N, Nakamura S, Tokuda Y et al. Serum HER2 levels determined by two methods in patients with metastatic breast cancer. Int. J. Clin. Oncol. 17(1), 55–62 (2012).
20
Witzel I, Loibl S, Von Minckwitz G et al. Monitoring serum HER2 levels during neoadjuvant trastuzumab treatment within the GeparQuattro trial. Breast Cancer Res. Treat. 123(2), 437–445 (2010).
21
Sorensen PD, Jakobsen EH, Langkjer ST et al. Serum HER2 concentrations for monitoring women with breast cancer in a routine oncology setting. Clin. Chem. Lab. Med. 47(9), 1117–1123 (2009).
22
Krainer M, Brodowicz T, Zeillinger R et al. Tissue expression and serum levels of HER-2/neu in patients with breast cancer. Oncology 54(6), 475–481 (1997).
23
Ardavanis A, Kountourakis P, Kyriakou F et al. Trastuzumab plus paclitaxel or docetaxel in HER-2negative/HER-2 ECD-positive anthracycline- and taxane-refractory advanced breast cancer. Oncologist 13(4), 361–369 (2008).
24
Mimura K, Kono K, Hanawa M et al. Trastuzumabmediated antibody-dependent cellular cytotoxicity against esophageal squamous cell carcinoma. Clinical Cancer Res. 11(13), 4898–4904 (2005).
25
Pietras RJ, Pegram MD, Finn RS, Maneval DA, Slamon DJ. Remission of human breast cancer xenografts on therapy with humanized monoclonal antibody to HER-2 receptor and DNA-reactive drugs. Oncogene 17(17), 2235–2249 (1998).
8
Ross JS. Update on HER2 testing for breast and upper gastrointestinal tract cancers. Biomark. Med. 5(3), 307–318 (2011).
9
Leyland-Jones B, Smith BR. Serum HER2 testing in patients with HER2-positive breast cancer: the death knell tolls. Lancet Oncol. 12(3), 286–295 (2011).
•
Major review of the relationship between serum HER2 and breast cancer.
10
Liu PC, Liu X, Li Y et al. Identification of ADAM10 as a major source of HER2 ectodomain sheddase activity in HER2 overexpressing breast cancer cells. Cancer Biol. Ther. 5(6), 657–664 (2006).
11
Codony-Servat J, Albanell J, Lopez-Talavera JC, Arribas J, Baselga J. Cleavage of the HER2 ectodomain is a pervanadate-activable process that is inhibited by the tissue inhibitor of metalloproteases-1 in breast cancer cells. Cancer Res. 59(6), 1196–1201 (1999).
••
The mechanism of HER2 extracellular domain production.
12
Xia W, Chen JS, Zhou X et al. Phosphorylation/ cytoplasmic localization of p21Cip1/WAF1 is associated with HER2/neu overexpression and provides a novel
future science group
www.futuremedicine.com
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27
670
Pietras RJ, Poen JC, Gallardo D, Wongvipat PN, Lee HJ, Slamon DJ. Monoclonal antibody to HER-2/neu receptor modulates repair of radiation-induced DNA damage and enhances radiosensitivity of human breast cancer cells overexpressing this oncogene. Cancer Res. 59(6), 1347–1355 (1999). Lipton A, Leitzel K, Ali SM et al. Human epidermal growth factor receptor 2 (HER2) extracellular domain levels are
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associated with progression-free survival in patients with HER2-positive metastatic breast cancer receiving lapatinib monotherapy. Cancer 117(21), 5013–5020 (2011). 28
Witzel I, Loibl S, Von Minckwitz G et al. Predictive value of HER2 serum levels in patients treated with lapatinib or trastuzumab – a translational project in the neoadjuvant GeparQuinto trial. Br. J. Cancer 107(6), 956–960 (2012).
