Strategies for Successful Lyophilization Process Scale-Up

Introduction Lyophilization, also termed as freeze-drying, is widely used in the pharmaceutical industry to prepare dry dosage forms. This unit operation involves three major phases: 1. Freezing, where liquid drug solution typically contained in glass vials is cooled and solidified, 2. Primary drying, where ice is removed from the frozen drug solution via sublimation through application of vacuum and heat, and 3. Secondary drying, where some of the residual noncrystalline water is removed via desorption. Several reports in the literature discuss approaches to formulation and process development for lyophilized products [1, 2, 3]. This article will focus on the lyophilization process aspects pertaining to large-scale manufacturing and considerations involved in scaling-up a lyophilization cycle from pilot scale to production scale. Basic lyophilization equipment is comprised of three major components: 1. A drying chamber, which typically consists of multiple shelves on which product containing vials are loaded, 2. A condenser, which is connected to the drying chamber through a vapor tube and functions as a water trap by condensing the water vapor from the drying chamber, and 3. Refrigeration, heating and vacuum system that is used to control the temperature of the shelves and the condenser, as well as the pressure in the chamber. These three components are responsible for controlling the overall heat transfer rate in and out of the product vials and mass transfer of water vapor away from the product vials. The rates of these heat and mass transport processes ultimately dictate the properties of the lyophilized product. Even though the user specified lyophilization cycle parameters (viz. shelf temperature, chamber pressure and time) may be the same, the heat and mass transfer rates may differ, depending on the design and performance of the lyophilizer. Therefore, during lyophilization scale-up, it is important to understand the differences in the equipment at two different scales and their impact on lyophilization process dynamics. What Exactly Changes During Lyophilization Scale-Up? Figure 1 illustrates the various parameters that influence the lyophilization process performance and the properties of a lyophilized product. Among the parameters listed, the Equipment Variables are hardest to match on different scales and often lead to adjustments to the Cycle Parameters to ensure equivalent product quality. Some of those variables are discussed in greater detail below.

Environmental Conditions in the Processing Area: Production scale machines are often operated in a clean room environment while testing on pilot scale is generally performed in a lab environment that has significantly higher

particulate loading. Particles from the environment can get introduced in the bulk solution when it is filled and loaded in the lyophilizer. These particles can act as nucleation sites leading to higher ice nucleation temperature on pilot scale versus the production scale during the freezing phase of a lyophilization cycle. It is widely reported in the literature that the nucleation temperature affects the ice crystal size lower the nucleation temperature smaller is the ice crystal size and greater is the ice interfacial area [4]. Small ice crystal size increases the mass transfer resistance offered by the dry cake during primary drying. As a result, the material produced in clean manufacturing environment tends to exhibit slower ice sublication and higher product temperature. Additionally, size of the ice crystals could potentially impact biochemical stability of some proteins due to degradation at ice/liquid or ice/void interface [5]. Shelf Surface Temperature: Shelf temperature during lyophilization is generally controlled based on the temperature of the heat transfer fluid (e.g. silicone oil) at the inlet of the shelf. On the other hand, what the product sees is the shelf surface temperature. Depending on the differences in the machine design and the shelf temperature control strategy, the shelf surface temperature may be different in different machines even though the heat transfer fluid temperature set point is kept the same. Furthermore, the production scale machines tend to have larger temperature variation across shelves, especially during heavy heat exchange requirement. Temperature difference of 2-5C between the inlet and outlet of a shelf is commonly observed during freezing ramp and early stages of primary drying. This could lead to greater variability in the temperature ramping rates, extent of annealing and sublimation rates among product vials from different regions of a shelf. The lyophilization cycle on production scale has to be appropriately designed to yield homogeneous product in spite of this variability. Alternatively, worst case locations need to be identified so that product from those locations can be monitored and tested as a part of validation and batch release analysis. Radiation Effects: For a product vial undergoing lyophilization, primary mode of heat transfer is gas phase conduction to or from the shelf on which the vial is placed. However, there is some contribution via radiation from shelves and walls of the chamber. The differences in the shelf spacing, the wall temperature and the proximity of the walls to the edge vials, could lead to varying degree of radiation contributions on different scales. This could, in turn, influence the product attributes, for the edge vials. Chamber Pressure Control: The pressure control strategy could also have an influence on process dynamics due to its impact on chamber gas composition during drying. When nitrogen is used for chamber pressure control, the mole fraction of water vapor during drying is less than unity. On the other hand, when the pressure control is carried out without any introduction of nitrogen, the drying chamber contains exclusively water vapor. As mentioned earlier, primary mode of heat transfer from shelves to vials is gas phase conduction. Since nitrogen has lower conductivity than water vapor [6], the differences in chamber gas composition could lead to differences in heat transfer rates and, subsequently, in primary drying durations. This is especially true with slow primary drying cycles and partial dryer loads. Type of Measuring Devices, Tolerances and Calibrations: Two most commonly used gauges for chamber pressure (vacuum) measurement during lyophilization are capacitance manometer type and Pirani type. The latter is not recommended for chamber pressure control since its response is greatly affected by the composition of gas in the chamber. It is important to use the same type of gauges on pilot and production scales to assure equivalent conditions for freeze drying. There could also be differences in the shelf temperature, chamber pressure control tolerances and transducer calibrations on the two scales which could lead to small differences in process dynamics. Although any one of the factors discussed here may not be enough to significantly affect the performance of a lyophilization process, the combination of these factors, if not accounted for upfront, could lead to significantly different process performance upon scale-up. Lyophilization Process Scale-Up Methodology The main objective of lyophilization process scale-up is to specify a robust lyophilization cycle that can be executed reproducibly and yields uniform product attributes consistent with the material manufactured on pilot or clinical scale. To be successful in achieving this objective, one of the keys is to have a well characterized formulation. The

