Current Gene Therapy , Volume 2, No. 3, 2002
Tropism-Modified Adenoviral and
Adeno-Associated Viral Vectors for Gene Therapy Pp.273-293
S.A.
Nicklin and A.H. Baker
Non-Viral
Approach toward Gene Therapy of Cystic Fibrosis Lung Disease Pp.295-305
A.
Bragonzi and M. Conese
Influence of the Bystander Effect on
HSV-tk/GCV Gene Therapy. A Review. Pp.307-322
I.J.
van Dillen, N.H. Mulder, W. Vaalburg, E.F.J. de Vries and G.A.P. Hospers
Transductional Targeting with Recombinant
Adenovirus Vectors
Pp.323-339
V.
Legrand, P. Leissner, A. Winter, M. Mehtali and M. Lusky
Gene Therapy Strategies to Prevent Autoimmune
Disorders Pp.341-354
M.
Trucco, P.D. Robbins, A.W. Thomson and N. Giannoukakis
Polycistronic Viral Vectors Pp.355-378
P.
de Felipe
Improvement of Nonviral Gene Therapy by
Epstein-Barr Virus (EBV)-based Plasmid Vectors Pp.379-392
O.
Mazda
[Back to top] Tropism-Modified Adenoviral and
Adeno-Associated Viral Vectors for Gene Therapy
S.A. Nicklin and A.H. Baker
One of the most
rapidly advancing areas of gene therapy is vector development. For the majority
of gene therapy procedures, efficient and selective transduction would provide
safe and more effective treatments at optimal vector doses. Advances in vector
targeting strategies have been rapid within the field of DNA-based viruses,
particularly adenovirus (Ad) and more recently adeno-associated virus (AAV)
based vectors. Vector targeting at the level of virus: cell interaction can be
achieved using both non-genetic and genetic methodology. Non-genetic approaches
typically utilise bi-specific antibodies that both neutralise wild-type virus
tropism and provide a new cell binding capacity. For genetic targeting
strategies, the virus capsid can be engineered to express foreign ligands that
target selected receptors in the absence or presence of additional modification
to ablate the virus’ natural tropism. This review covers technological advances
that have led to targeting of Ad and AAV and highlights the potential for these
‘designer’ viruses for future gene-based therapeutics.
[Back to top] Non-Viral
Approach toward Gene Therapy of Cystic Fibrosis Lung Disease
A.
Bragonzi and M. Conese
Since Cystic
Fibrosis (CF) is an autosomal recessive disorder due to mutations in the CFTR
(Cystic Fibrosis Transmembrane Conductance Regulator) gene, studies towards a
gene therapy approach to its treatment followed immediately upon the cloning of
the gene. It was demonstrated that the insertion of a single copy of the wild-type
gene restored the normal phenotype in CF cells in vitro. Encouraging results
were obtained in many in vivo model systems (CF transgenic mice) involving
viral as well as non-viral vectors, which demonstrated the recovery of CFTR
function in the airways. These results constituted the basis for human studies.
Of those with a non-viral approach, a total of seven clinical trials using
cationic lipids have reported data on efficiency, efficacy and safety. An
effective gene transfer approach for the treatment of CF lung disease is not
however imminent: low transfection efficiency and poor maintenance of gene
expression are so far the main obstacles on this therapeutic path. On the other
hand, no important adverse effects have been documented and repeated administration
in humans is possible.
The understanding
of tissue and cellular barriers is a prerequisite for the development of more
efficient non-viral gene therapy protocols for CF patients. While cationic
lipids have been shown to be blocked by the mucous airway barrier and not be
able to transfect differentiated respiratory epithelial cells, a new class of
non-viral vectors, cationic polymers, are endowed with chemical and biological
properties that make them more efficient in mediating gene transfer than
lipids. Cationic polymers, such as polyethylenimine, are promising vectors for
CF lung gene therapy.
[Back to top] Influence of the Bystander Effect on
HSV-tk/GCV Gene Therapy. A Review.
I.J. van Dillen, N.H. Mulder, W. Vaalburg, E.F.J. de
Vries and G.A.P. Hospers
Despite the development of new therapeutic strategies, cancer remains incurable in most patients with advanced disease. A recent potential improvement in therapeutic strategies is the concept of suicide gene therapy. After transfection with a suicide gene, cells can convert a harmless prodrug into its toxic metabolite, resulting in selective elimination of these cells. One of the most frequently studied therapeutic strategies is based on transfection with the herpes simplex virus thymidine kinase gene (HSV-tk), followed by ganciclovir administration. Despite promising results in vitro and in vivo, the antitumor effect in clinical trials remains poor, due to very low transfection efficiency. However, high percentages of transfected cells are not mandatory for complete eradication of a tumor in vivo. Transfected tumor cells appear to be capable of inducing the death of neighboring untransfected cells. This cell kill is called the "bystander effect". Various attempts have been made to increase this effect. A substantial bystander effect could overcome the limitations of low transfection efficiency and result in an enhanced and possibly clinically worthwhile antitumor effect in patients. This review is focused on the HSV-tk/GCV system and gives an overview of current knowledge on the bystander effect in vitro and in vivo. In addition, theories concerning its mechanisms and possible approaches to augment this effect are discussed. Finally, we give an overview of clinical trials using suicide gene therapy.
[Back to top] Transductional Targeting with Recombinant
Adenovirus Vectors
V.
