Novel Synthetic Adsorptive Depth Filter Media for CHO Harvest Clarification

John Amara, Ph.D. and Michael Dullen

Abstract
Recent advancements in the productivity of upstream cell culture processes for the manufacture of therapeutic proteins, including monoclonal antibodies (mAbs), have resulted in greatly elevated cell densities and higher levels of process-related impurities. Current depth filters may demonstrate an insufficient filtration capacity for these applications and often comprise naturally-derived materials which require increased flushing to remove undesired organic and inorganic extractable components.
We report the design and performance characteristics of a novel synthetic depth filtration media that demonstrates a greatly increased filtration capacity for the clarification of mAb from CHO-S harvests. When compared to commercial depth filtration media derived from natural components, this synthetic depth filter media is shown to have significantly decreased organic extractables and reduced WFI flushing requirements by TOC analysis. As an added benefit, the synthetic media is also shown to impart a modest HCP clearance capability through an adsorption mechanism.

Introduction
Objective: Develop a disposable synthetic depth filter media.

Key features:

• lower flushing requirements

• increased process capacity

• increased HCP & DNA impurity clearance

in three depth filter grades:

• primary clarification: D0SP

• secondary clarification: X0SP

• combined primary+secondary: C0SP

delivered in a disposable Millistak+^® Pod device format.

Materials

Media configuration details
Summary of depth filter media configurations for three synthetic depth filters, as compared to commercial Millistak+^® HC benchmark configurations.

Procedure
Six new synthetic depth filter media layers were prepared from proprietary mixtures of a polyacrylic fiber, a silica filteraid and a wet-strength binder resin. These depth filter media layers were subsequently incorporated into Millistak+^® micropod devices (23 cm² filtration area). The performance of the synthetic depth filters was evaluated in the primary and secondary clarification of CHO cell harvests and compared to commercial benchmark devices.

Results

Discussion
Multi-mode adsorption properties
Depth filter media components:

• Acrylic fiber pulp (hydrophobic surface)

• Silica gel filter-aid (~50% of silanols ionized at pH 7 and can contribute significantly to cation exchange mechanisms)¹

• Cationic wet-strength binder (anion exchange mechanism)

¹Ghose, S.; McNerney, T.M.; Hubbard, B. Biotechnology and Bioengineering, 2004, 87, 3, 413-423. Hair, M.L.; Hertl, W. J. Phys. Chem. 1970, 74, 1, 91-94.

DSP impact study
Performed direct harvest clarification of CHO-S feed (mAb05) using D0SP/X0SP 2/1 compared to D0HC/X0HC 2/1 benchmark

Clarified feeds were subsequently processed through capture chromatography (ProSep^® Ultra Plus resin). Equivalent mAb05 recovery from both filter trains

Direct harvest clarification using D0SP/X0SP 2/1 afforded a 50% increase in HCP clearance and 30% higher filtration capacity than the HC benchmark (early D0SP prototype configurations)

Increased protein A elution pool purity for SP clarified feedstream

Conclusion

1. Synthetic depth filters demonstrate a significantly increased filtration capacity and HCP impurity clearance compared to commercial cellulose/DE – based filters.

2. Tests using model proteins and solution additives reveal that a combination of electrostatic and hydrophobic adsorptive interactions with the silica filter-aid are largely responsible for the increased protein binding capacity

3. Positive impact of HCP impurity clearance during clarification, affording an increased mAb product purity in protein A and AEX F/T process steps. Further work is warranted to fully assess the advantages this feature brings to the downstream process economics.

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