Tumors and the danger model

This article reviews the evidence for the danger model in the context of immune response to tumors and the insufficiency of the immune system to eliminate tumor growth. Despite their potential immunogenicity tumors do not induce significant immune responses which could destroy malignant cells. According to the danger model, the immune surveillance system fails to detect tumor antigens because transformed cells do not send any danger signals which could activate dendritic cells and initiate an immune response. Instead, tumor cells or antigen presenting cells turn off the responding T cells and induce tolerance. The studies reviewed herein based on model tumor antigens, recombinant viral vectors and detection of tumor specific T cells by MHC/peptide tetramers underscore the critical role of tumor antigen presentation and the context in which it occurs. They indicate that antigen presentation only by activated but not by cancer or resting dendritic cells is necessary for the induction of immune responses to tumor antigens. It becomes apparent that the inability of dendritic cells to become activated provides a biological niche for tumor escape from immune destruction and seems to be a principal mechanism for the failure of tumor immune surveillance.     

Interactions between macrophages and T-lymphocytes: tumor sneaking through intrinsic to helper T cell dynamics

In a mathematical model of the cellular immune response we investigate immune reactions to tumors that are introduced in various doses. The model represents macrophage T-lymphocyte interactions that generate cytotoxic macrophages and cytotoxic T-lymphocytes. In this model antigens (tumors) can induce infinitely large T-lymphocyte effector populations because effector T-lymphocytes are capable of repeated proliferation and we have omitted immunosuppression. In this (proliferative) model small doses of weakly antigenic tumors grow infinitely large (i.e. sneak through) eliciting an immune response of limited magnitude. Intermediate doses of the same tumor induce larger immune responses and are hence rejected. Large doses of the tumor break through, but their progressive growth is accompanied by a strong immune response involving extensive lymphocyte proliferation. Similarly a more antigenic tumor is rejected in intermediate doses and breaks through in large doses. Initially small doses however lead to tumor dormancy. Thus although the model is devoid of explicit regulatory mechanisms that limit the magnitude of its response (immunosuppression is such a mechanism), the immune response to large increasing tumors may either be a stable reaction of limited magnitude (experimentally known as tolerance or unresponsiveness) or a strong and ever increasing reaction. Unresponsiveness can evolve because in this model net T-lymphocyte proliferation requires the presence of a minimum number of helper T cells (i.e. a proliferation threshold). Unresponsiveness is caused by depletion of helper T cell precursors.     

Distribution and morphologic characteristics of lymphoid cells in the canine popliteal lymph nodes in their normal state and under antigen effect

By means of morphometry, light and electron microscopy methods peculiarities in distribution of small, middle and large lymphocytes, as well as plasmocytes in various zones of the popliteal lymph nodes have been studied in normal and in dynamics up to one year after subcutaneous injection of BCG vaccine into the left hind paw. The antigen produces certain changes in density and morphological parameters of lymphoid cells both in the regional and in the contralateral lymph nodes. For them 3 periods are specific. During the first 3 days they are not antigen-dependent (stipulated by the stress reaction), during 7-24 days antigen-dependent processes of proliferation and differentiation of lymphocytes get into action. In 3 months a new wave of the immune response is observed.     

Macrophage T lymphocyte interactions in the anti-tumor immune response: a mathematical model

In this paper we present a model of the macrophage T lymphocyte interactions that generate an anti-tumor immune response. The model specifies i) induction of cytotoxic T lymphocytes, ii) antigen presentation by macrophages, which leads to iii) activation of helper T cells, and iv) production of lymphoid factors, which induce a) cytotoxic macrophages, b) T lymphocyte proliferation, and c) an inflammation reaction. Tumor escape mechanisms (suppression, antigenic heterogeneity) have been deliberately omitted from the model. This research combines hitherto unrelated or even contradictory data within the range of behavior of one model. In the model behavior, helper T cells play a crucial role: Tumors that differ minimally in antigenicity (i.e., helper reactivity) can differ markedly in rejectability. Immunization yields protection against tumor doses that would otherwise be lethal, because it increases the number of helper T cells. The magnitude of the cytotoxic effector cell response depends on the time at which helper T cells become activated: early helper activity steeply increases the magnitude of the immune response. The type of cytotoxic effector cells that eradicates the tumor depends on tumor antigenicity: lowly antigenic tumors are attacked mainly by macrophages, whereas large highly antigenic tumors can be eradicated by cytotoxic T lymphocytes only.     

