Pre-existing AAV capsid-specific CD8+ T cells are unable to eliminate AAV-transduced hepatocytes.

The goal of these studies was to test whether adeno-associated virus (AAV) capsid-specific CD8(+) T cells cause loss of hepatic AAV-mediated gene expression in experimental animals. Mice immunized with adenoviral vectors expressing AAV capsid or with AAV vectors developed CD8(+) T cells in blood, lymphatic tissues, and liver to epitopes shared between AAV2 and AAV8, and serotype-specific neutralizing antibodies. At the height of the T cells' effector phase, mice were infused with a heterologous AAV vector expressing human factor IX under a hepatocyte-specific promoter. Despite the presence of lytic CD8(+) T cells in the liver, hepatic Factor IX expression was sustained and comparable in AAV-preimmune and na?ve animals. These results suggest that, in mice, pre-existing CD8(+) T cells to AAV capsid do not affect the longevity of AAV-mediated hepatic gene transfer. These results are in contrast to the outcome of a recent gene therapy trial of hemophilia B patients who were treated by hepatic gene transfer of AAV2 vectors expressing Factor IX. The loss of Factor IX expression, accompanied by a rise in liver enzymes and detectable frequencies of circulating AAV capsid-specific T cells, suggested T-cell-mediated destruction of transduced hepatocytes following reactivation of AAV-specific T cells upon AAV transfer.     

Determination of specific CD4 and CD8 T cell epitopes after AAV2- and AAV8-hF.IX gene therapy.

The application of AAV2 or AAV8 vectors for delivery of human coagulation factor IX (hF.IX) is a promising gene therapy for hemophilia B. One major limitation of this therapy is the development of antibodies and a cytotoxic T lymphocyte (CTL) response against both the vector capsid and the transgene. We determined the class I and class II MHC peptide epitopes for AAV2, AAV8, and hF.IX after administration of AAV-2-hF.IX or AAV8-hF.IX in H2(b) (C57BL/6), H2(d) (BALB/c), or H2(k) (C3H) strains of mice. The results indicate that the AAV2 capsid peptide AA(373-381), the AAV8 capsid peptide AA(50-58), and the hF.IX transgene peptide AA(311-319) can elicit a CTL response as indicated by an IFN-gamma ELISPOT assay and an in vivo CTL assay. Furthermore, a strong H2(k) MHC II-restricted Th1 response can be elicited in C3H mice by the AAV8 capsid peptide AA(126-140) and the hF.IX peptide AA(108-122), whereas a strong Th2 response can be elicited by the AAV2 peptide AA(475-489). These results show that specific CTL responses are generated to both AAV capsid epitopes and hF.IX epitopes after injection of AAV-hF.IX, and MHC class II epitopes derived from AAV-hF.IX promote development of either Th1 or Th2 cells.     

Modulation of CD8+ T cell responses to AAV vectors with IgG-derived MHC class II epitopes

Immune responses directed against viral capsid proteins constitute a main safety concern in the use of adeno-associated virus (AAV) as gene transfer vectors in humans. Pharmacological immunosuppression has been proposed as a solution to the problem; however, the approach suffers from several potential limitations. Using MHC class II epitopes initially identified within human IgG, named Tregitopes, we showed that it is possible to modulate CD8⁺ T cell responses to several viral antigens in vitro. We showed that incubation of peripheral blood mononuclear cells with these epitopes triggers proliferation of CD4⁺CD25⁺FoxP3⁺ T cells that suppress killing of target cells loaded with MHC class I antigens in an antigen-specific fashion, through a mechanism that seems to require cell-to-cell contact. Expression of a construct encoding for the AAV capsid structural protein fused to Tregitopes resulted in reduction of CD8⁺ T cell reactivity against the AAV capsid following immunization with an adenoviral vector expressing capsid. This was accompanied by an increase in frequency of CD4⁺CD25⁺FoxP3⁺ T cells in spleens and lower levels of inflammatory infiltrates in injected tissues. This proof-of-concept study demonstrates modulation of CD8⁺ T cell reactivity to an antigen using regulatory T cell epitopes is possible.     

