Cover
Translational Oncology
Title page
Copyright page
List of Contributors
Series Foreword
Preface
1. Reference
Part 1: Delivery Systems
1. Chapter 1: Translational Cancer Research
  1. Adenovirus
  2. Oncolytic Adenoviruses for Treatment of Cancer
  3. Vaccinia Virus
  4. Herpes Virus
  5. Parvovirus
  6. Final Remarks
  7. References
2. Chapter 2: Retroviruses for Cancer Therapy
  1. Introduction
  2. Overview of Retroviral Vectors
  3. Gammaretroviral and Alpharetroviral Vectors
  4. Foamyviral (Spumaviral) Vectors
  5. HIV-1-Derived Lentiviral Vectors
  6. Evolution of HIV-1-Derived Lentiviral Vectors
  7. Retroviral Vector-Mediated RNAi Therapy: From Basic Science to Clinical Trial
  8. Concluding Remarks and Challenges
  9. Acknowledgments
  10. References
3. Chapter 3: DNA Plasmids for Non-viral Gene Therapy of Cancer
  1. Delivery
  2. Expression of Transgenes from DNA Plasmid
  3. Safety
  4. Therapeutic Applications
  5. Summary
  6. References
4. Chapter 4: Cancer Therapy with RNAi Delivered by Non-viral Membrane/Core Nanoparticles
  1. Introduction
  2. Barriers
  3. Overcoming Delivery Barriers
  4. Putting it All Together: Model Examples of Innovative and Successful NPs
  5. Conclusions
  6. Acknowledgments
  7. References
Part 2: Targeted Expression
1. Chapter 5: Cancer Gene Therapy by Tissue-specific and Cancer-targeting Promoters
  1. Introduction
  2. Cancer-Specific Promoters
  3. Cancer-Targeting Promoters
  4. Engineered Systems that Enhance Promoter Activity
  5. Conclusion
  6. References
2. Chapter 6: MicroRNAs as Drugs and Drug Targets in Cancer
  1. Introduction
  2. MiRNA and its Functioning Mechanism
  3. MiRNA and Cancers
  4. MiRNA in Other Diseases
  5. MiRNA Therapeutics
  6. Challenges
  7. Outlook
  8. References
Part 3: Principles of Clinical Trials in Gene Therapy
1. Chapter 7: Regulatory Issues for Manufacturers of Viral Vectors and Vector-transduced Cells for Phase I/II Trials
  1. Introduction
  2. Development of Good Manufacturing Practices
  3. cGMP Requirements
  4. cGMP Compliance
  5. Practical Compliance: One Center’s Experience
  6. Conclusions
  7. Acknowledgments
  8. References
2. Chapter 8: US Regulations Governing Clinical Trials in Gene Therapy
  1. Introduction
  2. NIH OBA and RAC
  3. IBC
  4. IRB
  5. FDA
  6. Protocol
  7. Consent Form
  8. Multisite Protocols
  9. Conclusion
  10. References
3. Chapter 9: Remaining Obstacles to the Success of Cancer Gene Therapy
  1. Introduction
  2. Drug-Development Pathways
  3. Broadening the Appeal of Complex Biologics
  4. Clinical Trial Design and Execution
  5. Conceptual Limitations
  6. Conclusions
  7. References
Index
End User License Agreement

List of Tables

Chapter 1
1. Table 1.1 The main characteristics of the viruses discussed in this chapter.
Chapter 3
1. Table 3.1 Ongoing clinical trials targeting B-cell malignancies infusing T cells (i) genetically modified ex vivo using the Sleeping Beauty (SB) system to express a CD19-specific chimeric antigen receptor and (ii) propagated on engineered artificial antigen-presenting cells.
Chapter 4
1. Table 4.1 Barriers that hinder delivery efficiency and necessitate strategies to overcome them.
2. Table 4.2 NP characteristics.
Chapter 6
1. Table 6.1 Overview of the major miRNAs involved in cancer.
Chapter 8
1. Table 8.1 Assessment of risk of delayed adverse events in a gene therapy study. Source: US Food and Drug Administration [26].
2. Table 8.2 Integration properties of current commonly used gene therapy vectors in clinical trials.Source: US Food and Drug Administration [26].
Chapter 9
1. Table 9.1 Therapeutic applications of cancer gene therapy.
2. Table 9.2 Advantages and disadvantages of cancer gene therapy vectors.

