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. 2021 Sep;118(9):3581-3592.
doi: 10.1002/bit.27864. Epub 2021 Jul 1.

Process intensification for the production of yellow fever virus-like particles as potential recombinant vaccine antigen

Renata G F Alvim  1 , Túlio M Lima  1   2 , Jerson L Silva  3 , Guilherme A P de Oliveira  3 , Leda R Castilho  1
Affiliations

Process intensification for the production of yellow fever virus-like particles as potential recombinant vaccine antigen

Renata G F Alvim et al. Biotechnol Bioeng. 2021 Sep.

Abstract

Yellow fever (YF) is a life-threatening viral disease endemic in parts of Africa and Latin America. Although there is a very efficacious vaccine since the 1930s, YF still causes 29,000-60,000 annual deaths. During recent YF outbreaks there were issues of vaccine shortage of the current egg-derived vaccine; rare but fatal vaccine adverse effects occurred; and cases were imported to Asia, where the circulating mosquito vector could potentially start local transmission. Here we investigated the production of YF virus-like particles (VLPs) using stably transfected HEK293 cells. Process intensification was achieved by combining sequential FACS (fluorescence-activated cell sorting) rounds to enrich the stable cell pool in terms of high producers and the use of perfusion processes. At shaken-tube scale, FACS enrichment of cells allowed doubling VLP production, and pseudoperfusion cultivation (with daily medium exchange) further increased VLP production by 9.3-fold as compared to batch operation mode. At perfusion bioreactor scale, the use of an inclined settler as cell retention device showed operational advantages over an ATF system. A one-step steric exclusion chromatography purification allowed significant removal of impurities and is a promising technique for future integration of upstream and downstream operations. Characterization by different techniques confirmed the identity and 3D-structure of the purified VLPs.

Keywords: FACS for high producers; HEK293 cells; perfusion cultivation; stable expression; virus-like particles (VLPs); yellow fever virus.

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References

REFERENCES

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