Bioreactors for high cell density and continuous multi-stage cultivations: options for process intensification in cell culture-based viral vaccine production

Abstract

With an increasing demand for efficacious, safe, and affordable vaccines for human and animal use, process intensification in cell culture-based viral vaccine production demands advanced process strategies to overcome the limitations of conventional batch cultivations. However, the use of fed-batch, perfusion, or continuous modes to drive processes at high cell density (HCD) and overextended operating times has so far been little explored in large-scale viral vaccine manufacturing. Also, possible reductions in cell-specific virus yields for HCD cultivations have been reported frequently. Taking into account that vaccine production is one of the most heavily regulated industries in the pharmaceutical sector with tough margins to meet, it is understandable that process intensification is being considered by both academia and industry as a next step toward more efficient viral vaccine production processes only recently. Compared to conventional batch processes, fed-batch and perfusion strategies could result in ten to a hundred times higher product yields. Both cultivation strategies can be implemented to achieve cell concentrations exceeding 107 cells/mL or even 108 cells/mL, while keeping low levels of metabolites that potentially inhibit cell growth and virus replication. The trend towards HCD processes is supported by development of GMP-compliant cultivation platforms, i.e., acoustic settlers, hollow fiber bioreactors, and hollow fiber-based perfusion systems including tangential flow filtration (TFF) or alternating tangential flow (ATF) technologies. In this review, these process modes are discussed in detail and compared with conventional batch processes based on productivity indicators such as space-time yield, cell concentration, and product titers. In addition, options for the production of viral vaccines in continuous multi-stage bioreactors such as two- and three-stage systems are addressed. While such systems have shown similar virus titers compared to batch cultivations, keeping high yields for extended production times is still a challenge. Overall, we demonstrate that process intensification of cell culture-based viral vaccine production can be realized by the consequent application of fed-batch, perfusion, and continuous systems with a significant increase in productivity. The potential for even further improvements is high, considering recent developments in establishment of new (designer) cell lines, better characterization of host cell metabolism, advances in media design, and the use of mathematical models as a tool for process optimization and control.

Alternating flow filtration as an alternative to internal spin filter based perfusion process: Impact on productivity and product quality

“…Also, the quality of the produced protein must fulfill well‐established requirements and specifications. ATF technology was widely evaluated for cell culture‐based viral vaccine production . Also, in silico comparisons between ISF and ATF cell retention systems were published from economic, environmental and operational perspectives .…”

“…Therefore, in order to improve cost‐effectiveness ratio, perfusion systems should be optimized to reach higher cell densities with minimum cell‐specific perfusion rates and to allow longer runs …”

“…It supports high cell densities, with moderate equipment cost . This external cell‐separation system is able to maintain continuous culture for extended periods of time and offers the capability of rapid filter change without compromising the culture run . The ATF system allows increased volumetric productivity and reduced bioreactor size…”

High-cell-density cultivations to increase MVA virus production

“…To further increase virus titers, high-cell-density (HCD) processes can be established [17]. While virus propagation in HCD often results either in a decrease in cell-specific yields [18,19] the so-called ''cell-density effect" [20,21] -or in low volumetric productivity [22], there are reports on positive effects on virus titers and cell-specific yields if certain medium feeding/exchange strategies prior to or during virus propagation are being applied.…”

Hybrid simulation algorithm for efficient numerical solution of population balance equations

“…Multicellular systems are found in many bioprocesses, for example vaccine production (Müller et al, 2013;Tapia et al, 2016), biopolymer production (Franz et al, 2011(Franz et al, , 2012, biological waste water treatment (Nopens et al, 2015)), and they play an important part in many physiological processes like cell differentiation (Bessonnard et al, 2014). Individual cells can be characterized by certain properties, e.g.…”