CD70 EXPRESSION IN ENDOMETRIAL ENDOMETRIOID CARCINOMA: AN IMMUNOHISTOCHEMICAL STUDY

Dina F. El-Yasergy 1 , Moustafa A. Abousarie 2 and Lobna O. Abdel-Salam 1 . 1. Department of Pathology, Faculty of Medicine, Cairo University, Giza, Egypt. 2. Department of Pathology, Faculty of Medicine, Fayoum University, Fayoum. ...................................................................................................................... Manuscript Info Abstract ......................... ........................................................................ Manuscript History Received: 11 February 2019 Final Accepted: 13 March 2019 Published: April 2019


Introduction:-
Uterine cancer is globally one of the top-ranking female cancers (Ferlay et al., 2015). Endometrial carcinoma arises from the uterine endometrium, and it represents 90% of uterine malignancy, followed by sarcoma that arises from the myometrium (8%) and less frequent cancer types (2%) (Torre et al., 2015). Endometrial carcinoma represents the most common gynecological malignancy in developed countries and the second most common gynecological malignancy in developing countries after cervical cancer (Jemal et al., 2011). It includes two major classes, Type I (endometrioid, the majority) and Type II (non-endometrioid). EECA is the prototypical Type I (Doll et al., 2008). Surgical treatment only is not sufficient for treatment of advanced cases of endometrial adenocarcinoma. It needs also adjuvant treatments which are employed depending on the tumor site, type and stage. Surgical treatment is total abdominal hysterectomy + bilateral salpingo-oopherectomy (TAH + BSO) with removal of pelvic and para-aortic lymph nodes. Adjuvant treatment modalities include chemotherapy and/or radiotherapy. Unfortunately, these treatment modalities have systemic toxicity in most cases. This has inspired the clinicians to search for targeted therapy directed specifically to the malignant cells. Targeting the ligands and receptors of the tumor necrosis factor (TNF) superfamily seems to be promising in treatment of advanced endometrial carcinoma  CD70 is aberrantly upregulated in hematological malignancies and some solid malignancies, including RCC, brain tumors, thymic carcinoma, melanoma, pancreatic, nasopharyngeal, lung, ovarian and colon carcinoma CD70 is found to be a poor prognostic marker in many cancers. As an example, it plays an important role in melanoma progression; increased CD70 expression increases invasiveness of melanoma cell through activation of MAPK pathway, overexpression of RhoE, and modulation of the cytoskeleton

Study design
A retrospective cohort study, with 60 cases of pan-hysterectomy specimens of EEC, collected as paraffin blocks from archives of Pathology Department, Faculty of Medicine, Cairo University during the time period between January 2013 and December 2018. Complete clinicopathological data of all cases were collected from the pathology reports, including age, grade, depth of myometrial invasion, lympho-vascular emboli, TILs, lymph node status and FIGO stage. The tumors were graded histologically according to the revised FIGO grading system (Kandoth et al., 2013). The tumors were staged according to FIGO staging system (Amin et al., 2017). Cases lacking proper data were excluded. The evaluation of immunohistochemical staining results was done by two different pathologists. Approval of institutional review board was obtained before data collection.

Immunohistochemical study
Paraffin-embedded specimens were cut into 4 μm thick sections. The slides were dewaxed through heating at 60 °C for 60 min followed by deparaffinization using xylene and rehydration in graded alcohol. The slides were put afterwards in Citrate Buffer solution (pH 6.0) and microwaved for 10 min at low power for antigen retrieval. Deparaffinized sections were stained with mouse monoclonal anti-CD70 primary antibody (clone 301731 diluted 1:40; R&D Systems Inc., Minneapolis, Md., USA) and DAB detection using the EnVision FLEX+ kit (Dako) according to the instructions of the manufacturer. Slides were counterstained with hematoxylin.

Statistical analysis
Data were analyzed using Statistical Package for the Social Sciences (SPSS software) program, version 21. Its presentation was in the form of mean, SD and percentage. The Fisher exact test was used to compare CD70 expression and clinicopathological parameters. P< 0.05 was considered statistically significant.

