LGCmain
Indian Journal of Pathology and Microbiology
Home About us Instructions Submission Subscribe Advertise Contact e-Alerts Ahead Of Print Login 
Users Online: 5708
Print this page  Email this page Bookmark this page Small font sizeDefault font sizeIncrease font size


 
  Table of Contents    
ORIGINAL ARTICLE  
Year : 2014  |  Volume : 57  |  Issue : 2  |  Page : 201-204
Inter-observer agreement in reporting HER 2 Neu protein over expression by immunohistochemistry


1 Department of Pathology, Sultan Qaboos University, Muscat, Oman
2 Public Health and Family Medicine, Sultan Qaboos University, Muscat, Oman

Click here for correspondence address and email

Date of Web Publication19-Jun-2014
 

   Abstract 

Introduction: HER 2 Neu protein overexpression and its detection by immunohistochemistry (IHC) has become quiet critical because of its relevance in regards to Herceptin treatment. This peer review was done at a tertiary care center, which aimed at determining the inter-observer variation among five pathologists and evaluating the degree of agreement between them. Aims: The aim of our study was to determine the reproducibility of HER 2 Neu system of reporting in breast cancer cases and determine inter-observer variability among five pathologists at a tertiary care center. To compare the results with similar studies done at other centers. Settings and Design: Retrospective descriptive study. Materials and Methods: Hematoxylin and Eosin (H and E) and IHC stained slides of 104 cases of carcinoma breast, on which HER 2 Neu status had been reported were reviewed. The time period for selection was from January 2010 to December 2011 (2 year period). Five pathologists reviewed the H and E and IHC slides independently and scored the results on a specially designed work sheet. Kappa values for inter-observer variation and Cornbach's alpha for internal consistency were calculated. Statistical Analysis Used: SPSS 20.0 (IBM). not known. Results: Complete agreement was seen between all five pathologists in 70 cases (70/104) = 67%. Agreement between four pathologists was seen in 78 cases (78/104) = 75%. Agreement between three pathologists was seen in 92 cases (92/104) = 88%. The global value for kappa co efficient for agreement between two pathologists was 0.706 and Cornbach alpha for internal consistency of reporting in the department was 0.987. Conclusion/Key Messages: Our departmental peer review indicated that there is good inter-observer concordance (agreement between two pathologists) and there is strong overall internal consistency of reporting for HER 2 Neu reporting by IHC. Our results are comparable to International reported data of similar studies.

Keywords: HER 2 Neu, immunohistochemistry, peer review

How to cite this article:
Al Haddabi I, Qureshi A, Saparamadu A, Al Hamdani A, Al Riyami M, Ganguly S. Inter-observer agreement in reporting HER 2 Neu protein over expression by immunohistochemistry. Indian J Pathol Microbiol 2014;57:201-4

How to cite this URL:
Al Haddabi I, Qureshi A, Saparamadu A, Al Hamdani A, Al Riyami M, Ganguly S. Inter-observer agreement in reporting HER 2 Neu protein over expression by immunohistochemistry. Indian J Pathol Microbiol [serial online] 2014 [cited 2022 Jan 20];57:201-4. Available from: https://www.ijpmonline.org/text.asp?2014/57/2/201/134663



   Introduction Top


The role of the pathologist in management of breast cancer cannot be undermined, now most of the breast units in the world are running a breast clinic where a tru-cut biopsy is done to perform estrogen and progesterone receptor and HER 2 Neu stains. [1],[2] HER 2 Neu belongs to the family of epidermal growth factors receptors for which protein overexpression is estimated by immunohistochemistry (IHC) and gene amplification is seen by florescent in situ hybridization (FISH). [3]

HER 2 Neu is a surface bound receptor present on the cell membrane involved in activation of path ways leading to cell growth and differentiation. [4] College of American Pathologists (CAP) recommends a set guideline for the interpretation of HER 2 Neu and American Society of Clinical Oncology outlines an algorithm for interpretation of Her 2 Neu by IHC. [5] In addition to its prognostic role, HER 2 Neu has become more important as a predictive marker to assess response to treatment by trastuzumab. [6] The response to therapy depends on accurate reporting of HER 2 Neu results. Many antibodies are used of which Food and Drug Administration approved Ventanna Hercept is the most reliable. [7] There is an acceptable range of inter-observer variability in HER 2 Neu reporting, which can be multifactorial. The determination of staining intensity and percentage of cells with complete membrane staining is subjective.

