Description
This open access volume gathers a variety of models, delivery systems, and approaches that can be used to assess RNA technology for exploiting antisense as a therapeutic intervention. Beginning with a section on the design of antisense technology and their delivery, the book continues by covering model systems developed to evaluate efficacy, both in vivo and in vitro, as well as methods to evaluate preclinically the toxicity associated with these new potential drugs, and intellectual property considerations. Written for the highly successful Methods in Molecular Biology series, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Authoritative and practical, Antisense RNA Design, Delivery, and Analysis provides basic knowledge and a large collection of methods to facilitate the work of newcomers to this vibrant and expanding field. This book was conceived thanks to the network DARTER (Delivery of Antisense RNA Therapeutics). DARTER is funded by the EU Cooperation of Science and Technology (COST), which aims to enhance interaction and collaborations between researchers in Europe and other countries. Part I: Overview 1. Introduction and History of the Chemistry of Nucleic Acids Therapeutics Michael J. Gait and Sudhir Agrawal 2. Antisense RNA Therapeutics: A Brief Overview Virginia Arechavala-Gomeza and Alejandro Garanto Part II: Design and Formulation of Antisense Technology 3. Design of Bifunctional Antisense Oligonucleotides for Exon Inclusion Haiyan Zhou 4. Design and Delivery of SINEUP: A New Modular Tool to Increase Protein Translation Michele Arnoldi, Giulia Zarantonello, Stefano Espinoza, Stefano Gustincich, Francesca Di Leva, and Marta Biagioli 5. How to Design U1 snRNA Molecules for Splicing Rescue Liliana Matos, Juliana I. Santos, M Francisca Coutinho, and Sandra Alves 6. Conjugation of Nucleic Acids and Drugs to Gold Nanoparticles Paula Miln-Rois, Ciro Rodriguez-Diaz, Milagros Castellanos, and lvaro Somoza 7. Determination of Optimum Ratio of Cationic Polymers and Small Interfering RNA with Agarose Gel Retardation Assay Omer Aydin, Dilek Kanarya, Ummugulsum Yilmaz, and Cansu Umran Tunc 8. Generation of Protein-Phosphorodiamidate Morpholino Oligomer Conjugates for Efficient Cellular Delivery via Anthrax Protective Antigen Valentina Palacio-Castaeda, Roland Brock, and Wouter P.R. Verdurmen Part III: In Vitro Model Systems 9. Development and Use of Cellular Systems to Assess and Correct Splicing Defects Nuria Surez-Herrera, Tomasz Z. Tomkiewicz, Alejandro Garanto, and Rob W.J. Collin 10. Modeling Splicing Variants Amenable to Antisense Therapy by Use of CRISPR-Cas9-Based Gene Editing in HepG2 Cells Arstides Lpez-Mrquez, Ainhoa Martnez-Pizarro, Beln Prez, Eva Richard, and Lourdes R. Desviat 11. In Vitro Models for the Evaluation of Antisense Oligonucleotides in Skin Jeroen Bremer and Peter C. van den Akker 12. In Vitro Delivery of PMOs in Myoblasts by Electroporation Remko Goossens and Annemieke Aartsma-Rus 13. Rapid Determination of MBNL1 Protein Levels by Quantitative Dot Blot for Evaluation of Antisense Oligonucleotides in Myotonic Dystrophy Myoblasts Nerea Moreno-Cervera, Irene Gonzalez-Martinez, Rubn Artero, and Estefana Cerro-Herreros 14. Evaluation of Exon Skipping and Dystrophin Restoration in In Vitro Models of Duchenne Muscular Dystrophy Andrea Lpez-Martnez, Patricia Soblechero-Martn, and Virginia Arechavala-Gomeza 15. Generation of Human iPSC-Derived Myotubes to Investigate RNA-Based Therapies In Vitro Pablo Herrero-Hernandez, Atze J. Bergsma, and W.W.M. Pim Pijnappel 16. Eye on a Dish Models to Evaluate Splicing Modulation Kwan-Leong Hau, Amelia Lane, Rosellina Guarascio, and Michael E. Cheetham 17. Establishment of In Vitro Brain Models for AON Delivery Elena Daoutsali and Ronald A.M. Buijsen Part IV: In Vivo Model Systems 18. Considerations for Generating Humanized Mouse Models to Test Efficacy of Antisense Oligonucleotides Irene Vzquez-Domnguez and Alejandro Garanto 19. Generation of Humanized Zebrafish Models for the In Vivo Assessment of Antisense Oligonucleotide-Based Splice Modulation Therapies Renske Schellens, Erik de Vrieze, Ralph Slijkerman, Hannie Kremer, and Erwin van Wijk 20. Use of Small Animal Models for Duchenne and Parameters to Assess Efficiency upon Antisense Treatment Ngoc Lu-Nguyen, Alberto Malerba, and Linda Popplewell 21. In Vivo Models for the Evaluation of Antisense Oligonucleotides in Skin Jeroen Bremer and Peter C. van den Akker 22. Delivery of Antisense Oligonucleotides to the Mouse Retina Alejandro Garanto 23. Delivery of Antisense Oligonucleotides to the Mouse Brain by Intracerebroventricular Injections Tom Metz, Elsa C. Kuijper, and Willeke M.C. van Roon-Mom 24. Intrathecal Delivery of Therapeutic Oligonucleotides for Potent Modulation of Gene Expression in the Central Nervous System Zachary Kennedy, James W. Gilbert, and Bruno M.D.C. Godinho Part V: Safety and Toxicology 25. Preclinical Safety Assessment of Therapeutic Oligonucleotides Patrik Andersson 26. Preclinical Evaluation of the Renal Toxicity of Oligonucleotide Therapeutics in Mice Luca Echevarra and Aurelie Goyenvalle 27. Protocol for Isolation and Culture of Mouse Hepatocytes (HCs), Kupffer Cells (KCs), and Liver Sinusoidal Endothelial Cells (LSECs) in Analyses of Hepatic Drug Distribution Kjetil Elvevold, Ingelin Kyrrestad, Karen Kristine Srensen, and Brd Smedsrd Part VI: Intellectual Property 28. Patent Considerations When Embarking on New Antisense Drug Programs Laurence D.S. Gainey
