Chromatographic Approaches in the Purification of Plasmid DNA for Therapy and Vaccination

Abstract

The need for large scale plasmid DNA purification processes has increased in the last few years due to the rapid evolution of gene therapy and DNA vaccines. Chromatographic purification of plasmid DNA has specific problems which are mostly related to the structural nature of plasmid DNA, the resemblance of plasmid DNA to some impurities, and the lack of capacity and selectivity of the traditional bead adsorbents. In this paper, plasmid DNA purification using chromatographic processes is reviewed. Initially the most relevant structural and chemical properties of supercoiled plasmid DNA and related impurities are described, and the purification objectives are specified. The most significant properties are highlighted and their possible exploitation for plasmid DNA adsorption is described. After a brief description of currently used chromatographic purification strategies, novel approaches centered on the use and development of new stationary phases with higher capacity utilization are described.

Poly(hydroxyethyl methacrylate) based affinity cryogel for plasmid DNA purification

“…Commercially available carriers are designed primarily for protein binding rather than large diameter macromolecules and nanoparticles such as pDNA [33]. It has been reported that these large macromolecules are excluded from particle pores smaller than 500 nm.…”

Binding and purification of plasmid DNA using multi-layered carbon nanotubes

“…These nucleic acids are non-active admixtures in the preparations of supercoiled p-DNA which may induce immune reactions (Ferreira, 2005;Stadler et al, 2004). We did not study the adsorption of other immunogenic bacteria components such as proteins onto CNTs.…”

“…chromosomal DNA and RNAs. These latter forms are immunogenic and therefore their removal is a prerequisite step in preparing pharmacologically pure plasmid DNA for clinical applications (Ferreira, 2005;Stadler et al, 2004).…”

“…This biotechnological problem particularly covers large-scale isolation of plasmid DNA for gene therapy and vaccination (Ferreira, 2005). Plasmid DNA produced by bacteria and some eukaryotes has compact supercoiled structure which considerably differs from that of other forms of nucleic acids, e.g.…”

Adsorption of plasmid DNA on ceramic hydroxyapatite chromatographic materials

“…Numerous plasmid purification protocols including precipitation and chromatographic processes have been developed [2][3][4][5]. Chromatographic methods for plasmid DNA (pDNA) purification are characterized by high selectivity and high resolution (HR), but the stationary phases used for plasmid purification are limited because of the large size of plasmids.…”