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Serum HER2 extracellular domain as a potential alternative for tissue HER2 status in metastatic gastric cancer patients
Aim: We investigated whether serum concentrations of the HER2 extracellular domain (ECD) can be used as an alternative to test tissue HER2 status in metastatic gastric cancer. Materials & methods: A total of 133 cases of metastatic gastric cancer were included in present study. Serum HER2 ECD was measured by chemiluminescence immunoassay (CLIA). Receiver operating characteristic curve analysis was used to determine optimal serum HER2 ECD concentrations for differentiation between positive and negative HER2 status. Results: The median level of serum HER2 ECD was 9.6 ng/ml in metastatic gastric cancer patients. There was a significant relationship between serum and tissue levels of HER2 protein (p < 0.001). Area under the curve for serum HER2 ECD was 0.771 (95% CI: 0.682–0.860). Conclusion: Levels of serum HER2 ECD are highly correlated with tissue HER2 status in metastatic gastric cancer. Serum HER2 ECD assay can be considered as a potential alternative for tissue HER2 status. Keywords: ECD • extracellular domain • HER2 • metastatic gastric cancer • serum
The human epidermal growth factor receptor-2 proto-oncogene, also known as HER2 and c-erbB-2, encoding a growth factor receptor, has been found to play an important role in gastric cancer [1] . HER2 has been shown to be overexpressed and amplified in 6–35% of gastric tumors [2] . The Phase III ToGA trial showed that adding the HER2targeted humanized monoclonal antibody trastuzumab to chemotherapy significantly improved survival of patients with advanced gastric cancer. Moreover, the study indicated that patients with immunohistochemistry (IHC) 2+/FISH+ and IHC 3+ (HER2 positive) had an increased benefit from trastuzumab treatment [3] . HER2-positive patients are eligible for HER2-targeted (trastuzumab) treatment, although the methods of assessment and prognostic value of HER2 have been subject to debate [4] . However, both the methods IHC and FISH for detecting HER2 status have limitations. One such limitation is the lack of ‘real-time’ follow-up due to the dependency of both assays on tumor biopsies. In many
10.2217/BMM.14.10 © 2014 Future Medicine Ltd
Zhi Peng1, Yi Liu2, Yanyan Li1, Xiaotian Zhang1, Jun Zhou1, Ming Lu1, Qingqing Li1 & Lin Shen*,1 Department of Gastrointestinal Oncology, Key Laboratory of Carcinogenesis & Translational Research (Ministry of Education), Peking University Cancer Hospital & Institute, Beijing, China 2 Translational Medicine Center, Laboratory of Oncology, Affiliated Hospital of Academy of Military Medical Sciences, Beijing, China *Author for correspondence Tel.: +86 10 8819 6561 Fax: +86 10 8819 6561 lin100@ medmail.com.cn 1
patients with metastatic gastric cancer, the re-evaluation of HER2 status by a tissue biopsy of the metastatic lesion is not feasible due to location of the metastatic site. Another important issue in HER2-targeted therapy is HER2 heterogeneity [5] . The heterogeneity may result in the inaccurate assessment of the HER2 status and the consequent rendering patients unsuitable for treatment of trastuzumab [6] . In addition, HER2 status may vary between different metastatic sites and also change during treatment [7] . New tests have been continuing development. To find more convenient, reproducible and accurate detection to identify patients with gastric cancer for anti-HER2 therapy is particularly urgent [8] . Serum extracellular domain (ECD) fragment of HER2 represents a noninvasive, relatively fast, reproducible and quantifiable biomarker that could supplement existing HER2 testing [9] . This extracellular fragment of the receptor is released from the surface of tumor cells into the circulation by the ADAM proteases [10,11] . It retains its sig-
Biomarkers Med. (2014) 8(5), 663–670
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Research Article Peng, Liu, Li et al. naling activity and, upon phosphorylation, can contribute to disease progression [12] . Studies have found elevated levels of HER2 ECD in the serum of many breast cancer patients, and in most cases, high levels of serum HER2 ECD are associated with higher relapse rates and worse prognosis [13] . The association between serum HER2 ECD and tissue HER2 status in gastric cancer patients has been investigated in a number of studies, too [14–17] . However, most of these studies included different groups of patients and used different methods. In the present study, we evaluated the correlation of HER2 tissue expression with serum levels of soluble HER2 ECD in patients with metastatic gastric cancer. Furthermore, we investigated whether serum HER2 ECD could be an alternative method to choose patients who are suitable for anti-HER2 therapy. Materials & methods Subjects
