Advanced Delivery and Therapeutic Applications of RNAi

Professor Kun Cheng, Division of Pharmaceutical Sciences, School of Pharmacy, University of Missouri-Kansas City, USA Professor Cheng's research interests include the delivery and therapeutic applications of RNAi for breast cancer, prostate cancer, liver diseases, and type I diabetes. He has been working in the area of nucleic acids since 2002. He is co-editor of Advanced Drug Delivery with Dr. Ashim Mitra and Dr. Chi Lee (Wiley, 2012). He has also co-edited a themed issue entitled "siRNA Delivery" with Dr. Ram Mahato for the journal Molecular Pharmaceutics, and has developed and taught a graduate course entitled "Protein and Nucleic Acid Drug Delivery". Professor Ram I. Mahato, Department of Pharmaceutical Sciences, School of Pharmacy, University of Tennessee Health Science Center, USA Professor Mahato is a full professor of Pharmaceutics and Drug/Gene Delivery at the University of Tennessee Health Science Center Memphis. He is the author or co-author of 85 peer reviewed articles and book chapters. He has also edited/written five books and six special journal issues. He is a Special Features Editor of Pharmaceutical Research and on the editorial board of several journals. His research interest includes delivery and targeting of small molecules, oligonucleotides, siRNA and genes.

Preface xvii Contributors xix About the Editors xxiii Part 1 Introduction and Basics of RNAi 1 1 Mechanisms and Barriers to RNAi Delivery 3 Jiehua Zhou and John J. Rossi 1.1 Introduction 3 1.2 Barriers to Systemic RNAi Delivery 5 1.3 Rational Design to Improve RNAi Efficacy 6 1.4 Chemical Modifications to Enhance siRNA Stability and Reduce Immune Response 7 1.5 Cellular Uptake and Intracellular Release of siRNA 7 1.6 Combinatorial Targeting for Targeted RNAi Delivery 8 1.7 Cell-Specific Aptamer-Functionalized Nanocarriers for RNAi Delivery 9 1.8 The Clinical Development and Challenges of siRNAs Therapeutics 10 1.9 Conclusion and Perspectives 12 References 12 Analysis of siRNA Delivery Using Various Methodologies 19 Yi Pei 2.1 Introduction 19 2.2 Checkpoints for Analyzing siRNA Delivery 20 2.2.1 Circulation Checkpoint 22 2.2.2 Organ or Tissue Checkpoint 22 2.2.3 Cellular Checkpoint 22 2.2.4 RISC Checkpoint 23 2.2.5 Target mRNA Knockdown (Indirect Checkpoint) 24 2.2.6 Protein and Outcome (Indirect Checkpoint) 25 2.2.7 Safety (Indirect Checkpoint) 26 2.3 Methods for Analysis of siRNA 26 2.3.1 General Considerations 26 2.3.2 Hybridization-Based (Non-Imaging) Methods 28 2.3.3 Non-Hybridization-Based (Non-Imaging) Methods 34 2.3.4 Imaging-Based (Non-Hybridization) Methods 35 2.3.5 Imaging-Based (Hybridization) Methods 37 2.4 Case Study for siRNA Delivery Analysis 38 References 39 3 Challenges and Opportunities in Bringing RNAi Technologies from Bench to Bed 45 Sandesh Subramanya and Lance Ford 3.1 Introduction 45 3.2 RNAi Mediator (siRNA or shRNA) 45 3.2.1 siRNA 45 3.2.2 Vector-derived shRNA 47 3.2.3 miRNAs 49 3.3 Safety Issues of RNAi Mediators 50 3.3.1 Immune Stimulation 50 3.3.2 RNAi Over expression 52 3.4 Efficacy of RNAi Mediators 52 3.4.1 Therapeutic Response 52 3.5 RNAi Mediators in Clinical Trials 53 3.6 Conclusion 54 References 55 Nonclinical Safety Assessments and Clinical Pharmacokinetics for Oligonucleotide Therapeutics: A Regulatory Perspective 63 Shwu-Luan Lee, Paul Brown, Jian Wang and Robert T. Dorsam 4.1 Introduction 63 4.2 Unique Properties of Oligonucleotide-based Therapeutics 63 4.3 Regulation of Oligonucleotide-Based Therapeutics 65 4.3.1 Submission to the FDA 65 4.3.2 Review Process for Non-clinical Studies 67 4.3.3 