Stay in line: Achieving cost-effective small molecule medicines

Engin Ayturk PhD, senior R&D manager at Pall Life Sciences reviews the challenges of downstream process optimisation and advantages of single pass tangential flow filtration (SPTFF), looking at a broad range of results with inline concentration.

A growing need for downstream process optimisation

The fast-growing global biopharmaceutical market is both attractive and daunting for pharmaceutical firms. While large molecule drugs have the potential to generate significantly higher revenues than traditional small molecule medicines, producing these complex molecules in a cost-effective and timely manner is a difficult challenge.

To address this, industry has sought to make steps in the biopharmaceutical production processes as efficient as possible. For example, significant improvements in upstream monitoring technologies and culturing techniques have increased productivity and efficiency and allowed manufacturers to achieve high titers.

Unfortunately, for efficiency-focused biopharmaceutical firms, advances in downstream technologies have been slower to emerge. Consequently, the focus now is on improving how the products of upstream processes feed into later purification steps.

Continuous bioprocessing is viewed by many as a potential solution, and this approach is attracting significant attention because switching from batch to continuous manufacturing has been shown in many industry sectors to lead to significant savings in cost and time, greater efficiency and improved product quality and consistency.

With integrated, automated continuous processing, unit operations are directly connected to one another with better process controls, eliminating intermediate hold steps and reducing waste and the likelihood of human errors and equipment downtime; this results in more efficient and quality operations with smaller physical and environmental footprints.

Technologies that can facilitate continuous processing, most notably single-use solutions that enable the integration of downstream unit processes, are of interest. Suppliers have been working to deliver solutions that enable continuous processes. In particular, inline concentration of dilute solutions prior to various downstream purification steps, such as continuous chromatography, facilitates continuous manufacturing while increasing productivity and reducing operating costs.

Enabling efficiency with concentration

The efficiency with which the products of upstream processes are captured and concentrated affects the efficiency of the overall production process. Traditionally, concentration in biomanufacturing involved direct and/or normal flow filtration operations by running feed streams through a membrane or bed that traps the solids in the filter and allows the filtrate to pass through. Such processes can be used to concentrate product to a limited efficiency; however, they require significant additional handling and frequent replacement of blocked filters.

In contrast, widespread adoption of tangential flow filtration (TFF) enabled the feed stream to be fed across the surface of the filter rather than into it. One advantage of the approach is that filter life is extended. However, because it involves the recirculation of the process fluid, TFF can lead to degradation and/or aggregation of fragile (shear-sensitive) biologic molecules.

Single-pass TFF (SPTFF), in which the recirculation loop is eliminated by a unique flow path design and the staging of cassettes in a serial and parallel configuration, overcomes the limitation of conventional TFF. Using this approach, bioprocess fluids pass only once through the pump and module; the reduced residence time and shear exposure eliminates mixing and foaming issues and can lead to reduced risk of product damage or aggregation. In addition, if pressurised vessels are used to flow the process fluid through an SPTFF module, concerns over sensitivity to pumping can also be eliminated.

Reducing the volume and increasing the titer of bioprocess fluids enables the adoption of smaller tubing and equipment sizes for unit operations downstream of the SPTFF concentration step. Perhaps most notably, the adoption of single-use technologies is often possible, which is attractive because they can provide significant savings in both capital and operating expenses, allow for reduced setup and switching times between processes, and reduce the risk of cross-contamination.

Moving toward continuous processing

In particular, continuous processes provide the greater flexibility in manufacturing needed today to accommodate the shift to more targeted, niche, smaller-volume biopharmaceutical drugs, as well as the constantly changing market needs. An increasing number of branded and generic biologics manufacturers and contract service providers have pursued the concept of semi-continuous processing on a commercial scale, and have sought to integrate these processes over the next several years. From this perspective, it is possible that within five years fully integrated bioprocesses will be implemented at the production scale.

The application of the SPTFF technique allows for better control of the purification steps by normalising the protein concentration from cell culture to chromatography and maintains the process productivity and economical gains afforded by continuous chromatography. Furthermore, it enables significant process cost reduction without the need to improve the upstream perfusion process, thus facilitating the move from batch to continuous mode and increasing speed to market.

