title: A cDNA Clone-Launched Platform for High-Yield Production of Inactivated Zika Vaccine.
abstract: A purified inactivated vaccine (PIV) using the Zika virus (ZIKV) Puerto Rico strain PRVABC59 showed efficacy in monkeys, and is currently in a phase I clinical trial. High-yield manufacture of this PIV is essential for its development and vaccine access. Here we report an infectious cDNA clone-launched platform to maximize its yield. A single NS1 protein substitution (K265E) was identified to increase ZIKV replication on Vero cells (a cell line approved for vaccine production) for both Cambodian FSS13025 and Puerto Rico PRVABC59 strains. The NS1 mutation did not affect viral RNA synthesis, but significantly increased virion assembly through an increased interaction between NS1 and NS2A (a known regulator of flavivirus assembly). The NS1 mutant virus retained wild-type virulence in the A129 mouse model, but decreased its competence to infect Aedes aegypti mosquitoes. To further increase virus yield, we constructed an infectious cDNA clone of the clinical trial PIV strain PRVABC59 containing three viral replication-enhancing mutations (NS1 K265E, prM H83R, and NS3 S356F). The mutant cDNA clone produced >25-fold more ZIKV than the wild-type parent on Vero cells. This cDNA clone-launched manufacture platform has the advantages of higher virus yield, shortened manufacture time, and minimized chance of contamination.
authors.affiliation: Department of Biochemistry & Molecular Biology, University of Texas Medical Branch, Galveston, TX, USA; College of Animal Science and Technology, Southwest University, Chongqing, China.
authors.forename: Yujiao
authors.initials: Y
authors.lastname: Yang
authors.affiliation: Department of Biochemistry & Molecular Biology, University of Texas Medical Branch, Galveston, TX, USA.
authors.forename: Chao
authors.initials: C
authors.lastname: Shan
authors.affiliation: Department of Biochemistry & Molecular Biology, University of Texas Medical Branch, Galveston, TX, USA.
authors.forename: Jing
authors.initials: J
authors.lastname: Zou
authors.affiliation: Institute for Human Infections & Immunity, Galveston, TX, USA; Institute for Translational Science, Galveston, TX, USA.
authors.forename: Antonio E
authors.initials: AE
authors.lastname: Muruato
authors.affiliation: Department of Biochemistry & Molecular Biology, University of Texas Medical Branch, Galveston, TX, USA; Seção de Arbovirologia e Febres Hemorrágicas, Instituto Evandro Chagas, Ministério da Saúde, Ananindeua, Pará, Brazil.
authors.forename: Diniz Nunes
authors.initials: DN
authors.lastname: Bruno
authors.affiliation: Department of Biochemistry & Molecular Biology, University of Texas Medical Branch, Galveston, TX, USA; Seção de Arbovirologia e Febres Hemorrágicas, Instituto Evandro Chagas, Ministério da Saúde, Ananindeua, Pará, Brazil.
authors.forename: Barbosa
authors.initials: B
authors.lastname: de Almeida Medeiros Daniele
authors.affiliation: Seção de Arbovirologia e Febres Hemorrágicas, Instituto Evandro Chagas, Ministério da Saúde, Ananindeua, Pará, Brazil; Department of Pathology, Pará State University, Belém, Brazil.
authors.forename: Pedro F C
authors.initials: PFC
authors.lastname: Vasconcelos
authors.affiliation: Institute for Human Infections & Immunity, Galveston, TX, USA; Department of Pathology, Center for Biodefense & Emerging Infectious Diseases, Galveston, TX, USA.
authors.forename: Shannan L
authors.initials: SL
authors.lastname: Rossi
authors.affiliation: Institute for Human Infections & Immunity, Galveston, TX, USA; Institute for Translational Science, Galveston, TX, USA; Department of Microbiology & Immunology, Galveston, TX, USA; Sealy Center for Vaccine Development, Galveston, TX, USA; Sealy Center for Structural Biology & Molecular Biophysics, University of Texas Medical Branch, Galveston, TX, USA.
authors.forename: Scott C
authors.initials: SC
authors.lastname: Weaver
authors.affiliation: Department of Biochemistry & Molecular Biology, University of Texas Medical Branch, Galveston, TX, USA. Electronic address: xuxie@utmb.edu.
authors.forename: Xuping
authors.initials: X
authors.lastname: Xie
authors.affiliation: Department of Biochemistry & Molecular Biology, University of Texas Medical Branch, Galveston, TX, USA; Institute for Translational Science, Galveston, TX, USA; Sealy Center for Structural Biology & Molecular Biophysics, University of Texas Medical Branch, Galveston, TX, USA; Department of Pathology, Pará State University, Belém, Brazil; Department of Pharmacology & Toxicology, University of Texas Medical Branch, Galveston, TX, USA. Electronic address: peshi@utmb.edu.
authors.forename: Pei-Yong
authors.initials: PY
authors.lastname: Shi
mesh_terms: D000818:Animals; D002522:Chlorocebus aethiops; D006224:Cricetinae; D003412:Cricetulus; D018076:DNA, Complementary / Q000235:genetics; D057809:HEK293 Cells; D006801:Humans; D051379:Mice; D020125:Mutation, Missense; D019444:Vaccines, DNA / Q000235:genetics* / Q000276:immunology; D015164:Vaccines, Inactivated / Q000235:genetics / Q000276:immunology; D014709:Vero Cells; D017361:Viral Nonstructural Proteins / Q000235:genetics / Q000276:immunology; D000071244:Zika Virus / Q000235:genetics / Q000276:immunology*
chemical_list: D018076:DNA, Complementary; D019444:Vaccines, DNA; D015164:Vaccines, Inactivated; D017361:Viral Nonstructural Proteins
keywords: Flavivirus NS1; Flavivirus assembly; Zika virus vaccine