Hélène Pora

Quality assurance (QA) is vital for vaccine manufacturers, typically accounting for 70% of total vaccine development and production time.

  Compounding this challenge are the time-to-market issues vaccine manufacturers face in protecting populations from emerging diseases such as avian flu. For these and other reasons, total US vaccine manufacturers dropped from 25 in 1973, to approximately five at present.

  The introduction of good manufacturing practices (GMPs) in 1980, which caused manufacturing costs to skyrocket, was also responsible for the exodus of companies from the market.

  The attempt in 1993–1994 to provide universal healthcare in the US, which forced 50% discounts on vaccine makers in exchange for government-purchasing of one-third of the nation's supply, caused the vaccine supplier market to shrink even further.

However, the tides have turned, especially for the five biggest vaccine makers. This is partly because of the pipeline of new prophylactic and therapeutic vaccines, particularly those targeting cancer and AIDS. Companies involved in vaccine manufacture would now agree that the rewards exceed the challenges in this market. 

This sentiment is echoed in the following statement concerning one of the vaccine industry's five dominant players: "Vaccines now represent an important element of GlaxoSmithKline's growth strategy, and were, uncharacteristically, the main focus of a pipeline update during the summer of 2005."

While larger manufacturers with extensive experience and the economies of scale to profit from the lower margins associated with this class of drug have certain advantages over smaller makers, vaccine producers of any size can benefit from reducing cleaning/cleaning validation steps, and improving separation and purification processes. 

Wide-scale adoption of disposable technologies represents one of the most sweeping changes to take place in the biopharmaceutical industry. This is because disposables eliminate the need for cleaning and cleaning validation, processes that are time-consuming, expensive and wrought with potential compliance pitfalls. In fact, 30–50% of all FDA warning letters are cleaning related.

  Considering the time vaccine manufacturers dedicate to QA, the complete omission of this particularly challenging task can drastically speed up production.  

Disposable technologies are used in numerous steps throughout the vaccine production process. At connection points, disposable technologies enable two fluid pathways to be connected in a matter of seconds without the need for a laminar air-flow cabinet or tubing welder. 

Disposable filters and chromatography membranes minimize opportunities for operator error, and can be disposed of after use to avoid cleaning and cleaning validation. When combined with single-use bags and tubing, these technologies form fully-integrated disposable systems that can be supplied preassembled and presterilized by gamma irradiation to further reduce process steps. 

Advances in drug development and production not only come from new technologies, but also from the application of different technologies to existing processes. One example is favouring ultrafiltration over size exclusion chromatography (SEC) to separate and purify conjugate vaccines, which protect infants and young children from infectious diseases such as meningitis and invasive pneumonia. There are at least 30 steps involved in conjugate vaccine production, and the use of ultrafiltration instead of SEC during fractionation for protein and saccharide components can significantly reduce this number and the complexity of the separation step. 

Capture and purification steps in virus- and plasmid-based gene therapy and vaccine purification applications are additional areas of inefficiency in drug production. A disposable, prepacked membrane column can remove contaminants of up to 100 times the speed of resin-based chromatography, while providing efficient capture of large molecules. This is because conventional chromatography is highly inefficient and limited because of the speed that large molecules, such as plasmids, diffuse through the media's pore structure. However, anion-exchange membranes that use a microporous membrane with active quaternary ammonium chemistries linked directly to the membrane surface can overcome these inefficiencies.  

By using three-dimensional structures with open (0.8 μm) pores, active chemistry groups are immediately available for binding. With over 200 novel viral and DNA-based vaccines in development today, this technology has the potential to change the timetable for bringing these new vaccines to market. 

While egg-based vaccine production has certain limitations in terms of development time, downstream processing can be a significant source of new efficiencies. The move towards cell-based vaccine development will further speed the production timetable, especially as disposable technologies can be used for a greater number of steps in this process. While single-use technologies can be used for filtration, storage and at connection points in egg-based vaccine production, their use multiplies in cell-based applications. For example, disposable technologies can be used for media preparation, clarification, buffer preparation, capture and polishing chromatography steps, filling in downstream processes, and for venting and cell harvesting in upstream processes.  

As newer vaccine production methods take hold and more effective technologies are applied to these processes, the industry will be better equipped to handle vaccine demand, whether to stem a pandemic outbreak, protect against bioterrorism or ensure that everyone receives their annual flu shot.  

 is marketing director for biopharmaceuticals at Pall Life Sciences (France).