Endogenous presentation of CD8+ T cell epitopes from Epstein-Barr virus-encoded nuclear antigen 1

Epstein-Barr virus (EBV)-encoded nuclear antigen (EBNA)1 is thought to escape cytotoxic T lymphocyte (CTL) recognition through either self-inhibition of synthesis or by blockade of proteasomal degradation by the glycine-alanine repeat (GAr) domain. Here we show that EBNA1 has a remarkably varied cell type-dependent stability. However, these different degradation rates do not correspond to the level of major histocompatibility complex class I-restricted presentation of EBNA1 epitopes. In spite of the highly stable expression of EBNA1 in B cells, CTL epitopes derived from this protein are efficiently processed and presented to CD8+ T cells. Furthermore, we show that EBV-infected B cells can readily activate EBNA1-specific memory T cell responses from healthy virus carriers. Functional assays revealed that processing of these EBNA1 epitopes is proteasome and transporter associated with antigen processing dependent. We also show that the endogenous presentation of these epitopes is dependent on the newly synthesized protein rather than the long-lived stable EBNA1. Based on these observations, we propose that defective ribosomal products, not the full-length antigen, are the primary source of endogenously processed CD8+ T cell epitopes from EBNA1.     

Reactivity of V beta 17a+ CD8+ T cell hybrids. Analysis using a new CD8+ T cell fusion partner

Tolerance to IE molecules leads to deletion of V beta 17a-bearing T cells. Both, the CD4+ as well as the CD8+ T cell subsets are affected. A large percentage of CD4+ V beta 17a+ T cell hybrids recognize IE molecules. We now have investigated the reactivity for IE antigens of CD8+ V beta 17a+ T cell hybrids. Using a transfection approach, we have introduced the murine CD8 molecule into different V beta 17a+ T cell hybrids. Furthermore, the CD8 cDNA was transfected into the BW5147 alpha-beta- fusion partner. This allowed us to generate a large number of V beta 17a+ T cell hybrids by fusion with the appropriate T cells. Only 6% of T cell hybrids were stimulated to produce IL-2 upon incubation with IE+ cells. However, in those, the CD8 molecule seemed not to contribute to the IE reactivity of the hybrid, since mAbs against the CD8 molecule failed to inhibit their reactivity. This low percentage of V beta 17a+ CD8+ IE-reactive T cell hybrids contrasts with the strong reduction of CD8+ V beta 17a+ T cells in IE+ mice, strongly suggesting that elimination of such cells in the thymus occurs when they are coexpressing CD4 and CD8. This view was confirmed by the occasional expression of CD4 in some hybrids in which case IE reactivity was detected. Furthermore, we demonstrated the functional integrity of the introduced CD8 molecule by: (a) reconstitution of the IL-2 response in a class I-restricted TNP-specific T cell hybrid; and (b) by generation of alloreactive class I-restricted T cell hybrids using the new CD8+ fusion cell line. This CD8+ fusion partner, BWLyt2-4, should prove useful to study antigen processing and antigen presentation requirements of class I-restricted T cells.     

Regulatory CD4+CD25+ T cells restrict memory CD8+ T cell responses

CD4+ T cell help is important for the generation of CD8+ T cell responses. We used depleting anti-CD4 mAb to analyze the role of CD4+ T cells for memory CD8+ T cell responses after secondary infection of mice with the intracellular bacterium Listeria monocytogenes, or after boost immunization by specific peptide or DNA vaccination. Surprisingly, anti-CD4 mAb treatment during secondary CD8+ T cell responses markedly enlarged the population size of antigen-specific CD8+ T cells. After boost immunization with peptide or DNA, this effect was particularly profound, and antigen-specific CD8+ T cell populations were enlarged at least 10-fold. In terms of cytokine production and cytotoxicity, the enlarged CD8+ T cell population consisted of functional effector T cells. In depletion and transfer experiments, the suppressive function could be ascribed to CD4+CD25+ T cells. Our results demonstrate that CD4+ T cells control the CD8+ T cell response in two directions. Initially, they promote the generation of a CD8+ T cell responses and later they restrain the strength of the CD8+ T cell memory response. Down-modulation of CD8+ T cell responses during infection could prevent harmful consequences after eradication of the pathogen.     

CD4+CD25+ T cells regulate virus-specific primary and memory CD8+ T cell responses

Naturally occurring CD4+CD25+ regulatory T cells appear important to prevent activation of autoreactive T cells. This article demonstrates that the magnitude of a CD8+ T cell-mediated immune response to an acute viral infection is also subject to control by CD4+CD25+ T regulatory cells (Treg). Accordingly, if natural Treg were depleted with specific anti-CD25 antibody before infection with HSV, the resultant CD8+ T cell response to the immunodominant peptide SSIEFARL was significantly enhanced. This was shown by several in vitro measures of CD8+ T cell reactivity and by assays that directly determine CD8+ T cell function, such as proliferation and cytotoxicity in vivo. The enhanced responsiveness in CD25-depleted animals was between three- and fourfold with the effect evident both in the acute and memory phases of the immune response. Surprisingly, HSV infection resulted in enhanced Treg function with such cells able to suppress CD8+ T cell responses to both viral and unrelated antigens. Our results are discussed both in term of how viral infection might temporarily diminish immunity to other infectious agents and their application to vaccines. Thus, controlling suppressor effects at the time of vaccination could result in more effective immunity.     

