Cell Culture Engineering

Offers a comprehensive overview of cell culture engineering, providing insight into cell engineering, systems biology approaches and processing technology In Cell Culture Engineering: Recombinant Protein Production, editors Gyun Min Lee and Helene Faustrup Kildegaard assemble top class authors to present expert coverage of topics such as: cell line development for therapeutic protein production; development of a transient gene expression upstream platform; and CHO synthetic biology. They provide readers with everything they need to know about enhancing product and bioprocess attributes using genome-scale models of CHO metabolism; omics data and mammalian systems biotechnology; perfusion culture; and much more. This all-new, up-to-date reference covers all of the important aspects of cell culture engineering, including cell engineering, system biology approaches, and processing technology. It describes the challenges in cell line development and cell engineering, e.g. via gene editing tools like CRISPR/Cas9 and with the aim to engineer glycosylation patterns. Furthermore, it gives an overview about synthetic biology approaches applied to cell culture engineering and elaborates the use of CHO cells as common cell line for protein production. In addition, the book discusses the most important aspects of production processes, including cell culture media, batch, fed-batch, and perfusion processes as well as process analytical technology, quality by design, and scale down models. -Covers key elements of cell culture engineering applied to the production of recombinant proteins for therapeutic use -Focuses on mammalian and animal cells to help highlight synthetic and systems biology approaches to cell culture engineering, exemplified by the widely used CHO cell line -Part of the renowned "Advanced Biotechnology" book series Cell Culture Engineering: Recombinant Protein Production will appeal to biotechnologists, bioengineers, life scientists, chemical engineers, and PhD students in the life sciences.

Autorentext
Gyun Min Lee, PhD, is Professor at the Department of Biological Sciences at KAIST, South Korea, and heads the Animal Cell Engineering Laboratory. He is also Scientific Director at the Novo Nordisk Foundation Center for Biosustainability at the Technical University of Denmark.

Helene Faustrup Kildegaard, PhD, is a senior researcher and Co-PI for the CHO Cell Line Engineering and Design section at the Novo Nordisk Foundation Center for Biosustainability at the Technical University of Denmark (DTU).

Inhalt

About the Series Editors xvii

1 Platform Technology for Therapeutic Protein Production **1
Tae Kwang Ha, Jae Seong Lee, and Gyun Min Lee

1.1 Introduction 1

1.2 Overall Trend Analysis 3

1.2.1 Mammalian Cell Lines 3

1.2.2 Brief Introduction of Advances and Techniques 5

1.3 General Guidelines for Recombinant Cell Line Development 6

1.3.1 Host Selection 6

1.3.2 Expression Vector 7

1.3.3 Transfection/Selection 7

1.3.4 Clone Selection 8

1.3.4.1 Primary Parameters During Clone Selection 8

1.3.4.2 Clone Screening Technologies 9

1.4 Process Development 9

1.4.1 Media Development 10

1.4.2 Culture Environment 10

1.4.3 Culture Mode (Operation) 10

1.4.4 Scale-up and Single-Use Bioreactor 11

1.4.5 Quality Analysis 12

1.5 Downstream Process Development 12

1.5.1 Purification 12

1.5.2 Quality by Design (QbD) 13

1.6 Trends in Platform Technology Development 14

1.6.1 Rational Strategies for Cell Line and Process Development 14

1.6.2 Hybrid Culture Mode and Continuous System 15

1.6.3 Recombinant Human Cell Line Development for Therapeutic Protein Production 16

1.7 Conclusion 17

Acknowledgment 17

Conflict of Interest 17

References 17

2 Cell Line Development for Therapeutic Protein Production **23
Soo Min Noh, Seunghyeon Shin, and Gyun Min Lee

2.1 Introduction 23

2.2 Mammalian Host Cell Lines for Therapeutic Protein Production 25

2.2.1 CHO Cell Lines 25

2.2.2 Human Cell Lines 26

2.2.3 Other Mammalian Cell Lines 27

2.3 Development of Recombinant CHO Cell Lines 27

2.3.1 Expression Systems for CHO Cells 28

2.3.2 Cell Line Development Process Using CHO Cells Based on Random Integration 28

2.3.2.1 Vector Construction 29

2.3.2.2 Transfection and Selection 30

2.3.2.3 Gene Amplification 30

2.3.2.4 Clone Selection 31

2.3.3 Cell Line Development Process Using CHO Cells Based on Site-Specific Integration 32

2.4 Development of Recombinant Human Cell Lines 34

2.4.1 Necessity for Human Cell Lines 34

2.4.2 Stable Cell Line Development Process Using Human Cell Lines 35

2.5 Important Consideration for Cell Line Development 36

2.5.1 Clonality 36

2.5.2 Stability 36

2.5.3 Quality of Therapeutic Proteins 37

2.6 Conclusion 38

References 38

3 Transient Gene Expression-Based Protein Production in Recombinant Mammalian Cells **49
Joo-Hyoung Lee, Henning G. Hansen, Sun-Hye Park, Jong-Ho Park, and Yeon-Gu Kim

3.1 Introduction 49

3.2 Gene Delivery: Transient Transfection Methods 50

3.2.1 Calcium Phosphate-Based Transient Transfection 50

3.2.2 Electroporation 51

3.2.3 Polyethylenimine-Based Transient Transfection 52

3.2.4 Liposome-Based Transient Transfection 52

3.3 Expression Vectors 53

3.3.1 Expression Vector Composition and Preparation 53

3.3.2 Episomal Replication 53

3.3.3 Coexpression Strategies 54

3.4 Mammalian Cell Lines 54

3.4.1 HEK293 Cell-Based TGE Platforms 55

3.4.2 CHO Cell-Based TGE Platforms 56

3.4.3 TGE Platforms Using Other Cell Lines 58

3.5 Cell Culture Strategies 58

3.5.1 Culture Media for TGE 58

3.5.2 Optimization of Cell Culture Processes for TGE 59

3.5.3 *q*p-Enhancing Factors in TGE-Based Culture Processes 59

3.5.4 Culture Longevity-Enhancing Factors in TGE-Based Culture Processes 59

3.6 Large-Scale TGE-Based Protein Production 60

3.7 Concluding Remarks 62

References 62 **4 Enhancing Product and Bioprocess Attributes Using Genome-Scale Models of CHO Metabolism </...