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Volume 1 Number 2 2007 .jpg)
CONTENTS AND ABSTRACTS
Dominic J. Glover (Australia), Ljudmila Glouchkova, Hans J. Lipps (Germany), David A. Jans (Australia) Overcoming Barriers to Achieve Safe, Sustained and Efficient Non-Viral Gene Therapy (pp 126-140)
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ABSTRACT
Invited Review: Adverse events in using viral-based vectors to deliver and integrate therapeutic DNA into patients in recent clinical trials have revitalized efforts to improve the efficacy of the alternative, the much safer non-viral vectors. Non-viral vectors are generally orders of magnitude less efficient than viral counterparts due their inability to overcome numerous biological barriers to DNA delivery, but deeper understanding of these barriers, and formulation of new approaches to transcend them, has enabled the generation of non-viral vectors for DNA delivery that comprise multiple functional modules mimicking the ability of viruses to traffic through the cell to the nucleus. These novel, multifunctional vectors may be combined with strategies for episomal replication and faithful expression of therapeutic DNA bereft of the potential dangers of integration into the host genome, with enormous potential for future clinical as well as research applications.
Frank Alderuccio, James Chan, Ban-Hock Toh (Australia) Haematopoietic Stem Cell Based Gene Therapy as a Strategy to Treat Autoimmune Disease (pp 141-149)
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ABSTRACT
Invited Review: Autoimmune diseases affect 5-6% of the Caucasian population and represent a major burden on an individual’s life-style and the nation’s health budget. There have been no cures described with current treatment based on replacement therapy and/or broad immunosuppressive agents. A challenge for research in this field is to devise strategies aimed at specifically abrogating the immune response to self-antigens while retaining an immune system that can combat foreign pathogens. The ability of the bone marrow compartment to confer immunological tolerance to a recipient is well established and the use of allogeneic and autologous haematopoietic stem cell transplantation (HSCT) to treat autoimmune disease have been reported for a number of autoimmune diseases. Results vary between trials and diseases. A critical observation is the significant rate of relapses with autologous HSCT in humans with autoimmune disease that suggest that this may not be a curative strategy. However, the ability to easily harvest autologous haematopoietic stem cells and genetically manipulate them, opens up avenues that can be evaluated in the quest to enhance HSCT as a curative strategy. The use of gene therapy to target autoantigen to HSCs in the autoimmune diseases to establish molecular chimeras and antigen-specific tolerance is still in its infancy. Nonetheless, the few experimental studies that have directly used this approach, coupled with approaches to clear the periphery of self-reactive T cells indicates that genetically-engineered HSCT may be a feasible and viable option for the curative treatment of the autoimmune diseases.
Matthew J. Mellon, Theodore F. Logan, Thomas A. Gardner (USA) Current Gene Therapy Strategy for Renal Cell Carcinoma (pp 150-160)
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ABSTRACT
Invited Review: Renal cell carcinoma (RCC) is the third most common urologic neoplasm and its age-adjusted incidence has been increasing annually. Despite advances in cancer research and current therapy strategies, the effective treatment of advanced RCC remains elusive. Over the past two decades, clinical and genetic studies have shown that kidney cancer is not a single disease entity, rather a spectrum of different forms. This understanding has translated into therapies targeted against specific genetic targets. In this review, we highlight the scientific principles, current applications, and future direction of renal cell cancer gene therapy.
Josué K. Mfopou, Luc Bouwens (Belgium) Milestones of Pancreatic Beta Cell Differentiation from Embryonic Stem Cells (pp 161-171)
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ABSTRACT
Invited Review: Transplantation of pancreatic islets of Langerhans has proven effective in setting diabetic patients insulin-free for periods longer than a year, resulting in a net improvement in generic measures of quality of life. Although very promising, this treatment cannot yet be generally applied because of shortage in brain-dead heart-beating islet donors. Several non-endocrine pancreatic cells and non-pancreatic cells are presently investigated for their possible use as alternative sources of islets needed for diabetes cell therapy. Among them, embryonic stem (ES) cells are valued for their high proliferative capacity and their potential to generate progenies of all three developmental germ layers. Till date, several attempts to generate pancreatic epithelial cells, namely insulin-producing beta cells from rodent and primate ES cells have been initiated by different groups and resulted in diverse success rates. One of the most striking limiting factors has been the lack of a comprehensive efficient and reproducible protocol taking into account the main features of normal pancreas development in vivo. This review focuses on the successive achievements in this field, with special credits to the ideological milestones of the so far proposed protocols for generation of beta cells from ES cells.