CD8+ T cell tolerance and cancer immunotherapy

To provide protection against all foreign pathogens one can possibly encounter during their lifetime, the T cell repertoire has to be as diverse as possible. At the same time, it is desirable that the T cell repertoire remains unresponsive towards healthy tissues. To realize this self/nonself discriminatory property, T cells undergo tightly controlled selection processes during maturation in the thymus. The key parameter determining the outcome of these selection processes is the avidity of the T cells for self-MHC/self-peptide complexes expressed in the thymus; low avidity interactions result in positive selection, whereas high avidity interactions lead to negative selection. Despite the selection processes, self-tolerance is far from absolute. In many cases, this is due to the presence of self-antigen in the thymus at a level that is too low to induce thymic deletion. In addition, T cells with a low avidity for ubiquitously expressed self-antigens can escape clonal deletion and enter the periphery. A thorough understanding of the self-specific T cell repertoire is important because many potential targets for cancer immunotherapy are self-proteins. In this review, the authors discuss the impact of self-antigen expression on the CD8+ T cell repertoire. An overview of the fate and functional capacities of self-specific T cells with specificity for tissue-restricted self-antigens and for ubiquitously expressed self-antigens is provided. Furthermore, the authors discuss the influence of negative selection on the antitumor reactivity of the mature T cell repertoire.     

Influence of translation efficiency of homologous viral proteins on the endogenous presentation of CD8+ T cell epitopes

A significant proportion of endogenously processed CD8(+) T cell epitopes are derived from newly synthesized proteins and rapidly degrading polypeptides (RDPs). It has been hypothesized that the generation of rapidly degrading polypeptides and CD8(+) T cell epitopes from these RDP precursors may be influenced by the efficiency of protein translation. Here we address this hypothesis by using the Epstein-Barr virus-encoded nuclear antigen 1 protein (EBNA1), with or without its internal glycine-alanine repeat sequence (EBNA1 and EBNA1DeltaGA, respectively), which display distinct differences in translation efficiency. We demonstrate that RDPs constitute a significant proportion of newly synthesized EBNA1 and EBNA1DeltaGA and that the levels of RDPs produced by each of these proteins directly correlate with the translation efficiency of either EBNA1 or EBNA1DeltaGA. As a consequence, a higher number of major histocompatibility complex-peptide complexes can be detected on the surface of cells expressing EBNA1DeltaGA, and these cells are more efficiently recognized by virus-specific cytotoxic T lymphocytes compared to the full-length EBNA1. More importantly, we also demonstrate that the endogenous processing of these CD8(+) T cell epitopes is predominantly determined by the rate at which the RDPs are generated rather than the intracellular turnover of these proteins.     

Identification of H-2Db-specific CD8+ T-cell epitopes from mouse VEGFR2 that can inhibit angiogenesis and tumor growth

Vascular endothelial growth factor receptor 2 (VEGFR2/KDR) plays a crucial role in tumor-associated angiogenesis and vascularization. It has been established that monoclonal antibodies against VEGFR2 can inhibit angiogenesis. In this study, two naturally processed CD8 T-cell epitopes (VILTNPISM and FSNSTNDILI) were identified from murine KDR. Cytotoxic T lymphocytes targeting endothelial cells could be directly monitored by KDR2 and KDR3 Elispots or major histocompatibility complex class I tetramer staining. Immunization with these two peptides effectively reduced angiogenesis and inhibited tumor growth in mouse models. Thus, vaccination with KDR peptides alone or in combination with other anti-angiogenesis agents may afford a novel immunotherapy for inhibition of tumor growth.     