List of Illustrations

Chapter 1
1. Figure 1.1 Schematic diagram representing the different kinds of adenovirus-derived vectors used for gene therapy. (A) Wild-type adenovirus is able to replicate and kill all permissive cells. (B) The E1 gene is replaced by the expression cassette; this vector can infect all permissive cells but they cannot replicate unless E1 is not transcomplemented by the packaging cell line. (C) All viral genes are deleted except ITRs and the packaging signal. These vectors can infect all permissive cells but they cannot replicate. (D) Oncolytic adenoviruses. These viruses have been engineered to selectively replicate in and kill cancer cells.
2. Figure 1.2 Transcriptional targeting. Simplified schematic illustrating the strategies used to achieve transcriptional targeting of tumor cells. (A) For example, a viral genome is modified to not be able to counteract the defense mechanisms that a normal cell turns on following a viral infection. (B) Tumor cells are defective of such mechanisms hence the virus can have its normal life cycle. (C) Tumor-specific promoter can initiate virus DNA replication, starting its life cycle.
3. Figure 1.3 Transductional targeting. Schematic diagram representing the strategies used for transductional targeting. (A) Tumor expressing a specific receptor can be infected and killed by an adenovirus that infects through the same receptor, e.g. adenovirus serotype 5 and Coxackie adenovirus receptor-expressing tumor. (B) Some tumors do not express Coxackie adenovirus receptors hence they are not susceptible to infection by Ad5. (C) Viruses bearing the knob from a different serotype (chimera viruses) are used to overcome the lack of Coxackie adenovirus receptors.
Chapter 2
1. Figure 2.1 Schematic representations of retrovirus genomes and HIV-1 life cycle. (A) A simple retrovirus genomic RNA. (B) HIV-1 genomic RNA. In addition to three indispensable retroviral genes (gag, pol, and env), the HIV-1 genome also contains six accessory genes (tat, rev, vpr, vif, vpu, and nef). PBS, primer-binding site. (C) Life cycle of HIV-1. Early stage: the HIV-1 envelope protein recognizes specific receptors on the surface of target cells to mediate entry and release of HIV-1 virion into the cytoplasm. After entry, the viral RNA is reverse transcribed into cDNA, imported into the host cell nucleus, and integrated into the host genome to become a provirus. Late stage: using the host cell transcription and translation machinery, the provirus is transcribed and the transcripts are exported to cytoplasm where they are translated into viral packaging proteins or used to generate the viral genome of next-generation virus. These viral components are then assembled into mature virions that exit the host cell through budding, to become next generation viruses.
2. Figure 2.2 Scheme of lentiviral vectors derived from HIV-1. (A) The first-generation vector consists of three plasmid constructs. HIV-1 envelope glycoprotein is substituted by VSV-G. (B) On the basis of the first-generation vectors, the second-generation vectors exclude the four accessory genes (vpr, vif, vpu, and nef) of HIV-1. (C) The third-generation vector further deletes the HIV-1 accessory gene tat and is composed of four plasmid constructs. Rev is expressed by a separate plasmid construct and SIN LTRs are included in transgene plasmid construct.
3. Figure 2.3 Schematic representation of lentiviral vector production and transduction. Transgene plasmid construct and packaging plasmids are cotransfected into 293 T cells for viral production. Then viral particles are collected by ultracentrifugation, resuspended, and titrated to determine the needed amount for target cell infection.
Chapter 3
1. Figure 3.1 Model of electroporation leading to channel formation and DNA transfer. (A) DNA is present in the proximity or in direct contact with cell membranes prior to the application of an electric field. (B) Upon application of an electric current, the cells elongate along electric field lines. Consequently, the curvature radii at the apices of the aligned cells increase and the internal and external charges redistribute to form a transmembrane potential. Subsequent discharge of electric potential across the membrane leads to the formation of conical pores that permit ion flux. (C) Under the continuous influence of electric field currents, the pores coalesce to form larger inverted hydrophilic channels that facilitate the passage of nucleic acid. Intracellular transport of nucleic acid may be passive or may be facilitated by the application of electrophoretic currents.
