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Modifying Tumor Cell Surface with Immune Molecules and Preliminary Study on Its Anti-tumor Function

Author: ChengShaoHui
Tutor: ShenBeiFen
School: PLA Military Academy of Medical Sciences
Course: Molecular Immunology
Keywords: Immune molecules Eukaryotic expression vector Anchoring protein Inhibitory effect Exogenous genes Fusion gene Tumor cells Membrane surface Modification of a Melanoma
CLC: R392
Type: PhD thesis
Year: 2003
Downloads: 71
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Abstract

How to break the tumor immune evasion mechanisms, and to re-awaken the identification of the immune system against tumor cells and killing capability, is the key to cancer immunotherapy. The purpose of this paper \The main subject of doing the work of the following aspects: 1 GPI anchor modification of eukaryotic expression vector and vector reconstructive GPI anchor eukaryotic expression vector pCI-GPI in our laboratory has been constructed, based on the carrier reconstructive work. Connected to the carrier source of decay accelerating factor (DAF), the GPI-modified signal sequence and human IgGFc fusion gene segment, exogenous gene via the GPI signal peptide modifications may be such that the nascent protein and GPI (Glycosylphosphatidy-1-inositol, glycosylation phosphatidyl inositol) structure, including the endoplasmic reticulum connected to form the GPI-anchored proteins, and integrated into the surface of the cell membrane and its lipid tail. Human IgG Fc fragment and GPI modification signal peptide fusion gene fragment directed inserted in pCI-dhfr ~ eukaryotic expression vector EcoRI and XbaI restriction sites between the name of the carrier the pCI-GPI carrier. On this basis a certain carrier reconstructive work, EcoRI / NotI double digested Fc-GPI fusion gene fragment, and is connected on the pCI-neo eukaryotic expression vector, this vector has a neomycin resistance gene , on the carrier at the same time connected to the secretion signal peptide sequence of the V-J2-C region of the mouse Igκ chain, in order to facilitate the secretion expression of the exogenous gene. 2 mGM-CSF anchor modification and preliminary study of B16 mouse melanoma inhibitory effect in order to prove that the the immune molecules After anchoring modified, whether retain biological activity and immune molecules anchor expression if there is a better inhibitory effect, we anchored modified GM-CSF in B16 mouse melanoma cell surface, through animal studies observed inhibitory effect of GM-CSF anchored modified to lay the foundation for a series of studies in other immune molecules anchored modified. By RT-PCR from mouse spleen lymphocytes fishing take mGM-CSF length gene amplified fragment connected to pGMT-easy vector, sequencing, insert PcDNA3.1 eukaryotic expression vector, named PcDNA3.1 -mGM. In addition mGM-CSF gene was amplified NheI / EcoRI digested connected to the eukaryotic expression vector pCI-GPI enzyme digestion confirmed the connection in the upstream of the Fc-GPI fusion gene, named pCI-GPI-with mGM. Liposomal transfection of the two plasmids were transfected B16 mouse melanoma

Modifying Tumor Cell Surface with Immune Molecules and Preliminary Study on Its Anti-tumor Function

Abstract

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