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Therapeutic Strategy for the Prevention of Pseudorabies Virus Infection in C57BL/6 Mice by 3D8 scFv with Intrinsic Nuclease Activity.

Lee G, Cho S, Hoang PM, Kim D, Lee Y, Kil EJ, Byun SJ, Lee TK, Kim DH, Kim S, Lee S - Mol. Cells (2015)

Bottom Line: The observed therapeutic effect of 3D8 scFv against PRV was also supported by results from quantitative reverse transcription polymerase chain reaction, southern hybridization, and immunohistochemical assays.Intraperitoneal injection of 5 and 10 μg 3D8 scFv resulted in no detectable toxicity.The results indicate that 3D8 scFv could be utilized as an effective antiviral agent in several animal models.

View Article: PubMed Central - PubMed

Affiliation: Department of Genetic Engineering, Sungkyunkwan University, Suwon 440-746, Korea.

ABSTRACT
3D8 single chain variable fragment (scFv) is a recombinant monoclonal antibody with nuclease activity that was originally isolated from autoimmune-prone MRL mice. In a previous study, we analyzed the nuclease activity of 3D8 scFv and determined that a HeLa cell line expressing 3D8 scFv conferred resistance to herpes simplex virus type 1 (HSV-1) and pseudorabies virus (PRV). In this study, we demonstrate that 3D8 scFv could be delivered to target tissues and cells where it exerted a therapeutic effect against PRV. PRV was inoculated via intramuscular injection, and 3D8 scFv was injected intraperitoneally. The observed therapeutic effect of 3D8 scFv against PRV was also supported by results from quantitative reverse transcription polymerase chain reaction, southern hybridization, and immunohistochemical assays. Intraperitoneal injection of 5 and 10 μg 3D8 scFv resulted in no detectable toxicity. The survival rate in C57BL/6 mice was 9% after intramuscular injection of 10 LD50 PRV. In contrast, the 3D8 scFv-injected C57BL/6 mice showed survival rates of 57% (5 μg) and 47% (10 μg). The results indicate that 3D8 scFv could be utilized as an effective antiviral agent in several animal models.

No MeSH data available.


Related in: MedlinePlus

Antiviral mechanism by 3D8 scFv protein. (A) This model shows the pseudorabies virus (PRV) replication cycle. Entry begins with attachment or binding of the virus particle to the cell surface. The viral DNA enclosed by nucleocapsid delivers into nucleus. Viral DNA synthesis begins after the expression of the beta proteins and the temporal program of viral gene expression ends with the appearance of the gamma or late proteins, which constitute the structural proteins of the virus. Finally, the virus initiates a lytic cycle. (B) Mice were injected intraperitoneally with purified 3D8 scFv protein. The treated 3D8 scFv proteins penetrate the cell via a caveolae-mediated endocytosis mechanism. Consequentially, 3D8 scFv proteins are localized in cytoplasm. (C) 3D8 scFv proteins localized in cytoplasm degrade viral RNA by RNase activity and inhibit the expression of the viral proteins. The RNase activity of 3D8 scFv exhibits the therapeutic effects against PRV infection.
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f6-molce-38-9-773: Antiviral mechanism by 3D8 scFv protein. (A) This model shows the pseudorabies virus (PRV) replication cycle. Entry begins with attachment or binding of the virus particle to the cell surface. The viral DNA enclosed by nucleocapsid delivers into nucleus. Viral DNA synthesis begins after the expression of the beta proteins and the temporal program of viral gene expression ends with the appearance of the gamma or late proteins, which constitute the structural proteins of the virus. Finally, the virus initiates a lytic cycle. (B) Mice were injected intraperitoneally with purified 3D8 scFv protein. The treated 3D8 scFv proteins penetrate the cell via a caveolae-mediated endocytosis mechanism. Consequentially, 3D8 scFv proteins are localized in cytoplasm. (C) 3D8 scFv proteins localized in cytoplasm degrade viral RNA by RNase activity and inhibit the expression of the viral proteins. The RNase activity of 3D8 scFv exhibits the therapeutic effects against PRV infection.

Mentions: In conclusion, as an extension of our previous studies in which the antiviral effect of 3D8 scFv was demonstrated by in vitro experiments, we further showed that nuclease activity of 3D8 scFv could be effectively employed in vivo as an antiviral therapeutic agent using the mouse model system. Purified 3D8 scFv protein from E. coli was i.p. injected into PRV infected mice and localized to the cytoplasm by caveolae-mediated endocytosis (Jun et al., 2010). Then, the PRV transcriptome was degraded by 3D8 scFv with intrinsic RNase activity in cytoplasm (Fig. 6). The finding that 3D8 scFv nuclease activity acts on both DNA and RNA demonstrates the utility of 3D8 scFv as a multifaceted, broad-spectrum antiviral agent that works against DNA and RNA viruses.


