Biomedical subjects
M Kuroki
Publications and source records attributed to M Kuroki.
Carcinoma-associated antigens MK-1 and CEA in urological cancers.
BACKGROUND: The MK-1 antigen, recognized by monoclonal antibody FU-MK-1, is widely associated with human carcinomas. However, the expression and distribution of MK-1 in urological cancers is not well known. MATERIALS AND METHODS: We examined the expression of MK-1 in 10 urological tumor cell lines using flow cytometry and reverse transcription-polymerase chain reaction (RT-PCR) and in 15 cancer tissue specimens by immunohistochemical staining, and then compared it with that of carcinoembryonic antigen (CEA). RESULTS: When analyzed by flow cytometry, MK-1 was positive in 2 out of 3 bladder, 3 out of 3 prostate and one out of 4 renal tumor cell lines, whereas CEA was negative in all the 10 tumor cell lines. RT-PCR confirmed the presence of MK-1 mRNA in all the six MK-1-positive tumor cell lines. An immunohistochemical study demonstrated that MK-1 was positive in 2 out of 5 bladder, 2 out of 5 prostate and one out of 5 renal cancer tissues. Again, however, CEA was negative in all the 15 urological cancer tissues tested. CONCLUSION: These findings suggest that MK-1 seems to be a useful biological marker for malignant urological tumors, especially in cases of bladder and prostate cancer.
Specific targeting strategies of cancer gene therapy using a single-chain variable fragment (scFv) with a high affinity for CEA.
Two specific targeting strategies of cancer gene therapy using carcinoembryonic antigen (CEA) as a target are briefly reviewed here. One method is the specific targeting of suicide genes to CEA-expressing tumor cells by a retrovector displaying anti-CEA single-chain variable fragment (scFv). We reconstructed a recombinant retroviral vector that displays both anti-CEA scFv-expressing chimeric and normal envelope proteins and carries the inducible nitric oxide synthase (iNOS) gene. This recombinant retrovirus specifically bound, infected and killed only CEA-expressing tumor cells, indicating the cell specific retroviral vector delivery of the iNOS gene. Another novel method is the specific redirecting of cytotoxic T-cells to CEA-expressing tumor cells through chimeric receptors. We also reconstructed a chimeric receptor gene which encoded an anti-CEA scFv antibody and the zeta-chain of human TCR/CD3 complex and expressed it on T-cell surface. When incubated with CEA-expressing tumor cells in vitro, the transduced T-cells tended to make a rosette-like formation around the tumor cells, suggesting the cell specific targeting of T-cells.
Targeting of LAK activity to CEA-expressing tumor cells with an anti-CEA scFv/IL-2 fusion protein.
BACKGROUND: Fusion of tumor-specific monoclonal antibody (MAb) and cytokines has proved to be an efficient way to target cytokines to tumor cells and hence focuses the killing activity of effector cells to the target cells. We previously produced a high affinity MAb, F11-39, against carcinoembryonic antigen (CEA), which is often overexpressed on the surface of various tumor cells. MATERIALS AND METHODS: To target the cytotoxicity of effector cells to CEA-expressing tumor cells, we employed recombinant DNA techniques to fuse recombinant human interleukin-2 (rhIL-2) to a single chain variable fragment (scFv) antibody derived from F11-39. The resulting fusion protein, designated F39scFv/IL-2, was expressed in the Sp2/0-Ag14 mouse hybridoma cells, purified by CEA-affinity chromatography and characterized for the CEA-binding specificity and the IL-2 biological activity. RESULTS: F39scFv/IL-2 protein effectively targeted rhIL-2 onto the surface of CEA-expressing tumor cells and consequently introduced a specific cytotoxicity of lymphokine-activated killer cells to the tumor cells. CONCLUSIONS: This approach may be used for in vivo administration to localize IL-2 to tumor tissues, maximizing the immune response to CEA-expressing tumors while keeping systemic side effects to a minimum.
[Topical application of antibiotics-aminoacids for healing in dorsal rat skin. Scanning electron microscope study].
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Initial histopathology of the autovein graft in late occlusion after arterial reconstruction.
