(222d) A Modular, Antibody Conjugate Dual-Delivery Strategy for Enhancing Endosomal Escape of Delivered siRNA

Title: A Modular, Antibody Conjugate Dual-Delivery
Strategy for Enhancing Endosomal Escape of Delivered siRNA

RNA
interference (RNAi) has emerged over the last 15 years as a specific and potent
means of targeted gene silencing at the mRNA level. Since its initial discovery
in C. elegans, researchers have sought
to harness this mechanism, by way of short interfering RNAs (siRNA), as an
efficient route to silencing disease-causing genes. However, inefficient
cytosolic localization of delivered siRNA continues to be a key challenge in
RNAi therapeutic development. The most widely used delivery strategy currently
involves cationic polymers or lipid nanoparticles to encapsulate the siRNA
cargo, deliver it to the cell, and facilitate endosomal release to the
cytoplasm. However, the design of a single polymer or lipid formulation that
can overcome the multiple biological barriers to delivery has proven difficult.

In
this talk, we introduce a modular dual-delivery strategy that decouples cellular
internalization from endosomal escape through the use of antibody conjugates. Cellular
internalization of these conjugates will be dictated solely by the antibody
while endosomal escape will be attained with an endosomal escape agent (EAA). Decoupling
internalization from escape will enable us to screen EEAs that disrupt
membranes for cytosolic translocation of siRNA independently of its endocytic
pathway. Two bioconjugates, an antibody-siRNA and antibody-EEA, will be
prepared using the same antibody to ensure compartmental co-localization within
the cell, then co-delivered to initiate RNAi.

Toward
this aim, I will introduce a novel method for the assembly of both
RNA-transporting and EEA-transporting antibody conjugates. This method enables efficient
and reagent free bioconjugate assembly at sub-micromolar concentrations,
typical of commercial antibodies, without significant disruption of antibody binding
affinity. The modularity of this system allows us to then explore the effect of
entry pathway on gene silencing by substituting the targeting antibody. Furthermore,
I will present in vitro data on novel
biomaterials with lytic properties for use as EEAs. This novel dual-delivery
approach will be used as a platform to elucidate structure-activity
relationships of potent EEAs and effects of entry pathway on the efficient
delivery of siRNAs.