Preface ........................................................ xv
Acknowledgements ............................................. xvii
1 Biopharmaceuticals Today ................................... 1
1.1 Industrial Context ......................................... 1
1.2 Overview of Biopharmaceutical History ...................... 2
1.2.1 Protein drugs ....................................... 2
1.2.2 Vaccines ............................................ 4
1.3 Biopharma Business Environment ............................. 6
1.3.1 Biopharma pipeline: the promise ..................... 6
1.3.2 Launched biopharmaceuticals: delivering on
promises ............................................ 7
1.3.3 Chromatography products used for making
biopharmaceuticals .................................. 9
1.4 Key Business Issues ....................................... 10
1.4.1 Prime challenge, time and cost of new drug
development ........................................ 10
1.4.2 Other significant business challenges .............. 11
1.4.3 Biosafety and general drug safety issues ........... 14
1.4.4 Regulatory issues .................................. 15
1.5 Process Chromatography within an Industrial Context ....... 16
1.5.1 High-level managerial strategies ................... 16
1.5.2 Development project throughput ..................... 17
1.5.3 Manufacturing strategies ........................... 17
1.5.4 Technology platforms ............................... 18
1.5.5 Constraints on technology and product choices ...... 19
1.6 Summary ................................................... 20
References ................................................ 21
2 Process Capability and Production Scenarios ............... 23
2.1 Process Capability ........................................ 23
2.1.1 The process scientist's perspective ................ 23
2.1.2 The production manager's perspective ............... 23
2.1.3 How much product needs to be made? ................. 25
2.2 Production Setups ......................................... 27
2.2.1 Production setup for several hundred grams up to
ten-kilogram scale ................................. 30
2.2.2 Production setup for several tens of kilograms up
to multi-ton scale ................................. 30
Upstream process ................................... 30
Downstream process ................................. 33
2.2.3 Multi-product facilities ........................... 36
2.3 Process Capability Conclusions ............................ 37
References ................................................ 39
3 Process-Design Concepts ................................... 41
3.1 Typical Process Design for Biopharmaceuticals ............. 41
3.2 Management Framework for Process Design ................... 43
3.2.1 Defining structure and workflow for process
design ............................................. 43
3.3 Production Cells and Typical Product Characteristics ...... 47
3.3.1 Production cells and their selection ............... 47
Mammalian cells .................................... 49
Escherichia coli ................................... 49
Yeast .............................................. 50
Filamentous fungi .................................. 50
Insect cells ....................................... 50
Transgenic plants .................................. 51
Transgenic animals ................................. 51
3.3.2 Typical impurity profiles .......................... 51
Product-related impurities ......................... 51
Process-related impurities ......................... 52
3.3.3 Knowing the target molecule and its stability
window ............................................. 54
3.4 Risk Analysis and Risk Mitigation ......................... 55
3.4.1 Categories of risks, prioritization and
mitigation strategies .............................. 56
3.4.2 Mitigation of safety risks ......................... 58
3.5 Downstream Processing ..................................... 60
3.5.1 Recovery process ................................... 60
3.5.2 Purification process ............................... 61
Selection of industrial raw materials ..................... 61
3.5.3 The Capture, purification, polishing (CPP)
concept ............................................ 63
Capture stage ...................................... 66
Purification stage ................................. 66
Polishing stage .................................... 67
3.5.4 Process integration, combining steps for an
efficient process .................................. 67
3.6 Selected Downstream Processing Platform Examples .......... 69
3.6.1 Monoclonal antibodies .............................. 69
3.6.2 Plasmid DNA (pDNA) ................................. 71
3.7 Characterizing the Process, Process Understanding ......... 73
3.7.1 Design space concept, ICH Q8 ....................... 73
3.7.2 Statistical design of experiments (DoE) ............ 73
3.7.3 Process characterization ........................... 77
References ................................................ 78
4 Separation Technologies ................................... 81
4.1 Introduction .............................................. 81
4.2 Recovery .................................................. 81
4.2.1 Centrifugation ..................................... 82
4.2.2 Filtration ......................................... 83
4.2.3 Other techniques ................................... 84
Fluidized beds ..................................... 85
Protein crystallization ............................ 85
4.3 Purification .............................................. 86
4.3.1 Design principles .................................. 86
4.3.2 Chromatography resins .............................. 86
4.3.3 Selectivity and productivity of some popular
chromatographic techniques ......................... 88
Size exclusion chromatography ...................... 92
