Preclinical safety evaluation of hepatic arterial infusion of oncolytic poxvirus

Abstract

Purpose: Oncolytic poxvirus has shown promise in treating various solid tumors, such as liver cancer, and administration of oncolytic poxvirus via the hepatic artery may provide more survival benefits than other routes of administration. However, there is a lack of safety information to guide the application of hepatic arterial infusion (HAI) of oncolytic poxvirus in human studies. To investigate the acute and chronic toxicity of HAI administration of oncolytic poxvirus in animals and provide safety information for future human studies.

Methods: VV^tk-, a vaccinia poxvirus with inactivated thymidine kinase gene, was administered via HAI to rabbits with normal liver function under angiography (1×10⁸ or 1×10⁹ pfu), and rats with N-nitrosomorpholine-induced precancerous liver cirrhosis under open surgery (1×10⁸ pfu). Body weights and survival were monitored and blood samples were collected for hematological and biochemical tests. Distribution of A56 (a specific marker for poxvirus infection) in rabbit organs was evaluated using immunofluorescence assays.

Results: HAI of high doses of VV^tk- did not cause any acute or chronic changes in body weight, survival or in biochemical, hematological tests in the 2 animal models, and none of the changes showed dose dependency (in rabbit study), or were influenced by liver cirrhosis (in rat study). A56 was not detected in any of the major rabbit organs.

Conclusion: HAI may provide a safe alternative route of oncolytic poxvirus administration for human studies.

Keywords: hepatocellular carcinoma; liver cirrhosis; oncolytic virus; transhepatic angiography.

Figure 1

Selective hepatic artery angiography in a rabbit. Abbreviation: LHA, left hepatic artery.

Figure 2

Body weight of 8 rabbits with normal liver function, measured at baseline and post hepatic artery infusion (week) of a single low (1×10⁸ pfu; n=4) and a high (1×10⁹ pfu; n=4) dose of VV^tk−.

Figure 3

Organ functions of 8 rabbits with normal liver function, measured at baseline and post hepatic arterial infusion (week) of a single low (1×10⁸ pfu; n=4) and high (1×10⁹ pfu; n=4) dose of VV^tk−. Note: Distances between 2 green dash lines are reference ranges. Abbreviations: ALT, alanine transaminase; AST, aspartate aminotransferase; BUN, blood urea nitrogen; GGT, gamma-glutamyl transpeptidase; GOT, glutamic-oxalacetic transaminase; GPT, glutamic-pyruvic transaminase.

Figure 4

Hematological tests of 8 rabbits with normal liver function, measured at baseline and post hepatic artery infusion (week) of a single low (1×10⁸ pfu; n=4) and a high (1×10⁹ pfu; n=4) dose of VV^tk−. Note: Distances between 2 green dash lines are reference ranges.

Figure 5

Immunofluorescence assay detected no A56 expression in multiple organs of rabbits with normal liver function, after hepatic artery infusion of a single dose (1×10⁹ pfu) of VV^tk−. Notes: The expression of A56 in tumor tissue and other organs from humanized tumor mouse models (HT-29 and HCT-116) serves as positive and negative controls. Green: A56; blue: nuclei.

Figure 6

Study design (upper) and study results (lower) of the rat study. Masson Trichrome histological examination was conducted in liver tissue obtained from 2 rats (at week 0) that finished the NNM pretreatment, to confirm the formation of liver cirrhosis. Survival and body weight were monitored between week 0 and 26. Survival was not significantly different (P=0.388) between the 2 groups, P-value was obtained using log-rank test, and body weight increase (% of baseline) was significantly higher in the VV^tk− group than the PBS one, P-value was obtained using linear mixed model analysis (P<0.001). Abbreviation: NNM, N-nitrosomorpholine.

Figure 7

ALT measured before and after the second HAI (1×10⁸ pfu VV^tk− or PBS) in NNM-induced liver cirrhotic rats. Note: Comparison between the 2 groups was conducted using linear mix model analysis. Abbreviations: ALT, alanine transaminase; HAI, hepatic arterial infusion; NNM, N-nitrosomorpholin.

Figure 8

Liver tissue obtained from (A) rats without any treatment, (B) rats with NNM-induced precancerous liver cirrhosis after the second HAI of 1×10⁸ pfu VV^tk−, and (C) rats with NNM-induced precancerous liver cirrhosis after the second HAI of PBS. Abbreviations: HAI, hepatic arterial infusion; NNM, N-nitrosomorpholin.