Pulmonary tumors inefficiently prime tumor-specific T cells

The lung is a common site of metastatic and primary tumor growth, and has been shown to be an immunosuppressive environment. We tested the impact of the lung environment on the development of tumor-specific T cell responses against the CMS5 fibrosarcoma, and found a deficit in the efficacy of naive tumor-specific DUC18 T cells against tumors established in the lung. One hundred-fold more naive tumor-specific T cells were required to protect against tumor development or reject established tumors in the lung than an identical tumor challenge delivered s.c. in the flank. Importantly, CMS5 growing in the flank facilitated the rejection of tumors present in the lungs. In the presence of flank tumors, transferred T cells were not phenotypically altered but were present in much greater numbers in the parabronchial lymph nodes, bronchoalveolar lavage, and lung parenchyma than in mice bearing lung tumors alone. We hypothesized that APC present in the lung and skin draining lymph nodes were differentially initiating T cell proliferation, leading to differences in the size of the final effector populations. A direct comparison of DUC18 T cell proliferation against APC from flank or lung draining lymph nodes showed profoundly greater proliferation to flank draining lymph node APC. The impaired stimulation of naive T cell proliferation by lung draining APC provides one mechanistic explanation for the lower overall immune response, and inability to effectively reject tumors, in the lung.     

Phenotype and homing of CD4 tumor-specific T cells is modulated by tumor bulk

Technical difficulties in tracking endogenous CD4 T lymphocytes have limited the characterization of tumor-specific CD4 T cell responses. Using fluorescent MHC class II/peptide multimers, we defined the fate of endogenous Leishmania receptor for activated C kinase (LACK)-specific CD4 T cells in mice bearing LACK-expressing TS/A tumors. LACK-specific CD44(high)CD62L(low) CD4 T cells accumulated in the draining lymph nodes and had characteristics of effector cells, secreting IL-2 and IFN-gamma upon Ag restimulation. Increased frequencies of CD44(high)CD62L(low) LACK-experienced cells were also detected in the spleen, lung, liver, and tumor itself, but not in nondraining lymph nodes, where the cells maintained a naive phenotype. The absence of systemic redistribution of LACK-specific memory T cells correlated with the presence of tumor. Indeed, LACK-specific CD4 T cells with central memory features (IL-2(+)IFN-gamma(-)CD44(high)CD62L(high) cells) accumulated in all peripheral lymph nodes of mice immunized with LACK-pulsed dendritic cells and after tumor resection. Together, our data demonstrate that although tumor-specific CD4 effector T cells producing IFN-gamma are continuously generated in the presence of tumor, central memory CD4 T cells accumulate only after tumor resection. Thus, the continuous stimulation of tumor-specific CD4 T cells in tumor-bearing mice appears to hinder the systemic accumulation of central memory CD4 T lymphocytes.     

The kinetics of in vivo priming of CD4 and CD8 T cells by dendritic/tumor fusion cells in MUC1-transgenic mice

Previous work has demonstrated that dendritic/tumor fusion cells induce potent antitumor immune responses in vivo and in vitro. However, little is known about the migration and homing of fusion cells after s.c. injection or the kinetics of CD4+ and CD8+ T cell activation. In the present study, fluorescence-labeled dendritic/MUC1-positive tumor fusion cells (FC/MUC1) were injected s.c. into MUC1-transgenic mice. The FC/MUC1 migrated to draining lymph nodes and were closely associated with T cells in a pattern comparable with that of unfused dendritic cells. Immunization of MUC1-transgenic mice with FC/MUC1 resulted in proliferation of T cells and induced MUC1-specific CD8+ CTL. Moreover, CD4+ T cells activated by FC/MUC1 were multifunctional effectors that produced IL-2, IFN-gamma, IL-4, and IL-10. These findings indicate that both CD4+ and CD8+ T cells can be primed in vivo by FC/MUC1 immunization.     

T and B lymphocyte migration into syngeneic tumors.

