Induction of tumor-reactive T helper responses by a posttranslational modified epitope from tumor protein p53

Posttranslational modifications regulate the function and stability of proteins, and the immune system is able to recognize some of these modifications. Therefore, the presence of posttranslational modifications increases the diversity of potential immune responses to a determinant antigen. The stimulation of tumor-specific CD4⁺ helper T lymphocytes (HTLs) is considered important for the production of anti-tumor antibodies by B cells and for the generation and persistence of CD8⁺ cytotoxic T lymphocytes, and in some instances, HTLs can directly reduce tumor cell growth. Identification of MHC class II-restricted peptide epitopes from tumor-associated antigens including those generated from posttranslational protein modifications should enable the improvement of peptide-based cancer immunotherapy. We describe here an MHC class II binding peptide from the tumor protein p53, which possesses an acetylated lysine at position 120 (p53_{110-124/AcK120}) that is effective in eliciting CD4⁺ T cell responses specific for the acetylated peptide. Most importantly, the acetylated peptide-reactive CD4 HTLs recognized the corresponding naturally processed posttranslational modified epitope presented by either dendritic cells loaded with tumor cell lysates or directly on tumors expressing p53 and the restricting MHC class II molecules. Treatment of tumor cells with a histone deacetylase inhibitor augmented their recognition by the p53_{110-124/AcK120}-reactive CD4⁺ T cells. These findings prove that the epitope p53_{110-124/AcK120} is immunogenic for anti-tumor responses and is likely to be useful for cancer immunotherapy.     

Tumor-specific CD4+ T lymphocytes from cancer patients are required for optimal induction of cytotoxic T cells against the autologous tumor

This study focuses on the specific CD4+ T cell requirement for optimal induction of cytotoxicity against MHC class II negative autologous tumors (AuTu) collected from patients with various types of cancer at advanced stages. CD4+ T cells were induced in cultures of cancer patients' malignant effusion-associated mononuclear cells with irradiated AuTu (mixed lymphocyte tumor cultures (MLTC)) in the presence of recombinant IL-2 and recombinant IL-7. Tumor-specific CD4+ T cells did not directly recognize the AuTu cells, but there was an MHC class II-restricted cross-priming by autologous dendritic cells (DCs), used as APC. CD8+ CTL, also induced during the MLTC, lysed specifically AuTu cells or DCs pulsed with AuTu peptide extracts (acid wash extracts (AWE)) in an MHC class I-restricted manner. Removal of CD4+ T cells or DCs from the MLTC drastically reduced the CD8+ CTL-mediated cytotoxic response against the AuTu. AWE-pulsed DCs preincubated with autologous CD4+ T cells were able, in the absence of CD4+ T cells, to stimulate CD8+ T cells to lyse autologous tumor targets. Such activated CD8+ T cells produced IL-2, IFN-gamma, TNF-alpha, and GM-CSF. The process of the activation of AWE-pulsed DCs by CD4+ T cells could be inhibited with anti-CD40 ligand mAb. Moreover, the role of CD4+ T cells in activating AWE-pulsed DCs was undertaken by anti-CD40 mAb. Our data demonstrate for the first time in patients with metastatic cancer the essential role of CD4+ Th cell-activated DCs for optimal CD8+ T cell-mediated killing of autologous tumors and provide the basis for the design of novel protocols in cellular adoptive immunotherapy of cancer, utilizing synthetic peptides capable of inducing T cell help in vivo.     

Regulation of antitumour immunity by CD1d-restricted NKT cells

An understanding of the complex interactions occurring between tumours and the immune system is a prerequisite for the rational design of effective cancer immunotherapies. To date, attention has focused mainly on the role the adaptive immune system plays in controlling tumourigenesis, with conventional T cells, which recognize peptide antigens presented by classical MHC molecules, coming under close scrutiny. Accumulating reports now suggest that an additional T-cell subset, known as CD1d-restricted natural killer T (NKT) cells, also plays a pivotal role in modulating antitumour responses. Found in both humans and mice, CD1d-restricted NKT cells are a highly specialized cell type that, in contrast to conventional T cells, recognize lipid/glycolipid antigens presented by the non-classical MHC molecule CD1d. Several features of NKT cells, including their ability to rapidly produce large quantities of cytokines upon primary stimulation, make them ideal targets for developing anticancer immunotherapies. This intriguing cell type is the focus of this review.     