Viral vectors for inducing CD8+ T cell responses

CD8⁺ T cells (T_CD8+) can mediate protective immunity to intracellular pathogens and tumours. Viruses generate strong T_CD8+ responses and, therefore, represent attractive vectors for generating vaccines aimed at producing T_CD8+-mediated protective immunity. This review will examine the immunological properties of viruses that make them good candidates as vaccine vectors, as well as the manipulations of both vector and antigen that may be required to produce an effective vaccine. The areas addressed include virus infection of dendritic cells in vivo, stimulation of the innate immune response via intracellular and extracellular pattern recognition receptors, the effect of antigenic form on the pathways of antigen presentation and the requirement for elimination of viral genes that target various aspects of the innate and adaptive immune response.     

One naive T cell,multiple fates in CD8+ T cell differentiation

The mechanism by which the immune system produces effector and memory T cells is largely unclear. To allow a large-scale assessment of the development of single naive T cells into different subsets, we have developed a technology that introduces unique genetic tags (barcodes) into naive T cells. By comparing the barcodes present in antigen-specific effector and memory T cell populations in systemic and local infection models, at different anatomical sites, and for TCR–pMHC interactions of different avidities, we demonstrate that under all conditions tested, individual naive T cells yield both effector and memory CD8⁺ T cell progeny. This indicates that effector and memory fate decisions are not determined by the nature of the priming antigen-presenting cell or the time of T cell priming. Instead, for both low and high avidity T cells, individual naive T cells have multiple fates and can differentiate into effector and memory T cell subsets.Activation of naive antigen-specific T cells is characterized by a vigorous proliferative burst, resulting in the formation of a large pool of effector T cells. After pathogen clearance, ∼95% of activated T cells die, leaving behind a stable pool of long-lived memory cells (Williams and Bevan, 2007). Two fundamentally different mechanisms could give rise to the production of effector and memory T cells during an immune response. First, single naive T cells may be destined to produce either effector T cells or memory T cells, but not both (“one naive cell, one fate”). As an alternative, effector and memory T cells could derive from the same clonal precursors within the naive T cell pool (“one naive cell, multiple fates”). As the fate decisions that control T cell differentiation could either be taken during initial T cell priming (i.e., before the first cell division) or at later stages, at least four conceptually different models describing effector and memory T cell differentiation can be formulated (Fig. S1).A first model predicts a separate origin of effector and memory T cells as a result of differential T cell priming by APCs. In this scenario, fate decisions would be taken before the first cell division, and even though cells destined to become memory cells may transiently display traits associated with effector T cells (e.g., expression of granzyme B or IFN-γ; see the following paragraphs), their ability for long-term survival would be predetermined. In line with this model, several studies have provided evidence that the fate of CD8⁺ T cells may, to some extent, be programmed during initial activation (Kaech and Ahmed, 2001; van Stipdonk et al., 2003; Masopust et al., 2004; Williams and Bevan, 2007; Bannard et al., 2009).A second model, which relies on recent data from Chang et al. (2007), likewise suggests that the priming APC plays the crucial role in determining effector or memory T cell fate, but by a strikingly different mechanism and with an opposite prediction concerning the lineage relationship of effector and memory T cells. Specifically, analysis of T cell–APC conjugates has shown that the first division of activated T cells can be asymmetric, with the daughter T cell that is formed proximal to the APC being more likely to contribute to the effector T cell subset and the distal daughter T cell being more likely to generate memory T cells (Chang et al., 2007). Assuming that all primary daughter cells survive and yield further progeny, these data would predict that single naive T cells contribute to both the effector and the memory subset.In contrast to these two models that are based on a determining role of the priming APC, two other models predict that T cell fate is determined by the cumulative effect of signals that not only naive T cells but also their descendants receive. The first of these models, termed the “decreasing potential model,” argues that T cell progeny that receive additional stimulation after priming undergo terminal differentiation toward the effector subset, whereas descendants that do not encounter these signals may transiently display certain effector functions but will ultimately become memory T cells (Ahmed and Gray, 1996). In support of this model, it has been demonstrated that continued inflammatory signals (Badovinac et al., 2004; Joshi et al., 2007) and prolonged antigenic stimulation (Sarkar et al., 2008) can lead descendant CD8⁺ T cells to preferentially develop into effector cells.If the descendants of all individual naive T cells have an equal chance of receiving signals for terminal differentiation, the standard decreasing potential model predicts that memory and effector T cells will be derived from the same population of naive T cells. However, there is evidence that the environmental factors that promote either terminal differentiation or memory T cell development may alter over the course of infection (Sarkar et al., 2008). A fourth model therefore argues that the progeny of T cells that are activated early or late during infection will receive distinct signals and, hence, assume (partially) different fates (van Faassen et al., 2005; D’Souza and Hedrick, 2006; Quigley et al., 2007; Stemberger et al., 2007a).A large number of studies in which cell differentiation was analyzed at the population level have been informative in revealing which effector properties can be displayed by T cells that subsequently differentiate into memory T cells (for review see Jameson and Masopust, 2009). In particular, two recent studies using IFN-γ or granzyme B reporter mice have shown that memory T cells can arise from cells that have previously transcribed IFN-γ or granzyme B genes (Harrington et al., 2008; Bannard et al., 2009). However, it is important to realize that these studies reveal little with regard to the developmental potential of individual naive T cells. Specifically, the fact that T cells that have a particular effector capacity can become memory T cells does not indicate that all naive T cells yield such effector cells, nor does it indicate that all memory T cells have gone through an effector phase.To determine the developmental potential of naive T cells, it is essential to develop technologies in which T cell responses can be analyzed at the single naive T cell level. In early work that aimed to follow T cell responses at the clonal level, TCR repertoire analysis has been used to assess the kinship of T cell populations (Maryanski et al., 1996; Kedzierska et al., 2004). However, as several naive T cell clones can share the same TCR, it has been argued that such analyses do not necessarily monitor T cell fate at the single T cell level (Stemberger et al., 2007b; Obar et al., 2008). Recently, Stemberger et al. (2007a) have reported on a more elegant approach to address naive T cell potency. Using the transfer of single naive CD8⁺ T cells into mice, this study provides direct evidence that a single naive CD8⁺ T cell can form both effector and memory cell subsets. However, the statistical power of single-cell transfer studies obviously has limitations. In addition, if homeostatic proliferation would occur before antigen-driven proliferation in this system, this would limit the conclusions that can be drawn with regard to the pluripotency of a single naive T cell.In this study, we have developed a technology that allows the generation of naive T cells that carry unique genetic tags (barcodes), and we describe how this technology can be used for the large-scale assessment of the developmental potential of single naive T cells. Using physiological frequencies of barcode-labeled naive CD8⁺ T cells of different functional avidities, we demonstrate that in both systemic and local infection models, effector and memory CD8⁺ T cell subsets share the same precursors in the naive T cell pool. These data demonstrate that under all conditions analyzed, single naive T cells do not selectively yield effector or memory T cells. Rather, T cell differentiation into effector and memory T cell subsets occurs by a one naive cell, multiple fates principle.     

