Abstract
The prevalence of cytotoxic tumor infiltrating lymphocytes (TILs) has demonstrated prognostic value in multiple tumor types. In particular, CD8 counts (in combination with CD3 and CD45RO) have been shown to be superior to traditional UICC staging in colon cancer patients and higher total CD8 counts have been associated with better survival in breast cancer patients. However, immune infiltrate heterogeneity can lead to potentially significant misrepresentations of marker prevalence in routine histologic sections. We examined step sections of breast and colorectal cancer samples for CD8+ T cell prevalence by standard chromogenic immunohistochemistry to determine marker variability and inform practice of T cell biomarker assessment in formalin-fixed, paraffin-embedded (FFPE) tissue samples. Stained sections were digitally imaged and CD8+ lymphocytes within defined regions of interest (ROI) including the tumor and surrounding stroma were enumerated. Statistical analyses of CD8+ cell count variability using a linear model/ANOVA framework between patients as well as between levels within a patient sample were performed. Our results show that CD8+ T-cell distribution is highly homogeneous within a standard tissue sample in both colorectal and breast carcinomas. As such, cytotoxic T cell prevalence by immunohistochemistry on a single level or even from a subsample of biopsy fragments taken from that level can be considered representative of cytotoxic T cell infiltration for the entire tumor section within the block. These findings support the technical validity of biomarker strategies relying on CD8 immunohistochemistry.
Highlights
Breast and colorectal cancers are the second and third most common malignancies worldwide, respectively
Our results show that CD8+ T cell numbers are highly homogenous throughout colon and breast cancer tissue section and that a single virtual core biopsy can potentially adequately represent tumor-associated cytotoxic T cell prevalence, though estimates improve with increasing sample number
In order to determine whether sampling error contributes to variation in intratumoral T cell counts, we sectioned through and enumerated CD8+ T cells in 13 primary colorectal adenocarcinoma and 12 breast carcinoma (6 ductal adenocarcinoma, 6 medullary carcinoma) cases (1 FFPE block per case)
Summary
Breast and colorectal cancers are the second and third most common malignancies worldwide, respectively. Between 2008 and 2015, breast cancer incidence has increased by more than 20%, while mortality has increased by 14% and it represents one in four of all cancers in women [1]. Colorectal cancer, at the same time, has decreased 3.2% per year in incidence mortality has remained stable [2]. Heterogeneity of CD8 T cells in breast and colon cancer adherence to PLOS ONE policies on sharing data and materials
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