Generation of T cell help through a MHC class I-restricted TCR

CD4+ T cells that are activated by a MHC class II/peptide encounter can induce maturation of APCs and promote cytotoxic CD8+ T cell responses. Unfortunately, the number of well-defined tumor-specific CD4+ T cell epitopes that can be exploited for adoptive immunotherapy is limited. To determine whether Th cell responses can be generated by redirecting CD4+ T cells to MHC class I ligands, we have introduced MHC class I-restricted TCRs into postthymic murine CD4+ T cells and examined CD4+ T cell activation and helper function in vitro and in vivo. These experiments indicate that Ag-specific CD4+ T cell help can be induced by the engagement of MHC class I-restricted TCRs in peripheral CD4+ T cells but that it is highly dependent on the coreceptor function of the CD8beta-chain. The ability to generate Th cell immunity by infusion of MHC class I-restricted Th cells may prove useful for the induction of tumor-specific T cell immunity in cases where MHC class II-associated epitopes are lacking.     

Characterization of human CD4 helper T cell responses against Aurora kinase A

Aurora kinase A (Aurora-A) is a cell cycle-associated serine–threonine kinase that is overexpressed by various types of cancer and is highly associated with poor prognosis. Since the expression of Aurora-A in normal tissues has been shown to be significantly lower as compared to tumor cells, this protein is being considered as a potential tumor-associated antigen for developing immunotherapies. The goal in the present study was to identify CD4 helper T lymphocyte (HTL) epitopes for Aurora-A for the design of T cell-based immunotherapies against Aurora-A-expressing tumors. Synthetic peptides corresponding to potential HTL epitopes were identified from Aurora-A and used to stimulate CD4 T lymphocytes in vitro to generate antigen-specific HTL clones that were evaluated for antigen specificity, MHC restriction and for their ability to interact with Aurora-A-expressing tumor cells. The results show that two peptides (Aurora-A_161–175 and Aurora-A_233–247) were effective in generating HTL responses that were restricted by more than one MHC class II allele (i.e., promiscuous responses). The CD4 HTL clones were able to directly recognize Aurora-A-expressing tumor cells in an antigen-specific and MHC class II-restricted manner and some of the clones displayed cytolytic activity toward Aurora-A + tumor cells. Both of these peptides were capable of stimulating in vitro T cell responses in patients with bladder cancer.     

Genetic approaches for the induction of a CD4+ T cell response in cancer immunotherapy

Recently, it has become more and more obvious that not only CD8+ cytotoxic T lymphocytes, but also CD4+ T helper cells are required for the induction of an optimal, long-lasting anti-tumor immune response. CD4+ T helper cells, and in particular IFN-gamma-secreting type 1 T helper cells, have been shown to fulfill a critical function in the mounting of a cancer-specific response. Consequently, targeting antigens into MHC class II molecules would greatly enhance the efficacy of an anti-cancer vaccine. The dissection of the MHC class II presentation pathway has paved the way for rational approaches to achieve this goal: novel systems have been developed to genetically manipulate the MHC class II presentation pathway. First, different genetic approaches have been used for the delivery of known epitopes into the MHC class II processing pathway or directly onto the peptide-binding groove of the MHC molecules. Second, several strategies exist for the targeting of whole tumor antigens, containing both MHC class I and class II restricted epitopes, to the MHC class II processing pathway. We review these data and describe how this knowledge is currently applied in vaccine development.     

CD8 is needed for development of cytotoxic T cells but not helper T cells.

A mutant mouse strain without CD8 (Lyt-2 and Lyt-3) expression on the cell surface has been generated by disrupting the Lyt-2 gene using embryonic stem cell technology. In these mice, CD8+ T lymphocytes are not present in peripheral lymphoid organs, but the CD4+ T lymphocyte population seems to be unaltered. Cytotoxic response of T lymphocytes from these mice against alloantigens and viral antigens is dramatically decreased. Proliferative response against alloantigens and in vivo help to B lymphocytes, however, are not affected. These data suggest that CD8 is necessary for the maturation and positive selection of class I MHC restricted cytotoxic T lymphocytes but is not required on any of the intermediate thymocyte populations (CD8+CD4-TcR- or CD4+CD8+TcRlow) during the development of functional class II MHC restricted helper T cells.     

