Labeling of Adenovirus Particles with PARACEST Agents

Abstract

Recombinant adenovirus type 5 particles (AdCMVLuc) were labeled with two different bifunctional ligands capable of forming stable complexes with paramagnetic lanthanide ions. The number of covalently attached ligands varied between 630 and 1960 per adenovirus particle depending upon the chemical reactivity of the bifunctional ligand (NHS ester versus isothiocyanide), the amount of excess ligand added, and the reaction time. The bioactivity of each labeled adenovirus derivative, as measured by the ability of the virus to infect cells and express luciferase, was shown to be highly dependent upon the number of covalently attached ligands. This indicates that certain amino groups, likely on the surface of the adenovirus fiber protein where cell binding is known to occur, are critical for viral attachment and infection. Addition of 177 Lu 3+ to chemically modified versus control viruses demonstrated a significant amount of nonspecific binding of 177 Lu 3+ to the virus particles that could not be sequestered by addition of excess DTPA. Thus, it became necessary to implement a prelabeling strategy for conjugation of preformed lanthanide ligand chelates to adenovirus particles. Using preformed Tm 3+ -L2, a large number of chelates having chemical exchange saturation transfer (CEST) properties were attached to the surface residues of AdCMVLuc without nonspecific binding of metal ions elsewhere on the virus particle. The potential of such conjugates to act as PARACEST imaging agents was tested using an on-resonance WALTZ sequence for CEST activation. A 12% decrease in bulk water signal intensity was observed relative to controls. This demonstrates that viral particles labeled with PARACEST-type imaging agents can potentially serve as targeted agents for molecular imaging.

Macromolecules, Dendrimers, and Nanomaterials in Magnetic Resonance Imaging: The Interplay between Size, Function, and Pharmacokinetics

“…The externally modified virion was measured to have an ion relaxivity of 30.7 mM −1 s −1 , while that of the internally modified virion was 41.6 mM −1 s −1 , which are among the highest values reported so far for viral-based macromolecular contrast agents. Finally, Vasalatiy and co-workers reported the labeling of adenovirus particles (AdCMVLuc) with Tm 3+ and 177 Lu 3+ chelates of 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetra(methylcarbonylamide) (DTMA), the former a PARACEST agent and the latter a radioactive reporter, while retaining the capability of the virus to infect cells and express luciferase 368. Though between 630 and 1960 ligands could be loaded onto the adenovirus, a limit of ~800 attached ligands was determined to maintain a 75–80% bioactivity.…”

Dendrimer-based magnetic resonance imaging agents for brain cancer

“…The incorporation of metal ions results in paramagnetic CEST (PARACEST) agents, which have chemical shifts that are further away from water [32]. PARACEST agents have also been covalently linked to nanocarriers, including linear polymers, dendrimers and other macromolecules [133][134][135][136], to further improve the sensitivity of detection in vivo. For example, Ali et al [137] conjugated the PARACEST agent Eu-DOTA-Gly 4 to a PAMAM dendrimer of generation 5 (Fig.…”

Suppression of Nicotine-Induced Pathophysiology by an Adenovirus Hexon-Based Antinicotine Vaccine

“…Through conjugation to free lysines in the Ad capsid proteins, molecules such as polymers, lipids, biotin, fluorophores, and metal nanoparticles have been covalently linked to Adhexons (Kramp et al, 1979;Singh and Kostarelos, 2009). Trials have used Ad viral capsid proteins to deliver macromolecules in therapeutic applications, such as MRI contrast agents, radiation sensitizers, and antigenic peptides (HIVTat) for vaccine development (Liepold et al, 2007;Vasalatiy et al, 2008;Yoshioka et al, 2008;Singh and Kostarelos, 2009). The high seropositivity to several strains of Ad in the human population, including Ad 2 and Ad5, has limited the success of Ad-based vaccine trials, such as HIV STEP clinical trial (Matthews et al, 2010).…”