A discrete model for immune surveillance, tumor immunity and cancer.

In this paper we propose a model of tumor immunity in terms of discrete automata where each automation describes the concentration of one particular type of cell involved in immune response. In contrast to the earlier models of normal immune response, there is more than one type of cell surface antigen in this model. As a consequence, the tumor can evade destruction through humoral response by changing its identity. However, the tumor can be killed by the killer cells through cell-mediated response unless protected by a high concentration of the suppressor T cells.     

Targeting activity of a TCR/IL-2 fusion protein against established tumors

We have previously reported that a single-chain T cell receptor/IL-2 fusion protein (scTCR-IL2) exhibits potent targeted antitumor activity in nude mice bearing human tumor xenografts that display cognate peptide/HLA complexes. In this study, we further explore the mechanism of action of this molecule. We compared the biological activities of c264scTCR-IL2, a scTCR-IL2 protein recognizing the aa264–272 peptide of human p53, with that of MART-1scTCR-IL2, which recognizes the MART-1 melanoma antigen (aa27–35). In vitro studies showed that c264scTCR-IL2 and MART-1scTCR-IL2 were equivalent in their ability to bind cell-surface IL-2 receptors and stimulate NK cell responses. In mice, MART-1scTCR-IL2 was found to have a twofold longer serum half-life than c264scTCR-IL2. However, despite its shorter serum half-life, c264scTCR-IL2 showed significantly better antitumor activity than MART-1scTCR-IL2 against p53⁺/HLA-A2⁺ tumor xenografts. The more potent antitumor activity of c264scTCR-IL2 correlated with an enhanced capacity to promote NK cell infiltration into tumors. Similar differences in antigen-dependent tumor infiltration were observed with activated splenocytes pre-treated in vitro with c264scTCR-IL2 or MART-1scTCR-IL2 and then transferred into p53⁺/HLA-A2⁺ tumor bearing recipients. The data support a model where c264scTCR-IL2 activates immune cells to express IL-2 receptors. Following stable interactions with cell-surface IL-2 receptors, c264scTCR-IL2 fusion molecule enhances the trafficking of immune cells to tumors displaying target peptide/HLA complexes where the immune cells mediate antitumor effects. Thus, this type of fusion molecule could be used directly as a targeted immunotherapeutic or in adoptive cell transfer approaches to activate and improve the anti-cancer activities of immune cells by providing them with pre-selected antigen recognition capability.     

Tumor-Immune Interaction, Surgical Treatment, and Cancer Recurrence in a Mathematical Model of Melanoma

Malignant melanoma is a cancer of the skin arising in the melanocytes. We present a mathematical model of melanoma invasion into healthy tissue with an immune response. We use this model as a framework with which to investigate primary tumor invasion and treatment by surgical excision. We observe that the presence of immune cells can destroy tumors, hold them to minimal expansion, or, through the production of angiogenic factors, induce tumorigenic expansion. We also find that the tumor–immune system dynamic is critically important in determining the likelihood and extent of tumor regrowth following resection. We find that small metastatic lesions distal to the primary tumor mass can be held to a minimal size via the immune interaction with the larger primary tumor. Numerical experiments further suggest that metastatic disease is optimally suppressed by immune activation when the primary tumor is moderately, rather than minimally, metastatic. Furthermore, satellite lesions can become aggressively tumorigenic upon removal of the primary tumor and its associated immune tissue. This can lead to recurrence where total cancer mass increases more quickly than in primary tumor invasion, representing a clinically more dangerous disease state. These results are in line with clinical case studies involving resection of a primary melanoma followed by recurrence in local metastases.     

Danger signals and nonself entity of tumor antigen are both required for eliciting effective immune responses against HER-2/neu positive mammary carcinoma: implications for vaccine design

