sCD40L induced greater PD-1 expression in vitro on T cells from cancer patients (CP) than T cells from healthy donors (HD). (A-B) Induction of CD25 and CD70 on T cells. PBMCs from 6 age-matched HD and 5 metastatic prostate CP were incubated in medium containing 25 U/mL of IL-2 with or without 2 μg/mL of sCD40L for 4 days. Cells were then analyzed by FACS. (C) Comparison of PD-1–expressing T cells in PBMCs between HD and CP after the addition of sCD40L to the culture. The samples were also evaluated for PD-1 expression on CD3⁺ T cells. (D) Differential enhancement of PD-1 expression on CD4⁺ and CD8⁺ T cells after the addition of sCD40L to cell cultures in HD and CP. PD-1 expression on CD4⁺ and CD8⁺ T cells was evaluated. All results are representative of 2 individual repeats. Statistical analysis was performed using a paired t test. *P < .05. NS indicates not significant.