Five patients developed local stenosis of autologous vein grafts implanted for femoro-popliteal arterial occlusive lesions. This stenosis occurred in the vein grafts 2 months to 5 years after the initial operation. Histopathologic study revealed that the stenotic segments had a thickened intima with a prominent proliferation of smooth muscle cells associated with fibrous extracellular matrix. There were no findings showing deposition of mural thrombi, as has heretofore been reported. The intimal thickening due to excessive fibromuscular proliferative response of the vein graft may possibly play an important role in the development of late graft occlusion.
Occluded persistent sciatic artery.
The case of a patient with ischemic symptoms due to occlusion of a persistent left sciatic artery is presented. Femoral arteriography revealed a tapering hypoplastic superficial femoral artery terminating as small branches in the distal thigh and a hyperplastic profunda femoris artery. Also, translumbar aortography demonstrated an abrupt occlusion of a hyperplastic left hypogastric artery just distal to the inferior gluteal artery. The popliteal artery was found to be patent and perfused by collaterals. A bypass graft using a composite E-PTFE prosthesis and an autogenous vein graft was successfully performed.
A mouse/human-chimeric bispecific antibody reactive with human carcinoembryonic antigen-expressing cells and human T-lymphocytes.
A mouse/human-chimeric bispecific antibody, designated CBA-CEACD3, with dual specificities for carcinoembryonic antigen (CEA) and CD3, was generated by chemical cross-linking of a chimeric antibody specific for CEA to another chimeric antibody against CD3. Flow cytometric analysis showed that CBA-CEACD3 can bind specifically to cells expressing CEA and to normal human peripheral blood mononuclear cells (HPBMCs) bearing CD3, respectively. Furthermore, a cell to cell adhesion analysis by a colorimetric assay using the dye 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) demonstrated that CBA-CEACD3 is able to bind CEA-producing cells to CD3-expressing cells, suggesting that both arms of CBA-CEACD3 are simultaneously working and can retarget T-cells to the tumor. In an additional colorimetric assay using MTT, this antibody was shown to effectively mediate CEA-expressing tumor cell killing by freshly isolated HPBMCs. Together these results demonstrate that this chimeric bispecific antibody may serve as a potentially useful immunotherapeutic reagent for human CEA-producing cancers.
Tumor growth suppression by a mouse/human chimeric anti-CEA antibody and lymphokine-activated killer cells in vitro and in SCID mouse xenograft model.
The mouse/human chimeric antibody Ch F11-39, recently generated by ourselves, shows the same high specificity and affinity for carcinoembryonic antigen (CEA) as those of its parental mouse monoclonal antibody. Ch F11-39 is capable of mediating antibody-dependent cell-mediated cytotoxicity (ADCC) with human peripheral blood lymphocytes (PBL). Interleukin-2 (IL-2) modulates the function of immunocytes, in particular inducing lymphokine-activated killer (LAK) cells and enhancing ADCC. In the present study, we therefore tested the combination immunotherapy of Ch F11-39 with LAK cells in vitro and in severe combined immunodeficiency (SCID) mice bearing human CEA-producing tumors. In vitro experiments using human gastric tumor cell lines, Ch F11-39 effectively mediated ADCC against CEA-positive MKN-45 cells, but not against CEA-negative cells. The specificity of ADCC for Ch F11-39 was demonstrated by experiments with irrelevant target cells or irrelevant antibody. ADCC activity of PBL with Ch F11-39 was enhanced by double after preincubation with IL-2 at 10 U/ml. The concentration of Ch F11-39 required for 50% maximal cell killing was about 0.25 microgram/ml at 10 U/ml of IL-2. Increasing ADCC was triggered by IL-2 earlier (1 day) than the generation of LAK cells (3 days). Control human IgG blocked the ADCC, suggesting that the enhancement of ADCC by IL-2 may be caused by activation of effector cells expressing Fc receptors. In vivo anti-tumor activity of combined immunotherapy was estimated using SCID mice inoculated s.c. with 1 x 10(7) MKN-45 cells. The i.v. administration of LAK cells and i.p. administration of Ch F11-39 and IL-2 produced a marked growth inhibition of MKN-45 tumors in SCID mice (about 50% reduction in tumor size as compared to the control untreated group, measured 15 days after treatment). In summary, the enhanced antitumor activity of Ch F11-39 with LAK cells suggests that it might be a useful immunotherapeutic reagent for CEA-expressing tumors.