Scalability of Cadence Inline Concentrator Modules for Bovine IgG Processing

Transcription

1 Application Note USD3004 Scalability of Cadence Inline Concentrator Modules for Bovine IgG Processing Catherine Casey, M.Sc., and Engin Ayturk, Ph.D., BioPharm Applications R&D

2 . Introduction Pall s single-pass tangential flow filtration (SPTFF) technology is revolutionizing bioprocessing, through process productivity, flexibility and yield improvements and enabling continuous bioprocessing. SPTFF utilizes a staged flow path design to achieve significantly higher concentration factors than a conventional TFF system in one pump pass [-4]. The single pump pass minimizes shear exposure and eliminates foaming/ mixing concerns. Additionally, pump and tubing sizes are reduced for increased cost savings. System hold-up volumes are also reduced to maximize product recovery with minimal dilution, while expensive hold tanks and large system footprints, characteristic of a conventional TFF system, are eliminated. Pall s Cadence Inline Concentrator (ILC) Module is an extension of the SPTFF product line. The Cadence ILC module is a holderless SPTFF device with a built-in restrictor that is designed to achieve concentration factors of 2 to 4X (or higher) [5]. It can be placed virtually anywhere within a given bioprocess to enable inline volume reduction and concentration. The ILC module requires only a feed pressure source (i.e., pump or pressure vessel) and feed pressure measurement device, and an optional flow meter further reducing system requirements (Figure ). Its reduced system footprint eases facility fit concerns where manufacturing floor space is at a premium. Figure Cadence Inline Concentrator Set-up Feed tank PERMEATE FEED F P RETENTATE (to break tank or inline to next unit operation) A few of potential applications of the Cadence ILC module include: inline volume reduction, concentration prior to capture step, concentration in between chromatography steps, and replacement of an existing conventional TFF concentration step. Table shows the eight Cadence ILC modules that are currently available. The ILC modules are available in four format sizes (T0, T02, T2 and T06), which correspond to membrane areas ranging from to 3.5 m 2. They are available with Delta membrane (regenerated cellulose) in both 0 and 30 kda MWCOs. 2

3 Table Available Cadence Inline Concentrator Modules Membrane & MWCO Format Size Membrane Area (m 2 ) Part Number Delta 0 kda T ILD00T00407 T ILD00T T2 0.7 ILD00T20407 T ILD00T Delta 30 kda T ILD030T00407 T ILD030T T2 0.7 ILD030T20407 T ILD030T It is important that the modules be scalable so the end-user will be able model their process at lab scale and directly scale-up to process or manufacturing scales. Parameters such as fluxes and concentration factors need to be scalable within an acceptable range, as they are important determinants of processing times, membrane area, and tank sizes required for the scaled up process. In this concept, the main objective of this study was to evaluate the scalability of the performance between commercially available ILC formats and cutoffs over a wide range of processing conditions. 2. Materials and Methods The smallest (T0, m 2 ) and largest (T06, 3.5 m 2 ) ILC modules were tested for scalability with both Delta 0 kda and 30 kda membrane. Prior to processing with protein, the system was set-up and the ILC module were pre-conditioned. The holderless design of the ILC module greatly simplified the system set-up by allowing for a plug and play installation. After sanitizing, flushing and buffer conditioning the module, processing was carried out with 2 to 0 g/l of polyclonal bovine IgG feedstocks (Lee Biosciences) in PBS buffer. Although the ILC module can potentially be positioned at several locations within a given bioprocess, this concentration range was chosen to represent the majority of case studies where the Cadence ILC module would typically be implemented between primary clarification and final concentration steps. ILC modules with 0 and 30 kda membranes were tested over a feed pressure range of 20 to 60 psig in order to generate performance maps, consisting of volumetric concentration factors (VCF) and respective fluxes (i.e., feed, retentate, permeate) for the 2, 5, and 0 g/l polyclonal bovine IgG feedstocks formulated in PBS buffer. Process information was captured at each set-point starting from high-to-low throughput run conditions to minimize hysteresis due to concentration polarization (i.e., high-to-low PFEED). Also, multiple data points were taken at each set-point to verify steady-state and process stability with subsequent sampling to confirm and quantify the performance. It should be noted that the aforementioned performance maps were generated in total recirculation mode in order to conserve solution. In a typical end-user application, the target molecule is processed in single-pass, as soon as the target test conditions are finalized. Feed concentrations and respective feed volumes per ILC module utilized for the scalability study is summarized in Table

