The rapidly expanding field of therapeutic nucleic acids (TNAs) has led to an increased urgency in the development of chemical approaches that can broaden their clinical application. In just the last 3 years the US Food and Drug Administration has approved two TNA based drugs that treat diseases that previously had little option for treatment. Despite these recent achievements, many clinical trials involving TNAs result in termination due to limited efficacy, a result that is often attributed to poor delivery owing to the highly polar and charged character of TNAs that limits accessibility to mRNA in the cytosol of cells. Many platforms deliver nucleic acids using nanoparticle-based approaches that are internalized into cells through endocytosis mechanisms. Therefore, a central bottleneck to their effective delivery is escaping endosomal compartments and gaining access to the cytosol. The central vision of my research program is to address these challenges associated with endosomal escape and TNA delivery to the cytosol at the molecular level.
We aim to address these challenges through the synthesis of nucleic acid surfactant conjugates that we have recently shown can successfully target mRNA for gene silencing in vitro. As their design is chemically tunable, we aim to systematically assess the role of the surfactant as it relates to the extent of endosomal escape by evaluating the importance of hydrophobic character, net charge and size of the conjugates. We will also develop a new class of fluorescent surfactant probes which can be used to monitored the delivery and stability of the surfactant conjugates as they are trafficked through cells, thereby helping us to quantify the effects changes in the chemical character of the conjugates have on cytosolic delivery. In conjunction with these studies we will determine the net effect of chemically modifying the nucleic acid surfactant conjugates on the efficacy of gene knockdown in vitro using DNAzymes that result in mRNA cleavage. By controlling the molecular design of individual DNAzyme-surfactant conjugates we can better understand their mechanism of cellular entry and the properties that lead to cytosolic access. Through successful realization of our program we will not only contribute to our understanding of the properties that are necessary for nucleic acids to successfully gain access to the cytosol of cells but also design a platform that will have immediate therapeutic value suitable for future translational applications.
Therapeutic nucleic acids have recently gained FDA approval for their ability to silence genes and correct mRNA splicing events, showing their promise for treating traditionally difficult to target disease pathways. Despite their great potential for clinical applications, efforts to develop broadly applicable strategies for their cellular delivery have been limited due to the charged nature of nucleic acids that limits both their cellular uptake and accessibility to their mRNA targets. Here we propose the synthesis of a new class of chemically tunable nucleic acid surfactant conjugates as an innovative way to improve nucleic acid cytosolic delivery along with novel fluorescent surfactant probes that can help us quantify their delivery to enable this promising class of biomolecules to achieve greater clinical application.
