Cancer Res Treat.  2020 Jan;52(1):309-319. 10.4143/crt.2019.161.

Characterization of Oncolytic Vaccinia Virus Harboring the Human IFNB1 and CES2 Transgenes

• Cho E
• Islam SMBU
• Jiang F
• Park JE
• Lee B
• Kim ND
• Hwang TH

• KMID: 2501223
• DOI: http://doi.org/10.4143/crt.2019.161

Abstract

Purpose
The purpose of this study was to assess characteristics of SJ-815, a novel oncolytic vaccinia virus lacking a functional thymidine kinase-encoding TK gene, and instead, having two human transgenes: the IFNB1 that encodes interferon β1, and the CES2 that encodes carboxylesterase 2, which metabolizes the prodrug, irinotecan, into cytotoxic SN-38.
Materials and Methods
Viral replication and dissemination of SJ-815 were measured by plaque assay and comet assay, respectively, and compared to the backbone of SJ-815, a modified Western Reserve virus named WI. Tumor cytotoxicity of SJ-815 (or mSJ-815, which has the murine IFNB1 transgene for mouse cancers) was evaluated using human and mouse cancer cells. Antitumor effects of SJ-815, with/without irinotecan, were evaluated using a human pancreatic cancer-bearing mouse model and a syngeneic melanoma-bearing mouse model. The SN-38/ irinotecan ratios in mouse melanoma tissue 4 days post irinotecan treatment were compared between groups with and without SJ-815 intravenous injection.
Results
SJ-815 demonstrated significantly lower viral replication and dissemination, but considerably stronger in vitro tumor cytotoxicity than WI. The combination use of SJ-815 plus irinotecan generated substantial tumor regression in the human pancreatic cancer model, and significantly prolonged survival in the melanoma model (hazard ratio, 0.11; 95% confidence interval, 0.02 to 0.50; p=0.013). The tumor SN-38/irinotecan ratios were over 3-fold higher in the group with SJ-815 than those without (p < 0.001).
Conclusion
SJ-815 demonstrates distinct characteristics gained from the inserted IFNB1 and CES2 transgenes. The potent antitumor effects of SJ-815, particularly when combined with irinotecan, against multiple solid tumors make SJ-815 an attractive candidate for further preclinical and clinical studies.

Keyword

Oncolytic viruses; Vaccinia virus; Carboxylesterase 2; Interferon β1; Irinotecan

Figure

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  Fig. 1. The activity of human proteins interferon β1 (IFN-β1) and carboxylesterase 2 (CES2) expressed by three final selected plaques of SJ-815 was demonstrated using HEK-Blue IFN-α/β cell assay (A), and p-nitrophenyl acetate assay (B).

• 

  Fig. 2. Viral replication of SJ-815 and other viruses in human (A, B) and mouse (C) cancer cell lines. Replication of Western Reserve (WR), modified Western Reserve (WI) (backbone), and SJ-815 in HeLa and HeLa S3 cells, with infection virus dose of 3 pfu/cell, at 2, 24, 48, and 72 hours (A); and in HeLa cells, with infection virus dose of 0.001, 0.01, 0.1, and 1 pfu/cell, at 24 hours (B). **p < 0.01, ***p < 0.001; WI (backbone) or SJ-815 vs. WR; the p-values were obtained using paired t tests. (C) Replication of WI (backbone), SJ-815, and mSJ-815 in MC-38 and B16-F10 mouse cancer cells, with infection virus dose of 0.1 pfu/cell at 72 hours (left), or 1 pfu/cell at 24 hours (right). *p < 0.05, **p < 0.01, ***p < 0.001; WI (backbone) or SJ-815 vs. mSJ-815; the p-values were obtained using paired t tests.

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  Fig. 3. Comet assays conducted on U-2 OS cells after infection with Western Reserve (WR) virus (A), modified Western Reserve (WI) (backbone) virus (B), or SJ-815 (C) for 2 hours first and then culture with Dulbecco's modified Eagle medium containing 2.5% fetal bovine serum for 72 hours; or after infection with WI (backbone) virus (D) for 2 hours first, and then culture with 1,000 U interferon β1 (IFN-β1), SJ-815 supernatant, or WR supernatant for 72 hours; or after infection with SJ-815 (E) for 2 hours first, and then culture with WR supernatant for 72 hours. The virus supernatants were collected after the U-2 OS cells were infected with the viruses: WR, WI (backbone), or SJ-815 for 16 hours, and filtered three times and stored at 4°C until use.

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  Fig. 4. (A) Comparison of tumor cytotoxicity (EC50) of SJ-815 and modified Western Reserve (WI) (backbone) in 12 human cancer cell lines. Each data was presented as mean±standard deviation of two EC50, which were calculated from two sets of dose-response data. (B) Tumor cytotoxicity (EC50) of mSJ-815 in 5 mouse cancer cell lines. Each data was presented as 95% confidence interval was EC50, which was calculated from only one set of dose-response data. *p < 0.05, **p < 0.01, ***p < 0.001; SJ-815 vs. WI (backbone); the p-values were obtained using paired t tests.

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  Fig. 5. Changes in tumor volume from day 0 to day 41 (sacrifice day) in human pancreatic cancer (MIA PaCa-2)-bearing mice, treated with phosphate-buffered saline (PBS), SJ-815 (1×106 pfu; intratumoral; day 7 and 14), irinotecan (25 mg/kg; intravenous; day 3, 10, and 17), or SJ-815 (1×106 pfu; intratumoral; day 7 and 14)+irinotecan (25 mg/kg; intravenous; day 3, 10, and 17). Statistical analyses were conducted to compare tumor volume on the sacrifice day. **p < 0.01, SJ-815 vs. SJ-815+irinotecan, ***p < 0.001, PBS vs. SJ-815+irinotecan; the p-values were obtained using t tests. Irinotecan was not compared because one mouse died before the sacrifice day.

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  Fig. 6. Percent survival (A) and body weight change (B) of B16-F10 melanoma-bearing mice, treated with phosphate-buffered saline (PBS), mSJ-815 (3×107 pfu; intratumoral; day 7 and 14), irinotecan (25 mg/kg; intravenous; day 3, 10, and 17), or mSJ-815 (3×107 pfu; intratumoral; day 7 and 14)+irinotecan (25 mg/kg; intravenous; day 3, 10, and 17). The dashed lines indicate the normal range of body weight change. The p-value was obtained using a log-rank test.

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  Fig. 7. Ratio of SN-38/irinotecan in tumor tissue, obtained on day 7 from B16-F10 melanoma-bearing mice, treated with phosphate-buffered saline, irinotecan (25 mg/kg; intravenous; day 3), modified Western Reserve (WI; 1×107 pfu; intravenous; day 0)+irinotecan (25 mg/kg; intravenous; day 3), or SJ-815 (1×107 pfu; intravenous; day 0) +irinotecan (25 mg/kg; intravenous; day 3). The p-value was obtained using an ANOVA test.

Reference

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