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IRES-driven expression of the capsid protein of the Venezuelan equine encephalitis virus TC-83 vaccine strain increases its attenuation and safety.

Guerbois M, Volkova E, Forrester NL, Rossi SL, Frolov I, Weaver SC - PLoS Negl Trop Dis (2013)

Bottom Line: Here we describe a second generation of the recombinant TC-83 in which the subgenomic promoter is maintained and only the capsid protein gene is translated from the IRES.This VEEV/IRES/C vaccine candidate did not infect mosquitoes, was stable in its attenuation phenotype after serial murine passages, and was more attenuated in newborn mice but still as protective as TC-83 against VEEV challenge.Thus, by using the IRES to modulate TC-83 capsid protein expression, we generated a vaccine candidate that combines efficient immunogenicity and efficacy with lower virulence and a reduced potential for spread in nature.

View Article: PubMed Central - PubMed

Affiliation: Institute for Human Infections and Immunity, Sealy Center for Vaccine Development, and Department of Pathology, University of Texas Medical Branch, Galveston, Texas, United States of America.

ABSTRACT
The live-attenuated TC-83 strain is the only licensed veterinary vaccine available to protect equids against Venezuelan equine encephalitis virus (VEEV) and to protect humans indirectly by preventing equine amplification. However, TC-83 is reactogenic due to its reliance on only two attenuating point mutations and has infected mosquitoes following equine vaccination. To increase its stability and safety, a recombinant TC-83 was previously engineered by placing the expression of the viral structural proteins under the control of the Internal Ribosome Entry Site (IRES) of encephalomyocarditis virus (EMCV), which drives translation inefficiently in insect cells. However, this vaccine candidate was poorly immunogenic. Here we describe a second generation of the recombinant TC-83 in which the subgenomic promoter is maintained and only the capsid protein gene is translated from the IRES. This VEEV/IRES/C vaccine candidate did not infect mosquitoes, was stable in its attenuation phenotype after serial murine passages, and was more attenuated in newborn mice but still as protective as TC-83 against VEEV challenge. Thus, by using the IRES to modulate TC-83 capsid protein expression, we generated a vaccine candidate that combines efficient immunogenicity and efficacy with lower virulence and a reduced potential for spread in nature.

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Protection against challenge following vaccination of infant mice.Six-day-old mice were inoculated SC with 5×104 PFU of VEEV TC-83 or IRES-based viruses. Animals were challenged 6 weeks post-vaccination with 104 PFU SC of VEEV IC strain 3908 and monitored daily for survival (A) and weight change (B), with no deaths recorded after day 8 post-challenge. *** = P<0.0001.
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pntd-0002197-g008: Protection against challenge following vaccination of infant mice.Six-day-old mice were inoculated SC with 5×104 PFU of VEEV TC-83 or IRES-based viruses. Animals were challenged 6 weeks post-vaccination with 104 PFU SC of VEEV IC strain 3908 and monitored daily for survival (A) and weight change (B), with no deaths recorded after day 8 post-challenge. *** = P<0.0001.

Mentions: The ability of the new VEEV/IRES/C vaccine candidate to induce neutralizing antibodies and to protect against a lethal VEEV challenge was evaluated in neonatal and adult mouse models and compared to VEEV/mutSG/IRES/1 and TC-83. Animals that survived the single SC inoculation with VEEV/mutSG/IRES/1 and VEEV/IRES/C at 6 days of age were held for 6 weeks post-infection before sera were collected and tested by PRNT. Seroconversion was detected in 6 of 7 (85%) animals vaccinated with VEEV/IRES/C and in 6 of 10 (60%) animals vaccinated with VEEV/mutSG/IRES/1, with mean PRNT80 titers of 26±8 and 57±22, respectively (Table 2). Challenge was performed on these animals 3 weeks later with virulent VEEV strain 3908, a human isolate from the last major VEE epidemic [40], at a SC dose of 104 PFU (ca. 104 LD50). All sham-vaccinated animals died between days 6 and 8, whereas 30% mortality was recorded for the animals that received VEEV/mutSG/IRES/1, and all animals vaccinated with VEEV/IRES/C survived challenge (Fig. 8A). No weight loss was observed in the VEEV/IRES/C-vaccinated cohort after challenge, whereas the VEEV/mutSG/IRES/1- and sham-vaccinated animals lost an average of 6.5% and 19.4% of pre-challenge weight by day 6 post-challenge, respectively (Fig. 8B).


