Recent studies have found that tumor-infiltrating lymphocytes (TIL) expressing PD-1 can recognize autologous tumor cells, suggesting that cells derived from PD-1 TILs can be used in adoptive T-cell therapy (ACT). However, no study thus far has evaluated the antitumor activity of PD-1-selected TILs In two mouse models of solid tumors, we show that PD-1 allows identification and isolation of tumor-specific TILs without previous knowledge of their antigen specificities. Importantly, despite the high proportion of tumor-reactive T cells present in bulk CD8 TILs before expansion, only T-cell products derived from sorted PD-1, but not from PD-1 or bulk CD8 TILs, specifically recognized tumor cells. The fold expansion of PD-1 CD8 TILs was 10 times lower than that of PD-1 cells, suggesting that outgrowth of PD-1 cells was the limiting factor in the tumor specificity of cells derived from bulk CD8 TILs. The highly differentiated state of PD-1 cells was likely the main cause hampering expansion of this subset. Moreover, PD-1 precisely identified marrow-infiltrating, myeloma-specific T cells in a mouse model of multiple myeloma., only cells expanded from PD-1 CD8 TILs contained tumor progression, and their efficacy was enhanced by PDL-1 blockade. Overall, our data provide a rationale for the use of PD-1-selected TILs in ACT. .

<p>Supplementary Figures: (Fig. S1) Multi-parametric flow cytometric analysis of tumor-infiltrating T lymphocyte subsets from freshly excised MC38 tumors; (Fig. S2) Expression of co-inhibitory and co-stimulatory molecules over the course of tumor progression; (Fig. S3) Tumor-reactivity of CD137+ and PD-1+ TILs, freshly after isolation from tumors and after in vitro expansion; (Fig. S4) Kinetic of expansion of different CD8 TIL subsets isolated from MC38 tumors; (Fig. S5) CDR3 spectratype analysis. Supplementary Tables: (Table S1) Number of CDR3 fragments which dominance hierarchy changed after TIL expansion; (Table S2) Number of TCR-Vbeta families that lose, preserve or acquire new dominant CDR3 fragments after TIL expansion. Supplemental Material and Methods: Spectratyping.</p>

<div>Abstract<p>Recent studies have found that tumor-infiltrating lymphocytes (TIL) expressing PD-1 can recognize autologous tumor cells, suggesting that cells derived from PD-1⁺ TILs can be used in adoptive T-cell therapy (ACT). However, no study thus far has evaluated the antitumor activity of PD-1–selected TILs <i>in vivo</i>. In two mouse models of solid tumors, we show that PD-1 allows identification and isolation of tumor-specific TILs without previous knowledge of their antigen specificities. Importantly, despite the high proportion of tumor-reactive T cells present in bulk CD8 TILs before expansion, only T-cell products derived from sorted PD-1⁺, but not from PD-1⁻ or bulk CD8 TILs, specifically recognized tumor cells. The fold expansion of PD-1⁺ CD8 TILs was 10 times lower than that of PD-1⁻ cells, suggesting that outgrowth of PD-1⁻ cells was the limiting factor in the tumor specificity of cells derived from bulk CD8 TILs. The highly differentiated state of PD-1⁺ cells was likely the main cause hampering <i>ex vivo</i> expansion of this subset. Moreover, PD-1 precisely identified marrow-infiltrating, myeloma-specific T cells in a mouse model of multiple myeloma. <i>In vivo</i>, only cells expanded from PD-1⁺ CD8 TILs contained tumor progression, and their efficacy was enhanced by PDL-1 blockade. Overall, our data provide a rationale for the use of PD-1–selected TILs in ACT. <i>Cancer Res; 77(13); 3672–84. ©2017 AACR</i>.</p></div>

<p>Supplementary Figures: (Fig. S1) Multi-parametric flow cytometric analysis of tumor-infiltrating T lymphocyte subsets from freshly excised MC38 tumors; (Fig. S2) Expression of co-inhibitory and co-stimulatory molecules over the course of tumor progression; (Fig. S3) Tumor-reactivity of CD137+ and PD-1+ TILs, freshly after isolation from tumors and after in vitro expansion; (Fig. S4) Kinetic of expansion of different CD8 TIL subsets isolated from MC38 tumors; (Fig. S5) CDR3 spectratype analysis. Supplementary Tables: (Table S1) Number of CDR3 fragments which dominance hierarchy changed after TIL expansion; (Table S2) Number of TCR-Vbeta families that lose, preserve or acquire new dominant CDR3 fragments after TIL expansion. Supplemental Material and Methods: Spectratyping.</p>