This is a bioorthogonal reporter system that consists of a functionalized polyamidoamine (PAMAM) dendrimer capable of click chemistry used for monitoring of nanoparticle trafficking in complex biological systems. Dendrimers conjugated to bioactive molecules can be used for targeted delivery of therapeutic and imaging agents for many biological applications. Dendrimers and other nanoparticles can be directly labeled with fluorescent or radioactive probes to monitor their in vitro and in vivo trafficking and visualize targeted macromolecular structures. However, the use of such labels is limited by the inherent differences between labeled and unlabeled particles, and synthetic difficulties found in making these reporter molecules. Bioorthogonal chemical reporters that utilize click chemistry azide-alkyne cycloaddition reactions are known for their high efficiency, specificity, and small handle size, and have been used as reporters for DNA synthesis, proteomics, and metabolite profiling. Using dendrimers functionalized with click chemistry handles can provide important insights into nanoparticle trafficking and allow monitoring of targeted delivery of therapeutic agents.
Researchers at the University of Michigan Nanotechnology Institute for Medicine and Biological Sciences developed a method for preparing alkyne-conjugated PAMAM dendrimers and their use in biological applications. Using NMR and mass spectrometry, it was determined that the dendrimer scaffolds had ~12 alkyne handles on their surface. Folic acid and mannose were used as targeting moieties that exhibit high affinity for the folate receptor (FR) and the macrophage mannose receptor (MMR), respectively. The functionalized dendrimers were incubated with cells expressing the appropriate molecular target and then visualized using several azide-containing fluorophores. The signal-to-noise ratios were comparable to traditional fluorescent labeling techniques, and the same nanoparticle could be detected using several fluorophores. Subsequently, in vivo trafficking of folic acid targeted dendrimers was monitored in a murine model of inflammation. As expected, this study showed significant uptake of conjugated dendrimers by macrophages found in the mesenteric lymph nodes, the primary lymph organs draining the peritoneal cavity. These data support the use of dendrimers conjugated with click chemistry reporters in animal models to elucidate the relationship between nanoparticles and biological systems.