Multiproduct Resin Reuse for Clinical and Commercial Manufacturing-Methodology and Acceptance Criteria.

Chromatography resins used for purifying biopharmaceuticals are generally dedicated to a single product. In good manufacturing practice (GMP) facilities that manufacture a limited amount of any particular product, this practice can result in the resin being used for a fraction of its useful life. A methodology for extending resin reuse to multiple products is described. With this methodology, resin and column performance, product carryover, and cleaning effectiveness are continually monitored to ensure that product quality is not affected by multiproduct resin reuse (MRR). Resin and column performance is evaluated in terms of (a) system suitability parameters, such as peak-shape and transition, and height equivalent theoretical plate (HETP) data; (b) key operating parameters, such as flow rate, inlet pressure, and pressure drop across the column; and (c) process performance parameters, such as impurity profiles, product quality, and yield. Historical data are used to establish process capability limits (PCLs) for these parameters. Operation within the PCLs provides assurance that column integrity and binding capacity of the resin are not affected by MRR.Product carryover defined as the carryover of the previously processed product (A) into a dose of the subsequently processed product (B) (COA→B), should be acceptable from a predictive patient safety standpoint. A methodology for determining COA→B from first principles and setting acceptance limits for cleaning validation is described.Cleaning effectiveness is evaluated by performing a blank elution run after inter-campaign cleaning and prior to product changeover. The acceptance limits for product carryover (COA→B) are more stringent for MRR than for single-product resin reuse. Thus, the inter-campaign cleaning process should be robust enough to consistently meet the more stringent acceptance limits for MRR. Additionally, the analytical methods should be sensitive enough to adequately quantify the concentration of the previously processed product (A) and its degradants in the eluent.General considerations for designing small-scale chromatographic studies for process development are also described. These studies typically include process-cycling runs with multiple products followed by viral clearance studies with a panel of model viruses. Small-scale studies can be used to optimize cleaning parameters, predict resin performance and product quality, and estimate the number of multiproduct purification cycles that can be run without affecting product quality. The proposed methodology is intended to be broadly applicable; however, it is acknowledged that alternative approaches may be more appropriate for specific scenarios.LAY ABSTRACT: Chromatography resins used for purifying biopharmaceuticals are generally dedicated to a single product. In good manufacturing practice (GMP) facilities that make a limited amount of any particular product, this practice can result in the resin being used for a fraction of its useful life. A methodology for extending resin reuse to multiple products is described. With this methodology, resin and column performance, product carryover, and cleaning effectiveness are continually monitored to ensure that product quality is not affected by multiproduct resin reuse.General considerations for designing small-scale chromatographic studies for process development are described. These studies typically include process-cycling runs with multiple products followed by viral clearance studies with a panel of model viruses. Small-scale studies can be used to optimize cleaning parameters, predict resin performance and product quality, and estimate the number of multiproduct purification cycles that can be run without impacting product quality.The proposed methodology is intended to be broadly applicable; however, it is acknowledged that alternative approaches may be more appropriate for specific scenarios.

Structural and functional characteristics of virgin and fouled Protein A MabSelect resin cycled in a monoclonal antibody purification process.

The structural and functional characteristics of the Protein A MabSelect resin are determined for a virgin sample and for samples removed from a column that had been operated in an antibody capture process which had shown losses in product recovery over fewer than 20 cycles. Compared to the virgin resin, the cycled samples show reduced porosity and apparent pore size based on inverse size exclusion chromatography while transmission electron microscopy (TEM) shows accumulation of foulants on the cycled resin. Adsorption isotherms, batch adsorption kinetics, and batch desorption kinetics, obtained using the antibody in purified form, show that the cycled samples have about 10% lower binding capacity and slower mass transfer. Confocal scanning laser microscopy shows, however, that different degrees of fouling exist for different beads in the cycled samples, which may correspond to the existence of areas exposed to minimal or no flow in the process column. Replacing the standard cleaning procedure with an improved multi-step cleaning protocol prevented the accumulation of foulants in the resin beads, as evident from TEM, and resulted in a stable operation with high recovery.

Nature of foulants and fouling mechanism in the Protein A MabSelect resin cycled in a monoclonal antibody purification process.

The composition and origin of foulants and their spatial distribution within the particles of the Protein A MabSelect resin cycled in a mAb purification process are determined using electron and confocal microscopy techniques with gold and fluorescently labeled protein probes that associate with the foulants. The results show that the foulants are primarily related to the mAb product, are heterogeneously dispersed both on the outer surface and in the interior of the resin beads, and accumulate only when loading the conditioned CHO cell culture supernatant. Insignificant accumulation is seen if the process is run with purified mAb or with the null cell culture supernatant. When bound to the Protein A ligand, the mAb responsible for the observed fouling behavior is shown to associate with BSA and α-lactalbumin. This property is exploited using labeled versions of these lipophilic proteins to assess the effectiveness of improved resin cleaning processes and to elucidate the fouling mechanism. Resin fouling for this mAb appears to be consistent with the occurrence of conformational changes that occur upon binding, which, in turn, facilitate association of lipophilic proteins with the mAb. Upon desorption at low pH, these destabilized mAb complexes are deposited on and within the resin growing with each cycle and eventually leading to significant degradation of process performance.