Risks and Control Strategies of Scale-up in Purification Process

Before being launched on the market, the process development of biological products can be divided into three stages, namely, preclinical stage, phase I/II clinical stage, and phase III clinical and commercial manufacturing stage. Generally, the scales of the three stages are different, so the process scale-up is often accompanied by two process transfers.

Figure 1: Stages of Biological Product Process Development

As the main processes of downstream purification, tangential flow filtration and chromatography have mature scale-up strategies, which can reduce the risks of the process itself in the scale-up process. In addition, materials and equipment will also affect the process scale-up process. Therefore, it is essential to approach their introduction with a focus on the end goal, making choices that align with production requirements.

Risk Control of Materials

Select Manufacturing-Grade Materials

• The grade of materials should meet the manufacturing standards, and for human drug production, it is advisable to select GMP-grade materials as much as possible.

• The supply of materials needs to meet the requirements of manufacturing. Suppliers should have sufficient capacity and can ensure the continuity of production to meet the needs of the final manufacturing scale.

• The lead time of the material supply chain should be able to meet the timeline of process development to prevent delays in product launch due to material shortages.

Avoid Material Contamination

• Materials of non-animal origin should be used as much as possible, and the TSE and BSE certificates of the products should be checked.

• Materials of the corresponding grade should have certificates for sterility, endotoxin limit, and bacterial colony limit.

Select Materials Compatible with Products/Intermediates

• It is necessary to check the elution, chemical resistance, and absence of relevant enzymes of materials that come into contact with products/intermediates.

Select Scalable Materials

• The filters, cassettes, hollow fibers, and other consumables selected during process development should have models suitable for manufacturing scale. Chromatographic resins should have sufficient mechanical properties to support large-scale column packing.

Process Risk Control

Process Selection

During the process development of some products with precious samples (such as mRNA), in order to save samples during preclinical studies, the laboratory scale is usually small, and kits and other methods are often used in the process. Therefore, when scaling up for the first time, it is necessary to select a suitable alternative process. Usually, there are multiple optional technologies for manufacturing scale, and the best process selection is generally determined by the final manufacturing scale.

Process Scale-up Strategy

Linear scale-up is currently the most commonly used strategy for purification processes, including dead-end filtration, tangential flow filtration, and chromatography scale-up.

In tangential flow linear scale-up, it is necessary to maintain consistency in the flow path length and fluid dynamics within the flow path. The former requires that during the process development at laboratory scale, small filters with flow path lengths consistent with the cassettes or hollow fiber models used at the scaled-up scale should be selected. When processing samples with higher viscosity, the fluid dynamics may change, leading to unpredictable ultrafiltration behavior after scale-up. Therefore, the concentration factor used in the process should be avoided exceeding 50 as much as possible, and the diafiltration volume should be kept below 14 times.

During chromatography linear scale-up, the bed height should be maintained while expanding the diameter of the chromatography column. When the column diameter exceeds 30cm, it can lead to reduced support of the packing by the column wall, making the bed more prone to collapse at high flow rates. In addition to selecting resins with scaling-up cases, when determining process parameters such as flow rate at a small scale, the nature of the pressure-flow rate curve of the resin should be referenced.

It should be noted that the pressure-flow rate curve is usually measured using water or simple buffer solutions. If the liquid viscosity is relatively high at a certain stage of the chromatography step (such as the protein elution stage in the binding-elution mode, CIP alkaline cleaning stage, and complex sample loading stage), the flow rate should be further reduced.

Figure 2: Pressure-Flow Rate Curves of BioLink Chromstar® 6FF and MaXtar® ARPA

One of the disadvantages of linear scale-up is that the diameter of chromatography columns is not continuous, and in some cases, maintaining the bed height can result in a column volume significantly exceeding the actual amount of resin required, leading to waste. In affinity chromatography, ion exchange that does not rely on column efficiency, and hydrophobic chromatography, a scale-up strategy based on constant retention time can be used. In this strategy, the bed height will change, and particular attention should be paid to overpressure and bed collapse of the chromatography column, especially when the bed height is higher than that at the small scale.