The migration of splenic T and B lymphocytes into syngeneic tumors undergoing immunologic rejection was investigates. Spleen cells were obtained from normal BALC/c mice or BALB/c mice bearing tumors induced by murine sarcoma virus (MSV). Either whole spleen cells or immunoabsorbent purified T and B cells were radiolabeled with sodium chromate-51 and injected i.v. into normal or MSV inducted-tumor bearing syngeneic recipients. Twenty-four hours later the recipient mice were sacrificed and radioactivity was assessed for tumor, contralateral normal muscle, the lymph nodes draining the tumor and contralateral draining lymph nodes, peripheral lymph nodes, spleen, and liver. Both T and B lymphocytes from either normal or MSV tumor-bearing animals show greatly increased migration into the tumor when compared with normal muscle. Migration of T cells from both normal and MSV tumor bearers was 30 times that of migration to normal muscle. B cells from tumor-bearing mice, on the other hand, localized in the tumor itself only 50% as frequently as did B cells from normal animals. In addition, T cells from MSV tumor bearers were found in the highest proportion in the lymph node draining the tumor site. We conclude that T and B lymphocytes from either normal or tumor-bearing mice migrate to a syngeneic tumor undergoing immunologic rejection. In contrast, the migration of both T and B cells from tumor-bearing animals was decreased to the peripheral lymph nodes at the time of maximum tumor growth.     

CD40 ligand promotes priming of fully potent antitumor CD4(+) T cells in draining lymph nodes in the presence of apoptotic tumor cells.

The presence or absence of CD4(+) T cell help can determine the direction of adaptive immune responses toward either cross-priming or cross-tolerance. It has been demonstrated that interactions of CD40-CD40 ligand can replace CD4(+) T cell help and enable dendritic cells to prime cytotoxic T cells. Here, we demonstrate that antitumor reactivity induced in regional lymph nodes (LNs) by s.c. injection of CD40 ligand (CD40L)-transduced tumor (MCA205 CD40L) showed far superior therapeutic efficacy against established brain tumors of a weakly immunogenic fibrosarcoma, MCA205, when adoptively transferred. Coinjection of apoptotic, but not necrotic parental tumor cells with CD40L-expressing tumor cells caused a strong synergistic induction of antitumor reactivity in tumor-draining LNs. Freshly isolated T cells from LNs immunized with apoptotic parental tumor cells and MCA205 CD40L were capable of mediating regression of the parental tumor in vivo. In contrast, T cells derived from LNs immunized without MCA205 CD40L required ex vivo anti-CD3/IL-2 activation to elicit therapeutic activity. On anti-CD3/IL-2 activation, cells from LNs immunized with MCA205 CD40L exhibited superior per cell antitumor reactivity. An in vitro depletion study revealed that either CD4(+) or CD8(+) T cells could mediate therapeutic efficacy but that the antitumor efficacy mediated by CD4(+) T cells was far superior. Cytosolic flow cytometric analyses indicated that priming of CD4(+) cells in LNs draining CD40L-expressing tumors was polarized to the Th1 type. This is the first report that fully potent antitumor CD4(+) T cell priming was promoted by s.c. injection of CD40L-transduced tumor in the presence of apoptotic tumor cells.     

Identification of identical TCRs in primary melanoma lesions and tumor free corresponding sentinel lymph nodes

It is generally believed that priming of efficient T-cell responses takes place in peripheral lymphoid tissues. Although this notion has been rigidly proven for infectious diseases, direct evidence for lymph node priming of in vivo T-cell responses against tumors is still lacking. In the present study, we conducted a full and nonbiased comparison of T-cell clonotypes in melanoma lesions and corresponding sentinel lymph nodes. Whereas most tumor lesions comprised a high number of T-cell clonotypes, only a small number of clonally expanded T cells were detected in the draining lymph nodes. Comparative clonotype mapping demonstrated the presence of identical T-cell clonotypes in the tumors and the respective sentinel lymph nodes, only when tumor cells were present in the latter. However, taking advantage of clonotype specific PCR amplification, TCR sequences representing clonally expanded T cells at the tumor site could be detected in the lymph nodes draining the tumors even in the absence of tumor cells. Evidence for the tumor-specific characteristics of these cells was obtained by in situ staining with peptide/HLA class I complexes demonstrating the presence of MART-1/HLA-A2- and MAGE-3/HLA-A2-reactive T cells at the tumor site, as well as in the draining lymph node. Our data indicate that T-cell responses to melanoma are primed in the sentinel lymph node by cross presentation of tumor antigens by dendritic cells.     

