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Specificity of HercepTest in Determining HER-2/neu Status of Breast Cancers Using the United States Food and Drug Administration–Approved Scoring System

From the Department of Pathology, Beth Israel Deaconess Medical Center and Harvard Medical School, Boston, MA; and PhenoPath Laboratories and IRIS, Seattle, WA.

Address reprint requests to Stuart J. Schnitt, MD, Department of Pathology, Beth Israel Deaconess Medical Center–East Campus, 330 Brookline Ave, Boston, MA 02215; email sschnitt{at}caregroup harvard.edu.

ABSTRACT

PURPOSE: To evaluate the specificity of the HercepTest for Immunoenzymatic Staining (Dako Corp, Carpinteria, CA) for determining HER-2/neu protein expression in breast cancer.

MATERIALS AND METHODS: Forty-eight invasive breast cancers previously found to be HER-2/neu–negative by two different immunohistochemical (IHC) assays and not amplified for the HER-2/neu gene by fluorescence in situ hybridization were studied using the HercepTest kit. HercepTest was performed according to the manufacturer's guidelines, and the results were scored on a 0 to 3+ scale using the United States Food and Drug Administration (FDA)–approved grading system. In this system, cases scored as 2+ or 3+ are considered HER-2/neu–positive.

RESULTS: Among these 48 cases, the IHC score using the FDA-approved scoring system was 0 in four cases (8.3%), 1+ in 16 (33.3%), 2+ in 21 (43.8%), and 3+ in seven (14.6%). Therefore, 58.4% of these cases were categorized as HER-2/neu–positive, and the specificity of the HercepTest kit for HER-2/neu expression was 41.6%. However, with the use of a modified scoring system that took into account the level of staining of nonneoplastic epithelium, the specificity increased to 93.2%.

CONCLUSION: Our results indicate that the HercepTest kit, when used in accordance with the manufacturer's guidelines and the FDA-approved scoring system, results in a large proportion of breast cancers being categorized as positive for HER-2/neu protein expression and that many of these seem to be false-positives. Consideration of the level of staining of nonneoplastic epithelium resulted in improved specificity. The current FDA-approved scoring system for HercepTest results should be reevaluated before its widespread use in clinical practice.

THE HER-2/neu (c-erbB-2) oncogene encodes a 185-kda transmembrane protein (p185), which is overexpressed in 20% to 30% of invasive breast carcinomas.^1,2 Since 1987, when Slamon et al¹ first reported a significant relationship between amplification of the HER-2/neu oncogene and poor clinical outcome in breast cancer patients, numerous studies have examined the utility of HER-2/neu as a prognostic factor. Recent evidence also supports a role for HER-2/neu status of breast cancers as predictive of their sensitivity or resistance to various forms of systemic therapy. Most recently, HER-2/neu protein expression has been used to select patients for treatment with trastuzumab (Herceptin, Genentech, Inc, South San Francisco, CA), a monoclonal antibody to the HER-2/neu protein. Initial clinical trials have indicated that this therapy may be useful in prolonging the survival of patients with advanced, metastatic breast carcinoma.^3-5 Several studies have also indicated that tumors that overexpress HER-2/neu may show resistance to certain forms of chemotherapy (such as cyclophosphamide/methotrexate)^6-11 and sensitivity to others (such as doxorubicin).^12-15 Furthermore, some clinical studies have suggested that HER-2/neu overexpression is predictive of resistance to tamoxifen.^11,16-19

Therefore, analysis of the HER-2/neu status of breast cancer specimens is assuming increasing clinical relevance. Immunohistochemistry (IHC) is commonly used for evaluating HER-2/neu protein expression on formalin-fixed, paraffin-embedded samples of breast cancer.^20-22 However, given that various assay protocols, HER-2/neu antibodies, and scoring systems are currently in use, variability in HER-2/neu IHC results has become a matter of legitimate concern.^23-27 A standardized IHC kit for the evaluation of HER-2/neu protein expression (HercepTest for Immunoenzymatic Staining, Dako Corp, Carpinteria, CA) has recently been approved by the United States Food and Drug Administration (FDA). Of note, this release coincided with the FDA's approval of trastuzumab.²⁸ As a result of these developments, there is now great interest among both clinicians and pathologists in evaluating the ability of the HercepTest assay to accurately determine the HER-2/neu status of breast cancers. The specificity of the assay is of particular concern, because low specificity, manifested as a large number of false-positive results, could result in inappropriate use of trastuzumab.

