September 1, 2017 (Vol. 37, No. 15)
Kevin Marino Global Product Manager Pall Life Sciences
Peter Levison Ph.D. Senior Marketing Director Pall Life Sciences
Improving Process Economics and Speed to Market with Cross-Functional Inline Concentration
The biopharmaceutical industry produces highly complex and effective medicines to treat challenging diseases. However, changing dynamics in the industry are placing significant pressure on existing biomanufacturing approaches. Greater flexibility and robustness, shorter development times, and lower costs—which translate into increased efficiency and productivity—are all needed to meet future demands for existing products and advanced, next-generation biologic drugs. While the productivity of upstream bioprocesses has been improved to produce dramatically higher titers, similar advances have not yet been achieved in downstream processing.
Process intensification of downstream operations has, thus, become a focus of the biopharmaceutical industry. Drug manufacturers and equipment suppliers are working closely to develop new systems and solutions for purification of high-quality biologic drug substances. The overall goals are to reduce complexity, equipment size, and overall operating footprint, while offering increased capacity as well as lower energy consumption and waste generation.
Some of the issues that must be addressed in downstream processing include a lack of sufficient storage capacity, large buffer (and thus process tank) volumes, extended process times, and large hold-up times during final product concentration.
Enabling Process Intensification with Single-Pass TFF
Concentration of bioprocess fluids prior to many downstream operations is an effective approach for achieving process intensification. This can be a challenge, however; when stressed, many biologics undergo aggregation or degradation, leading to reduced product quality, safety, and efficacy.
An effective concentration technology must, therefore, operate under mild conditions while also being easy to use and must easily integrate with other downstream operations. Tangential-flow filtration (TFF) has been widely adopted in the biopharmaceutical industry for the concentration of both biologic drug substances and formulated drug products. However, because conventional TFF involves recirculation of the process fluid, it can lead to product quality issues.
Single-pass TFF (SPTFF), on the other hand, does not involve recirculation. With the Cadence® Inline Concentrator (ILC) from Pall (Figure), process fluid flows through a staged-flow path with a step-wise decrease in the number of long, thin feed-flow channels consisting of Delta regenerated cellulose membranes. Rather than transmembrane pressure control, flow ratio control is employed. Concentration is achieved in a single pump pass. The increased path length increases productivity for a given mass throughput, and aggregation is minimized, even for fragile and shear-sensitive biomolecules.
With no need for a recirculation loop, 5–10x lower feed-flow rates are possible, allowing for lower working volumes and the use of 2–3x smaller line sizes and less expensive pumps. Mixing and foaming issues are avoided, leading to easier and higher product recoveries, and required flush volumes are reduced. This results in an improved product yield, in addition to enabling production scale runs using development-scale equipment. Importantly, SPTFF allows for very high concentration factors and thus, can be used to effectively produce highly viscous fluids, such as those required for drug products administered by subcutaneous injection.
SPTFF technology using inline concentration was developed for volume reduction before or after several types of downstream unit operations. Compared with conventional TFF devices, Cadence ILC modules achieve typical concentration factors of 2–4x (or more) and provide high flux, high selectivity, and low protein binding with greatly reduced system working and hold volumes.
Both TFF and SPTFF technologies are self-contained, reusable units with no contact to a holder surface. The Cadence ILC can be applied as a single-use technology to allow for more rapid changeover between runs through elimination of the need for cleaning and cleaning-validation steps (thereby avoiding cross-contamination, as well).
Overall, the technology helps biopharmaceutical manufacturers achieve not only process intensification, but also reductions in capital expenses, operating costs, and processing times. By performing inline concentration at ideal process points, it is therefore possible to optimize downstream processing and achieve better facility fit.
Many Applications for Inline Concentration
SPTFF has applications well beyond its obvious use as an alternative to conventional TFF. Inline volume reduction/concentration can be achieved prior to capture chromatography to reduce the column load, and in many locations throughout downstream processing where facilities lack sufficient tank volumes. SPTFF is also ideal for achieving in-process dilution and de-salting, and it has applicability for ultrafiltration/diafiltration (buffer exchange). As mentioned above, it is ideal for the high-yield generation of high-concentration formulations and well suited for the processing of fragile molecules.
