Imaging of viral thymidine kinase gene expression by replicating oncolytic adenovirus and prediction of therapeutic efficacy

Abstract

Purpose: We have used a genetically attenuated adenoviral vector which expresses HSVtk to assess the possible additive role of suicidal gene therapy for enhanced oncolytic effect of the virus. Expression of TK was measured using a radiotracer-based molecular counting and imaging system. Materials and Methods: Replication-competent recombinant adenoviral vector (Ad-ΔE1B19/55) was used in this study, whereas replication-inompetent adenovirus (Ad-ΔE1A) was generated as a control. Both Ad-ΔE1B19/55-TK and Ad-ΔE1A-TK comprise the HSVtk gene inserted into the E3 region of the viruses. YCC-2 cells were infected with the viruses and incubated with 2′-deoxy-2′- fluoro-β-D-arabinofuranosyl-5-iodouracil (I-131 FLAU) to measure amount of radioactivity. The cytotoxicity of the viruses was determined, and gamma ray imaging of HSVtk gene was performed. MIT assay was also performed after GCV treatment. Results: On gamma counter-analyses, counts/minute (cpm)/μg of protein showed MOIs dependency with Δ E1B19/55-TK infection. On MIT assay, Ad-ΔE1B19/55-TK led to more efficient cell killing than Ad-ΔE1A-TK. On plate imaging by gamma camera, both Ad-ΔE1B19/ 55-TK and Ad-ΔE1A-TK infected cells showed increased I-131 FIAU uptake in a MOI dependent pattern, and with GCV treatment, cell viability of ΔE1B19/55-TK infection was remarkably reduced compared to that of Ad-ΔE1A-TK infection. Conclusion: Replicating Ad-ΔE1B19/55-TK showed more efficient TK expression even in the presence of higher-cancer cell killing effects compared to non-replicating Ad-ΔE1A-TK. Therefore, GCV treatment still possessed an additive role to oncolytic effect of Ad-ΔE1B19/ 55-TK. The expression of TK by oncolytic viruses could rapidly be screened using a radiotracer-based counting and imaging technique.