BACKGROUND: Elevated tumor-infiltrating lymphocytes (TILs) within the tumor 
microenvironment is a known positive prognostic factor in colorectal cancer 
(CRC). We hypothesized that since cytotoxic T cells release cytolytic proteins 
such as perforin (PRF1) and pro-apoptotic granzymes (GZMA) to attack cancer 
cells, a cytolytic activity score (CYT) would be a useful tool to assess 
anticancer immunity.
METHODS: Genomic expression data were obtained from 456 patients from The Cancer 
Genome Atlas (TCGA). CYT was defined by GZMA and PRF1 expression, and CIBERSORT 
was used to evaluate intratumoral immune cell composition.
RESULTS: High CYT was associated with high microsatellite instability (MSI-H), 
as well as high levels of activated memory CD4+T cells, gamma-delta T cells, and 
M1 macrophages. CYT-high CRC patients had improved overall survival (p = 0.019) 
and disease-free survival (p = 0.016) compared with CYT-low CRC patients, 
especially in TIL-positive tumors. Multivariate analysis demonstrated that CYT- 
high associates with improved survival independently after controlling for age, 
lymphovascular invasion, colonic location, microsatellite instability, and TIL 
positivity. The levels of immune checkpoint molecules (ICMs)-programmed death-1 
(PD-1), programmed death-ligand 1 (PD-L1), cytotoxic T-lymphocyte-associated 
protein 4 (CTLA4), lymphocyte-activation gene 3 (LAG3), T cell immunoglobulin 
and mucin domain 3 (TIM3), and indoleamine 2,3-dioxygenase 1 (IDO1)-correlated 
significantly with CYT (p < 0.0001); with improved survival in CYT-high and 
ICM-low patients, and poorer survival in ICM-high patients.
CONCLUSIONS: High CYT within CRC is associated with improved survival, likely 
due to increased immunity and cytolytic activity of T cells and M1 macrophages. 
High CYT is also associated with high expression of ICMs; thus, further studies 
to elucidate the role of CYT as a predictive biomarker of the efficacy of immune 
checkpoint blockade are warranted.