thermal characteristics of the formulation in frozen state, such as the collapse temperature, should be evaluated to determine the critical product temperature that shouldnt be exceeded during early stages of primary drying in order to avoid impact on product quality [7]. It is also important to characterize the final lyophilized product for the target moisture range, cake appearance, reconstitution time and any other critical attributes so that appropriate acceptance criteria can be developed for testing product from production scale. Another key to successful scale-up is to do small scale process development while taking into account the capabilities of the large scale machine. As was discussed in the previous section, large scale equipment tends to have greater variability in shelf surface temperature and also some limitations in terms of maximum achievable ramping rates. The pilot scale process parameters need to be appropriately selected to account for these limitations (e.g. longer primary drying duration, slower freezing ramp). Additionally, vial and stopper components should be identical and be pre-processed the same as that for the production scale runs. Once the lyophilization process parameters are established on pilot scale, one to four engineering runs should be performed on production scale to confirm that the equipment can execute the intended lyophilization cycle and yields product of acceptable quality and uniformity. If the pilot scale cycle has been properly designed and with adequate safety margin, no adjustments to the shelf temperature, chamber pressure set points and ramping rates for production-scale cycle should be necessary. Only adjustments to the drying durations may be required because of the factors described in the previous section. Often times, active product is not available in sufficient quantities to perform these engineering runs. In these situations, mixed load of active and a suitable surrogate can be used. To demonstrate that the product quality is uniform regardless of the location in the lyophilizer, groups of active vials can be dispersed throughout the lyophilizer and analyzed for cake appearance, moisture content and any other relevant product quality attributes. Engineering runs can also be utilized to rationalize the choice of critical alarm limits on production scale. This can generally be accomplished with two runs. In one of the runs, the lyophilizer cycle can be set at the upper limits of primary and secondary drying shelf temperature and chamber pressure, as well as secondary drying duration. While in the other run, the lyophilizer can be set at the lower limits of primary and secondary drying shelf temperature and chamber pressure, and secondary drying duration. For example, if the target conditions are 0C and 120 mHg for primary drying, one of the runs can be executed at +3C, 140 mHg and the second run at -3C, 100 mHg. This choice of parameters brackets the entire range of product temperatures, sublimation rates and residual moisture contents that may be observed within the tolerance limits of the intended lyophilization cycle [8]. Once the lyophilization cycle on production scale is established and confirmed to be robust, it can be validated [9]. At this point, having comprehensive data from engineering runs minimizes any uncertainty with the validation batches. If the product uniformity has been well-characterized during engineering runs, reduced level of sampling for validation testing may be justifiable. Key Considerations During Lyophlization Scale-Up Equipment Capabilities: For economic and operational reasons, the production scale machines generally deliver lower cooling and heating capacity per unit shelf area as compared to the pilot scale machine. One should be aware of the maximum ramping rates that can be executed on production scale with full product load. Lyophilizer manufacturers generally specify the maximum ramping rates for empty chamber. However, the performance with product load tends to be significantly slower and should be determined during qualification of the machine. Another equipment parameter that could be potentially limiting is dynamic condenser capacity of the production scale machine. This parameter dictates maximum rate at which water can be captured (kg water/hr) by the condenser while maintaining the process set points. Formulations with high collapse temperature (e.g. high concentration protein formulations) can be freeze dried using fairly aggressive primary drying conditions. However, if the condenser is not able to keep up with the resulting water vapor, then that could lead to loss of pressure control. The lyophilizer manufacturers often specify only the ultimate condenser capacity. The dynamic condenser capacity can be estimated experimentally by determining maximum achievable ice sublimation rate for water filled vials/trays before the lyophilizer starts to lose chamber pressure control [10]. Table 1 lists some of the performance parameters

for a large production scale machine under empty and fully loaded conditions.

