[Carcinoembryonic antigen (CEA)].
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Biomarkers, Tumor↗
Biomedical subjects
Tetsushi Kinugasa
Publications and source records attributed to Tetsushi Kinugasa.
Strategies to endow cytotoxic T lymphocytes or natural killer cells with antibody activity against carcinoembryonic antigen.
Cytotoxic T lymphocytes (CTLs) and natural killer (NK) cells are main effecter cells in cellular immunity against tumor cells. T-cell immunotherapy is based on the assumption that tumor(-associated) antigen (TA) peptides are correctly presented by HLA class I molecules on target tumor cells, and NK cell immunotherapy is based on the hypothesis that cell surface TAs or ligands for NK receptors are widely expressed in tumor cells. However, human tumor cells often lose HLA class I molecules, and target cell ligands for NK receptors are not always expressed in human tumor cells. These altered HLA class I phenotypes and non-ubiquitous expression of NK receptor ligands constitute the major tumor escape mechanism facing tumor-specific CTL and/or NK cell mediated responses. These facts also indicate that it is not easy to eliminate the target tumors only by activating tumor-specific CTLs or NK cells with cancer vaccine treatments. On the other hand, it is easily confirmed by immunohistochemistry whether or not antibody-recognized TAs exist on the cell surface of target tumor cells. Therefore, endowing CTLs or NK cells with antigen-binding specificity of anti-TA antibody is a promising approach for re-targeting the activities of these effector cells to tumor cells in an HLA-independent manner. This review summarizes the following four new strategies for re-targeting CTLs or NK cells to carcinoembryonic-antigen-expressing tumor cells: (1) bispecific antibody technology; (2) antibody-cytokine fusion protein technology; (3) chimeric immune receptor technology, and (4) antibody-HLA/peptide complex technology.
Binding Sites, Antibody↗
Cloning and sequencing of variable region cDNAs of a novel human monoclonal antibody to carcinoembryonic antigen, and generation of a single chain variable fragmented antibody.
BACKGROUND: Taking into consideration immunogenicity in humans, human antibodies and their derivatives have potential advantages for cancer immunotherapy and/or gene therapy over those from different species such as mouse. Recently, we generated 22 human monoclonal antibodies (HmAbs) specific for human carcinoembryonic antigen (CEA) using KM mouse, which carries human antibody genes. In the present study, we tried to clone the variable (V) region genes of the C2-45 HmAb, with the highest affinity for CEA and to generate a human single-chain variable fragmented (scFv) antibody. MATERIALS AND METHODS: Using RT-PCR methods, we cloned and sequenced cDNAs encoding V regions of C2-45, and constructed an scFv gene including a signal sequence for secreting into the periplasmic space of E. coli. RESULTS: Soluble scFv antibody, designated 45KHscFv, was secreted in the periplasmic space and purified by Ni2+ affinity column from the supernatant of E. coli after osmotic shock. When analyzed by SDS-PAGE, the 45KHscFv antibody showed a band corresponding to a calculated molecular weight of 30 kDa. The 45KHscFv antibody also retained a high reactivity with CEA in ELISA using immobilized CEA. CONCLUSION: These results indicate that the nonimmunogenic 45KHscFv antibody could be advantageously used for fusing with other human functional proteins such as cytokines.
Amino Acid Sequence↗
Tumor-targeting of viral vectors for cancer gene therapy by using antibodies or their genes against tumor-associated antigens.
Gene therapy is a promising modality for the treatment of malignant tumors for which conventional therapies are often inadequate. Various therapeutic genes have shown promise for tumor cell killing. However, successful gene therapy depends on the development of efficient and targeted gene transfer vectors. The aim of this review is to summarize the tumor-targeting viral vectors for cancer gene therapy by utilizing antibodies or genes against tumor-associated antigens.
Adenoviridae↗
Re-targeting of cytotoxic T lymphocytes and/or natural killer cells to CEA-expressing tumor cells with anti-CEA antibody activity.
Cellular immunity, in which cytotoxic T lymphocytes (CTLs) and natural killer (NK) cells are main effector cells, plays an important role in the antitumor defense mechanism. T cell immunotherapy is based on the assumption that tumor antigen (TA) peptides are correctly presented by HLA class I molecules on target tumor cells, while NK cell immunotherapy is based on the hypothesis that cell surface TAs or ligands for NK receptors are widely expressed in tumor cells. However, human tumor cells are well known to often lose HLA class 1 molecules, and target cell ligands for NK receptors are not always expressed in human tumor cells. This altered HLA class 1 expression and non-ubiquitous distribution of NK receptor ligands constitute the major tumor escape mechanism facing tumor-specific CTL and/or NK cell-mediated responses. These facts also indicate that it is not easy to eliminate the target tumors only by activating tumor-specific CTLs or NK cells. On the other hand, although the protective role of humoral immunity in cancer seems not to be imperative, it is easily confirmed by immunostaining whether or not antibody-recognized TAs such as carcinoembryonic antigen (CEA) exist on the cell surface of target tumor cells. Therefore, endowing CTLs or NK cells with antigen-binding specificity of anti-TA antibody is promising for re-targeting the activities of these effector cells to tumor cells in an HLA-independent manner. This mini-review provides a brief overview of the following four technologies for re-targeting CTLs or NK cells to CEA-expressing tumor cells with anti-CEA antibody activity: i) bispecific antibody technology, ii) antibody-cytokine fusion protein technology, iii) chimeric immune receptor technology, and iv) antibody-HLA/peptide complex technology.
