Treatment schedules for the two studies, baseline characteristics and clinical outcomes of both trials have already been described in detail. Clinical proto cols were approved by local regulatory authorities, sellekchem while the present translational research protocol was approved by the Bioethics Committee of the Aristotle University of Thessaloniki, School Inhibitors,Modulators,Libraries of Medicine, under the general title Molecular investigation of the predictive and or prognostic role of important signal transduction path ways in breast cancer. All patients signed a study specific written informed consent before randomization, which in addition to providing consent for the trial allowed the use of their biological material for future research purposes. In total, 271 patients from the HE10 97 trial and 669 from the HE10 00 trial were included in this study, based on tissue availability.
Primary tumor diameter, ax illary nodal status and Inhibitors,Modulators,Libraries tumor grade were obtained from the pathology report in each case. TMA construction Formalin fixed paraffin embedded tissue samples from patient tumors were collected retro spectively in the first trial and prospectively in the second. The present study was carried out on tissue microarrays. Representative hematoxylin eosin stained sections from the tissue blocks were re viewed by two experienced in breast cancer pathologists and the most representative tumor areas were marked for the construction of U blocks with a manual arrayer. Each tumor was represented by 2 tissue cores, 1.
5 mm in diam eter, which were obtained from different and most repre sentative areas of primary invasive or in some cases from synchronous axillary lymph node metastases and re embedded in recipient paraffin blocks. TMAs also contained control cores from different tumors and non neoplastic tissues. Inhibitors,Modulators,Libraries In total, 51 TMAs were created. Cases not represented, damaged or inadequate on the TMA sections were re cut from the original blocks, where tissue material was still available. Immunohistochemistry for breast cancer subtyping IHC for ER, PgR, HER2, Ki67, CK5, CK14, CK17 and EGFR was performed on serial 2 micron thick sections, using the Bond Max and Bond III autostainers, as previously described. IHC was performed centrally at the Laboratory of Molecular On cology of the Hellenic Inhibitors,Modulators,Libraries Foundation for Cancer Research, Aristotle University of Thessaloniki School of Medicine.
Vimentin and cytokeratin 8 18 were used as immunoreactivity controls and for the identification Inhibitors,Modulators,Libraries of tumor cells. Tissue samples negative with the above two antibodies were ex cluded from the study. The evaluation of all IHC sections was done by experienced breast cancer pathologists, blinded to the patient clinical characteristics and survival data. All tumors included in this study were nevertheless classified based on ER, PgR, HER2, Ki67, CK5 and EGFR, as Luminal A, Luminal B, luminal HER2, HER2 enriched and TNBC.