Madhusudan P Dabhole
Group Manager – BioProcess,
Richcore Life Sciences Ltd, Bangalore, Karnataka, India
The manufacturing of recombinant products by fermentation and purification in stainless steel vessels has seen the transition from small scale to large scale and further to single use disposable technology. The requirement to develop and modulate the process has arisen from the cost and manufacturers need to move the facility on mobile platforms. The review describes the strategies and considerations for Single Use Disposable Technology. Recombinant proteins are widely used for treatment of various diseases and disorders. Single Use Disposable Technology makes it promising to produce and formulate these proteins from bench scale to commercial level in a shorter span of time so that it can reach the physician and patients.
REFERENCE ID: PHARMATUTOR-ART-2268
The production of recombinant proteins can be broadly divided into four general steps: Cloning the DNA of interest in a suitable vector under an adequate promoter; transforming and stabilizing the host cells; biosynthesis of the desired protein under controlled conditions; and recovery and purification of the r-product and comparison with its native counterpart. For an adequate industrial production, all four steps have to incorporate safety and regulatory issues into an integral process.
The demand for recombinant products has increased exponentially during the last fifteen years and the trend should continue in the present decade. In 1998 the market for medicines derived from r-DNA technologies was worth over $13 billion and had a growth rate of 14 per cent per year, whereas the average growth rate of the whole pharmaceutical market was only 6 per cent. Today almost 1200 new biopharmaceutical products are in clinical trials and many of them are expected to reach the market within this decade. The total market of recombinant products is expectedto increase 12 per cent annually during the next 8 years and reach sales for over $300 x 109 by year 2050. In this expanding scenario, the markets of specialty chemicals and agricultural products derived from biotechnology are expected to grow at the fastest rates. The specialty chemicals sector is very active innew r-product development, and due to less regulatory burdens, more r-products are expected to be approved in the near future. In terms of number of products the specialty chemical sector is the most important since it produces for industrial applications more than 50 enzymes, many genetically engineered. Furthermore, industrial applications of r-products usually require very large quantities of protein; therefore very large-scale operations must be performed. Accordingly, highly productive processes are particularly required in this sector for maintaining economic viability
Biopharmaceutical manufacturing typically involves a limited number of similar unit operations and processes — the primary differences among processes being the number, size and sequence of the unit operations. However, the facilities are large, complex and capital-intensive, with multiple interacting systems and operational preferences, so making comparisons between SU/DT and traditional systems is difficult and subjective. Such an evaluation, therefore, needs to take a whole-facility approach.
Single-use/disposable technology (SU/DT) has emerged over the past decade as a cost-effective and flexible basis for biopharmaceutical manufacturing. It has moved beyond the limited applications of culture bags, liquid storage bags, and sampling devices, and now includes more unit-operation-based capabilities, such as cartridgefiltration, depth filtration, ultra filtration, and chromatography.
SU/DT is now widely considered state-of-the-art for applications involving the bioreactor train (volumes up to 500 L), cartridge filtration (sizes up to single 30-in. elements), depth filtration (size depends on element type, limited to single-element configurations), and process hold (disposable bags with volumes up to 1,000 L for portable configurations or 2,500 L for fixed configurations). Pre-packed chromatography columns, microfiltration and ultrafiltration systems are being developed rapidly, but are still considered to be in a developmental stage. In addition, several hybrid systems combine both reusable and disposable components for instance, the emerging cassette style depth filters that have a disposable filter element and a reusable base, and septum-style samplers that employ disposable sample bags attached to a reusable vessel.
The single use disposable technology will be viable for multiproduct in a facility as compared to one or even two products for recombinant proteins depending on the volumes and output taking profit margin into consideration.
The timing of market it takes to bring new drugscontinues to be an important goal for pharmaceutical companies. There are several approaches that are taken with the overall goal of bringing more drugs into commercialization phase. Single-use (SU) technology is one of the strategies being adopted to reduce overall drug development time. SU technology brings significant advantages of reduced capital costs, faster construction and installation, reduced processing cycle times, and elimination of the need for post-use equipment cleaning and verification.Starting with the use of plastic ware such as pipettes, petri dishes, and t-flasks, disposable components are being increasingly incorporated in the laboratory environment. With the development of SU bioreactors at the 2,000 L scale (Xcellerex XDR), chromatography systems, and systems for micro- and ultrafiltration, disposable equipment continues to replace fixed stainless steel equipment in manufacturing plants.
Criteria for Evaluating SU UF–DF Systems
Currently there are several SU UF/DF systems available. An evaluation was made to choose an appropriate system based on GMP production requirements, operational needs, budget, and timeline. These criteria included:
- Operation: The system should have the capacity to handle 5 m2 membrane size, 10–50 L retentate tank working volume, >80 L/h/m2 (LHM) feeding flux, and differential pressure (ΔP) and transmembrane pressure (TMP) controls at 0–20 psi. The retentate tank should have a mixing device to avoid localized concentration during the operation.
- Process monitoring, control, and data management: The process should be able to be controlled by constant TMP and ΔP. The pressures, flow rates, process phases, and other process parameters should be able to be monitored and recorded in real time. Data management should meet 21 CFR Part 11 compliance.
- Disposable parts: There should be no or a limited number of bio-compatibility issues for any disposable parts that contact product and process buffers. The levels of leachables and extractables should be in the acceptable safety range for clinical drug substances.
- Equipment availability and vendor support: The product should be readily available. An integrated and off-the-shelf system is preferred because it saves time on equipment validation and meets short project timelines. The manufacturer should have a good record for on-time equipment delivery and reliable technical support. The manufacturer should have the capability to consistently supply high quality accessories and consumable items.
- Cost: The price of the equipment and disposable items should be reasonable to reduce the overall cost of production when implementing a new UF–DF system in a GMP production facility.
