Comparison of Vectors
Objective studies comparing the relative merits of different delivery systems
are rarely carried out, making comparisons difficult. Lozier
et al (1997), compared the merits of retroviruses,
adenoviruses & liposomes for transducing gut epithelial cells & found
that all three methods efficiently expressed the transgene. The ideal vector combines
features of all the delivery systems described above. Preparation of this ideal
vector would be relatively simple (e.g. not involving multiple steps such as attachment
of a ligand targeting a particular cell type) & result in high vector concentrations
(>108 particles/ml). To allow subsequent readministration &
avoid undesired host reactions there would be no significant immune response to
any component of the vector. The lack of an immune response may allow transgene
expression to be sufficiently prolonged from episomal systems, such that readministration
is not necessary. Alternatively, integration into the host genome, preferably
in a site specific location, would ensure that the transgene is not lost during
the lifetime of the cell. Whatever the means of delivery, transcriptional control
as exerted by the host cell or by the physician, would enable greater tissue specificity
& regulation of the transgene expression. As yet no one vector has all these
requirements, but improvements are being made to all systems, e.g. immunological
problems of adenoviruses, production problems of AAV, the short duration of retroviruses,
etc. The genomes of viral vectors are being progressively reduced to contain the
transgene & a minimum of maintenance genes, whilst developments to molecular
conjugates & liposomes are making them more like viruses.
It is likely that all delivery systems will find a niche, either as dictated by biological needs, e.g. HSV-1 for neuronal applications, or because the expense of developing a system to replace an existing functional method would be prohibitive. A very large number of research groups have invested considerable effort in developing a given system & are unlikely to abandon it before a therapeutic application has been found.
Promoter Control:
A recurring theme throughout this review has been the value of tissue specific promoters, allowing a second level of control over transgene expression, in addition to that of selective transduction by the vector. The most frequently used promoters are viral in origin, often derived from a different virus than the vector backbone, for example cytomegalovirus promoters have been used in all vector systems. Viral promoters have the advantages of being smaller, stronger & better understood than most human promoter sequences. One of the drawbacks of viral promoters is their tendency to be silenced when there is an immunological response. Qin et al (1997) showed that a range of commonly used viral promoters were silenced by IFN-gamma & TNF-alpha at the level of mRNA stability. By contrast a constitutive human promoter, beta actin, was less effected.
A range of cellular promoters have been developed for specific tissues including the liver (albumin; Miyatake et al, 1997), muscle (myosin light chain 1; Shi et al, 1997) & endothelial cells (von Willebrandt; Ozaki et al, 1996 & smooth muscle 22a; Kim et al, 1997). Tumour specific promoters are also being used in developing cancer therapies, including tyrosine kinase for B16 melanoma (Diaz et al, 1998), DF3/MUC1 for certain breast cancers (Wen et al, 1993) & afetoprotein for hepatomas (Chen et al, 1995). The temporal expression of the transgene construct can be controlled by drug inducible promoters, for example by including cAMP response element enhancers in a promoter, cAMP modulating drugs can be used (Suzuki et al, 1996). Alternatively repressor elements can prevent transcription in the presence of the drug (Hu et al, 1997). Spatial control of expression has been developed by using ionising radiation (radiotherapy) in conjunction with the erg1 gene promoter (Hallaham et al, 1995).
Despite the plethora of different promoters that have been identified, their behaviour in the context of a vector can be very difficult to predict (Harris & Lemoine, 1996). Regulatory elements may be distant from the promoter sequence & when included within a smaller distance may fail to operate. With greater understanding of the human genome, consensus sequences can be constructed to improve the strength of cellular promoters (Suzuki et al, 1996).
<<< Previous Section | Next Section >>>
© MicrobiologyBytes 2004.