Researchers have recently shown that delivery of the zinc chelating compound TPEN can alleviate the adverse effects of axon injury however there is effective way to currently deliver TPEN for therapeutic applications. Here we report a fluorescence-based DNAzyme assay for monitoring the release and chelation of zinc ions by TPEN from a nanomaterial delivery system in an effort to more effectively delivery TPEN to diseased cells. To this end, TPEN encapsulated nucleic acid nanocapsule (NANs) functionalized with a zinc specific DNAzyme were synthesized. The NANs have the potential to enable cellular uptake without using transfecting agents and can enzymatically trigger the release of TPEN cargo for chelating zinc ions in living cells. The zinc sensing DNAzyme is prepared with a complementary substrate strand modified with a fluorophore and a quencher. In the presence of zinc ions, the DNAzyme is activated and cleaves the substrate producing fluorescence. The extent of cleavage and the intensity of fluorescence produced acts as a measure of zinc concentration. We show here that the reduction in fluorescence caused by payload release can be used to quantify the amount of zinc chelated by TPEN and indicates the successful enzyme-dependent release of TPEN from the NANs. Release of TPEN alongside therapeutic nucleic acids has previously been shown to result in synergistic gene regulation in injured axons. Future work will center on using the TPEN-loaded NANs to co-deliver TPEN alongside a gene regulating siRNA to achieve maximal axon regeneration.

Towards intracellular zinc quantification and chelation by targeted release of N, N, N′, N′-tetrakis (2-pyridinylmethyl)-1,2-ethanediamine (TPEN)

ACS Spring Meeting 2021

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