AbstractColitis is a common, but poorly understood, adverse event of immune checkpoint inhibitors that are standard-of-care for an expanding range of cancer types. This explorative study aimed to describe the immune infiltrates in the colon from individuals developing checkpoint inhibitor colitis and compare them to well-known immunophenotypes of acute graft-versus-host disease, ulcerative colitis, and Crohn’s disease. Colon biopsies (n = 20 per group) of patients with checkpoint inhibitor colitis, acute graft-versus-host disease, ulcerative colitis and Crohn’s disease, all colitis treatment-naïve, and of individuals with a normal colon were analyzed using immunohistochemistry: CD8 for cytotoxic T cells, CD4 for T helper cells, and CD68 to identify cells of macrophage lineage. CD8 + T cell, CD4 + T cell, and CD68 + cell counts were performed. Cell infiltration was scored as scattered/patchy or band-like in the superficial and deep gut mucosa. Checkpoint inhibitor colitis was found to be heavily infiltrated by CD8 + T cells. Comparative analysis between groups showed that both CD8 + T cell counts (P < 0.01) and immune cell infiltration patterns in checkpoint inhibitor colitis were most similar to those observed in ulcerative colitis, with a deep band-like CD4 + T cell infiltration pattern and a superficial band-like CD68 + cell infiltration pattern in both. In conclusion, this is the first immunohistopathological study comparing infiltrate characteristics of checkpoint inhibitor colitis, acute graft-versus-host disease, ulcerative colitis, and Crohn’s disease. Checkpoint inhibitor colitis samples are heterogeneous, heavily infiltrated by CD8 + T cells, and show an immune cell infiltration pattern that is more similar to ulcerative colitis than to colonic acute graft-versus-host disease or colonic Crohn’s disease.