Existence of MHC class I-restricted alloreactive CD4+ T cells reacting with peptide transporter-deficient cells

It is generally accepted that as the result of positive thymic selection, CD8-expressing T cells recognize peptide antigens presented in the context of MHC class I molecules and CD4-expressing T cells interact with peptide antigens presented by MHC class II molecules. Here we report the generation of TCRalpha/beta(+), CD3(+), CD4(+), CD8(-), MHC class I-restricted alloreactive T-cell clones which were induced using peripheral blood mononuclear cells from healthy individuals following in vitro stimulation with transporter associated with antigen processing (TAP)-deficient cell lines T2. The CD4(+) T-cell clones showed an HLA-A2.1-specific proliferative response against T2 cells which was inhibited by anti-CD3 and anti-CD4 monoclonal antibodies. These results suggest that interaction of the TCR with peptide-bound HLA class I molecules contributes to antigen-specific activation of these co-receptor-mismatched T-cell clones. Antigen recognition by alloreactive MHC class I-restricted CD4(+) T cells was inhibited by removing peptides bound to HLA molecules on T2 cells suggesting that the alloreactive CD4(+) T cells recognize peptides that bind in a TAP-independent manner to HLA-A2 molecules. The existence of such MHC class I-restricted CD4(+) T cells which can recognize HLA-A2 molecules in the absence of TAP function may provide a basis for the development of immunotherapy against TAP-deficient tumor variants which would be tolerant to immunosurveillance by conventional MHC class I-restricted cytotoxic lymphocytes.     

Cancer Immunotherapy Employing an Innovative Strategy to Enhance CD4+ T Cell Help in the Tumor Microenvironment

Chemotherapy and/or radiation therapy are widely used as cancer treatments, but the antitumor effects they produce can be enhanced when combined with immunotherapies. Chemotherapy kills tumor cells, but it also releases tumor antigen and allows the cross-presentation of the tumor antigen to trigger antigen-specific cell-mediated immune responses. Promoting CD4+ T helper cell immune responses can be used to enhance the cross-presentation of the tumor antigen following chemotherapy. The pan HLA-DR binding epitope (PADRE peptide) is capable of generating antigen-specific CD4+ T cells that bind various MHC class II molecules with high affinity and has been widely used in conjunction with vaccines to improve their potency by enhancing CD4+ T cell responses. Here, we investigated whether intratumoral injection of PADRE and the adjuvant CpG into HPV16 E7-expressing TC-1 tumors following cisplatin chemotherapy could lead to potent antitumor effects and antigen-specific cell-mediated immune responses. We observed that treatment with all three agents produced the most potent antitumor effects compared to pairwise combinations. Moreover, treatment with cisplatin, CpG and PADRE was able to control tumors at a distant site, indicating that our approach is able to induce cross-presentation of the tumor antigen. Treatment with cisplatin, CpG and PADRE also enhanced the generation of PADRE-specific CD4+ T cells and E7-specific CD8+ T cells and decreased the number of MDSCs in tumor loci. The treatment regimen presented here represents a universal approach to cancer control.     

Identification of a ras oncogene peptide that contains both CD4(+) and CD8(+) T cell epitopes in a nested configuration and elicits both T cell subset responses by peptide or DNA immunization

Mutations in ras proto-oncogenes are commonly found in a diversity of malignancies and may encode unique, non-self epitopes for T cell-mediated antitumor activity. In a BALB/c (H-2(d)) murine model, we have identified a single peptide sequence derived from the ras oncogenes that contained both CD8(+) and CD4(+) T cell epitopes in a nested configuration. This peptide reflected ras sequence 4-16, and contained the substitution of Gly to Val at position 12 ?i.e., 4-16(Val12)?. Mice immunized with this 13-mer peptide induced a strong antigen (Ag)-specific CD4(+) proliferative response in vitro. In contrast, mice inoculated with the wild-type ras sequence failed to generate a peptide-specific T cell response. Additionally, mice immunized with the ras 4-16(Val12) peptide concomitantly displayed an Ag-specific CD8(+) cytotoxic T lymphocyte (CTL) response, as determined by lysis of syngeneic tumor target cells incubated with the nominal 9-mer nested epitope peptide ?i.e., 4-12(Val12)?, as well as lysis of tumor target cells expressing the corresponding ras codon 12 mutation. Analysis of the Valpha- and Vbeta-chains of the T cell receptor (TCR) expressed by these CTL revealed usage of the Valpha1 and Vbeta9 subunits, consistent with the TCR phenotype of anti-ras Val12 CTL lines produced by in vivo immunization with the nominal peptide epitope alone. Moreover, immunization with the nested epitope peptide, as compared to immunization with either the 9-mer CTL peptide alone or an admixture of the 9-mer CTL peptide with an overlapping 13-mer CD4(+) T cell helper peptide ?i.e., 5-17(Val12)? lacking the class I N-terminus anchor site, enhanced the production of the CD8(+) T cell response. Finally, immunization with plasmid DNA encoding the ras 4-16(Val12) sequence led to the induction of both Ag-specific proliferative and cytotoxic responses. Overall, these results suggested that a single peptide immunogen containing nested mutant ras-specific CD4(+) and CD8(+) T cell epitopes: (1) can be processed in vivo to induce both subset-specific T lymphocyte responses; and (2) leads to the generation of a quantitatively enhanced CD8(+) CTL response, likely due to the intimate coexistence of CD4(+) help, which may have implications in peptide- or DNA-based immunotherapies.     