4. Presentation by H. Pora at Phacilitate Conference, January 2005. 

Articles

While single-use technologies can be used for filtration, storage and at connection points in egg-based vaccine production, their use multiplies in cell-based applications.

Improving speed and quality in vaccine production

The medical industry was the first to understand the benefits of using disposable devices, such as needles and syringes, to prevent risks of cross contamination. The technology was then extended to blood transfusion activities, and it was only 10–15 years ago that the biopharmaceutical industry started to use disposables. Initially, most of the applications were limited to storage, involving bags, tubing and filter capsules. Since then, significant progress has been made in the polymer and plastics industry; in particular, a number of organic polymers have been developed that are resistant to gamma irradiation, autoclaving and even sterilization-in-place, rendering the technology attractive and usable by the biopharmaceutical industry. Now, the industry is moving beyond storage-focused disposable technologies to more complex processing applications. 

The adoption of single-use technologies has considerably increased with the emergence of a strong biotech sector. Disposable technology is seen as a boon to this industry because of the potential speed of implementation; the relative ease of the facility design; the reduction in validation requirements; and the decrease in capital investment and footprint. The idea is to adopt disposable bioprocessing operations very early in process development, which can be scaled up to large volumes. The scale at which disposable technology can be used has been questioned for a long time, but now the availability of bags, filters and devices that can handle thousands of litres is setting new capacity limits.  			

Suppliers are helping to drive the success of the disposables paradigm by keeping construction materials consistent across product and scale ranges. This reduces revalidation requirements and simplifies scale-up. The result is that disposable technology is now seen as a valid possibility from the initial manufacturing of clinical batches to full production scale, and is poised to be the technology of choice for all specific patient therapies.  

Disposable technologies play a key role in minimizing any risk of cross contamination, which has been a strong driver for adoption by contract manufacturing organizations (CMOs). However, the time and cost benefits associated with the elimination of steam sterilization validation, cleaning and cleaning validation, as well as the reduction in the number of necessary connections may be seen as the strongest benefits for the broader pharmaceutical industry.  

Cleaning validation is seen as a major hurdle in the biopharmaceutical industry. In fact, a high proportion of warning letters issued by FDA contain at least one remark pertaining to cleaning or cleaning validation (e.g., in 2002, this was close to 50% of cases). 

Despite the relative ease of use of disposables, adopting and implementing this processing model requires careful evaluation. Table 1 is designed to help biotech/biopharm companies evaluate both process and cost models, as well as weigh the advantages/disadvantages and potential risks of adopting this model. Successful implementation also requires a high degree of commitment from management, quality assurance and processing teams. Some additional considerations to note include selection of the right materials and components, assembly design, developing a solid relationship with suppliers, training users, and developing an understanding of quality requirements.  

 Table 1. Comparison of hard pipe versus disposable factory. 			

For a given bioprocessing unit, materials and components must be carefully selected. Product compatibility must be evaluated, and process parameters such as temperature and pressure along with sterilization methods need to be clearly defined. It is also important to look at in process measurements so that the appropriate sensor technology is chosen.  

Criteria for establishing proper disposable system ergonomics include: 					

Tubing — length, quality, attachment to the equipment through cable ties or improved methods.  

Elements to be integrated — for example, bags, filters and chromatography units. 

Connections — how to minimize the number of sterile connections, connection and disconnection capabilities. 

Sampling requirements during the process.

Despite the fact that single-use systems significantly reduce validation requirements, they still need testing and validating. Initial testing involves checking the suitability of all components for the application and their relative position in the system. For example, operators should confirm that cable ties, where used, are in the right position and that the general assembly is strong enough to withstand handling and operating requirements. The entire system should also be tested for leaks. 

Care should also be taken with packaging. Risk of damage during the irradiation and shipping processes must be carefully assessed too. Shipment studies can be conducted to ensure safe delivery at the user site. 

Once packaging has been defined, and assuming that gamma irradiation is chosen as the sterilization method, it will be necessary to validate the irradiation procedure. The minimum required irradiation dose is 25 Kgy, and for most components the maximum is 45 Kgy. Dose mapping of the system is necessary to ensure that irradiation is falling within the requested specifications. Shelf life of the total assembly is also an important consideration when using irradiation as a sterilization technique as it will have a significant impact on inventory management at the user's manufacturing site.  

