摘要:

The characterization of tumor-associated antigens and of human leukocyte antigen (HLA) class I molecule-binding peptide epitopes derived from these antigens has prompted the initiation of various vaccination trials aimed at inducing tumor-specific CD8+T cells in persons with cancer. Sensitive and easy-to-perform T-cell assays that assess the frequency of tumor-reactive T cells are crucial for the evaluation and further development of vaccination approaches. This review focuses on a novel ELISPOT technique that allows quantification of tumor-specific T lymphocytes from peripheral blood by detecting antigen-induced cytokine secretion. Various ELISPOT methods using different antigen-presenting cells and different cytokines as read-out are described. T-cell analyses performed using the standard chromium release assay and the ELISPOT assay are also compared. Results from various clinical trials, including peptide and whole tumor cell vaccination and cytokine treatment, are now available and show the suitability of the ELISPOT assay for monitoring T-cell responses. To establish a basis for standardization and to further improve this technique, the first comparative quality assurance studies analyzing T-cell frequencies in different laboratories with the ELISPOT assay are being performed.

ISSN:1524-9557    

出版商:OVID    

年代:2000

数据来源: OVID

摘要:

Peptide antigens available for use in specific immunotherapy of patients with cancer have not been fully determined. Although the authors have reported the SART1 gene encoding epitopes recognized by HLA-A2601–restricted and tumor-specific cytotoxic T lymphocytes (CTLs), the HLA-A26 allele is mainly subdivided into A2601, A2602, and A2603 subtypes. In this study, the authors attempted to determine whether the SART1-derived peptide at position 736-744 (KGSGKMKTE) is suitable to induce HLA-A26–restricted and tumor-specific CTLs in patients with cancer who have these subtypes. This peptide induced the HLA-A26 subtype-restricted and tumor-specific CTLs in HLA-A2601+or HLA-A2603+peripheral blood mononuclear cells, respectively. It also induced the HLA-A26–restricted CTL activity in HLA-A2602+peripheral blood mononuclear cells. Therefore, this peptide could be useful for specific immunotherapy of patients with cancer who have any of the three HLA-A26 subtypes.

ISSN:1524-9557    

出版商:OVID    

年代:2000

数据来源: OVID

摘要:

Recently, highly efficient natural killer-like T immunologic effector cells called cytokine-induced killer (CIK) cells have been described. Most interestingly, CIK cells have been shown to eradicate established human lymphoma cells in a severe combined immunodeficient (SCID) mouse xenograft model in vivo. The current study was aimed at increasing the sensitivity of leukemia and lymphoma cells to CIK cells. In particular, the authors wanted to target CIK cells to leukemia and lymphoma cells via reverse antibody-dependent cellular cytotoxicity. Binding of an anti-CD3 monoclonal antibody to CIK cell cultures derived from patients with lymphoma was shown using flow cytometric analysis. For the target side, several B-cell lines were found to express CD19 on the cell surface. There was an impressive increase in sensitivity to CIK-mediated lysis of various lymphoma and leukemia cell lines by preincubation of the targets with a monoclonal antibody against CD3. This increase could be partially blocked by preincubation with anti-CD16 (Fc receptor III) and anti-CD32 (Fc receptor II) antibodies. These data suggest that the increase in cytotoxic activity is caused by Fc receptor-mediated antibody binding. Cytotoxic activity could be further increased by adding an anti-CD28 antibody in addition to anti-CD3. Finally, there was a further increase in sensitivity to CIK-mediated lysis of CD19+malignant cells using the bispecific OKT3xHD37 antibody with specificity against CD3 and CD19. Interestingly, preincubation of malignant cells with an anti-CD3 monoclonal antibody followed by addition of the bispecific OKT3xHD37 antibody led to a further increase of cytotoxic sensitivity compared with the addition of the bispecific antibody alone. In conclusion, these data suggest that cytotoxic activity of immunologic effector cells can be increased not only by using the bispecific antibody OKT3xHD37 in vitro but also by preincubation of CD19+leukemia and lymphoma cells with a monoclonal antibody against CD3. In addition, the immunostimulatory effect of the bispecific antibody OKT3xHD37 can be further increased by adding a monoclonal antibody against CD3.

