In this retrospective study, formalin-fixed, paraffin-embedded tissue blocks of 62 hysterectomy specimens that were diagnosed as endometrioid carcinoma were retrieved from (March 2016 to January 2017) archives in the pathology department of teaching laboratories in Medical City complex at Bagdad. From each block representing endometrial mass, two histological sections were done for routine haematoxylin and eosin staining and for immunohistochemical staining with HER2/neu by application of the same standardised immunohistochemical protocols used for HER2/neu expression in breast cancer [12]. For detection of HER2/neu antigen polyclonal rabbit anti-human c-erb-2 oncoprotein (Code A0485) manufactured by DAKO with a dilution of (1:1000) was used. The antibody was used with the buffer supplied by the manufacturer and the primary antibody was diluted and, after incubation for 30 minutes, visualisation of antigen was done with envision detection kit and after counterstaining with haematoxylin and eosin the sections were cover slipped. Positive and negative controls were included with each run, positive control was breast tissue known to be positive for HER2/neu and negative control was prepared by omitting the primary antibody. Appropriate grading and staging was done for each case according to the FIGO grading and staging system that is widely used now-a-days for evaluation and reporting endometrial carcinoma [9,10]. Age of women was the only available clinical information that was extracted from the patient records. Regarding HER2/neu immunohistochemical interpretation, this study used the previously laid down HER2/neu scoring criteria for breast carcinoma. A 3+ was rendered if complete, strong membranous staining was present in more than 10% of tumour cells and 2+ if complete, moderately intense staining was seen in more than 10% of tumour cells [13].

All histopathological observations were done by three pathologists using a light microscope (Leica DM500). This study was approved by the ethical committee of the national centre for educational laboratories (ethical clearance file number 32/18) and done in accordance with the institutional policy in which patient consent was taken at the beginning of consultation to teaching laboratories for any future retrospective studies. Statistical analysis was done using Chi-square test analysis. Photomicrographs were taken for this study using the camera (Leica Icc 50E).

This study enrolled 62 cases of endometrioid carcinoma, the age range from 35 to 72 years with a mean age of 56.16 years. According to FIGO grading system 40 (64.51%) were grade I, 10 (16.12) were grade II, 12 (19.3%) were grade III tumours. According to FIGO staging system, 44 (70.96%) were stage IA, 8 (12.90%) were stage IB, 4 (6.45%) were stage II, 6 (9.67%) were stage III tumours.

Out of 62 cases 30 (48.38%) were positive for HER2/neu [Table/Fig-1,2] and the remaining cases were negative [Table/Fig-3].

[Table/Fig-1]:

Histological section of grade I endometrioid carcinoma showing positive membranous staining pattern for HER2/neu antibody (x100).

[Table/Fig-2]:

Histological section of grade II endometrioid carcinoma showing positive membranous staining pattern for HER2/neu antibody (x400).

[Table/Fig-3]:

Histological section of grade II endometrioid carcinoma showing negative staining for HER2/neu antibody (x200).

The distribution of HER2/neu immunohistochemical status according to FIGO stage of endometrial carcinoma is shown in [Table/Fig-4].

Association between FIGO grading of endometrioid cancer and HER2/neu immunohistochemical status revealed a highly significant statistical relationship with p-value<0.001 [Table/Fig-5].

Association between patient age and HER2/neu immunohistochemical status revealed a significant statistical relationship with p-value 0.002 [Table/Fig-6].

International Federation of Gynaecology and Obstetrics (FIGO) established a surgical staging and grading system widely used nowadays for endometrial cancer and carries an important prognostic implication in which histological grading relies on both architectural and nuclear features in three tier system while the staging system relies mainly on depth of myometrial and adjacent structures or organ invasion [10]. Treatment is mainly surgical, the extent of which depends on the stage of disease [14,15]. In general, the prognosis of endometrioid carcinoma is favorable because over 80% of disease is diagnosed in the early stages [16].