Viral vectors for inducing CD8+ T cell responses

CD8(+) T cells (T(CD8+)) can mediate protective immunity to intracellular pathogens and tumours. Viruses generate strong T(CD8+) responses and, therefore, represent attractive vectors for generating vaccines aimed at producing T(CD8+)-mediated protective immunity. This review will examine the immunological properties of viruses that make them good candidates as vaccine vectors, as well as the manipulations of both vector and antigen that may be required to produce an effective vaccine. The areas addressed include virus infection of dendritic cells in vivo, stimulation of the innate immune response via intracellular and extracellular pattern recognition receptors, the effect of antigenic form on the pathways of antigen presentation and the requirement for elimination of viral genes that target various aspects of the innate and adaptive immune response.     

Essential roles of CD8+CD122+ regulatory T cells in the maintenance of T cell homeostasis

Regulation of immune system is of paramount importance to prevent immune attacks against self-components. Mice deficient in the interleukin (IL)-2/IL-15 receptor beta chain, CD122, are model animals of such immune attacks and characteristically have a high number of abnormally activated T cells. Here, we show that the transfer of CD8+CD122+ cells into CD122-deficient neonates totally prevented the development of abnormal T cells. Furthermore, recombination activating gene-2-/- mice that received wild-type mice-derived CD8+CD122- cells died within 10 wk after cell transfer, indicating that normal CD8+CD122- cells become dangerously activated T cells in the absence of CD8+CD122+ T cells. CD8+CD122+ cells could control activated CD8+ or CD4+ T cells both in vivo and in vitro. Our results indicate that the CD8+CD122+ population includes naturally occurring CD8+ regulatory T cells that control potentially dangerous T cells.     

Both CD8+ and CD16+ human T cell clones can provide B cell help for immunoglobulin production.

