Nigel J. Titchener-Hooker

Decision-support tool for assessing biomanufacturing strategies under uncertainty: Stainless steel versus disposable equipment for clinical trial material preparation

This paper presents the application of a decision‐support tool, SimBiopharma, for assessing different manufacturing strategies under uncertainty for the production of biopharmaceuticals. SimBiopharma captures both the technical and business aspects of biopharmaceutical manufacture within a single tool that permits manufacturing alternatives to be evaluated in terms of cost, time, yield, project throughput, resource utilization, and risk. Its use for risk analysis is demonstrated through a hypothetical case study that uses the Monte Carlo simulation technique to imitate the randomness inherent in manufacturing subject to technical and market uncertainties. The case study addresses whether start‐up companies should invest in a stainless steel pilot plant or use disposable equipment for the production of early phase clinical trial material. The effects of fluctuating product demands and titers on the performance of a biopharmaceutical company manufacturing clinical trial material are analyzed. The analysis highlights the impact of different manufacturing options on the range in possible outcomes for the project throughput and cost of goods and the likelihood that these metrics exceed a critical threshold. The simulation studies highlight the benefits of incorporating uncertainties when evaluating manufacturing strategies. Methods of presenting and analyzing information generated by the simulations are suggested. These are used to help determine the ranking of alternatives under different scenarios. The example illustrates the benefits to companies of using such a tool to improve management of their R&D portfolios so as to control the cost of goods.

Micro biochemical engineering to accelerate the design of industrial-scale downstream processes for biopharmaceutical proteins.

The article examines how a small set of easily implemented micro biochemical engineering procedures combined with regime analysis and bioprocess models can be used to predict industrial scale performance of biopharmaceutical protein downstream processing. This approach has been worked on in many of our studies of individual operations over the last 10 years and allows preliminary evaluation to be conducted much earlier in the development pathway because of lower costs. It then permits the later large scale trials to be more highly focused. This means that the risk of delays during bioprocess development and of product launch are reduced. Here we draw the outcomes of this research together and illustrate its use in a set of typical operations; cell rupture, centrifugation, filtration, precipitation, expanded bed adsorption, chromatography and for common sources, E. coli, two yeasts and mammalian cells (GS‐NSO). The general approach to establishing this method for other operations is summarized and new developments outlined. The technique is placed against the background of the scale‐down methods that preceded it and complementary ones that are being examined in parallel. The article concludes with a discussion of the advantages and limitations of the micro biochemical engineering approach versus other methods. Biotechnol. Bioeng. 2008;100: 473–487.

Bioprocess Engineering Issues That Would Be Faced in Producing a DNA Vaccine at up to 100 m3 Fermentation Scale for an Influenza Pandemic

The risk of a pandemic with a virulent form of influenza is acknowledged by the World Health Organization (WHO) and other agencies. Current vaccine production facilities would be unable to meet the global requirement for vaccine. As a possible supplement a DNA vaccine may be appropriate, and bioprocess engineering factors bearing on the use of existing biopharmaceutical and antibiotics plants to produce it are described. This approach addresses the uncertainty of timing of a pandemic that precludes purpose‐built facilities. The strengths and weaknesses of alternative downstream processing routes are analyzed, and several gaps in public domain information are addressed. The conclusion is that such processing would be challenging but feasible.

The use of windows of operation as a bioprocess design tool

Bioprocess design problems are frequently multivariate and complex. However, they may be visualised by a graphical representation of the design constraints and correlations governing both the process and system under consideration, namely windows of operation. Windows of operation exist at all stages of process design and find use both in the identification of key constraints from limited information, and also, with more detailed knowledge, the sensitivity of a process to design or operating changes. In this way windows of operation may be used to help understand and optimise a bioprocess design. In this paper the formulation, development and application of windows of operation is discussed for a range of biological processes including fermentation, protein recovery and biotransformation.

Modelling biopharmaceutical manufacture: Design and implementation of SimBiopharma

Abstract This paper presents the implementation of a conceptual framework, for modelling a biopharmaceutical manufacturing plant, into a prototype decision-support tool, S im B iopharma . The tools scope covers the ability to evaluate manufacturing alternatives in terms of cost, time, yield, resource utilisation and risk. Incorporating uncertainty means that investment appraisal can be based on both the expected outputs and the likelihood of achieving them. A hierarchical approach to represent the key activities in a manufacturing process is introduced. Emphasis is placed on how a closer integration of bioprocess and business process modelling can be achieved by capturing common information in an object-oriented environment, G2 (Gensym Corporation, Cambridge, MA). The key features of S im B iopharma are highlighted; these include interactive graphics, task-oriented representation and dynamic simulation which create a much more flexible environment for modelling processes. Examples of typical outputs generated by S im B iopharma , when addressing the impact of manufacturing options on strategic operational and financial indicators, are given.