This retrospective study included unselected consecutive 133 patients who were diagnosed with metastatic gastric cancer at the Department of Gastroenterology Oncology in Peking University Cancer Hospital in China from February 2010 to May 2013. All patients were collected as following criteria: patients had histologically confirmed metastatic gastric cancer with serum and primary gastric tumor specimens available; patients were >18 years old. Their peripheral bloods before chemotherapy were drawn, after signed a written informed consents for their blood to be used for research. Demographic and clinicopathologic features such as age, gender, disease site, metastatic site and number of metastases were retrospectively obtained from the patient records. This study was approved by the medical ethics committee of Peking University Cancer Hospital and was performed according to the Declaration of Helsinki Principles. Tissue expression of HER2: IHC & FISH
Tumor biopsies of primary lesion were fixed in 10% neutral-buffered formalin overnight and embedded in paraffin blocks. Paraffin-embedded tissues were cut into 4-µm thick slices, and IHC was performed using HercepTest II™ (Dako, Glostrup, Denmark) according to manufacturer instructions. All IHC specimens were stained at the Department of Pathology, Peking University Cancer Hospital. The scoring criteria used in the ToGA trial were applied to give IHC scores for HER2 expression. Furthermore, FISH was performed with Abbott-Vysis PathVysion™ (Abbott Laboratories, IL, USA) according to manufacturer
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instructions if HER2 protein expression in IHC 2+ cases. Tumor specimens with a HER2:CEP17 signal ratio ≥2.0 were considered HER2 FISH positive. Serum HER2 levels using CLIA
Venous blood specimens were collected into separator tubes before treatment. Serum specimens were recovered by centrifuging blood specimens at 3000 rpm for 10 min and stored at -70°C. All samples stored in a standardized fashion and were detected at the same time. Serum HER2 levels were measured using ADVIA Centaur CP Immunoassay System (Siemens Healthcare Diagnostics, Erlangen, Germany) according to manufacturer instructions. Quality control was ensured by assaying the two levels of control sera supplied with the kit in each series. The intra-assay coefficient of variation (CV) was less than 6% and the interassay CV was 10%. Serum HER2 ECD measurement was carried out in a blinded manner without knowledge about the results of HER2 IHC and FISH. Statistical analysis
We used SAS software (version 8.0, SAS Institute Inc., NC, USA) for statistical analysis. The χ 2 test was used to assess the association between clinical and pathological features and the different levels of expression of HER2 ECD. Wilcoxon rank sum test or Kruskal–Wallis H test was used to compare the serum HER2 concentration in the patients with different tissue HER2 status. p-values less than 0.05 were considered statistically significant. To estimate sample size, we carried out a preliminary experiment first. We calculated that group sample sizes of 37 and 74 achieve 80% power to detect a difference of 0.4 between the null hypothesis that both group (HER2 positive and HER2 negative) means are 2.5 (log conversion) and the alternative hypothesis that the mean of HER2 negative group is 2.1 with estimated standard deviations of 0.8 and 0.4 and with a significance level (alpha) of 0.05 using a two-sided t wo-sample t-test. Based on the above calculation, over 111 samples are required to be involved in this study. Therefore, we choose the latest consecutive periods including over 111 patients to screen subjects. The receiver operating characteristic (ROC) curve was analyzed to evaluate the discriminating ability of serum HER2 and to arrive at a serum HER2 cutoff for predicting tissue HER2 positivity. To obtain appropriate serum level cutoff values, we calculated the total sensitivity and specificity for each cutoff value and then chose the cutoff values that maximized the sum of s ensitivity plus 1-specificity.
