Engineering BV to efficiently deliver large genetic payloads

Summary: Efficient in vivo delivery of large DNA constructs encoding RNA and proteins will provide a powerful
tool for a wide range of applications in health, including the use of genome engineering for understanding and
controlling biological functions, and gene therapies for treating human disease. Existing in vivo delivery vehicles,
such as adeno-associated airal vectors (AAVs) and lipid nanoparticles (LNPs) lack the ability to deliver large
DNA constructs in vivo. Baculoviral (BV) vectors offer a potential solution to this unmet need. Derived from an
insect virus, BV vectors have an extraordinary DNA payload packing capacity (up to 300 kb), can infect both
dividing and nondividing cells, only replicate in insect cells thus are considered safe and nonpathogenic to
humans. However, there are major challenges for BV to become a vehicle for in vivo gene delivery. BVs delivered
systemically in vivo are often inactivated by the innate complement system, hindering their ability to transduce
cells efficiently in vivo. BVs also have weak interactions with mammalian cell surface proteins, and those
internalized can be trapped in the endosomes, further reducing transduction efficiency. This proposed study aims
to engineer BV vectors for efficient in vivo delivery of large DNA constructs, by screening diverse BV surface-
displayed factors including cell adhesion, endosomal escape and complement protection factors. We
hypothesize that, thorough the expression of an optimal set of BV surface-displayed factors, BVs can be
programmed to deliver large therapeutic DNA payloads in vivo with high efficiency. This hypothesis is based on
our preliminary data demonstrating that in vivo delivery efficiency of engineered BV vectors can be synergistically
enhanced by co-expression of endosomal escape and complement protection factors. In Aim 1 studies we will
carry out high-throughput screening to identify combinations of cell adhesion and endosomal escape factors that
improve the transduction efficiency of BVs. In Aim 2 studies we will identify complement protection factors and
combine them with the optimal cell adhesion and endosomal escape factors, and determine if the in vivo delivery
efficiency of large therapeutic DNA cargos can be enhanced significantly. Successfully completion of this work
will result in gene delivery vectors capable of efficiently expressing large DNA payloads in vivo, enabling safter
and more effective therapeutic strategies for a wider array of diseases.

Grant Number: 1R21EB037939-01
NIH Institute/Center: NIH
Principal Investigator: Caleb Bashor