Dendritic cell-based immunotherapy of prostate cancer: immune monitoring of a phase II clinical trial.

We assessed both non- and peptide-specific immune responses in prostate cancer patients before and after immunotherapy with dendritic cells exogenously pulsed with the prostate-specific membrane antigen-derived peptides, PSM-P1 and PSM-P2. For all subjects, we observed that clinical responses were strongly associated with two indicators of immunocompetence: skin test responses to recall antigens and cytokine secretion by T cells after nonspecific stimulation. In a subset of responders, we observed cytokine secretion or cytotoxicity against the immunizing peptides or an immunodominant epitope from an influenza recall antigen. The clinical results support the use of monitoring for overall immunocompetence to help determine why a patient has or has not responded to therapy. Moreover, it could be useful as an inclusion criterion to select those more likely to benefit from treatment.

Expression and purification of prostate-specific membrane antigen in the baculovirus expression system and recognition by prostate-specific membrane antigen-specific T cells.

Antigen-specific immunotherapy of cancer depends on a consistent source of well-defined protein antigen. Production of recombinant protein offers the obvious solution to this problem but few comparisons of recombinant and native proteins in cellular immune assays have been reported. We report expression of a putative immunotherapy antigen, prostate-specific membrane antigen (PSMA), in insect cells using a baculovirus vector. T cells stimulated with recombinant PSMA or native PSMA derived from the LNCaP cell line recognized both native PSMA and recombinant, baculoviral PSMA. These data indicate that PSMA produced in Sf9 cells is immunologically cross-reactive with native PSMA and therefore suitable for immunotherapy as it is recognized by both cellular and humoral immune responses.

Dendritic cell-based immunotherapy for prostate cancer.

Dendritic cells are unique in their ability to stimulate naive T cells. These investigators have developed a prostate cancer vaccine using autologous dendritic cells as a vehicle to present prostate antigens to T cells in vivo.

Use of cellular and cytokine adjuvants in the immunotherapy of cancer.

Cellular and cytokine adjuvants, often immune effector cells and soluble factors, respectively, are supplemental and/or follow-up treatments of human origin for cancer patients who have unsatisfactory clinical responses to conventional chemotherapy, radiotherapy, and surgery. Since many human studies with these reagents are in their infancy, extensive data collection is only now being performed to determine which strategy provides the greatest therapeutic benefit. Research published in the literature since the genesis of this approach to cancer treatment is summarized in this report. Methodologies attempting to generate anticancer responses by provoking or enhancing the patient's own immune system are new compared with the other standard types of cancer treatment. Although a few encouraging human studies can be discussed, many of the most promising techniques are only now being transferred from the laboratory to the clinic. The administration of immune effector cells in combination with immunomodulators, such as interferons or interleukins, often enhances clinical outcome. The literature cited in this report indicate that immune-cell- and cytokine-based therapies hold promise in our attempts to improve the quality and duration of life in those with cancer. With each report reaching the literature, more effective clinical trials are being designed and implemented.

Report of immune monitoring of prostate cancer patients undergoing T-cell therapy using dendritic cells pulsed with HLA-A2-specific peptides from prostate-specific membrane antigen (PSMA).

BACKGROUND: In this paper we describe our program for the immune monitoring of phase II participants given dendritic cell (DC)/prostate-specific membrane antigen (PSMA)-based immunotherapy, and we also present some initial findings. METHODS: Phase II subjects received six administrations of autologous dendritic cells exogenously pulsed with two peptides derived from PSMA. Prior to the initial infusion, and following each treatment, peripheral blood mononuclear cells (PBMC) were collected for the generation of dendritic cells as well as for comprehensive immune monitoring. RESULTS: Thus far, an increase in PSMA-peptide-specific as well as overall cellular reactivity has been observed in several patients receiving DC plus PSM-P1 and -P2, as measured by delayed-type hypersensitivity (DTH) test and enzyme-linked immunosorbant assay (ELISA). CONCLUSIONS: Our initial observations using an ELISA and DTH test indicate that we are enhancing cellular immunity in prostate cancer patients following infusion with DC plus PSMA-derived peptides. Several methods are underway to comprehensively monitor both cell-mediated and humoral immune responsiveness, including: determining anti-PSMA serum antibody titers, testing immunogen-restricted responder-cell proliferation and cytotoxicity, assessing aberrations in signal transduction, antigen processing, and presentation, and measuring soluble factors that may promote tumor outgrowth.

Dendritic cell-based immunotherapy of prostate cancer.

The immunotherapy of cancer, based on eliciting or enhancing the body's own capacity to mount an effective antitumor response, has produced encouraging early results in the areas of melanoma and renal-cell carcinoma. Such treatments utilizing dendritic cells (DC), immune cells that are excellent antigen presenters, are especially promising. We performed a phase I clinical trial assessing the administration of autologous DC pulsed with HLA-A0201-specific prostate-specific membrane antigen (PSMA) for the treatment of 51 men with hormone-refractory prostate cancer. Participants were divided into five groups receiving four or five infusions of peptides alone (PSM-P1 or PSM-P2; group 1 and 2, respectively), autologous DC (group 3), or DC pulsed with PSM-P1 or P2 (group 4 and 5, respectively). No significant toxicity was observed. Immune reactivity against PSM-P2 was detected in HLA-A2+ patients infused with DC pulsed with PSM-P1 or -P2 (group 4 and 5). An average decrease in PSA was observed only in group 5. Seven partial responders were identified based on NPCP criteria + PSA. The excellent tolerance of this treatment approach, as well as the enhanced cellular responses, decreased PSA levels, and partial clinical responses in some patients suggests that it holds great potential in prostate cancer therapy.

