Yasuhisa Imai

Sequence analysis of the MAGE gene family encoding human tumor-rejection antigens

The MAGE multigene family, which includes the MAGE-1 and -3 genes that encode tumor-rejection antigens on HLA-A1 recognized by cytotoxic T-lymphocytes (CTL), is preferentially expressed at the mRNA level on human malignant cells, but not on normal cells. However, little is known about the MAGE-4, -41 and -6 genes. In this study, we have amplified 1040 bp (MAGE-1), 1061 bp (MAGE-3 and -6) and 1064 bp (MAGE-4 and -41) cDNA fragments, including the entire coding sequences (927-951 bp), using the reverse transcription-polymerase chain reaction (RT-PCR) method followed by nucleotide (nt) sequencing. One member had greater than 80 or 66% homology with the other members at the nt or deduced amino acid (aa) levels, respectively. Higher homology was found between MAGE-3 and -6 (98% at the nt level) and also between MAGE-4 and -41 (98%). The results of this investigation demonstrated high homology, as well as the clear differences between the members of the MAGE family at the coding sequence level.

Expression of the SART-1 tumor rejection antigen in breast cancer

We investigated in breast cancers the expression of the SART-1 gene encoding tumor rejection antigens. SART-1 mRNA was expressed in all of the samples tested. The SART-1(800) antigen was detectable in 20 of 50 (40%) breast cancer tissues and all breast cancer cell lines tested, but not in normal breast tissues. The SART-1(800)+ breast cancer cells transfected with HLA-A2601 or HLA-A2402 cDNA were recognized by the HLA-A26-restricted and SART-1-specific cytotoxic T lymphocytes (CTLs) or the HLA-A24-restricted and SART-1-specific CTLs, respectively. Among the 20 SART-1(800)+ tumors, 9 or 8 tumors expressed estrogen receptor or progesterone receptor, respectively. Therefore, the patients with HLA-A26 or -A24 haplotype might be appropriate candidates for specific immunotherapy with the SART-1 peptides independently or in combination with hormone therapy.

DEVELOPMENT OF CANCER VACCINE BY TUMOR REJECTION ANTIGENS

Identification of the MAGE genes allowed us the molecular approach to identify genes encoding tumor rejection antigens expressed on human cancer cells. MAGE-1 proteins are normal tissue antigens compartmentalized in the particular testicular cells playing an important role in the early phase of the spermatogenesis. The MAGE-1, -2, -3, -4 and -6 genes are preferentially expressed in many different cancers at both the mRNA and protein levels. The MAGE genes, particularly MAGE-1, became positive in relatively advanced stages of cancers and recurrent cancers. Approximately one-third to half of human cancers except for myelo-monocytic leukemia expressed at least one of these MAGE genes. The MAGE gene products shall be appropriate target molecules for development of new cancer vaccine.

Higher susceptibility of erythropoietin-producing renal cell carcinomas to lysis by lymphokine-activated killer cells

Erythropoietin production by renal cell carcinoma (RCC) is reported to be a potential marker for interleukin-2/interferon-alpha-responding tumor. We have investigated whether erythropoietin of RCC cells is involved in the immune recognition by lymphokine-activated killer (LAK) cells. Cells from primary culture of RCC cells expressing erythropoietin-mRNA or producing erythropoietin were more susceptible to lysis by LAK cells than those not expressing or producing it, respectively. RCC cells transfected with erythropoietin-cDNA became more susceptible to lysis by LAK cells than their erythropoietin-negative parental cells. These results indicate higher susceptibility of erythropoietin-producing RCC cells to lysis by LAK cells, suggesting that erythropoietin of RCC cells is involved in the immune recognition by LAK cells.

Sequence Analysis of Genes Encoding Rodent Homologues of the Human Tumor‐rejection Antigen SART‐1

Human SART‐1 (hSART‐1) gene encodes a 125 kD protein with a leucine‐zipper motif expressed in the nucleus of all proliferating cells, and a 43 kD protein expressed in the cytosol of most epithelial cancers. In this study, two rodent genes (rSART‐1 and mSART‐1) homologous to hSART‐1 were cloned from cDNA libraries of murine brain and a rat tumor cell line, respectively. mSART‐1 and rSART‐1 were highly homologous to hSART‐1 with 86% and 84% identity at the nucleotide level, and 95% and 91% at the protein level, respectively. The leucine zipper domain and two basic amino acid portions that bind DNA, as well as peptide sequences recognized by human cyto‐toxic T lymphocytes (CTLs), were all conserved in these rodent genes. Nuclear protein homologous to the 125 kD hSART‐1800 protein, but not to the 43 kD cytosol SART‐1259 protein, was detectable with specific antibody in the nuclear fractions of rodent tumor cell lines, and normal rodent fetal liver and testis. These rodent genes should be a novel tool for studies on the biological roles of the SART‐1 gene, and also in the construction of animal models of specific immuno‐therapy using SART‐1 gene products.

Changes in the mononuclear cell subpopulations of rat cardiac transplant recipients administered FK506 for the treatment of ongoing rejection

The inhibitory effect on ongoing rejection and the changes that occurred in mononuclear cell subpopulations were compared between four groups of rats treated with FK506 or steroids. Group 1 was given no immunosuppressive drugs, group 2 was given FK506 from the day of grafting, group 3 was commenced on FK506 on the 4th day after grafting, and group 4 was commenced on methylprednisolone (MP) on the 4th day after grafting. The garft survival times in groups 2 and 3 were significantly longer than those in groups 1 and 4, and there were fewer CD3+ and CD4+ T lymphocytes in the peripheral blood in the groups treated with immunosuppressive drugs than in group 1. In group 4, the levels in both the peripheral blood and thymus were significantly lower than those in the groups treated with FK506 despite the fact that graft rejection occurred soon after the discontinuation of steroid administration. Moreover, the levels of interleukin-2 receptors and macrophages in groups 2, 3, and 4 were significantly lower than that in group 1 postoperatively; however, the number of macrophages in groups 2 and 3 was significantly lower than that in group 4 on the 10th day after transplantation. The findings of this study demonstrated that FK506, even if administered after rejection has begun, might inhibit the subsequent extensive allograft rejection more specifically and effectively than steroids, and that the measurement of a marker for macrophages in the peripheral blood could be useful for the detection of rejection following allograft transplantation in rats.