1999 Fiscal Year Final Research Report Summary
Molecular targeting against growth factor receptors by a novel drug compord of human fusion proteins
| Project/Area Number |
09470258
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
General surgery
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| Research Institution | Keio University |
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Principal Investigator |
UEDA Masakazu Keio Univ., Surgery, Assistant Professor, 医学部, 講師 (50142419)
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| Co-Investigator(Kenkyū-buntansha) |
KAMEYAMA Noriaki Keio Univ., Surgery, Assistant, 医学部, 助手 (50276268)
WATANABE Yasuo Keio Univ., Surgery, Assistant, 医学部, 助手 (20265846)
OZAWA Soji Keio Univ., Surgery, Assistant Professor, 医学部, 講師 (10169287)
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| Project Period (FY) |
1997 – 1999
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| Keywords | Missils therapy / growth factor / growth factor receplor / IL-2 / EGF / IL-2 receptor / fusion protein |
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| Research Abstract |
To develop a new Missile therapy targeted at growth factor receptor, fusion proteins with a human Rnase and monoclonal antibody against growth factor receptors or growth factor. A hybrid human protein was engineered by fusing the genes encoding human pancreatic Rnase 1 and human IL-2, was isolated and refolded from E.coli inclusion bodies, and was purified to homogeneity. The hpRNase1-hIL-2 inhibited protein synthesis in HTLV-1 infected, malignant T lymphocytes, hyperproducing high affinity IL-2 receptors, with an IC50 of 2x10-8M, whereas no inhibition was detectable for receptor deficient control cells or hpRNase 1 alone had an IL50 of almost 10-3M.. A molar excess of hIL-2 locked the protein synthesis inhibition dose dependently. In a human mixed lymphocyte culture, hpRNase 1-hIL-2 inhibited the proliferation of responder cells with potency comparable to that of cyclosporine, while minimal doses of FK506 importantly improved its potency. Concentrations comparable to the IC50 in vitro could be safely achieved in animals, and could be escalated without any toxic side effects. The murine monoclonal antibody (528) against the human epidermal growth factor receptor (EGFR) was conjugated with mammalian pancreatic Rnase via SPDP and 2-IT. The conjugate showed dose-dependent cytotoxicity against EGFR-producing squamous cancer cells and no detectable cytotoxicity against EGFR-deficient small cell lung cancer cells. The cytotoxicity of the conjugate was positively correlated with the EGFR numbers of each cell line. The addition of excess 528 antibody to the medium rotected A431 cells from the conjugate cytotoxicity. This immunoconjugate might be useful for targeted treatment of squamous cell carcinomas hyperexpressing EGFR.
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