Project Summary/Abstract Oligonucleotides face several biopharmaceutical difficulties, including stability and delivery issues as well as non-hybridization activities such as coagulopathy and unwanted activation of the immune system. We have developed a unique oligonucleotide delivery system, termed pacDNA, which uses a high-density bottlebrush polymer to provide oligonucleotides with binding selectivity. The polymer amounts to an entropic barrier, reducing access to the oligonucleotide by various proteins (and thus side effects) but still allows for unhindered hybridization. This novel strategy not only improves nuclease stability, preserves target-binding capability, and minimizes off-target side effects, but also massively enhances plasma pharmacokinetics, tissue retention, and antisense potency in vivo. Our current studies also reveal that the pacDNA’s pharmacological properties are intimately related to the bottlebrush backbone. In addition, the pacDNA appears to be uniquely capable of evading anti-carrier adaptive immunity, which is useful for therapies that requires long-term/frequent dosing. Finally, the pacDNA deposits into tissues and organs that lack mature delivery technologies for, such as the skin, the skeletal muscle, and the heart. These surprising and enabling discoveries will be the basis for investigations in the next funding period, in which we will 1) explore the property space of the pacDNA structure using a combinatorial polymer library with specific backbone compositions and monomer sequences, and 2) probe in vivo properties of the pacDNA in mice. We anticipate that accomplishment of these objectives will yield significant fundamental understanding of this class of materials and bring us much closer to clinical evaluation of pacDNA.