Platforms for delivery of macromolecules to sites of DNA double-strand break repair.

ABSTRACT

Double-strand break (DSB) repair foci are important therapeutic targets. Here we describe platforms for delivery of macromolecules, nanomaterials and nanomedicines to repair foci. The strategy is based on the high affinity of the human 53BP1 protein for modified chromatin present at sites of DNA damage. As proof of concept, we created, expressed, and purified an engineered fragment of 53BP1 and coupled it to fluorescent streptavidin, a model cargo with no intrinsic affinity for repair foci. This binary complex was in turn coupled to the iron carrier protein, transferrin, which engages a high-affinity cell surface receptor. In a different version of the complex, transferrin was omitted and a protein transduction domain was incorporated directly into the primary structure of the 53BP1. These complexes were efficiently taken up into human osteosarcoma cells and synchronously released from endocytic vesicles by brief exposure to far-red light in the presence of the photosensitizer, disulfonated aluminum phthalocyanine. Upon release, the streptavidin cargo entered the nucleus and was recruited to repair foci. 53BP1-based platforms provide a method for targeted, temporally controlled delivery of macromolecular agents to sites of double-strand break repair. With the delivery platforms, we are capable to visualize, modify and redirect DSB repair pathways by coupling various nanomaterials to study machinery or manipulate for therapy purpose in the future.