Construction of chimeric IL-21 recombinant oncolytic influenza virus and mechanisms for its antitumor efficacy against hepatocellular carcinoma
Objective To rescue a recombinant oncolytic influenza virus chimeric with IL-21 and evaluate its inhibitory effects and safety against hepatocellular carcinoma (HCC) both in vitro and in vivo, and to explore the mechanism by which this virus enhances antitumor effects when combined with anti-program...
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Editorial Office of Journal of Army Medical University
2024-12-01
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| Series: | 陆军军医大学学报 |
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| Online Access: | https://aammt.tmmu.edu.cn/html/202407051.htm |
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| author | CAO Rui ZENG Guineng TIAN Yuying TIAN Yuying |
| author_facet | CAO Rui ZENG Guineng TIAN Yuying TIAN Yuying |
| author_sort | CAO Rui |
| collection | DOAJ |
| description | Objective To rescue a recombinant oncolytic influenza virus chimeric with IL-21 and evaluate its inhibitory effects and safety against hepatocellular carcinoma (HCC) both in vitro and in vivo, and to explore the mechanism by which this virus enhances antitumor effects when combined with anti-programmed cell death protein 1(anti-PD-1) antibody. Methods The IL-21 gene fragment was inserted into the nonstructural protein (NS) sequence of the influenza virus PR8 using reverse genetics (RG) technology to rescue the recombinant oncolytic influenza virus rOV-IL-21-NS.The virus titer and virulence were determined using the 50% tissue culture infectious dose (TCID50) and hemagglutination assays.The successful insertion of the exogenous gene into the NS sequence was verified using RT-qPCR, gel electrophoresis, and sequencing analysis.Viral morphology and size were observed with transmission electron microscopy.The impact of the virus on the viability of hepatocellular carcinoma cells was assessed with CCK-8 assay.A subcutaneous tumor model of HCC was established in 45 female C57BL/6 mice (8 weeks old, weighing 16~20 g), and then the mice were randomly assigned into PBS, PR8, anti-PD-1, rOV-IL-21-NS, and the rOV-IL-21-NS+anti-PD-1 treatment groups, with 9 mice in each group, to evaluate the anti-tumor effects of monotherapy versus combination therapy.Flow cytometry was conducted to assess the regulatory effects of monotherapy and combination therapy on the tumor immune microenvironment. Results RG technology successfully rescued the recombinant oncolytic influenza virus rOV-IL-21-NS.Sequencing confirmed the successful insertion of IL-21 into the target sequence, and the obtained virus could be stably propagated, with its sixth passage reaching a hemagglutination titer of 211, and a viral titer of 6 Log10(TCID50/mL).Oncolytic virus rOV-IL-21-NS, at a multiplicity of infection (MOI) of 3, selectively reduced the viability of HCC cells without significantly affecting normal liver cells.Compared to the control group, the combination of rOV-IL-21-NS and anti-PD-1 antibodies significantly inhibited tumor growth (P < 0.001) and increased the proportions of CD4+ T and CD8+ T cells in the spleen tissue of the mouse model of subcutaneous HCC tumor (P < 0.001). Conclusion The recombinant oncolytic influenza virus rOV-IL-21-NS chimeric with IL-21 can effectively and safely exert targeted killing to HCC cells, enhance T cell activation by synergizing with anti-PD-1 antibodies, and improve the immune microenvironment.
