Capsid-Labeled Oncolytic Adenovirus, Targeted to Tumor-Specific Marker Her2/neu

Oncolytic adenoviruses (Ads) hold enormous potential for treatment of metastatic cancers as novel targeted therapeutics. To date specificity of tumor targeting, crucial for achieving clinical efficacy of oncolytic Ads upon their systemic delivery, has been a problem due to technical limitations of the virus tropism modification technology. Furthermore, clinical improvement of oncolytic Ads has also been limited by the lack of adequate non-invasive virus imaging strategies. Objective: To designed a new Ad with tumor-specific replication and transduction potential and fluorescent properties, allowing direct visualization of viral particles.Methods: Using polymerase chain reaction (PCR) and DNA homologous recombination in bacteria we replaced the Ad fiber protein carboxy-terminal “knob” domain with a fibritin-affibody fusion, possessing engineered binding specificity for the Her2/neu receptor. To allow tracking of viral particles by non-invasive spectral imaging, we also incorporated the fluorescent reporter mCherry in the Ad capsid by its genetic fusion to the minor capsid protein pIX. Results: The new virus was successfully rescued and propagated in Her2/neu-positive target DY36T2 cancer cells and its genetic identity verified at the molecular level. Efficient incorporation of mCherry in the capsid was evidenced by strong fluorescence of the purified viral particles as well as the target cells following Her2/neu-mediated binding of the virus at 4^oC. The fast growth kinetics along with efficient and specific killing of cancer cells in vitro, observed for the new Ad, demonstrate that its biological characteristics are not adversely affected by the genetic modifications. Conclusions: In vitro validation of the new oncolytic Ad suggests its utility for tumor targeting and justifies future experiments in various animal models of cancer. Its potential utility for systemic application opens a unique possibility for diagnostics and virotherapy of human oral, head and neck, breast and other disseminated forms of human cancer. Support: T35-HL007473 & UAB SOD Faculty Research Award/IOHR.