Truly continuous low pH viral inactivation for biopharmaceutical process integration

Abstract

Continuous virus inactivation (VI) has received little attention in the efforts to realize fully continuous biomanufacturing in the future. Implementation of continuous VI must assure a specific minimum incubation time, typically 60 min. To guarantee the minimum incubation time, we implemented a packed bed continuous viral inactivation reactor (CVIR) with narrow residence time distribution (RTD) for low pH incubation. We show that the RTD does not broaden significantly over a wide range of linear flow velocities—which highlights the flexibility and robustness of the design. Prolonged exposure to acidic pH has no impact on bed stability, assuring constant RTD throughout long term operation. The suitability of the packed bed CVIR for low pH inactivation is shown with two industry‐standard model viruses, that is xenotropic murine leukemia virus and pseudorabies virus. Controls at neutral pH showed no system‐induced VI. At low pH, significant VI is observed, even after only 15 min. Based on the low pH inactivation kinetics, the continuous process is equivalent to traditional batch operation. This study establishes a concept for continuous low pH inactivation and, together with previous reports, highlights the versatility of the packed bed reactor for continuous VI, regardless of the inactivation method.

A common framework for integrated and continuous biomanufacturing

“…For the systems that use continuous inactivation, a PFR is required. Many PFR designs have been published for low pH virus inactivation (Gillespie et al, 2019; Klutz et al, 2016b; Martins et al, 2020; Parker et al, 2018). Since the amount of acid and base changes with product concentration, the stream might require dilution with water to maintain constant ionic strength.…”

Developments and opportunities in continuous biopharmaceutical manufacturing

“… A subset of proposed technologies for continuous biopharmaceutical manufacturing. (a) Perfusion cell culture with alternating tangential flow filtration (ATF) for cellular retention, (b) 3-column periodic countercurrent chromatography for product capture 44 and a two-column solvent gradient purification method for product polishing, 45 (c) a continuous packed bed viral inactivator, 46 , 47 and (d) hollow fiber dialysis module for continuous ultrafiltration 48 and diafiltration. 49 …”

“…This can also be performed continuously, as shown recently using a continuous reactor with a narrow RTD, where significant viral inactivation was observed after just 15 mins. 46 To ensure an appropriate minimum residence time, the FDA recommends evaluating the RTD for continuous viral inactivation. 85 To this end, three viral inactivation reactor designs have been proposed: a coiled flow inverter, 86 , 87 a tubular reactor called jig-in-a-box, 88 , 89 and a packed-bed reactor 46 , 47 ( Figure 1c ).…”

“…However, challenges remain to be overcome in the integration of continuous viral inactivation reactors with multicolumn chromatography methods that process sample periodically. 46 The variation in the pH and concentration of the affinity eluate over time may also hinder the performance of continuous viral inactivation, requiring an additional hold step. 46 In addition to viral inactivation, viral filtration may also be carried out continuously 90 under a lower pressure over an extended duration.…”

“… 46 The variation in the pH and concentration of the affinity eluate over time may also hinder the performance of continuous viral inactivation, requiring an additional hold step. 46 In addition to viral inactivation, viral filtration may also be carried out continuously 90 under a lower pressure over an extended duration. Filters appropriate for such an operation must be carefully chosen, 90 and multiple filter set-ups may be considered.…”