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Herpes simplex type 2 virus deleted in glycoprotein D protects against vaginal, skin and neural disease.

Petro C, González PA, Cheshenko N, Jandl T, Khajoueinejad N, Bénard A, Sengupta M, Herold BC, Jacobs WR - Elife (2015)

Bottom Line: Subunit vaccines comprised of glycoprotein D (gD-2) failed to prevent HSV-2 highlighting need for novel strategies.The antibodies elicited cell-mediated cytotoxicity, but little neutralizing activity.These studies demonstrate that non-neutralizing Fc-mediated humoral responses confer protection and support advancement of this attenuated vaccine.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, Albert Einstein College of Medicine, New York, United States.

ABSTRACT
Subunit vaccines comprised of glycoprotein D (gD-2) failed to prevent HSV-2 highlighting need for novel strategies. To test the hypothesis that deletion of gD-2 unmasks protective antigens, we evaluated the efficacy and safety of an HSV-2 virus deleted in gD-2 and complemented allowing a single round of replication on cells expressing HSV-1 gD (ΔgD(-/+gD-1)). Subcutaneous immunization of C57BL/6 or BALB/c mice with ΔgD(-/+gD1) provided 100% protection against lethal intravaginal or skin challenges and prevented latency. ΔgD(-/+gD1) elicited no disease in SCID mice, whereas 1000-fold lower doses of wild-type virus were lethal. HSV-specific antibodies were detected in serum (titer 1:800,000) following immunization and in vaginal washes after intravaginal challenge. The antibodies elicited cell-mediated cytotoxicity, but little neutralizing activity. Passive transfer of immune serum completely protected wild-type, but not Fcγ-receptor or neonatal Fc-receptor knock-out mice. These studies demonstrate that non-neutralizing Fc-mediated humoral responses confer protection and support advancement of this attenuated vaccine.

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Characterization of the ΔgD−/− virus.(A) Alignment of the upstream and downstream regions of gD located within the HSV-1 BamHI J fragment encoded in VD60 cells and within the genome of ΔgD−/− using LALIGN (ExPASy) (Myers and Miller, 1988). Global alignments assess end-to-end sequences and local pairwise alignments search for regions with high identity. (B) Western blots of dextran gradient-purified virus isolated 24 hr after infection of VD60 (ΔgD−/+gD−1) and Vero (ΔgD−/−) cells. Protein expression was assessed for viral glycoproteins B (gB, UL27), gC (UL44), gD (Us6), gE (Us8), gH (Us22), gL (UL1), VP5 (UL19) and VP16 (UL48).DOI:http://dx.doi.org/10.7554/eLife.06054.003
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fig1: Characterization of the ΔgD−/− virus.(A) Alignment of the upstream and downstream regions of gD located within the HSV-1 BamHI J fragment encoded in VD60 cells and within the genome of ΔgD−/− using LALIGN (ExPASy) (Myers and Miller, 1988). Global alignments assess end-to-end sequences and local pairwise alignments search for regions with high identity. (B) Western blots of dextran gradient-purified virus isolated 24 hr after infection of VD60 (ΔgD−/+gD−1) and Vero (ΔgD−/−) cells. Protein expression was assessed for viral glycoproteins B (gB, UL27), gC (UL44), gD (Us6), gE (Us8), gH (Us22), gL (UL1), VP5 (UL19) and VP16 (UL48).DOI:http://dx.doi.org/10.7554/eLife.06054.003

Mentions: We reasoned that deletion of gD-2, an envelope glycoprotein required for viral entry and cell-to-cell spread, would restrict infection to a single round of viral replication and thus provide a safe, highly attenuated candidate vaccine. Therefore, we constructed a variant of HSV-2(G) deleted for US6 (gD-2) (Cheshenko et al., 2013). The virus was subsequently subjected to three rounds of plaque purification and grew to high titers (108 pfu/ml) when phenotypically complemented by growing the virus on VD60 cells, which express HSV-1 gD under the control of the gD-1 promoter. No plaques were observed when three independent plaque-purified isolates were grown to 108 pfu on VD60 cells and then plated on non-complementing Vero cells. Moreover, no recombinants were detected by PCR, most likely reflecting differences in the regions flanking gD in VD60 cells and in HSV-2 (Figure 1A). Western blots show similar levels of expression of other HSV viral proteins when the deletion virus is grown on VD60 or Vero cells (Figure 1B).10.7554/eLife.06054.003Figure 1.Characterization of the ΔgD−/− virus.