Enhancement of antitumor immunity by prolonging antigen presentation on dendritic cells

Vaccination with dendritic cells (DCs) pulsed with antigenic peptides derived from various tumor antigens has great, but as yet significantly unrealized, potential in cancer treatment. Here, we describe a strategy for prolonged presentation of an MHC class I-restricted self-peptide on DCs through linkage of it to a cell penetrating peptide (CPP). DCs loaded with a peptide derived from tyrosinase-related protein 2 (TRP2) covalently linked to a CPP1 sequence retained full capacity to stimulate T cells for at least 24 h, completely protected immunized mice from subsequent tumor challenge, and significantly inhibited lung metastases in a 3-day tumor model. DCs pulsed with TRP2 alone failed to provide any of these protections. In addition, we demonstrate that both CD4+ and CD8+ T cells were required for potent antitumor immunity. This CPP-based approach may be generally applicable to enhance the efficacy of DC-based peptide vaccines against cancer and other diseases.     

Cutting edge: CD4+ T cell control of CD8+ T cell reactivity to a model tumor antigen

Neoantigens resulting from the inherent genomic instability of tumor cells generally do not trigger immune recognition. Similarly, transfection of tumors with model Ags often fails to elicit CD8+ T cell responses or alter a tumor's growth rate or lethality. We report here that the adoptive transfer of activated Th1-type CD4+ T cells specific for a model tumor Ag results in the de novo generation of CD8+ T cells with specificity to that Ag and concomitant tumor destruction. The anti-tumor effects of the CD4+ T cells required the presence of both MHC class I and class II on host cells, as evidenced by experiments in knockout mice, suggesting that CD4+ T cells enhanced the ability of host APC to activate endogenous CD8+ T cells. These results indicate that the apparent inability of tumor cells expressing highly immunogenic epitopes to activate tumor-specific CD8+ T cells can be altered by activated CD4+ T cells.     

H2-M3-restricted T cells participate in the priming of antigen-specific CD4+ T cells

H2-M3-restricted CD8+ T cells provide early protection against bacterial infections. In this study, we demonstrate that activated H2-M3-restricted T cells provide early signals for efficient CD4+ T cell priming. C57BL/6 mice immunized with dendritic cells coated with the MHC class II-restricted listeriolysin O peptide LLO(190-201) (LLO) generated CD4+ T cells capable of responding to Listeria monocytogenes (LM) infection. Inclusion of a H2-M3-restricted formylated peptide fMIGWII (fMIG), but not MHC class Ia-restricted peptides, during immunization with LLO significantly increased IFN-gamma-producing CD4+ T cell numbers, which was associated with increased protection against LM infection. Studies with a CD4+ T cell-depleting mAb indicate that the reduction in bacterial load in fMIG plus LLO immunized mice is likely due to augmented numbers of LLO-specific CD4+ T cells, generated with the help of H2-M3-restricted CD8+ T cells. We also found that augmentation of LLO-specific CD4+ T lymphocytes with H2-M3-restricted T cells requires presentation of LLO and fMIG by the same dendritic cells. Interestingly, the augmented CD4+ T cell response generated with fMIG also increased primary LM-specific responses by MHC class Ia-restricted CD8 T cells. Coimmunization with LLO and fMIG also increases the number of memory Ag-specific CD4+ T cells. We also demonstrate that CD8 T cells restricted to another MHC class Ib molecule, Qa-1, whose human equivalent is HLA-E, are also able to enhance Ag-specific CD4+ T cell responses. These results reveal a novel function for H2-M3- and Qa-1-restricted T cells; provision of help to CD4+ Th cells during the primary response.     

