Recombinant nanocapsid for targeted theranostic delivery

Abstract

Developments in diagnostic and therapeutic delivery are trending towards molecular level targeting with nano-platforms. Targeted delivery reduces generalized distribution by localizing diagnostic and/or therapeutic (theranostic) molecules to an intended target site. The first section of this thesis proposes Hepatitis E Virus (HEV) nanocapsids as a vector to stabilize and target theranostic delivery. Derived from the capsid protein of HEV, a feco-orally transmitted virus, HEV-like particles self-assemble in to non-infectious, nanocapsids that can withstand harsh protease and pH conditions in the mucosal system. The flexible nanocapsid surface protrusion domain is amenable to substantial modification. Chemical modulation of nanocapsids was achieved through surface conjugation to single solvent exposed cysteine sites. In this thesis, nanocapsids chemically modulated with tumor-targeting ligands exhibit cancer cell-specific binding and internalization, as well as in vivo tumor detection. We also used cysteine sites to conjugate thiolate-protected gold nanoclusters (AuNCs), which have molecule like qualities distinct from colloidal gold nanoparticles (AuNPs). Specifically, Au102(pMBA)44 (Au102) and maleimide-linked Au102 (Au102_C6MI) were conjugated to nanocapsid cysteine through place exchange and maleimide-thiol coupling, respectively. Au102_C6MI-bound nanocapsids were imaged in cryoEM and a 3D structure was determined. The resolved structure of AuNC-bound nanocapsid revealed a 5-fold ring density attributable to AuNC densities. Rigid modelling supported this finding. In the last section of this thesis, a recombinant enterovirus was engineered to distinguish the role of structural vs. non-structural proteins in Enterovirus B infection kinetics, replication and infection persistency. The results indicated cell-receptor binding likely triggered lytic vs. non-lytic infection, providing insight in to adaptive mechanisms of native virus cell delivery.