Therapeutic Strategy for the Prevention of Pseudorabies Virus Infection in C57BL/6 Mice by 3D8 scFv with Intrinsic Nuclease Activity.

Lee G, Cho S, Hoang PM, Kim D, Lee Y, Kil EJ, Byun SJ, Lee TK, Kim DH, Kim S, Lee S - Mol. Cells (2015)

Antiviral mechanism by 3D8 scFv protein. (A) This model shows the pseudorabies virus (PRV) replication cycle. Entry begins with attachment or binding of the virus particle to the cell surface. The viral DNA enclosed by nucleocapsid delivers into nucleus. Viral DNA synthesis begins after the expression of the beta proteins and the temporal program of viral gene expression ends with the appearance of the gamma or late proteins, which constitute the structural proteins of the virus. Finally, the virus initiates a lytic cycle. (B) Mice were injected intraperitoneally with purified 3D8 scFv protein. The treated 3D8 scFv proteins penetrate the cell via a caveolae-mediated endocytosis mechanism. Consequentially, 3D8 scFv proteins are localized in cytoplasm. (C) 3D8 scFv proteins localized in cytoplasm degrade viral RNA by RNase activity and inhibit the expression of the viral proteins. The RNase activity of 3D8 scFv exhibits the therapeutic effects against PRV infection.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4588720&req=5

f6-molce-38-9-773: Antiviral mechanism by 3D8 scFv protein. (A) This model shows the pseudorabies virus (PRV) replication cycle. Entry begins with attachment or binding of the virus particle to the cell surface. The viral DNA enclosed by nucleocapsid delivers into nucleus. Viral DNA synthesis begins after the expression of the beta proteins and the temporal program of viral gene expression ends with the appearance of the gamma or late proteins, which constitute the structural proteins of the virus. Finally, the virus initiates a lytic cycle. (B) Mice were injected intraperitoneally with purified 3D8 scFv protein. The treated 3D8 scFv proteins penetrate the cell via a caveolae-mediated endocytosis mechanism. Consequentially, 3D8 scFv proteins are localized in cytoplasm. (C) 3D8 scFv proteins localized in cytoplasm degrade viral RNA by RNase activity and inhibit the expression of the viral proteins. The RNase activity of 3D8 scFv exhibits the therapeutic effects against PRV infection.
Mentions: In conclusion, as an extension of our previous studies in which the antiviral effect of 3D8 scFv was demonstrated by in vitro experiments, we further showed that nuclease activity of 3D8 scFv could be effectively employed in vivo as an antiviral therapeutic agent using the mouse model system. Purified 3D8 scFv protein from E. coli was i.p. injected into PRV infected mice and localized to the cytoplasm by caveolae-mediated endocytosis (Jun et al., 2010). Then, the PRV transcriptome was degraded by 3D8 scFv with intrinsic RNase activity in cytoplasm (Fig. 6). The finding that 3D8 scFv nuclease activity acts on both DNA and RNA demonstrates the utility of 3D8 scFv as a multifaceted, broad-spectrum antiviral agent that works against DNA and RNA viruses.

Bottom Line: The observed therapeutic effect of 3D8 scFv against PRV was also supported by results from quantitative reverse transcription polymerase chain reaction, southern hybridization, and immunohistochemical assays.Intraperitoneal injection of 5 and 10 μg 3D8 scFv resulted in no detectable toxicity.The results indicate that 3D8 scFv could be utilized as an effective antiviral agent in several animal models.

View Article: PubMed Central - PubMed

Affiliation: Department of Genetic Engineering, Sungkyunkwan University, Suwon 440-746, Korea.

ABSTRACT
3D8 single chain variable fragment (scFv) is a recombinant monoclonal antibody with nuclease activity that was originally isolated from autoimmune-prone MRL mice. In a previous study, we analyzed the nuclease activity of 3D8 scFv and determined that a HeLa cell line expressing 3D8 scFv conferred resistance to herpes simplex virus type 1 (HSV-1) and pseudorabies virus (PRV). In this study, we demonstrate that 3D8 scFv could be delivered to target tissues and cells where it exerted a therapeutic effect against PRV. PRV was inoculated via intramuscular injection, and 3D8 scFv was injected intraperitoneally. The observed therapeutic effect of 3D8 scFv against PRV was also supported by results from quantitative reverse transcription polymerase chain reaction, southern hybridization, and immunohistochemical assays. Intraperitoneal injection of 5 and 10 μg 3D8 scFv resulted in no detectable toxicity. The survival rate in C57BL/6 mice was 9% after intramuscular injection of 10 LD50 PRV. In contrast, the 3D8 scFv-injected C57BL/6 mice showed survival rates of 57% (5 μg) and 47% (10 μg). The results indicate that 3D8 scFv could be utilized as an effective antiviral agent in several animal models.

No MeSH data available.


Related in: MedlinePlus