Gene therapy retroviral

Fig. 22.2. Schematic for retroviral gene therapy. Retroviral particles containing RNA that encodes the gene of interest are prepared and added to the recipient cells. The retroviral vector is internalized and then releases its genetic material into the cytoplasm. Reverse transcriptase uses the viral template to produce a DNA copy of the desired gene and this gene can then be integrated into the recipient cell s genome. The integrated gene then serves as a template for transcription into mRNA and translation into protein.

Gene therapy uses cloned DNA in adeno or retroviral vectors to transduce host cells for longterm therapy of certain genetic disorders with defined... 

A nuclear receptor that is a key transcription factor in adipocytes. It plays a critical role in the control of adipocyte differentiation and is involved in the regulation of the expression of specific adipokines, including leptin and adiponectin. It has anti-inflammatory actions and is the target of the thiazolidinedione drugs. The preintegration complex is a complex of retroviral DNA and proteins that translocates from the cytosol into the nucleus prior to integration. Gene Therapy... 

Once assimilated by the cell, the exogenous nucleic acid must now travel/be delivered to the nucleus. In some cases, the mechanism by which this transfer occurs is understood, at least in part (e.g. in the case of retroviral vectors). In other cases (e.g. use of liposome vectors or naked DNA), this process is less well understood. At a practical level, gene therapy protocols may entail one of three different strategies (Figure 14.3). 

Some 24 per cent of all gene therapy clinical trials undertaken to date have employed retroviral vectors as gene delivery systems. Retroviruses are enveloped viruses. Their genome consists of ssRNA of approximately 5-8 kb. Upon entry into sensitive cells, the viral RNA is reverse transcribed and eventually yields double-stranded DNA. This subsequently integrates into the host cell genome (Box 14.1). The basic retroviral genome contains a minimum of three structural genes ... 

The theoretical complications posed by random chromosomal integration became a medical reality in 2002, when two children who had received retroviral-based gene therapy 2 years previously developed a leukaemic-like condition. The initial clinical trial aimed to treat X-linked severe combined immunodeficiency (SCID-X1), a hereditary disorder in which T-lymphocytes and NK cells in particular do not develop, due to a mutation in the gene coding for the yc cytokine receptor subunit. The clinical consequence is near abolition of a functional immune system. 

The trial entailed retroviral-mediated ex vivo transduction of haematopoietic stem cells from 10 young SCID-X1 sufferers, with subsequent re-infusion of the treated cells. A marked and prolonged clinical response in which the condition was essentially reversed was observed in 9 out of the 10 patients. The prolonged response was likely due to the transduction of pluripotent progenitor cells with self-renewal capacity (Chapter 10). However, the two youngest patients (1 and 3 months old at the time of treatment) developed uncontrolled proliferation of mature T-lymphocytes 30 months and 34 months after gene therapy respectively. 

An alternative system, which effectively results in the introduction of antisense oligonucleotides into the cell, entails application of gene therapy. In this case, a gene, which when transcribed yields (antisense) mRNA of appropriate nucleotide sequence, is introduced into the cell by a retroviral or other appropriate vector. This approach, as applied to the treatment of cancer and AIDS, is being appraised in a number of trials. 

Gene therapy for this metabolic defect may become available within the next few years. In vitro studies have demonstrated the feasibility of retroviral-mediated gene transfer of both the El-a and E2 subunits of the branched-chain decarboxylase complex [16,18],... 

Anson DS. The use of retroviral vectors for gene therapy - what are the risks A review of retroviral pathogenesis and its relevance to retroviral vector-mediated gene delivery. Genet Vaccines Ther 2004 2. 

Yi Y. Retroviral gene therapy safety issues and possible solutions. Curr Gene Ther 2005 5(l) 25-35. 

Table 11.2. Vector systems used to deliver genes into mammalian cells. To date, the majority of clinical trials undertaken have utilized retroviral vector systems. Non-viral systems have generally been employed least often, although some, e.g. nucleic acid-containing liposomes, may be used more extensively in the future. Some of the methods tested, e.g. calcium phosphate precipitation, electroporation and particle acceleration, are unlikely to be employed to any great extent in gene therapy protocols...

Figure 11.6. Large-scale manufacture of retroviral vectors for use for gene therapy-based clinical protocols. Refer to text for details...

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