Identification of cyclophilin B-derived peptides capable of inducing histocompatibility leukocyte antigen-A2-restricted and tumor-specific cytotoxic T lymphocytes.

We recently suggested that cyclophilin B (Cyp-B) is a tumor antigen recognized by histocompatibility leukocyte antigen (HLA)-A24-restricted and tumor-specific cytotoxic T lymphocytes (CTLs). In this study, we tried to identify Cyp-B-derived epitopes, which can induce HLA-A2-restricted and tumor-specific CTLs in cancer patients. The tumor-infiltrating lymphocytes (TILs) from an HLA-A0207 patient with colon cancer were found to respond to COS-7 cells when co-transfected with the Cyp-B gene and either HLA-A0201, -A0206, or -A0207 cDNA. These TILs contained CTLs capable of recognizing either the Cyp-B(129 - 138) or the Cyp-B(172 - 179) peptide among 28 different peptides, all of which were prepared based on the HLA-A2 binding motif. Both Cyp-B peptides possessed the ability to induce tumor-specific CTLs in HLA-A2(+) cancer patients. Cyp-B(172 - 180 (V)), which is a 9-mer peptide with valine added at the C terminus, showed no clear superiority over the parental Cyp-B(172 - 179) peptide in an in vitro sensitization experiment. In vitro-sensitized T cells with these peptides responded to cancer cells in an HLA-A2-restricted manner. These two Cyp-B peptides could be useful for specific immunotherapy of HLA-A2(+) cancer patients.

A new tumor-rejection antigen recognized by cytotoxic T lymphocytes infiltrating into a lung adenocarcinoma.

Lung cancer is the most commonly occurring malignancy worldwide and one of the few that continues to show an increasing incidence. To understand the molecular basis of host immunity against lung cancer, we investigated tumor antigens recognized by HLA-A24-restricted CTLs established from T cells infiltrating into lung adenocarcinoma and report a new gene coding for antigens recognized by the CTLs. The mRNA of this gene was expressed at different levels in all of the malignant cells tested (high in adenocarcinomas and gliomas and low in esophageal cancers and malignant hematological disease). It was also expressed at the different levels in each of a panel of normal tissues (high in the thymus, low in peripheral blood mononuclear cells, and lowest in the stomach, small intestine, and skeletal muscle). This gene encodes a Mr 60,000 nuclear protein with 414 deduced amino acids. The three peptides at positions 158-165, 170-179, and 188-196 were recognized by the CTLs. One peptide at position 188-196 had the ability to induce HLA-A24-restricted and tumor-specific CTLs in peripheral blood mononuclear cells of lung cancer patients. These CTLs, however, did not lyse HLA-A24+ PHA-activated T cells in which the mRNA of this gene was highly expressed, even in the presence of excess amounts of a corresponding peptide in culture. These results suggest that this gene product and peptide could be applicable to specific immunotherapy of lung cancer patients.

Expression of SART3 tumor-rejection antigen in gastric cancers.

We previously reported SART3 as a tumor-rejection antigen recognized by histocompatibility leukocyte antigen (HLA)-A24-restricted cytotoxic T lymphocytes (CTLs). In this study, we investigated the expression of the SART3 antigen in gastric cancers, as a candidate for use in specific immunotherapy. The SART3 antigen was detected in 9 of 10 (90%) gastric cancer cell lines, 35 of 52 (67.3%) gastric cancer tissues, and 0 of 20 non-tumorous gastric tissues. SART3-derived peptides corresponding to positions 109- 118 and 315-323 induced HLA-A24-restricted and tumor-specific CTLs from peripheral blood mononuclear cells (PBMCs) of gastric cancer patients. These peptide-induced CTLs recognized HLA-A24(+) SART3(+) gastric cancer cells, but not HLA-A24(+) SART3(-) or HLA-A24(-) SART3(+) gastric cancer cells. Therefore, the SART3 peptides could be useful in specific immunotherapy of gastric cancer patients.

Cytokines required for induction of histocompatibility leukocyte antigen-class I-restricted and tumor-specific cytotoxic T lymphocytes by a SART1-derived peptide.

