title: Good Manufacturing Practices production and analysis of a DNA vaccine against dental caries.
abstract: OBJECTIVE
  To prepare a clinical-grade anti-caries DNA vaccine pGJA-P/VAX and explore its immune effect and protective efficacy against a cariogenic bacterial challenge.
  
  
  METHODS
  A large-scale industrial production process was developed under Good Manufacturing Practices (GMP) by combining and optimizing common unit operations such as alkaline lysis, precipitation, endotoxin removal and column chromatography. Quality controls of the purified bulk and final lyophilized vaccine were conducted according to authoritative guidelines. Mice and gnotobiotic rats were intranasally immunized with clinical-grade pGJA-P/VAX with chitosan. Antibody levels of serum IgG and salivary SIgA were assessed by an enzyme-linked immunosorbent assay (ELISA), and caries activity was evaluated by the Keyes method. pGJA-P/VAX and pVAX1 prepared by a laboratory-scale commercial kit were used as controls.
  
  
  RESULTS
  The production process proved to be scalable and reproducible. Impurities including host protein, residual RNA, genomic DNA and endotoxin in the purified plasmid were all under the limits of set specifications. Intranasal vaccination with clinical-grade pGJA-P/VAX induced higher serum IgG and salivary SIgA in both mice and gnotobiotic rats. While in the experimental caries model, the enamel (E), dentinal slight (Ds), and dentinal moderate (Dm) caries lesions were reduced by 21.1%, 33.0%, and 40.9%, respectively.
  
  
  CONCLUSIONS
  The production process under GMP was efficient in preparing clinical-grade pGJA-P/VAX with high purity and intended effectiveness, thus facilitating future clinical trials for the anti-caries DNA vaccine.
authors.affiliation: Key Laboratory for Oral Biomedical Engineering of Ministry of Education, School and Hospital of Stomatology, Wuhan University, Wuhan, China.
authors.forename: Ya-ping
authors.initials: YP
authors.lastname: Yang
authors.forename: Yu-hong
authors.initials: YH
authors.lastname: Li
authors.forename: Ai-hua
authors.initials: AH
authors.lastname: Zhang
authors.forename: Lan
authors.initials: L
authors.lastname: Bi
authors.forename: Ming-wen
authors.initials: MW
authors.lastname: Fan
mesh_terms: D000818:Animals; D048271:Chitosan / Q000737:chemistry*; D003731:Dental Caries / Q000276:immunology / Q000382:microbiology / Q000517:prevention & control*; D004345:Drug Industry / Q000592:standards; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D017408:Guidelines as Topic; D007071:Immunoglobulin A, Secretory / Q000276:immunology; D007074:Immunoglobulin G / Q000276:immunology; D051379:Mice; D008807:Mice, Inbred BALB C; D010957:Plasmids; D011786:Quality Control; D051381:Rats; D017208:Rats, Wistar; D015203:Reproducibility of Results; D012463:Saliva / Q000276:immunology; D019444:Vaccines, DNA / Q000276:immunology* / Q000592:standards
chemical_list: D007071:Immunoglobulin A, Secretory; D007074:Immunoglobulin G; D019444:Vaccines, DNA; C530535:pGJA-P VAX vaccine; D048271:Chitosan