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Research Article
Table 1. Clinicopathologic characteristics and tissue HER2 status of 133 patients with metastatic gastric cancer. Characteristic
All patients, n (%)
HER2 positive, n (%)
HER2 negative, n (%)
Total
133 (100)
43 (32.3)
90 (67.7)
Age (years): – >60 – ≥60
73 (54.9) 60 (45.1)
28 (21.15) 15 (11.3)
45 (33.8) 45 (33.8)
Gender: – Male – Female
102 (76.7) 31 (23.3)
36 (27.1) 7 (5.3)
66 (49.6) 24 (18.0)
Primary site: – GEJ – Non-GEJ
48 (36.1) 85 (63.9)
13 (9.8) 30 (22.6)
35 (26.3) 55 (41.3)
Tumor type (Laurén classification): – Diffuse type 34 (25.6) – Intestinal type 67 (45.9) – Mixed type 32 (24.0)
10 (7.5) 23 (17.3) 10 (7.5)
24 (18.0) 44 (33.1) 22 (16.5)
Metastatic site: – Liver – Lung – Lymph node
62 (47.4) 23 (20.3) 96 (72.2)
17 (12.8) 12 (12.0) 27 (20.3)
45 (34.6) 11 (8.3) 69 (51.9)
Metastasis (n): –1 –2 – ≥3
15 (11.3) 52 (35.3) 66 (53.4)
6 (4.5) 16 (12.0) 21 (15.8)
9 (6.8) 36 (27.1) 45 (33.8)
GEJ: Gastroesophageal junction.
Results Patient characteristics
A total of 133 unselected consecutive patients (102 males and 31 females) with metastatic gastric cancer were enrolled in this study. Patient characteristics are summarized in Table 1. All patients were diagnosed as stage IV gastric cancer according to the American Joint Committee on Cancer (AJCC) 7th edition at the time of initial diagnosis. The median age was 59 years (range: 24–79 years). For the selection bias, we further analyzed the demographic and clinicopathologic features, no statistically differences were found in this study (data not shown). Therefore, it was considered that these subjects could represent the patients treated in our department. There were 66 IHC 1+/0 cases, 31 IHC 2+ cases and 36 IHC 3+ cases. Seven cases were HER2 gene amplification in IHC 2+ cases (Figure 1). According to ToGA trial criteria for HER2 positivity (IHC 3+ or IHC 2+ plus FISH+), 43 cases were positive of tissue HER2 expression and 90 cases were negative of tissue HER2 expression. Correlation between serum HER2 ECD levels & tissue HER2 status
The median serum HER2 level was 9.6 ng/ml in all metastatic gastric cancer patients (interquartile range:
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8.3–13.3 ng/ml; range: 4.4–350.0 ng/ml). Significant differences was found in serum HER2 ECD levels between patients with 3+ and those with 2+ and 0/1+ (χ2: 26.505; p < 0.0001) (Figure 2). Serum HER2 ECD levels were significantly higher in patients with 3+ (median: 13.3 ng/ml; interquartile range: 9.5–22.6 ng/ml; range: 6.9–350.0 ng/ml) than those in patients with 2+ (median: 10.1 ng/ml; interquartile range: 8.5–13.0 ng/ml; range: 4.8–23.7 ng/ml) and with 0/1+ (median: 8.9 ng/ml; interquartile: 7.5–10.3 ng/ml; range: 4.4–18.6 ng/ml). There was a significant relationship between serum and tissue levels of HER2 protein (p < 0.001). We defined IHC 3+ and IHC 2+ plus FISH positive as HER2 positive according to the ToGA trial. The HER2 ECD levels of HER2 positive (median: 13.0 ng/ml; interquartile range: 9.5–21.3 ng/ml; range: 5.5–350.0 ng/ml) were significantly higher than HER2 negative (median: 9.0 ng/ml; interquartile range: 7.7–10.3 ng/ml; range: 4.4–23.7 ng/ml; p < 0.0001) (Figure 3). The ROC curves plotted for serum HER2 ECD in patients with metastatic gastric cancer versus tissue status are shown in Figure 4. The area under the curve generated from the serum HER2 ECD was 0.771 (95% CI: 0.682–0.860) for all 133 patients with metastatic gastric cancer. The set point of serum HER2 ECD at 10.65 ng/ml to identify tissue HER2 status has
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A
B
Figure 1. HER2 gene amplification evaluated by FISH. (A) FISH-positive case; (B) FISH-negative case (magnification ×1000). Please see color figures online at www.futuremedicine.com/doi/pdf/10.2217/bmm.14.10.
