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Novel gene therapy viral vector using non-oncogenic lymphotropic herpesvirus.

Shimizu A, Kobayashi N, Shimada K, Oura K, Tanaka T, Okamoto A, Kondo K - PLoS ONE (2013)

Bottom Line: In the present study, we have altered the cell specificity of the resulting recombinant HHV-6 by knocking out the U2-U8 genes.Furthermore, HHV-6 vectors containing short hairpin RNAs against CD4 and HIV Gag remarkably inhibited the production of these proteins and HIV particles.Here we demonstrate the utility of HHV-6 as a new non-carcinogenic viral vector for immunologic diseases and immunotherapy.

View Article: PubMed Central - PubMed

Affiliation: Department of Virology, The Jikei University School of Medicine, Tokyo, Japan.

ABSTRACT
Despite the use of retroviral vectors, efficiently introducing target genes into immunocytes such as T cells is difficult. In addition, retroviral vectors carry risks associated with the oncogenicity of the native virus and the potential for introducing malignancy in recipients due to genetic carryover from immortalized cells used during vector production. To address these issues, we have established a new virus vector that is based on human herpesvirus 6 (HHV-6), a non-oncogenic lymphotropic herpesvirus that infects CD4(+) T cells, macrophages, and dendritic cells. In the present study, we have altered the cell specificity of the resulting recombinant HHV-6 by knocking out the U2-U8 genes. The resulting virus proliferated only in activated cord blood cells and not in peripheral blood cells. Umbilical cord blood cells produced replication-defective recombinant virus in sufficiently high titer to omit the use of immortalized cells during vector production. HHV-6 vectors led to high rates (>90%) of gene transduction in both CD4(+) and CD8(+) T cells. These viruses showed low-level replication of viral DNA that supported greater expression of the induced genes than that of other methods but that was insufficient to support the production of replication-competent virus. Furthermore, HHV-6 vectors containing short hairpin RNAs against CD4 and HIV Gag remarkably inhibited the production of these proteins and HIV particles. Here we demonstrate the utility of HHV-6 as a new non-carcinogenic viral vector for immunologic diseases and immunotherapy.

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Related in: MedlinePlus

H6R28LEP activity in human serum.H6R28LEP viral solution was mixed with RPMI containing 10% FBS or human serum at a 10∶1 ratio, and the virus titer was assayed after the mixture was left to stand at 37°C for 0, 30, 60, and 120 min. The human serum was sampled from three healthy donors already infected with HHV-6. Data are given as mean ±1 standard deviation.
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pone-0056027-g004: H6R28LEP activity in human serum.H6R28LEP viral solution was mixed with RPMI containing 10% FBS or human serum at a 10∶1 ratio, and the virus titer was assayed after the mixture was left to stand at 37°C for 0, 30, 60, and 120 min. The human serum was sampled from three healthy donors already infected with HHV-6. Data are given as mean ±1 standard deviation.

Mentions: As most individuals are infected with HHV-6 during their childhood and the virus remains dormant in the body, neutralizing antibodies are likely to be present in the blood. For this reason, HHV-6 vectors can only be used ex vivo to infect target cells. However, this also means there is little risk that contaminating virus could replicate by infecting cells other than the target cells. As the experiments described above suggested changes in the H6R28LEP particle structure, we investigated whether serum from healthy individuals could neutralize H6R28LEP. The results revealed that H6R28LEP is rapidly neutralized by human serum (Fig. 4). We concluded that there is no new risk from contaminating virus.


Novel gene therapy viral vector using non-oncogenic lymphotropic herpesvirus.

Shimizu A, Kobayashi N, Shimada K, Oura K, Tanaka T, Okamoto A, Kondo K - PLoS ONE (2013)

H6R28LEP activity in human serum.H6R28LEP viral solution was mixed with RPMI containing 10% FBS or human serum at a 10∶1 ratio, and the virus titer was assayed after the mixture was left to stand at 37°C for 0, 30, 60, and 120 min. The human serum was sampled from three healthy donors already infected with HHV-6. Data are given as mean ±1 standard deviation.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3569415&req=5

pone-0056027-g004: H6R28LEP activity in human serum.H6R28LEP viral solution was mixed with RPMI containing 10% FBS or human serum at a 10∶1 ratio, and the virus titer was assayed after the mixture was left to stand at 37°C for 0, 30, 60, and 120 min. The human serum was sampled from three healthy donors already infected with HHV-6. Data are given as mean ±1 standard deviation.
Mentions: As most individuals are infected with HHV-6 during their childhood and the virus remains dormant in the body, neutralizing antibodies are likely to be present in the blood. For this reason, HHV-6 vectors can only be used ex vivo to infect target cells. However, this also means there is little risk that contaminating virus could replicate by infecting cells other than the target cells. As the experiments described above suggested changes in the H6R28LEP particle structure, we investigated whether serum from healthy individuals could neutralize H6R28LEP. The results revealed that H6R28LEP is rapidly neutralized by human serum (Fig. 4). We concluded that there is no new risk from contaminating virus.

Bottom Line: In the present study, we have altered the cell specificity of the resulting recombinant HHV-6 by knocking out the U2-U8 genes.Furthermore, HHV-6 vectors containing short hairpin RNAs against CD4 and HIV Gag remarkably inhibited the production of these proteins and HIV particles.Here we demonstrate the utility of HHV-6 as a new non-carcinogenic viral vector for immunologic diseases and immunotherapy.

View Article: PubMed Central - PubMed

Affiliation: Department of Virology, The Jikei University School of Medicine, Tokyo, Japan.

ABSTRACT
Despite the use of retroviral vectors, efficiently introducing target genes into immunocytes such as T cells is difficult. In addition, retroviral vectors carry risks associated with the oncogenicity of the native virus and the potential for introducing malignancy in recipients due to genetic carryover from immortalized cells used during vector production. To address these issues, we have established a new virus vector that is based on human herpesvirus 6 (HHV-6), a non-oncogenic lymphotropic herpesvirus that infects CD4(+) T cells, macrophages, and dendritic cells. In the present study, we have altered the cell specificity of the resulting recombinant HHV-6 by knocking out the U2-U8 genes. The resulting virus proliferated only in activated cord blood cells and not in peripheral blood cells. Umbilical cord blood cells produced replication-defective recombinant virus in sufficiently high titer to omit the use of immortalized cells during vector production. HHV-6 vectors led to high rates (>90%) of gene transduction in both CD4(+) and CD8(+) T cells. These viruses showed low-level replication of viral DNA that supported greater expression of the induced genes than that of other methods but that was insufficient to support the production of replication-competent virus. Furthermore, HHV-6 vectors containing short hairpin RNAs against CD4 and HIV Gag remarkably inhibited the production of these proteins and HIV particles. Here we demonstrate the utility of HHV-6 as a new non-carcinogenic viral vector for immunologic diseases and immunotherapy.

Show MeSH
Related in: MedlinePlus