Thursday, November 21, 2024
11/21/2024
PROJECT ABSTRACT/SUMMARY DNA nanotechnology offers near-atomic control for building structures, with precise positioning of guest molecules such as antibodies, fluorophores and ligands that make them potentially useful in a number of biological applications such as biosensing, drug delivery, cell modulation and bioimaging. However, there are still many challenges that need to be addressed for DNA nanotechnology to reach its full potential for practical applications. In this proposal, we focus on two main areas of development in DNA nanotechnology to address these challenges: (1) Creating a robust, multifunctional drug delivery platform for treating multisystemic diseases, and (2) designing 3D DNA crystals as scaffolds for X-ray structure determination and characterization of such 3D lattices using the new technique of Serial Femtosecond X-ray Crystallography (SFX). For drug delivery, we will develop DNA polyhedra as drug carriers for delivering a new class of modified polycyclic compounds (MPCs) to multiple organ systems and enhancing drug candidate screening using myotonic dystrophy type 1 (DM1) as a testbed disease. Our work will provide quantifiable loading of these minor groove binding drugs and thorough validation of drug delivery efficiency from cell culture to preclinical DM1 mouse models, establishing cell internalization, lack of toxicity and immune response, cell- and disease-specific targeting, bioavailability and biodistribution of the drug-loaded DNA nanostructures. For developing DNA nanostructures as structural scaffolds, we will design and construct DNA motifs that assemble into 3D DNA crystals with different cavity sizes that allow hosting guests of different sizes ranging from nanoparticles to proteins. We will improve resolution of the crystals by programming crystal contacts and incorporating chemical modifications and demonstrate macromolecular scaffolding of proteins using triplex forming oligonucleotides (TFOs) as tethers and peptides using PNA linkers. We will develop methods to grow microcrystals of these DNA motifs for structural analysis using SFX, where diffraction data is collected using high intensity X-ray free-electron lasers, that eliminate the need for large single crystals, freezing, and radiation damage associated with traditional crystallography. This proposed research extends beyond a single disease or health issue, making this work well-suited for the R35 Maximizing Investigators’ Research Award (MIRA) at the NIGMS. In the long-term, I envision that our modular, platform technology using DNA nanostructures can be adopted by other labs for different disease treatments and drug screening (drug delivery) and to obtain crystallographic information of hard-to-crystallize molecules (macromolecular scaffolds).
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