Development of a Purification Process for Adenovirus: Controlling Virus Aggregation to Improve the Clearance of Host Cell DNA

Konz, John; Lee, Ann L.; Lewis, John A.; Sagar, Sangeetha L.

doi:10.1021/bp049644r

Cited by 66 publications

(46 citation statements)

References 28 publications

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“…Several unit operations have been described for the removal of solid matter from virus-containing cell culture supernatant: microfiltration [42], depth filtration [43,44] and centrifugation [11,43,45,46]. In the present study, clarification in batch centrifuges was evaluated as a model system for the clarification in manufacturing-scale continuous centrifuges.…”

Section: Centrifugationmentioning

confidence: 99%

“…Although highly effective, reduction in the specific DNA content by precipitation alone will not be sufficient to replace other unit operations for DNA depletion, e.g., chromatography [10] or treatment with Benzonase [11]. However, the significant reduction in the level of contaminating DNA will not only considerably facilitate design and optimization of succeeding chromatography steps, but would also allow to reduce the total amount of Benzonase used for DNA degradation.…”

Section: Dna Precipitationmentioning

confidence: 99%

“…Konz et al [11] reported the successful clearance of DNA in an adenovirus purification process, comprising of enzymatic digestion with Benzonase, ultrafiltration, and AEC. Removal of DNA was also achieved by adsorption to polymer beads cross-linked with cationic dimethylaminopropylacrylamide (DMAPAA) and polyethyleneimine (PEI) [7].…”

Section: Introductionmentioning

confidence: 99%

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DNA Depletion by Precipitation in the Purification of Cell Culture‐Derived Influenza Vaccines

et al. 2010

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A pilot study for the purification of cell culture-derived human influenza virus is presented, which focuses on the early removal of DNA by precipitation. Strains of influenza virus were propagated using Madin Darby canine kidney cells as a host. A harvesting time of about 72 h postinfection was chosen to minimize the level of impurities (host cell DNA and protein). Cell culture supernatant was clarified by centrifugation and the performance of this operation was characterized on the basis of Sigma theory. An average clarification efficiency of 93 % (based on turbidity) and a product yield of 85 % (based on hemagglutination activity) were obtained at a load of 1.6 · 10 -8 m s -1. Furthermore, the applicability of Sigma theory for scale-up studies using two different laboratory centrifuges was verified. Selective precipitation of DNA was achieved by the addition of polyethyleneimine (PEI). Full factorial design was applied to optimize selectivity considering pH, ionic strength, and the concentration and molecular weight of PEI. Under optimized conditions, treatment with PEI resulted in a reduction of DNA to 15 % of the initial amount, while 86 % of virions (based on neuraminidase activity) were recovered. The subsequent concentration of virions was realized by tangentialflow ultrafiltration. Recovery based on hemagglutination activity was determined to 63 % on average. Including the previous precipitation step, overall reduction in DNA after tangential-flow ultrafiltration was 500-fold. These results indicate that the suggested unit operations are suited for the early depletion of DNA in cell culture-derived influenza vaccine production.

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Section: Centrifugationmentioning

confidence: 99%

Section: Dna Precipitationmentioning

confidence: 99%

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DNA Depletion by Precipitation in the Purification of Cell Culture‐Derived Influenza Vaccines

et al. 2010

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“…Physical and chemical factors associated with the production and processing of recombinant viruses can alter the stability of the virus capsid and facilitate aggregate formation (Konz, 2005). Administration of a preparation containing aggregates can potentiate a severe immune response and, if the aggregates are sufficiently large, can facilitate clot formation and cause phlebitis (Brange, 1997;Braun, 1997;Thornton, 1993;Tomazic-Jezic, 2001).…”

Section: Effect Of Photosenstizers On Aggregation Of Adenoviral Partimentioning

confidence: 99%

Controlled inactivation of recombinant viruses with vitamin B2

Callahan

Wonganan

Obenauer-Kutner³

et al. 2008

Journal of Virological Methods

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Inactivated viruses are important tools for vaccine development and gene transfer. 8-methoxypsoralen (8-MOP) and long-wavelength ultraviolet irradiation (LWUVI) inactivates many viruses. Toxicity limits its use in animals and humans. Toxicological and photosensitizing properties of riboflavin make it suitable for virus inactivation in preparations for biological use. Viruses expressing beta-galactosidase were mixed with either 8-MOP (1.5 mM) or riboflavin (50 μM) and exposed to LWUVI (365 nm) for 2 hours. Virus activity was determined by limiting dilution. The half-life of the adenovirus preparation treated with 8-MOP was 8.28 nanoseconds −1 (ns −1 ) and 36.5 ns −1 after treatment with riboflavin. Despite the difference in half-life, both preparations were completely inactivated within 45 minutes. In contrast, the half-lives for adeno-associated virus (AAV) preparations were similar (63 ns −1 8-MOP vs. 67 ns −1 riboflavin). Each AAV preparation was fully inactivated within 90 minutes. The half-life of lentivirus was 193.4 ns −1 after treatment with 8-MOP and 208 ns −1 after exposure to riboflavin. Virus treated with riboflavin was inactivated within 20 minutes. Virus exposed to 8-MOP was inactivated in 90 minutes. DNA and RNA viruses can be inactivated by riboflavin and LWUVI and used in physiological systems sensitive to other photochemicals.

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“…the loss of the 5 protein functional activity after the treatment. Although antibody-based chromatography can avoid this, it is too specific and always leads to failure in binding the target viruses [4].…”

Section: Introductionmentioning

confidence: 99%

Nanoceramics for blood-borne virus removal

Zhao

Sugiyama

Miller

et al. 2008

Expert Review of Medical Devices

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20The development of nanoscience and nanotechnology in the field of ceramics has brought new opportunities for the development of virus removal techniques. A number of nanoceramics, including nanostructured alumina, titania, zirconia etc. have been introduced for the applications in virus removal/separation. Filtration or adsorption of viruses and thus the removal of viruses through nanoceramics, such as nanoporous/mesoporous ceramic 25 membranes, ceramic nanofibres, and ceramic nanoparticles will make it possible to produce an efficient system for virus removal from blood and with excellent chemical/thermal stability. Currently nanoceramic membranes and filters based on sol-gel alumina membranes and NanoCeram® nanofibre filters have been commercialized and applied to remove viruses from the blood. Nevertheless, filtration using nanoprous filters is limited to the removal of 30 only free viruses in the bloodstream.

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Development of a Purification Process for Adenovirus: Controlling Virus Aggregation to Improve the Clearance of Host Cell DNA

Cited by 66 publications

References 28 publications

DNA Depletion by Precipitation in the Purification of Cell Culture‐Derived Influenza Vaccines

DNA Depletion by Precipitation in the Purification of Cell Culture‐Derived Influenza Vaccines

Controlled inactivation of recombinant viruses with vitamin B2

Nanoceramics for blood-borne virus removal

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