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Anti-gp120 minibody gene transfer to female genital epithelial cells protects against HIV-1 virus challenge in vitro.

Abdel-Motal UM, Sarkis PT, Han T, Pudney J, Anderson DJ, Zhu Q, Marasco WA - PLoS ONE (2011)

Bottom Line: Accordingly, a recombinant AAV vector that encodes human b12 anti-HIV gp120 BnAb as a single-chain variable fragment Fc fusion (scFvFc), or "minibody" was constructed.Furthermore, cervico-vaginal epithelial stem cells were found to be efficiently transduced by the optimal AAV serotype mediated expression of GFP.This study provides the foundation for a novel microbicide strategy to protect against sexual transmission of HIV-1 by AAV transfer of broadly neutralizing antibody genes to cervico-vaginal epithelial stem cells that could replenish b12 BnAb secreting cells through multiple menstrual cycles.

View Article: PubMed Central - PubMed

Affiliation: Department of Cancer Immunology and AIDS, Dana-Farber Cancer Institute, Harvard Medical School, Boston, Massachusetts, United States of America.

ABSTRACT

Background: Although cervico-vaginal epithelial cells of the female lower genital tract provide the initial defense system against HIV-1 infection, the protection is sometimes incomplete. Thus, enhancing anti-HIV-1 humoral immunity at the mucosal cell surface by local expression of anti-HIV-1 broadly neutralizing antibodies (BnAb) that block HIV-1 entry would provide an important new intervention that could slow the spread of HIV/AIDS.

Methods and findings: This study tested the hypothesis that adeno-associated virus (AAV)-BnAb gene transfer to cervico-vaginal epithelial cells will lead to protection against HIV-1. Accordingly, a recombinant AAV vector that encodes human b12 anti-HIV gp120 BnAb as a single-chain variable fragment Fc fusion (scFvFc), or "minibody" was constructed. The secreted b12 minibody was shown to be biologically functional in binding to virus envelope protein, neutralizing HIV-1 and importantly, blocking transfer and infectivity of HIV-1(bal) in an organotypic human vaginal epithelial cell (VEC) model. Furthermore, cervico-vaginal epithelial stem cells were found to be efficiently transduced by the optimal AAV serotype mediated expression of GFP.

Conclusion: This study provides the foundation for a novel microbicide strategy to protect against sexual transmission of HIV-1 by AAV transfer of broadly neutralizing antibody genes to cervico-vaginal epithelial stem cells that could replenish b12 BnAb secreting cells through multiple menstrual cycles.

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Related in: MedlinePlus

Inhibition of HIV-1 transfer and activity by b12 minibodies in the human VEC organotypic model tissues.After a 1 h pre-incubation of b12 minibodies or full-length b12 IgG (10 µg/ml) with HIV-1bal (50 ng), medium from the basal chambers were collected at different time points and tested for inhibition of HIV-1bal transfer by measuring p24 content by ELISA (A) and for inhibition of virus infectivity by incubation on TZM-bl target cells (B). Note that media collected at 3 and 6 h from tissue samples treated with HIV-1bal and b12 IgG1 antibodies or with b12 minibodies had almost completely lost their ability to infect TZM-bl cells. Irrelevant 11 A minibodies served as negative controls.
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pone-0026473-g005: Inhibition of HIV-1 transfer and activity by b12 minibodies in the human VEC organotypic model tissues.After a 1 h pre-incubation of b12 minibodies or full-length b12 IgG (10 µg/ml) with HIV-1bal (50 ng), medium from the basal chambers were collected at different time points and tested for inhibition of HIV-1bal transfer by measuring p24 content by ELISA (A) and for inhibition of virus infectivity by incubation on TZM-bl target cells (B). Note that media collected at 3 and 6 h from tissue samples treated with HIV-1bal and b12 IgG1 antibodies or with b12 minibodies had almost completely lost their ability to infect TZM-bl cells. Irrelevant 11 A minibodies served as negative controls.

Mentions: As the b12 minibody was confirmed to have the same capacity as full-length b12 IgG1 to neutralize HIV-1 infectivity (Fig. 4B), its activity was further compared with the full-length b12 IgG1 in preventing HIV-1 transfer and subsequent infectivity. An organotypic vaginal epithelial cells transwell tissue model (VEC tissue) was used as described in Materials and Methods. After confirming the integrity of the VEC tissues using exclusion of 70 kDa Dextran-Rhodamine from the lower chamber, b12 minibodies, full-length b12 IgG1 or irrelevant control minibodies (each 10 µg/ml) were incubated with or without HIV-1bal virus (50 ng) and then added to the apical surface of the organotypic VEC tissue in transwell inserts. The media from the lower chambers were then collected at different time points to measure the amounts of viral particles that have crossed the VEC tissues and to determine if they were infectious. The amounts of viral particles were measured by p24 ELISA (Fig. 5A), and infectivity was tested by incubation of the samples with the TZM-bl cells and measuring luciferase gene activity as discussed previously (Fig. 5B).