The purpose of this study was to evaluate the HercepTest kit in a series of breast cancers previously shown to be negative for HER-2/neu protein expression by two other IHC assays and nonamplified for the HER-2/neu gene by fluorescence in situ hybridization (FISH).

MATERIALS AND METHODS

Study Design
We previously studied 100 consecutive cases of invasive breast cancer for HER-2/neu protein overexpression using two IHC methods and for HER-2/neu gene amplification by FISH.^29,30 Of note, the two prior IHC assays employed different methodology and scoring systems but used the same primary antibody (Dako rabbit antihuman polyclonal antibody to c-erbB-2 oncoprotein, code number A0485; Dako Corp),^29,30 which is a concentrate of the same anti–HER-2/neu antibody provided in prediluted form in the HercepTest kit. In these prior analyses, 22.9% of the cases showed HER-2/neu protein overexpression by one of the IHC methods, 23.7% showed HER-2/neu overexpression by the second IHC method, and 25.8% showed HER-2/neu gene amplification by FISH. Among these 100 cases, there was sufficient tissue remaining in the paraffin block for further IHC analysis in 48 cases that lacked HER-2/neu overexpression by both IHC assays and lacked HER-2/neu gene amplification by FISH. These 48 cases constitute the population for this study. All cases had been accessioned at Beth Israel Deaconess Medical Center (BIDMC), Boston, MA, between July 24, 1997, and February 18, 1998. The tissue from these cases was fixed initially in alcoholic formalin (Anatech, Ltd, Battle Creek, MI) followed by fixation in 10% neutral buffered formalin. For each case, 4-µm thick tissue sections were cut from a representative paraffin block and applied to positively charged slides.

In the first of the prior IHC assays, performed at PhenoPath Laboratories (PPL), Seattle, WA, tissue sections were subjected to heat-induced epitope retrieval (HIER) by immersion in citrate buffer (pH 6.0) preheated to greater than 90°C and heating in a Black & Decker vegetable steamer (Black & Decker Corp, Towson, MD) for 20 minutes before incubation with the anti–HER-2/neu antibody on a Dako Autostainer. Primary antibody was localized using the LSAB+ Detection System (labeled streptavidin biotin immunoperoxidase; Dako Corp) according to the manufacturer's instructions using the Dako Autostainer. Membrane staining intensity and pattern were evaluated using a 0 to 4+ scale (0, completely negative; 1+, faint membranous positivity; 2+, moderate membranous positivity; 3+, strong, circumferential membranous positivity; and 4+, extremely strong, circumferential membranous positivity). For a score of 2+ to 4+, membrane staining in the majority of the tumor cells was required to be present. Cytoplasmic immunostaining was noted but not incorporated into the final scoring. For each case, infiltrating carcinoma and adjacent normal epithelium (if available) were separately scored. A final subtracted score of the tumor minus normal epithelium was used to correct for variability in background staining of normal epithelium (which should not overexpress the HER-2/neu protein). Either a final subtracted score of 2 or tumor cell staining of 3+ or greater was required to categorize a case as HER-2/neu–positive.

In the second of the previous IHC assays, performed at BIDMC, tissue sections were subjected to HIER by heating in a microwave oven in citrate buffer (pH = 6) for a total of 10 minutes before immunostaining using the Ventana 320 automated immunostainer (Ventana Medical Systems, Tucson, AZ). The primary antibody to the HER-2/neu oncoprotein was used at a 1:500 dilution, and diaminobenzidine (DAB, Sigma Chemicals, St. Louis, MO) was used as the chromogen. HER-2/neu staining was considered positive when the tumor cells showed intense circumferential cell membrane staining, easily identified with a 10x objective. In all of these cases, staining was observed in the majority (> 50%) of the tumor cells. Tumors in which there was cytoplasmic staining without distinct cell membrane staining were scored as negative.

