Active Uptake and Trafficking of Nucleoside Triphosphates In Vivo

Modified nucleoside triphosphates (NTPs) are powerful probes and medicines, but their anionic character impedes membrane permeability. As such, invasive delivery techniques, transport carriers, or prodrug strategies are required for their in vivo use. Here, we present a fluorescent 2'-deoxyribonucleoside triphosphate "TAMRA-dATP" that exhibits surprisingly high bioavailability in vivo. TAMRA-dATP spontaneously forms nanoparticles in Mg⁺²-containing buffers that are taken into the vesicles of living cells and animals by energy-dependent processes. In cell cultures, photochemical activation with yellow laser light (561 nm) facilitated endosomal escape of TAMRA-dATP, resulting in its metabolic incorporation into DNA in vitro. In contrast, in vivo studies revealed that TAMRA-dATP is extensively trafficked by active pathways into cellular DNA of zebrafish (Danio rerio) and Caenorhabditis elegans where DNA labeling was observed in live animals, even without photochemical release. Metabolic labeling of DNA in whole, living animals can therefore be achieved by simply soaking animals in a buffer containing TAMRA-dATP or a structurally related compound, Cy3-dATP.