2. Figure 3.2 Mechanism of hydrodynamic gene delivery. (A) Tight junctions between endothelial cells form the vascular wall surrounded by hepatocytes in undisrupted liver tissue. (B) Upon hydrodynamic injection of naked DNA solution, the rapid increase in vascular volume disrupts the tight endothelial cell junctions, placing the hepatocytes in direct contact with the circulation. Transient pores form in the cell membranes allowing naked DNA in circulation to enter the cytosol. Other models suggest entry of DNA into the cytosol via vesicles created by macropinocytosis.
3. Figure 3.3 Non-viral genomic integration systems for stable expression of therapeutic genes. These systems are based on the cotransfection of expression cassettes and transposase genes to facilitate genomic integration. These genes may be on separate vectors, as shown, or encoded on the same plasmid. In the Sleeping Beauty system, the expression cassette is flanked by inverted terminal repeat/direct repeat (IR/DR) sequences. Transposase binds to the IR/DR elements forming a synaptic complex that is transposed into genomic TA dinucleotide sites. The piggyBac expression cassette is flanked by inverted terminal repeat (ITR) elements that target TTAA sequences in the target cell genome. The phiC31 integrase system is based on recombination between two attachment (att) sites: attB on the vector and so-called pseudo-attP genomic sites that are similar to native bacterial attP sequences. The stably integrated expression cassette is flanked by the newly recombined att sites (attR and attL).
Chapter 4
1. Figure 4.1 Cytoplasmic double-stranded DNA binds to Dicer, which cleaves it into a short oligonucleotide. Upon binding with R2D2/TRBP and Ago2, one RNA strand is discarded and the other is used as the template strand that is complementary to the target mRNA. mRNA then binds to this guide strand and is cleaved and released, while the siRNA guide strand remains a part of the RNA-induced silencing complex (RISC) and is able to bind and cleave additional mRNA.
2. Figure 4.2 Depictions of mushroom, brush, and collapsed conformations. (A) When s > 2r, PEG molecules do not sterically hinder each other and are free to spread out close to the NP surface. (B) When s < 2r, the PEG molecules are closer together and their interaction causes them to extend out from the NP. (C) When the NP curvature is increased, the same PEG density on the NP surface provides less shielding because the distal ends of each PEG molecule are now farther from each other. (D) High-density PEG may collapse on the surface of NPs.
3. Figure 4.3 Mechanisms of endosomal escape. (A) Ion-pair formation. Cationic lipids in the NP form ion pairs with anionic lipids in the endosome, leading to destabilization and fusion of both membranes. (B) Electrostatic interaction. Highly charged cationic lipids (or other particles) in the NP disrupt the anionic endosome. (C) The proton sponge effect. A proton sponge molecule (e.g. PEI) acts as a buffer while proton pumps in the endosomal membrane actively pump in H+ and Cl−. Therefore, the pH in the endosome does not drop, and more H+ and Cl− are pumped in, followed by water moving with its osmotic gradient. The increased pressure in the endosome ruptures it and allows the release of siRNA. (D) Osmotic rupture. A solid precipitate (e.g. calcium phosphate, shown by CaP) dissolves in the acidic endosome, increasing the osmotic pressure in the endosome, and ushering in water moving with its osmotic gradient. The increased pressure in the endosome ruptures it and allows the release of siRNA.
4. Figure 4.4 Hypothetical diagram of LCP with a collapsed and entangled PEG layer. A hollow calcium phosphate core (middle) can encapsulate siRNA, DNA, drug, or peptide. An asymmetric lipid bilayer, where the lipids in the inner leaflet differ from those in the outer leaflet, surrounds the core and is functionalized with PEG (green) and targeting ligands (yellow). Ligands may also be entangled, allowing only a fraction to be available for receptor binding. Source: B. DiPrete, unpublished diagram.
5. Figure 4.5 Schematic diagram of an AuNP complex. (A) The fully functionalized NP in systemic circulation. (B) Degradation of ketal linkages causes individual PEG-functionalized AuNPs to release and dissipate. (C) Exposed cationic core facilitates enhanced endocytosis and proton sponge effect, and is further degraded in the endosome to release siRNA. Source: Adapted from Shim 2010 [55]. Copyright 2012 American Chemical Society.
Chapter 5
1. Figure 5.1 Schematic of transcriptional amplification modules. (A) The TSTA system comprises of a two-step transcription process. In the first step of the system, a synthetic transcription factor, GAL4–VP2 is expressed under a tissue-/cancer-specific promoter. This transcription activator then binds to five GAL4 DNA-binding sites in the second step to direct transcription of the downstream therapeutic or reporter gene. Adapted from Figueiredo et al. [135]. (B) The VISA system is comprised of the TSTA module plus WPRE. The combination of transcription amplification plus RNA stabilization enhances and prolongs expression of the target gene.