The following economic analysis of an ultrafiltration process demonstrates the savings that can be achieved when using single-use filtration cassettes instead of reusable ones. The analysis takes into consideration the cost of consumables, such as water, CIP solutions, and buffer solutions, as well as labor and overhead costs associated with running a TFF process in a cGMP process. These costs can vary from site to site, so individual site costs can be input to create a customized model.
The membrane surface area of the filtration cassettes chosen for this model was 2.5 m2, an area sufficient to process batch sizes up to approximately 1,000 L. This amount of membrane surface area typically would be used for one ultrafiltration process step in clinical-scale production. In clinical-scale and contract manufacturing, often only four to six batches of product are processed per campaign and then the reusable cassettes are either placed in storage or discarded. This scenario is represented in this particular economic analysis.
Consumables, including the filtration cassettes, buffers, water, and CIP solutions used during the process, also are accounted for in this model.
Cassette costs vary from different manufactures, so an average cost of the reusable cassettes was used. Typically reusable cassettes cost approximately $3,600 per m2. By way of comparison, single-use cassettes are approximately 20% of the cost of reusable cassettes or approximately $800 per m2.
In addition to the cassettes, various buffer solutions are used for the filtration process. Some of the buffers are used for the membrane equilibration and diafiltration operations. More importantly, with reusable TFF, a significant amount of these buffers are used for the CIP portion of the process. In addition to these CIP solutions, purified water is used for flushing the reusable cassettes. These CIP solutions and the need for purified water for flushing the cassettes are eliminated when single-use cassettes are used.
Lastly, labor and overhead costs are considered. These labor and overhead costs (also referred to as factory overhead, factory burden, and manufacturing support costs) refer to both direct and indirect factory-related costs that are incurred when the product is manufactured. Along with costs such as direct material, the cost of labor and overhead must be assigned to each batch produced so that the cost of goods are valued and reported accurately. The overhead includes such things as the electricity used to operate the factory equipment, depreciation on the factory equipment and building, factory supplies, and factory personnel not included as direct labor. Once these costs have been tabulated, they have a significant impact on the cost of the ultrafiltration process
Implementing single-use cassettes can eliminate many of the non-value–added steps of pre-use and post-use CIP and as a result the overall time of the ultrafiltration process is reduced. As a result, there is increased productivity and the overall cost of manufacturing is reduced.
The major concerns for biological manufacturers in the case of a new facilitydesign or indeed within their existing processes are
Safety: The main GMP (Good Manufacturing Practice) deficiency reported from biopharmaceutical plant audits is linked to cross contamination, which represents 15% of total deficiencies.
Maintenance: Biologicals plants are extremely complex and require expensive maintenance.
Long construction time:Average construction time is 2 - 3 years or more followed by extensive validation - the commissioning phasemaytake several months. Companies maylose the ground to get their drug on the market aheadof the competition.
Process costs:One of the biggest costs in bio manufacturing is the cost of transfer ofsterile fluids (such as product and reagents) through different process steps located in different parts of the facility: traditionally, thelogistics of fluid transfer has been handled through product piping, stainless steel vessels,routing manifolds and valves. All this equipment has to be cleaned and sterilized.Equipment validation is required before re-use.
The first single-use bioreactors, known as wave reactors, were developed in the 1970s (DiBlasi et al., 2006). Although these systems are efficient and easy to use, they lack the common Geometry seen in the stirred tank reactors, which are the systems of choice in industry (Nienow, 2006).
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The second single use bioreactor usually referred to as a “liner style” and functions as a typical stirred tank bioreactor and was first used in 2006 by ThermoFisher Scientific (Selker and Paldus, 2008). A disposable bag is typically comprised of mostly polyethylene is used as a Liner in a cylindrical steel tank. The bag has an integrated impeller for mixing as well as sampling ports and sparger system. The steel tank is conventionally open at the top and view ports can be added to optically monitor the culture.
Figure 1: A Biostat STR 200 Bioreactor (source:Anne Weber et al, Sartorius, BioprocessInternational 11(4) s April 2013)
One of the studies consideredduring single use disposable technology of the several aspects of the environmental footprint, including carbon output and the usage of water and land. The disposables-based facility reduces the overall environmental impact despite the creation of solid plastic waste. As a consequence, there is a substantial decrease in carbon footprint.
It can be seen that the greatest impact of disposables results from reduced water requirements. For the stainless steel facility, water usage is one of the most significant contributors to the carbon footprint (excluding driving to work). The key consequence of the extensive use of disposables, therefore, is the removal of significant requirements for high quality water by eliminating clean-in-place (CIP) operations. This gives rise to one of the key impacts in terms of reducing the facility's overall carbon footprint: water use.
It is instructive to see that by far the largest contributor to carbon footprint of all categories is workers driving to work. This is significant because disposables lower headcount and therefore reduce carbon footprint by reducing the number of workers driving to work. This suggests that if we are truly going to reduce the impact of these facilities, we must seek more energy-efficient transportation systems. Further developments from single-use suppliers, such as working with recyclable or separable materials and revising current packaging methods for single-use systems, will reduce further environmental impact of disposables.
Single Use disposable technology is one of the promising innovative processes in upstream and downstream production. It can be concluded that the land and environmental footprint will reduce considerably by adapting this technology. Cost saving in terms of manufacturing and man power will be achieved only if the process yields are as per the desired expectations. The Single Use systems need to be continuously and thoroughly tested over a period of batches to accomplish both the physical robustness of the system and the product safety.
Dr Madhusudan P Dabhole holds a Ph.D in Microbiology, Mumbai University and PGDBA from Symbiosis, Pune with 15 years of industrial fermentation experience in classical and recombinant products.
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