Regulatory Issues 74 4.3.4 Clinical Pharmacokinetics 76 4.4 Conclusion 79 Disclaimer 79 Appendix 79 References 80 Role of Promoters and MicroRNA Backbone for Efficient Gene Silencing 83 Feng Li and Ram I. Mahato 5.1 Introduction 83 5.2 Promoters for shRNA Expression 84 5.2.1 Constitutive Promoters 84 5.2.2 Inducible Promoters 87 5.2.3 Site Specific Promoters 93 5.3 miRNA-based shRNAs 96 5.3.1 miRNA-based shRNA Enhances Gene Silencing 96 5.3.2 miRNA-based shRNA Reduces Toxicities 97 5.3.3 Application of miRNA-based shRNA for Combination Gene Therapy 98 5.4 Concluding Remarks 100 References 101 Part 2 RNAi Delivery Strategies 109 6 Bioconjugation of siRNA for Site-specific Delivery 111 Bin Qin, Wei Jin and Kun Cheng 6.1 Introduction 111 6.2 Conjugation Strategy 112 6.2.1 RNA Chemical Modification 112 6.2.2 Site of Conjugation 114 6.2.3 Conjugation Chemistry 115 6.3 Bioconjugates for Site-specific Delivery 120 6.3.1 Antibody-siRNA Bioconjugates 120 6.3.2 Aptamer-siRNA Bioconjugates 122 6.3.3 Peptide-siRNA Bioconjugates 124 6.3.4 Lipid-siRNA Bioconjugates 126 6.3.5 Others 128 6.4 Conclusion 129 References 129 7 Multifunctional RNAi Delivery Systems 137 China Malakondaiah Kummitha, Anthony S. Malamas and Zheng-Rong Lu 7.1 Introduction 137 7.1.1 Chapter Objectives 139 7.2 Lipid-Based Delivery Systems 139 7.2.1 Cationic Lipids 139 7.2.2 Ionizable Cationic Lipids 140 7.2.3 Lipid-Like Materials 140 7.2.4 pH-sensitive Surfactants as Multifunctional siRNA Carriers 142 7.3 Polymeric Multifunctional siRNA Delivery Systems 150 7.3.1 Polyethylenimine 150 7.3.2 Chitosan 151 7.3.3 Cyclodextrins 152 7.3.4 Dendrimers 152 7.3.5 Polyalkylacrylic Acid-based pH-sensitive Polymers 153 7.3.6 Other pH-sensitive Polymers 156 7.4 Conclusion 157 References 157 8 Dendrimers in RNAi Delivery 163 Jose Luis Jimenez Fuentes, Paula Ortega, Sara Ferrando-Martynez, Rafael Gomez, Manuel Leal, Javier de la Mata and MaAngeles Munoz-Fernandez 8.1 Introduction 163 8.2 Challenges in RNAi Delivery 164 8.3 Dendrimers as Non Viral Vectors 166 8.3.1 Dendritic Architectures 166 8.3.2 Synthesis of Dendrimers 168 8.3.3 Types of Dendrimers in Drug Delivery 169 9 Development of Pharmaceutically Adapted Mesoporous Silica Nanoparticles for siRNA Delivery 187 Wilson X. Mai, Tian Xia and Huan Meng 9.1 Introduction 187 9.2 Mesoporous Silica Nanoparticles as Novel Inorganic Nanocarriers for siRNA Delivery 188 9.2.1 Discovery and Synthesis 188 9.2.2 Surface Modification of MSNP for Nucleic Acid Delivery 190 9.2.3 MSNP for Dual siRNA and Drug Delivery 191 9.2.4 Improving in vivo Implementation of MSNP-Based Delivery Platform 196 9.2.5 Design of Pharmaceutically Adapted MSNP via the Knowledge Generated by Discoveries at the Nano/Bio Interface 197 9.3 Safety Assessment of Nanocarrier and Design of Safe MSNP Carrier 199 9.3.1 Safety of Nanocarriers 199 9.3.2 Safe Design of MSNP Carrier 201 References 179 9.4 Summary References 202 10 Environmentally-Responsive Nanogels for siRNA Delivery 207 Atsushi Tamura and Yukio Nagasaki 10.1 Introduction 207 10.1.1 siRNA Delivery System 207 10.1.2 Crosslinked Nanogels for siRNA Delivery 208 10.2 Reductive Environment-Responsive Disulfide Crosslinked Nanogels 209 10.3 Temperature-Responsive Nanogels 211 10.4 pH-Responsive Nanogels 212 10.4.1 Acid-degradable Nanogels for Intracellular Release of siRNA 212 10.4.2 Design of pH-Responsive PEGylated Nanogels with Endosomal Escape Ability 212 10.4.3 Cytoplasmic Delivery of PEGylated Nanogel/siRNA Complexes 214 10.5 PEGylated and Partially Quaternized Polyamine Nanogels 216 10.5.1 Design of Quaternized Polyamine Nanogels 216 10.5.2 Enhanced Cellular Uptake of siRNA by Quaternized Polyamine Nanogels 216 10.5.3 Enhanced Gene-Silencing Activity of Quaternized Polyamine Nanogel/siRNA Complexes 219 10.6 Conclusions 220 References 220 11 Viral-Mediated Delivery of shRNA and miRNA 225 Fredric P. Manfredsson 11.1 Introduction 225 11.2 RNAi -- A Brief Overview 226 11.3 shRNA or miRNA? 