Peter A. Savage

Characterization of circulating T cells specific for tumor- associated antigens in melanoma patients

We identified circulating CD8+ T-cell populations specific for the tumor-associated antigens (TAAs) MART-1 (27-35) or tyrosinase (368-376) in six of eleven patients with metastatic melanoma using peptide/HLA-A*0201 tetramers. These TAA-specific populations were of two phenotypically distinct types: one, typical for memory/effector T cells; the other, a previously undescribed phenotype expressing both naive and effector cell markers. This latter type represented more than 2% of the total CD8+ T cells in one patient, permitting detailed phenotypic and functional analysis. Although these cells have many of the hallmarks of effector T cells, they were functionally unresponsive, unable to directly lyse melanoma target cells or produce cytokines in response to mitogens. In contrast, CD8+ T cells from the same patient were able to lyse EBV-pulsed target cells and showed robust allogeneic responses. Thus, the clonally expanded TAA-specific population seems to have been selectively rendered anergic in vivo. Peptide stimulation of the TAA-specific T-cell populations in other patients failed to induce substantial upregulation of CD69 expression, indicating that these cells may also have functional defects, leading to blunted activation responses. These data demonstrate that systemic TAA-specific T-cell responses can develop de novo in cancer patients, but that antigen-specific unresponsiveness may explain why such cells are unable to control tumor growth.

A Kinetic Basis For T Cell Receptor Repertoire Selection during an Immune Response

The basis for T cell antigen receptor (TCR) repertoire selection upon repeated antigenic challenge is unclear. We evaluated the avidity and dissociation kinetics of peptide/major histocompatibility complex (MHC) tetramer binding to antigen-specific T lymphocytes isolated following primary or secondary immunization. The data reveal a narrowing of the secondary repertoire relative to the primary repertoire, largely resulting from the loss of cells expressing TCRs with the fastest dissociation rates for peptide/MHC binding. In addition, T cells in the secondary response express TCRs of higher average affinity for peptide/MHC than cells in the primary response. These results provide a link between the kinetics and affinity of TCR-peptide/MHC interactions and TCR sequence selection during the course of an immune response.

Aire-Dependent Thymic Development of Tumor-Associated Regulatory T Cells

On the Origin of Tumor Tregs The tumor microenvironment is often seeded with regulatory T cells (Tregs), which inhibit antitumor immunity. Using mice with genetically driven prostate cancer, Malchow et al. (p. 1219; see the Perspective by Joshi and Jacks) found a population of Tregs that were enriched in the prostate of tumor-bearing mice. Surprisingly, these cells were also present in female mice and were found to be specific, not for a tumor-specific antigen, but rather for an antigen normally expressed in the prostate. Prostate antigen–specific Tregs arose in the thymus and their selection was dependent on Aire, a protein that drives the expression of tissue-specific antigens in the thymus. Thus, Tregs that seed tumors likely arise in the thymus, are not necessarily tumor-specific, and are recruited and/or expand in an organ when a tumor arises. Prostate tumors in mice recruit thymus-derived regulatory T cells that are specific for tissue autoantigens. [Also see Perspective by Joshi and Jacks] Despite considerable interest in the modulation of tumor-associated Foxp3+ regulatory T cells (Tregs) for therapeutic benefit, little is known about the developmental origins of these cells and the nature of the antigens that they recognize. We identified an endogenous population of antigen-specific Tregs (termed MJ23 Tregs) found recurrently enriched in the tumors of mice with oncogene-driven prostate cancer. MJ23 Tregs were not reactive to a tumor-specific antigen but instead recognized a prostate-associated antigen that was present in tumor-free mice. MJ23 Tregs underwent autoimmune regulator (Aire)–dependent thymic development in both male and female mice. Thus, Aire-mediated expression of peripheral tissue antigens drives the thymic development of a subset of organ-specific Tregs, which are likely coopted by tumors developing within the associated organ.