Val-BoroPro Accelerates T Cell Priming via Modulation of Dendritic Cell Trafficking Resulting in Complete Regression of Established Murine Tumors

Although tumors naturally prime adaptive immune responses, tolerance may limit the capacity to control progression and can compromise effectiveness of immune-based therapies for cancer. Post-proline cleaving enzymes (PPCE) modulate protein function through N-terminal dipeptide cleavage and inhibition of these enzymes has been shown to have anti-tumor activity. We investigated the mechanism by which Val-boroPro, a boronic dipeptide that inhibits post-proline cleaving enzymes, mediates tumor regression and tested whether this agent could serve as a novel immune adjuvant to dendritic cell vaccines in two different murine syngeneic murine tumors. In mice challenged with MB49, which expresses the HY antigen complex, T cell responses primed by the tumor with and without Val-boroPro were measured using interferon gamma ELISPOT. Antibody depletion and gene-deficient mice were used to establish the immune cell subsets required for tumor regression. We demonstrate that Val-boroPro mediates tumor eradication by accelerating the expansion of tumor-specific T cells. Interestingly, T cells primed by tumor during Val-boroPro treatment demonstrate increased capacity to reject tumors following adoptive transfer without further treatment of the recipient. Val-boroPro -mediated tumor regression requires dendritic cells and is associated with enhanced trafficking of dendritic cells to tumor draining lymph nodes. Finally, dendritic cell vaccination combined with Val-boroPro treatment results in complete regression of established tumors. Our findings demonstrate that Val-boroPro has antitumor activity and a novel mechanism of action that involves more robust DC trafficking with earlier priming of T cells. Finally, we show that Val-boroPro has potent adjuvant properties resulting in an effective therapeutic vaccine.     

T-cell death and cancer immune tolerance

Cancer patients mount adaptive immune responses against their tumors. However, tumor develops many mechanisms to evade effective immunosurveillance. T-cell death caused by tumor plays a critical role in establishing tumor immunotolerance. Chronic stimulation of T cells by tumors leads to activation-induced cell death. Abortive stimulation of T cells by tolerogenic antigen-presenting cells loaded with tumor antigens leads to autonomous death of tumor-specific T cells. Therapeutic approaches that prevent T-cell death in the tumor microenvironment and tumor draining lymph nodes, therefore, should boost adaptive immune responses against cancer.     

NK and CD4 cells collaborate to protect against melanoma tumor formation in the brain

NK cells represent a potent immune effector cell type that have the ability to recognize and lyse tumors. However, the existence and function of NK cells in the traditionally "immune-privileged" CNS is controversial. Furthermore, the cellular interactions involved in NK cell anti-CNS tumor immunity are even less well understood. We administered non-Ag-loaded, immature dendritic cells (DC) to CD8alpha knockout (KO) mice and studied their anti-CNS tumor immune responses. DC administration induced dramatic antitumor immune protection in CD8alpha KO mice that were challenged with B16 melanoma both s.c. and in the brain. The CNS antitumor immunity was dependent on both CD4+ T cells and NK cells. Administration of non-Ag-loaded, immature DC resulted in significant CD4+ T cell and NK cell expansion in the draining lymph nodes at 6 days postvaccination, which persisted for 2 wk. Finally, DC administration in CD8alpha KO mice was associated with robust infiltration of CD4+ T cells and NK cells into the brain tumor parenchyma. These results represent the first demonstration of a potent innate antitumor immune response against CNS tumors in the absence of toxicity. Thus, non-Ag-loaded, immature DC administration, in the setting of CD8 genetically deficient mice, can induce dramatic antitumor immune responses within the CNS that surpass the effects observed in wild-type mice. Our results suggest that a better understanding of the cross-talk between DC and innate immune cells may provide improved methods to vaccinate patients with tumors located both systemically and within the CNS.     

