Oligonucleotide-Based Drugs and Therapeutics

Preclinical and Clinical Considerations for Development

Nicolay Ferrari, Rosanne Seguin

• Publisher: Wiley
• Year: 2018
• ISBN:
  9781119070306
• Edition: 1
• Topic: Medicine
• Subtopic: Pharmacology

A comprehensive review of contemporary antisense oligonucleotides drugs and therapeutic principles, methods, applications, and research

Oligonucleotide-based drugs, in particular antisense oligonucleotides, are part of a growing number of pharmaceutical and biotech programs progressing to treat a wide range of indications including cancer, cardiovascular, neurodegenerative, neuromuscular, and respiratory diseases, as well as other severe and rare diseases. Reviewing fundamentals and offering guidelines for drug discovery and development, this book is a practical guide covering all key aspects of this increasingly popular area of pharmacology and biotech and pharma research, from the basic science behind antisense oligonucleotides chemistry, toxicology, manufacturing, to safety assessments, the design of therapeutic protocols, to clinical experience.

Antisense oligonucleotides are single strands of DNA or RNA that are complementary to a chosen sequence. While the idea of antisense oligonucleotides to target single genes dates back to the 1970's, most advances have taken place in recent years. The increasing number of antisense oligonucleotide programs in clinical development is a testament to the progress and understanding of pharmacologic, pharmacokinetic, and toxicologic properties as well as improvement in the delivery of oligonucleotides. This valuable book reviews the fundamentals of oligonucleotides, with a focus on antisense oligonucleotide drugs, and reports on the latest research underway worldwide.

• Helps readers understand antisense molecules and their targets, biochemistry, and toxicity mechanisms, roles in disease, and applications for safety and therapeutics

• Examines the principles, practices, and tools for scientists in both pre-clinical and clinical settings and how to apply them to antisense oligonucleotides

• Provides guidelines for scientists in drug design and discovery to help improve efficiency, assessment, and the success of drug candidates

• Includes interdisciplinary perspectives, from academia, industry, regulatory and from the fields of pharmacology, toxicology, biology, and medicinal chemistry

Oligonucleotide-Based Drugs and Therapeutics belongs on the reference shelves of chemists, pharmaceutical scientists, chemical biologists, toxicologists and other scientists working in the pharmaceutical and biotechnology industries. It will also be a valuable resource for regulatory specialists and safety assessment professionals and an important reference for academic researchers and post-graduates interested in therapeutics, antisense therapy, and oligonucleotides.

1
Mechanisms of Oligonucleotide Actions

Annemieke Aartsma‐Rus¹, Aimee L. Jackson², and Arthur A. Levin³

¹ Department of Human Genetics, Leiden University Medical Center, Leiden, The Netherlands

² miRagen Therapeutics, Boulder, CO, USA

³ Avidity Biosciences, La Jolla, CA, USA

1.1 Introduction

The promise of antisense oligonucleotide (ASO) therapeutics is the ability to design drugs that are specific inhibitors of the expression solely on the basis of Watson and Crick base‐pairing rules. The premise is that treatment of a patient with a DNA‐like oligonucleotide complementary to a disease‐related RNA (usually a messenger RNA) results in the formation of a heteroduplex that inhibits the function (generally translation) of that target RNA. Although antisense RNAs were first described in 1978 [1, 2], until recently the promise of selectivity and efficacy has always remained slightly out of reach for various reasons. Oligonucleotides are large molecules leading to synthesis and delivery issues. In addition, natural DNA and RNA oligonucleotides are rapidly degraded and cleared after systemic delivery. Over time many of the issues that have challenged developers of oligonucleotide‐based therapeutics have been addressed: Synthesis costs have been reduced by orders of magnitude over the past two decades, allowing more investigators to use the technology. Stability issues were addressed partially with the introduction of phosphorothioate backbones (reviewed in Ref. [3]) and later sugar modifications (reviewed in Ref. [4]), and, as a result, oligonucleotides now used clinically and preclinically have more conventional drug‐like properties [5]. In addition, fundamental discoveries have improved our understanding of the antisense mechanisms. We now know that target RNA structure and accessibility impacts activity of oligonucleotide therapeutics [6] and therefore pharmacologic activity. Apparently small changes in oligonucleotide chemistry can also have large pharmacologic effects as analyzed at the phenomenological level [7] and at the quantum level [8].