4 Table 2 Materials and Feed Conditions Feed Concentration Feed Volume per ILC Module: T0 (0.065 m 2 ) T06 (3.5 m 2 ) 2 g/l 5L 00L 5 g/l 2L 40L 0 g/l L 20L The scalability metrics that were evaluated in this study were feed flux (L/m 2 /hr), retentate flux (L.m -2.h) and VCF with a criterion of 5% that is set to elucidate the performance dissimilarities between the smallest and largest ILC modules. 3. Results and Discussion The major trends between feed pressure, feed flux and VCF coupled with the performance benchmark between ILC modules with Delta 0 and 30 kda membranes (T06, 3.5 m 2 ) is given for the 5 g/l IgG processing case in Figure 2. Compared to the ILC Delta 0 kda modules, the Delta 30 kda modules exhibited higher feed and permeate fluxes, thus wider range of VCFs (i.e, ~5%), due to relative differences in pore size distributions and membrane permeability. As captured in Figure 2, with the decreasing feed pressure and subsequently the feed flux, the feed solution resides longer in the module, allowing for a larger percentage of the flow to permeate and higher concentration factors to be achieved. Therefore, the tradeoff between the operating fluxes (processing time) and the achievable VCF ranges (volume reduction), as well as membrane cut-offs, is of great importance to optimize target bioprocessing conditions. Figure 2 ILC Performance Map and Benchmark for Delta 0 and 30 kda T06 (3.5 m 2 ) Modules Tested with 5 g/l IgG in PBS Feed Flux, J [L/m 2 /hr] J 30 kda J 0 kda VCF 30 kda VCF 0 kda VCF [X] Feed Pressure [psig] 4

5 Figure 3 shows the resulting flux and VCF ranges for the Delta 0 kda and Delta 30 kda, T0 (0.065 m 2 ) and T06 (3.5 m 2 ) modules, over the feed concentration and operational feed pressure range of 2 to 0 g/l IgG in PBS and 20 to 60 psig, respectively. As depicted in Figure 3, both the fluxes and the VCFs for the T0 and T06 modules were similar and within the scalability target of 5% over the entire feed pressure and feed concentration ranges tested. Figure 3 ILC [a] Delta 0 kda and [b] Delta 30 kda Scalability for T0 (0.065 m 2 ) and T06 (3.5 m 2 ) Modules During Processing with 2, 5, and 0 g/l Bovine IgG in PBS Over the Feed Pressure Operational Window of psig [a] ILC Delta 0 kda T0 Module (0.065 m 2 ) 00 0 VCF for: Retentate & Feed fluxes for: VCF J RETENTATE T0 (0.065 m 2 ) J FEED T06 (3.5 m 2 ) 0 00 ILC Delta 0 kda T06 Module (3.5 m 2 ) vs. [b] ILC Delta 30 kda T0 Module (0.065 m 2 ) 00 0 VCF for: Retentate & Feed fluxes for: VCF J RETENTATE J FEED T0 (0.065 m 2 ) T06 (3.5 m 2 ) 0 00 ILC Delta 30 kda T06 Module (3.5 m 2 ) vs. 5

6 It is noteworthy to re-emphasize that the feed flux is a critical determinant for scalability because it regulates processing time, which in turn dictates the membrane area required for a given bioprocess volume. Similarly, the scalability of the VCFs is critical in order to appropriately determine the required tank sizes for product pooling and/or quantify the variants in inlet conditions, if ILC outlet is coupled with another unit operation. In addition to robustness of the performance between the process development (PD), pilot and manufacturing scale operations, the volume reduction with ILC will further result in the downsizing of respective downstream unit operations, enabling further cost savings and footprint reduction. Also important to note that the slight differences in module performance, shown in Figure 3[a] and [b], were attributed to minor variations in process temperatures and feed concentrations, differences in mass and/or volumetric loadings (g/m 2 and/or L/m 2 IgG feed) between T0 and T06 runs and the feed channel pressure drop of the individual cassettes used to build the respective modules. Nevertheless, scalability of the process fluxes and VCFs is confirmed by the good agreement between the smallest and biggest available ILC modules tested over the entire pressure (20 to 60 psig) and concentration (2 to 0 g/l) range. The scalable performance of the ILC modules will allow the process developers to perform benchmarking runs with PD scale ILC modules, while being able to gauge the performance of the scaled-up process without further qualification and/or validation runs at pilot and/or manufacturing scales, thus minimizing expensive feedstock needs and saving valuable development time and efforts. The end-user will be able to confidently scale up the ILC fluxes and VCFs in order to calculate tank sizes, processing times, and membrane areas required for their scaled-up process. For example, the batch processing capacity of ILC Delta 0 and 30 kda modules is calculated for all the commercially available format sizes, as shown in Figure 4. The case study assumes an 8 hour processing window for concentrating a 5 g/l batch by 2.5 to 3 fold when processed at a feed pressure of 60 psig. As high inlet pressures resulting in high feed fluxes, representing the highest throughput case, data shown in Figure 4 suggests that the 3.5 m 2 ILC modules are capable of processing 3000L to 4000L batches during the specified target processing time. Similar calculations can be performed to evaluate the impact of other process variables such as membrane area and tank size. Figure 4 ILC Delta 0 and 30 kda Batch Processing Capacity at CFEED=5 g/l and PFEED=60 psig Within an 8 Hour Window for all Available Cadence ILC Module Format Sizes Liters Processed in 8 Hours Delta 0 kda Delta 30 kda T0 T02 T2 T06 [0.065 m 2 ] [0.3m 2 ] [0.7m 2 ] [3.5m 2 ] Format Size and Total Membrane Area 6

