الفهرس 
INTRODUCTIONOnly 14 pages are availabe for public view
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Abstract
Breast cancer is a major concern and one of the leading causes of cancer-related death worldwide. In Egypt, breast cancer came as number 1 in ranking of malignant tumors constituting 17.5% of total malignancies in both sexes. Molecular classification of breast cancer is a subject of debate for many researchers.
The aim of this study was to evaluate the immunohistochemical expression of CK 8/18 and Ki 67 in breast carcinoma of Egyptian patients to identify its categories according to molecular classification with the help of ER, PR and HER2/neu. Then investigate the impact of this categorization on available clinicopathologic features.
The material of this study consisted of archival paraffin blocks of 70 specimens from breast carcinoma. They are composed of 61(87.1%) invasive duct carcinoma no special type (IDC, NST), 4 (5.7%)invasive lobular carcinoma (ILC) and other types including 3 medullary and 2 mixed tubular carcinomas.
Histologic type, tumor grade, tumor stage, Nottingham Prognostic index (NPI), stroma, lymphovascular invasion (LVI), spontaneous tumor necrosis, Paget’s disease, mitotic and apoptotic counts were assessed in haematoxylin and eosin (H&E) stained sections. Moreover, Estrogen receptor (ER), Progesterone receptor (PR) and Her2neu statuses were assessed in immunohistochemical stained sections. Then, all cases were investigated for Cytokeratin 8/18 (CK 8/18) expression and Ki 67 labelling index (LI) with classification of grade II tumors according to the proliferation marker Ki 67 into grade IIa and grade IIb.
Forty six cases (65.71%) were ER positive while 40 cases 57.14% were PR positive. As regards to hormonal status, we had 3 groups: a)- forty cases (57.14%) were both ER and PR positive, b)- Six cases(8.57%) were ER positive but PR negative, c)- Twenty four cases
(34.29%) were both ER and PR negative. Twenty two out of 70 cases(31.43%) were Her2neu positive, 9 cases (12.86%) were triple positive while 13 cases (18.58%) were triple negative.
As regards to CK 8/18, all cases showed positivity of CK 8/18 in malignant cells. We did not find any significant association between CK 8/18 positivity and clinicopathologic parameters. In tumor cells, 49 cases (70%) showed predominant cytoplasmic expression, while 21 cases (30%) showed both patterns of expression (membranocytoplasmic).
Cytokeratin 8/18 was also expressed in adjacent normal breast tissue in all cases and showed membranocytoplasmic pattern variable in expression ranging from mild to strong. CK 8/18 H score ranged between 20 and 290.
Predominant cytoplasmic pattern (without membranous pattern) of CK 8/18 was significantly associated with higher tumor grade (P=0.05),
higher mitotic count (P=0.03), higher apoptotic count (P=0.05), lower value of PR H score (P=0.02) and triple negative phenotype (P=0.005).
Lower CK 8/18 H score showed significant association with advanced stage (P=0.04) and poor NPI (P=0.04). So, predominant cytoplasmic pattern of expression of CK 8/18 as well as lower values of CK 8/18 H score was associated with poor prognostic factors.
Regarding Ki 67, all cases showed nuclear expression of Ki 67. Ki 67 index ranged between 2% and 98%. According to the cutoff point of 14%, tumors were classified into low proliferative which represent 26 cases (37.14%), while high proliferative tumors were 44 cases
(62.86%).
High proliferative tumors were associated with higher tumor grade (P=0.001), presence of spontaneous tumor necrosis (P=0.03), higher mitotic count (P=0.001) and higher apoptotic count (P=0.008). It was found that 95.8% of ER negative cases and 80% of PR negative tumors as well as 100% of Her2neu positive cases were of high proliferative rate.
Histological grade II breast cancer was stratified into either (a) low(Ki 67-LI<14%), or (b) high proliferative (Ki 67-LI > 14%) groups:
GIIa, GIIb respectively. There was a near significant association between Ki 67 labelling index related subgroups of grade II tumors and both mitotic and apoptotic counts (P=0.06 for each parameter).
A highly statistical significant association was found between Ki 67 proliferation index and IHC molecular typing of breast carcinoma cases(P<0.001), since all cases of luminal B and Her2neu positive types belonged to high proliferative Ki 67 index.
According to the immunohistochemical profile of ER, PR and Her2neu, we classified the studied breast carcinoma cases according to the molecular classification into luminal, Her2neu positive and triple negative equivalent subtypes. Then according to Ki67 LI, we further classified luminal subtype into luminal A and luminal B subtypes.
It was found that 65.71% belonging to luminal subtype, (of which;
76.1% cases were of luminal A subtype while the remaining 23.91% cases were of the luminal B subtype), 18.57% of cases were triple negative and 15.71% cases were of Her2neu positive subtype.
Classification of luminal B tumors into: luminal B1 (ER and/or PR positive, Her2neu negative and high Ki 67 LI) and luminal B2 (ER and/or PR positive, Her2neu positive and any value for Ki 67 LI)revealed that all cases of luminal B subtype in the current study-
belonged to luminal B2 as they had positive expression of Her2neu and high proliferative rate as expressed by high Ki 67 LI.
Tumor grade II (P<0.001), absence of spontaneous tumor necrosis(P<0.001) and low mitotic count (P=0.002) were significantly associated with luminal type of breast carcinoma. High mean value of ki 67 LI as well as high proliferation group ki 67 LI were associated with non-luminal type of breast carcinoma (P=0.002 and P<0.001 respectively). There was a statistical significant association between luminal B subtype and both higher tumor grade and higher mitotic count (P=0.03 and P=0.05 respectively). Tumors belonging to luminal B subtype were also associated with high Ki 67 labelling index (P=0.005)and all belonged to high Ki 67 proliferative group (P<0.001). Her2neu positive tumors were highly associated with presence of necrosis(P<0.001), higher tumor grade and higher mitotic count (P=0.007 and P=0.039) respectively. These tumors were also associated with higher Ki 67 LI (P=0.018) and high proliferation group (P=0.001). Her2neu positive tumors were also highly associated with membrano-cytoplasmic pattern of CK 8/18 (P=0.001). Triple negative tumors were associated with higher tumor grade (P=0.003), higher mitotic count (P=0.011) and higher value of NPI (P=0.05). They were also significantly associated with higher Ki 67 LI (P=0.04).
According to the survival analysis, the univariate Kaplan-Meier analysis revealed absent skin manifestations (P=0.03), grade IIa (P=0.02), absence of Paget’s disease (P=0.02), positive ER status(P=0.04), positive PR status (P= 0.03), positive hormonal status(P=0.03), three marker status (triple positive) (P=0.02) and non-triple negative class (P=0.03) had statistically significant association with longer overall survival. Luminal subtype of breast carcinoma had a tendency to be associated with longer survival, but the association was near significant (P=0.06). Cox regression analysis indicated that Hormonal status (ER, PR positive) and PR status (positive) were independent prognostic factors affecting OS.