Lyophilization, or freeze-drying, is a solution for sensitive therapeutic molecules, particularly complex dynamic biologics, to overcome physical and chemical instability and achieve desired serviceable life. In many cases, lyophilization is the only means to produce protein, peptide and vaccine products with pharmaceutically acceptable stability, shelf life, and bioactivity.
The increasing R&D investment for novel biologics and vigorous expansion of the global biopharmaceutical market are fueling the growth of the lyophilization market. Valued at $1.97 billion in 2014, the global lyophilization market for pharmaceutical and biotechnology products is expected to reach $2.66 billion by 2019 at a compound annual growth rate of (CAGR) of 6.2%.1 Currently, lyophilized products account for about 25% of newly approved parenteral drugs.2 The number of approved biologics and biosimilars has been steadily increasing in recent years. This trend is expected to propel more new lyophilized products onto the market.
Nevertheless, lyophilization, a downstream fill-finish operation, is a complex process to develop. It presents unique challenges to the manufacturer on product design, formulation and process engineering. To meet patients’ needs and regulatory requirements, the industry has shifted from traditional trial-and-error practice to a science-based Quality by Design (QbD) approach for lyophilization process development. By implementing QbD principles in drug discovery and development, product quality can be predicted, controlled, and consistently delivered through translating in-depth product and process knowledge into manufacturing sophistication.
The majority of lyophilized molecules are proteins, peptides, liposomes, and oligonucleotides. Each molecule is unique in terms of physical, chemical, biological, thermal, and stability characteristics. Therefore, for each lyophilized product, the formulation must be designed for that specific active ingredient and aimed to maximize its safety and efficacy profile. Further, the lyophilization cycle should be designed to accommodate this formulation. The QbD approach provides a systemic and efficient platform to develop a robust formulation fundamental to produce a quality, lyophilized product.
The usage of a combination of QbD tools including process analytical technology (PAT), risk assessment and design space, plays an important role in achieving QbD goals. The QbD framework starts with defining the quality target product profile (QTPP) and critical quality attributes (CQAs), which lay out the product and quality goals for the formulation and process to achieve. Some lyophilization-specific CQAs include potency, product related impurities, cake elegance, residual moisture content, appearance, reconstitution time, clarity after reconstitution and long-term stability.3 The specification for residual moisture is required in the data package for regulatory filing and regulated by 21 CFR (Code of Federal Regulations) 610.13 (a) Purity.4
A typical lyophilization formulation contains the active ingredient and a variety of excipients including bulking agents (e.g. mannitol), cryoprotectants (e.g. PEG), lyoprotectants (e.g. disaccharides), tonicity adjusters (e.g. glycerol) and buffers (e.g. histidine).5 For lyophilized products, one important aspect during extensive pre-formulation and formulation studies is to understand how the formulation, and every component in it, reacts to the freeze-drying cycle and their impacts on the CQAs. In this respect, the key critical material attributes (CMAs) to be determined include the glass transition temperature of the freeze concentrate (Tg) at which liquid changes to solid, and the collapse temperature (Tc) for amorphous products or the eutectic temperature(Teu) for crystalline materials.6 Traditionally, differential scanning calorimetry (DSC) thermogram has been used to assess the thermal profile of a material for decades. Tg is usually determined by DSC. Due to the complex nature of biological products, glass transition often occurs over a range of temperatures.7 Freeze-dry microscopy (FDM) represents an advanced tool that is used to determine Tc and Teu and simulate freeze-drying cycle in microscale. These data are then used to determine the critical formulation temperature (Tcrit), which sets the upper limit of the temperature during freezing.6
Once the initial characterization of the product is established, the next step is to define the design of experiment (DoE) for formulation and conduct a conservative lyophilization cycle based on the thermal profile of the formulation. Through this exercise, further insights are gained on formulation variables, their influence on CQAs, and formulation optimization. As the knowledge of the product and process accumulates along with the advancement of product development, the second DoE can be conducted to further optimize the formulation.3
A robust formulation must account for the freeze-drying conditions to ensure that the product can withstand an aggressive lyophilization cycle.3 For example, salts (e.g. phosphates) that undergo pH change during freezing are avoided in the lyophilization formulation. Likewise, a robust lyophilization process can only be developed based on the profound understanding and characterization of the formulation. The impact of the freeze-drying cycle on the product attributes and process parameters is evaluated throughout the process development studies. The knowledge gained is used to define design space and optimize the formulation and process.