Activation of dendritic cells that cross-present tumor-derived antigen licenses CD8+ CTL to cause tumor eradication

The fate of naive CD8(+) T cells is determined by the environment in which they encounter MHC class I presented peptide Ags. The manner in which tumor Ags are presented is a longstanding matter of debate. Ag presentation might be mediated by tumor cells in tumor draining lymph nodes or via cross-presentation by professional APC. Either pathway is insufficient to elicit protective antitumor immunity. We now demonstrate using a syngeneic mouse tumor model, expressing an Ag derived from the early region 1A of human adenovirus type 5, that the inadequate nature of the antitumor CTL response is not due to direct Ag presentation by the tumor cells, but results from presentation of tumor-derived Ag by nonactivated CD11c(+) APC. Although this event results in division of naive CTL in tumor draining lymph nodes, it does not establish a productive immune response. Treatment of tumor-bearing mice with dendritic cell-stimulating agonistic anti-CD40 mAb resulted in systemic efflux of CTL with robust effector function capable to eradicate established tumors. For efficacy of anti-CD40 treatment, CD40 ligation of host APC is required because adoptive transfer of CD40-proficient tumor-specific TCR transgenic CTL into CD40-deficient tumor-bearing mice did not lead to productive antitumor immunity after CD40 triggering in vivo. CpG and detoxified LPS (MPL) acted similarly as agonistic anti-CD40 mAb with respect to CD8(+) CTL efflux and tumor eradication. Together these results indicate that dendritic cells, depending on their activation state, orchestrate the outcome of CTL-mediated immunity against tumors, leading either to an ineffective immune response or potent antitumor immunity.     

Intralesional injection of adenovirus encoding CC chemokine ligand 16 inhibits mammary tumor growth and prevents metastatic-induced death after surgical removal of the treated primary tumor

The CC chemokine ligand (CCL)16 exerts chemotactic activity on human monocytes and lymphocytes. Although no murine homologous has been defined, the TSA mouse adenocarcinoma cells engineered to express human CCL16 are rapidly rejected by syngenic mice. An adenovirus encoding CCL16 (AdCCL16) was generated using a Cre-Lox-based system and was used to determine whether this chemokine might also block pre-existing tumors. Both recombinant and viral CCL16 showed in vitro chemotactic activity for murine CD4(+) and CD8(+) lymphocytes and dendritic cells (DC). AdCCL16, but not the control empty vector, when injected in established nodules significantly delayed tumor growth. Immunohistochemistry revealed accumulation of CD4(+) and CD8(+) T cells and DC in the treated tumors as well as in draining lymph nodes. DC from such lymph nodes stimulated IFN-gamma by a T cell clone specific for the known TSA tumor-associated Ag (TAA), suggesting the tumor origin of these cells. Lymphocytes from the same nodes showed specific CTL activity against TSA tumor cells and their immunodominant TAA peptide. Antitumor activity required CD4, CD8, and IFN-gamma production, as shown using subset-depleted and knockout mice. Despite the robust and rapid immune response triggered by intratumoral injection of AdCCL16, the lesions were not completely rejected; however, the same treatment given before surgical excision of primary lesions prevented metastatic spread and cured 63% of mice bearing the 4T1 mammary adenocarcinoma, which is perhaps the most compelling model of spontaneous metastasis.     