Animals↗
Preparation of human IgG and IgM monoclonal antibodies for MK-1/Ep-CAM by using human immunoglobulin gene-transferred mouse and gene cloning of their variable regions.
For antibody-based therapy of cancer, monoclonal antibodies (mAbs) of human origin are superior to mouse, mouse/human chimeric or humanized mAbs, because of their minimum immunogenicity to humans and their efficient collaboration with human effector cells. In the present study, human mAbs were prepared against a pancarcinoma antigen, MK-1 (Ep-CAM), using a genetically-engineered mouse (KM mouse) that contains the human immunoglobulin genes. Spleen cells from KM mice, immunized with recombinant MK-1, were fused with P3-U1 mouse myeloma cells. Of 44 anti-MK-1 clones analyzed, two were of IgG4 and the others of IgM clones. Although the two IgG4 clones were suggested to recognize the same antigenic determinant or two closely located determinants, their VK regions were encoded by different light-chain genes while their VH sequences were identical. The two IgG4 and one of the IgM clones tested revealed antibody-dependent cell-mediated cytotoxicity and complement-dependent cytotoxicity, respectively, against MK-1-expressing cells in vitro, suggesting that these fully human mAbs produced against MK-1 and their V-region genes, which are applicable for the preparation of engineered antibody fragments that may be useful for antibody-based therapy of cancer.
Amino Acid Sequence↗
IgG isotype conversion of a novel human anti-carcinoembryonic antigen antibody to increase its biological activity.
BACKGROUND: The IgG isotype of antibodies is very important for their biological functions such as complement-dependent cytotoxicity (CDC) and antibody-dependent cellular cytotoxicity (ADCC). To increase the biological activity of a novel human monoclonal antibody (C2-45) against carcinoembryonic antigen (CEA), we tried to genetically convert its isotype from IgG4 to IgG1. MATERIALS AND METHODS: VH and VL genes were cloned from the parental antibody C2-45 (IgG4) and inserted into the pAc-kappa-CH3 expression vector which contained the constant region gene of human IgG1. The recombinant gene was transfected into Sf9 insect cells to produce recombinant protein. The resulting recombinant protein, designated C2-45 (cIgG1), in the culture medium was purified by affinity chromatography and characterized for its CEA binding activity and biological activity. RESULTS: The converted C2-45 (cIgG1) retained the original antigen-binding activity and showed significantly higher CDC and ADCC activities against CEA-expressing tumor cells than did the original C2-45 (IgG4). CONCLUSION: C2-45 (cIgG1) may be useful for antibody-based immunotherapy of human CEA-expressing tumors.
Animals↗
Possible applications of antibodies or their genes in cancer therapy.
In this review article the possible applications of anti-tumor-associated antigen (TAA) antibodies in the therapy of cancer have been summarized. First, recombinant monoclonal antibodies (MAbs) are increasingly being used as therapeutic agents, especially in combination with anti-cancer drugs. Second, conjugation of antibody therapy with toxins or radioisotopes offers more therapeutic approaches. Third, development of cytotoxic T-lymphocyte (CTL) or natural killer (NK)-cell populations with anti-TAA antibody activity may be important for the success of cancer immunotherapy because the downregulated HLA class I molecules and the non-ubiquitous expression of NK receptor ligands in tumor tissues constitute the major tumor escape mechanism facing tumor-specific CTL- and/or NK-cell-mediated responses. Finally, in cancer gene therapy, the strategies to target viral vectors carrying therapeutic genes to tumor tissues by modifying the tropisms with MAbs or their genes against TAAs are also very promising.
Animals↗
A fully human chimeric immune receptor for retargeting T-cells to CEA-expressing tumor cells.
BACKGROUND: Recombinant chimeric immune receptors (CIRs) with anti-CEA specificity can retarget grafted T-cells to CEA-expressing tumors in an HLA-independent manner. To reduce the immunogenicity of conventional CIR in humans, an attempt was made to generate a CIR encoded by all human genes. MATERIALS AND METHODS: A single-chain variable fragmented (scFv) antibody gene was prepared from variable region genes of the C2-45 human mAb clone specific for CEA. The scFv gene was connected to a gene construct comprised of the cDNAs for the human CD8a hinge region, the human CD28 transmembrane and cytoplasmic domains, and the human CD3zeta intracellular domain. The resulting human CIR gene, designated L45scFv-CIR, was inserted into the pcDNA3.1 expression vector and transfected into human primary T-cells. RESULTS: Flow cytometric analysis using allophycocyanin-labeled CEA demonstrated the expression of the L45scFv-CIR protein on the T-cells and its specific antigen binding activity. CONCLUSION: This L45scFv-CIR gene, consisting of four human genes, may be a useful tool for eradication of CEA-expressing but HLA-downregulated tumor cells.
CD28 Antigens↗