Gene Therapy for Viral Infections

Patrick Arbuthnot

• Publisher: Academic Press
• Year: 2015
• Print ISBN: 9780124105188
• eBook ISBN: 9780124114524
• Topic: Biological Sciences
• Subtopic: Genetics & Genomics
• Index: Biological Sciences

Gene Therapy for Viral Infections provides a comprehensive review of the broader field of nucleic acid and its use in treating viral infections. The text bridges the gap between basic science and important clinical applications of the technology, providing a systematic, integrated review of the advances in nucleic acid-based antiviral drugs and the potential advantages of new technologies over current treatment options.Coverage begins with the fundamentals, exploring varying topics, including harnessing RNAi to silence viral gene expression, antiviral gene editing, viral gene therapy vectors, and non-viral vectors.Subsequent sections include detailed coverage of the developing use of gene therapy for the treatment of specific infections, the principles of rational design of antivirals, and the hurdles that currently face the further advancement of gene therapy technology.- Provides coverage of gene therapy for a variety of infections, including HBV, HCV, HIV, hemorrhagic fever viruses, and respiratory and other viral infections- Bridges the gap between the basic science and the important medical applications of this technology- Features a broad approach to the topic, including an essential overview and the applications of gene therapy, synthetic RNA, and other antiviral strategies that involve nucleic acid engineering- Presents perspectives on the future use of nucleic acids as a novel class of antiviral drugs- Arms the reader with the cutting-edge information needed to stay abreast of this developing field

Chapter 1

Essentials of Viruses and their Suitability for Treatment Using Gene Therapy

Abstract

Viruses constitute the simplest life forms and are a major cause of disease. They are also very abundant and make up a reservoir of enormous genetic diversity. As obligate intracellular parasites, viruses orchestrate intricate sets of reactions that recruit and usurp cellular functions in such a way as to facilitate their replication. Highly efficient host antiviral mechanisms, particularly innate and adaptive immune responses, are triggered in response to an infection. To control these effects, viruses have evolved ways of evading or partly disabling host antiviral responses. Treatment of patients with small-molecule antivirals, immunomodulators, and immunoprophylaxis by vaccination are currently licensed therapeutic antiviral measures. Harnessing gene therapy offers the potential for developing versatile and effective new intervention strategies. A rational design approach, which only requires basic knowledge of the viral nucleic acid sequences, is particularly advantageous for antiviral gene therapies. Strategies have generally entailed inactivation of viral sequences and host factors. Gene transfer may also be used to induce antiviral immunity. Several approaches to disabling gene function have been used; the use of RNA interference activators and engineering designer nucleases are particularly promising. Because candidate gene therapy drugs are complex, successful implementation of antiviral gene therapy faces significant challenges. Ensuring specificity, efficient delivery to target tissue, and adequate therapeutic effects without toxicity are crucial for the advancement of antiviral gene therapy to clinical use.

Keywords

Adaptive immunity; Antivirals; Escape mutation; Gene therapy; Innate immunity; Vaccination; Viral capsid

1.1. Gene Therapy

The term “gene therapy” was coined in 1972 to explain the use of procedures that are intended to treat or alleviate disease by genetically modifying the cells of a patient [1]. The concept of gene therapy was developed after publication of the first reports demonstrating that it was possible to alter gene expression in cultured cells. These early studies showed that gene expression in murine or human cells could be modified by transfection with DNA expressing herpes simplex virus thymidine kinase [2] or hexose-1-phosphate uridylyltransferase [3], respectively. Initial definitions of gene therapy referred exclusively to the use of genes to treat disease, but the meaning has now become broader [4–6]. Currently, gene therapy is defined by the use of nucleic acids, which may include DNA, RNA, or chemically modified derivatives, to alter gene function and treat disease. Because abnormalities of gene function underlie many disease processes, including those caused by viral infections, interventions using gene therapy potentially have wide-ranging applicability.

Gene therapy has many applications and may be used for restoring the health of diseased cells, killing of malignant tissue, and induction of immune responses to gene-encoded proteins. To treat diseased cells, gene therapy may entail repairing damaged genes or silencing “rogue” genetic elements that are expressed by viral pathogens. With the advent of recombinant DNA technology, polymerase chain reaction, and sophisticated nucleic acid sequencing procedures, insights into molecular biology and the fundamental mechanisms causing disease processes have greatly progressed. These developments have had a profound enabling effect on the rational design of gene therapy approaches.

Different methods of therapeutic inhibition of gene function have been used to counter viral infections. These include silencing of virus-encoded genes (Chapter 2) and introduction of targeted disabling mutations into viral genes or host factors (HFs) (Chapter 3). Using gene transfer to augment patients’ immune responses to virus infections has been another way of achieving preventative or therapeutic antiviral therapeutic effects (Chapter 11).

Exploiting gene therapy to counter virus replication has advanced considerably, and several viruses have now been shown to be candidates for treatment using this approach. However, there is no universal method of using gene therapy for viral infections. Individual viruses have particular characteristics, and this necessitates that viral gene therapy be tailored to specific infections. In developing viral gene therapy, tissue tropism, the acute or chronic nature of an infection, and the efficiency with which antiviral sequences can be delivered to infected tissues are important considerations (Table 1.1). Both synthetic nucleic acids and DNA expression cassettes are being developed for viral gene therapy. Expressed antiviral sequences may be more useful for countering chronic viral infection whereas synthetic antiviral nucleic acids are better suited to inhibiting acute viral infections. Preventing viral escape from gene therapy is important, and overcoming this problem by simultaneous targeting of multiple viral sites or suppressing HFs has shown promise.