Infiltration of a mesothelioma by IFN-gamma-producing cells and tumor rejection after depletion of regulatory T cells

Depletion of CD4+CD25+Foxp3+ regulatory T cells (CD25+ T(reg)) with an anti-CD25 Ab results in immune-mediated rejection of tolerogenic solid tumors. In this study, we have examined the immune response to a mesothelioma tumor in mice after depletion of CD25+ cells to elucidate the cellular mechanisms of CD25+ T(reg), a subject over which there is currently much conjecture. Tumor rejection was found to be primarily due to the action of CD8+ T cells, although CD4+ cells appeared to play some role. Depletion of CD25+ cells resulted in an accumulation in tumor tissue of CD4+ and CD8+ T cells and NK cells that were producing the potent antitumor cytokine IFN-gamma. Invasion of tumors by CD8+ T cells was partially dependent on the presence of CD4+ T cells. Although a significant increase in the proliferation and number of tumor-specific CD8+ T cells was observed in lymph nodes draining the tumor of anti-CD25-treated mice, this effect was relatively modest compared with the large increase in IFN-gamma-producing T cells found in tumor tissue, which suggests that the migration of T cells into tumor tissue may also have been altered. Depletion of CD25+ cells did not appear to modulate antitumor CTL activity on a per cell basis. Our data suggests that CD25+ T(reg) limit the accumulation of activated T cells producing IFN-gamma in the tumor tissue and, to a lesser extent, activation and/or rate of mitosis of tumor-specific T cells in lymph nodes.     

Recovery of activated cytotoxic T cells from minor H incompatible tumor graft rejection sites

In this study we determined whether minor H-specific cytotoxic T cells and their precursors (pTc) are present at the site of rejection of minor H disparate tumor allografts. Lymphocytes were retrieved from eyes of BALB/c mice that received subconjunctival injections of minor H-incompatible P815 tumor cells. The lymphocytes were then assayed for direct cytotoxic activity as well as precursor frequency by limiting dilution. Similar assays were conducted on cells obtained from the draining lymph nodes and from the spleen. As expected, tumor rejection was accompanied by significant clonal expansion of minor H-specific pTc within the draining lymph node and the spleen. A correspondingly high frequency of pTc was also detected at the graft site. More importantly, fully functional cytotoxic T cells were recovered from the tumor graft site during rejection, but no similarly active cells were found in either the draining nodes or spleen. We conclude that, after Ag stimulation, pTc are generated in draining central lymphoid compartments. From this generative site, the precursor cells then disseminate systemically, gradually reaching and infiltrating the tumor graft site. A further activation step, dependent upon Ag and T cell help, permits these cells to mature into fully active cytotoxic cells which can then effect tumor rejection. We propose that the terminal stage(s) of pTc activation is promoted by lymphokines released locally from TDH cells that are also generated during the alloimmune response and simultaneously infiltrate the site.     

Incomplete differentiation of antigen-specific CD8 T cells in tumor-draining lymph nodes

CD8 T cells lacking effector activity have been recovered from lymphoid organs of mice and patients with progressing tumors. We explored the basis for lack of effector activity in tumor-bearing mice by evaluating Ag presentation and CD8 T cell function in lymphoid organs over the course of tumor outgrowth. Early after tumor injection, cross-presentation by bone marrow-derived APC was necessary for T cell activation, inducing proliferation and differentiation into IFN-gamma-producing, cytolytic effectors. At later stages of outgrowth, tumor metastasized to draining lymph nodes. Both cross- and direct presentation occurred, but T cell differentiation induced by either modality was incomplete (proliferation without cytokine production). T cells within tumor-infiltrated nodes differentiated appropriately if Ag was presented by activated, exogenous dendritic cells. Thus, activated T cells lacking effector function develop through incomplete differentiation in the lymph nodes of late-stage tumor-bearing mice, rather than through suppression of previously differentiated cells.     

