Rapid Process Development for High Yield Plasmid DNA Fed-batch Fermentation - How to reduce plasmid-mediated metabolic burden for higher yields. - BioPharm International

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Rapid Process Development for High Yield Plasmid DNA Fed-batch Fermentation
How to reduce plasmid-mediated metabolic burden for higher yields.


BioPharm International
Volume 22, Issue 11

CONCLUSIONS

The temperature-inducible fermentation process has been successfully scaled-up to 300 L and used for GMP production of DNA vaccine plasmids. Fermentation cells are compatible with a variety of vectors,6–8 lysis,9 and downstream purification strategies.10–11 This low metabolic burden process is ideal as a generic plasmid DNA production platform to produce previously unstable and toxic plasmid DNA, as well as optimized plasmids at high yields in standard host strain DH5α, thus eliminating the need to use specialized stabilizing host strains. Contract manufacturers can use the base process with simple defined process optimization steps if necessary, to produce a wide range of customer-developed plasmids.

ACKNOWLEDGEMENTS

We thank Sheryl Anderson, Sarah Langtry, Justin Vincent, and Angela Schukar for their tireless efforts cleaning, batching, and operating fermenters. This paper described work supported by NIH grant R44GM072141.

James A. Williams is the vice president of research and development, Clague P. Hodgson is the president, and Aaron E. Carnes is the director of process development, all at Nature Technology Corporation, Lincoln, NE, 402.472.6530,

REFERENCES

1. Kutzler MA, Weiner DB. DNA vaccines: ready for prime time? Nat Rev Genet. 2008;9:776–88.

2. Carnes AE. Fermentation design for the manufacture of plasmid DNA. BioProcess Int. 2005;3(9):36–42.

3. Williams JA, Carnes A, Hodgson CP. Plasmid DNA vaccine vector design: impact on efficacy, safety and upstream production. Biotechnol Adv. 2009;27:353–70.

4. Williams JA, Luke J, Langtry S, Anderson S, Hodgson CP, Carnes AE. Generic plasmid DNA production platform incorporating low metabolic burden seed-stock and fed-batch fermentation processes. Biotechnol Bioeng. 2009;103:1129–43.

5. Carnes AE, Hodgson CP, Williams JA. Inducible Escherichia coli fermentation for increased plasmid DNA production. Biotechnol Appl Biochem. 2006;45:155–66.

6. Williams JA, Luke J, Johnson L, Hodgson C. pDNAVACCultra vector family: high throughput intracellular targeting DNA vaccine plasmids. Vaccine. 2006;24:4671–6.

7. Luke J, Carnes AE, Hodgson CP, Williams JA. Improved antibiotic-free DNA vaccine vectors utilizing a novel RNA based plasmid selection system. Vaccine. 2009. Epub.

8. Williams JA. Vectors and methods for genetic immunization. World Patent Application WO2008153733; 2008.

9. Carnes AE, Hodgson CP, Luke J, Vincent J, Williams JA. Plasmid DNA production combining antibiotic-free selection, inducible high yield fermentation, and novel autolytic purification. Biotechnol Bioeng. 2009;104:505–15.

10. Carnes AE, Williams JA. Plasmid DNA manufacturing technology. Recent Patents Biotechnol. 2007;1:151–66.

11. Hoare M, Levy MS, Bracewell DG, Doig SD, Kong S, Titchener-Hooker N, Ward JM, Dunnill P. Bioprocess engineering issues that would be faced in producing a DNA vaccine at up to 100 m3 fermentation scale for an influenza pandemic. Biotechnol Prog. 2005;21:1577–92.


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