Departmental peer review is an important, ongoing process and should be carried out in all histopathology departments to see consistency of reporting between all reporting pathologists so as to improve reporting as it directly affects patient care. Audit and peer reviews are important components of departmental educational activity and they are conducted regularly to compare departmental standards to international data.

This peer review has particular relevance to patients receiving a specific chemotherapeutic agent if the patient is given 3+ score she will get the treatment if she is reported negative she will be denied that specific treatment. Thereby it is important that all reporting pathologists are aware of the recent guidelines of reporting.

This peer review was carried out in accordance with the same principle, and it has helped us to find out intradepartmental concordance and learn statistical methods of analyzing histopathology results.


   Materials AND METHODS Top


Hematoxylin and Eosin (H and E) and IHC stained Slides of 104 cases of carcinoma breast on which HER 2 Neu status had been reported were pulled out from the archives of the pathology department. The time period for selection was from January 2010 to December 2011 (2 year period). Ethical approval was acquired from the hospital ethics committee.

Five pathologists reviewed the H and E and IHC slides independently and scored the results on a specially designed work sheet. One senior resident (R5) was also involved.

She also looked at the cases independently and helped in the compilation of results. The pathologists initials were replaced by numbers thereby giving the numbers 1-5.

The cases were reported as complete negative (0) to strong positive (3+) using the college of American pathologists guideline CAP. [8]

SPSS version 20.0 was used to calculate the statistics. Kappa values for inter-observer variation (to see agreement between two pathologists), and Cornbach's alpha for internal consistency (to see overall agreement in the department) were calculated.

Only cases on which Her 2 Neu was done on primary invasive tumor were selected, cases where the test was done on metastatic disease or in situ cancer were excluded from the study.


   Results Top


A total of 104 cases were analyzed (seen by all five pathologists).

Out of 104 cases there were 70 (67%) cases with complete agreement of all five pathologists. These included, 41 cases in which score 0 was given by all five, there were 22 cases in which all five gave a score of three, six cases in which a score of two was given by all five and only one case which received a score of one by all five.

Agreement between four pathologists was seen in 78 cases (78/104) = 75%.

Agreement between three pathologists was seen in 92 cases (92/104) = 88%.

The major area of discrepancy was between 0 and 1 (12 cases where no majority agreement was achieved) and 2-3 (12 cases where no majority agreement was reached).



The degree of agreement between two pathologists was calculated using kappa coefficient.

[Table 1] maximum agreement (concordance) was seen between pathologist four and two (0.805) and the global value for all five of them was 0.706. This is considered a good agreement (concordance) (061-0.80 is a good score).
Table 1: Kappa coefficient

Click here to view


To see overall consistency of reporting in the department that is to see internal consistency between five pathologists together at one time Cornbach alpha [Table 2] was calculated which gave an overall score of 0.987. Which excellent overall consistency (max score is 1.000).
Table 2: Cornbach alpha

Click here to view



   Discussion Top


Inter observer variability is an accepted caveat in reporting HER 2 Neu by IHC and the area of focus is more problematic when it comes to 2+ and 3+. In general, human perception varies among individuals [8],[9] can be affected by training, experience, physical differences and fatigue. Observer variability is known to exist in other fields of medicine.

For reporting HER 2 neu in cases of breast cancer here it is critical to tell what is clear cut negative (0) and what is clear cut positive (3+).

There are 70 cases in our study where there is complete agreement, and these include 41 cases where there is the complete absence of staining so all of the pathologists gave a score of zero (clear cut negative) and there are 22 cases with clear cut thick membranous staining so the pathologists gave a score of 3+, on which there is 100% agreement. The major problematic area is score 2 and there were only 6 cases, on which all pathologists agreed to give a score of 2. In our institute, all the cases on which a score of 2+ is given are sent for FISH testing and if turn out to be positive by FISH receive Herceptin therapy.

We found frequent discrepancies between the scores of 0 and 1+ (12 cases). However, the differences between the scores of 0 and 1+ had no clinical significance. These are not considered for Herceptin therapy so no harm done to the patients.

Score 3 are the cases which will go for Herceptin treatment so as per our concordance we have very little disagreement. Those which were clear cut 3+ cases all the five pathologists had reported them as 3+.