The TLR-7 agonist, imiquimod, enhances dendritic cell survival and promotes tumor antigen-specific T cell priming: relation to central nervous system antitumor immunity

Immunotherapy represents an appealing option to specifically target CNS tumors using the immune system. In this report, we tested whether adjunctive treatment with the TLR-7 agonist imiquimod could augment antitumor immune responsiveness in CNS tumor-bearing mice treated with human gp100 + tyrosine-related protein-2 melanoma-associated Ag peptide-pulsed dendritic cell (DC) vaccination. Treatment of mice with 5% imiquimod resulted in synergistic reduction in CNS tumor growth compared with melanoma-associated Ag-pulsed DC vaccination alone. Continuous imiquimod administration in CNS tumor-bearing mice, however, was associated with the appearance of robust innate immune cell infiltration and hemorrhage into the brain and the tumor. To understand the immunological mechanisms by which imiquimod augmented antitumor immunity, we tested whether imiquimod treatment enhanced DC function or the priming of tumor-specific CD8+ T cells in vivo. With bioluminescent, in vivo imaging, we determined that imiquimod dramatically enhanced both the persistence and trafficking of DCs into the draining lymph nodes after vaccination. We additionally demonstrated that imiquimod administration significantly increased the accumulation of tumor-specific CD8+ T cells in the spleen and draining lymph nodes after DC vaccination. The results suggest that imiquimod positively influences DC trafficking and the priming of tumor-specific CD8+ T cells. However, inflammatory responses induced in the brain by TLR signaling must also take into account the local microenvironment in the context of antitumor immunity to induce clinical benefit. Nevertheless, immunotherapeutic targeting of malignant CNS tumors may be enhanced by the administration of the innate immune response modifier imiquimod.     

Regulatory T cells prevent CD8 T cell maturation by inhibiting CD4 Th cells at tumor sites

Natural regulatory T cells (Tregs) are present in high frequencies among tumor-infiltrating lymphocytes and in draining lymph nodes, supposedly facilitating tumor development. To investigate their role in controlling local immune responses, we analyzed intratumoral T cell accumulation and function in the presence or absence of Tregs. Tumors that grew in normal BALB/c mice injected with the 4T1 tumor cell line were highly infiltrated by Tregs, CD4 and CD8 cells, all having unique characteristics. Most infiltrating Tregs expressed low levels of CD25Rs and Foxp3. They did not proliferate even in the presence of IL-2 but maintained a strong suppressor activity. CD4 T cells were profoundly anergic and CD8 T cell proliferation and cytotoxicity were severely impaired. Depletion of Tregs modified the characteristics of tumor infiltrates. Tumors were initially invaded by activated CD4(+)CD25(-) T cells, which produced IL-2 and IFN-gamma. This was followed by the recruitment of highly cytotoxic CD8(+) T cells at tumor sites leading to tumor rejection. The beneficial effect of Treg depletion in tumor regression was abrogated when CD4 helper cells were also depleted. These findings indicate that the massive infiltration of tumors by Tregs prevents the development of a successful helper response. The Tregs in our model prevent Th cell activation and subsequent development of efficient CD8 T cell activity required for the control of tumor growth.     

Identification of a highly immunogenic mouse breast cancer sub cell line, 4T1-S

Cancer vaccines serve as a promising clinical immunotherapeutic strategy that help to trigger an effective and specific antitumor immune response compared to conventional therapies. However, poor immunogenicity of tumor cells remains a major obstacle for clinical application, and developing new methods to modify the immunogenicity of tumor cells may help to improve the clinical outcome of cancer vaccines. 4T1 mouse breast cancer cell line has been known as poorly immunogenic and highly metastatic cell line. Using this model, we identified a sub cell line of 4T1—designated as 4T1-Sapporo (4T1-S)—which shows immunogenic properties when used as a vaccine against the same line. In 4T1-S-vaccinated mice, subcutaneous injection of 4T1-S resulted in an antitumor inflammatory response represented by significant enlargement of draining lymph nodes, accompanied with increased frequencies of activated CD8 T cells and a subpopulation of myeloid cells. Additionally, 4T1-S vaccine was ineffective to induce tumor rejection in nude mice, which importantly indicate that 4T1-S vaccine rely on T cell response to induce tumor rejection. Further analysis to identify mechanisms that control tumor immunogenicity in this model may help to develop new methods for improving the efficacies of clinical cancer vaccines.     