1.2. Essentials of Viruses

Viruses are the simplest life forms; not surprisingly, they are also very plentiful. Estimations place the number of viral particles in the biosphere to be between 10³¹ and 10³² [7–9]. In natural waters of the Earth, they are estimated to outnumber bacteria by an order of magnitude. Viruses are a major cause of disease and constitute a reservoir of enormous genetic diversity. They are highly varied with respect to their structure, genome replication mechanisms, and modes of interacting with their host organisms. Common and interrelated defining features of viruses are the following:

• They are obligate intracellular parasites that only reproduce within host cells and are incapable of independent replication.

• Viruses do not have the machinery required for translation of proteins. They use host protein synthesis mechanisms, with their own genetic material as template, to produce the components constituting intact infectious viral particles (virions).

• Viruses lack the mechanisms for generating the energy required to drive the biochemical processes required for their existence.

Viruses may essentially be considered as nucleic acid parasites that use virions to introduce their own genetic material into cells. Thereafter, the host cellular machinery is reprogramed for copying the viral genome and the formation of more virions to result in completion of the viral replication cycle. Viruses have evolved efficient mechanisms for introducing DNA or RNA genomes into cells, and this property has been exploited for development of viral gene therapy vectors. Ironically, in some cases these recombinant vectors are being developed as therapeutics to counter virus infections.

Table 1.1

Viral Characteristics That Influence the Gene Therapy Strategy

Viral characteristics	Implications for gene therapy of viral infections
DNA or RNA virus	Both DNA and RNA viruses are susceptible to RNA silencing mechanisms (e.g., by RNAi activators). DNA targeting, such as by sequence-specific nucleases, is only possible with DNA viruses.
Virus tropism	Delivery of gene therapies remains a challenging task for successfully implementing gene therapy of viral infections. Therefore, accessibility of infected tissues to gene therapy vectors is an important factor that currently influences success. For example, delivery of small interfering RNAs targeting respiratory syncytial virus after inhalation is easier than delivery to hepatocytes after systemic administration of gene therapy formulations.
Acute or chronic nature of infection	Acute infection may require a single dose of a therapeutic whereas chronic infection may require repeated administrations. Approaches that achieve sustained inhibition of viral replication may also be necessary for treating chronic infections.
Viral replication rate	Very high replication rate may overwhelm efficacy of antiviral gene therapy. Moreover, viral dormancy may also evade nucleic acid antivirals.
Predisposition to mutation	RNA viruses or viruses that use reverse transcription during genome duplication are prone to error and may introduce mutations that evade sequence-specific antivirals. Combinatorial approaches or host factor targeting may be necessary to provide a higher barrier to resistance.
Host immune response to virus	Host immunity may augment efficacy of antivirals. Although generally undesirable, induction of the innate immune response by viral gene therapies may augment their antiviral efficacy (e.g., 5′ triphosphate-containing small interfering RNAs against HBV infection).
Reliance on host factors (HFs) that may be silenced	Disabling HFs (e.g., CCR5 for HIV-1 infection and micro RNA 122 for HCV replication) that are required for viral replication may be used to inhibit viral replication. Successful use of this approach requires that inhibition of HF function is not toxic to cells.
Virus-encoded mechanisms of countering gene silencing	Viruses that are capable of inhibiting the RNAi pathway (e.g., adenovirus virus-associated RNAs) may result in attenuated inhibition of silencing efficacy.
Similarity of viral targets to host cellular sequences	Sequence homology between viral targets and cellular sequences may result in unintended harmful off-target effects of a gene therapy.

Whether viruses meet the basic requirements of what constitutes life has been a subject of debate. Definitions of life are themselves fraught [10], but viruses display at least some of the traits that characterize living entities. Key attributes of living entities include the ability to reproduce and evolve in response to external influences. Because viruses reproduce, albeit in a parasitic manner, they may be considered as living organisms. Moreover, adaptation of viruses to their environments through evolution, an important and clinically relevant property, is another argument in favor of viruses being classed as living entities. However, because viruses are obligate intracellular parasites that are incapable of independent replication, some have argued that they do not qualify as living organisms.

Defining what makes a virus a virus has also been the subject of some debate [11–13]. It has been argued that the disappearance and appearance of virions (disintegration and reconstitution) is the fundamental characteristic feature of viruses [13]. Disappearance of the virion occurs when the particle breaks down to release the viral genome into the cell, then reappearance happens when the intracellular machinery is used to propagate virion progeny. Although understanding the phenomenon of disappearance and appearance of viruses is useful, definin...

Table of contents

• Cover image
• Title page
• Table of Contents
• Copyright
• Dedication
• Acknowledgments
• Chapter 1. Essentials of Viruses and their Suitability for Treatment Using Gene Therapy
• Chapter 2. Harnessing RNAi to Silence Viral Gene Expression
• Chapter 3. Engineering Sequence-Specific DNA Binding Proteins for Antiviral Gene Editing
• Chapter 4. Viral Vectors for Delivery of Antiviral Sequences
• Chapter 5. Delivery of Antiviral Nucleic Acids with Nonviral Vectors
• Chapter 6. Gene Therapy for Chronic Hepatitis B Virus Infection
• Chapter 7. Gene Therapy for Hepatitis C Virus Infection
• Chapter 8. Gene Therapy for HIV-1 Infection
• Chapter 9. Gene Therapy for Respiratory Viral Infections
• Chapter 10. Gene Therapy for Infection with Hemorrhagic Fever Viruses
• Chapter 11. Gene Transfer for Prophylaxis and Therapy of Viral Infections
• Chapter 12. Antiviral Gene Therapy: Summary and Perspectives
• Index