As previously described [17 (link)], we evaluated ER, PR, HER2, and Ki67 expression using the following antibodies, ER (1:100 clone 6F11; Novocastra, Newcastle upon Tyne, UK), PR (clone 16; Novocastra), HER2 (4B5 rabbit monoclonal antibody; Ventana Medical Systems, Tucson, AZ, USA), and Ki-67 (MIB-1; Dako, Glostrup, Denmark). ER- and PR-positive IHC expression was defined according to the modified Allred system: positive, Allred score 3–8 and negative, Allred score 0–2. HER2 status was re-evaluated according to American Society of Clinical Oncology/College of American Pathologists guideline [18 (link)]. HER2 status was considered positive if the score was 3+, and was considered negative with a score of 0 or 1+. Tumors with a score of 2+ underwent FISH or SISH analysis, according to the manufacturer’s instructions (PathVysion kit; Vysis, Downers Grove, IL, USA or HER2 inform; Ventana). Ki-67 expression is presented as the percentage (range 0–100%) of positive tumor cells.
For the molecular subtyping, the following definitions were used: i) Luminal/HER2 negative: ER positive and/or PR positive and HER2 negative; ii) HER2 positive: HER2 positive regardless of ER and PR status; and iii) TNBC: ER negative, PR negative, and HER2 negative.
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