Protein-engineered nanostructures for targeted delivery
Proteins can serve as building blocks for the fabrication of nanoparticles to deliver various drug cargoes or as the therapeutic itself. Despite great interest and investment, protein therapeutics are limited by delivery challenges such as short circulation times and low efficacy preventing their widespread use as therapeutics and delivery carriers for intracellular targets. Engineering efforts to address delivery limitations often rely on modifying proteins through direct conjugation of polymers and peptides using reactive residues on amino acids. The key shortcoming of this method is the inability to modify a specific site within a protein, which can significantly reduce pharmacological action. Additionally, such approaches do not offer control over variables such as ligand clustering, which is an important determinant of targeting efficacy. Ongoing work looks to use molecular engineering approaches for the design of tunable protein nanocarriers for optimal intracellular enzyme and gene delivery.
Extracellular matrix (ECM)-triggered delivery
We have capitalized on a class of peptides known as collagen-mimetic peptides, or CMPs, that have been recognized for their unique affinity for native collagen, and we have explored these CMPs for applications in substrate-mediated gene transfer. CMPs incorporate themselves into the natural collagen triple helical structure via strand invasion, in a reversible process previously that has been used to modify extracted collagen in vitro and exclusively target remodeling collagen in vivo. In our studies, we have designed CMPs to act as adjustable tethers as well as adhesive/endocytic ligands for polyplexes within collagen-based matrices. This CMP-based approach maintains polyplex activity over multiple weeks in serum, and the level of transgene expression can be directly correlated to local proteolytic activity and the extent of collagen remodeling, demonstrating “on demand” release. The ability to tailor release over extended periods via physical attachments, combined with the ability to provide cell-triggered release and collagen-mediated uptake, make this approach very attractive for wound repair and other applications in regenerative medicine.
Nuclear targeting in gene delivery
Histones have received great interest as potential gene carriers for several decades due to their seminal role in chromatin packaging and gene transfer, yet therapeutic efforts with histones have lacked both a well-controlled materials approach and a deeper knowledge of cellular processing mechanisms. Hence, histone-based carriers have failed to reach clinical efficacy. We have capitalized on newly recognized and highly pivotal roles for histone tails in native gene regulatory control to develop a gene transfer method. In advancing our expertise in nanoparticle formulation, we are integrating our previously recognized histone tails peptide with lipid nanoparticles to facilitate highly effective neonatal lung delivery. Our endeavors have demonstrated a significant enhancement in transfection efficiency through the incorporation of this novel peptide. Currently, we are dedicated to comprehending and applying this advantageous mechanism in the design of our future nanoparticles.
Biomaterials and and the Microbiome
We are focussing on the design of microscale biomaterials, or ‘microgels,’ for the controlled delivery of gene therapies, protein therapies, and/or therapeutics for microbiome manipulation in healing tissues such as chronic wounds. Focus areas include the design of the microgel biomaterial matrix to enable tunable encapsulation and release of various cargoes with control over duration and dosing; surface modification strategies to enable decoration of the microgel surface with therapeutics, and/or to alter the adhesive properties of the microgels; and evaluation of the controlled release properties of the microgels in the presence of mammalian cells, mammalian tissue components, and/or microbes that mimic those found in the human wound bed.
DNA-gated drug delivery
We are developing strand displacement-inspired DNA biosensing designs coupled into polymeric and protein-based nanocarriers to effect the sequence- and concentration-dependent release of imaging and/or therapeutic agents in response to a versatile array of intracellular RNA structures. These approaches are ultimately aimed at the development of highly tailored DNA-polymer nanotheranostics, and should enable the elucidation of detailed structure/property/function relationships underlying recognition of composite and individualized signals in healthy vs. diseased tissues.
Photo-patterned RNA transfer
Precise and dynamic strategies to modulate gene expression can illuminate fundamental cell growth and repair processes and would have enormous benefits in a wide range of tissue engineering as well as developmental biology and drug screening applications. To meet this need, we have developed versatile and highly tunable light-based strategies to deploy nucleic acids with control at cellular length scales. In particular, we have demonstrated a new class of light-sensitive polymers that provide biocompatibility and rapid application with proven on/off initiation of gene silencing and the ability to locally “dial-in” the level of siRNA deployment and gene regulation. These simple polymer/siRNA formulations provide unusually robust siRNA binding, yet siRNA release and efficient gene silencing are easily triggered by application of cytocompatible doses of light. The ability to control the spatial placement and induction of biomolecular signaling programs through this on/off switch will be essential in developing and modeling complex tissues such as cardiovascular and neural networks.