All 48 cases had also been previously analyzed for HER-2/neu gene amplification using the Oncor/Ventana INFORM HER-2/neu Gene Detection System (Ventana Medical Systems; formerly sold by Oncor, Inc, Gaithersburg, MD) at BIDMC in a laboratory certified by Oncor as proficient in the procedure. The methodology and interpretation were in accordance with the guide accompanying the kit³¹ as previously described.²⁹ Briefly, tissue sections were digested with proteinase, denatured, and hybridized with Oncor biotinylated HER-2/neu DNA probe. Oncor Fluorescein-Labeled Avidin Detection Reagent and Oncor Anti-Avidin Antibody were used for probe detection. Nuclei were counterstained with 4'-6'-diamidino-2'-phenylindole (DAPI)/Antifade. Slides were examined using a fluorescence microscope. Twenty randomly selected invasive tumor cell nuclei in each of two separate, distinct microscopic areas were evaluated for HER-2/neu gene copy number (ie, a total of 40 nuclei per case). Cases were scored as amplified by FISH when the mean number of fluorescent signals per nucleus was greater than four.

HercepTest IHC Assay
In this study, HER-2/neu protein expression was evaluated using the HercepTest for Immunoenzymatic Staining at PPL according to the protocol described in the manufacturer's guide accompanying the kit. Tissue sections were deparaffinized in two 5-minute changes of xylene and were rehydrated through alcohols to distilled water. Subsequently, sections were subjected to HIER by immersing the slides in Dako Epitope Retrieval Solution (0.01 mol/L citrate buffer; pH = 6) preheated to 95°C, and then heated in waterbath at 95°C for a total of 40 minutes, followed by a 20-minute cooldown period at room temperature. Slides were incubated with the primary rabbit polyclonal antibody to the HER-2/neu oncoprotein (as supplied prediluted in the HercepTest kit) on a Dako Autostainer for 30 minutes at room temperature. Antibody was localized by incubating slides with the Dako Visualization Reagent (dextran polymer conjugated with horseradish peroxidase and goat antirabbit immunoglobulins) for 30 minutes using the Dako Autostainer. Diaminobenzidine (DAB) was used as the chromogen, and the sections were counterstained with hematoxylin. Positive controls were included in each staining run and consisted of freshly cut breast cancer cases known to express HER-2/neu and a control slide consisting of three pelleted, formalin-fixed, paraffin-embedded human breast cell lines with staining intensity scores of 0, 1+, and 3+ (supplied in the HercepTest kit). Negative controls consisted of substituting normal rabbit serum (Dako Negative Control Reagent) for the HER-2/neu primary antibody. Only membrane staining intensity and pattern were evaluated using the 0 to 3+ scale as illustrated in the HercepTest kit scoring guidelines. As defined in the HercepTest kit guide, scores of 0 or 1+ were considered negative for HER-2/neu overexpression, 2+ was weak positive, and 3+ was strong positive. To qualify for 2+ and 3+ scoring (ie, positive), complete membrane staining of more than 10% of tumor cells had to be observed.

We also used a modification of this scoring system that took into consideration the level of staining of nonneoplastic epithelium present on the same slide as the cancer. In this system, nonneoplastic epithelium was also graded on a 0 to 3+ scale using the same criteria used for assessment of tumor cell staining. Cases were considered HER-2/neu positive only when the difference between the tumor cell staining score and the nonneoplastic epithelial cell staining score was 2.