2. Figure 5.2 A simplified schematic representation of transcriptionally targeted gene expression by cancer-specific or cancer-targeting promoter. In the first step, genes that are highly expressed in specific cancer types are identified and considered candidates for cancer-specific or cancer-targeting promoters. Then, regulatory elements in the promoter region upstream of the transcriptional start site (TSS) are cloned and characterized. A region containing elements or a combination of regulatory elements carrying the highest transcriptional activities is cloned into a reporter expression vector to test for transcriptional activities in cancer cells. Expression of the reporter gene is limited to cancer cells by presence of specific transcription factors. Reporter gene expression is compared between cancer-specific/-targeting promoter alone, cancer-specific/-targeting promoter plus the VISA/TSTA module, and the CMV promoter. Ideally, to achieve the highest therapeutic window (++++), the promoter activity should be higher than the nonspecific CMV promoter (++++) and show little or no expression (+/−) in normal cells. CSP, cancer-specific promoter; CTP, cancer-targeting promoter.
Chapter 6
1. Figure 6.1 MiRNA biogenesis. MiRNAs are transcribed from the genome by RNA polymerase II (Pol II) into pri-miRNAs, which are then processed by Drosha into hairpin structures called pre-miRNAs. The pre-miRNAs are transported by exportin-5 from nucleus to the cytoplasm, where Dicer further processes them into miRNA duplex. The miRNA duplex unwinds and the mature miRNAs are incorporated onto the RISC. By base pairing with the target mRNAs, miRNA affects protein production either by interfering with protein translation or by changing the mRNA's stability.
2. Figure 6.2 Strategies of miRNA therapeutics. Generally, two strategies can be employed to modulate miRNA expression or function: using miRNA mimics or viral constructs to restore miRNA function, and using antisense oligonucleotides (including antagomirs and locked nucleic acids) or miRNA sponge constructs to block miRNA function.
3. Figure 6.3 The chemical structures of RNA and LNA. By forming a C3′-endo conformation by the introduction of a 2′-O,4′-C methylene bridge, the LNA modification greatly increases the stability and specificity of the antisense oligonucleotide.
4. Figure 6.4 The mechanism of miRNA sponges. MiRNA sponges are transcribed at high levels by using expression vectors in the nucleus. After release into the cytosol, these sponges compete with the miRNA-targeting mRNAs for the binding of miRNAs. As a result, the mRNAs are released from the inhibition by miRNAs, and consequently the expression levels of proteins translated by these mRNAs are upregulated.
Chapter 7
1. Figure 7.1 Producer line Master Cell Bank production and testing. Manufacturers are cautioned to contact the FDA to determine current testing requirements. QC, Quality Control; RCR, replication-competent retrovirus; USP, United States’ Pharmacopoeia.
2. Figure 7.2 Retrovirus producer working cell bank freezing and testing. Manufacturers are cautioned to contact the FDA to determine current testing requirements.* Indicates required for vaccine product. PERT, product-enhanced reverse transcriptase; QC, Quality Control; USP, United States’ Pharmacopoeia.
3. Figure 7.3 Major steps in the manufacture and testing of an adenoviral vector. Manufacturers are cautioned to contact the FDA to determine current testing requirements. In this schema part of the Master Virus Bank has been used as the final clinical product. TGB, Tris/glycerol buffer; USP, United States’ Pharmacopoeia; VP, viral particles.
4. Figure 7.4 Major steps in the manufacture and testing of a retroviral vector. Manufacturers are cautioned to contact the FDA to determine current testing requirements. RCR, replication-competent retrovirus; USP, United States’ Pharmacopoeia.
5. Figure 7.5 Floor plan of the cGMP facilities at the Baylor Center for Cell and Gene Therapy. The Vector Production Facility has unidirectional flow of staff, materials, and waste, whereas the Cell Processing Facility is a single-corridor design with multidirectional flow.
6. Figure 7.6 Vector manufacturing. Showing the interior of a Class 10,000 (ISO 7) manufacturing suite during retroviral vector production. Staff at the left-hand BSC are harvesting the supernatant. The staff member at the right-hand BSC is labeling the tubes for storage of the supernatant. The staff member in the foreground is performing in-production environmental monitoring.
Chapter 8
1. Figure 8.1 Flow chart for development and review of a gene therapy clinical trial. PI, Principal Investigator.
2. Figure 8.2 An example of the organizational chart developed to provide support for gene-transfer studies.