a sensitivity of 67.4% and a specificity of 78.9%. ROC curve analysis showed 94% specificity and 40% sensitivity for tissue HER2 positivity at 15.0 ng/ml serum HER2 ECD. Relationships between serum HER2 ECD levels and clinicopathological variables are summarized in Table 2. Gender, Laurén classification, live metastases and lung metastases were significantly associated with serum HER2 levels. There was no association between serum HER2 ECD levels and age, primary tumor site and number of metastases. Discussion This study indicated a strong correlation between serum HER2 ECD concentration and tissue IHC
Log HER2 ECD
3.0 2.5
1+/0 2+ 3+
2.0 1.5 1.0 0.5 1+/0
2+ HER2 IHC
3+
Figure 2. Serum HER2 extracellular domain concentration according to tissue HER2 status in 133 patients with metastatic gastric cancer. IHC staining intensity and serum HER2 ECD level. Please see color figures online at www.futuremedicine. com/doi/pdf/10.2217/bmm.14.10. ECD: Extracellular domain; IHC: Immunohistochemistry.
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HER2 test results in metastatic gastric cancer, which suggested detection of serum HER2 ECD concentration can be a potential alternative way to choose patients for trastuzumab treatment. Serum HER2 ECD detection in breast cancer patients has been discovered in many studies. The elevated serum HER2 ECD levels are more frequently observed in metastatic breast cancers. The concentration of serum HER2 ECD is dynamic during disease progression and treatment. However, detection of serum HER2 ECD and establishment of its potential clinical usefulness is still under debate. Monitoring of HER2 ECD will not replace FISH/IHC assays right now but definitely will complement the tissue assay to offer a real-time picture of HER2/neu status in breast cancer patients [8,12] . There are two methods for detecting serum HER2 ECD approved in breast cancer for monitoring patients with metastatic disease: enzyme immunoassay (EIA; Bayer Immuno 1, Siemens Healthcare Diagnostics, Erlangen, Germany) and chemiluminescence immunoassay (CLIA, ADVIA Centaur System, Siemens Healthcare Diagnostics) [18] . Previously, serum HER2 ECD levels were usually determined by EIA. However, trastuzumab therapy influences EIA results since the therapeutic monoclonal antibody acts as a competitive inhibitor. CLIA was a more sensitive method than EIA for measuring serum HER2 ECD levels in patients who received trastuzumab [19] . Bayer Immuno 1 and ADVIA Centaur assays (used in automated systems) are approved for monitoring patients with metastatic disease. To date, only one study tested serum HER2 ECD levels in gastric cancer patients by CLIA [17] . In this study, CLIA was used to detect the serum HER2 ECD. To our knowledge, four articles have reported the serum HER2 ECD in gastric cancer patients
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Serum HER2 extracellular domain in metastatic gastric cancer patients
up to 1 May 2013. However, the relationship between serum and tissue levels of HER2 remains controversial. Three of them detect serum HER2 levels in gastric cancer patients by EIA [14–16] , one by CLIA [17] . Three studies are not consistent with our result, which showed no correlation with tissue HER2 status [15–17] . One explanation might be the different characteristics of included patients. All of the four studies used different stages (I–IV) of gastric cancer. This is the first study to compare the consistency of serum HER2 ECD and tissue HER2 status in patients with metastatic gastric cancer. The cutoff value (10.65 ng/ml) of serum HER2 ECD in present study was lower than in other studies (15 ng/ml) [20,21] . However, in Witzel’s study, the cutoff value of HER2 ECD equal to 10 ng/ml led to better sensitivity and specificity instead of 15 ng/ml [20] . Moreover, the calculated cutoff value of the serum HER2 assay used in our study might be appropriate for Chinese patients with metastatic gastric cancer in general. Different cutoff values need to be established for each population for specific disease from the prospective randomized trial. Because of higher specificity and sensitivity of CLIA test relative to tissue HER2 status, it is recommended to use serum HER2 ECD level measurement as complementary of histological methods. The technique is rapid, cost effective and fully automated, thereby offering a high degree of standardization and reproducibility. Due to the correlation between serum HER2 ECD levels and HER2 expression in tumor tissue, patients could be considered for trastuzumab therapy even without tissue, just on the basis of high HER2 ECD levels. It has been indicated that a certain percentage of primary breast cancers with negative tissue HER2 status presented abnormal HER2 ECD levels in serum [22] . In our experiment, some cases with negative tissue HER2 also showed high serum HER2 ECD concentrations. Ardavanis et al. found that the HER2 tissue negative but serum positive patients can be benefited from trastuzumab therapy [23] . Thus, a potential utility of serum HER2 ECD is to identify patients who missed the opportunity of being treated with HER2-targeted therapies due to a lack of overexpression of HER2 in tumor tissues. Trastuzumab can bind to the HER2 ECD on the surface of tumor cells, eliciting an immune response via antibody-dependent cellular cytotoxicity, inhibiting receptor downstream signaling, inhibiting of tumor induced angiogenesis tumor cell repair after chemotherapy-induced and radiation-induced DNA damage [24–26] . A number of studies have monitored the serum HER2 ECD in response to HER2-tar[14–17]
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Research Article
3.0
Log HER2 ECD
2.5
HER2 negative HER2 positive
2.0 1.5 1.0 0.5 HER2 positive
HER2 negative
Figure 3. Serum HER2 extracellular domain concentration in metastatic gastric cancer patients tissue HER2 negative and positive. Tissue HER2 status and serum HER2 ECD level. ECD: Extracellular domain.