Anti-gp120 minibody gene transfer to female genital epithelial cells protects against HIV-1 virus challenge in vitro.

Abdel-Motal UM, Sarkis PT, Han T, Pudney J, Anderson DJ, Zhu Q, Marasco WA - PLoS ONE (2011)

Inhibition of HIV-1 transfer and activity by b12 minibodies in the human VEC organotypic model tissues.After a 1 h pre-incubation of b12 minibodies or full-length b12 IgG (10 µg/ml) with HIV-1bal (50 ng), medium from the basal chambers were collected at different time points and tested for inhibition of HIV-1bal transfer by measuring p24 content by ELISA (A) and for inhibition of virus infectivity by incubation on TZM-bl target cells (B). Note that media collected at 3 and 6 h from tissue samples treated with HIV-1bal and b12 IgG1 antibodies or with b12 minibodies had almost completely lost their ability to infect TZM-bl cells. Irrelevant 11 A minibodies served as negative controls.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3198777&req=5

pone-0026473-g005: Inhibition of HIV-1 transfer and activity by b12 minibodies in the human VEC organotypic model tissues.After a 1 h pre-incubation of b12 minibodies or full-length b12 IgG (10 µg/ml) with HIV-1bal (50 ng), medium from the basal chambers were collected at different time points and tested for inhibition of HIV-1bal transfer by measuring p24 content by ELISA (A) and for inhibition of virus infectivity by incubation on TZM-bl target cells (B). Note that media collected at 3 and 6 h from tissue samples treated with HIV-1bal and b12 IgG1 antibodies or with b12 minibodies had almost completely lost their ability to infect TZM-bl cells. Irrelevant 11 A minibodies served as negative controls.
Mentions: As the b12 minibody was confirmed to have the same capacity as full-length b12 IgG1 to neutralize HIV-1 infectivity (Fig. 4B), its activity was further compared with the full-length b12 IgG1 in preventing HIV-1 transfer and subsequent infectivity. An organotypic vaginal epithelial cells transwell tissue model (VEC tissue) was used as described in Materials and Methods. After confirming the integrity of the VEC tissues using exclusion of 70 kDa Dextran-Rhodamine from the lower chamber, b12 minibodies, full-length b12 IgG1 or irrelevant control minibodies (each 10 µg/ml) were incubated with or without HIV-1bal virus (50 ng) and then added to the apical surface of the organotypic VEC tissue in transwell inserts. The media from the lower chambers were then collected at different time points to measure the amounts of viral particles that have crossed the VEC tissues and to determine if they were infectious. The amounts of viral particles were measured by p24 ELISA (Fig. 5A), and infectivity was tested by incubation of the samples with the TZM-bl cells and measuring luciferase gene activity as discussed previously (Fig. 5B).

Bottom Line: Accordingly, a recombinant AAV vector that encodes human b12 anti-HIV gp120 BnAb as a single-chain variable fragment Fc fusion (scFvFc), or "minibody" was constructed.Furthermore, cervico-vaginal epithelial stem cells were found to be efficiently transduced by the optimal AAV serotype mediated expression of GFP.This study provides the foundation for a novel microbicide strategy to protect against sexual transmission of HIV-1 by AAV transfer of broadly neutralizing antibody genes to cervico-vaginal epithelial stem cells that could replenish b12 BnAb secreting cells through multiple menstrual cycles.

View Article: PubMed Central - PubMed

Affiliation: Department of Cancer Immunology and AIDS, Dana-Farber Cancer Institute, Harvard Medical School, Boston, Massachusetts, United States of America.

ABSTRACT

Background: Although cervico-vaginal epithelial cells of the female lower genital tract provide the initial defense system against HIV-1 infection, the protection is sometimes incomplete. Thus, enhancing anti-HIV-1 humoral immunity at the mucosal cell surface by local expression of anti-HIV-1 broadly neutralizing antibodies (BnAb) that block HIV-1 entry would provide an important new intervention that could slow the spread of HIV/AIDS.

Methods and findings: This study tested the hypothesis that adeno-associated virus (AAV)-BnAb gene transfer to cervico-vaginal epithelial cells will lead to protection against HIV-1. Accordingly, a recombinant AAV vector that encodes human b12 anti-HIV gp120 BnAb as a single-chain variable fragment Fc fusion (scFvFc), or "minibody" was constructed. The secreted b12 minibody was shown to be biologically functional in binding to virus envelope protein, neutralizing HIV-1 and importantly, blocking transfer and infectivity of HIV-1(bal) in an organotypic human vaginal epithelial cell (VEC) model. Furthermore, cervico-vaginal epithelial stem cells were found to be efficiently transduced by the optimal AAV serotype mediated expression of GFP.

Conclusion: This study provides the foundation for a novel microbicide strategy to protect against sexual transmission of HIV-1 by AAV transfer of broadly neutralizing antibody genes to cervico-vaginal epithelial stem cells that could replenish b12 BnAb secreting cells through multiple menstrual cycles.

Show MeSH
Related in: MedlinePlus