Clinical Immunoprofiling Reveals that High Numbers of CD8+and PD-1+Cells Predict Superior Patient Survival Across Major Cancer Types Independent of Major Risk Factors

ABSTRACT Background Numerous retrospective studies have shown associations between the number of intratumoral immune cells and patient outcomes for individual cancers treated with specific therapies. However, the clinical value of using a digital pathology platform to enumerate intratumoral immune biomarkers prospectively in the pan-cancer setting has not been established. Methods We developed ImmunoProfile, a clinical workflow combining automated multiplex immunofluorescence tissue staining, digital slide imaging, and machine learning-assisted scoring to quantify intratumoral CD8 + , PD-1 + , CD8 + PD-1 + , and FOXP3 + immune cells and PD-L1 expression in formalin-fixed, paraffin-embedded tissue samples in a standardized and reproducible manner. Over three years, we prospectively applied ImmunoProfile to biopsies collected from 2,023 unselected cancer patients in the clinical laboratory. We correlated the results with patient survival. Results In the pan-cancer cohort, patients with intratumoral CD8 + or PD-1 + cells in the top or middle tertiles had significantly lower risks of death than those in the bottom (CD8 + (high vs. low) HR 0.62 [95% CI 0.48 – 0.81], (middle vs. low) HR 0.82 [95% CI 0.67 - 0.99], Wald p=0.002]; PD-1 + (high vs. low) HR 0.65 [95% CI 0.51 - 0.83], (middle vs. low) HR 0.74 [95% CI: 0.60 - 0.90], p=0.0009) after controlling for risk factors, including cancer type. In subset analyses, patients with high intratumoral CD8 + , PD-1 + , and/or CD8 + PD-1 + cells had lower risks of death from non-small cell lung, colorectal, breast, esophagogastric, head and neck, pancreatic, and ovarian cancers after controlling for clinical risk factors, including stage, and despite distinct therapies (all p < 0.05). Conclusions Enumerating intratumoral CD8 + and PD-1 + cells with a clinically validated digital pathology platform predicts patient survival across cancer types independent of clinical stage and despite disparate therapies.