Calculation of HercepTest Assay Specificity
Specificity of the HercepTest was defined as the number of true-negative cases (ie, cases that were negative for HER-2/neu protein expression by HercepTest that were also negative for HER-2/neu protein expression by both prior IHC assays and negative for HER-2/neu gene amplification by the FISH assay) divided by the total number cases that were HER-2/neu–negative by both prior IHC assays and by the FISH assay (ie, true-negatives and false-positives by HercepTest). Specificity was expressed as a percentage.

RESULTS

Patient Data and Histologic Features of Carcinomas
The median age of the patients was 66 years (range, 36 to 89 years). Thirty-two of the 48 carcinomas (66.7%) were of infiltrating ductal type, seven (14.6%) were infiltrating lobular, four (8.3%) were invasive cancers with both ductal and lobular features, three (6.3%) were mucinous (colloid) carcinomas, and two (4.2%) were tubular carcinomas. The median size of the tumors was 15 mm (range, 6 to 90 mm). Histologic grading was performed using the Elston and Ellis³² modification of the Bloom-Richardson grading system. Sixteen of the 48 carcinomas (33.3%) were grade 1, 18 (37.5%) were grade 2, and 14 (29.2%) were grade 3. Twenty of the 48 patients (41.7%) were axillary lymph node-negative and 13 (27.1%) were node-positive. Fifteen patients did not undergo axillary lymph node dissection. Forty-one of the 48 cases (85.4%) were estrogen receptor (ER)-positive and six (12.5%) were ER-negative. ER status was not determined in one case (Table 1).

HER-2/neu Status
All 48 cases were negative for HER-2/neu protein expression by previous IHC assays at both PPL and BIDMC, and none were amplified for the HER-2/neu gene by FISH.^29,30 However, using the HercepTest IHC kit and the FDA-approved scoring system, 28 of these cases (58.4%) were interpreted as positive (score of 2+ or 3+), and 20 (41.6%) were interpreted as negative (score of 0 or 1+) (Table 2). Therefore, if the results of the three previous HER-2/neu assays performed on these cases are considered true-negative results, then under these circumstances, the specificity of the HercepTest kit for HER-2/neu protein expression was 41.6%.

In 44 of these cases, nonneoplastic epithelium was present on the same tissue sections as the cancer. The HercepTest score in the benign epithelium was 0 in five cases (11.4%), 1+ in 15 (34.1%), 2+ in 21 (47.7%), and 3+ in three (6.8%). The difference between the tumor cell score and the nonneoplastic epithelial cell score was 2 in only three cases, and these three cases were considered HER-2/neu–positive. Therefore, when the level of staining of nonneoplastic epithelium was taken into consideration, the specificity of the HercepTest increased to 93.2%.

DISCUSSION

In this study, the HercepTest kit, when used in accordance with the manufacturer's guidelines and FDA-approved scoring system, categorized as HER-2/neu–positive almost 60% of invasive breast cancers that were previously shown to lack both HER-2/neu protein expression and HER-2/neu gene amplification in prior assays. These findings are subject to a number of different interpretations. First, it could be argued that the results using the HercepTest kit accurately reflect the HER-2/neu protein expression status of these cases and that our prior results, in which these cases were categorized as HER-2/neu–negative, were incorrect. However, this interpretation seems unlikely for several reasons. First, the HER-2/neu positivity rate by IHC using the HercepTest kit in this selected series of cases was substantially higher than the 20% to 30% rate of positivity noted in unselected series of breast cancers reported in other studies.^1,2 Second, all of the cases in this study that were scored as positive using the HercepTest kit lacked HER-2/neu gene amplification as determined by a FISH assay. Although prior studies have clearly shown that breast cancers may exhibit HER-2/neu protein expression in the absence of gene amplification, this phenomenon has been observed in only 3% to 7% of cases.^33-35 Third, the HercepTest assay was performed and the results were scored strictly in accordance with the manufacturer's recommendations and FDA-approved scoring system. Therefore, neither technical nor interpretive deviations from the proscribed method are likely to explain these results.