The Cadence Inline Concentrator (ILC) system from Pall Life Sciences, based on SPTFF technology, incorporates Delta regenerated cellulose membranes that offer high flux, high selectivity, and low protein binding. Yields are higher due to improved product recovery and reduced working and hold-up volumes compared with TFF — the line sizes are 2–3 times smaller and the feed flow rates are 5–10 times lower. The modules can also help eliminate or reduce the size of intermediate storage tanks and the associated cleaning of tanks when used for in-process volume reduction while achieving typical concentration factors of 2–4 times (or more).

Broad applications for SPTFF

In addition to its use as an alternative to TFF and for the concentration of clarified cell-culture fluids prior to chromatography, SPTFF technology can be used to optimise other downstream processes by performing product concentration at ideal process points. Examples include concentration between different chromatographic purifications, in-process volume reduction, and in some cases, final biologic drug substance concentration. Product recovery of at least 99% can often be achieved after processing with inclusion of a buffer flush of a single system hold-up volume.

The scalability of the performance of Cadence ILC modules of different sizes has been demonstrated by evaluating fluxes and concentration factors, which determine the processing times, membrane areas and tank sizes required for larger processes, over a wide range of processing conditions. As a result, end users can directly scale up processes modeled in the laboratory to process development, pilot and production scales. Furthermore, although they are single-use systems, Pall has shown that Cadence ILC modules can be reused [5] several times with minimal change in performance in terms of seal integrity and volumetric concentration factors.

Advantages of SPTFF

To demonstrate the broad applicability of the SPTFF technology, the technical team conducted numerous studies with different feedstocks and with the ILC system coupled to other downstream unit operations.

In one study, the inline volume reduction of a highly viscous feed stream consisting of a suspension of nanoparticles used in therapeutic drug delivery applications was evaluated. The volume reduction was needed to enable the scale up of an ultrafiltration/diafiltration step from a 10 m2 membrane to a 40 m2 membrane without the need to scale the 400 L stainless steel hold tank due to floor space limitations. A 30 kDa Cadence ILC module was shown to provide a stable process at a feed pressure of 30 psig (2.0 barg), resulting in a feed flux of 22 LMH, a permeate flux of 16 LMH, a 3.7 times volumetric concentration factor, and an actual concentration factor of 4.1 times based upon solids content, which met the process target.

In another study, the combined use of SPTFF and multicolumn chromatography for continuous mAb capture from a clarified CHO cell culture fluid via Protein A affinity sorbent was demonstrated. Clarified feedstocks containing low MAb concentrations (≤1 g/L) were used to mimic a typical product titer achieved from perfusion cell culture.

In another case, a 1000 L batch of 0.4 g/L mAb in clarified CHO feedstock was concentrated using the Cadence ILC system before a capture step to reduce the size of the existing chromatography column and hardware (i.e. 60 cm column) yet retain a total chromatography processing time of four hours. A five-fold mAb concentration was achieved using a 17.5 m2 SPTFF module before loading onto the smaller size (i.e. 30 cm) column. The four-fold reduction in sorbent volume resulted in approximately $500,000 in savings. Further cost savings were realised through a four-fold reduction in buffer consumption, considering an identical number of CVs was used for the non-load (i.e. wash, CIP, re-equilibration, etc.) steps.

In another example [4], a 3.5 m2 SPTFF module was operated at a four-fold volume reduction capacity with a membrane chromatography step for capturing recombinant proteins at a 500 L scale. Inline concentration allowed a three-fold reduction in chromatography membrane costs and a two-fold reduction in the total processing time. Buffer consumption was also marginally reduced.

Next steps

It has long been recognised that continuous processing has the potential to revolutionise biopharmaceutical production. The problem has always been the technical challenge of implementing such processes on an industrial scale.

The major challenge has been that improvements in upstream culturing techniques and technologies have outpaced those downstream, creating a bottleneck and preventing the implementation of truly efficient continuous processes. Innovations like SPTFF are a significant step towards addressing this bottleneck and helping manufacturers optimise their downstream processes, reduce costs and bring products to market faster.