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Library of Congress Cataloging-in-Publication Data

Cancer gene therapy by viral and non-viral vectors / edited by Malcolm K. Brenner, Mien-Chie Hung.
p. ; cm. – (Translational oncology ; 4)
Includes bibliographical references and index.
ISBN 978-1-118-50162-7 (hardback)
I. Brenner, Malcolm K., 1951– editor of compilation. II. Hung, Mien-chie, editor of compilation.
[DNLM: 1. Neoplasms–therapy. 2. Genetic Therapy. 3. Genetic Vectors–therapeutic use. 4. Oncolytic Virotherapy. QZ 266]
RC268.4
616.99′4042–dc23

2013049552

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Printed in the United States of America

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List of Contributors

Malcolm K. Brenner, MB, BChir, PhD
Center for Cell and Gene Therapy
Baylor College of Medicine
Houston, TX, USA

John C. Burnett, PhD
Department of Molecular and Cellular Biology
Beckman Research Institute of City of Hope
Duarte, CA, USA

George A. Calin, MD, PhD
Department of Experimental Therapeutics
The University of Texas MD Anderson Cancer Center
Houston, TX, USA

Vincenzo Cerullo, PhD
Laboratory of Immunovirotherapy
Centre for Drug Research (CDR) and Division of
Pharmaceutical Bioscience
Faculty of Pharmacy
University of Helsinki
Helsinki, Finland

Laurence J.N. Cooper, MD, PhD
Division of Pediatrics
The University of Texas MD Anderson Cancer Center
Houston, TX, USA

Yue Ding, MSc
Department of Molecular and Cellular Biology
Irell and Manella Graduate School of Biological Sciences
Beckman Research Institute of City of Hope
Duarte, CA, USA

Adrian P. Gee, PhD
Center for Cell and Gene Therapy
Baylor College of Medicine
Houston, TX, USA

Bambi Grilley, RPh, RAC, CIP, CCRC, CCRP
Center for Cell and Gene Therapy
Baylor College of Medicine
Houston, TX, USA

Kilian Guse, PharmD, PhD
Cancer Gene Therapy Group
Haartman Institute
University of Helsinki
Helsinki, Finland

Akseli Hemminki MD, PhD
Cancer Gene Therapy Group
Haartman Institute
University of Helsinki
Helsinki, Finland

Jennifer L. Hsu, PhD
Department of Molecular and Cellular Oncology
The University of Texas MD Anderson Cancer Center
Houston, TX, USA

Yi-Hsin Hsu, PhD
Department of Molecular and Cellular Oncology
The University of Texas MD Anderson Cancer Center
Houston, TX, USA

