Therapeutic Targeting of Chitosan-PEG-Folate-Complexed Oncolytic Adenovirus for Active and Systemic Cancer Gene Therapy

Abstract

Adenovirus (Ad)-based cancer therapies have shown much promise. However, until now, Ad has only been delivered directly to primary tumors because the therapeutic efficacy of systemic delivery is limited by the immune response of the host, short blood circulation times, and non-specific liver uptake of Ad. In order to circumvent the issues regarding systemic delivery and to increase the safety and efficacy of Ad therapies, the surface of Hmt oncolytic Ad was coated with cationic polymer chitosan via ionic crosslinking (Hmt/chitosan), after which polyethylene glycol (PEG) and/or folate (FA) was chemically conjugated onto the surface of Hmt/chitosan, generating Hmt/chitosan-FA, Hmt/chitosan-PEG, and Hmt/chitosan-PEG-FA nanocomplex. The FA-coordinated Hmt nanocomplexes (Hmt/chitosan-FA & Hmt/chitosan-PEG-FA) elicited FR-selective cancer cell killing efficacy. In vivo administration of Hmt/chitosan-PEG or Hmt/chitosan-PEG-FA into mice demonstrated that PEGylation greatly increased blood circulation time, resulting in 9.0-fold and 48.9-fold increase at 24 hrs after injection compared with naked Hmt, respectively. In addition, generation of Ad-specific neutralizing antibodies in mice treated with Hmt/chitosan-PEG-FA was markedly decreased by 75.3% compared with naked Ad. The Quantitative PCR assay results showed 285.0-fold increase in tumor tissues and 378-fold reduction of Hmt/chitosan-PEG-FA in liver tissues compared with naked Hmt. Bioluminescence imaging study further supported the enhanced tumor-to-liver ratio of Hmt/chitosan-PEG-FA. Consequently, systemic delivery of Hmt/chitosan-PEG-FA significantly inhibited the growth of FR-positive tumor, decreasing 52.8% compared to the Hmt-treated group. Importantly, PEGylated oncolytic Ad nanocomplexes showed no elevation of both ALT and AST levels, demonstrating that systemically delivered Ad-related hepatic damage can be completely eliminated with PEG conjugation. In sum, these results demonstrate that conjugation of chitosan-PEG-FA to oncolytic Ad significantly improves antitumor efficacy and safety profiles, suggesting that Hmt/chitosan-PEG-FA has potential as a therapeutic agent to target FR-positive cancer via systemic administration.