ISSN:1524-9557    

出版商:OVID    

年代:2000

数据来源: OVID

摘要:

The authors previously showed that monocytes treated with calcium ionophore (CI) acquire characteristics of mature dendritic cells (DC) in part through a calcineurin-dependent pathway. In this study, the authors evaluated the ability of granulocyte-macrophage colony stimulating factor (GM-CSF), interleukin-2 (IL-2), and interleukin-12 (IL-12) alone or in combination with CI to induce DC characteristics in peripheral blood monocytes. Monocytes obtained by leukapheresis and countercurrent centrifugal elutriation were cultured with calcium, cytokines, or both, profiled by flow cytometry, and assessed for antigen uptake and sensitization of autologous CD8+T cells to antigen. Monocytes treated with the combination of GM-CSF, IL-2, and IL-12 resulted in immunophenotypic and antigen uptake profiles typical of immature DC, including loss of surface CD14 expression, de novo low-level expression of B7.1, negligible CD83 expression, marked enhancement of CD40 and ICAM-1, and high major histocompatibility complex class I and II levels. A high level of antigen uptake by macropinocytosis was observed. In contrast, CI treatment significantly upregulates B7.1, B7.2, CD40, CD54, and CD83 and substantially downregulates CD14 and macropinocytosis, a profile consistent with mature DC. Many CI-induced modulations, but none resulting from cytokine treatment alone, were inhibited by the calcineurin phosphatase inhibitor cyclosporin A. Compared with monocytes treated with CI alone, combined treatment of monocytes with GM-CSF, IL-2, IL-12, and CI augmented B7.1 and CD83 expression and enhanced sensitization of autologous CD8+T cells to melanoma-antigen–derived peptides. These results suggest that several independent pathways of DC activation can cooperatively enhance the function of monocyte-derived DC.

ISSN:1524-9557    

出版商:OVID    

年代:2000

数据来源: OVID

摘要:

The authors recently reported that tumoricidal activation of macrophages by a new synthetic bacterial lipopeptide, JBT 3002, can augment chemotherapy-mediated tumor-cell killing. The aim of this study was to identify the mechanism responsible for the destruction of metastatic cells. Three daily oral doses of JBT 3002 before once-weekly intraperitoneal injections of 100 mg/kg irinotecan for 3 weeks significantly increased the eradication of established CT-26 murine colon cancer liver metastases compared with treatment with irinotecan alone. Immunohistochemical analyses revealed that the hepatic metastases in mice given combination therapy contained infiltrating CD8+lymphocytes and a dense infiltrate of macrophages expressing both inducible nitric oxide synthase (iNOS) and interleukin-15. In vitro treatment of peritoneal macrophages with JBT 3002 plus interferon-&ggr; induced the expression of iNOS and the production of nitric oxide. In the presence of a low (subtoxic) dose of irinotecan, these activated macrophages produced significant lysis of CT-26 cells. The high level of cytotoxicity was inhibited by the specific inducible nitric oxide synthase inhibitor, NG-methyl-L-arginine. In contrast, irinotecan-mediated lysis of normal intestinal epithelial IEC-6 cells was not increased by activated macrophages. Scanning electron microscopy revealed that activated macrophages bound to CT-26 tumor cells but not to normal IEC-6 cells, confirming that nitric oxide–mediated cytotoxicity is specific for tumor cells. Collectively, the results suggest that augmentation of the therapeutic efficacy of irinotecan against colon cancer liver metastases by immunomodulation with JBT 3002 may be associated with elevated inducible nitric oxide synthase and endogenous interleukin-15 in tumor-infiltrating macrophages.

