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 + 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 + TILs and TCR β chain (TCRβ) clonotypic frequency in melanoma tumors to identify patient-specific repertoires of tumor-reactive CD8 + 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 + TILs identified the autologous tumor-reactive repertoire, including mutated neoantigen-specific CD8 + 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 + PD-1 + compared with CD8 + PD-1 -TIL populations. Furthermore, the most highly expanded TCRβ clonotypes in the CD8 + and the CD8 + PD-1 + 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 + 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.

Antibodies capable of neutralizing divergent influenza A viruses could form the basis of a universal vaccine. Here, from subjects enrolled in an H5N1 DNA/MIV-prime-boost influenza vaccine trial, we sorted hemagglutinin-cross reactive memory-B cells and identified three antibody classes, each capable of neutralizing diverse subtypes of group 1 and 2-influenza A viruses. Co-crystal structures with hemagglutinin revealed that each class utilized characteristic germline genes and convergent sequence motifs to recognize overlapping epitopes in the hemagglutinin stem. All six analyzed subjects had sequences from at least one multidonor class, and -in half the subjects- multidonor-class sequences were recovered from >40% of cross reactive-B cells. By contrast, these multidonor-class sequences were rare in published antibody datasets. Vaccination with a divergent hemagglutinin can thus increase the frequency of B cells encoding broad influenza A-neutralizing antibodies; we propose the sequence signature-quantified prevalence of these B cells as a metric to guide universal influenza A-immunization strategies.

SUMMARY Detailed studies of the broadly neutralizing antibodies (bNAbs) that underlie the best available examples of the humoral immune response to HIV are providing important information for the development of therapies and prophylaxis for HIV-1 infection. Here, we report a CD4-binding site (CD4bs) antibody, named N6, that potently neutralized 98% of HIV-1 isolates, including 16 of 20 that were resistant to other members of its class. N6 evolved a mode of recognition such that its binding was not impacted by the loss of individual contacts across the immunoglobulin heavy chain. In addition, structural analysis revealed that the orientation of N6 permitted it to avoid steric clashes with glycans, which is a common mechanism of resistance. Thus, an HIV-1-specific bNAb can achieve potent, near-pan neutralization of HIV-1, making it an attractive candidate for use in therapy and prophylaxis.

Human leukocyte antigen (HLA) B*27 and B*57 are associated with protection against HIV-1 disease progression, yet most persons expressing these alleles are unable to control HIV-1. Here we show that HLA-B*27-restricted CD8+ T cells in controllers and progressors differ in their ability to inhibit virus replication through targeting of the immunodominant Gag epitope. This is associated with distinct TCR clonotypes, characterized by superior control of HIV-1 replication in vitro, greater cross-reactivity against epitope variants, and enhanced perforin delivery. Clonotype-specific differences in antiviral efficacy were also observed for an immunodominant HLA-B*57 restricted response in controllers and progressors. Thus, the efficacy of protective alleles is modulated by specific TCR clonotypes selected in natural infection, providing a functional explanation for divergent HIV-1 outcomes.