1. The Systematic Production of Cells for Cell Therapies
Daniel C. Kirouac, Peter W. Zandstra 
Cell Stem Cell 
Volume 3, Issue 4, Pages (October 2008)
DOI: /j.stem
Copyright © 2008 Elsevier Inc. Terms and Conditions

2. Figure 1 Roadmap for the Development of Cell Therapies
Cell therapy development can be divided into four phases: discovery, process optimization, production, and therapeutic delivery. In the discovery phase, the main issue is product characterization. Functional assays and cellular and molecular profiling are useful tools in this phase. The process optimization phase involves quantifying the relationship between culture parameters and cell output. The rational design of experiments and high-throughput screening using microculture platforms can be used to generate empirical cell-based models, which may be integrated with molecular profiling technologies to develop more mechanistic, molecular-based models. Considerations in the production phase include the scale-up strategy and quality control. The final phase is the therapeutic delivery of the cell product. It should be noted that the development phases feed back on one another—during process optimization, biological discoveries may be made, and during production, the design space will become better defined.
Cell Stem Cell 2008 3, DOI: ( /j.stem )
Copyright © 2008 Elsevier Inc. Terms and Conditions

3. Figure 2 Multidimensional Optimization of Cell Production Processes
Multidimensional optimization problems are an intrinsic part of process design engineering wherein multiple (often conflicting) outputs must be simultaneously maximized/minimized as a function of multiple (nonlinear) input parameters (the “design space”). For cell therapy production processes, one desires to minimize cost (reagents, time, and operator requirements) while maximizing product yield (target cells/day) and quality (cell purity, functionality, sterility, etc.). Depicted is the response surface of a theoretical objective function to be minimized (cost) as a function of two closely linked process parameters, selective pressure (in this case the rate at which undesired genetic changes occur as a function of the culture conditions) and cell output (the rate at which the target cells are generated).
Cell Stem Cell 2008 3, DOI: ( /j.stem )
Copyright © 2008 Elsevier Inc. Terms and Conditions

4. Figure 3 Bioprocess Control Strategy
In order to maintain a bioprocess at optimal conditions, disturbances must be continually corrected by varying controllable input parameters. “Disturbances” for cell therapy production pertain to internal culture dynamics (nutrient depletion, endogenous protein secretion, etc.) and external uncontrolled parameters (biological variability, media fluctuations, etc.). Culture outputs (cell density, cell surface antigen expression, etc.) and process inputs (input cell populations) can be continually monitored using online sensors. Based on mathematical models relating cell outputs to culture parameters, controllable input parameters (feeding rate, air sparging, growth factor concentrations, etc.) can be adjusted based on feedback (FB) and feed-forward (FF) signals to maintain the culture environment within allowable ranges.
Cell Stem Cell 2008 3, DOI: ( /j.stem )
Copyright © 2008 Elsevier Inc. Terms and Conditions

5. Figure 4 A “Two-Phase” Bioreactor
A stirred tank bioreactor can be used to cultivate adherent cells or aggregates on engineered scaffolds or surfaces. Bulk macroenvironmental parameters (i.e., temperature, pH, pO2, glucose concentration, lactose concentration, etc.) are monitored and controlled using online sensors. Microenvironmental, or cell-level, parameters (endogenous factor concentrations and cell-cell and cell-ECM interactions) are regulated using the scaffold design. In this example, an immobilized ligand gradient within the scaffold directs tissue-level organization of the cells.
Cell Stem Cell 2008 3, DOI: ( /j.stem )
Copyright © 2008 Elsevier Inc. Terms and Conditions

6. Figure 5 In Vitro Cell-Based Functional Bioassay
Engineered tissue may serve as a powerful platform to assay cells produced in bioprocesses. t0: cells produced in the bioprocess are seeded onto a human tissue engineered construct. t1: the relevant cell population(s) adhere/integrate within the tissue construct. t2: at a defined time point following culture the functional activity of the resulting cell population is evaluated using morphological (i.e., cell number, distribution, and colony size), functional (i.e., electromechanical properties, enzyme activity), and/or biochemical (i.e., gene expression, protein secretion, cell surface antigen expression) readouts predictive of the human clinical response.
Cell Stem Cell 2008 3, DOI: ( /j.stem )
Copyright © 2008 Elsevier Inc. Terms and Conditions