Using parental FVB mice and their neu transgenic counterparts, FVBN202, we showed for the first time that dangerous hyperplasia of mammary epithelial cells coincided with breaking immunological tolerance to the neu "self" tumor antigen, though such immune responses failed to prevent formation of spontaneous neu-overexpressing mammary carcinoma (MMC) or reject transplanted MMC in FVBN202 mice. On the other hand, neu-specific immune responses appeared to be effective against MMC in parental FVB mice because of the fact that rat neu protein was seen as "nonself" antigen in these animals and the protein was dangerously overexpressed in MMC. Interestingly, low/intermediate expression of the neu "nonself" protein in tumors induced immune responses but such immune responses failed to reject the tumor in FVB mice. Our results showed that self-nonself (SNS) entity of a tumor antigen or danger signal alone, while may equally induce an antigen-specific immune response, will not warrant the efficacy of immune responses against tumors. On the other hand, entity of antigen in the context of dangerous conditions, i.e. abnormal/dangerous overexpression of the neu nonself protein, will warrant effective anti-tumor immune responses in FVB mice. This unified "danger-SNS" model suggests focusing on identification of naturally processed cryptic or mutated epitopes, which are considered semi-nonself by the host immune system, along with novel dangerous adjuvant in vaccine design.     

Combination of Id2 Knockdown Whole Tumor Cells and Checkpoint Blockade: A Potent Vaccine Strategy in a Mouse Neuroblastoma Model

Tumor vaccines have held much promise, but to date have demonstrated little clinical success. This lack of success is conceivably due to poor tumor antigen presentation combined with immuno-suppressive mechanisms exploited by the tumor itself. Knock down of Inhibitor of differentiation protein 2 (Id2-kd) in mouse neuroblastoma whole tumor cells rendered these cells immunogenic. Id2-kd neuroblastoma (Neuro2a) cells (Id2-kd N2a) failed to grow in most immune competent mice and these mice subsequently developed immunity against further wild-type Neuro2a tumor cell challenge. Id2-kd N2a cells grew aggressively in immune-compromised hosts, thereby establishing the immunogenicity of these cells. Therapeutic vaccination with Id2-kd N2a cells alone suppressed tumor growth even in established neuroblastoma tumors and when used in combination with immune checkpoint blockade eradicated large established tumors. Mechanistically, immune cell depletion studies demonstrated that while CD8+ T cells are critical for antitumor immunity, CD4+ T cells are also required to induce a sustained long-lasting helper effect. An increase in number of CD8+ T-cells and enhanced production of interferon gamma (IFNγ) was observed in tumor antigen stimulated splenocytes of vaccinated mice. More importantly, a massive influx of cytotoxic CD8+ T-cells infiltrated the shrinking tumor following combined immunotherapy. These findings show that down regulation of Id2 induced tumor cell immunity and in combination with checkpoint blockade produced a novel, potent, T-cell mediated tumor vaccine strategy.     

Brave Little World: Spheroids as an in vitro Model to Study Tumor-Immune-Cell Interactions

Multicellular tumor spheroids (MCTS) are a well established 3-D in vitro model system that reflects the pathophysiological in vivo situation in tumor microregions and of avascular micrometastatic sites. Because monocytes and other immune cells infiltrate into MCTS of different origin, such spheroid co-cultures are a valuable, still underestimated tool to systematically study heterologous interactions between tumor and immune cells. The present article gives a brief overview on work that has been published on tumor - immune cell interactions in MCTS and also summarizes mechanisms of immune suppression in the tumor milieu focussing on myeloid cells. Using the co-culture model, we recently demonstrated that tumor-derived lactic acid is a potent modulator of human monocyte as lactic acid inhibited the differentiation of monocytes (MO) into dendritic cells (DC) and also impaired antigen presentation. We show herein, that the capacity of various tumor cells in MCTS to secrete lactic acid differs up to tenfold, suggesting that this capacity is dependent on the tumor cell type. It is further demonstrated that lactic acid disturbs the migration of MO into MCTS as infiltration could be increased by blocking lactic acid production. We therefore discuss lactic acid which accumulates in many tumors and tumor microregions as a potent immune suppressor for MO/DC in the tumor milieu and conclude that these data are highly relevant for adoptive immunotherapy protocols with DC.     

Costimulation of antitumor immunity by the B7 counterreceptor for the T lymphocyte molecules CD28 and CTLA-4.

Interaction of the B7 molecule on antigen-presenting cells with its receptors CD28 and CTLA-4 on T cells provides costimulatory signals for T cell activation. We have studied the effects of B7 on antitumor immunity to a murine melanoma that expresses a rejection antigen associated with the E7 gene product of human papillomavirus 16. While this E7+ tumor grows progressively in immunocompetent hosts, cotransfection of its cells with B7 led to tumor regression by a B7-dependent immune response mediated by CD8+ cytolytic T lymphocytes. The immune response induced by E7+B7+ tumor cells also caused regression of E7+B7- tumors at distant sites and was curative for established E7+B7- micrometastases. Our findings suggest that increasing T cell costimulation through the CD28 and CTLA-4 receptors may have therapeutic usefulness for generating immunity against tumors expressing viral antigens.     