Chromatography Column Packing

The chromatography column packing method should be robust and reliable to ensure that the column efficiency measurement results meet the process requirements. The asymmetry factor is commonly used to assess the rationality of packing compression, and the acceptable range of the asymmetry factor should be within 0.8 to 1.8, while also meeting the separation requirements. Over-packing the column may cause the bed to crack, resulting in channeling and premature breakthrough of the sample. An overly loose bed may be re-compressed during use, creating liquid gaps and mixing within them, which can reduce separation efficiency. For chromatography processes that rely on the number of theoretical plates for separation, such as molecular sieving and peak-cutting ion exchange, the number of theoretical plates should also meet the process requirements.

Others

It is necessary to confirm the stability of the sample at room temperature; use a buffer whose pH value is not affected by temperature; and confirm the storage method of chromatography packing at a larger scale.

Equipments

There are often some differences between the equipment used after process scale-up and the equipment used on a small scale, even if they use the same principle and design.

The supplier should ensure that the equipment meets the requirements of hygienic design, including the polishing and welding quality of stainless steel components, as well as the surface treatment of material contact areas, to prevent potential cleaning dead zones and rust contamination after scale-up. The applicability of cleaning methods needs to be checked at the scaled-up scale.

Figure 3: BioLink Metal Processing Technology

On this basis, some details of the equipment will also have an impact on the process. For example, whether there is a change in the optical path of the UV detection flow cell in the chromatography system after scale-up; different lengths and diameters of outlet pipes may lead to more significant dilution and diffusion in the system; time delays caused by the response speed of the system control valves, combined with larger system dead volumes, may lead to errors in the starting and ending points of peak collection; the flow distribution of the chromatography column distributor may affect the separation effect, and so on. The main consideration for selecting a scaled-up chromatography system is to meet the flow rate requirements in the process steps. Generally, the process flow rate will be around the median of the maximum flow rate of the equipment. This is not the same as the consideration for selecting a laboratory chromatography system, which often has a wider flow rate range to cover different process conditions.

Figure 4: BioLink Lab-scale Chromatography System and Empty Chromatography Column

The laboratory column packing can be done using the axial compression method or the high flow rate method. However, the linear flow rate required for the high flow rate method often exceeds the upper limit of the most suitable chromatography system for process operation after scale-up. As the pressure resistance of the chromatography column decreases with its diameter, there are also hardware limitations for scaling up the high flow rate constant pressure method for column packing. Choosing a larger-scale chromatography system or a high-pressure chromatography column for column packing is not economical, so the axial compression method is usually adopted. For chromatography columns with a diameter of less than 300mm, a manual column can meet the packing requirements; while for columns with a diameter of 450mm or more, a chromatography column with electric axial compression function is required.

Figure 5: BioLink Chrom-LinX® Manual Chromatography Column and Verdot Ips2 InPlace Process-scale Chromatography Column

The impact of hardware often does not appear immediately, and selecting reliable and responsive suppliers to handle problems in a timely manner can reduce the potential risks faced by production.

Process scale-up in purification is a multi-faceted endeavor, and despite numerous successful cases, caution is still warranted when scaling up a new process. In particular, practical constraints such as limited resources and time often prevent the process development and scale-up from being carried out in an ideal manner. The above lists some common issues encountered in the process scale-up of purification. It is recommended to use tools such as GAP analysis and Failure Modes and Effects Analysis (FMEA) during the execution of process scale-up. These tools can help analyze the gaps between scales, assess process risks, and implement corresponding risk control measures tailored to specific process conditions, thereby enhancing the success rate of process scale-up.

Summary

The process scale-up in purification is a multi-faceted endeavor that requires caution even with numerous successful cases when scaling up a new process. In particular, due to practical constraints such as limited resources and time, the process development and scale-up cannot always be completed in an ideal manner. Above, we have outlined some common issues encountered in the process scale-up of purification. To enhance the success rate of process scale-up, it is recommended to use tools such as GAP analysis and Failure Modes and Effects Analysis (FMEA) during the execution process. These tools can help analyze the gaps between scales, assess process risks, and implement corresponding risk control measures tailored to specific process conditions.