Results:-
In this study, 60 paraffin-embedded EEC specimens were collected from archives of Pathology Department, Faculty of Medicine, Cairo University during the period from 2013 to 2018. All specimens were obtained by total abdominal hysterectomy and bilateral salpingo-oophorectomy with removal of regional lymph nodes. Thirty-nine cases (65%) were FIGO grade 2, 13 cases (21.7%) were FIGO grade 3, and 8 cases (13.3%) were FIGO grade 1. Thirty-three specimens (55%) showed tumor infiltration of more than half myometrial thickness, and the remaining 27 tumor specimens (45%) showed infiltration of less than half myometrial thickness. Lympho-vascular emboli were detected in 23 cases (38.3%), while absent in 37 cases (61.7%). Twenty cases (33.3%) were associated with regional lymph node metastasis, while the remaining 40 cases (66.7%) showed no tumor deposits in the dissected regional lymph nodes. TILs were subjectively assessed within the vicinity of the tumor, TILs were mild, moderate and marked in 19 cases (31.7%), 24 cases (40%), and 17 cases (28.3%) respectively.

Discussion:-
CD70 is a type II transmembrane protein belonging to the TNF super family. It is aberrantly upregulated in hematologic malignancies and some solid malignancies and its overexpression is associated with poor prognosis and progression of many cancer types. To our knowledge, CD70 expression was not assessed in EEC. In our study, among the 60 EEC cases, 20%, 26.7%, 33.3%, 20% showed negative, mild, moderate and marked expression, respectively. In this study, we correlated CD70 expression and other pathological variables including grade, stage, depth of myometrial invasion by the tumor tissue, the presence of lympho-vascular emboli, regional lymph node metastasis and TILs within the vicinity of the tumor. We found significant positive correlation between CD70 expression in EEC and each of tumor stage, presence of lympho-vascular emboli, regional lymph node metastasis and TILs within the vicinity of the tumor.

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In many research studies, CD70 expression is associated with poor prognostic variables and poor survival. For example, and not as a limitation, CD70 expression by GBM is associated with poor prognosis and poor overall survival due to involvement of CD70 expression in promoting tumor aggressiveness and immunosuppression (Ge et al., 2017). In hematological malignancies, overexpression of CD70 was found to be involved in proliferation and survival of tumor cells mediated through its interaction with CD27 (Nilsson et al., 2005). High expression of CD70 was associated with decreased survival rate in RCC, clear cell type (Jilaveanu et al., 2012). CD70 is significantly higher in renal sarcomatoid carcinoma (Jilaveanu et al., 2012) which has an aggressive behavior and poor prognosis (ElMouallem el al., 2018). 75% of NSCLC that express CD70 showed poor response to 1 st line medical treatment with less than 1-year survival (Jacobs et al., 2015).
In our study, although there was no significant relation between CD70 expression and the tumor grade, CD70 expression tends to be more marked in higher grade tumors. In this study, CD70 expression tends to be more in tumors invading more than half the myometrial thickness and this was statistically significant. This may be explained as depth of myometrial invasion is one of the parameters of the FIGO staging system which also showed significant positive correlation with CD70 expression.
In the current study, we found that CD70 expression tends to be more marked in tumors having greater density of TILs suggesting an interaction between CD70 on tumor cell with TILs and this was statistically significant. In agreement with us, Jacobs et al., 2015 stated that CD70 expression by tumor cells was found in 16.3% of NSCLC specimens by IHC and CD27-expressing TILs were found adjacent to the tumor cells, suggesting active CD70mediated signaling and this was statistically significant. Also, in GBM, CD70 expression was associated with immune cell infiltrates, such as T cells (Ge et al., 2017). In contrast to our study, De Meulenaere et al., 2016 found that high level of CD70 expression was associated with a lower TILs density within HNSCC.
In summary, our study describes CD70 expression in ECC. CD70 showed mild expression in 26.7% of cases, moderate expression in 33.3%, marked expression in 20%, while and negative in 20%. It displays a significant statistical association with poor pathological variables including stage, depth of myometrial invasion, lymphovascular emboli, regional lymph node metastasis. It also showed significant positive correlation with TILs, however, it shows a non-significant relation with tumor differentiation. These findings suggest that CD70 might be involved in ECC tumor biology and might be a good targeted therapy in ECC expressing CD70.
Further researches are warranted to clarify the function of CD70 expression in the pathophysiology of EEC, to elucidate the response to treatment with CD70 targeted therapy and the possibility of using CD70 as a predictive immunohistochemical marker for assessing the response to anti-CD70 targeted therapy.