Inhibition of lens fiber cell morphogenesis by expression of a mutant SV40 large T antigen that binds CREB-binding protein/p300 but not pRb

Simian virus (SV) 40 large T antigen can both induce tumors and inhibit cellular differentiation. It is not clear whether these cellular changes are synonymous, sequential, or distinct responses to the protein. T antigen is known to bind to p53, to the retinoblastoma (Rb) family of tumor suppressor proteins, and to other cellular proteins such as p300 family members. To test whether SV40 large T antigen inhibits cellular differentiation in vivo in the absence of cell cycle induction, we generated transgenic mice that express in the lens a mutant version of the early region of SV40. This mutant, which we term E107KDelta, has a deletion that eliminates synthesis of small t antigen and a point mutation (E107K) that results in loss of the ability to bind to Rb family members. At embryonic day 15.5 (E15.5), the transgenic lenses show dramatic defects in lens fiber cell differentiation. The fiber cells become post-mitotic, but do not elongate properly. The cells show a dramatic reduction in expression of their beta- and gamma-crystallins. Because CBP and p300 are co-activators for crystallin gene expression, we assayed for interactions between E107KDelta and CBP/p300. Our studies demonstrate that cellular differentiation can be inhibited by SV40 large T antigen in the absence of pRb inactivation, and that interaction of large T antigen with CBP/p300 may be enhanced by a mutation that eliminates the binding to pRb.     

Recombinant polyoma--vaccinia viruses: T antigen expression vectors and anti-tumor immunization agents

Live vaccinia virus recombinants expressing viral antigens have recently been developed as effective anti-viral vaccines. We have examined the possibility of extending this approach to specific anti-tumor immunity, using tumors induced by the polyoma virus (PyV) as a model system. Three recombinant vaccinia viruses, separately encoding the three early proteins of the polyoma virus (large, middle and small tumor (T) antigens) were constructed. Each recombinant efficiently expresses the appropriate T antigen, which exhibits biochemical properties and subcellular localization of the authentic PyV protein. The potential of the recombinants to elicit immunity towards PyV-induced tumors was assessed in rats by a challenge injection of syngeneic PyV-transformed cells. After prior immunization with the large-T or the middle-T viruses, small tumors developed, which later regressed and were eliminated in more than 50% of the animals. In contrast, the small-T virus failed to elicit tumor rejection. Established tumors could also be eliminated by curative vaccinations. No circulating antibodies directed against PyV large-T or middle-T antigens were detected in animals vaccinated with the large-T or middle-T viruses, suggesting that rejection may be due to a cell-mediated immune response.     

Humoral immunity directed against tumor-associated antigens as potential biomarkers for the early diagnosis of cancer

Over the past decade, it has been demonstrated that cancer is immunogenic, and multiple tumor antigens have been identified in cancer patients. It is now possible to potentially harness the immune response elicited by cancer growth as a potential diagnostic tool. Humoral immunity, or the development of autoantibodies against tumor-associated proteins, may be used as a marker for cancer exposure. Unlike circulating proteins that are shed by bulky tumors, serum autoantibodies are detectable even when antigen expression is minimal. This paper will review the methods used for tumor antigen discovery and overview what is known about autoantibodies targeting common cancer antigens with a focus on breast cancer. Data will be presented modeling the use of tumor antigen associated autoantibodies as a breast cancer diagnostic. The endogenous humoral immune response present in cancer patients may allow the identification of individuals exposed to the malignant transformation of somatic cells.     