Cancer immunotherapy for prostate cancer.

Our approach to prostate cancer immunotherapy involves two components dendritic cells as antigen-presenting cells; and the antigen used to target T-cell attack, HLA-A0201-associated peptides from prostate specific membrane antigen (PSMA). We have conducted a phase I dose-ranging study in 51 men with advanced prostate cancer, using dendritic cells pulsed with a PSMA peptide. no significant toxicity was observed. In that study, T-cell response was enhanced, with seven men meeting NCPC and PSA criteria for partial response. We are now conducting a phase II study with 67 men, who will receive 6 infusions of dendritic cells that have been pulsed with 2 PSMA peptides, at 6-week intervals. The phase II study design and rationale is described in this paper.

Regulation of microglial activation by TGF-beta, IL-10, and CSF-1.

Microglia are the resident macrophages of the brain and as such are active participants in immune responses in the central nervous system. Normal resting microglia express low levels of MHC class I and class II antigens and do not produce proinflammatory cytokines. However, microglial immune functions are induced in areas of infection or injury. To understand regulation of cytokines that are secreted by and act upon microglia, we examined production of interleukin (IL) -12, tumor necrosis factor-alpha (TNF-alpha), and nitric oxide (NO) by lipopolysaccharide (LPS)-stimulated microglia. We observed secretion of IL-12, TNF-alpha, and NO following stimulation of microglia with LPS. Addition of IL-10 suppressed TNF-alpha, IL-12, and NO production. Transforming growth factor-beta (TGF-beta) also inhibited TNF-alpha and NO but did not affect IL-12 secretion. IL-12 secretion became sensitive to TGF-beta inhibition when microglia were cultured in the absence of CSF-1. In addition to its effect on the response to TGF-beta, CSF-1 suppressed the response of microglia to LPS. These data suggest that CSF-1 may contribute to the immunologically privileged status of the central nervous system.

The vascular endothelial cell is central to xenogeneic immune reactivity.

With the demand for organ transplants increasing, xenograft transplantation is being considered. A major obstacle to a solution to fulfilling this demand is xenograft rejection. Histologic evidence indicates that the vascular endothelial cell (VEC) is involved in both humoral and cellular aspects of xenograft rejection. To study this phenomenon murine VEC and splenocytes were used in a mixed lymphocyte/endothelial cell culture and in a mixed lymphocyte culture with human peripheral blood mononuclear cells as responders. The VEC is a much better stimulator than the splenocyte. Removal of macrophages and B cells from the human peripheral blood mononuclear cells has no effect on the response to the VEC. The VEC acts as a target for humoral responses in the complement-dependent cytotoxicity and the antibody-dependent cell-mediated cytotoxicity. The VEC is killed in these two assays, which indicates the importance of the VEC in humoral rejection. These data indicate that the VEC is an important cell in xenogeneic immune reaction and may be pivotal in xenograft rejection.

Coxsackievirus B-3 myocarditis. Identification of different pathogenic mechanisms in DBA/2 and Balb/c mice.

DBA/2 and Balb/c mice, both H-2d, develop myocardial inflammation and necrosis when infected with a heart-adapted strain of coxsackievirus Group B, Type 3. Similar inoculation of C57Bl/6 (H-2b) animals results in minimal myocarditis despite equivalent heart virus titers in the three stains. Thus, the host's genetic constitution influences the pathogenesis of the infection. Anti-mouse thymocyte serum and monoclonal Iad antibody effectively prevent myocarditis induction in DBA/2 and Balb/c mice, which demonstrates the importance of the immune system in this disease. Cytolytic T lymphocytes lysing virus-infected and uninfected myocytes and heart-reactive autoantibodies occur in both myocarditis-susceptible strains. Cellular immunity causes the myocardial injury in Balb/c mice. Complement depletion of Balb/c mice using cobra venom factor fails to alter the disease. Similar treatment of DBA/2 animals abrogates inflammation and necrosis, which suggests that heart-reactive antibodies in this strain are primarily responsible for initiating myocardial damage.

Cardiac injury in myocarditis induced by Coxsackievirus group B, type 3 in Balb/c mice is mediated by Lyt 2 + cytolytic lymphocytes.

Male Balb/c mice inoculated with a heart-adapted variant of Coxsackievirus, group B, type 3 (CVB3) develop severe myocarditis 7 days later. The lesions are characterized by mononuclear cell inflammation and myocyte necrosis. Infected T-lymphocyte-deficient mice show either minimal or no cardiac injury, although virus concentrations in the hearts of T-cell-deficient and -sufficient animals are similar. Adoptive transfer of 2 X 10(6) CVB3 immune Thy 1+ cells into CVB3-infected T-cell-deficient mice effectively restored myocarditis to levels observed in intact animals. Similar reconstitution with immune Ig+ cells or serum resulted in only a minimal increase in cardiac injury. To determine whether T-lymphocyte-dependent humoral or cellular immunity was responsible for myocarditis. T lymphocytes were obtained from Balb/c mice 6 days after infection with CVB3, separated into Lyt 1+2- (helper) and Lyt 1-2+ (cytolytic/suppressor) cell populations, and 2 X 10(6) of the enriched helper and cytolytic cells were adoptively transfused into infected T-cell-deficient recipients. Animals receiving the immune Lyt2+ cells developed severe myocarditis, had cytolytic T lymphocytes to both CVB3-infected and uninfected myocytes, but lacked a detectable IgG antibody response. Recipients of the Lyt 1+ cells failed to develop either myocarditis or cytolytic T cells but had normal serum IgG antibody titers to the virus. These results demonstrate that cardiac myocarditis is the product of cellular immune mechanisms.