geted therapies such as trastuzumab and lapatinib in patients with breast cancer [20,27–28] . It can be anticipated that the change of serum HER2 ECD might be a new prognostic biomarker and predict response to HER2-targeted therapy in metastatic gastric cancer. Conclusion & future perspective The present study documented an association between serum HER2 ECD and tissue HER2 status in patients with metastatic gastric cancer. Determination of serum HER2 ECD concentration in metastatic gastric cancer may provide a useful alternative of tissue HER2 status, especially in light of its relaROC curve
1.0
Sensitivity
0.8 0.6 0.4 0.2 0.0 0.0
0.2
0.4
0.6
0.8
1.0
1-specificity Figure 4. ROC curve of serum HER2 extracellular domain concentration for tissue HER2-negative and -positive patients. The area under the curve for serum HER2 extracellular domain was 0.771 (95% CI: 0.682–0.860). Specificities and sensitivity for tissue HER2 positivity were 78.9 and 67.4%, respectively, at 10.65 ng/ml serum HER2 extracellular domain. ROC: Receiver operating characteristic.
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Table 2. Relationship between clinicopathological factors and serum HER2 extracellular domain levels. Characteristic
Patients, n (%)
Median serum HER2 (interquartile range), ng/ml
p-value
Age (years): – >60 – ≥60
73 (54.9) 60 (45.1)
9.60 (8.3–12.0) 9.65 (8.3–13.1)
0.643
Gender: – Male – Female
102 (76.7) 31 (23.3)
9.9 (8.5–13.1) 8.6 (7.6–10.2)
0.005
Primary site: – GEJ – Non-GEJ
8 (36.1) 85 (63.9)
9.85 (8.2–13) 9.6 (8.4–11.4)
0.568
Tumor type (Laurén classification): – Intestinal type – Diffuse or mixed
67 (50.4) 66 (49.6)
9.7 (8.8–14.2) 8.85 (7.7–10.8)
0.0048
Live metastases: – Yes – No
62 (46.6) 71 (53.4)
10.45 (9.0–14.2) 8.7 (7.7–11.0)
0.0008
Lung metastases: – Yes – No
23 (17.3) 110 (82.7)
13 (9.0–23.4) 9.3 (8.1–11.0)
0.0033
Metastases (n): – 1–2 – ≥3
67 (50.4) 66 (49.6)
9.6 (8.4–11.4) 9.7 (8.1–13.5)
0.6805
GEJ: Gastroesophageal junction.
tively easy, inexpensive and reproductive methodology and the simplicity of sample collection. Future study, especially prospective clinical trials to explore serum HER2 ECD in clinical applications such as determining the precise cutoff value, choosing suitable patients for anti-HER2 therapy and monitoring the treatment response, are needed.
dation (2013M530494). The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript apart from those disclosed. No writing assistance was utilized in the production of this manuscript.
