Current Gene Therapy - Volume 9, Issue 3, 2009

Dysfunction in or lack of osteoclasts result in osteopetrosis, a group of rare but often severe, genetic disorders characterized by an increase in bone mass, skeletal malformations and bone marrow failure that may be fatal. Several of the underlying defects have lately been characterized in humans and in animal disease models. In humans, these defects often involve mutations in genes expressing proteins involved in the acidification of the osteoclast sub-cellular compartment, a process necessary for proper bone resorption. So far, the only cure for children with severe osteopetrosis is allogeneic hematopoietic stem cell transplantation (SCT). However, the characterization of the genetic defects opens up the possibility for gene replacement therapy as an alternative to SCT. Recently, gene therapy targeting hematopoietic stem cells (HSC) in a mouse model of infantile malignant osteopetrosis was shown to correct many aspects of the disease. Here we review important aspects of this group of diseases and discuss the prospects for development of gene therapy of osteopetrosis.

Nuclear factor (NF)-κB is regarded as one of the most important transcription factors and plays an essential role in the transcriptional activation of pro-inflammatory cytokines, cell proliferation and survival. NF-κB can be activated via two distinct NF-κB signal transduction pathways, the so-called canonical and non-canonical pathways, and has been demonstrated to play a key role in a wide range of inflammatory diseases and various types of cancer. Much effort has been put in strategies to inhibit NF-κB activation, for example by the development of pharmacological compounds that selectively inhibit NF-κB activity and therefore would be beneficial for immunotherapy of transplantation, autoimmune and allergic diseases, as well as an adjuvant approach in patients treated with chemotherapy for cancer. Gene therapy targeting NF-κB is a promising new strategy with the potential of long-term effects and has been explored in a wide variety of diseases, ranging from cancer to transplantation medicine and autoimmune diseases. In this review we discuss recent progress made in the development of NF-κB targeted gene therapy and the evolution towards clinical application.

Infectious diseases caused by virulent bacteria are a significant cause of morbidity and mortality worldwide, especially in developing countries. However, attenuated strains derived from pathogenic bacteria, such as Salmonella, are highly immunogenic and can be used as vaccines to promote immunity against parental pathogenic bacteria strains. Further, they can be genetically manipulated to either express foreign antigens or deliver exogenous DNA, in order to induce immunity against other pathogens or antigens. Contrarily, specific structural modifications in attenuated Salmonella have allowed the generation of strains that can be well tolerated by the immune system and reduce inflammatory responses. It is thought that those strains could be considered as vectors to promote specific immune tolerance for certain auto-antigens or allergens and reduce unwanted or self-reactive immune responses. In addition, some structural features of Salmonella can contribute to defining the nature and type of polarization of the adaptive immune response induced after immunization, which can be considered as a tool to modulate antigen-specific immunity. In this article we discuss recent advances in the understanding of immune system modulation by molecular components of bacteria and their exploitation for the rational induction of pathogen immunity or antigen-specific tolerance.

Allergic diseases, such as allergic asthma, allergic rhinitis, atopic dermatitis, conjunctivitis, urticaria, food allergy, and/or anaphylaxis, are associated with the skewing of immune responses towards a T helper 2 (TH2) phenotype, resulting in eosinophilic inflammation. TH2 cytokines, such as interleukin (IL)-4, IL-5 and IL-13, promote IgE production, mast cell differentiation, and eosinophil growth, migration and activation which then lead to the pathologic abnormalities in allergic diseases. Moreover, the impaired function of regulatory T cells has been noted in allergic diseases. To date, treatments for allergic diseases, such as antihistamines, corticosteroids, bronchodilators and some allergen-specific immunotherapy, are effective but costly and require long-term and recurrent drug administration. Gene therapy has been shown to be an easy, effective, and convenient treatment by delivering the allergen or the therapeutic protein in the form of plasmid DNA in vivo to modulate allergic immune responses. We summarize here the recent advances of gene therapy in allergic diseases and discuss the challenges in clinical application.

A decade after its discovery, RNA interference has proven to be an instant success both in fundamental research and clinical applications. Lentiviral delivery of shRNAs is one of the most popular approaches to study gene functionalities in both developmental biology and disorders. During the past 10 years, several adaptations and novel techniques have emerged to improve (conditional) transgene expression and to meet researchers' needs. However, due to this magnitude of diversity, it is sometimes difficult to select the most suitable approach for a specific experimental setup. Here, we summarize the different systems and techniques available for every step in the generation of shRNA-bearing lentiviruses. The most crucial point is inevitably the selection of the target sequence itself. A good shRNA design is indispensable and determines almost completely the success of the experiments. In addition, an adequate promoter that drives the shRNA expression has to be chosen depending on its strength, inducibility, tissue-specificity, … At this point, the researcher has also to decide whether the expression of the shRNA should be inducible or not. Another point one has to keep in mind is the choice of lentiviral vector in which the silencing cassette will be incorporated; single- or double-copy vectors are available. The last 2 years, shRNA multiplex approaches in which several targets are silenced with one vector have emerged and have shown a lot of potential in complex studies (like HIV-1). Finally, in the last section, we will discuss the possible induction of an immune response by short dsRNA molecules.

Over the past ten years, a variety of imaging techniques have been developed that allow non-invasive detection of gene expression within the brain of intact mammals, ranging from mouse to man. The basic concepts of these imaging techniques, including positron emission tomography, single photon emission computed tomography, magnetic resonance imaging and spectroscopy, bioluminescence imaging and fluorescent imaging, are discussed. The expression of imaging reporter genes can be detected and quantified by these imaging techniques, which allow to unravel the temporospatial dynamics of gene expression within the intact living animal. Different imaging reporter genes have been developed each with their specific use in the basic and clinical neurosciences. Applications of reporter gene imaging can be found in neurooncology, infectious disease of the central nervous system, brain gene transfer, neural cellular therapy and in transgenic mice. Strategies that aim to image gene expression based on detection of mRNA levels have also been developed. We anticipate that these techniques will have a strong impact on preclinical neuroscience and will be of utmost importance in the implementation of gene and cell therapy for diseases of the brain.