Although there have been several reports on peptides of human tumor-rejection antigens capable of inducing histocompatibility leukocyte antigen (HLA)-class I-restricted and tumor-specific cytotoxic T lymphocytes (CTLs), it is not yet clear which cytokines are required for CTL induction. This study has investigated the cytokine combinations required for optimal induction of CTLs by SART1(690-698) peptide, which is capable of inducing HLA-A24-restricted and tumor-specific CTLs in peripheral blood mononuclear cells (PBMCs). Pretreatment of PBMCs as a source of antigen-presenting cells (APCs) with interferon (IFN)-gamma, or to some extent with IFN-alpha, but not with any of the other cytokines tested, augmented the peptide-induced CTL activity in HLA-A24 heterozygotes, but not in HLA-A24 homozygotes. This IFN-gamma-mediated augmentation was inhibited by either interleukin (IL)-4 or IL-10. IL-2 alone in culture, along with weekly stimulation by peptide-pulsed APCs, was sufficient for the differentiation and proliferation of CTLs for the initial several weeks of culture. This IL-2-mediated activation of CTLs was inhibited by the addition of IFN-gamma, IL-4, or IL-10 to the IL-2 culture. For further expansion of the CTLs, dendritic cells (DCs) induced from PBMCs with IL-4 and granulocyte macrophage colony-stimulating factor (GM-CSF) were required as APCs. These results indicate that IFN-gamma and IL-2 are important in the activation of APCs and CTLs, respectively, while GM-CSF and IL-4 are needed for the induction of DCs, which in turn are required for further expansion of mature CTLs. These results are important in allowing for a better understanding of the cellular and molecular basis of tumor-specific immunity, and also for the development of peptide-based specific immunotherapy.

A cyclophilin B gene encodes antigenic epitopes recognized by HLA-A24-restricted and tumor-specific CTLs.

We have studied Ags recognized by HLA class I-restricted CTLs established from tumor site to better understand the molecular basis of tumor immunology. HLA-A24-restricted and tumor-specific CTLs established from T cells infiltrating into lung adenocarcinoma recognized the two antigenic peptides encoded by a cyclophilin B gene, a family of genes for cyclophilins involved in T cell activation. These two cyclophilin B peptides at positions 84-92 and 91-99 induced HLA-A24-restricted CTL activity against tumor cells in PBMCs of leukemia patients, but not in epithelial cancer patients or in healthy donors. In contrast, the modified peptides at position 2 from phenylalanine to tyrosine, which had more than 10 times higher binding affinities to HLA-A24 molecules, could induce HLA-A24-restricted CTL activity against tumor cells in PBMCs from leukemia patients, epithelial cancer patients, or healthy donors. PHA-activated normal T cells were resistant to lysis by the CTL line or by these peptide-induced CTLs. These results indicate that a cyclophilin B gene encodes antigenic epitopes recognized by CTLs at the tumor site, although T cells in peripheral blood (except for those from leukemia patients) are immunologically tolerant to the cyclophilin B. These peptides might be applicable for use in specific immunotherapy of leukemia patients or that of epithelial cancer patients.

Polymorphism of the 5'-flanking region of the tumor necrosis factor (TNF)-alpha gene and susceptibility to human T-cell lymphotropic virus type I (HTLV-I) uveitis.

The genetic background of human T-cell lymphotropic virus type I (HTLV-I) uveitis (HU) was investigated by studying the distribution of 5 polymorphisms of the 5'-flanking promoter/enhancer region of the tumor necrosis factor (TNF)-alpha gene in patients with HU, together with patients with adult T-cell leukemia (ATL), asymptomatic HTLV-I carriers, and healthy controls. The frequencies of the -1,031C allele (T-->C transition at position -1,031) and -863A allele (C-->A transition at position -863) in the HU patients, but neither in the ATL patients nor in the carriers, were significantly higher than those in the controls. The -1,031C and -863A alleles, in the absence of the HLA B61 or the DRB1*0901 allele which is in linkage disequilibrium with these alleles, were associated with increased susceptibility to HU. These results suggest that the -1,031C and -863A alleles might be genetic risk factors for HU.