Financial & competing interests disclosure
Ethical conduct of research
This work was supported by National Natural Science Foundation of China (No. 81172110), National High Technology Research and Development Program (No. 2012AA 02A 504), Beijing Municipal Science & Technology Commission Program (No. Z11110706730000), Beijing Natural Science Foundation (7142034) and China Postdoctoral Science Foun-
The authors state that they have obtained appropriate institutional review board approval or have followed the principles outlined in the Declaration of Helsinki for all human or animal experimental investigations. In addition, for investigations involving human subjects, informed consent has been obtained from the participants involved.
Executive summary Background • The correlation between serum HER2 extracellular domain (ECD) and tissue HER2 status in gastric cancer is still not well known.
Results • A significant difference was found in serum HER2 ECD levels between patients with HER2 IHC 3+ and those with 2+ and 0/1+. • There was a significant relationship between serum and tissue levels of HER2 protein.
Conclusion • This study suggests that the serum HER2 ECD assay can be considered as a potential alternative for tissue HER2 status in patients with metastatic gastric cancer.
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Serum HER2 extracellular domain in metastatic gastric cancer patients
combination predictor for poor prognosis in breast cancer patients. Clinical Cancer Res. 10(11), 3815–3824 (2004).
References Papers of special note have been highlighted as: • of interest; •• of considerable interest 1
Bang YJ. Advances in the management of HER2-positive advanced gastric and gastroesophageal junction cancer. J. Clin. Gastroenterol. 46(8), 637–648 (2012).
2
Ruschoff J, Hanna W, Bilous M et al. HER2 testing in gastric cancer: a practical approach. Modern Pathol. 25(5), 637–650 (2012).
•
Evaluation of HER2 testing in gastric cancer.
3
Bang YJ, Van Cutsem E, Feyereislova A et al. Trastuzumab in combination with chemotherapy versus chemotherapy alone for treatment of HER2-positive advanced gastric or gastro-oesophageal junction cancer (ToGA): a Phase III, open-label, randomised controlled trial. Lancet 376(9742), 687–697 (2010).
Research Article
13
Lam L, McAndrew N, Yee M, Fu T, Tchou JC, Zhang H. Challenges in the clinical utility of the serum test for HER2 ECD. Biochim. Biophys. Acta 1826(1), 199–208 (2012).
•
Major review of the relationship between serum HER2 and breast cancer.
14
Kono K, Naganuma H, Sekikawa T et al. Serum level of HER-2/neu in patients with gastric cancer: correlation with HER-2/neu overexpression in gastric carcinoma tissue. Tumour Biol. 21(3), 139–144 (2000).
15
Takehana T, Kunitomo K, Kono K et al. Status of c-erbB-2 in gastric adenocarcinoma: a comparative study of immunohistochemistry, fluorescence in situ hybridization and enzyme-linked immuno-sorbent assay. Int. J. Cancer 98(6), 833–837 (2002).
16
Tsigris C, Karayiannakis AJ, Syrigos KN et al. Clinical significance of soluble c-erbB-2 levels in the serum and urine of patients with gastric cancer. Anticancer Res. 22(5), 3061–3065 (2002).
•
Describes how entry into the trial was guided by centralized HER2 testing using both immunohistochemistry and FISH.
4
De Vita F, Giuliani F, Silvestris N et al. Current status of targeted therapies in advanced gastric cancer. Expert Opin. Ther. Targets 16(Suppl. 2), S29–S34 (2012).
17
5
Grabsch H, Sivakumar S, Gray S, Gabbert HE, Muller W. HER2 expression in gastric cancer: rare, heterogeneous and of no prognostic value – conclusions from 924 cases of two independent series. Cellular Oncol. 32(1–2), 57–65 (2010).
Narita T, Seshimo A, Suzuki M, Murata J, Kameoka S. Status of tissue expression and serum levels of HER2 in gastric cancer patients in Japan. Hepatogastroenterology 60(125), 1083–1088 (2013).
18
6
Warneke VS, Behrens HM, Boger C et al. Her2/neu testing in gastric cancer: evaluating the risk of sampling errors. Ann. Oncol. 24(3), 725–733 (2013).
Carlson RW, Moench SJ, Hammond ME et al. HER2 testing in breast cancer: NCCN Task Force report and recommendations. J. Natl Compr. Canc. Netw. 4(Suppl. 3), S1–S22; quiz S23–S24 (2006).
