Process Design and Optimization towards Digital Twins for HIV-Gag VLP Production in HEK293 Cells, including Purification

Abstract

Despite great efforts to develop a vaccine against human immunodeficiency virus (HIV), which causes AIDS if untreated, no approved HIV vaccine is available to date. A promising class of vaccines are virus-like particles (VLPs), which were shown to be very effective for the prevention of other diseases. In this study, production of HI-VLPs using different 293F cell lines, followed by a three-step purification of HI-VLPs, was conducted. The quality-by-design-based process development was supported by process analytical technology (PAT). The HI-VLP concentration increased 12.5-fold while >80% purity was achieved. This article reports on the first general process development and optimization up to purification. Further research will focus on process development for polishing and formulation up to lyophilization. In addition, process analytical technology and process modeling for process automation and optimization by digital twins in the context of quality-by-design framework will be developed.

Digital Twin for HIV‐Gag VLP Production in HEK293 Cells

“…All simulations were performed using Aspen Custom Modeler (AspenTechnology Inc., Bedford, MA, USA). The materials and methods used to generate the experimental data are described in detail in [23].…”

“…Few methods for the production of HIV-based VLPs in mammalian cells-particularly for suspension cultures-have been described in the literature [18]. Unlike non-enveloped VLPs, enveloped nanoparticles such as HIV-1-Gag VLPs are very sensitive to shear stress, pH variation, and osmotic pressure, rendering their production and purification especially challenging [8,23].…”

“…Designof-experiments (DoE) methods are traditionally used to reduce the experimental effort [47]. The final steps of QbD-based process development include the development of a Process Analytical Technology (PAT)-assisted control strategy and continuous improvement, which are considered in detail for the production of HIV-1 gag VLPs by Helgers et al [23,45,48]. In the first step, the QTPPs are defined.…”

“…In addition, this leads to a reduction in the workload of personnel due to lower cleaning and sterilization efforts. A process developed according to QbD guidelines allows better utilization of cost-intensive raw materials such as cultivation medium or feed and reduces the logistical effort [23,28,[49][50][51].…”

“…Therefore, the aim of this study was to develop a digital twin for the process developed and optimized by Helgers et al [23] to produce HI-VLPs in HEK293 cells as illustrated in Figure 3. To this end, simulation studies were performed for cultivation, cell separation by depth filtration, product purification, and concentration by ultra-and diafiltration (UF/DF) anion exchange chromatography (AEX).…”