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| format | Article |
| id | doaj-art-0fbabbbaae6e45d99cf2cd5817ba69b1 |
| institution | Kabale University |
| issn | 2097-0927 |
| language | zho |
| publishDate | 2024-12-01 |
| publisher | Editorial Office of Journal of Army Medical University |
| record_format | Article |
| series | 陆军军医大学学报 |
| spelling | doaj-art-0fbabbbaae6e45d99cf2cd5817ba69b12024-12-29T00:27:03ZzhoEditorial Office of Journal of Army Medical University陆军军医大学学报2097-09272024-12-0146242736274410.16016/j.2097-0927.202407051Construction of chimeric IL-21 recombinant oncolytic influenza virus and mechanisms for its antitumor efficacy against hepatocellular carcinomaCAO Rui0 ZENG Guineng1TIAN Yuying2TIAN Yuying3School of Basic Medical Sciences, Inner Mongolia Medical University, Hohhot, Inner Mongolia Autonomous Region, 010110Faculty of Hepato-Biliary-Pancreatic Surgery, First Medical Center, Chinese PLA General Hospital, Beijing, 100853, ChinaSchool of Basic Medical Sciences, Inner Mongolia Medical University, Hohhot, Inner Mongolia Autonomous Region, 010110 Faculty of Hepato-Biliary-Pancreatic Surgery, First Medical Center, Chinese PLA General Hospital, Beijing, 100853, ChinaObjective To rescue a recombinant oncolytic influenza virus chimeric with IL-21 and evaluate its inhibitory effects and safety against hepatocellular carcinoma (HCC) both in vitro and in vivo, and to explore the mechanism by which this virus enhances antitumor effects when combined with anti-programmed cell death protein 1(anti-PD-1) antibody. Methods The IL-21 gene fragment was inserted into the nonstructural protein (NS) sequence of the influenza virus PR8 using reverse genetics (RG) technology to rescue the recombinant oncolytic influenza virus rOV-IL-21-NS.The virus titer and virulence were determined using the 50% tissue culture infectious dose (TCID50) and hemagglutination assays.The successful insertion of the exogenous gene into the NS sequence was verified using RT-qPCR, gel electrophoresis, and sequencing analysis.Viral morphology and size were observed with transmission electron microscopy.The impact of the virus on the viability of hepatocellular carcinoma cells was assessed with CCK-8 assay.A subcutaneous tumor model of HCC was established in 45 female C57BL/6 mice (8 weeks old, weighing 16~20 g), and then the mice were randomly assigned into PBS, PR8, anti-PD-1, rOV-IL-21-NS, and the rOV-IL-21-NS+anti-PD-1 treatment groups, with 9 mice in each group, to evaluate the anti-tumor effects of monotherapy versus combination therapy.Flow cytometry was conducted to assess the regulatory effects of monotherapy and combination therapy on the tumor immune microenvironment. Results RG technology successfully rescued the recombinant oncolytic influenza virus rOV-IL-21-NS.Sequencing confirmed the successful insertion of IL-21 into the target sequence, and the obtained virus could be stably propagated, with its sixth passage reaching a hemagglutination titer of 211, and a viral titer of 6 Log10(TCID50/mL).Oncolytic virus rOV-IL-21-NS, at a multiplicity of infection (MOI) of 3, selectively reduced the viability of HCC cells without significantly affecting normal liver cells.Compared to the control group, the combination of rOV-IL-21-NS and anti-PD-1 antibodies significantly inhibited tumor growth (P < 0.001) and increased the proportions of CD4+ T and CD8+ T cells in the spleen tissue of the mouse model of subcutaneous HCC tumor (P < 0.001). Conclusion The recombinant oncolytic influenza virus rOV-IL-21-NS chimeric with IL-21 can effectively and safely exert targeted killing to HCC cells, enhance T cell activation by synergizing with anti-PD-1 antibodies, and improve the immune microenvironment. https://aammt.tmmu.edu.cn/html/202407051.htmoncolytic virusreverse genetic technologycombination therapyil-21anti-programmed cell death protein 1 antibodyhepatocellular carcinoma |
| spellingShingle | CAO Rui ZENG Guineng TIAN Yuying TIAN Yuying Construction of chimeric IL-21 recombinant oncolytic influenza virus and mechanisms for its antitumor efficacy against hepatocellular carcinoma 陆军军医大学学报 oncolytic virus reverse genetic technology combination therapy il-21 anti-programmed cell death protein 1 antibody hepatocellular carcinoma |
| title | Construction of chimeric IL-21 recombinant oncolytic influenza virus and mechanisms for its antitumor efficacy against hepatocellular carcinoma |
| title_full | Construction of chimeric IL-21 recombinant oncolytic influenza virus and mechanisms for its antitumor efficacy against hepatocellular carcinoma |
| title_fullStr | Construction of chimeric IL-21 recombinant oncolytic influenza virus and mechanisms for its antitumor efficacy against hepatocellular carcinoma |
| title_full_unstemmed | Construction of chimeric IL-21 recombinant oncolytic influenza virus and mechanisms for its antitumor efficacy against hepatocellular carcinoma |
| title_short | Construction of chimeric IL-21 recombinant oncolytic influenza virus and mechanisms for its antitumor efficacy against hepatocellular carcinoma |
| title_sort | construction of chimeric il 21 recombinant oncolytic influenza virus and mechanisms for its antitumor efficacy against hepatocellular carcinoma |
| topic | oncolytic virus reverse genetic technology combination therapy il-21 anti-programmed cell death protein 1 antibody hepatocellular carcinoma |
| url | https://aammt.tmmu.edu.cn/html/202407051.htm |
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