Current Gene Therapy (v.11, #4)

Advances and Future Challenges in Adenoviral Vector Pharmacology and Targeting by Reeti Khare, Christopher Y. Chen, Eric A. Weaver, Michael A. Barry (241-258).
Adenovirus is a robust vector for therapeutic applications, but its use is limited by our understanding of itscomplex in vivo pharmacology. In this review we describe the necessity of identifying its natural, widespread, and multifacetedinteractions with the host since this information will be crucial for efficiently redirecting virus into target cells. Inthe rational design of vectors, the notion of overcoming a sequence of viral “sinks” must be combined with re-targeting totarget populations with capsid as well as shielding the vectors from pre-existing or toxic immune responses. It must alsobe noted that most known adenoviral pharmacology is deduced from the most commonly used serotypes, Ad5 and Ad2.However, these serotypes may not represent all adenoviruses, and may not even represent the most useful vectors for allpurposes. Chimeras between Ad serotypes may become useful in engineering vectors that can selectively evade substantialviral traps, such as Kupffer cells, while retaining the robust qualities of Ad5. Similarly, vectorizing other Ad serotypesmay become useful in avoiding immunity against Ad5 altogether. Taken together, this research on basic adenovirus biologywill be necessary in developing vectors that interact more strategically with the host for the most optimal therapeuticeffect.

Antisense oligomers initially showed promise as compounds to modify gene expression, primarily throughRNaseH induced degradation of the target transcript. Expansion of the field has led to new chemistries capable of invokingdifferent mechanisms, including suppression of protein synthesis by translational blockade and gene silencing usingshort interfering RNAs. It is now apparent that the majority of the eukaryotic genome is transcribed and non-protein codingRNAs have been implicated in the regulation of gene expression at many levels. This review considers potentialtherapeutic applications of antisense oligomers to modify gene expression, primarily by interfering with the process ofexon recognition and intron removal during gene transcript splicing. While suppression of gene expression will be necessaryto address some conditions, it is likely that antisense oligomer splice modification will have extensive clinical application.Pre-mRNA splicing is a tightly co-ordinated, multifactorial process that can be disrupted by antisense oligomers ina highly specific manner to suppress aberrant splicing, remove exons to by-pass nonsense or frame-shifting mutations orinfluence exon selection to alter spliceoform ratios. Manipulation of splicing patterns has been applied to a diverse rangeof conditions, including ..-thalassemia, Duchenne muscular dystrophy, spinal muscular atrophy and certain cancers. Alternativeexon usage has been identified as a major mechanism for generating diversity from a limited repertoire of genesin higher eukaryotes. Considering that the majority of all human primary gene transcripts are reportedly alternativelyspliced, intervention at the level of pre-mRNA processing is likely to become increasingly significant in the fight againstgenetic and acquired disorders.

Prospects and Limitations of T Cell Receptor Gene Therapy by Annelies Jorritsma, Remko Schotte, Miriam Coccoris, Moniek A. de Witte, Ton N.M. Schumacher (276-287).
Adoptive transfer of antigen-specific T cells is an attractive means to provide cancer patients with immune cellsof a desired specificity and the efficacy of such adoptive transfers has been demonstrated in several clinical trials. Becausethe T cell receptor is the single specificity-determining molecule in T cell function, adoptive transfer of TCR genes intopatient T cells may be used as an alternative approach for the transfer of tumor-specific T cell immunity. On theoreticalgrounds, TCR gene therapy has two substantial advantages over conventional cellular transfer. First, it circumvents thedemanding process of in vitro generation of large numbers of specific immune cells. Second, it allows the use of a set ofparticularly effective TCR genes in large patient groups. Conversely, TCR gene therapy may be associated with a numberof specific problems that are not confronted during classical cellular therapy. Here we review our current understanding ofthe potential and possible problems of TCR gene therapy, as based on in vitro experiments, mouse model systems andphase I clinical trials. Furthermore, we discuss the prospects of widespread clinical application of this gene therapy approachfor the treatment of human cancer.

Polycation-Based Gene Therapy: Current Knowledge and New Perspectives by Marcio J. Tiera, Qin Shi, Francoise M. Winnik, Julio C. Fernandes (288-306).
At present, gene transfection insufficient efficiency is a major drawback of non-viral gene therapy. The 2 maintypes of delivery systems deployed in gene therapy are based on viral or non-viral gene carriers. Several non-viral modalitiescan transfer foreign genetic material into the human body. To do so, polycation-based gene delivery methods mustachieve sufficient efficiency in the transportation of therapeutic genes across various extracellular and intracellular barriers.These barriers include interactions with blood components, vascular endothelial cells and uptake by the reticuloendothelialsystem. Furthermore, the degradation of therapeutic DNA by serum nucleases is a potential obstacle for functionaldelivery to target cells. Cationic polymers constitute one of the most promising approaches to the use of viral vectors forgene therapy. A better understanding of the mechanisms by which DNA can escape from endosomes and traffic to enterthe nucleus has triggered new strategies of synthesis and has revitalized research into new polycation-based systems. Theobjective of this review is to address the state of the art in gene therapy with synthetic and natural polycations and the latestadvances to improve gene transfer efficiency in cells.

Adenoviral Vector Immunity: Its Implications and Circumvention Strategies by Yadvinder S. Ahi, Dinesh S. Bangari, Suresh K. Mittal (307-320).
Adenoviral (Ad) vectors have emerged as a promising gene delivery platform for a variety of therapeutic andvaccine purposes during last two decades. However, the presence of preexisting Ad immunity and the rapid developmentof Ad vector immunity still pose significant challenges to the clinical use of these vectors. Innate inflammatory responsefollowing Ad vector administration may lead to systemic toxicity, drastically limit vector transduction efficiency and significantlyabbreviate the duration of transgene expression. Currently, a number of approaches are being extensively pursuedto overcome these drawbacks by strategies that target either the host or the Ad vector. In addition, significant progresshas been made in the development of novel Ad vectors based on less prevalent human Ad serotypes and nonhumanAd. This review provides an update on our current understanding of immune responses to Ad vectors and delineates variousapproaches for eluding Ad vector immunity. Approaches targeting the host and those targeting the vector are discussedin light of their promises and limitations.

Immune Responses to AAV in Clinical Trials by Federico Mingozzi, Katherine A. High (321-330).
Findings in the first clinical trial in which an adeno-associated virus (AAV) vector was introduced into the liverof human subjects highlighted an issue not previously identified in animal studies. Upon AAV gene transfer to liver, twosubjects developed transient elevation of liver enzymes, likely as a consequence of immune rejection of transduced hepatocytesmediated by AAV capsid-specific CD8+ T cells. Studies in healthy donors showed that humans carry a populationof antigen-specific memory CD8+ T cells probably arising from wild-type AAV infections. The hypothesis formulated atthat time was that these cells expanded upon re-exposure to capsid, i.e. upon AAV-2 hepatic gene transfer, and clearedAAV epitope-bearing transduced hepatocytes. Other hypotheses have been formulated which include specific receptorbindingproperties of AAV-2 capsid, presence of capsid-expressing DNA in AAV vector preparations, and expression ofalternate open reading frames from the transgene; emerging data from clinical trials however fail to support these competinghypotheses. Possible solutions to the problem are discussed, including the administration of a short-term immunosuppressionregimen concomitant with gene transfer, or the development of more efficient vectors that can be administered atlower doses. While more studies will be necessary to define mechanisms and risks associated with capsid-specific immuneresponses in humans, monitoring of these responses in clinical trials will be essential to achieving the goal of longtermtherapeutic gene transfer in humans.