(612f) Continuous Downstream Process of Antibody Developed Based on the Process Analysis and Understanding

Downstream process (DSP) of monoclonal antibody (mAb) includes several batch chromatography steps. As the efficiency of DSP is low, various strategies have been developed to improve the efficiency such as continuous operation. In this paper, we will present a continuous DSP process of mAb developed by the MAB consortium*[1].

A typical platform DSP includes three chromatography steps. For the first “capture” step, protein A chromatography is normally used. A multi-column operation called “periodic counter-current chromatography (PCCC)” can be employed as a continuous operation for the capture step. The efficiency of PCCC is strongly dependent on the operating conditions, which should be determined properly [2, 3]. We decided to use a PCCC system with two-protein A columns.

The recovered pool of the capture step should be kept at low pH for a specified time for virus inactivation (VI). Although this step can be carried out continuously by using a flow reactor, our analysis showed that the scale-down is difficult. So we decided to use a batch reactor, which can work also as a surge tank.

The polishing step usually includes two chromatography columns of different separation modes. We decided to use a pseudo continuous operation method known as flow-through chromatography (FTC) [4]. In order to eliminate the buffer exchange between two FTC operations, the same buffer solution was employed so that the two columns (mixed mode and cation exchange) can be connected in series. Virus filter (VF) was also connected at the exit of the second column.

After 24-h operation of PCCC, the recovered pool was transferred to the batch VI process, which performed one hour incubation at an assigned low pH. The solution pH and the conductivity were then adjusted for the polishing FTC step. Then, the FTC step started. PCCC and FTC were operated in parallel. After each operation for 24 hours, the lines of PCCC setup were cleaned, and the columns and/or the VF were replaced for the polishing step.

The operating conditions including the mobile phase properties for PCCC and 2-column connected FTC processes were determined based on the experimental data. The continuous operations for five days were carried out with different mAb concentrations (1–3.2 g/L), feed flow rates (0.5–10 L/d) and column volumes (1–100 mL). High yield and purity were accomplished for all runs ; yield >80%, purity determined by SEC > 95% [1].

*Manufacturing Technology Association of Biologics http://cho-mab.or.jp/english/

References

[1] Konoike, F., Taniguchi, M., Yamamoto, S. Continuous downstream process of antibody - Process development, optimization and verification (in Japanese). 52th SCEJ Autumn Meeting, Paper No. LF120, 2021.

[2] Chen，C-S., Yoshimoto, N., Yamamoto, S. Prediction of the performance of capture chromatography processes of proteins and its application to the repeated cyclic operation optimization. J. Chem. Eng. Jpn., 53, 689-697 (2020)

[3] Chen，C-S., Konoike, F., Yoshimoto, N., Yamamoto, S. A regressive approach to the design of continuous capture process with multi-column chromatography for monoclonal antibodies J. Chromatogr. A 1658, 46260 (2021)

[4] Hasegawa. S., Chen，C-S., Yoshimoto, N., Yamamoto, S. Optimization of flow-through chromatography of proteins. J.Chem.Eng. Jpn., 53, 214-221(2020)