An alternative explanation for our findings is that the HercepTest assay, when used according to the manufacturer's FDA-approved guidelines, has low specificity for the detection of HER-2/neu protein expression. This interpretation is in agreement with the recent findings of Roche and Ingle.³⁶ These investigators noted a HER-2/neu positivity rate of 54% using the HercepTest kit in 59 cases that were found to be HER-2/neu–negative using another IHC assay. It could be argued that this comparison is not entirely valid, because these authors compared results of the HercepTest assay to an assay that uses a different antibody (monoclonal antibody CB11). However, our prior negative IHC results on the cases evaluated in the current study were obtained using the same primary anti–HER-2/neu antibody supplied in the HercepTest kit. Therefore, it seems unlikely that the observed discrepancies in IHC results between the HercepTest and other IHC assays is related to the nature of the primary antibody alone. It is possible that other methodologic aspects of the assay contributed to the low specificity observed in the study of Roche and Ingle³⁶ and in our study. However, our results strongly suggest that the low specificity is in large part related to the use of the recommended scoring system, because a dramatic improvement in specificity was noted when the level of staining of nonneoplastic epithelium was taken into account in our modified scoring system.

Previous studies have highlighted a number of potential problems in the use of IHC assays for HER-2/neu, including variability in tissue fixation and processing, variable sensitivity and specificity of commercially available antibodies, and differences in scoring criteria.^23-25 In our experience, variations in the type of fixative, length of tissue fixation, and details of tissue processing can result in differences in the intensity of specific staining for HER-2/neu in tumor cells as well as in variable levels of staining of nonneoplastic epithelium. In particular, fixatives that contain alcohol (including alcoholic formalin) can result in prominent staining of nonneoplastic epithelium in some cases. Any scoring system must, therefore, take into account the immutable fact that different fixation and processing protocols will be used in different laboratories. Our results suggest that consideration of the level of staining of nonneoplastic epithelium helps to "normalize" the level of HER-2/neu staining by serving as an internal control and may help to compensate for interlaboratory differences in tissue fixation and processing. However, one potential limitation of this approach is the lack of nonneoplastic epithelium in association with some primary tumors and in metastatic lesions.

The development of standardized methods for HER-2/neu IHC is clearly an important goal. However, our results suggest that the HercepTest kit, the first such proposed standardized assay, has low specificity for HER-2/neu protein expression when used in accordance with the manufacturer's guidelines and FDA-approved scoring system. Pathologists who perform assays for HER-2/neu and clinicians who use this information in formulating therapeutic recommendations need to be aware of these issues. In particular, the current FDA-approved scoring system for HercepTest should be re-evaluated before widespread use of the scoring system in clinical practice.

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Submitted February 8, 1999; accepted April 9, 1999.

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				G. Cornolti, M. Ungari, M. L. Morassi, F. Facchetti, E. Rossi, D. Lombardi, and P. Nicolai Amplification and Overexpression of HER2/neu Gene and HER2/neu Protein in Salivary Duct Carcinoma of the Parotid Gland Arch Otolaryngol Head Neck Surg, October 1, 2007; 133(10): 1031 - 1036. [Abstract] [Full Text] [PDF]

				A. Lee, W. C. Park, H. W. Yim, M. A. Lee, G. Park, and K. Y. Lee Expression of c-erbB2, cyclin D1 and Estrogen Receptor and their Clinical Implications in the Invasive Ductal Carcinoma of the Breast Jpn. J. Clin. Oncol., September 1, 2007; 37(9): 708 - 714. [Abstract] [Full Text] [PDF]

				A. Mavi, T. F. Cermik, M. Urhan, H. Puskulcu, S. Basu, J. Q. Yu, H. Zhuang, B. Czerniecki, and A. Alavi The Effects of Estrogen, Progesterone, and C-erbB-2 Receptor States on 18F-FDG Uptake of Primary Breast Cancer Lesions J. Nucl. Med., August 1, 2007; 48(8): 1266 - 1272. [Abstract] [Full Text] [PDF]