Recognition of a Ubiquitous Self Antigen by Prostate Cancer-Infiltrating CD8+ T Lymphocytes

Substantial evidence exists that many tumors can be specifically recognized by CD8+ T lymphocytes. The definition of antigens targeted by these cells is paramount for the development of effective immunotherapeutic strategies for treating human cancers. In a screen for endogenous tumor-associated T cell responses in a primary mouse model of prostatic adenocarcinoma, we identified a naturally arising CD8+ T cell response that is reactive to a peptide derived from histone H4. Despite the ubiquitous nature of histones, T cell recognition of histone H4 peptide was specifically associated with the presence of prostate cancer in these mice. Thus, the repertoire of antigens recognized by tumor-infiltrating T cells is broader than previously thought and includes peptides derived from ubiquitous self antigens that are normally sequestered from immune detection.

Basic principles of tumor-associated regulatory T cell biology

Due to the critical role of forkhead box (Fox)p3(+) regulatory T cells (Tregs) in the regulation of immunity and the enrichment of Tregs within many human tumors, several emerging therapeutic strategies for cancer involve the depletion or modulation of Tregs, with the aim of eliciting enhanced antitumor immune responses. Here, we review recent advances in understanding of the fundamental biology of Tregs, and discuss the implications of these findings for current models of tumor-associated Treg biology. In particular, we discuss the context-dependent functional diversity of Tregs, the developmental origins of these cells, and the nature of the antigens that they recognize within the tumor environment. In addition, we highlight critical areas of focus for future research.

PD-1 regulates extrathymic regulatory T-cell differentiation

Treg cells and the programed death‐1/programed death ligand‐1 (PD‐1/PD‐L1) pathway are both critical for maintaining peripheral tolerance to self‐Ags. A significant subset of Treg cells constitutively expresses PD‐1, which prompted an investigation into the role of PD‐1/PD‐L1 interactions in Treg‐cell development, function, and induction in vivo. The phenotype and abundance of Treg cells was not significantly altered in PD‐1‐deficient mice. The thymic development of polyclonal and monospecific Treg cells was not negatively impacted by PD‐1 deficiency. The suppressive function of PD‐1−/− Treg cells was similar to their PD‐1+/+ counterparts both in vitro and in vivo. However, in three different in vivo experimental settings, PD‐1−/− conventional CD4+ T cells demonstrated a strikingly diminished tendency toward differentiation into peripherally induced Treg (pTreg) cells. Our results demonstrate that PD‐1 is dispensable for thymic Treg‐cell development and suppressive function, but is critical for the extrathymic differentiation of pTreg cells in vivo. These data suggest that Ab blockade of the PD‐1/PD‐L1 pathway may augment T‐cell responses by acting directly on conventional T cells, and also by suppressing the differentiation of pTreg cells.

Dendritic Cells Coordinate the Development and Homeostasis of Organ-Specific Regulatory T Cells

Although antigen recognition mediated by the T cell receptor (TCR) influences many facets of Foxp3(+) regulatory T (Treg) cell biology, including development and function, the cell types that present antigen to Treg cells in vivo remain largely undefined. By tracking a clonal population of Aire-dependent, prostate-specific Treg cells in mice, we demonstrated an essential role for dendritic cells (DCs) in regulating organ-specific Treg cell biology. We have shown that the thymic development of prostate-specific Treg cells required antigen presentation by DCs. Moreover, Batf3-dependent CD8α(+) DCs were dispensable for the development of this clonotype and had negligible impact on the polyclonal Treg cell repertoire. In the periphery, CCR7-dependent migratory DCs coordinated the activation of organ-specific Treg cells in the prostate-draining lymph nodes. Our results demonstrate that the development and peripheral regulation of organ-specific Treg cells are dependent on antigen presentation by DCs, implicating DCs as key mediators of organ-specific immune tolerance.

Shaping the repertoire of tumor-infiltrating effector and regulatory T cells.