Choice of Surrogate Formulation: As discussed in the previous section, during engineering runs on production scale, the lyophilizer can be filled mostly with surrogate vials with groups of active vials dispersed across the lyophilizer. For this approach to be valid, the surrogate should mimic the freeze drying behavior of the active product. Accordingly, pilot scale studies should be performed to demonstrate that the sublimation rates and product temperature profiles of the surrogate vials are comparable to that of the actives. Examples of sublimation rate measurements for an antibody formulation (active), the corresponding formulation buffer (placebo) and a surrogate formulation are shown in Figure 2. Protein to formulation-excipient weight ratio for this particular antibody formulation was about 1:1. Hence, when just the formulation buffer was freeze dried, the mass transfer resistance to sublimation was significantly lower than that for active and the corresponding sublimation rate was significantly higher. On the other hand, the surrogate where the protein in the active formulation was replaced by the same weight concentration of sugar, yielded sublimation rate that was comparable to that of the active. Use of such a surrogate during engineering runs ensures that the active vials experience the same conditions and the lyophilizer experiences the same thermal load as it would when the chamber is filled with all active vials.

Lyophilization cycle Monitoring: As a part of the engineering runs, it is important to collect product temperature data during the lyophilization cycle. This allows one to confirm that the product is exposed only to intended temperatures (e.g. not exceeding the collapse temperature) during various stages of lyophilization. Additionally, based on the data, one can confirm that the product temperatures have reached shelf temperature prior to start of the

secondary drying ramp. If the product temperature information is available in real-time, adjustments can also be made to the primary drying duration on the fly, to accommodate any ice sublimation rate differences that may be present between the pilot and production scales. Often times on production scale, especially in machines with automatic loading/unloading systems, use of product temperature probes on routine basis is not possible. In such situations, one could rely on other measurements such as condenser temperature, chamber condenser pressure differential, nitrogen bleed flow rate and pressure rise measurement (PRM) to assess the progress of the primary drying phase. All these parameters except PRM can be monitored continuously and are indirect measures of sublimation rate. See Figure 3a for an example of these measurements along with product temperature data on production scale. During early stages of primary drying when the sublimation rate is high, the condenser temperature is high due to large water load, the chamber-condenser pressure differential is high due to high volumetric flowrate of water vapor and the nitrogen bleed rate is low since the chamber pressure is being maintained mostly by the water vapor. As the primary drying gets completed and the ice sublimation rate decreases, the condenser temperature and the chamber-condenser pressure differential decrease and the nitrogen bleed flow rate increases.

A PRM, on the other hand, is a discrete measurement where the rate of pressure increase in the chamber is measured while it is temporarily isolated by closing the valve between the chamber and the condenser. The rate of pressure rise measured in this manner is an indication of the amount of residual water in the product. Figure 3b shows an example of PRM results as a function of primary drying duration; as the primary drying progresses and the product temperatures approach the shelf temperature (Figure 3a), the rate of pressure rise rapidly decreases. It should be noted that during PRMs in Figure 3b, the chamber pressure was allowed to increase only by 20-30 mHg and the measurements lasted only for a few seconds. This is important because, if the pressure is allowed to increase excessively while the ice sublimation phase is not complete, the product temperature could significantly increase and cause cake collapse.

The condenser temperature, chamber-pressure differential, nitrogen bleed rate and PRM data in Figure 3 correlate well with the product temperature data and demonstrate that they can be useful in monitoring the progress of a lyophilization cycle during routine production. This type of correlation can be established during engineering runs on production scale so that their use during routine production can be justified. Concluding Remarks Lyophilization involves numerous parameters pertaining to the process setpoints, equipment, manufacturing environment and product configuration. The key to successful scale-up lies in understanding the interdependencies among these parameters, knowing the formulation and the target product properties and executing a scientifically sound methodology that demonstrates production of a lyophilized product with uniform and acceptable quality. If the pilot-scale lyophilization cycle has been designed with the knowledge of production scale equipment capabilities, any surprises during scale-up can be minimized.