				L. Verlinden, I. Vanden Bempt, G. Eelen, M. Drijkoningen, I. Verlinden, K. Marchal, C. De Wolf-Peeters, M.-R. Christiaens, L. Michiels, R. Bouillon, et al. The E2F-Regulated Gene Chk1 Is Highly Expressed in Triple-Negative Estrogen Receptor /Progesterone Receptor /HER-2 Breast Carcinomas Cancer Res., July 15, 2007; 67(14): 6574 - 6581. [Abstract] [Full Text] [PDF]

				C. A. Hudis Trastuzumab -- Mechanism of Action and Use in Clinical Practice N. Engl. J. Med., July 5, 2007; 357(1): 39 - 51. [Full Text] [PDF]

				G Valabrega, F Montemurro, and M Aglietta Trastuzumab: mechanism of action, resistance and future perspectives in HER2-overexpressing breast cancer Ann. Onc., June 1, 2007; 18(6): 977 - 984. [Abstract] [Full Text] [PDF]

				C. Barrett, H. Magee, D. O'Toole, S. Daly, and M. Jeffers Amplification of the HER2 gene in breast cancers testing 2+ weak positive by HercepTest immunohistochemistry: false-positive or false-negative immunohistochemistry? J. Clin. Pathol., June 1, 2007; 60(6): 690 - 693. [Abstract] [Full Text] [PDF]

				K. L Henriksen, B. B Rasmussen, A. E Lykkesfeldt, S. Moller, B. Ejlertsen, and H. T Mouridsen Semi-quantitative scoring of potentially predictive markers for endocrine treatment of breast cancer: a comparison between whole sections and tissue microarrays J. Clin. Pathol., April 1, 2007; 60(4): 397 - 404. [Abstract] [Full Text] [PDF]

				L. Dal Lago, V. Durbecq, C. Desmedt, R. Salgado, T. Verjat, L. Lespagnard, Y. Ma, I. Veys, A. Di Leo, C. Sotiriou, et al. Correction for chromosome-17 is critical for the determination of true Her-2/neu gene amplification status in breast cancer. Mol. Cancer Ther., October 1, 2006; 5(10): 2572 - 2579. [Abstract] [Full Text] [PDF]

				S Banerjee, J S Reis-Filho, S Ashley, D Steele, A Ashworth, S R Lakhani, and I E Smith Basal-like breast carcinomas: clinical outcome and response to chemotherapy J. Clin. Pathol., July 1, 2006; 59(7): 729 - 735. [Abstract] [Full Text] [PDF]

				N. L. Henry and D. F. Hayes Uses and abuses of tumor markers in the diagnosis, monitoring, and treatment of primary and metastatic breast cancer. Oncologist, June 1, 2006; 11(6): 541 - 552. [Abstract] [Full Text] [PDF]

				M. S. Lyall, S. R. Dundas, S. Curran, and G. I. Murray Profiling Markers of Prognosis in Colorectal Cancer Clin. Cancer Res., February 15, 2006; 12(4): 1184 - 1191. [Abstract] [Full Text] [PDF]

				U. Vinatzer, B. Dampier, B. Streubel, M. Pacher, M. J. Seewald, C. Stratowa, K. Kaserer, and M. Schreiber Expression of HER2 and the Coamplified Genes GRB7 and MLN64 in Human Breast Cancer: Quantitative Real-time Reverse Transcription-PCR as a Diagnostic Alternative to Immunohistochemistry and Fluorescence In situ Hybridization Clin. Cancer Res., December 1, 2005; 11(23): 8348 - 8357. [Abstract] [Full Text] [PDF]

				E. Li-Ning-T, R. Ronchetti, C. Torres-Cabala, and M. J. Merino Role of Chromogenic in Situ Hybridization (CISHTM) in the Evaluation of HER2 Status in Breast Carcinoma: Comparison with Immunohistochemistry and Fish International Journal of Surgical Pathology, October 1, 2005; 13(4): 343 - 351. [Abstract] [PDF]