Membrane structures involved in auto-tumor recognition

Tumor patients' blood lymphocytes have the capacity to recognize autologous tumor cells in vitro. A consequence of this recognition is the proliferation of small-size, high-density, resting T cells. Both helper (CD4+) and cytotoxic/suppressor (CD8+) T lymphocytes proliferate in the mixed lymphocyte-tumor cell cultures. In contrast to the autologous mixed lymphocyte cultures, both the auto-erythrocyte rosetting and non-rosetting (AE+ and AE-) T cells participate in the auto-tumor response. In contrast to stimulation by virus-infected or hapten-modified cells, DR antigen expression is not essential for stimulation by autologous tumor cells. In a proportion of cancer patients, blood lymphocytes have the capacity to lyse the patients' own tumor cells in vitro. There are two populations of lymphocytes with auto-tumor cytotoxic function. The first is characterized by low buoyant density and by non-adaptive cytotoxicity. In contrast to the recognition of hapten-modified or virus-infected target cells by the CTL, recognition of autologous tumor cells by the cytotoxic LD cells occurs even when the MHC class I antigens are blocked by mAb. The CD3 complex is also not involved in LD-mediated lysis. The other population with auto-tumor cytotoxic function comprises high-density, resting T cells. Recognition of autologous tumor cells by cytotoxic HD lymphocytes shares the characteristics of CTLs, i.e., their function is abrogated by pretreatment of the effectors with mAbs directed to the T3 receptor complex and by preincubation of the targets with mAb to the MHC class I antigens. Cytotoxicity of HD cells is restricted to the autologous tumor cells. This selectivity and the characteristics shared with CTL suggest that the auto-tumor reactivity of HD lymphocytes reflects an immune response against the autologous tumor.     

Identification of MHC class II-restricted tumor antigens recognized by CD4+ T cells

CD4+ T cells play a central role in orchestrating host immune responses against cancer as well as autoimmune and infectious diseases. Identification of major histocompatibility complex (MHC) class II-restricted helper T peptides is important for development of effective vaccines. The lack of effective methods to identify such T-cell peptides is a major hurdle in the use of antigen-specific CD4+ T cells in cancer vaccines. Here we describe a genetic targeting expression system for cloning genes encoding for MHC class II-restricted tumor antigens recognized by tumor-reactive CD4+ T cells. Helper T peptides are subsequently identified by using synthetic peptides to test their ability to stimulate CD4+ T cells.     

Mouse Hepa-1 tumor is rejected by H-2Db-restricted CTL despite decreased MHC class I antigen expression

It has recently been hypothesized that tumor cells with reduced levels of MHC class I antigens are more susceptible to NK-mediated lysis and are rejected by NK cells, whereas tumor cells with normal levels of class I are rejected by tumor-specific CTL. We have tested this hypothesis using a mouse hepatoma system. The Hepa-1 tumor is a spontaneous H-2Kb loss variant that arose from the BW7756 tumor, when BW7756 was adapted to growth in culture. Our studies have shown that despite the loss of H-2Kb antigen, Hepa-1 is not more susceptible to NK lysis than its H-2Kb-transfected variants. These studies also suggested that NK cells were not responsible for rejection of the Hepa-1 tumor. The Hepa-1 tumor, therefore, appears to contradict the hypothesized linkage of MHC levels and NK susceptibility. Because NK cells are not involved in immunity to this tumor, we have sought to identify the effector cell responsible for Hepa-1 rejection. Cytotoxic T lymphocyte assays demonstrate that in vitro, Hepa-1 cells are lysed by Hepa-1-specific H-2Db-restricted CD4-CD8+ T lymphocytes. Footpad assays demonstrate that in vivo, Hepa-1 rejection requires CD4+CD8- and CD4-CD8+ Hepa-1-primed splenocytes. These results indicate that immunity to Hepa-1 is T cell mediated. Hepa-1 is therefore an example of an unusual tumor in that down-regulation of MHC class I antigen expression is associated with increased CTL susceptibility.     

CD4+ tumor-infiltrating lymphocytes in cervical cancer recognize HLA-DR-restricted peptides provided by human papillomavirus-E7.

Human papillomavirus (HPV)-encoded proteins may provide targets for CD8+ or CD4+ T lymphocytes infiltrating into cervical cancer. We established an MHC class II-restricted CD4+ T cell line from a patient with cervical cancer that recognizes autologous (HPV35+, HPV59+) cervical cancer cells and the HLA-DR4-matched cervical cancer cell line Me180 (HPV68+) as determined by TNF-alpha secretion. Expression of different HPV-E7 genes in autologous B cells revealed that this T cell line defines a DR4-presented T cell epitope that is shared among the E7 genes of HPV59 and HPV68. MHC class II-presented peptides may be implemented to augment T cell responses directed against autologous tumor cells, particularly if cancer cells lack MHC class I expression, which is a frequent event in the evolution of cervical cancer.     