Biocompatibility is the next aspect that should be addressed following system design. To ensure biological safety, products should be manufactured from biologically inert materials and individual components that are in contact with the fluid path should be tested against United States Pharmacopeia (USP) Class VI. Most suppliers will also provide information related to potential extractables and leachables. However, in many cases there will still be a need for further testing with the process fluid under worst case process conditions, especially in instances where there is a long contact time and/or a high surface contact area.  

For instance, during the validation of disposable sterilization filtration sets, to accurately represent process conditions, all materials must be gamma irradiated (if that is the chosen mode of sterilization) prior to extractables, adsorption and microbial challenge studies. This is crucial because even though materials such as polyethersulfone (PES), polyvinylidenedifluoride (PVDF), nylon and stabilized polypropylene are known for their low level of extractables, the sterilization method may impact the overall value.  

The validation strategy for a disposable membrane chromatography device is much simpler compared with, for example, standard chromatography using gel in a column. The process efficiency will need to be validated as well as leachables. However, neither column packing nor reuse studies must be performed, greatly simplifying this validation step. One rule of thumb users should keep in mind during validation is that leachables testing must be performed for each product and application.  

The purpose of the following validation study on a disposable filtration set was to measure and identify the extractables in a process involving the sterile filtration of a solution through a sterilizing grade filter and subsequent storage over a period of time in a bag. Parameters that were examined include:  					

To achieve this, a scaled-down version of the full size system (Figure 1) was built. One of the objectives was to make sure that tubing lengths for all different storage time points were kept the same. The disposable set was gamma irradiated at a minimum dose of 25 Kgy. The four different aqueous solutions (water for injection [WFI] pH 9.0; WFI pH 2.0; WFI pH 7.0; and 1.0 M NaOH) were tested and extractables were measured using conventional HPLC methods.  

Figure 1. Design of the extractables design study for a filter-bag assembly.			

The results obtained (Figure 2) showed that whatever the storage time, the extractables were very low. In all cases, extractables were shown to be <120 ppb. Of the four solutions tested, a high proportion of the extractables was found in bag 1, corresponding to the bag that received most of the extractables coming from the filter. This is logical because this filter is the component in the assembly with the highest surface area.  

Figure 2. Total extractables from a filter-bag assembly.			

With all solutions tested, the level of extractables varied slightly with time. More than 70 chemical compounds were identified during this study. The relative level of each compound depended on the solution that was tested as well as storage time.  

This study helped to understand how to design validation studies related to extractables with complex disposable assemblies. A scaled-down system was found to be the best approach, and the four solvents selected in the study were representative of a number of buffer solutions that were evaluated. It also demonstrated that the longest contact time could be chosen as the only test point. Given the extreme sensitivity of the analytical methods used in this test, it is essential to define a reasonable detection limit.  

Filtration of a radiolabelled product 			

The availability of disposable technology (Figure 3) on the upstream and downstream side of products from cell culture, and from small-scale to large-scale marks a new era for the manufacture of biopharmaceutical drugs. These technologies serve to speed time-to-market of new products, while enabling continuous improvement of existing processes through retrofitting, which can be done with a minimum of re-engineering and often with little validation because of material consistency.  

Figure 3. Typical biotech process - where to use disposables. 			

On the upstream side, the availability of disposable sterilizing grade filters with mycoplasma removal capabilities has created new opportunities for significant time and cost savings; serum and media manufacturers largely use this technology. A study has compared processing times between a single-use filtration system and a stainless steel assembly using multiple 30 in filter elements.

  The results showed that the disposable filters not only reduced the operation time by more than half (from 19 h to 8 h), but they also showed that the filtration line could be set up in less than 40 days compared with the 6–8 months it would have taken with a standard approach.  

				A steady flow of innovative technologies for downstream applications is also being introduced, which includes					

direct flow filtration (DFF) filters from clarification/prefiltration down to virus removal applications 

tangential flow filtration (TFF) cassettes 

The medical industry was the first to understand the benefits of using disposable devices, such as needles and syringes, to prevent risks of cross contamination. The technology was then extended to blood transfusion activities, and it was only 10–15 years ago that the biopharmaceutical industry started to use disposables. Initially, most of the applications were limited to storage, involving bags, tubing and filter capsules. Since then, significant progress has been made in the polymer and plastics industry; in particular, a number of organic polymers have been developed that are resistant to gamma irradiation, autoclaving and even sterilization-in-place, rendering the technology attractive and usable by the biopharmaceutical industry. Now, the industry is moving beyond storage-focused disposable technologies to more complex processing applications.