226 11.4 Rational Design 227 11.5 Viral Vectors 227 11.5.1 Recombinant Adeno-associated Virus (rAAV) 229 11.5.2 Retrovirus (RV) 230 11.5.3 Lentivirus (LV) 230 11.5.4 Adenovirus (AD) 231 11.5.5 Herpes Simplex Virus (HSV) 231 11.5.6 Baculovirus (BV) 232 11.5.7 Poxvirus 232 11.6 Tissue-specific Transduction 233 11.6.1 CNS 233 11.6.2 Ocular 234 11.6.3 Respiratory System 235 11.6.4 Liver 236 11.6.5 Skeletal Muscle 237 11.6.6 Heart 237 11.6.7 Systemic 238 11.6.8 Ex Vivo 238 11.6.9 Cell Culture 238 11.6.10 Transcription Cassettes 239 11.7 Applications of Virally Expressed shRNAs 241 11.7.1 Virally Mediated "Knockouts" 241 11.7.2 Concomitant Expression of Therapeutic Genes 241 11.8 Viral Gene Therapy in the Clinic 241 11.9 Conclusion 242 References 242 12 The Control of RNA Interference with Light 255 Simon H. Friedman 12.1 Introduction 255 12.2 The Importance of Gene Expression 255 12.3 Light Control of Gene Expression 257 12.4 Why Use RNA Interference as a Basis for Light Control of Gene Expression? 258 12.5 Light Activated RNA Interference (LARI), the work of Friedman and Co-Workers 259 12.6 Work of McMaster and Co-Workers, 50 Antisense Phosphate Block 262 12.7 Work of Heckel and Co-Workers, Nucleobase Block 263 12.8 Use of 20 FsiRNA, work of Monroe and Co-Workers 264 12.9 Photochemical Internalization 265 12.10 Future Directions and Conclusions 266 Acknowledgments 267 References 267 Part 3 Applications of RNAi in Various Diseases 269 13 RNAi in Cancer Therapy 271 Cristian Rodriguez-Aguayo, Arturo Chavez-Reyes, Gabriel Lopez-Berestein and Anil K. Sood 13.1 Introduction 271 13.2 Therapeutic Opportunities for Noncoding RNAs 274 13.3 RNAs as Drugs 277 13.4 Overcoming Anatomical and Physiologic Barriers 278 13.4.1 Intravascular Degradation 279 13.4.2 Tissue and Intracellular Delivery 280 13.4.3 Immune-mediated Toxic Effects 281 13.4.4 Nanocarrier-mediated Toxic Effects 282 13.5 Advanced Delivery 283 13.5.1 Localized siRNA Delivery 285 13.5.2 Systemic siRNA Delivery 288 13.5.3 Targeted siRNA Delivery 291 13.5.4 Monitoring Delivery and Therapeutic Response 293 13.6 Clinical Experience 294 13.7 The Next Steps 298 Acknowledgments 298 References 298 14 Adenovirus-mediated siRNA Delivery to Cancer 309 Chae-Ok Yun 14.1 Introduction 309 14.1.1 shRNA-expressing Vectors 310 14.1.2 Adenovirus Vectors 311 14.2 shRNA-expressing Adenoviruses: Cancer Biological Studies and Therapeutic Implications 312 14.2.1 Oncogene-targeted shRNA-expressing Ads 312 14.2.2 shRNA-expressing Adenoviruses that Target Anti-apoptotic Genes 314 14.3 Exploiting Oncolytic Adenovirus for siRNA Expression 315 14.4 Current Limitations of Adenovirus-mediated siRNA Therapy and Future Directions: Smart Adenovirus Nanocomplexes Expressing siRNA for Systemic Administration 318 14.5 Conclusion 320 References 321 15 RNAi in Liver Diseases 327 Jiang Li, Jianqin Lu, Yifei Zhang, Mohammed Ghazwani, Peng Zhang, Xiang Gao and Song Li 15.1 Introduction 327 15.2 RNAi in Viral Hepatitis 327 15.2.1 Hepatitis B 328 15.2.2 RNAi of HBV Infection via siRNA/shRNA 329 15.2.3 RNAi of HBV Infection via miRNAs 330 15.2.4 Hepatitis C 332 15.2.5 RNAi of HCV Infection via siRNA/shRNA 333 15.2.6 RNAi of HCV Infection via miRNAs 335 15.3 RNAi in Hepatocellular Carcinoma 336 15.3.1 RNAi of HCC via siRNA/shRNA 