Leaf Huang, PhD
Division of Molecular Pharmaceutics
Eshelman School of Pharmacy
UNC-NCSU Joint Department of Biomedical Engineering
University of North Carolina at Chapel Hill
Chapel Hill, NC, USA

Mien-Chie Hung, PhD
Department of Molecular and Cellular Oncology
The University of Texas MD Anderson Cancer Center
Houston, TX, USA

Longfei Huo, PhD
Department of Molecular and Cellular Oncology
The University of Texas MD Anderson Cancer Center
Houston, TX, USA

Chia-Wei Li, PhD
Department of Molecular and Cellular Oncology
The University of Texas MD Anderson Cancer Center
Houston, TX, USA

Hui Ling, MD, PhD
Department of Experimental Therapeutics
The University of Texas MD Anderson Cancer Center
Houston, TX, USA

Zhuyong Mei, MD
Center for Cell and Gene Therapy
Baylor College of Medicine
Houston, TX, USA

Judy S.E. Moyes, MB, BChir
Division of Pediatrics
The University of Texas MD Anderson Cancer Center
Houston, TX, USA

Amer M. Najjar, PhD
Cancer Systems Imaging
The University of Texas MD Anderson Cancer Center
Houston, TX, USA

John Rossi, PhD
Department of Molecular and Cellular Biology
Beckman Research Institute of City of Hope
Duarte, CA, USA

Andrew B. Satterlee, BS
Division of Molecular Pharmaceutics
Eshelman School of Pharmacy
UNC-NCSU Joint Department of Biomedical Engineering
University of North Carolina at Chapel Hill
Chapel Hill, NC, USA

Markus Vähä-Koskela, PhD
Cancer Gene Therapy Group
Haartman Institute
University of Helsinki
Helsinki, Finland

Yan Wang, PhD
Department of Molecular and Cellular Oncology
The University of Texas MD Anderson Cancer Center
Houston, TX, USA

Jiehua Zhou, PhD
Department of Molecular and Cellular Biology
Beckman Research Institute of City of Hope
Duarte, CA, USA

Series Foreword

While our knowledge of cancer at a cellular and molecular level has increased exponentially over the last decades, progress in the clinic has been more gradual, largely depending upon empirical trials using combinations of individually active anticancer drugs to treat the average patient. The challenge for the immediate future is to accelerate the pace of progress in clinical cancer care by enhancing the bidirectional interaction between laboratory and clinic. Our new understanding of human cancer biology and the heterogeneity of cancers at a molecular level must be used to identify novel targets for therapy, prevention, and detection focused on each individual. Barriers must be removed to facilitate the flow of targeted agents and fresh approaches from the laboratory to the clinic, while returning relevant human specimens, images, and data from the clinic to the laboratory for further analysis.

In this title we will provide a brief overview of our current understanding of human cancer biology that is driving interests in targeted therapy and personalized management. Further development of molecular diagnostics should facilitate earlier detection, more precise prognostication and prediction of response across the spectrum of cancer development. Targeted therapy has already had a dramatic impact on several forms of cancer and strategies are being developed to identify small groups of patients who would benefit from novel targeted drugs in combination with each other or with more conventional surgery, radiotherapy or chemotherapy. Development of personalized interventions – whether preventive or therapeutic in nature – will require multidisciplinary teams of investigators and the infrastructure to match patient samples and agents in real time.

To accelerate translational cancer research, greater alignment will be required between academic institutions, the US National Cancer Institute, the US Food and Drug Administration, foundations, pharma, and community oncologists. Ultimately, new approaches to prevention, detection, and therapy must be sustainable. In the long run, translational research and personalized management can reduce the cost of cancer care which has escalated in recent years. More accurate and specific identification of at-risk members and risk stratification will be helpful to minimize the risks of overdiagnosis and overtreatment, while maximizing the benefits of screening, early detection, and preventive intervention. Patients who would benefit most can be identified and funds saved by avoiding treatment in those whose cancers would not respond. Participation and education of community oncologists will be required, as will modification of practice patterns. For progress in the clinic to occur at an optimal pace, leaders of translational teams must envision a clear path to bring new concepts and new agents from the laboratory to the clinic, to complete pharmaceutical or biological development, to obtain regulatory approval, and to bring new strategies for detection, prevention, and treatment to patients in the community.