Herpes simplex type 2 virus deleted in glycoprotein D protects against vaginal, skin and neural disease.

Petro C, González PA, Cheshenko N, Jandl T, Khajoueinejad N, Bénard A, Sengupta M, Herold BC, Jacobs WR - Elife (2015)

Characterization of the ΔgD−/− virus.(A) Alignment of the upstream and downstream regions of gD located within the HSV-1 BamHI J fragment encoded in VD60 cells and within the genome of ΔgD−/− using LALIGN (ExPASy) (Myers and Miller, 1988). Global alignments assess end-to-end sequences and local pairwise alignments search for regions with high identity. (B) Western blots of dextran gradient-purified virus isolated 24 hr after infection of VD60 (ΔgD−/+gD−1) and Vero (ΔgD−/−) cells. Protein expression was assessed for viral glycoproteins B (gB, UL27), gC (UL44), gD (Us6), gE (Us8), gH (Us22), gL (UL1), VP5 (UL19) and VP16 (UL48).DOI:http://dx.doi.org/10.7554/eLife.06054.003
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4352706&req=5

fig1: Characterization of the ΔgD−/− virus.(A) Alignment of the upstream and downstream regions of gD located within the HSV-1 BamHI J fragment encoded in VD60 cells and within the genome of ΔgD−/− using LALIGN (ExPASy) (Myers and Miller, 1988). Global alignments assess end-to-end sequences and local pairwise alignments search for regions with high identity. (B) Western blots of dextran gradient-purified virus isolated 24 hr after infection of VD60 (ΔgD−/+gD−1) and Vero (ΔgD−/−) cells. Protein expression was assessed for viral glycoproteins B (gB, UL27), gC (UL44), gD (Us6), gE (Us8), gH (Us22), gL (UL1), VP5 (UL19) and VP16 (UL48).DOI:http://dx.doi.org/10.7554/eLife.06054.003
Mentions: We reasoned that deletion of gD-2, an envelope glycoprotein required for viral entry and cell-to-cell spread, would restrict infection to a single round of viral replication and thus provide a safe, highly attenuated candidate vaccine. Therefore, we constructed a variant of HSV-2(G) deleted for US6 (gD-2) (Cheshenko et al., 2013). The virus was subsequently subjected to three rounds of plaque purification and grew to high titers (108 pfu/ml) when phenotypically complemented by growing the virus on VD60 cells, which express HSV-1 gD under the control of the gD-1 promoter. No plaques were observed when three independent plaque-purified isolates were grown to 108 pfu on VD60 cells and then plated on non-complementing Vero cells. Moreover, no recombinants were detected by PCR, most likely reflecting differences in the regions flanking gD in VD60 cells and in HSV-2 (Figure 1A). Western blots show similar levels of expression of other HSV viral proteins when the deletion virus is grown on VD60 or Vero cells (Figure 1B).10.7554/eLife.06054.003Figure 1.Characterization of the ΔgD−/− virus.

Bottom Line: Subunit vaccines comprised of glycoprotein D (gD-2) failed to prevent HSV-2 highlighting need for novel strategies.The antibodies elicited cell-mediated cytotoxicity, but little neutralizing activity.These studies demonstrate that non-neutralizing Fc-mediated humoral responses confer protection and support advancement of this attenuated vaccine.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, Albert Einstein College of Medicine, New York, United States.

ABSTRACT
Subunit vaccines comprised of glycoprotein D (gD-2) failed to prevent HSV-2 highlighting need for novel strategies. To test the hypothesis that deletion of gD-2 unmasks protective antigens, we evaluated the efficacy and safety of an HSV-2 virus deleted in gD-2 and complemented allowing a single round of replication on cells expressing HSV-1 gD (ΔgD(-/+gD-1)). Subcutaneous immunization of C57BL/6 or BALB/c mice with ΔgD(-/+gD1) provided 100% protection against lethal intravaginal or skin challenges and prevented latency. ΔgD(-/+gD1) elicited no disease in SCID mice, whereas 1000-fold lower doses of wild-type virus were lethal. HSV-specific antibodies were detected in serum (titer 1:800,000) following immunization and in vaginal washes after intravaginal challenge. The antibodies elicited cell-mediated cytotoxicity, but little neutralizing activity. Passive transfer of immune serum completely protected wild-type, but not Fcγ-receptor or neonatal Fc-receptor knock-out mice. These studies demonstrate that non-neutralizing Fc-mediated humoral responses confer protection and support advancement of this attenuated vaccine.

Show MeSH
Related in: MedlinePlus