Industrial scale production of plasmid DNA for vaccine and gene therapy: plasmid design, production, and purification

Prather, Kristala Jones; Sagar, Sangeetha L.; Murphy, Jason C.; Chartrain, Michel

doi:10.1016/s0141-0229(03)00205-9

Cited by 187 publications

(178 citation statements)

References 132 publications

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“…Progress in gene therapy and genetic vaccination led to an increased demand in highly purified plasmid DNA (pDNA); estimations are in the range of several kilograms per year [1]. Initial protocols for pDNA purification were designed for molecular biology purpose, i. e. small quantities without specialized demands in purity and quality.…”

Section: Introductionmentioning

confidence: 99%

Adsorption of plasmid DNA on anion exchange chromatography media

Tarmann

Jungbauer²

2008

J of Separation Science

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Original PaperAdsorption of plasmid DNA on anion exchange chromatography media Anion exchange chromatography (AEC) is a useful and effective tool for DNA purification, but due to average pore sizes between 40 and 100 nm most AEC resins lack truly useful binding capacities for plasmid DNA (pDNA). Equilibrium binding capacities and uptake kinetics of AEC media including conventional media (Source 30 Q, Q Sepharose HP), a polymer grafted medium (Fractogel EMD DEAE (M)), media with large pores (Celbeads DEAE, PL SAX 4000 30 lm) and a monolithic medium (CIM-DEAE) were investigated by batch uptake or shallow bed experiments at two salt concentrations. Theoretical and experimental binding capacities suggest that the shape of the pDNA molecule can be described by a rod with a length to diameter ratio of 20:1 and that the molecule binds in upright position. The arrangement of DNA like a brush at the surface can be considered as entropy driven, kind of selfassembly process which is inherent to highly and uniformly charged DNA molecules. The initial phase of adsorption is very fast and levels off, associated with a change in mass transfer mechanism. Feed concentrations higher than 0.1 mg/mL pDNA pronounce this effect. Monolithic media showed the fastest adsorption rate and highest binding capacity with 13 mg pDNA per mL.

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Section: Introductionmentioning

confidence: 99%

Adsorption of plasmid DNA on anion exchange chromatography media

Tarmann

Jungbauer²

2008

J of Separation Science

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“…The foreign protein elicits a cellular immune response by triggering the maturation and proliferation of cytotoxic T-lymphocytes (CTL). The foreign antigen can also make contact with CD4 T cells and naive B cells, and initiates a classical humoral immune response cascade (25).…”

Section: Discussionmentioning

confidence: 99%

“…One possible solution for safer and more efficacious vaccine strategies is the development of DNA vaccines (15). DNA vaccines mimic the natural intracellular pathogen gene expression pathways, which trigger both cellular and humoral responses (25), and are simple to make and deliver.…”

mentioning

confidence: 99%

Protective Immunity against Bordetella pertussis by a Recombinant DNA Vaccine and the Effect of Coinjection with a Granulocyte‐Macrophage Colony Stimulating Factor Gene

Zhu

Chu

et al. 2006

Microbiology and Immunology

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A recombinant pertussis DNA vaccine was described here with its immunogenicity and the ability to induce protection against B. pertussis infection in mice. Three immunodominant antigen gene fragments of pertussis, pertussis toxin subunit 1 (pts1), fragments of pertactin (prn) and filamentous hemagglutinin (fha), were recombined as fragment pts1‐prn‐fha named ppf, and it was cloned to plasmid pVAX1 as pVAX1/ppf. Compared to those injected with pVAX1, the mice injected with pVAX1/ppf significantly elicited more antigen specific antibody anti‐PTS1, anti‐PRN, anti‐FHA and cytokine IL‐10, IFN‐γ. When pGM‐CSF was coinjected with pVAX1/ppf, the mice showed significantly increases of the three antibodies and cytokine IL‐10, IL‐4, IFN‐γ and TNF‐α compared to those injected with pVAX1 only. The mice in group pVAX1/ppf & pGM‐CSF, in particular; induced much more anti‐PTS1, IL‐4 and TNF‐α than those in group pVAX1/ppf. In the intracerebral mouse protection test, the mice immunized with pVAX1/ppf or pVAX1/ppf & pGM‐CSF induced protection to a lethal dose of B. pertussis. The results indicate that recombinant DNA vaccine and pGM‐CSF coinjection can induce protective immunity against B. pertussis, demonstrating a valuable method to prevent pertussis.

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“…This effect is more pronounced when larger beads are used ( Figure 7B). The R mt -value of 14.16 mg/g dcw suggests that all the plasmid content may be released if proper conditions are used [39]. However, the R msc1 -value of 8.98 mg/g dcw obtained at a low mill frequency represents the maximum concentration attainable and a lower pDNA(sc) purity.…”

Section: Rsm Analysismentioning

confidence: 99%

Efficient Disruption of Escherichia coli for Plasmid DNA Recovery in a Bead Mill

et al. 2017

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Featured Application: In general, the alkaline lysis is more acceptable than the mechanical lysis for pDNA recovery due to the risk of product degradation when the latter is used. In this work, the experimental design allowed us to explore mechanical disruption conditions under which such an effect was minimized. The application of this technique is important for novel pDNA-vaccine process development, since bead milling is one of the most preferable mechanical cell lysis methods on an industrial scale due to its scalability, ease of operation, controllability, and ability to load concentrated cell slurry. Abstract:The release kinetics of pDNA in a bead mill was studied. Samples taken during the process were analyzed to determine total pDNA (pDNA(t)) and supercoiled pDNA (pDNA(sc)) concentration. In order to identify important variables of the process and to develop an empirical model for optimal pDNA(t) and pDNA(sc) release, a two level 2 3 factorial design was used with variables: mill frequency, cell concentration, and bead size. The results were analyzed by response surface methodology. The optimized conditions for pDNA(t) yield 13.26 mg/g dcw (93.41% recovery), with a mill frequency of 30 Hz, a bead size of 0.10-0.25 mm, and a cell concentration of 20 g wcw/L. However, the optimized conditions for pDNA(sc) yield 7.65 mg/g dcw (92.05% recovery), with a mill frequency of 15 Hz, a bead size of 0.10-0.25 mm, and a cell concentration of 10 g wcw/L. Cell disruption in a bead mill was proved efficient for the release of pDNA(t) and pDNA(sc) compared to the alkaline treatment. The results obtained suggest a compromise between pDNA(sc) purity and recuperation in the process development.

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Industrial scale production of plasmid DNA for vaccine and gene therapy: plasmid design, production, and purification

Cited by 187 publications

References 132 publications

Adsorption of plasmid DNA on anion exchange chromatography media

Adsorption of plasmid DNA on anion exchange chromatography media

Protective Immunity against Bordetella pertussis by a Recombinant DNA Vaccine and the Effect of Coinjection with a Granulocyte‐Macrophage Colony Stimulating Factor Gene

Efficient Disruption of Escherichia coli for Plasmid DNA Recovery in a Bead Mill

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