Lucía Alonso-González, Stefan Wagner, Götz Laible (New Zealand) Emerging Gene and Cell-Based Therapies and Their Prospects for the Treatment of Animal Diseases (pp 172-185)
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ABSTRACT
Invited Review: Animal diseases caused by pathogens such as viruses, bacteria and nematodes account for major losses in livestock production and can have dramatic socioeconomic consequences in agriculture-dependant countries. Moreover, in recent years, the unpredictable worldwide outbreaks of pandemic infections such as mad-cow disease and bird flu, and the risks that these diseases pose for the human population, have raised public awareness about the importance of developing strategies for their treatment. The rapid development that cell and molecular genetics has experienced in the past 25 years has provided scientists with powerful new weapons which, unlike the more traditional medical approaches, can directly target the underlying molecular causes of disease. In this review, we will outline the most recent advances in cell and gene therapy applied to the treatment of animal diseases, with a particular focus on livestock animals. Relevant examples have been used to illustrate these novel treatment options and current restrictions, and future prospects for their application to combat animal diseases will be discussed.
Valeria Casotti, Marie-Luise Syren, Giorgia Dina, Elena Copreni, Sante Di Gioia, Donatella Piro, Massimo Conese (Italy) Gene Transfer in Differentiated Primary Rat Tracheal Epithelial Cells by Non-Viral Vectors (pp 186-191)
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ABSTRACT
Research Note: The cellular barriers to efficient gene transfer into the airways are still poorly understood. Rat tracheal epithelial (RTE) cells faithfully reproduce the mucociliary, pseudostratified characteristics of the in vivo human airway epithelium and are a suitable model for studying major pathways of airway cell differentiation. We studied the efficiency of various non-viral cationic vectors in the primary culture of RTE cells grown in different conditions. When we grew RTE cells on collagenated plastic dishes for 4 days in basal conditions (25 ng/ml EGF), a poorly-differentiated (PD) phenotype appeared, whether or not retinoic acid (RA) was present in culture medium. The cells were transfected with a plasmid encoding a luciferase reporter gene under the control of the cytomegalovirus promoter. The highest luciferase levels were obtained with a cationic lipid (DOTAP) and a cationic polymer (PEI). When cells were grown for 12 days on collagen-coated inserts (Transwells®) at the air-liquid interface in the presence of RA, the differentiation was towards a well-differentiated phenotype, as assessed by electron microscopy. Conversely, in the absence of RA, the cells acquired a flattened undifferentiated phenotype. In the absence of RA, the transfection levels were 3 to 8.8-fold higher than in the presence of RA. However, in differentiated cells, PEI gave transfection levels closer to those obtained in PD cells than DOTAP. These results support our view that PEI can overcome the barriers imposed by a differentiated phenotype.
Michèle Amouyal (France) Use of Histones to Increase the Frequency of Recombinant Plasmid Formation during Molecular Cloning (pp 192-197)
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ABSTRACT
Research Note: By compacting DNA in eukaryotes, histones bring closer two distant regions on DNA. This is likely to facilitate the work of transcriptional enhancers at a long distance in gene regulation (Amouyal 1991). Since the classical strategy for plasmid engineering involves a step where the recombinant fragment is circularized, this has prompted the author to test this hypothesis by plasmid engineering. In fact, the unfractionated mixture of calf thymus histones facilitates the in vitro ligation of two fragments, resulting in an increased number of transformed E. coli cells under the appropriate conditions for about 7 and 12 kb large recombinant plasmids. This property can be exploited for plasmid engineering, when inserting a DNA fragment into a plasmid vector becomes increasingly difficult as the recombinants get larger. The same effect was obtained with calf thymus histone H2B by itself, but was less marked with the linker histone H1. |