Comparative analysis of IFN-gamma B7.1 and antisense TGF-beta gene transfer on the tumorigenicity of a poorly immunogenic metastatic mammary carcinoma

Cancer progression is attributed in part to immune evasion strategies that include lack of co-stimulation, down-regulation of cell surface MHC molecules, and secretion of immunosuppressive factors, such as transforming growth factor-beta (TGF-beta). Gene therapy has been employed to counter these mechanisms of immune evasion by transference of B7.1, IFN-gamma or antisense TGF-beta genes into tumor cells, resulting in cell surface expression of B7.1, upregulation of MHC class I and class II molecules, or elimination of tumor-derived TGF-beta, respectively. Although each of these transgenes has been shown to alter tumorigenicity in murine models, a direct comparison of their efficacy has not been performed. In this study, we have employed a very aggressive, poorly immunogenic and highly metastatic mammary model, 4T1, to compare the efficacy of B7.1, IFN-gamma and antisense TGF-beta gene transfer in stimulating an anti-tumor response. We demonstrate that both IFN-gamma and antisense TGF-beta gene expression significantly reduced the tumorigenicity of these cells compared to mock transduced cells, with IFN-gamma having a greater effect. In contrast, B7.1 gene transfer did not affect the tumorigenicity of 4T1 cells. The anti-tumor response directed against antisense TGF-beta-expressing 4T1 tumors was mediated by CD4+ and CD8+ T cells. However, CD8+ T cells, and not CD4+ T cells, appeared to mediate the anti-tumor response against IFN-gamma-expressing tumors. Treatment of tumor-bearing animals with IFN-gamma or antisense TGF-beta gene-modified tumor cell vaccines reduced the number of clonogenic metastases to the lungs and liver compared to treatment with mock-transduced cells. Finally, in a residual disease model in which the primary tumor was excised and mice were vaccinated with irradiated tumor cells, treatment of mice with vaccinations consisting of 4T1 cells expressing both antisense TGF-beta and IFN-gamma genes was the most effective in prolonging survival.     

Adoptive tumor therapy with T lymphocytes enriched through an IFN-gamma capture assay

Successful adoptive T-cell therapy has been demonstrated in viral disease and selected forms of cancer. However, it is limited by the difficulty to efficiently isolate and amplify autologous tumor-reactive T-cell clones. Tetramers of major histocompatibility complex (MHC) class I and peptide have facilitated the characterization of CD8+ T cells specific for tumor-associated antigens. However, for adoptive T-cell therapy, MHC-tetramers have limitations: they require knowledge of tumor antigens, which is often not available; they select T cells with a single specificity, thereby posing risk for selection of tumor escape variants; they do not select for function, so that T cells may be anergic when isolated from cancer patients; and they do not allow the isolation of CD4+ T cells that can be essential for tumor rejection. Because interferon (IFN)-gamma is essential for tumor rejection, we isolated live T cells based on their IFN-gamma production. IFN-gamma secreted by previously activated T cells is retained on the cell surface, allowing their specific isolation and expansion. We show here that IFN-gamma+ but not IFN-gamma- T cells from tumor-immunized mice are cytolytic and mediate tumor rejection upon adoptive transfer. Importantly, tumor-specific T cells can be enriched from lymphocytes infiltrating human renal cell carcinoma by the IFN-gamma capture assay.     

Lactobacilli secreting a tumor antigen and IL15 activates neutrophils and dendritic cells and generates cytotoxic T lymphocytes against cancer cells

Lactobacillus rhamnosus GG (LGG) has been used to successfully induce tumor regression in an orthotopic model of bladder cancer. Increased infiltration of neutrophils and macrophages into the tumor mass was observed after therapy. This study evaluates the potential of LGG to induce a directed anti-tumor response. Lactobacilli were modified to secrete the prostate specific antigen (PSA) or IL15 and PSA (IL-15-PSA). Neutrophils and DC were exposed to LGG for 2 h as in clinical therapy for bladder cancer. Recombinant LGG activated neutrophils (elevated MHC class I expression) induced DC maturation (increased expression of CD86, CD80, CD40, MHC II and CD83), T cell proliferation and PSA specific cytotoxic T lymphocytes (CTL) activity. IL15 enhanced direct DC activation of CTL. Thus LGG secreting tumor antigens may activate antigen specific immune responses when instilled intravesically and IL15 could enhance this response.