The functional properties of two CD4+CD8-CD16-, five CD4-CD8+CD16- and three CD4-CD8-CD16+ human T cell clones were compared. All CD4- T cell clones displayed strong cytolytic activity in the lectin-dependent lytic assay against the P815 murine mastocytoma cell line, but only the CD4-CD8-CD16+ T cell clones exhibited lytic activity against the natural killer-sensitive K562 cell line. Upon activation with anti-CD3 monoclonal antibody, all T cell clones were able to support IgM and IgA synthesis in autologous B cells. Both CD4+ and CD4- T cell clones required cell-to-cell interaction with the B cells in order to exert their helper activity for immunoglobulin production. However, unlike CD4+, CD4-CD8+CD16- and CD4-CD8-CD16+ T cell clones provided helper function for immunoglobulin synthesis only when low T/B cell ratios were used in culture. At higher T/B cell ratios, there was a decline in the B cell helper activity of CD4- T cell clones that was probably related to the expression of cytolytic capacity against the antigen-presenting B cell. These data support the notion that under certain experimental conditions even cytotoxic T lymphocytes and natural killer cells may provide B cell helper function.     

TCR stimulation with modified anti-CD3 mAb expands CD8+ T cell population and induces CD8+CD25+ Tregs

Modified anti-CD3 mAbs are emerging as a possible means of inducing immunologic tolerance in settings including transplantation and autoimmunity such as in type 1 diabetes. In a trial of a modified anti-CD3 mAb [hOKT3gamma1(Ala-Ala)] in patients with type 1 diabetes, we identified clinical responders by an increase in the number of peripheral blood CD8+ cells following treatment with the mAb. Here we show that the anti-CD3 mAb caused activation of CD8+ T cells that was similar in vitro and in vivo and induced regulatory CD8+CD25+ T cells. These cells inhibited the responses of CD4+ cells to the mAb itself and to antigen. The regulatory CD8+CD25+ cells were CTLA4 and Foxp3 and required contact for inhibition. Foxp3 was also induced on CD8+ T cells in patients during mAb treatment, which suggests a potential mechanism of the anti-CD3 mAb immune modulatory effects involving induction of a subset of regulatory CD8+ T cells.     

CD30 expression by CD8+ T cells producing type 2 helper cytokines. Evidence for large numbers of CD8+CD30+ T cell clones in human immunodeficiency virus infection

A large panel of CD8+ T cell clones generated from peripheral blood lymphocytes (PBL) of healthy donors or human immunodeficiency virus (HIV)-infected individuals were assessed for both cytokine secretion profile and CD30 expression and release. The great majority of CD8+ T cell clones generated from healthy individuals showed the ability to produce interferon gamma (IFN-gamma), but not interleukin 4 (IL-4), and none of them either expressed membrane CD30 or released substantial amounts of soluble CD30 (sCD30) in their supernatant. In contrast, high numbers of CD8+ T cell clones generated from HIV-infected individuals, which produced IL-4 (and IL-5) in addition to IFN-gamma or IL-4 (and IL- 5) alone, expressed membrane CD30 and released detectable amounts of sCD30 in their supernatants. Indeed, CD30 expression appeared to be positively correlated with the ability of CD8+ T cell clones to produce IL-4 and IL-5 and inversely correlated with their ability to produce IFN-gamma, whereas no correlation between CD30 expression and production of IL-10 was observed. These data suggest that CD30 is a marker for CD8+ T cells that have switched to the production of type 2 helper cytokines.     

Autoimmune melanocyte destruction is required for robust CD8+ memory T cell responses to mouse melanoma

A link between autoimmunity and improved antitumor immunity has long been recognized, although the exact mechanistic relationship between these two phenomena remains unclear. In the present study we have found that vitiligo, the autoimmune destruction of melanocytes, generates self antigen required for mounting persistent and protective memory CD8+ T cell responses to melanoma. Vitiligo developed in approximately 60% of mice that were depleted of regulatory CD4+ T cells and then subjected to surgical excision of large established B16 melanomas. Mice with vitiligo generated 10-fold larger populations of CD8+ memory T cells specific for shared melanoma/melanocyte antigens. CD8+ T cells in mice with vitiligo acquired phenotypic and functional characteristics of effector memory, suggesting that they were supported by ongoing antigen stimulation. Such responses were not generated in melanocyte-deficient mice, indicating a requirement for melanocyte destruction in maintaining CD8+ T cell immunity to melanoma. Vitiligo-associated memory CD8+ T cells provided durable tumor protection, were capable of mounting a rapid recall response to melanoma, and did not demonstrate phenotypic or functional signs of exhaustion even after many months of exposure to antigen. This work establishes melanocyte destruction as a key determinant of lasting melanoma-reactive immune responses, thus illustrating that immune-mediated destruction of normal tissues can perpetuate adaptive immune responses to cancer.