ISSN:1524-9557    

出版商:OVID    

年代:2000

数据来源: OVID

摘要:

Chimeric immunoglobulin T-cell receptors (IgTCR) join the antigen-binding portion of an antibody to one of the signaling chains of the TCR. A previous report described the construction and functional testing of an IgTCR gene directed against the carcinoembryonic tumor antigen (CEA). These preclinical studies showed the proper assembly and cell surface expression of anti-CEA IgTCR molecules, specific target antigen binding, and activation of T-cell effector functions. Although IgTCR-modified T cells function well in vitro, therapeutic applications in humans may be complicated by various factors, such as the availability of appropriate T-cell cytokines, high systemic levels of antagonistic soluble CEA, and antigenic diversity in tumor cell populations. The current study analyzes tumor cell killing by IgTCR-modified human T cells under conditions that more closely model those that may be encountered in persons with cancer. This analysis shows that 1) depriving IgTCR-modified T cells of interleukin-2 does not diminish anti-CEA cytotoxic T lymphocyte activity, but does eliminate killing by lymphokine-activated killer cells; 2) high levels of soluble CEA do not significantly inhibit tumor cell killing even when approximately 80% of the chimeric receptors are blocked; and 3) CEA+tumor cells that can downregulate cell surface CEA evade immune destruction by IgTCR-modified T cells. These results have important implications for application strategies and protocol design considerations for early clinical testing of IgTCR anti-tumor therapies.

ISSN:1524-9557    

出版商:OVID    

年代:2000

数据来源: OVID

摘要:

Peptide vaccination of homozygous mice against syngeneic tumors using single major histocompatibility complex (MHC) class I–restricted cytotoxic T lymphocyte (CTL) epitopes elicits effective immune responses against metastatic growth. So far, single-peptide vaccination of patients against their autologous tumors seems to elicit less satisfactory results. In this study, the authors tried to determine whether effective anti-metastatic immunity requires the presentation of peptides restricted by the two parental class I major histocompatibility complex alleles in heterozygous hosts. The immune response against the H-2b–derived 3LL Lewis lung carcinoma was evaluated in heterozygous recombinant congenic F1 mice (Kk× Kb) and (Kd× Kb). Vaccination of such heterozygous animals with dendritic cells expressing the two parental H-2K alleles, pulsed with total tumor extract, elicited a potent anti-metastatic response. A comparable response was obtained after vaccination with tumor cells genetically modified to express the two class I alleles. In contrast, vaccination of the heterozygous mice with dendritic cells expressing only one of the parental F1 H-2K alleles or with tumors expressing only one H-2K allele failed to elicit effective immunity against tumor metastasis in recombinant congenic F1 mice. It appears, therefore, that to achieve effective anti-metastatic immunotherapy in heterozygous organisms, presentation of cytotoxic T lymphocyte epitopes restricted by the two parental class I alleles is required.

ISSN:1524-9557    

出版商:OVID    

年代:2000

数据来源: OVID

摘要:

Melanoma cells are unusual because, unlike most epithelial tumors, constitutive expression of human leukocyte antigen (HLA) class II molecules is common. To elucidate the role of HLA class II expression in the immunopathogenesis of melanoma, the authors compared HLA class II+melanoma cells to autologous B cells with respect to their ability to stimulate primary (naïve) histoincompatible lymphocytes and T-cell clones (antigen experienced). Using primary lymphocytes (peripheral blood lymphocytes [PBLs]), melanoma cells were nonstimulatory when compared to autologous B cells. To determine whether this was caused by defective antigen processing, the authors used alloreactive T-cell clones, which require alloantigen presentation by a histocompatible stimulator cell but not costimulation. Melanoma cells stimulated the alloreactive T-cell clones in two of three clones tested, indicating that they processed and presented alloantigen. To determine whether the failure of melanoma cells to stimulate primary lymphocytes was caused by their inability to costimulate the T cells, the authors transduced the melanoma cells with B7.1 and achieved stable expression in more than 95% of the cells. The transduced cells were highly stimulatory, eliciting a 17-to 25-fold increase in proliferation by the peripheral blood lymphocytes compared with controls. Indeed, B7-expressing melanoma cells were more stimulatory than autologous B cells, which elicited an 11-to 15-fold increase compared with controls. These data indicate that melanoma cells fail to stimulate primary lymphocytes because they do not deliver costimulatory signals. Engineering HLA class II+melanoma cells to express high levels of B7.1 may provide a way to elicit primary T-cell responses to melanoma-associated antigens.