7 4. Conclusions Pall s Cadence Delta 0 kda and 30 kda Inline Concentrator modules are holderless, single-pass TFF modules that utilize a staged flow path design and a built-in restrictor to achieve concentration factors suitable for inline volume reduction applications in a single module pump pass. The ILC modules are designed to concentrate typical mab feedstocks (i.e., IgG) by a factor of 2-4X (or higher); however, the exact concentration factors are dependent on the properties of the feed streams, with varying concentration, and viscosity profiles in various buffer types. The ILC modules offer reliable and scalable performance and available in format sizes suitable for R&D, process development, pilot and production scales and also enable continuous processing by coupling DSP opeartions.. The VCFs and feed fluxes of the smallest (T0, m 2 ) and largest (T06, 3.5 m 2 ) available format sizes in both 0 and 30 kda MWCOs met the scalability criterion of 5% over the entire pressure and concentration range evaluated. The proof of scalability is a key factor to allow process developers evaluate multiple products at R&D and/or PD scale and be able to accurately and confidently predict processing times, membrane areas, and tank sizes required for scaling-up target bioprocess. 5. References [] Leon Mir and Gaston de los Reyes. Method and apparatus for the filtration of biological solutions. US Patents 7,384,549 B2 (June 0, 2008), 7,682,5 B2 (March 23, 200), 7,967,987 B2 (June 28, 20) and 8,57,999 B2 (April 7, 202). [2] Casey, C., Gallos, T., Alekseev, Y., Ayturk, E., and Pearl, S. Protein concentration with Single-Pass Tangential Flow Filtration, Journal Membrane Science, 384(-2) (20) [3] Application Note (USD2789): Cadence Systems Employ New Single-Pass TFF Technology to Simplify Processes and Lower Costs. Pall Life Sciences. [4] Dizon-Maspat, J., Bourret, J., D Agostini, A. and Li, F. Single Pass Tangential Flow Filtration to Debottleneck Downstream Processing for Therapeutic Antibody Production. Biotechnology and Bioengineering, 09(4) (202) [5] Casey, C. and Ayturk, E. Application Note (USTR293): Volume Reduction and Process Optimization with Cadence Inline Concentrator. Visit us on the Web at us at biopharm@pall.com Corporate Headquarters Port Washington, NY, USA toll free (USA) phone biopharm@pall.com European Headquarters Fribourg, Switzerland +4 (0) phone LifeSciences.EU@pall.com Asia-Pacific Headquarters Singapore phone sgcustomerservice@pall.com International Offices Pall Corporation has offices and plants throughout the world in locations such as: Argentina, Australia, Austria, Belgium, Brazil, Canada, China, France, Germany, India, Indonesia, Ireland, Italy, Japan, Korea, Malaysia, Mexico, the Netherlands, New Zealand, Norway, Poland, Puerto Rico, Russia, Singapore, South Africa, Spain, Sweden, Switzerland, Taiwan, Thailand, the United Kingdom, the United States, and Venezuela. Distributors in all major industrial areas of the world. To locate the Pall office or distributor nearest you, visit The information provided in this literature was reviewed for accuracy at the time of publication. Product data may be subject to change without notice. For current information consult your local Pall distributor or contact Pall directly. 205, Pall Corporation. Pall,, and Cadence are trademarks of Pall Corporation. indicates a trademark registered in the USA and TM indicates a common law trademark. Filtration.Separation.Solution. is a service mark of Pall Corporation. /5, PDF, GN USD3004