The type of PD-L1 assay was anonymized, and PD-L1 expression was evaluated in both ICs and TCs with no special training other than following the manufacturer's instructions (Ventana PD-L1 assay interpretation guideline). The ICs were scored as the proportion of tumor area covered with any discernible PD-L1 staining of any intensity. The ICs that were counted included lymphocytes, macrophages, dendritic cells, and granulocytes. The tumor area was defined as the area occupied by TCs, as well as their associated intratumoral and contiguous peritumoral stroma. For TCs, positive PD-L1 staining was defined as complete and/or partial circumferential linear cellular membrane staining at any intensity that could be differentiated from the background and diffuse cytoplasmic staining, as previously described [15]. The ICs and TCs were scored in both continuous scores (0%–100%) and five categorical scores (< 1%, 1%–4%, 5%–9%, 10%–49%, and ≥ 50%). For 22C3, the combined positive score (CPS) was also calculated by dividing the number of PD-L1-stained cells (TCs and ICs) by the total number of viable TCs and multiplying the value by 100 [16].

To evaluate the intraobserver reproducibility and the impact of training, six participating pathologists, who were trained in scoring SP142 in TNBC after the first assessment, re-evaluated the SP142 assays. Training consisted of presentation covering the biology of PD-L1, development of the assay, cellular expression, and demonstration of PD-L1 interpretation in the clinical samples of TNBC in a half day.

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