Isolation of immune cells from primary tumors

Tumors create a unique immunosuppressive microenvironment (tumor microenvironment, TME) whereby leukocytes are recruited into the tumor by various chemokines and growth factors. However, once in the TME, these cells lose the ability to promote anti-tumor immunity and begin to support tumor growth and down-regulate anti-tumor immune responses. Studies on tumor-associated leukocytes have mainly focused on cells isolated from tumor-draining lymph nodes or spleen due to the inherent difficulties in obtaining sufficient cell numbers and purity from the primary tumor. While identifying the mechanisms of cell activation and trafficking through the lymphatic system of tumor bearing mice is important and may give insight to the kinetics of immune responses to cancer, in our experience, many leukocytes, including dendritic cells (DCs), in tumor-draining lymph nodes have a different phenotype than those that infiltrate tumors. Furthermore, we have previously demonstrated that adoptively-transferred T cells isolated from the tumor-draining lymph nodes are not tolerized and are capable of responding to secondary stimulation in vitro unlike T cells isolated from the TME, which are tolerized and incapable of proliferation or cytokine production. Interestingly, we have shown that changing the tumor microenvironment, such as providing CD4(+) T helper cells via adoptive transfer, promotes CD8(+) T cells to maintain pro-inflammatory effector functions. The results from each of the previously mentioned studies demonstrate the importance of measuring cellular responses from TME-infiltrating immune cells as opposed to cells that remain in the periphery. To study the function of immune cells which infiltrate tumors using the Miltenyi Biotech isolation system, we have modified and optimized this antibody-based isolation procedure to obtain highly enriched populations of antigen presenting cells and tumor antigen-specific cytotoxic T lymphocytes. The protocol includes a detailed dissection of murine prostate tissue from a spontaneous prostate tumor model (TRansgenic Adenocarcinoma of the Mouse Prostate -TRAMP) and a subcutaneous melanoma (B16) tumor model followed by subsequent purification of various leukocyte populations.     

Experimental manipulations of afferent immune responses influence efferent immune responses to brain tumors

Tumors grow more readily in the brain than in the periphery, in part due to immune privilege. Differences in both afferent and efferent components of the immune response contribute to this lower level of responsiveness. On the afferent side, despite the lack of lymphatic vessels in the brain, antigens from brain arrive in lymph nodes and spleen by several routes, and the route taken may influence the type of response generated. Work with viruses and soluble antigens in mice has shown that the intracerebral location and the volume of the inoculation influence the strength of the cytotoxic T cell response. We examined whether these factors influence the T cell response against experimental brain tumors in mice. Placement of tumor cells in the cerebral ventricles instead of the parenchyma generated an immune response sufficient to increase survival time. A large volume of an intraparenchymal infusion of tumor cells caused spread of cells to the ventricles, and resulted in longer survival time relative to a small volume infusion. Infusion of the same dose of radiolabeled tumor cells in either a small volume or a large volume allowed tracking of potential tumor antigens to the periphery. Both modes of infusion resulted in similar levels of radioactivity in blood, spleen and kidney. Unexpectedly, cells infused intraparenchymally in a small volume, compared to a large volume, resulted in (1) more radioactivity in cervical lymph nodes (parotid and deep cervical lymph nodes), (2) a greater number of CD11b+/Gr1+ myeloid suppressor cells in the tumors, and (3) fewer CD8+ cells within the tumor mass. Consistent with these observations, providing a stronger afferent stimulus by giving a concurrent subcutaneous injection of the same tumor cells infused into the brain increased CD8+ T cell infiltration of the tumor in the brain. These results suggest that the immune response elicited by antigens that drain predominantly to the cervical lymph nodes may be less effective than responses elicited at other lymph nodes, perhaps due to immunosuppressive cells. Directing therapies to the optimal peripheral sites may improve immune responses against brain tumors.     

Mathematical model of immune processes

In the model the time lags of the antibody production and immune memory formation are taken into account explicitly. The antibody-antigen reaction is supposed to be very fast. The cases of a reproducing antigen as well as that of a non-reproducting antigen are considered. The conditions of the infinite increase of the antigen quantity and of the antigen elimination are obtained. For the rapidly reproducing antigen the latter condition includes the requirement for the time lag of the immune response to be not too short or not too long. In the case of the poorly catabolized non-reproducing antigen the cyclic appearance of the antibody producing cells due to the immune memory is described in the frame-work of the model.The mathematical structure of the model is similar to that of the Volterra-Lotka jequations. The only difference is the presence of the time lags in the non-linear terms. The time lags lead to the instability of the stationary state. In the prolonged reaction the antigen quantity may perform several oscillations before the elimination of the antigen.     