Legrand, P. Leissner, A. Winter, M. Mehtali and M. Lusky
Replication-deficient adenoviruses are considered as gene delivery vectors for the genetic treatment of a variety of diseases. The ability of such vectors to mediate efficient expression of therapeutic genes in a broad spectrum of dividing and non-dividing cell types constitutes an advantage over alternative gene transfer vectors. However, this broad tissue tropism may also turn disadvantageous when genes encoding potentially harmful proteins (e.g. cytokines, toxic proteins) are expressed in surrounding normal tissues. Therefore, specific restrictions of the viral tropism would represent a significant technological advance towards safer and more efficient gene delivery vectors, in particular for cancer gene therapy applications. In this review, we summarize various strategies used to selectively modify the natural tropism of recombinant adenoviruses. The advantages, limitations and potential impact on gene therapy operations of such modified vectors are discussed.
[Back to
top] Gene Therapy Strategies to Prevent Autoimmune
Disorders
M. Trucco, P.D. Robbins, A.W. Thomson and N.
Giannoukakis
Autoimmunity accounts for a significant percentage of human disease and remains a challenging syndrome to treat. While systemic immunosuppression can be beneficial, the associated toxicity of the pharmacologic agents necessitates an antigen-specific approach to silence, eradicate or prevent the genesis of autoreactive immune cells. Gene therapy offers the possibility of providing precise antigen-targeted therapies, thereby sparing the patient the significant toxicity associated with lifelong commitment to chemical immunosuppressives. Gene-based therapies could include, but are not limited to the manipulation of immune networks of tolerance by antigen presentig cell engineering, pro-inflammatory cytokine blockade using soluble antagonists expressed from viral vectors as well as modulation of immune regulatory networks. The potential utility of gene therapy strategies promoting tolerance in two model autoimmune disorders, type I diabetes mellitus and rheumatoid arthritis are discussed in this review.
[Back to top]
Polycistronic Viral Vectors
P.
de Felipe
Traditionally,
vectors for gene transfer/therapy experiments were mono- or bicistronic. In the
latter case, vectors express the gene of interest coupled with a marker gene.
An increasing demand for more complex polycistronic vectors has arisen in
recent years to obtain complex gene transfer/therapy effects. In particular,
this demand is stimulated by the hope of a more powerful effect from combined
gene therapy than from single gene therapy in a process whose parallels lie in
the multi-drug combined therapies for cancer or AIDS. In the 1980’s we had only
splicing signals and internal promoters to construct such vectors: now a new
set of biotechnological tools enables us to design new and more reliable
bicistronic and polycistronic vectors.
This article
focuses on the description and comparison of the strategies for co-expression
of two genes in bicistronic vectors, from the oldest to the more recently
described: internal promoters, splicing, reinitiation, IRES, self-processing
peptides (e. g. foot-and-mouth disease virus 2A), proteolytic cleavable sites
(e.g. fusagen) and fusion of genes. I propose a classification of these
strategies based upon either the use of multiple transcripts (with
transcriptional mechanisms), or single transcripts (using
translational/post-translational mechanisms).
I also examine the
different attempts to utilise these strategies in the construction of
polycistronic vectors and the main problems encountered. Several potential uses
of these polycistronic vectors, both in basic research and in therapy-focused
applications, are discussed.
The importance of the study of viral gene expression strategies and the need to transfer this knowledge to vector design is highlighted.
[Back to top] Improvement of Nonviral Gene Therapy by Epstein-Barr Virus (EBV)-based
Plasmid Vectors
O. Mazda
The nonviral gene
transfer technologies include naked DNA administration, electrical or
particle-mediated transfer of naked DNA, and administration of DNA-synthetic
macromolecule complex vectors. Each method has its advantage, such as low
immunogenicity, inexpensiveness, ease in handling, etc., but the common
disadvantage is that the transfection efficiency has been relatively poor as
far as conventional plasmid vectors are involved. To improve the nonviral gene
transfer systems, Epstein-Barr virus
(EBV)-based plasmid vectors (also referred to EBV-based episomal vectors) have
been employed. These vectors contain the EBNA1 gene and oriP element that
enable high transfer efficiency, strong transgene expression and long term
maintenance of the expression. In the current article, I review recent preclinical
gene therapy studies with the EBV plasmid vectors conducted against various
diseases. For gene therapy against malignancies, drastic tumor suppression was
achieved by gancyclovir administrations following an intratumoral injection
with an EBV plasmid vector encoding the HSV1-TK suicide gene. Equiping the
plasmid with carcinoembryonic antigen (CEA) promoter sequences enabled targeted
killing of CEA-positive tumor cells, which was not accomplished by conventional
plasmid vectors without the EBV genetic elements. Transfection with an
apoptosis-inducing gene was also effective in inhibiting tumors. Interleukin
(IL)-12 and IL-18 gene transfer, either local or systemic, induced therapeutic
antitumoral immune responses including augmentation of the cytotoxic T
lymphocyte (CTL) and natural killer (NK) activities, while an autologous tumor
vaccine engineered to secrete Th1 cytokines via the EBV system also induced
growth retardation of tumors. Non-EBV conventional plasmids were much less
effective in eliciting these therapeutic outcomes. Intracardiomuscular transfer
of the b-adrenergic receptor gene induced a significant elevation in cardiac
output in cardiomyopathic animals, suggesting the usefulness of the EBV system
in treating heart failure. The EBV-based nonviral delivery also worked as
genetic vaccine that triggered prophylactic cellular and humoral immunity
against acute lethal viral infection. All the nonviral delivery vehicles so far
tested showed an improved transfection rate when combined with the EBV-plasmids.
Collectively, the EBV-based plasmid vectors may greatly contribute to nonviral
gene therapy against a variety of disorders, including malignant, congenital,
chronic and infectious diseases.