Mimicking live flavivirus immunization with a noninfectious RNA vaccine

Abstract

Flaviviruses are human pathogens of world-wide medical importance. They have recently received much additional attention because of their spread to new regions (such as West Nile virus to North America), highlighting their potential as newly emerging disease agents. Using tick-borne encephalitis virus, we have developed and evaluated in mice a new genetic vaccine based on self-replicating but noninfectious RNA. This RNA contains all of the necessary genetic information for establishing its replication machinery in the host cell, thus mimicking a natural infection. However, genetic modifications in the region encoding the capsid protein simultaneously prevent the assembly of infectious virus particles and promote the secretion of noninfectious subviral particles that elicit neutralizing antibodies. These characteristics demonstrate that a new generation of flavivirus vaccines can be designed that stimulate the same spectrum of innate and specific immune responses as a live vaccine but have the safety features of an inactivated vaccine.

Nonviral delivery of self-amplifying RNA vaccines

“…Hence, mRNA vaccines have been formulated with synthetic delivery vehicles such as liposomes (16) and cationic polymers (17) to increase potency. There are limited published data on the in vivo delivery of self-amplifying RNA using nonviral delivery strategies (3,14,18), and none has taken advantage of the recently developed, clinically suitable delivery systems for siRNA (19,20). There has been extensive work on viral delivery of self-amplifying RNA using viral replicon particles (VRPs) (4,11,(21)(22)(23)(24).…”

“…Although plasmid DNA (pDNA) vaccines have proven to be a flexible platform and are broadly effective in small animal models, they have generally lacked potency in human clinical trials (2). Recombinant viral vector technologies have the advantage of efficient delivery of the nucleic acid payload, but their utility is often hampered by antivector immunity, production limitations, and safety concerns (3,4). In 1990, Wolff et al (5) demonstrated that direct injection ("naked delivery") of messenger RNA (mRNA) or pDNA into the skeletal muscle of a mouse resulted in expression of the encoded protein.…”

“…After immunization, replication and amplification of the RNA molecule occurs exclusively in the cytoplasm of the transfected cells (Fig. S1), thereby eliminating risks of genomic integration and cell transformation, which pose safety hurdles for recombinant DNA, viral vectors, and pDNA vaccines (3,4). Moreover, the barrier of nuclear delivery, which is thought to be a rate-limiting step for nonviral delivery of pDNA, is circumvented.…”

“…1A). The structural protein genes were replaced with genes encoding protein antigens, which are abundantly expressed from a subgenomic mRNA in the cytoplasm of cells transfected with these self-amplifying RNAs (3,4,10,11). The RNA was produced in vitro by an enzymatic transcription reaction from a linear pDNA template using a T7 RNA polymerase, thereby avoiding safety concerns and complex manufacturing issues associated with cell culture production of live viral vaccines, recombinant subunit proteins, and viral vectors.…”

Construction of yellow fever virus subgenomic replicons by yeast-based homologous recombination cloning technique

“…Thereafter, replicons were described for several flaviviruses including DENV-2 (Ng et al 2007), DENV-3 (Mosimann et al 2010), YFV (Jones et al 2005), WNV and TBEV (Kofler et al 2004).…”