Conference Dates

June 12 – 17, 2022


HIV-1-based virus-like particles (VLPs) have high potential as scaffold for the development of chimeric or multivalent vaccines by functionalizing them with specific antigens [1], [2]. The obtention of vaccine formulations independent of cold chain is desirable to facilitate transportation and administration worldwide [3]. Recently, efforts are being made to develop cost-effective scalable processes to obtain these particles. The present study aims to compare some of the most used downstream processing (DSP) technologies for capture and purification of VLPs, taking especial consideration in using technologies enabling operation at large scale [4]. First, suspension adapted HEK 293 cells cultured in chemically defined cell culture media were used to produce the Gag-eGFP VLPs. Then, some steps of the purification process were studied, including a primary and secondary clarification by depth filtration and filtration respectively, an intermediate step by tangential flow filtration (TFF) or multimodal chromatography (MC), a capture step by ion exchange (IEC), heparin affinity (AC) and hydrophobic interaction chromatography (HIC), a polishing step by size exclusion chromatography (SEC) and a finally lyophilization step by freeze-drying process. Different operation units were tested for each step. Finally, a complete DSP train was implemented using the best results obtained in each stage. A concentration of 2.2 ± 0.8·109 VLPs/mL in the lyophilized samples was obtained after its storage at room temperature for 2 months. The morphology and structural integrity were further assessed by cryo-TEM. These first results in enveloped VLP lyophilization offer great promise to overcome the difficulty to distribute vaccines in poorly served remote rural areas and increase vaccine stability until their administration. Likewise, the purification methodologies proposed here could be easily scaled up and applied to purify similar enveloped viruses and vesicles.

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