Atomic-Layer Deposition Processes Applied to Phage λ and a Phage-like Particle Platform Yield Thermostable, Single-Shot Vaccines

Abstract

Especially in developing countries, the impact of vaccines can be limited by logistical obstacles associated with multiple dose regimens, pathogen variants, and challenges imposed by requirements for maintaining vaccines at low temperatures during shipping and storage. Thus, there is a need for vaccines that can be flexibly modified to address evolving pathogen landscapes, are stable outside of narrow "cold-chain" temperatures and require administration of only single doses. Here we demonstrate in proof-of-concept studies a vaccine platform that addresses these impediments to more widespread use of vaccines. The platform relies on bacteriophage-derived phage-like-particles (PLPs) that utilize a "plug-and-play" antigen delivery system that allows for fast, easy alteration of antigens on the surface of the PLPs. Thermostability of PLP-based vaccines can be achieved by embedding the PLPs within glassy particles produced by spray drying, and nanoscopic aluminum oxide layers applied using atomic layer deposition (ALD) can serve to control release of antigen in vivo, yielding vaccine formulations that elicit strong immune responses after administration of single doses. Bacteriophage λ was stabilized by spray drying to form powders that were incubated at 37 °C for up to a year without loss of infectious activity. PLPs derived from bacteriophage λ were expressed and purified from E. coli cultures, and an in vitro conjugation strategy was used to decorate specific PLP surface sites with T4-lysozyme, a model vaccine antigen. The resulting T4-lysozyme:PLP complexes (Lys-PLPs) were embedded in glassy dry powders formed by spray drying and coated with nanometer-thick layers of alumina deposited by ALD in a fluidized bed reactor. Alumina-coated Lys-PLP vaccines were stable for a least a month at 50 °C, and single doses of the alumina-coated vaccines elicited immune responses that were indistinguishable from responses generated by conventional two-dose, prime-and-boost dosing regimens of alum-adjuvanted Lys-PLP vaccines.

A Designer Nanoparticle Platform for Controlled Intracellular Delivery of Bioactive Macromolecules: Inhibition of Ubiquitin-Specific Protease 7 in Breast Cancer Cells

“…Herein, we demonstrate that λ PLPs (i) can be simultaneously decorated with trastuzumab and UbV.7.2 in rigorously defined surface densities, (ii) that the preparations are structurally and physically stable, and (iii) that the particles are homogeneous and amenable for pharmaceutical applications. Within this context, we recently demonstrated that antigen-decorated PLPs can be formulated in glassy powders that are stable and retain antigenicity for over 1 month at 50 °C, which is an important feature for a viable pharmaceutical preparation …”

“…Within this context, we recently demonstrated that antigen-decorated PLPs can be formulated in glassy powders that are stable and retain antigenicity for over 1 month at 50 °C, which is an important feature for a viable pharmaceutical preparation. 14 …”

“…Within this context, we recently demonstrated that antigen-decorated PLPs can be formulated in glassy powders that are stable and retain antigenicity for over 1 month at 50 °C, which is an important feature for a viable pharmaceutical preparation. 14 The Trz-decorated particles specifically target and are internalized by HER2-overexpressing cells, thus delivering the UbV biologic to the cell interior. While our data do not permit a mechanistic description of internalization and cell trafficking of the particles, it appears that they are taken into an endosomal compartment and slowly released.…”

“…A number of bacteriophage systems have been adapted as biologic platforms, each possessing strengths and weaknesses. − We have developed a semi-synthetic “designer nanoparticle” system based on the bacteriophage λ capsid, − which offers particular advantages in particle design. − Briefly, when co-expressed in Escherichia coli, the λ major capsid and scaffolding proteins self-assemble into native icosahedral shells. These phage-like particles (PLPs) differ from bone fide capsids in that they do not contain a portal complex.…”

Phage-like particle vaccines are highly immunogenic and protect against pathogenic coronavirus infection and disease

“…Importantly, the decorated particles are monodisperse, stable, and possess physiochemical properties amenable for pharmaceutical formulation and as required to address regulatory hurdles 53,63,64 . Indeed, we have recently demonstrated that antigen-decorated PLPs formulated in glassy dry powders are stable at 50 ˚C for more than one month 65 . Therefore, the lambda system described here allows for substantial flexibility and rigorous control in vaccine design and development, setting the stage for the construction of an "all-inone" vaccine platform.…”