Immunohistochemical expression of HER2/neu has been used most successfully in mammary cancer as a potent predictor to select patients who will most likely respond to anti-HER2 therapies such as trastuzumab [17]. There were limited studies worldwide on HER2/neu expression in endometrial cancer in general and endometrioid carcinoma in particular (unlike in breast cancer) and no such Iraqi study was done earlier.

HER2 is a 185-kDa member of the human epidermal growth factor receptor (HER) family of transmembrane tyrosine kinase receptor, it is also known as HER2/neu, ERBB2, and CD340 [11,12]. The HER family (to which HER2/neu belong) consists of three other tyrosine kinase receptors (HER1/EGFR, HER3, and HER4) [13,18].

There are no well established standardised criteria currently available for the immunohistochemical methods (i.e., tissue handling/fixation, antibodies used, controls, and artifacts) and for interpretation and scoring of HER2/neu immunohistochemical results. In the majority of studies and in this study, the original FDA approved criteria (previous breast HER2/neu scoring criteria based on more than 10 years experience in breast cancer) with positivity accepted as 3+ and +2 were used [13].

The present study showed that the mean age of endometrioid carcinoma was in the sixth decade of life which agreed with study done by Sharon N et al., and the majority of endometriod cancers were well differentiated grade I which agreed with an Iraqi study done by the study of Sharon N et al., and Abeer et al., [19,20]. This study showed that 48.38% of endometrioid carcinoma were immunohistochemicaly positive for HER2/neu suggestive of its role in the pathogenesis of endometrioid carcinoma. There is a great variation in the reported percentages of HER2/neu overexpression in endometriod carcinoma ranging between 1% to 47% [13,21-24]. There are many factors accounting for this difference such as variations in sample size, methods of testing, detection systems used, immunohistochemical interpretation and scoring criteria with interobservational variations.

The current study also showed that there is a significant statistical relationship between HER2/neu immunohistochemical results and histological grade in which the expression is closely associated with a high grade cancer. Variable results have been reported in the literature in which some studies are in congruence with current results [23,25] while other studies differ [22,26]. Such wide variations in the reported results again are due to lack of standardised protocols of testing HER2/neu in endometrial cancer. Histological grade is regarded as one of the prognostic factors applied in clinical decisions regarding treatment [27].

A significant statistical relationship between the patient age and HER2/neu immunohistochemical results were seen in this study in which the positivity closely associated with patients younger than 40 years. However, Srijaipracharoen S et al., failed to find such an association [27]. Because the cancer treatment field progressed to an era of targeted therapy which has the advantage of overcoming undesirable serious side effects of chemo radiotherapy, search for therapeutic targets by using accurate and reliable immunohistochemical test especially for common cancers worldwide is the first critical step. Our results demand standardisation of HER2/neu testing in endometrioid cancer is mandatory step in order to accurately identify patients who might benefit from HER2/neu-targeted therapy, and further for prognostication because most studies in the literature proved that both HER2/neu over expression and amplification are associated with poor prognosis and low overall patient survival in endometrial carcinoma [13].

A recent study on the use of (Trastuzumab for Gastric Cancer) shows the guidelines of immunohistochemical interpretation in a biologically and pathogenically unrelated different tumour category and couldn’t be simply applied, because this may lead to false interpretation of HER2/neu immunohistochemical results [28].

This is the first study in Iraq and may serve as the starting point towards the awareness about the probable use of HER2/neu targeted therapy in endometrioid carcinoma. However, large scale future immunohistochemical studies as well as genetic studies on HER2/neu amplification associated with clinical therapeutic response to targeted drugs in endometrioid carcinoma and establishing the specific HER2/neu scoring criteria and testing guidelines in endometrial carcinoma are required.

The present study was conducted in a single centre with small number of cases included in the study. There was limited clinical information about cases present in the records. In addition, no standardised testing protocols were available on HER2/neu expression in endometrioid carcinoma.

HER2/neu expression in endometrioid carcinoma is significantly associated with higher grade of tumour and younger age of patients. The study concludes that young patients with a high grade endometrioid carcinoma may hopefully benefit from the application of targeted anti-HER2/neu therapy in future.