The effects of material properties and fluid flow intensity on plasmid DNA recovery during cell lysis

The disruption of recombinant E. coli cells containing a 76.8 kb plasmid DNA was achieved by the chemical lysis method in a coaxial cylinder rheometer which allowed in situ measurements of rheological changes to be carried out as the lysis reaction proceeded. For the cases studied the cell lysis time was found to be approximately 30 s. Moreover, the release of intracellular material produced a mixture with shear thinning flow properties, the extent of non-Newtonian flow was found to depend on the shear rate used during the cell lysis operation. On neutralisation, the lysate produced a highly flocculated and shear sensitive gel which floated on the top of the liquor containing the plasmid DNA. Small amplitude oscillatory data were obtained showing the viscoelastic properties of the gel matrix. Experimental data were also obtained on the shear sensitivity of the plasmid DNA recovered using a purpose-built rotating disk shear device. Shear rates of the order of 106 s-1 were generated in the device and were confirmed by CFD analysis of the prevailing flow field. Tests carried out with 20 and 29 kb plasmid DNA showed that both plasmids were susceptible to shear damage. The extent of shear damage increased with plasmid size and as the ionic strength of solution decreased.

A physical model of high-pressure disruption of bakers' yeast cells

A model is presented for the disruption of microbial cells in a high-pressure homogeniser. It assumes that cell deformation and breakage due to elongational stresses occur in the high-stress zones close to the surfaces of the valve rod and the impact ring of the homogeniser. The equation based on the proposed model predicts an inverse cube-root dependency of the maximum diameter of cells that remain unbroken upon the pressure drop across the valve, and suggests that the maximum whole cell diameter will increase with an increase in the mechanical integrity of the cell walls. Experimental data are provided for the disruption of batch and continuously grown cells of Saccharomyces cerevisiae. The results from these experiments support, and are interpreted in terms of the predictions from, the proposed model.

Application of a decision-support tool to assess pooling strategies in perfusion culture processes under uncertainty.

Biopharmaceutical manufacture is subject to numerous risk factors that may affect operational costs and throughput. This paper discusses the need for incorporating such uncertainties in decision‐making tools in order to reflect the inherent variability of process parameters during the operation of a biopharmaceutical plant. The functionalities of a risk‐based prototype tool to model cost summation, perform mass balance calculations, simulate resource handling, and incorporate uncertainties in order to evaluate the potential risk associated with different manufacturing strategies are demonstrated via a case study. The case study is based upon the assessment of pooling strategies in the perfusion culture of mammalian cells to deliver a therapeutic protein for commercial use. Monte Carlo simulations, which generate random sample behaviors for probabilistic factors so as to imitate the uncertainties inherent in any process, have been applied. This provides an indication of the range of possible output values and hence enables trends or anomalies in the expected performance of a process to be determined.

GROWTH-INDEPENDENT BREAKAGE FREQUENCY OF PROTEIN PRECIPITATES IN TURBULENTLY AGITATED BIOREACTORS

Abstract A model is provided to describe the influence of hydrodynamic conditions on the breakage of protein precipates in turbulent suspension. To examine the applicability of the model, experiments are performed to measure the breakage of soya protein precipitates in 0.29 m diameter vessel at impeller Reynolds numbers ranging from 3.6 × 10 4 to 5.6 × 10 5 and with initial protein concentrations of 0.35, 3.5 and 35 kg/m 3 . An analysis of the experimental results based on the proposed model shows that the initial frequency of breakage of protein precipitates is a unique function of the energy dissipation rate in the vessel, encompassing both effects of impeller diameter and impeller speed. The model accurately decribes the influence of both energy dissipation rate and protein concentration on the initial breakage frequency of the aggregates.

Adapted Ultra Scale-Down Approach for Predicting the Centrifugal Separation Behavior of High Cell Density Cultures

The work presented here describes an ultra scale‐down (USD) methodology for predicting centrifugal clarification performance in the case of high cell density fermentation broths. Existing USD approaches generated for dilute systems led to a 5– to 10‐fold overprediction of clarification performance when applied to such high cell density feeds. This is due to increased interparticle forces, leading to effects such as aggregation, flocculation, or even blanket sedimentation, occurring in the low shear environment of a laboratory centrifuge, which will not be apparent in the settling region of a continuous‐flow industrial centrifuge. A USD methodology was created based upon the dilution of high solids feed material to ∼2% wet wt/vol prior to the application of the clarification test. At this level of dilution cell‐cell interactions are minimal. The dilution alters the level of hindered settling in the feed suspensions, and so mathematical corrections are applied to the resultant clarification curves to mimic the original feed accurately. The methodology was successfully verified: corrected USD curves accurately predicted pilot‐scale clarification performance of high cell density broths of Saccharomyces cerevisiae and Escherichia coli cells. The USD method allows for the rapid prediction of large‐scale clarification of high solids density material using millilitre quantities of feed. The advantages of this method to the biochemical engineer, such as the enabling of rapid process design and scale‐up, are discussed.