In a series of additional volumes regarding translational cancer research, several topics are explored in greater depth, including Biomarkers, Targeted Therapy, Immunotherapy, and this volume concerning Cancer Gene Therapy by Viral and Non-Viral Vectors. The purpose of these books has been not only to describe different strategies for particular forms of cancer, but also to identify some of the barriers to translation using different reagents or different strategies around common therapeutic or diagnostic modalities. Potential barriers include not only the need for a deeper understanding of science, methods to overcome the challenge of tumor heterogeneity, the development of targeted therapies, the availability of patients with an appropriate phenotype and genotype within a research center with the investigators, research teams, and infrastructure required for clinical/translational research and the design of novel trials, but also adequate financial support, a viable connection to diagnostic and pharmaceutical development, and a strategy for regulatory approval, as well as for dissemination in the community.

Cancer Gene Therapy by Viral and Non-Viral Vectors considers many of these areas, including the strengths and limitations of the several types of viral and non-viral delivery systems, the potential importance of tumor-specific promoter systems, examples of where gene therapy has succeeded, the challenge of targeting all cancer cells, the advantages of targeting the tumor stroma and immunocytes, and the logistic barriers to preparation of materials required for clinical trials. The need for substantial antitumor activity and the importance of clinical responses in phase I–II trials are also highlighted. Overall, this volume provides substantial perspective regarding the translational potential of cancer gene therapy.

Robert C. Bast
Maurie Markman
Ernest Hawk

Preface

The idea of gene therapy was first proposed to correct errors associated with genetic disease by supplementing defective or missing genes. Advances in DNA technology and in understanding the basis of genetic diseases gave high hopes that gene therapy would be the next big breakthrough in medicine. However, the journey ahead was not without challenges and roadblocks. In 1999, a tragedy occurred when an 18-year-old gene therapy trial participant, Jessie Gelsinger, died 4 days after receiving adenoviral treatment for a genetic disorder from a massive immune response that led to multiple organ failure. This incident caused a major setback in the gene therapy field and the US Food and Drug Administration placed a hold on active gene therapy trials. Yet another event that followed brought more bad news to the field. In 2003, five patients who received CD34+ hematopoietic (bone marrow) stem cells transduced with a retrovirus carrying the interleukin-2 receptor γ chain gene to treat inherited X-linked severe combined immunodeficiency (SCID-XI) developed T-cell leukemia. One patient later died. Despite the dismissal of promising hopes of gene therapy in the early days as a result of these events, there is now optimism as more current research data have shown substantial progress in the clinical development of gene therapy after years of intense investigations to improve vector design and safety. Several successful gene therapy trials, including treatment of an inherited eye disease (Leber’s congential amaurosis), Parkinson’s disease, blood disorders, SCID-XI, adenosine deaminase-deficient SCID, and Siemerling–Creutzfeldt disease (X-linked adrenoleukodystrophy), have been reported in the last few years. Most recently, two studies published in July 2013 in Science reported clinical efficacy in lentivirial-mediated gene therapy to treat metachromatic leukodystrophy and Wiskott–Aldrich symdrome (see Chapter 2 for more details). In addition to human trials, studies conducted in canines showed that achromatopsia (an inherited form of total color blindness), diabetes, and Duchenne muscular dystrophy were successfully treated by gene therapy; these encouraging findings will undoubtedly continue to pave the way for conducting human clinical trials to develop new drugs to treat these diseases.

While no gene therapy has yet been approved in the USA, two have been approved for use in other parts of the world. The Chinese State Federal Drug Administration approved the world’s first gene therapy to treat head and neck cancer using Gencidine, an adenoviral vector expressing tumor suppressor p53. However, concerns about the therapeutic efficacy have been raised [1], and there are no further reported clinical outcomes after a decade of approval. In 2012, the European Commission approved the first gene therapy product (Glybera) in the Western world to treat lipoprotein lipase deficiency, a rare inherited disease of fat metabolism. The company uniQure is currently seeking regulatory approval in the USA, Canada, and other countries.