Ion exchange chromatography ........................ 96
Reversed phase chromatography ..................... 103
Hydrophobic interaction chromatography ............ 106
Affinity chromatography ........................... 109
Other modes of chromatography ..................... 113
4.3.4 Scale-up of chromatographic purifications ......... 114
Guidelines ........................................ 114
System factors .................................... 116
Examples of scale-up .............................. 116
Non-chromatographic scale factors ................. 118
4.3.5 Ultrafiltration ................................... 118
Optimization of ultrafiltration ................... 119
4.3.6 Virus filters ..................................... 120
4.4 Equipment ................................................ 121
4.5 Selecting Tools from R&D to Production ................... 122
References ............................................... 122
5 Analysis ................................................. 127
5.1 Introduction ............................................. 127
5.2 Proteins ................................................. 127
5.2.1 Identity .......................................... 127
5.2.2 Purity ............................................ 128
Structural analysis ............................... 129
Process impurities ................................ 130
Contaminants ...................................... 132
5.2.3 Quantity .......................................... 133
5.2.4 Potency ........................................... 134
5.2.5 Stability ......................................... 134
5.2.6 Assays for monoclonal antibodies .................. 135
5.3 Nucleic Acid Products .................................... 136
5.3.1 Identity .......................................... 136
5.3.2 Purity ............................................ 136
Process impurities ................................ 137
Contaminants ...................................... 138
5.3.3 Quantity .......................................... 139
5.3.4 Potency ........................................... 139
5.3.5 Stability ......................................... 139
5.4 Comparability ............................................ 140
5.5 Setting Specifications and Reference Standards ........... 140
5.6 Method Validation ........................................ 141
5.7 Process Analytical Technologies (PAT) .................... 141
References ............................................... 142
6 Cleaning and Sanitization ................................ 147
6.1 Introduction ............................................. 147
6.2 Cleaning ................................................. 147
6.2.1 Resins ............................................ 147
6.2.2 Filter media ...................................... 151
6.2.3 Equipment ......................................... 152
6.3 Decontamination of Transmissible Spongiform
Encephalopathy Agents .................................... 153
6.3.1 Resins and filter media ........................... 153
6.3.2 Equipment ......................................... I54
6.4 Sanitization ............................................. 154
6.4.1 Resins ............................................ 155
6.4.2 Filter media ...................................... 157
6.4.3 Equipment ......................................... 157
References ............................................... 158
7 Validation ............................................... 161
7.1 Introduction ............................................. 161
7.1.1 Validation terminology ............................ 161
7.2 What to do When? ......................................... I62
7.2.1 Toxicology studies ................................ 163
7.2.2 Human clinical trials ............................. 163
7.3 Validation of Downstream Processes ....................... 164
7.3.1 General considerations ............................ 164
7.3.2 Raw materials and process tools ................... 165
Acceptance criteria for chromatography resins ............ 165
7.3.3 Equipment ......................................... 168
Equipment qualification ........................... 168
Installation qualification ........................ 169
Operational qualification ......................... 170
Automated equipment qualification ................. 170
Column packing and qualification .................. 172
7.3.4 Process validation ................................ 172
Storage ........................................... 174
Leachables ........................................ 175
Cleaning and sanitization validation .............. 177
Resin and membrane lifetime ....................... 180
Small and manufacturing scales .................... 182
Clearance studies ................................. 184
7.4 Making Changes ........................................... I84
7.5 Summary .................................................. 185
Acknowledgements ......................................... 185
References ............................................... 186
8 Economics ................................................ 189
8.1 Economics: An Educational Excursion ...................... 190
8.1.1 Costs as seen from a corporate level .............. 190
8.1.2 Costs as seen from a manufacturing management
level ............................................. 192
8.1.3 Costs as seen with an interesting novel
technology in mind ................................ 196
8.2 LEAN Manufacturing, Removal of Unproductive Activities ... 197
8.3 Cost Model: Monoclonal Antibody Downstream Process ....... 199
8.4 Cost Improvement Options ................................. 202
8.4.1 Facility utilization .............................. 202
8.4.2 Cell culture: product titer and culture time ...... 204
8.4.3 Process yield ..................................... 206
