Systemic cancer therapy with a tumor-selective vaccinia virus mutant lacking thymidine kinase and vaccinia growth factor genes

• Published in: Cancer research (Chicago, Ill.) Ill.), 2001-12, Vol.61 (24), p.8751-8757
• Main Authors: MCCART, J. Andrea, WARD, Jerrold M, LEE, John, YUN HU, ALEXANDER, H. Richard, LIBUTTI, Steven K, MOSS, Bernard, BARTLETT, David L
• Format: Article
• Language: English
• Subjects: 3T3 Cells; Anesthesia. Intensive care medicine. Transfusions. Cell therapy and gene therapy; Animals; Applied cell therapy and gene therapy; Biological and medical sciences; Cytopathogenic Effect, Viral; Female; Gene Deletion; Genetic Therapy - methods; Genetic Vectors - genetics; Green Fluorescent Proteins; Haplorhini; HeLa Cells; Humans; Luminescent Proteins - biosynthesis; Luminescent Proteins - genetics; Medical sciences; Mice; Mice, Inbred C57BL; Mice, Nude; Peptides - genetics; Poxviridae Infections - virology; Thymidine Kinase - genetics; Transfusions. Complications. Transfusion reactions. Cell and gene therapy; Tumor Cells, Cultured; Vaccinia virus - enzymology; Vaccinia virus - genetics; Vaccinia virus - pathogenicity; Vaccinia virus - physiology; Virus Replication
• ISSN: 0008-5472; 1538-7445

We have demonstrated previously the oncolytic effects of a systemically delivered, replicating vaccinia virus. To enhance the tumor specificity of this vector, we have developed a combined thymidine kinase-deleted (TK-) and vaccinia growth factor-deleted (VGF-) vaccinia virus and investigated its properties in vitro and in vivo. The gene for enhanced green fluorescent protein (EGFP) was inserted into the TK locus of a VGF- vaccinia virus by homologous recombination creating a double-deleted mutant vaccinia virus (vvDD-GFP). Infection of resting and dividing NIH3T3 cells with vvDD-GFP yielded reduced viral recovery compared with wild-type (WT), TK-, or VGF- viruses from resting cultures but equivalent virus recovery from dividing cultures. Eight days after nude mice were injected i.p. with 10(7) plaque-forming units (pfu) of WT, TK-, VGF-, or vvDD-GFP vaccinia virus, tissues and tumor were harvested for viral titer determination. No virus was recovered from the brains of mice injected with vvDD-GFP compared with the other viruses, which ranged from 130 to 28,000 pfu/mg protein; however, equivalent amounts were recovered from tumor. There was no toxicity from vvDD-GFP because nude mice receiving 10(8) pfu of IP vvDD-GFP lived >100 days, whereas mice receiving WT, VGF-, or TK- virus had median survivals of only 6, 17, and 29 days, respectively. Similar results were seen when 10(9) pfu of vvDD-GFP were given. Nude mice bearing s.c. murine colon adenocarcinoma (MC38) had significant tumor regression after treatment with 10(9) pfu of systemic (i.p.) vvDD-GFP compared with control (mean tumor size, 180.71 +/- 35.26 mm(3) versus 2796.79 +/- 573.20 mm(3) 12 days after injection of virus). Our data demonstrate that a TK- and VGF- mutant vaccinia virus is significantly attenuated in resting cells in vitro and demonstrates tumor-specific replication in vivo. It is a promising vector for use in tumor-directed gene therapy, given its enhanced safety profile, tumor selectivity, and the oncolytic effects after systemic delivery.