337 15.3.2 RNAi of HCC via miRNAs 338 15.4 RNAi in Liver Fibrosis 340 15.4.1 RNAi of Liver Fibrosis via siRNA/shRNA 341 15.4.2 RNAi of Liver Fibrosis via miRNAs 343 15.5 Delivery Systems in RNAi 345 15.5.1 Liver Anatomy 346 15.5.2 Viral Delivery Systems 346 15.5.3 Non-Viral Delivery Systems 347 15.5.4 Cell-specific Targeting Strategies 348 15.5.5 Cellular Events after the Uptake of Nucleic Acid-Carrier Complexes 349 15.5.6 Lipid-based Delivery Systems 350 15.5.7 Polymer-Based Systems 350 15.5.8 Calcium Phosphate-Lipid Hybrid System 351 15.5.9 Hydrophobitized Nucleic Acid Derivatives 351 15.5.10 Targeted Delivery to Tumor Blood Vessels 351 15.6 Conclusion 352 Acknowledgments 353 References 353 16 Approaches to Delivering RNAi Therapeutics that Target Hepatitis B Virus 367 Carol Crowther, Mohube Betty Mowa, Abdullah Ely and Patrick Arbuthnot 16.1 Introduction 367 16.1.1 RNAi Therapeutics 368 16.1.2 Hepatitis B Virus as a Target for RNAi-based Gene Silencing 369 16.2 Vectors Suitable for Hepatic Delivery of HBV Gene Silencers 369 16.2.1 Viral Vectors 370 16.2.2 Nonviral Vectors 377 16.3 Conclusions 381 Acknowledgments 382 References 382 17 RNAi in Respiratory Diseases 391 Ciara Kelly, Awadh B. Yadav, Paul J. McKiernan, Catherine M. Greene and Sally-Ann Cryan 17.1 Introduction 391 17.2 Respiratory Disease and RNA Interference 392 17.2.1 RNAi in Lung Cancer 393 17.2.2 RNAi to Treat Respiratory Infections 393 17.2.3 RNAi in Inflammatory Lung Disease 394 17.3 Delivery and Development of RNAi Therapies for Respiratory Disease 397 17.3.1 Inhalation of RNA-medicines 397 17.3.2 Chemical Modifications of siRNA 399 17.3.3 RNAi Vectors 400 17.3.4 RNAi Therapy In Vivo 405 17.4 Conclusions 408 Acknowledgements 408 References 408 18 RNAi in Ocular Diseases 417 Andrey Turchinovich, Georg Zoidl and Rolf Dermietzel 18.1 Introduction 417 18.2 The Principle of RNAi 418 18.3 In vivo Delivery of siRNA 419 18.4 Delivery of siRNA into the Eye 420 18.4.1 Routes for Ocular Delivery of siRNA 420 18.4.2 Delivery of Naked siRNA 421 18.4.3 Delivery of siRNA Using Carriers 425 18.4.4 Viral Delivery of shRNA 429 18.5 Conclusions 431 Abbreviations 432 References 432 19 micro RNAs as Therapeutic Agents and Targets 439 D.S. Karolina and K. Jeyaseelan 19.1 Introduction 439 19.2 miRNA Therapeutics 440 19.2.1 Therapeutic miRNA Inhibition 443 19.2.2 Therapeutic miRNA Mimicry 446 19.3 MicroRNAs and Cancer 447 19.4 MicroRNAs in Stroke 450 19.5 MicroRNAs in Heart Diseases 452 19.6 MicroRNAs in Diabetes Mellitus 454 19.7 MicroRNAs in Liver Diseases 457 19.8 MicroRNAs and Ocular Diseases 461 19.9 MicroRNAs and Respiratory Diseases 462 19.10 MicroRNAs and Stem Cell Research 465 19.11 Conclusion 468 References 469 20 Delivery of Micro RNA Sponges for Interrogation of MicroRNA Function In Vitro and In Vivo 483 Jiakai Lin and Shu Wang 20.1 MicroRNA Loss-of-Function Studies 483 20.2 Considerations in MicroRNA Sponge Design 486 20.2.1 Vector 486 20.2.2 Promoter 487 20.2.3 Reporter Gene 488 20.2.4 MicroRNA Binding Sites 488 20.3 Advantages and Limitations of MicroRNA Sponge over Other MicroRNA Loss-of-Function Strategies 489 20.4 Interrogating MicroRNA Function via Transient MicroRNA Sponge Expression 493 20.5 Interrogating MicroRNA Function via Stable MicroRNA Sponge Expression 494 20.5.1 MicroRNA and Cell Differentiation 494 20.5.2 MicroRNAs in Disease Development 495 20.6 Utility of MicroRNA Sponge in Living Organisms 496 20.6.1 MicroRNA Knockdown in Plants 496 20.6.2 MicroRNA Knockdown in Mouse 497 20.6.3 MicroRNA Knockdown in Drosophila Melanogaster 498 20.7 Future Perspectives 498 References 499 Index 505