8.4.4 Use of the latest resin technology ................ 208
8.4.5 Re-use strategies ................................. 209
8.4.6 Buffer consumption and cleaning buffers ........... 211
8.4.7 Exchange of one step against a cheaper one ........ 212
8.5 Impact from R&D, Platform Strategies and Technology
Outlook .................................................. 214
8.6 Conclusions, the Improvement Hierarchy ................... 215
References ............................................... 216
9 Basic Properties of Peptides, Proteins, Nucleic Acids
and Virus Particles ...................................... 219
9.1 Introduction ............................................. 219
9.2 Peptides ................................................. 219
9.2.1 Amino acid composition ............................ 220
9.2.2 Structure of peptides ............................. 221
9.2.3 Surface properties of peptides .................... 221
9.2.4 Characterization methods of peptides .............. 221
9.3 Proteins ................................................. 222
9.3.1 Structure of proteins ............................. 222
9.3.2 Surface properties of proteins .................... 223
9.3.3 Characterization methods of proteins .............. 226
9.3.4 Properties of human antibodies and antibody
fragments ......................................... 226
9.4 Nucleic Acids ............................................ 228
9.4.1 Basic structure of DNA and RNA .................... 228
9.4.2 Surface properties of nucleic acids ............... 229
9.4.3 Characterization methods of nucleic acids ......... 230
9.5 Viruses .................................................. 231
9.5.1 Structure of virus particles ...................... 231
9.5.2 Surface properties of virus particles ............. 232
9.5.3 Characterization methods for virus particles ...... 232
References ............................................... 234
10 Optimization of Chromatographic Separations .............. 237
10.1 Introduction ............................................. 237
10.2 Basic Relationships ...................................... 238
10.2.1 Resolution ........................................ 238
10.2.2 Retention ......................................... 239
10.2.3 Zone broadening ................................... 240
10.2.4 Mass transfer ..................................... 243
10.2.5 Flow resistance of packed beds .................... 243
10.3 Purification Principles .................................. 245
10.3.1 Gel filtration/size exclusion chromatography,
SEC ............................................... 245
Retention in SEC .................................. 245
Zone broadening in SEC ............................ 247
Resolution in SEC ................................. 248
Influence of experimental parameters in SEC ....... 248
10.3.2 Ion exchange chromatography, IEC .................. 252
Retention in IEC .................................. 252
Zone broadening in IEC ............................ 253
Resolution in IEC ................................. 254
Influence of experimental parameters in IEC ....... 254
10.3.3 Reversed-phase chromatography, RPC ................ 258
Retention in RPC .................................. 258
Zone broadening in RPC ............................ 258
Resolution in RPC ................................. 259
Influence of experimental parameters in RPC ....... 259
10.3.4 Hydrophobic interaction chromatography, HIC ....... 262
Retention in HIC .................................. 262
Zone broadening in HIC ............................ 262
Resolution in HIC ................................. 263
Influence of experimental parameters in HIC ....... 263
10.3.5 Affinity chromatography, AC ....................... 265
Retention in AC ................................... 265
Zone broadening in AC ............................. 266
Resolution in AC .................................. 266
Influence of experimental parameters in AC ........ 266
10.3.6 Other modes of chromatography ..................... 267
10.4 Adsorption ............................................... 267
10.4.1 Adsorption isotherms ............................. 269
Linear chromatography ............................. 271
Non-linear chromatography ......................... 271
10.5 Elution Modes ............................................ 272
10.5.1 Frontal chromatography ............................ 272
10.5.2 Elution chromatography ............................ 272
10.5.3 Displacement chromatography ....................... 273
10.5.4 Sample displacement ............................... 273
10.6 Bed Configuration ........................................ 274
10.6.1 Packed beds ....................................... 274
10.6.2 Fluidized beds .................................... 274
10.6.3 Moving beds ....................................... 275
10.7 Experimental Determination of Basic Parameters ........... 276
10.7.1 Retention ......................................... 276
Retention volume, VR .............................. 276
Mobile-phase volume, VM ........................... 278
Mobile-phase composition at elution ............... 278
10.7.2 Zone broadening ................................... 278
Peak width and plate number ....................... 278
Residence time distribution, RDT .................. 279
Vessel dispersion number .......................... 280
10.7.3 Resolution ........................................ 281
10.7.4 Mass transfer ..................................... 281
10.7.5 Capacity .......................................... 281
Ionic capacity .................................... 281
Solute capacity ................................... 281
10.8 Modelling of Chromatographic Purifications ............... 284
10.8.1 Mass transfer in chromatography ................... 285
Mass transfer in the mobile phase ................. 286
