Title:A Novel Recombinant Modified Vaccinia Ankara Virus expressing
Interleukin-13 Receptor α2 Antigen for Potential Cancer
Immunotherapy
Volume: 24
Issue: 6
关键词:
MVA，修饰的安卡拉牛痘，IL-13Rα2，白细胞介素-13受体α2，绿色荧光蛋白，IL13-PE，白细胞介素-13融合到假单胞菌外毒素突变形式。
摘要:
Background: Genetically altered recombinant poxviruses hold great
therapeutic promise in animal models of cancer. Poxviruses can induce effective cellmediated
immune responses against tumor-associated antigens. Preventive and
therapeutic vaccination with a DNA vaccine expressing IL-13Rα2 can mediate partial
regression of established tumors in vivo, indicating that host immune responses against
IL-13Rα2 need further augmentation.
Objective: The aim of the study is developing a recombinant modified vaccinia Ankara
(MVA) expressing IL-13Rα2 (rMVA-IL13Rα2) virus and study in vitro infectivity and
efficacy against IL-13Rα2 positive cell lines.
Methods: We constructed a recombinant MVA expressing IL-13Rα2 and a green
fluorescent protein (GFP) reporter gene. Purified virus titration by infection of target cells
and immunostaining using anti-vaccinia and anti-IL-13Rα2 antibodies was used to
confirm the identity and purity of the rMVA-IL13Rα2.
Results: Western Blot analysis confirmed the presence of IL-13Rα2 protein (~52 kDa).
Flow cytometric analysis of IL-13Rα2 negative T98G glioma cells when infected with
rMVA-IL13Rα2 virus demonstrated cell-surface expression of IL-13Rα2, indicating the
infectivity of the recombinant virus. Incubation of T98G-IL13Rα2 cells with varying
concentrations (0.1-100 ng/ml) of interleukin-13 fused to truncated Pseudomonas
exotoxin (IL13-PE) resulted in depletion of GFP+ fluorescence in T98G-IL13Rα2 cells.
IL13-PE (10-1000 ng/ml) at higher concentrations also inhibited the protein synthesis in
T98G-IL13Rα2 cells compared to cells infected with the control pLW44-MVA virus. IL13-
PE treatment of rMVA-IL13Rα2 infected chicken embryonic fibroblast and DF-1 cell line
reduced virus titer compared to untreated cells.
Conclusion: rMVA-IL13Rα2 virus can successfully infect mammalian cells to express
IL-13Rα2 in a biologically active form on the surface of infected cells. To evaluate the
efficacy of rMVA-IL13Rα2, immunization studies are planned in murine tumor models.