Mechanical Disruption of Tumors by Iron Particles and Magnetic Field Application Results in Increased Anti-Tumor Immune Responses

The primary tumor represents a potential source of antigens for priming immune responses for disseminated disease. Current means of debulking tumors involves the use of cytoreductive conditioning that impairs immune cells or removal by surgery. We hypothesized that activation of the immune system could occur through the localized release of tumor antigens and induction of tumor death due to physical disruption of tumor architecture and destruction of the primary tumor in situ. This was accomplished by intratumor injection of magneto-rheological fluid (MRF) consisting of iron microparticles, in Balb/c mice bearing orthotopic 4T1 breast cancer, followed by local application of a magnetic field resulting in immediate coalescence of the particles, tumor cell death, slower growth of primary tumors as well as decreased tumor progression in distant sites and metastatic spread. This treatment was associated with increased activation of DCs in the draining lymph nodes and recruitment of both DCs and CD8(+)T cells to the tumor. The particles remained within the tumor and no toxicities were observed. The immune induction observed was significantly greater compared to cryoablation. Further anti-tumor effects were observed when MRF/magnet therapy was combined with systemic low dose immunotherapy. Thus, mechanical disruption of the primary tumor with MRF/magnetic field application represents a novel means to induce systemic immune activation in cancer.     

NKT cells inhibit antigen-specific effector CD8 T cell induction to skin viral proteins

We recently demonstrated that CD1d-restricted NKT cells resident in skin can inhibit CD8 T cell-mediated graft rejection of human papillomavirus E7-expressing skin through an IFN-γ-dependent mechanism. In this study, we examined the role of systemically derived NKT cells in regulating the rejection of skin grafts expressing viral proteins. In lymph nodes draining transplanted skin, Ag-specific CD8 T cell proliferation, cytokine production, and cytotoxic activity were impaired by NKT cells. NKT cell suppression was mediated via CD11c(+) dendritic cells. Inhibition of CD8 T cell function did not require Foxp3(+) regulatory T cells or NKT cell-secreted IFN-γ, IL-10, or IL-17. Thus, following skin grafting or immunization with human papillomavirus-E7 oncoprotein, NKT cells reduce the capacity of draining lymph node-resident APCs to cross-present Ag to CD8 T cell precursors, as evidenced by impaired expansion and differentiation to Ag-specific CD8 T effector cells. Therefore, in the context of viral Ag challenge in the skin, systemic NKT cells limit the capacity for effective priming of adaptive immunity.     

Anti-tumor activity and trafficking of self, tumor-specific T cells against tumors located in the brain

It is commonly believed that T cells have difficulty reaching tumors located in the brain due to the presumed "immune privilege" of the central nervous system (CNS). Therefore, we studied the biodistribution and anti-tumor activity of adoptively transferred T cells specific for an endogenous tumor-associated antigen (TAA), gp100, expressed by tumors implanted in the brain. Mice with pre-established intracranial (i.c.) tumors underwent total body irradiation (TBI) to induce transient lymphopenia, followed by the adoptive transfer of gp100(25-33)-specific CD8+ T cells (Pmel-1). Pmel-1 cells were transduced to express the bioluminescent imaging (BLI) gene luciferase. Following adoptive transfer, recipient mice were vaccinated with hgp100(25-33) peptide-pulsed dendritic cells (hgp100(25-33)/DC) and systemic interleukin 2 (IL-2). This treatment regimen resulted in significant reduction in tumor size and extended survival. Imaging of T cell trafficking demonstrated early accumulation of transduced T cells in lymph nodes draining the hgp100(25-33)/DC vaccination sites, the spleen and the cervical lymph nodes draining the CNS tumor. Subsequently, transduced T cells accumulated in the bone marrow and brain tumor. BLI could also detect significant differences in the expansion of gp100-specific CD8+ T cells in the treatment group compared with mice that did not receive either DC vaccination or IL-2. These differences in BLI correlated with the differences seen both in survival and tumor infiltrating lymphocytes (TIL). These studies demonstrate that peripheral tolerance to endogenous TAA can be overcome to treat tumors in the brain and suggest a novel trafficking paradigm for the homing of tumor-specific T cells that target CNS tumors.