PD-1 identifies the patient-specific [CD8.sup.+] tumor-reactive repertoire infiltrating human tumors

Adoptive transfer of tumor-infiltrating lymphocytes (TILs) can mediate regression of metastatic melanoma; however, TILs are a heterogeneous population, and there are no effective markers to specifically identify and select the repertoire of tumor-reactive and mutation-specific [CD8.sup.+] lymphocytes. The lack of biomarkers limits the ability to study these cells and develop strategies to enhance clinical efficacy and extend this therapy to other malignancies. Here, we evaluated unique phenotypic traits of [CD8.sup.+] TILs and TCR P chain (TCRβ) clonotypic frequency in melanoma tumors to identify patient-specific repertoires of tumor-reactive [CD8.sup.+] lymphocytes. In all 6 tumors studied, expression of the inhibitory receptors programmed cell death 1 (PD-1; also known as CD279), lymphocyte-activation gene 3 (LAG-3; also known as CD223), and T cell immunoglobulin and mucin domain 3 (TIM-3) on [CD8.sup.+] TILs identified the autologous tumor-reactive repertoire, including mutated neoantigen-specific [CD8.sup.+] lymphocytes, whereas only a fraction of the tumor-reactive population expressed the costimulatory receptor 4-1BB (also known as CD137). TCRβ deep sequencing revealed oligoclonal expansion of specific TCRβ clonotypes in [CD8.sup.+][PD-1.sup.+] compared with [CD8.sup.+][PD-1.sup.-] TIL populations. Furthermore, the most highly expanded TCRβ clonotypes in the [CD8.sup.+] and the [CD8.sup.+][PD-1.sup.+] populations recognized the autologous tumor and included clonotypes targeting mutated antigens. Thus, in addition to the well-documented negative regulatory role of PD-1 in T cells, our findings demonstrate that PD-1 expression on [CD8.sup.+] TILs also accurately identifies the repertoire of clonally expanded tumor-reactive cells and reveal a dual importance of PD-1 expression in the tumor microenvironment.