Many tumors express antigens that can be specifically or selectively recognized by T lymphocytes, suggesting that T‐cell‐mediated immunity may be harnessed for the immunotherapy of cancer. However, since tumors originate from normal cells and evolve within the context of self‐tissues, the immune mechanisms that prevent the autoimmune attack of normal tissues function in parallel to restrict anti‐tumor immunity. In particular, the purging of autoreactive T cells and the development of immune‐suppressive regulatory T cells (Tregs) are thought to be major barriers impeding anti‐tumor immune responses. Here, we discuss current understanding regarding the antigens recognized by tumor‐infiltrating T‐cell populations, the mechanisms that shape the repertoire of these cells, and the role of the transcription factor autoimmune regulator (Aire) in these processes. Further elucidation of these principles is likely to be critical for optimizing emerging cancer immunotherapies, and for the rational design of novel therapies exhibiting robust anti‐tumor activity with limited toxicity.

Recipient NK cell inactivation and intestinal barrier loss are required for MHC-matched graft-versus-host disease

Pretransplant conditioning promotes minor mismatch GVHD by causing intestinal barrier loss and depleting NK cells. Double Trouble Before Transplant Graft-versus-host disease, a condition where transplanted immune cells attack the body of the transplant recipient, is a common complication of bone marrow or stem cell transplant. Although there are treatments available for treating graft-versus-host disease, they don’t always work well. Nalle et al. have now developed several different mouse models of this condition to provide insight into its causes. The authors used these mouse models to pinpoint two different ways that pretransplant conditioning contributes to graft-versus-host disease. This knowledge should help scientists search for new treatments against this disease, as well as identify safer ways to prepare patients for transplant. Previous studies have shown a correlation between pretransplant conditioning intensity, intestinal barrier loss, and graft-versus-host disease (GVHD) severity. However, because irradiation and other forms of pretransplant conditioning have pleiotropic effects, the precise role of intestinal barrier loss in GVHD pathogenesis remains unclear. We developed GVHD models that allowed us to isolate the specific contributions of distinct pretransplant variables. Intestinal damage was required for the induction of minor mismatch [major histocompatibility complex (MHC)–matched] GVHD, but was not necessary for major mismatch GVHD, demonstrating fundamental pathogenic distinctions between these forms of disease. Moreover, recipient natural killer (NK) cells prevented minor mismatch GVHD by limiting expansion and target organ infiltration of alloreactive T cells via a perforin-dependent mechanism, revealing an immunoregulatory function of MHC-matched recipient NK cells in GVHD. Minor mismatch GVHD required MyD88-mediated Toll-like receptor 4 (TLR4) signaling on donor cells, and intestinal damage could be bypassed by parenteral lipopolysaccharide (LPS) administration, indicating a critical role for the influx of bacterial components triggered by intestinal barrier loss. In all, the data demonstrate that pretransplant conditioning plays a dual role in promoting minor mismatch GVHD by both depleting recipient NK cells and inducing intestinal barrier loss.

Immune reconstitution after combined haploidentical and umbilical cord blood transplant

Abstract Umbilical cord blood (UCB) stem cells are frequently employed for allogeneic stem cell transplant, but delayed myeloid and lymphoid immune reconstitution leads to increased risk of infections. We recently reported the clinical results of 45 patients enrolled on a pilot study combining UCB with a human leukocyte antigen (HLA)-haploidentical donor with reduced-intensity conditioning who showed rapid neutrophil and platelet recovery. We report here preliminary immune reconstitution data of these patients. Patients were assessed for lymphocyte subsets, T-cell diversity, Cylex ImmuKnow assay and serological response to pneumococcal vaccination. Natural killer (NK)-cell and B-cell reconstitution were rapid at 1 month and 3 months, respectively. T-cell recovery was delayed, with a gradual increase in the number of T-cells starting around 6 months post-transplant, and was characterized by a diverse polyclonal T-cell repertoire. Overall, immune reconstitution after haplo-cord transplant is similar to that seen after cord blood transplant, despite infusion of much lower cord blood cell dose.