				A. Onn, D. H. Choe, R. S. Herbst, A. M. Correa, R. F. Munden, M. T. Truong, A. A. Vaporciyan, T. Isobe, M. Z. Gilcrease, and E. M. Marom Tumor Cavitation in Stage I Non-Small Cell Lung Cancer: Epidermal Growth Factor Receptor Expression and Prediction of Poor Outcome Radiology, October 1, 2005; 237(1): 342 - 347. [Abstract] [Full Text] [PDF]

				R Ainsworth, J M S Bartlett, J J Going, E A Mallon, A Forsyth, J Richmond, W Angerson, A Watters, and B Dunne IHC for Her2 with CBE356 antibody is a more accurate predictor of Her2 gene amplification by FISH than HercepTestTM in breast carcinoma J. Clin. Pathol., October 1, 2005; 58(10): 1086 - 1090. [Abstract] [Full Text] [PDF]

				M. F. Press, G. Sauter, L. Bernstein, I. E. Villalobos, M. Mirlacher, J.-Y. Zhou, R. Wardeh, Y.-T. Li, R. Guzman, Y. Ma, et al. Diagnostic Evaluation of HER-2 as a Molecular Target: An Assessment of Accuracy and Reproducibility of Laboratory Testing in Large, Prospective, Randomized Clinical Trials Clin. Cancer Res., September 15, 2005; 11(18): 6598 - 6607. [Abstract] [Full Text] [PDF]

				A. K. Witkiewicz, S. Varambally, R. Shen, R. Mehra, M. S. Sabel, D. Ghosh, A. M. Chinnaiyan, M. A. Rubin, and C. G. Kleer {alpha}-Methylacyl-CoA Racemase Protein Expression Is Associated with the Degree of Differentiation in Breast Cancer Using Quantitative Image Analysis Cancer Epidemiol. Biomarkers Prev., June 1, 2005; 14(6): 1418 - 1423. [Abstract] [Full Text] [PDF]

				Z-H Xian, S-H Zhang, W-M Cong, W-Q Wu, and M-C Wu Overexpression/amplification of HER-2/neu is uncommon in hepatocellular carcinoma J. Clin. Pathol., May 1, 2005; 58(5): 500 - 503. [Abstract] [Full Text] [PDF]

				Y. H. Kim, G. Ishii, K. Goto, K. Nagai, K. Tsuta, S. Shiono, J. Nitadori, T. Kodama, Y. Nishiwaki, and A. Ochiai Dominant Papillary Subtype Is a Significant Predictor of the Response to Gefitinib in Adenocarcinoma of the Lung Clin. Cancer Res., November 1, 2004; 10(21): 7311 - 7317. [Abstract] [Full Text] [PDF]

				H. Yaziji, T. S. Barry, and A. M. Gown HER-2 Testing in Breast Cancer--Reply JAMA, October 20, 2004; 292(15): 1818 - 1818. [Full Text] [PDF]

				R. R. Tubbs and D. G. Hicks HER-2 Testing in Breast Cancer JAMA, October 20, 2004; 292(15): 1817 - 1818. [Full Text] [PDF]

				G. Arpino, S. J. Green, D. C. Allred, D. Lew, S. Martino, C. K. Osborne, and R. M. Elledge HER-2 Amplification, HER-1 Expression, and Tamoxifen Response in Estrogen Receptor-Positive Metastatic Breast Cancer: A Southwest Oncology Group Study Clin. Cancer Res., September 1, 2004; 10(17): 5670 - 5676. [Abstract] [Full Text] [PDF]

				L. Zhu, L. W. C. Chow, W. T. Y. Loo, X.-Y. Guan, and M. Toi Her2/neu Expression Predicts the Response to Antiaromatase Neoadjuvant Therapy in Primary Breast Cancer: Subgroup Analysis from Celecoxib Antiaromatase Neoadjuvant Trial Clin. Cancer Res., July 15, 2004; 10(14): 4639 - 4644. [Abstract] [Full Text] [PDF]