In general, the lyophilization cycle is a three-step process; freezing, primary drying and secondary drying, where a product is first dried by sublimation, followed by desorption post freezing. The thermal characteristics of a formulation provide the knowledge needed to define the critical process parameters for the lyophilization cycle. For example, in the freezing stage, a rule of thumb is that the product is cooled well below the critical temperature (Tcrit) of the formulation to ensure it is completely frozen. In addition, the rate of freezing can greatly impact the CQAs.
Primary drying is conducted at a lower pressure and slightly higher temperature compared to the freezing stage, but still below Tcrit to promote sublimation, while the secondary drying is conducted at the minimal pressure and a higher temperature.8 The operating conditions for primary and secondary drying are mainly dominated by three variables: temperature of the heat transfer fluid, chamber pressure, and time of operation.7 It is critical to define the combination of these variables so that the final product can deliver desired CQAs.7 The optimal combination can be found through extended experimental campaign, sequential analysis, or DoEs, all of which can be time-consuming.7 Among the three stages of lyophilization process, primary drying is considered the most critical stage since it generally requires the longest operating time and poses the highest risk to product quality.6
One key element in QbD is to define the design space, the range of multidimensional combination, interaction of input variables and process parameters within which acceptable product quality attributes can be achieved.9 Design space can be specified for both the formulation and process although the industry focuses more on process design space.6 Typically, the process design space is identified by statistical DoEs and mathematical modeling. Process analytical technology tools provide measurement of critical process parameters (CPPs) that is essential to develop design space. The main PAT used in design space is the tunable diode laser absorption spectroscopy (TDLAS), a near-infrared technology that provides noninvasive in-line monitoring of the mass flow rate of water vapor during primary drying.6 The data obtained in TDLAS is comparable to another PAT tool, SMART technology, which can measure the product temperature at the ice sublimation interface (Tp).10,11
Another important component in design space development is to understand the edge of failure. For the product, it is the process conditions under which the product is no longer acceptable, commonly seen as a collapsed product for amorphous formulations. For the equipment, the edge of failure is determined by equipment capability including refrigeration, condenser, vacuum, and heating capacity.10 For lyophilization, there are distinct differences in equipment capability among laboratory, pilot, and production scale lyophilizers. Understanding the equipment is also fundamental in lyophilization cycle development. The knowledge on the edge of failure is useful in setting a safety margin within the design space, which leads to a robust lyophilization cycle that accommodates a range of variation of the material, process and equipment parameters without compromising the quality of the final product.
The benefits of QbD are evident with respect to regulatory filing, scale-up, process improvement, and troubleshooting. However, it requires extensive analytical and characterization studies upfront, which can add a burden to many companies, especially small-sized companies. Another challenge in implementing QbD is lack of PAT tools for production-scale lyophilization. As an integral part of QbD, PAT is used for real-time process monitoring and controlling and should be applicable for all scales of the equipment. Several PAT tools, such as TDLAS, can accurately measure process parameters at the lab-scale, but the measurement at production-scale is less reliable.6 To cope with this issue, it will be useful to understand the scalability of the critical product and process parameters. In addition, new PAT tools are in development including novel sensors distributed within the lyophilizer that can provide instantaneous readout of the system conditions during the freeze-drying cycle.2 The advancement in PAT will in turn promote the implementation of QbD in lyophilization manufacturing.
Kshitij (TJ) has been a part of Nice Insight since 2014. TJ’s role involves research design and operations, developing and maintaining syndicated studies, business intelligence data analysis, content development and article writing on the latest developments in the biopharmaceutical industry. Prior to market research, TJ spent time in academia research working on a broad range of subject matter, including pharmacoeconomics, drug delivery and genetics. TJ holds a masters of biotechnology degree from the University of Pennsylvania.