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. 2014 Jan;22(1):123-31.
doi: 10.1038/mt.2013.238. Epub 2013 Oct 9.

Immunogenic HSV-mediated oncolysis shapes the antitumor immune response and contributes to therapeutic efficacy

Samuel T Workenhe  1 , Graydon Simmons  1 , Jonathan G Pol  1 , Brian D Lichty  1 , William P Halford  2 , Karen L Mossman  1
Affiliations

Immunogenic HSV-mediated oncolysis shapes the antitumor immune response and contributes to therapeutic efficacy

Samuel T Workenhe et al. Mol Ther. 2014 Jan.

Abstract

Within the oncolytic virus field, the extent of virus replication that is essential for immune stimulation to control tumor growth remains unresolved. Using infected cell protein 0 (ICP0)-defective oncolytic Herpes simplex virus type 1 (HSV-1) and HSV-2 viruses (dICP0 and dNLS) that show differences in their in vitro replication and cytotoxicity, we investigated the inherent features of oncolytic HSV viruses that are required for potent antitumor activity. In vitro, the HSV-2 vectors showed rapid cytotoxicity despite lower viral burst sizes compared to HSV-1 vectors. In vivo, although both of the dICP0 vectors initially replicated to a similar level, HSV-1 dICP0 was rapidly cleared from the tumors. In spite of this rapid clearance, HSV-1 dICP0 treatment conferred significant survival benefit. HSV-1 dICP0-treated tumors showed significantly higher levels of danger-associated molecular patterns that correlated with higher numbers of antigen-presenting cells within the tumor and increased antigen-specific CD8+ T-cell levels in the peripheral blood. This study suggests that, at least in the context of oncolytic HSV, the initial stages of immunogenic virus replication leading to activation of antitumor immunity are more important than persistence of a replicating virus within the tumor. This knowledge provides important insight for the design of therapeutically successful oncolytic viruses.

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Figures

Figure 1
Figure 1
Herpes simplex virus (HSV)-1 and HSV-2 ICP0 mutants show differences in cytopathic effect, cellular metabolism and viral burst size in vitro. Bright field and green fluorescent protein fluorescence images of TUBO cells infected with (a) HSV-1 or (b) HSV-2 ICP0 mutants for 24 hours. Cellular metabolism of TUBO cells 24 hours following infection with HSV-1 or HSV-2 ICP0 mutants at (c) MOI 1 and (d) MOI 5. Burst sizes of HSV-1 and HSV-2 ICP0 mutants in TUBO cells 48 hours following infection with an (e) MOI of 0.5. The average and standard deviations are from three independent experiments each run with three biological replicates. The statistical significance of the mean differences between groups was analyzed using ANOVA. ***P < 0.0005, **P < 0.005.
Figure 2
Figure 2
HSV-2 dICP0–infected TUBO cells show an early apoptotic cell death. Western blot analysis of cleaved caspase 3 in TUBO cells treated with (a) 62.5 nm staurosporin for 16 hours or (b) following mock treatment or infection with HSV wild-type (MOI 0.5) or ICP0 mutant (MOI 2.5) viruses. The blots were used to calculate the ratio of cleaved caspase 3 to actin by densitometry using ImageJ software. Shown is a representative of three independent experiments. (c) HMGB1 levels in the supernatant after infection with different MOIs of HSV-1 and HSV-2 mutants for 24-hour period. The plots are values generated from an ELISA using pooled samples from triplicate independent experiments.
Figure 3
Figure 3
Schematic diagram of experimental procedures performed. Illustrated are the relative time frames of tumor implantation, oncolytic virus treatment, harvesting of tumors for protein and virus quantitation, IVIS imaging, and blood collection for intracellular cytokine staining.
Figure 4
Figure 4
Oncolytic HSV ICP0 mutants show different levels of therapeutic efficacy in vivo. A total of 5 × 105 TUBO cells were implanted into BALB/c mice by subcutaneous injection into the left flank. When tumors reached treatable size, 1 × 107 total pfu of each virus was administered intratumorally three times every 36 hours. (a) Kaplan–Meier survival analysis following treatment. (b) Fold change in tumor volume relative to the tumor volume at the start of treatment. **P < 0.005.
Figure 5
Figure 5
Locally administered HSV-1 dICP0 is rapidly cleared from subcutaneous tumors. Replication of HSV dICP0 mutants in subcutaneous tumors of TUBO cells in BALB/c mice. (a) Log of total flux calculated following IVIS imaging of tumor-bearing mice treated with firefly luciferase expressing oncolytic HSV dICP0 viruses. Luciferase activity was measured following injection of mice with D-luciferin (n = 5 for each treatment). (b) Viral titers from tumors that were resected and homogenized at the indicated times posttreatment.
Figure 6
Figure 6
HSV-1 dICP0 treatment leads to a higher expression of immunomodulatory molecules compared to HSV-2 dICP0. In vivo assessment of immunogenic cell death in tumor-bearing BALB/c mice treated with HSV dICP0 oncolytic viruses. Tumors (three per group) were harvested at (a) 36, (b) 72, and (c) 96 hours after the first oncolytic virus treatment, and the levels of cleaved caspase 3 and HSP-70 were measured by western blot analyses. (d) Serum levels of HMGB1 were measured by ELISA 24 hours after the first treatment with HSV dICP0 oncolytic viruses.
Figure 7
Figure 7
HSV-1 dICP0–treated tumors have higher infiltration of antigen-presenting cells and neutrophils. Tumors were treated with three doses of HSV dICP0. Ten days after treatment, the tumors were resected and CD45+ immune cells isolated using EasySep magnetic bead positive selection. Cells were directly stained with the indicated surface markers. The average and standard deviation of cell numbers were generated from five mice per each treatment group. ns, not significant, *P < 0.05.
Figure 8
Figure 8
HSV-1 dICP0–treated tumor-bearing mice have higher antigen-specific cytotoxic CD8+ T cells. (a) Percentage of HER-2–specific CD8+ T cells in the peripheral blood of treated tumor-bearing BALB/c mice. Blood samples were processed, and cells were restimulated with the HER-2–immunodominant epitope peptide and stained for the cell surface marker CD8 and intracellular IFN-γ to identify HER-2–specific CD8+ T cells (n = 5 for each treatment). The χ2 test was used to test the statistical significance of the differences in proportion of antigen-specific CD8+ T cells. (b) Viability assessed by measurement of cellular metabolism in TUBO cells left untreated or cocultured with CD8+ T splenocytes harvested from tumor-bearing mice treated with PBS or HSV dICP0. The values are average of three wells that used pooled splenocytes from five mice. ***P < 0.0005.
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References

    1. Vähä-Koskela MJ, Heikkilä JE, Hinkkanen AE. Oncolytic viruses in cancer therapy. Cancer Lett. 2007;254:178–216. - PMC - PubMed
    1. Russell SJ, Peng KW, Bell JC. Oncolytic virotherapy. Nat Biotechnol. 2012;30:658–670. - PMC - PubMed
    1. Martuza RL, Malick A, Markert JM, Ruffner KL, Coen DM. Experimental therapy of human glioma by means of a genetically engineered virus mutant. Science. 1991;252:854–856. - PubMed
    1. Mossman KL, Smiley JR. Herpes simplex virus ICP0 and ICP34.5 counteract distinct interferon-induced barriers to virus replication. J Virol. 2002;76:1995–1998. - PMC - PubMed
    1. Mossman KL, Saffran HA, Smiley JR. Herpes simplex virus ICP0 mutants are hypersensitive to interferon. J Virol. 2000;74:2052–2056. - PMC - PubMed

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