19
7
Fusco N, Rocco EG, Del Conte C et al. HER2 in gastric cancer: a digital image analysis in pre-neoplastic, primary and metastatic lesions. Modern Pathol. 26(6), 816–824 (2013).
Hayashi N, Nakamura S, Tokuda Y et al. Serum HER2 levels determined by two methods in patients with metastatic breast cancer. Int. J. Clin. Oncol. 17(1), 55–62 (2012).
20
Witzel I, Loibl S, Von Minckwitz G et al. Monitoring serum HER2 levels during neoadjuvant trastuzumab treatment within the GeparQuattro trial. Breast Cancer Res. Treat. 123(2), 437–445 (2010).
21
Sorensen PD, Jakobsen EH, Langkjer ST et al. Serum HER2 concentrations for monitoring women with breast cancer in a routine oncology setting. Clin. Chem. Lab. Med. 47(9), 1117–1123 (2009).
22
Krainer M, Brodowicz T, Zeillinger R et al. Tissue expression and serum levels of HER-2/neu in patients with breast cancer. Oncology 54(6), 475–481 (1997).
23
Ardavanis A, Kountourakis P, Kyriakou F et al. Trastuzumab plus paclitaxel or docetaxel in HER-2negative/HER-2 ECD-positive anthracycline- and taxane-refractory advanced breast cancer. Oncologist 13(4), 361–369 (2008).
24
Mimura K, Kono K, Hanawa M et al. Trastuzumabmediated antibody-dependent cellular cytotoxicity against esophageal squamous cell carcinoma. Clinical Cancer Res. 11(13), 4898–4904 (2005).
25
Pietras RJ, Pegram MD, Finn RS, Maneval DA, Slamon DJ. Remission of human breast cancer xenografts on therapy with humanized monoclonal antibody to HER-2 receptor and DNA-reactive drugs. Oncogene 17(17), 2235–2249 (1998).
8
Ross JS. Update on HER2 testing for breast and upper gastrointestinal tract cancers. Biomark. Med. 5(3), 307–318 (2011).
9
Leyland-Jones B, Smith BR. Serum HER2 testing in patients with HER2-positive breast cancer: the death knell tolls. Lancet Oncol. 12(3), 286–295 (2011).
•
Major review of the relationship between serum HER2 and breast cancer.
10
Liu PC, Liu X, Li Y et al. Identification of ADAM10 as a major source of HER2 ectodomain sheddase activity in HER2 overexpressing breast cancer cells. Cancer Biol. Ther. 5(6), 657–664 (2006).
11
Codony-Servat J, Albanell J, Lopez-Talavera JC, Arribas J, Baselga J. Cleavage of the HER2 ectodomain is a pervanadate-activable process that is inhibited by the tissue inhibitor of metalloproteases-1 in breast cancer cells. Cancer Res. 59(6), 1196–1201 (1999).
••
The mechanism of HER2 extracellular domain production.
12
Xia W, Chen JS, Zhou X et al. Phosphorylation/ cytoplasmic localization of p21Cip1/WAF1 is associated with HER2/neu overexpression and provides a novel
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www.futuremedicine.com
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Research Article Peng, Liu, Li et al. 26
27
670
Pietras RJ, Poen JC, Gallardo D, Wongvipat PN, Lee HJ, Slamon DJ. Monoclonal antibody to HER-2/neu receptor modulates repair of radiation-induced DNA damage and enhances radiosensitivity of human breast cancer cells overexpressing this oncogene. Cancer Res. 59(6), 1347–1355 (1999). Lipton A, Leitzel K, Ali SM et al. Human epidermal growth factor receptor 2 (HER2) extracellular domain levels are
Biomarkers Med. (2014) 8(5)
associated with progression-free survival in patients with HER2-positive metastatic breast cancer receiving lapatinib monotherapy. Cancer 117(21), 5013–5020 (2011). 28
Witzel I, Loibl S, Von Minckwitz G et al. Predictive value of HER2 serum levels in patients treated with lapatinib or trastuzumab – a translational project in the neoadjuvant GeparQuinto trial. Br. J. Cancer 107(6), 956–960 (2012).
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