On the contrary, there are 12 cases where there was major disagreement in all the pathologists to call these as 2 or 3+. Those which were finally labeled as 2+ they were sent for FISH analysis and of those only 4 came out as positive for gene amplification. These cases also did not miss the chance of getting Herceptin therapy. The kappa value for this is 0.561-0.738.

Hsu et al. published their data to compare results between five pathologists for 46 cases with a kappa concordance of 0.82-0.96 and for discordance between 2 and 3 there κ = 0.38. [10]

Lacroix-Triki et al. showed good concordance for score 3 κ = 0.81, also for 0/1 group κ = 0.85 and poor concordance for score 2 κ = 0.38. [10] These results are reflective of similar problems as seen in our study. [11]

Turashvili et al. combined visual method to automated image analysis "Using data from unequivocal positive (3+) or negative (0, 1+) results, both visual and automated scores were highly consistent: There was excellent concordance between two pathologists (κ = 1.000, 95% confidence interval [CI]: 1-1), between two machines (κ = 1.000, 95% CI: 1-1), and between both visual and both machine scores." [12]

Tsuda et al. also displayed excellent concordance when scoring for strong positive that is 3+ cases. They have also compared the results using multiple antibodies and two different envision detection systems. [13]

Our indices to ensure good agreement (concordance) between two pathologists which is kappa concordance shows that the best concordance is seen between pathologist 2 and 4 which is 0.806 and the weakest is seen between pathologist 1 and 3 which is 0.614 which is also in the good range of acceptance. Our global value is 0.706 which is in a good range of acceptance. [14]

In order to get consensus opinion, difficult cases should be reviewed on multi-head microscope. Any case which falls in a 2+ category, a second opinion can be sought from a fellow colleague. In any case if a case falls in 2+ category, it is sent for FISH analyses so that the patient does not miss the chance of getting Herceptin therapy. If the staining is weak or dubious the procedure can be repeated after checking the controls. [15]

As such there are no limitations of this study as the sample number is good and all the pathologists reviewed the cases independently, the only short coming is that this is a single center study so diversity of the sample is not taken into account (staining and reporting done at the same center).


   Conclusion Top


Departmental peer review is an important, ongoing process and should be carried out in all histopathology departments to see consistency of reporting between all reporting pathologists. Our peer review indicates that there is good inter-observer concordance (agreement between two pathologists) and there is strong overall internal consistency of reporting for HER 2 Neu reporting by IHC. Our results are comparable to International reported data of similar studies.

Furthermore all cases with 2+ staining pattern were sent for FISH analyses so that none of the patient misses a chance to get targeted therapy.

 
   References Top

1.Tímár J. Role of contemporary pathological diagnostics in the personalized treatment of cancer. Magy Onkol 2013;57:26-32.  Back to cited text no. 1
    
2.Manton DJ, Chaturvedi A, Hubbard A, Lind MJ, Lowry M, Maraveyas A, et al. Neoadjuvant chemotherapy in breast cancer: Early response prediction with quantitative MR imaging and spectroscopy. Br J Cancer 2006;94:427-35.  Back to cited text no. 2
    
3.Perez EA, Press MF, Dueck AC, Jenkins RB, Kim C, Chen B, et al. Immunohistochemistry and fluorescence in situ hybridization assessment of HER2 in clinical trials of adjuvant therapy for breast cancer (NCCTG N9831, BCIRG 006, and BCIRG 005). Breast Cancer Res Treat 2013;138:99-108.  Back to cited text no. 3
    
4.Chia YH, Ellis MJ, Ma CX. Neoadjuvant endocrine therapy in primary breast cancer: Indications and use as a research tool. Br J Cancer 2010;103:759-64.  Back to cited text no. 4
    
5.Perez EA, Romond EH, Suman VJ, Jeong JH, Davidson NE, Geyer CE Jr, et al. Four-year follow-up of trastuzumab plus adjuvant chemotherapy for operable human epidermal growth factor receptor 2-positive breast cancer: Joint analysis of data from NCCTG N9831 and NSABP B-31. J Clin Oncol 2011;29:3366-73.  Back to cited text no. 5
    
6.Criscitiello C, Metzger-Filho O, Saini KS, de Castro G Jr, Diaz M, La Gerche A, et al. Targeted therapies in breast cancer: Are heart and vessels also being targeted? Breast Cancer Res 2012;14:209.  Back to cited text no. 6
    
