VC TILs demonstrated immune compositions and T cell phenotype similar to those of CC TILs
To characterize the components and phenotype of expanded VC TILs, we analyzed the immune subsets as well as markers involved in T cell activation, differentiation and function. At the end of preREP, all TIL samples are mainly composed of CD3+CD56- T cells (mean>50%) with a small fragment of CD3-CD56+ NK cells (around 10%) (Figure 2A). The percentage of CD8+ and CD4+ T cells showed high variance among different individuals in both cancer types (Figure 2B). Both VC and CC TILs showed high expression of CD28, a marker associated with T cell activation and antitumor function(17), suggesting a functional phenotype of TILs in preREP with our preparation procedure (Figure 2C, 2D).
At the end of REP, T cells were further expanded in culture (most>90%) and the NK population decreased to a minimal level (Figure 3A). CD8+ or CD4+ subset in the T cell population still can still be observed in several samples in both types of cancers (Figure 3B). Similar to TILs in the preREP, the CD28+ T cells remained at a high percentage in the REP, while CD28+CD8+ T cells in VC TILs showed an even higher percentage compared to CC TILs (Figure 3C). We further analyzed the expression of inhibitory receptors TIM-3 and LAG-3, both of which are involved in T cell exhaustion on TILs(18). Both CC and VC TILs highly expressed TIM-3, but VC TILs showed lower expression of LAG-3 on both CD8+ and CD4+ T cell populations (Figure 3D). Tregs in both VC and CC TILs are under 5% (Figure 3E). Taken together, both VC and CC TILs are predominantly composed of T cells at the end of REP, but VC TILs may have a more active CD8+ T cell population as indicated by higher CD28 and lower LAG-3 expression (Figure 3F).