Immunohistochemistry staining was performed as we have described previously.(24 (link)) Tissue was processed for routine IHC staining using the antibodies against Deptor and c-Myc (Santa Cruz Biotechnology, Santa Cruz, CA), phospho-S6 (pS235/236), phospho-Akt (S473), β-catenin (Cell Signaling, Beverly, MA) and Cd8α (Thermo Fisher Scientific, San Diego, CA). Negative controls (including no primary antibody or isotype-matched mouse immunoglobulin G) were used in each assessment. Fifty-six clinical specimens of primary CRC with matching normal colonic mucosa were analyzed for expression of Deptor (Cell Signaling, Beverly, MA). Blind scoring performed by a pathologist used a semi-quantitative method. The extent of expression score was assessed on a scale of 0 to 3 (no positive cells = 0, <10% = 1, 10%–50% = 2, positive staining of >50% = 3); the intensity score was also measured on a scale of 0 to 3 (negative = 0, weak = 1, moderate = 2, strong = 3). Multiplication of the values for intensity and extent of expression provided a score for immunoreactivity as we have described previously.(26 (link))