Date on Master's Thesis/Doctoral Dissertation

5-2020

Document Type

Doctoral Dissertation

Degree Name

Ph. D.

Department

Microbiology and Immunology

Degree Program

Microbiology and Immunology, PhD

Committee Chair

Lukashevich, Igor S.

Committee Co-Chair (if applicable)

Mitchell, Thomas C.

Committee Member

Mitchell, Thomas C.

Committee Member

Sokoloski, Kevin J.

Committee Member

Chung, Donghoon

Committee Member

Steinbach-Rankins, Jill M.

Committee Member

Palmer, Kenneth E.

Author's Keywords

Vaccine; Lassa virus; Venezuelan equine encephalitis virus; small animal model development; biodefense vaccine; arenavirus

Abstract

Lassa Virus (LASV) and Venezuelan Equine Encephalitis Virus (VEEV) are two single stranded RNA viruses belonging to the Arenavirus and Alphavirus families, respectively. Both are emerging pathogens without approved vaccines or treatments. VEEV is an important biothreat pathogen because it has been weaponized and is extremely infectious as an aerosol. VEEV causes biphasic febrile illness that can progress to a viral encephalitis with low mortality, but high morbidity. LASV causes a viral hemorrhagic fever, Lassa Fever (LF), which is endemic in West Africa, and responsible for between 100-500k annual infections with a 1-2% overall mortality rate. ML29 and VEEV TC-83 are well-described live attenuated vaccines based on LASV and VEEV, respectively. This work describes strategies to further attenuate and enhance the safety of ML29 and TC-83, an important step in preclinical development. VEEV V4020 was designed based on the stabilization of an E2 mutation, and rearrangement of the structural genes of TC-83. Here, VEEV V4020 is shown to be more attenuated than TC-83 based on IC inoculation in mice, and more phenotypically stable, in terms of pathogenicity, during serial passaging. ML29 is a reassortant virus combining the immune dominant glycoprotein (GPC) and nucleoprotein (NP) of LASV with the replicative machinery of to Mopeia virus (MOPV), a nonpathogenic relative of LASV. We provide evidence ML29 is safer and more immunogenic than MOPV in STAT-1 deficient (STAT-1-/-) mice and Hartley guinea pigs. Additionally, Defective Interfering Particles (DIPs) from ML29 enhance vaccine immunity and attenuation in STAT-1-/- mice, and in intracranial inoculations of CBA/J mice. A potential mechanism of attenuation for ML29 is presented based on small RNAs detected from the ML29 L-segment by Northern Blot (NB). Furthermore, a unique RNA band was associated with DIP enriched ML29 was detected. A L-segment based Minigenome System (MG) for arenaviruses is described, which can be used for future analysis of the mechanism of replication for reassortant arenaviruses. The advanced safety data for both ML29 and VEEV V4020, described in this dissertation, combined with non-human primate efficacy studies described elsewhere, supports the advancement of both of these experimental vaccines to human clinical trials.

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