In many of these applications, >99% product recovery can be achieved after processing with one hold-up volume flush.
Easy Application for Improved Performance
The single-use, pre-assembled, and pre-flushed device has a built-in retentate resistor. Back pressure is generated at a fixed restriction, that is variable based on the specific flow rate and product viscosity, allowing concentrations of 2–4x to be achieved. Retentate is collected off the top plate and permeate can be removed from the top and bottom.
No holder is required for the Cadence ILC, requiring only attachment to a feed pressure source (pump or pressure can) and a feed pressure measurement device. It can also be easily connected to other downstream unit operations, allowing for integrated bioprocessing. Different module sizes with demonstrated scalable performance facilitate process development and commercialization. The consistent performance across scales also allows for accurate prediction of needed membrane areas, tank sizes, and process times.
Pre-conditioning only involves sanitization, flushing of the storage solutions, determination of the hold-up volume, integrity testing, and buffer conditioning. It is recommended that a process-optimization step be performed to determine the optimal operating conditions for a given process. Typically feed pressure excursions in the range of 1.4–4.1 barg (20–60 psig) are conducted in total recirculation mode. Alternatively, feed flux excursions can be performed (keeping the operating feed pressure below 4.1 barg (60 psig). Higher feed pressures lead to higher permeate and feed fluxes, but lower volume concentration factors (VCFs). Each process will have an optimum trade-off between the fluxes and VCF value. Once the process conditions are determined, it is preferable to run the process in single-pass mode to confirm that a stable process can be achieved.
The Cadence ILC has been used to improve many downstream operations. A variety of feedstocks, including monoclonal antibodies (mAbs), antibody fragments (fAbs), bovine immunoglobulin (IgG), and α-amylase, have been concentrated four- to fivefold. Stable process conditions were achieved in each case, as well as a range of concentration factors and retentate flow rates.
- A 1,000-L batch of clarified Chinese hamster ovary (CHO) feedstock containing 0.4 g/L of a mAb was concentrated using the Cadence ILC to allow for use of a smaller chromatography column, while maintaining an overall chromatography processing time of 4 hours. The mAb concentration was increased fivefold using a 17.5 m2 ILC module, allowing reduction of the column size from 60 to 30 cm. A saving of approximately $500,000 was achieved due to a factor-of-4 reduction in the sorbent volume. Buffer consumption was also reduced by a factor of 4.
- Clarified mAb feedstocks containing low mAb concentrations (≤1 g/L) to mimic typical product titers achieved via perfusion cell culture were also successfully concentrated using the ILC.
- A fourfold volume reduction of process fluid containing a recombinant protein was achieved using a 3.5 m2 Cadence ILC module prior to membrane chromatography. As a result, the membrane cost and processing time were reduced by factors of 3 and 2, respectively.
- A VCF of 3.7x (actual concentration factor of 4.1x based upon solids content) was achieved for a highly viscous suspension of nanoparticles at a feed pressure of 2.0 barg (30 psig). The stable process had a feed flux of 22 LMH and a permeate flux of 16 LMH. Notably, the step was performed prior to ultrafiltration/diafiltration, enabling scale up of this operation from a 10 m2 membrane to a 40 m2 membrane without the need to scale the 400-L stainless-steel hold tank. Use of inline concentration therefore provided capital savings and addressed a potential manufacturing footprint issue.
Single-use, SPTFF technology enables process intensification for downstream purification of biologic drug substances and final formulation of drug products. Use of the Cadence ILC for SPTFF allows for simplification of many downstream unit operations, leading to cost and time savings. The modules are easy to install and use, and provide target volume concentration factors with high product recoveries. With reduced feed fluxes, line sizes, and working volumes compared with conventional TFF, the technology enables biopharmaceutical manufacturers to achieve better facility fit with improved process productivity at the R&D, process development, pilot, and production scales.