Trogocytosis of MHC-I/peptide complexes derived from tumors and infected cells enhances dendritic cell cross-priming and promotes adaptive T cell responses

The transporter associated with antigen processing (TAP) and the major histocompatibility complex class I (MHC-I), two important components of the MHC-I antigen presentation pathway, are often deficient in tumor cells. The restoration of their expression has been shown to restore the antigenicity and immunogenicity of tumor cells. However, it is unclear whether TAP and MHC-I expression in tumor cells can affect the induction phase of the T cell response. To address this issue, we expressed viral antigens in tumors that are either deficient or proficient in TAP and MHC-I expression. The relative efficiency of direct immunization or immunization through cross-presentation in promoting adaptive T cell responses was compared. The results demonstrated that stimulation of animals with TAP and MHC-I proficient tumor cells generated antigen specific T cells with greater killing activities than those of TAP and MHC-I deficient tumor cells. This discrepancy was traced to differences in the ability of dendritic cells (DCs) to access and sample different antigen reservoirs in TAP and MHC-I proficient versus deficient cells and thereby stimulate adaptive immune responses through the process of cross-presentation. In addition, our data suggest that the increased activity of T cells is caused by the enhanced DC uptake and utilization of MHC-I/peptide complexes from the proficient cells as an additional source of processed antigen. Furthermore, we demonstrate that immune-escape and metastasis are promoted in the absence of this DC 'arming' mechanism. Physiologically, this novel form of DC antigen sampling resembles trogocytosis, and acts to enhance T cell priming and increase the efficacy of adaptive immune responses against tumors and infectious pathogens.     

Topoisomerase inhibitors modulate expression of melanocytic antigens and enhance T cell recognition of tumor cells

While there are many obstacles to immune destruction of autologous tumors, there is mounting evidence that tumor antigen recognition does occur. Unfortunately, immune recognition rarely controls clinically significant tumors. Even the most effective immune response will fail if tumors fail to express target antigens. Importantly, reduced tumor antigen expression often results from changes in gene regulation rather than irrevocable loss of genetic information. Such perturbations are often reversible by specific compounds or biological mediators, prompting a search for agents with improved antigen-enhancing properties. Some recent findings have suggested that certain conventional chemotherapeutic agents may have beneficial properties for cancer treatment beyond their direct cytotoxicities against tumor cells. Accordingly, we screened an important subset of these agents, topoisomerase inhibitors, for their effects on antigen levels in tumor cells. Our analyses demonstrate upregulation of antigen expression in a variety of melanoma cell lines and gliomas in response to nanomolar levels of certain specific topoisomerase inhibitors. To demonstrate the ability of CD8+ T cells to recognize tumors, we assayed cytokine secretion in T cells transfected with T cell receptors directed against Melan-A/MART-1 antigen. Three days of daunorubicin treatment resulted in enhanced antigen expression by tumor cells, in turn inducing co-cultured antigen-specific T cells to secrete Interleukin-2 and Interferon-γ. These results demonstrate that specific topoisomerase inhibitors can augment melanoma antigen production, suggesting that a combination of chemotherapy and immunotherapy may be of potential value in the treatment of otherwise insensitive cancers.     

Expression of human prostate-specific antigen (PSA) in a mouse tumor cell line reduces tumorigenicity and elicits PSA-specific cytotoxic T lymphocytes

 Human prostate-specific antigen (PSA) has a highly restricted tissue distribution. Its expression is essentially limited to the epithelial cells of the prostate gland. Moreover, it continues to be synthesized by prostate carcinoma cells. This makes PSA an attractive candidate for use as a target antigen in the immunotherapy of prostate cancer. As a first step in characterizing the specific immune response to PSA and its potential use as a tumor-rejection antigen, we have incorporated PSA into a well-established mouse tumor model. Line 1, a mouse lung carcinoma, and P815, a mouse mastocytoma, have been transfected with the cDNA for human PSA. Immunization with a PSA-expressing tumor cell line demonstrated a memory response to PSA which protected against subsequent challenge with PSA-expressing, but not wild-type, tumors. Tumor-infiltrating lymphocytes could be isolated from PSA-expressing tumors grown in naive hosts and were specifically cytotoxic against a syngeneic cell line that expressed PSA. Immunization with tumor cells resulted in the generation of primary and memory cytotoxic T lymphocytes (CTL) specific for PSA. The isolation of PSA-specific CTL clones from immunized animals further demonstrated that PSA can serve as a target antigen for antitumor CTL. The immunogenicity studies carried out in this mouse tumor model provide a rationale for the design of methods to elicit PSA-specific cell-mediated immunity in humans. Received: 4 April 1996 / Accepted: 31 May 1996