Harris, Kimberley J.

Abstract B7 homolog-1 ( B7-H1) expressed by antigen-presenting cells or tumor cells is a negative regulator of T cell responses, however its function on T cells is not clear. Here we show that B7-H1 was upregulated in CD8 T cells during immunization and its expression was maintained on memory CD8 T cells. Although B7-H1 deficient naïve CD8 T cells responded normally to antigen stimulation and underwent normal antigen-independent homeostatic proliferation, the maximal expansion of CD8 T cells was blocked due to their high susceptibility to apoptosis. B7-H1-deficient effector CD8 T cells showed weaker effector function and cytotoxicity against tumor cells compared to that of normal CD8 T cells. Once activated, B7-H1-deficient CD8 T cells generated less memory T cells. This effect was independent of the primary expansion of the antigen-specific CD8 T cell population. Our results revealed previously unknown function of B7-H1 on CD8 T cells in mediating survival and differentiation of effector CD8 T cells into memory T cells.

The expression of B7‐H1 on tumor infiltrating lymphocytes usually correlates with poor prognosis in cancer patients. It has also been observed that B7‐H1 can deliver signals into T cells, and therefore we hypothesized that B7‐H1 on T cells reduces T cell‐mediated anti‐tumor immunity. B7‐H1 knockout (KO) mice, when compared to their WT counterparts, have increased numbers of CD8 T cells with memory effector phenotype, the population of T cells that quickly respond upon antigen restimulation. In addition, B7‐H1 KO CD8 T cells proliferate more rapidly during homeostatic expansion. Once activated, these cells exhibit improved CTL function, as demonstrated by increased degranulation and IFN‐gamma production. Blocking the B7‐H1/PD‐1 pathway not only improved the CTL function of CD8 T cells when mixed with PD‐1+ tumors in vitro but also improved their ability to reject established tumors in vivo. This novel functional role for B7‐H1 on CD8 T cells provides us with a new approach to manipulate anti‐tumor T cell response and to improve current tumor immunotherapy. (This work is funded by Mayo Foundation).

Abstract Development of effective vaccination approaches to treat established tumors represents a focus of intensive research because such approaches offer the promise of enhancing immune system priming against tumor Ags via restimulation of pre-existing (memory) antitumoral helper and effector immune cells. However, inhibitory mechanisms, which function to limit the recall responses of tumor-specific immunity, remain poorly understood and interfere with therapies anticipated to induce protective immunity. The mouse renal cell carcinoma (RENCA) tumor model was used to investigate variables affecting vaccination outcomes. We demonstrate that although a whole cell irradiated tumor cell vaccine can trigger a functional antitumor memory response in the bone marrows of mice with established tumors, these responses do not culminate in the regression of established tumors. In addition, a CD103+ regulatory T (Treg) cell subset accumulates within the draining lymph nodes of tumor-bearing mice. We also show that B7-H1 (CD274, PD-L1), a negative costimulatory ligand, and CD4+ Treg cells collaborate to impair the recall responses of tumor-specific memory T cells. Specifically, mice bearing large established RENCA tumors were treated with tumor cell vaccination in combination with B7-H1 blockade and CD4+ T cell depletion (triple therapy treatment) and monitored for tumor growth and survival. Triple treatment therapy induced complete regression of large established RENCA tumors and raised long-lasting protective immunity. These results have implications for developing clinical antitumoral vaccination regimens in the setting in which tumors express elevated levels of B7-H1 in the presence of abundant Treg cells.

Abstract Adoptive γδ T cell immunotherapy has moved briskly into clinical trials prompted by several small studies suggesting abundant accumulation of γδ T cells within renal cell carcinoma (RCC). In this study, we re-examined levels of γδ T cells within RCC tumors and correlated levels of these cells with pathologic features and outcome associated with this form of cancer. Tissues from 248 consecutive clear cell RCC tumors obtained from 2000 to 2003 were stained and quantified for total CD3+ and γδ T cells per mm2. Wilcoxon rank sum and Kruskal-Wallis tests were used to evaluate associations between T cell amounts and prognostic factors (age, gender, tumor size, stage, grade, tumor necrosis). Cox models were used to assess associations with RCC-specific death. Median numbers of total CD3+ and γδ T cells were 281/mm2 (interquartile range (IQR): 149–536) and 2.6/mm2 (IQR: 1.3–4.6), respectively. The median percentage of CD3+ T cells that were γδ T cells was 1.0% (IQR: 0.4–1.9). This low percentage of intratumoral γδ T cells was diluted even further with rising CD3+ T cell infiltration. Percentages of γδ T cells were not associated with even one single clinicopathologic feature examined. Median follow-up for this study was 3.1 years (48 patients died of RCC) and Cox analysis failed to demonstrate that γδ T cells (hazard ratio = 1.02, p = 0.25) were predictive of RCC-specific death. γδ T cells are rare and not recruited nor expanded within RCC tumors. Percentages of γδ T cells fail to correlate with any prognostic features of RCC nor specific death. As such, the role of γδ T cells in RCC immunobiology remains questionable.

Abstract An immunoinhibitory role of B7 homologue 1 (B7-H1) expressed by non-T cells has been established; however, the function of B7-H1 expressed by T cells is not clear. Peak expression of B7-H1 on Ag-primed CD8 T cells was observed during the contraction phase of an immune response. Unexpectedly, B7-H1 blockade at this stage reduced the numbers of effector CD8 T cells, suggesting B7-H1 blocking Ab may disturb an unknown function of B7-H1 expressed by CD8 T cells. To exclusively examine the role of B7-H1 expressed by T cells, we introduced B7-H1 deficiency into TCR transgenic (OT-1) mice. Naive B7-H1–deficient CD8 T cells proliferated normally following Ag stimulation; however, once activated, they underwent more robust contraction in vivo and more apoptosis in vitro. In addition, B7-H1–deficient CD8 T cells were more sensitive to Ca-dependent and Fas ligand-dependent killing by cytotoxic T lymphocytes. Activation-induced Bcl-xL expression was lower in activated B7-H1–deficient CD8 T cells, whereas Bcl-2 and Bim expression were comparable to the wild type. Transfer of effector B7-H1–deficient CD8 T cells failed to suppress tumor growth in vivo. Thus, upregulation of B7-H1 on primed T cells helps effector T cells survive the contraction phase and consequently generate optimal protective immunity.

Abstract Agonists of TLR have been explored as vaccine adjuvants for tumor immunotherapy. However, their immunological consequences are not fully understood. Although TLR signaling increases the functional potential of dendritic cells (DCs) for priming T cells, coinduction of potentially negative immunoregulatory capacities may impair effector T cell generation. We examined the expression and function of B7 family costimulatory molecules on DCs after activation with the TLR3 agonist, polyinosinic:polycytidylic acid. We demonstrated that polyinosinic:polycytidylic acid consistently up-regulated both B7-2 and B7-H1 molecules on resident, migratory DCs from spleen and lymph nodes. Depletion or blockade of B7-H1 on activated DCs increased the magnitude of effector CD8 T cell expansion. DC-based or protein-based tumor vaccines, in combination with B7-H1 blockade, induced strong effector CD8 T cell responses, resulting in protective immunity against newly established tumors. Our studies suggest that TLR3 signaling has the potential to up-regulate both positive and negative coregulatory molecules on APCs. Selective blockade of negative regulatory molecules in combination with TLR3 agonist may be an effective strategy for increasing the efficacy of tumor vaccines.