IRES-driven expression of the capsid protein of the Venezuelan equine encephalitis virus TC-83 vaccine strain increases its attenuation and safety.

Guerbois M, Volkova E, Forrester NL, Rossi SL, Frolov I, Weaver SC - PLoS Negl Trop Dis (2013)

Protection against challenge following vaccination of infant mice.Six-day-old mice were inoculated SC with 5×104 PFU of VEEV TC-83 or IRES-based viruses. Animals were challenged 6 weeks post-vaccination with 104 PFU SC of VEEV IC strain 3908 and monitored daily for survival (A) and weight change (B), with no deaths recorded after day 8 post-challenge. *** = P<0.0001.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3649961&req=5

pntd-0002197-g008: Protection against challenge following vaccination of infant mice.Six-day-old mice were inoculated SC with 5×104 PFU of VEEV TC-83 or IRES-based viruses. Animals were challenged 6 weeks post-vaccination with 104 PFU SC of VEEV IC strain 3908 and monitored daily for survival (A) and weight change (B), with no deaths recorded after day 8 post-challenge. *** = P<0.0001.
Mentions: The ability of the new VEEV/IRES/C vaccine candidate to induce neutralizing antibodies and to protect against a lethal VEEV challenge was evaluated in neonatal and adult mouse models and compared to VEEV/mutSG/IRES/1 and TC-83. Animals that survived the single SC inoculation with VEEV/mutSG/IRES/1 and VEEV/IRES/C at 6 days of age were held for 6 weeks post-infection before sera were collected and tested by PRNT. Seroconversion was detected in 6 of 7 (85%) animals vaccinated with VEEV/IRES/C and in 6 of 10 (60%) animals vaccinated with VEEV/mutSG/IRES/1, with mean PRNT80 titers of 26±8 and 57±22, respectively (Table 2). Challenge was performed on these animals 3 weeks later with virulent VEEV strain 3908, a human isolate from the last major VEE epidemic [40], at a SC dose of 104 PFU (ca. 104 LD50). All sham-vaccinated animals died between days 6 and 8, whereas 30% mortality was recorded for the animals that received VEEV/mutSG/IRES/1, and all animals vaccinated with VEEV/IRES/C survived challenge (Fig. 8A). No weight loss was observed in the VEEV/IRES/C-vaccinated cohort after challenge, whereas the VEEV/mutSG/IRES/1- and sham-vaccinated animals lost an average of 6.5% and 19.4% of pre-challenge weight by day 6 post-challenge, respectively (Fig. 8B).

Bottom Line: Here we describe a second generation of the recombinant TC-83 in which the subgenomic promoter is maintained and only the capsid protein gene is translated from the IRES.This VEEV/IRES/C vaccine candidate did not infect mosquitoes, was stable in its attenuation phenotype after serial murine passages, and was more attenuated in newborn mice but still as protective as TC-83 against VEEV challenge.Thus, by using the IRES to modulate TC-83 capsid protein expression, we generated a vaccine candidate that combines efficient immunogenicity and efficacy with lower virulence and a reduced potential for spread in nature.

View Article: PubMed Central - PubMed

Affiliation: Institute for Human Infections and Immunity, Sealy Center for Vaccine Development, and Department of Pathology, University of Texas Medical Branch, Galveston, Texas, United States of America.

ABSTRACT
The live-attenuated TC-83 strain is the only licensed veterinary vaccine available to protect equids against Venezuelan equine encephalitis virus (VEEV) and to protect humans indirectly by preventing equine amplification. However, TC-83 is reactogenic due to its reliance on only two attenuating point mutations and has infected mosquitoes following equine vaccination. To increase its stability and safety, a recombinant TC-83 was previously engineered by placing the expression of the viral structural proteins under the control of the Internal Ribosome Entry Site (IRES) of encephalomyocarditis virus (EMCV), which drives translation inefficiently in insect cells. However, this vaccine candidate was poorly immunogenic. Here we describe a second generation of the recombinant TC-83 in which the subgenomic promoter is maintained and only the capsid protein gene is translated from the IRES. This VEEV/IRES/C vaccine candidate did not infect mosquitoes, was stable in its attenuation phenotype after serial murine passages, and was more attenuated in newborn mice but still as protective as TC-83 against VEEV challenge. Thus, by using the IRES to modulate TC-83 capsid protein expression, we generated a vaccine candidate that combines efficient immunogenicity and efficacy with lower virulence and a reduced potential for spread in nature.

Show MeSH
Related in: MedlinePlus