Adaptive laboratory evolution of stable insect cell lines for improved HIV-Gag VLPs production.

Adaptive laboratory evolution (ALE) has been extensively used to modulate the phenotype of industrial model organisms (e.g. Escherichia. coli and Saccharomyces cerevisae) towards a specific trait. Nevertheless, its application to animal cells, and in particular to insect cell lines, has been very limited. In this study, we describe employing an ALE method to improve the production of HIV-Gag virus-like particles (VLPs) in stable Sf-9 and High Five cell lines. Serial batch transfer was used for evolution experiments. During the ALE process, cells were cultured under controlled hypothermic conditions (22 °C instead of standard 27 °C) for a prolonged period of time (over 3 months), which allowed the selection of a population of cells with improved phenotype. Adapted cells expressed up to 26-fold (Sf-9 cells) and 10-fold (High Five cells) more Gag-VLPs than non-adapted cells cultured at standard conditions. The production of HIV Gag-VLPs in adapted, stable insect Sf-9 cell lines was successfully demonstrated at bioreactor scale. The Gag-VLPs produced at 22 °C and 27 °C were comparable, both in size and morphology, thus confirming the null impact of adaptation process and hypothermic culture conditions on VLP's quality. This work demonstrates the suitability of ALE as a powerful method for improving yields in stable insect cell lines producing VLPs.