7.Kay EW, Walsh CJ, Cassidy M, Curran B, Leader M. C-erbB-2 immunostaining: Problems with interpretation. J Clin Pathol 1994;47:816-22.  Back to cited text no. 7
    
8.Wolff AC, Hammond ME, Schwartz JN, Hagerty KL, Allred DC, Cote RJ, et al. American Society of Clinical Oncology/College of American Pathologists guideline recommendations for human epidermal growth factor receptor 2 testing in breast cancer. Arch Pathol Lab Med 2007;131:18-43.  Back to cited text no. 8
    
9.Cell Markers And Cytogenetics Committees College Of American Pathologists. Clinical laboratory assays for HER-2/neu amplification and overexpression: Quality assurance, standardization, and proficiency testing. Arch Pathol Lab Med 2002;126:803-8.  Back to cited text no. 9
    
10.Hsu CY, Ho DM, Yang CF, Lai CR, Yu IT, Chiang H. Interobserver reproducibility of Her-2/neu protein overexpression in invasive breast carcinoma using the DAKO HercepTest. Am J Clin Pathol 2002;118:693-8.  Back to cited text no. 10
    
11.Lacroix-Triki M, Mathoulin-Pelissier S, Ghnassia JP, Macgrogan G, Vincent-Salomon A, Brouste V, et al. High inter-observer agreement in immunohistochemical evaluation of HER-2/neu expression in breast cancer: A multicentre GEFPICS study. Eur J Cancer 2006;42:2946-53.  Back to cited text no. 11
    
12.Turashvili G, Leung S, Turbin D, Montgomery K, Gilks B, West R, et al. Inter-observer reproducibility of HER2 immunohistochemical assessment and concordance with fluorescent in situ hybridization (FISH): Pathologist assessment compared to quantitative image analysis. BMC Cancer 2009;9:165.  Back to cited text no. 12
    
13.Tsuda H, Sasano H, Akiyama F, Kurosumi M, Hasegawa T, Osamura RY, et al. Evaluation of interobserver agreement in scoring immunohistochemical results of HER-2/neu (c-erbB-2) expression detected by HercepTest, Nichirei polyclonal antibody, CB11 and TAB250 in breast carcinoma. Pathol Int 2002;52:126-34.  Back to cited text no. 13
    
14.Cross S. Observer accuracy in estimating proportions in images: Implications for the semi quantitative assessment of staining reactions and a proposal for a new system. J Clin Pathol 2001;54:385-90.  Back to cited text no. 14
    
15.Schnitt SJ, Jacobs TW. Current status of HER2 testing: Caught between a rock and a hard place. Am J Clin Pathol 2001;116:806-10.  Back to cited text no. 15
    

Top
Correspondence Address:
Asim Qureshi
Sultan Qaboos University Hospital, Postal Code 123, Al Khod, Muscat
Oman
Login to access the Email id

Source of Support: None, Conflict of Interest: None


DOI: 10.4103/0377-4929.134663

Rights and Permissions



 
 
    Tables

  [Table 1], [Table 2]

This article has been cited by
1 The FDA-Approved Breast Cancer HER2 Evaluation Kit (HercepTest; Dako) May Miss Some HER2-Positive Breast Cancers
Frank Schneider,Yulan Jin,Kevin Van Smaalen,Evin H Gulbahce,Rachel E Factor,Xiaoxian Li
American Journal of Clinical Pathology. 2019; 151(5): 504
[Pubmed] | [DOI]
2 Her2 immunohistochemical evaluation by traditional microscopy and by digital analysis, and the consequences for FISH testing
Thomas Marcuzzo,Fabiola Giudici,Elisa Ober,Clara Rizzardi,Cristina Bottin,Fabrizio Zanconati
Pathology - Research and Practice. 2016; 212(10): 911
[Pubmed] | [DOI]



 

Top
 
 
  Search
 
    Similar in PUBMED
   Search Pubmed for
   Search in Google Scholar for
 Related articles
    Email Alert *
    Add to My List *
* Registration required (free)  


    Abstract
   Introduction
    Materials AND ME...
   Results
   Discussion
   Conclusion
    References
    Article Tables

 Article Access Statistics
    Viewed2869    
    Printed71    
    Emailed1    
    PDF Downloaded122    
    Comments [Add]    
    Cited by others 2    

Recommend this journal