Mass transfer in the stationary phase ............. 286
10.8.2 Models for mass transfer .......................... 289
Rate model (mass balance model) ................... 289
Plate model ....................................... 290
10.8.3 Computer modelling of chromatographic
purifications ..................................... 291
10.9 Simulation of Separations ................................ 291
10.9.1 Calculation of process economy .................... 291
10.9.2 Simulations using the supplied software ........... 292
References ........................................ 292
11 Equipment ................................................ 299
11.1 Guidelines for Selecting Pilot Plant and Production
Chromatography Equipment ................................. 299
11.1.1 Dimensioning data ................................. 299
11.1.2 Functional specifications ......................... 300
11.1.3 Chemical specifications ........................... 300
11.1.4 Pressure specifications ........................... 303
11.1.5 Hygienic design ................................... 304
11.1.6 Zone spreading .................................... 304
11.1.7 Documentation ..................................... 306
11.2 Selection of Components .................................. 306
11.2.1 Columns ........................................... 306
11.2.2 Valves ............................................ 309
11.2.3 Pumps ............................................. 310
Cleaning .......................................... 310
Chemical resistance ............................... 310
Pressure/flow rate ................................ 311
Temperature tolerance ............................. 311
Shearing .......................................... 311
Speed control ..................................... 311
Pulsations ........................................ 312
11.2.4 Monitors, meters and sensors ...................... 312
UV monitors ....................................... 312
Conductivity monitors ............................. 312
pH monitors ....................................... 313
Flow meters ....................................... 313
Air sensors ....................................... 314
11.2.5 Tubing or piping .................................. 314
11.2.6 Fraction collectors ............................... 314
11.3 Automation ............................................... 314
11.3.1 Advantages of automation .......................... 315
11.3.2 Control systems ................................... 315
Dedicated control system .......................... 315
Programmable logical controller system ............ 315
Personal computer system .......................... 316
Distributed control systems and networking ........ 316
11.3.3 Hardware and software specifications .............. 316
References ............................................... 319
12 Column Packing ........................................... 321
12.1 Introduction ............................................. 321
12.2 Theory ................................................... 321
12.3 Preparation of Column and System ......................... 323
12.4 Packing the Column ....................................... 324
12.4.1 Packing solution .................................. 324
12.4.2 Preparation of the slurry ......................... 324
12.4.3 Determination of packing parameters ............... 325
12.4.4 Packing methods ................................... 325
12.5 Evaluating Column Packing Quality ........................ 326
12.5.1 The step method ................................... 326
12.5.2 The pulse method .................................. 327
12.6 Scale Up ................................................. 329
References ............................................... 330
Appendix A: Symbols and Definitions in Liquid Chromatography .. 331
A.l Introduction ............................................. 331
A.2 Symbols Used in Liquid Chromatography .................... 331
A.3 Definitions of Chromatographic Parameters and Equations .. 333
References ............................................... 336
Appendix B: Dimensionless Numbers ............................. 337
B.l Introduction ............................................. 337
References ............................................... 339
Appendix C: Activities for Biopharmaceutical Production from
Genetically Engineered Mammalian Cells ........................ 341
C.l Introduction ............................................. 341
C.2 Activities Chart from Toxicology to License Application .. 341
Appendix D: Simulations Using the Supplied Software ........... 345
D.l Introduction ............................................. 345
D.2 How to Use the Software? ................................. 345
D.3 Pressure Drop ............................................ 346
D.3.1 Calculations ...................................... 346
D.4 Sample Volume ............................................ 347
D.4.1 Calculations ...................................... 347
D.5 Resolution in SEC ........................................ 348
D.5.1 Calculations ...................................... 348
D.6 Resolution in IEC, RPC and HIC ........................... 348
D.6.1 Calculations ...................................... 348
D.7 Isocratic Separation ..................................... 349
D.7.1 Entry of parameters ............................... 349
D.7.2 Calculations ...................................... 349
D.8 Yield and Purity ......................................... 350
D.8.1 Calculations ...................................... 350
D.9 Langmuir Isotherm ........................................ 350
D.9.1 Calculations ...................................... 351
D.10 About this Software ...................................... 351
Subject Index ................................................. 353
|
