Polishing Complex Therapeutic Proteins - A new downstream purification platform using a salt-tolerant membrane adsorber. - BioPharm International


Polishing Complex Therapeutic Proteins
A new downstream purification platform using a salt-tolerant membrane adsorber.

BioPharm International
Volume 26, Issue 1, pp. 42-51

pH and conductivity are critical parameters for Sartobind STIC operation

Table III: Full factorial design of experiments (DOE) screening study shows the effect of pH and conductivity on yield and host-cell protein (HCP) clearance. N.D. = not detected.
To evaluate how pH and conductivity variations may affect Sartobind STIC performance, a full factorial screening DOE study was designed in JMP with a pH range of 6.8–7.4, and a conductivity range of 36–42 mS/cm at 5 C. Aliquots from Bay-A001 immunoaffinity eluate were adjusted to pH and conductivity targets, as listed in Table III. Each aliquot was then loaded to a Sartobind STIC LP15 device. Step yield, DNA, and HCP clearances from each run were also listed in Table III. A contour plot showing trends of yield and HCP clearance in response to pH and conductivity changes was generated in JMP (see Figure 5). As expected from any anion-exchange flow-through operation, increasing pH decreased product yield but increased HCP clearance, while increasing conductivity increased product yield but decreased HCP clearance. Robust DNA clearance was observed because DNA levels in STIC FT from all four runs were either at or below the limit of detection. Careful control of pH and conductivity is, therefore, crucial for ensuring robust performance of Sartobind STIC.

The performance of Sartobind STIC with Bay-A002

Figure 5: Contour plot of pH and conductivity on Sartobind STIC yield and host-cell protein (HCP) clearance. Red lines and numbers represent predicted yield, and green lines and numbers represent predicted HCP fold clearance. The data used to generate this contour plot are listed in Table III. DNA levels in Sartobind STIC FT are either below or close to the limit of detection of 2.5 pg/mL, so no contour plot was generated for DNA clearance.
To demonstrate that Sartobind STIC can be a platform unit operation, performance in processing another Bayer recombinant protein product, Bay-A002, was tested. As outlined in the platform purification process (see Figure 1b), Bay-A002 is also captured from cell-culture harvest using a large-scale Q MA in bind-and-elute mode, followed by purification using an immunoaffinity column. An ion-exchange column and a Q MA flow-through step were used to further polish the product. The process was tested using Sartobind STIC in flow-through mode to polish the high salt immunoaffinity eluate, which was diluted to a conductivity of 39 mS/cm at 5 C for STIC loading. A laboratory-scale purification run showed that Sartobind STIC gave an excellent yield of 96%. It reduced HCP to 0.1 g/dose, a six-fold reduction, and reduced DNA to below the limit of detection. These HCP and DNA levels were comparable to those in the MA flow-through in the current process, indicating that Sartobind STIC can replace both the ion-exchange column and the Q MA steps (see Figure 6). Because anion-exchange is a versatile purification technique, the authors believe Sartobind STIC can be easily adapted to processing various proteins by finding the optimal pH and conductivity settings for each protein, thus making it a true platform technology.

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