Our ability to design effective oligonucleotide‐based drugs has also been enhanced by studies of the molecular mechanisms of these agents. This chapter reviews the mechanisms of action, the chemistry, and the clinical applications of three broad categories of oligonucleotide therapeutics: antisense agents, splicing modifiers, and gene silencers that activate the RNA interference (RNAi) pathway.

Antisense technology has now produced dozens of clinical stage drugs and two approvals. That hybridization of an oligonucleotide to a pre‐mRNA could modulate the splicing of that RNA was described in 1993 [9], and the therapeutic potential of that mechanism is being exploited to treat Duchenne muscular dystrophy and now other diseases (see below). Running at first behind but more recently in parallel with applications of single‐stranded ASO agents is the use of double‐stranded RNA‐like molecules that activate the RNA‐induced silencing complex (RISC) to cleave targeted mRNAs or interfere with their translation. Synthetic small interfering RNA (siRNA) therapeutics relies on the same mechanism that is used by eukaryotic cells to control mRNA translation by endogenous microRNAs (miRNAs).

1.2 Antisense Oligonucleotide Therapeutics

1.2.1 Antisense Activity Mediated by RNase H

Zamecnik and Stephenson [1, 2] were the first to describe that a DNA strand complementary to a sequence of an mRNA prevented translation. They observed that an ASO prevented the accumulation of Rous sarcoma virus by inhibiting the translation of proteins encoded by the viral mRNA. A whole new potential field of therapeutics was launched with a single (understated) sentence: “It might also be possible to inhibit the translation of a specific cell protein” [1]. That RNase H was responsible for the inhibitor effects on translation was a conclusion reached by multiple investigators over a period of time. An elegant proof of the role of this specific enzyme in antisense activity was provided by Wu et al. in 1999 [10]; these authors showed that modulation of RNase H levels in cells or animals produces a coordinate change in antisense activity.

1.2.2 The RNase H Mechanism

Members of the RNase H family are ubiquitously expressed. The endonuclease mechanism of action and the crystal structure have been reviewed [10–15]. RNase H is approximately 20 kDa, and the isoforms in mammalian cells are known to have distinct functions. RNase H1 is necessary for transcription, and RNase H2 is thought to remove RNA primers in the replication of DNA [16]. The RNA binding domain of these enzymes is located at the N‐terminus. The catalytic activity is located in a C‐terminal domain and depends upon the presence of the 2′ hydroxyl on the ribose sugar for cleavage. The specificity of the enzyme is imparted by heteroduplex formation between a DNA and the targeted RNA. Thus, the enzyme does not cleave single‐stranded RNA in the absence of a heteroduple...

Table of contents

• Cover
• Table of Contents
• Preface
• 1 Mechanisms of Oligonucleotide Actions
• 2 The Medicinal Chemistry of Antisense Oligonucleotides
• 3 Cellular Pharmacology of Antisense Oligonucleotides
• 4 Pharmacokinetics and Pharmacodynamics of Antisense Oligonucleotides
• 5 Tissue Distribution, Metabolism, and Clearance
• 6 Hybridization‐Independent Effects
• 7 Hybridization‐Dependent Effects
• 8 Class‐Related Proinflammatory Effects
• 9 Exaggerated Pharmacology
• 10 Genotoxicity Tests for Novel Oligonucleotide‐Based Therapeutics
• 11 Reproductive and Developmental Toxicity Testing Strategies for Oligonucleotide‐Based Therapeutics
• 12 Specific Considerations for Preclinical Development of Inhaled Oligonucleotides
• 13 Lessons Learned in Oncology Programs
• 14 Inhaled Antisense for Treatment of Respiratory Disease
• 15 Antisense Oligonucleotides for Treatment of Neurological Diseases
• 16 Nucleic Acids as Adjuvants
• 17 Splice‐Switching Oligonucleotides
• 18 CMC Aspects for the Clinical Development of Spiegelmers
• Index
• End User License Agreement