Cancer, cardiovascular, and infectious diseases, among many others, are also targets of gene therapy. Adenovirus remains the most popular type of vector used in gene therapy clinical trials worldwide, followed by retrovirus, naked/plasmid DNA, vaccinia virus, and lipofection in the top five. For viral vectors, the important parts of the virus required for gene delivery are kept and those that are not required are deleted, and the development of self-inactivating integrating viruses such as retrovirus and lentivirus eliminates the transactivation of neighboring genes after integration. Current investigations also continue to broaden the viral vectors’ cell host range. For non-viral vectors, improvements have focused on the delivery system for therapeutic agents, including plasmid DNA, RNA interference (RNAi), and microRNAs, by increasing cellular uptake, protecting against microphage digestion, and optimizing nucleic acid payload release.

In the USA, clinical studies must be reviewed by regulatory committees such as the Institutional Review Board (IRB), Food and Drug Administration (FDA), Institutional Biosafety Committee (IBC), and Recombinant DNA Advisory Committee (RAC). Moreover, manufacture must also comply with Good Manufacturing Practice (GMP) guidelines set out by the FDA. The development of sufficient manufacturing capacity to meet the clinical demands after gene therapy attains approval is another concern.

The discovery of monoclonal antibodies brought much excitement as a new treatment modality in early 1980s. However, the lack of efficacy and the rapid clearance of murine monoclonal antibodies due to the development of human antimouse antibodies in patients led to the failure of many clinical trials. Nonetheless, perseverance allowed the development of technological improvements resulting in the eventual clinical success of monoclonal antibodies, which are now a standard approach for producing therapeutics targeting cell surface receptors. In a similar way, further improvements in gene therapy may allow this approach to follow the successful journey of monoclonal antibodies.

To ensure that gene therapy can be successfully developed into new drugs following the fate of monoclonal antibodies, there are several areas needing critical improvement, including efficient delivery, specificity, and well-designed clinical trials. In this book, we have invited experts to discuss the current updates on cancer gene therapy. The opening chapter by Cerullo et al. describes various types of viral therapy, particularly DNA viruses (adenovirus, vaccinia virus, herpes virus, parvovirus) and provides examples of their use in clinical studies. The following chapter by Zhou et al. focuses on the principal types and evolution of lentiviruses in cancer and HIV therapy with special interest in gene silencing by RNAi. The next two chapters describe non-viral delivery systems. First, in Chapter 3 Najjar et al. review various methods of plasmid DNA delivery, optimization of gene expression, and their application for therapy including cancer. Satterlee and Huang then explain in Chapter 4 the design and challenges of nanoparticles to deliver therapeutic RNAi. In the second part, starting with Chapter 5, Hsu et al. provide an introduction to the clinical applications of tissue-specific and cancer-targeting promoters in cancer gene therapy. As aberrant microRNA expression has been implicated in promoting and initiating carcinogenesis, in Chapter 6 Ling and Calin present an overview of the role of microRNAs in cancer and other diseases and discuss examples of anti-microRNA therapeutics.

The last part of the book provides some insight on the regulatory compliance of gene therapy clinical trials focusing on manufacturing regulations of viral vectors by Gee and Mei in Chapter 7 and review processes and requirements prior to obtaining FDA approval by Grilley in Chapter 8. In the closing chapter, Brenner discusses the tasks that must be accomplished to make gene therapy drugs more broadly applicable and the improvements in clinical trial design, as the development pathway of cancer gene therapy is distinct from and more complex than the traditional pharmaceutical model. It is our hope that this book can facilitate the maturation of gene therapy for its clinical application.

Malcolm K. Brenner
Mien-Chie Hung

Cancer Gene Therapy by Viral and Non-viral Vectors

TRANSLATIONAL ONCOLOGY

List of Contributors

Series Foreword

Preface

Reference

Part 1
Delivery Systems