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Small interfering RNA (siRNA) is an emerging methodology in reverse genetics. However, the challenging hurdle of achieving tissue and cell-specific targeting of siRNA therapeutics remains to be solved completely [21]. To expand the range of applications for siRNA therapeutics, an effective vehicle of much smaller scale should be envisioned and efforts should be directed in that aspect. Efficient vehicles for delivering the siRNA with better transfection efficiency and protecting it from degradation must be developed. siRNA mediated gene silencing holds the potential and the mechanism to unveil the complex functioning of the gene. siRNA can be the key for controlling and preventing chemotaxis in cancer. Early experiments in plants and lower animals showed that this post-transcriptional gene silencing could be effected by the introduction into cells of one of the intermediates in the RNAi pathway. In 2001, it was shown that RNAi could be successfully applied to mammalian cell culture. This proved a giant step towards revolutionizing discovery biology and drug development strategies. More recent studies have expanded the use of RNAi beyond cell culture, showing that gene silencing can be demonstrated in vivo in mammals and that siRNAs can be targeted to specific tissues [22]. RNAi-induced gene silencing is now commonly used by scientists as a tool to characterize the individual biological roles of specific genes and to illuminate their participation in clinically important pathways or mechanisms (e.g. insulin metabolism, the cell cycle, apoptosis etc.) Thermodynamic stability profiles of siRNAs duplexes led to the identification of key specific locations in the molecule where certain nucleotide base pairs provide high degree of probability of ensuring siRNA functionality [21]. To further refine the pool of functional siRNA candidtae, bioinformatic screens can be used to identify unique siRNA sequences that will be specifically target a gene without producing unintended silencing of other genes [20]. Therapeutic Applications of RNAi The development and application of chemical modification patterns to the siRNA molecule that further enhance potency, mRNA target specificity, and in vivo stability provide additional promise for the use of RNAi in the development of therapeutics. Current drug discovery and development of program are fed by fast-paced genome sequencing projects. These projects define the critical sets of genes that delineate normal biological function and lead to ]understanding of how genetic mutations or pathogens interfere with this normal function [19]. However, the drug-development process and more specifically the target validation, is often hampered by the plethora of sequence information, much of which remains to be fully characterized. while structure and function may be predicted from this genomic data, validation of candidate genes as suitable targets requires reliable, practical approaches to performing screens for functional analysis. Therefore, even with complete sequence information in hand, characterization of individual genes can be an involved and daunting task [20]. The serendipitous discovery of RNAi could not have been more opportune for the pharmaceutical industry, as the rapid output of functional information made possible by RNAi-based strategies alleviates the bottleneck of target validation [21]. Some recent high-throughput analytical approaches include combining siRNA-mediated gene silencing with sophisticated microarray assays, complex cell-based assays and comprehensive bioinformatics [22]. The widespread use of RNAi therapeutics for disease prevention and treatment requires the development of clinically suitable, safe and effective drug delivery vehicle [21]. Unmodified siRNA can be potent triggers of innate immune response. This represents a significant barrier to the therapeutic development of siRNA due to toxicity and off-target gene effects associated with this inflammatory response. Non-inflammatory siRNA, containing less than 20% modified nucleotides can be readily generated without disrupting their gene-silencing activity. Therapeutic application of the recently discovered small-interfering RNA (siRNA) gene silencing phenomenon is dependent on improvements in molecule bio-stability, specificity and delivery. Great care should be taken when considering length variations of dsRNA molecules for RNAi experimentation, especially in therapeutic applications, as induction of the interferon response is cell-type and duplex-length dependent [23]. RNAi can block a pathophysiological pain responseand provide relief from neuropathic pain by down-regulating an endogenous, neuronally expressed gene. If properly designed, low dosages of inhaled siRNA might offer a fast, potent and easily administrable antiviral regimen against respiratory viral diseases in humans [24]. Synthesis siRNAs formulated in non-viral delivery vehicles can be potent inducers of interferons and inflammatory cytokines [25]. siRNAs are attractive candidates for the active component of a microbicide designed to prevent viral infection or transmission. RNAi-mediated gene silencing can be employed in order to overcome the apoptosis resistance of cancer cells [26]. An adenoviral vector harbouring a tandem-type siRNA expression unit was proved to be a modality having a promising tool for cancer therapy. Sever acute respiratory syndrome (SARS) viral infection treatment using short interfering RNA inhibitors exemplifies a powerful new means to combat emerging infectious diseases [27, 28]. Delivery of small RNAs There are several types of vectors developed for efficient transport of small RNAs into the target cell. A chitosan-based siRNA nanoparticle delivery system for RNA interference in vitro and in vivo has potential applications in systemic and mucosal diseases [28]. Several DNA-based plasmid vectors have been developed that direct trascription of small hairpin RNAs, which are processes into functional siRNAs by cellular enzymes. These vectors have certain advantages over chemically synthesized siRNA [36]. Retrovirus-delivered siRNA provides significant advancement over previously available methods by providing efficient, uniform delivery and immediate selection of stable "knock-down" cells [29]. A prospective application of siRNA expressing a vector of recombinant adenovirus system was also found to be efficient [30]. The improvement of non-viral based gene delivery systems is of prime importance for the future of gene and anti-sense therapies [31]. Self-assembling nanoparticles with siRNA were constructed as means to target tumour neovasculature expressing integrins and used to deliver siRNA inhibiting tumour angiogenesis [8]. Efficient delivery of siRNA by single-walled carbon nanotubes achieved more functionality than conventional transfection agents, having applications in gene and protein therapy [32]. Conjugate MPP-siRNAs efficiently reduced transient and stable expressions of reporter genes and demonstrated better delivery characteristics [33]. Compartmental modelling is used to show that the primary advantage of targeted siRNA nanoparticles is associated with processes involved in cellular uptake in tumour cells [34]. Development of aptamer-siRNA chimeric RNAs capable of cell-type-specific binding and delivery of functional siRNAs into cells had proved to be very efficient [35]. Downloads | Glossary | References | Contact © 2012, Saie Mogre. All Rights Reserved. |