ISSN:1524-9557    

出版商:OVID    

年代:2000

数据来源: OVID

摘要:

A monoclonal antibody specific for the human analog of the murine T-cell activation molecule 4-1BB was generated and is shown here to react selectively with activated human CD4+and CD8+T lymphocytes. Treatment of these T cells in a one-way mixed lymphocyte culture with the anti–h4-1BB antibody enhanced the cell proliferation of the allostimulated lymphocytes. Previous studies in the mouse have shown that treatment of tumor-bearing mice with antibodies to 4-1BB augments anti-tumor immunity that is mediated by both CD4+and CD8+T cells. The authors consider the possibility that a similar approach may be efficacious for human cancer immunotherapy. This question was addressed by evaluating the effect of an anti–h4-1BB monoclonal antibody on human lymphocyte-mediated suppression of a human tumor xenograft in SCID mice. Mice treated with a control antibody and co-injected with the tumor and peripheral blood lymphocytes exhibited a lymphocyte dose-dependent suppression of tumor growth. In mice treated with the anti–h4-1BB antibody, the lymphocyte-mediated tumor suppression was completely eliminated and tumors grew progressively (as was observed in mice inoculated with tumors without lymphocytes). This monoclonal antibody specific for anti–h4-1BB, which augments the proliferation of allostimulated cells in vitro, blocks T-cell anti-tumor activity in vivo. These results suggest that although 4-1BB plays a role in the human peripheral blood lymphocyte–mediated suppression of tumor growth, antibodies to this molecule on human cells fail to stimulate anti-tumor activity, as was observed in tumor-bearing mice treated with an antibody to murine 4-1BB.

ISSN:1524-9557    

出版商:OVID    

年代:2000

数据来源: OVID

摘要:

The combination of interferon-&agr; (IFN-&agr;) plus interleukin (IL-2) has been accepted in the treatment of metastatic renal cell carcinoma (MRCC), whereas vaccines based on IL-12 or dendritic cells (DCs) are still being investigated. Here the authors analyzed 1) the feasibility to generate functional monocyte-derived DCs (MDDCs) from patients treated with biological response modifiers (BRMs) who have MRCC, 2) the phenotypic modulations of these MDDCs during BRM treatment. Eight and 13 MRCC patients received IL-2 plus IFN-&agr; or IL-12 immunotherapy, respectively. The adherent fraction of mononuclear cells from patients' blood drawn before, during, and after immunotherapy was incubated in clinically approved culture medium supplemented with 5% autologous serum, rhu granulocyte macrophage colony-stimulating factor, and rhuIL-4 for a week. At day 7 or 8 of culture, floating cells were examined in flow cytometric and functional assays (alloreactivity, proliferation assays in the presence of tetanus toxoid or tumor peptides, IL-12 secretion). In all patients except two, MDDCs could be generated but at a lower rate compared with healthy volunteers. Morphologic and phenotypical analyses revealed immature DCs with low levels of CD1a or CD83 expression throughout therapy with BRMs. Capacities in mixed leukocyte reactions were similar to those of healthy volunteers and stable during immunotherapy, whereas presentation of major histocompatibility complex class II tetanus toxoid peptide complexes was slightly enhanced during and after IL-12 therapy. IL-12 expression levels under IFN-&ggr; and CD40L stimulation were significantly lower in MDDC cultures from patients with MRCC compared with healthy volunteers. Overall, peripheral blood mononuclear cells from a cohort of 21 patients with metastatic disease who were treated with BRMs maintained their ability to differentiate into functional MDDCs with no selective quantitative or qualitative advantage.

ISSN:1524-9557    

出版商:OVID    

年代:2000

数据来源: OVID