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RNA interference: a new therapy for neuropathic pain?

Published online by Cambridge University Press:  01 July 2008

B. P. Sweeney*
Affiliation:
Department Anaesthesia, Poole and Royal Bournemouth Hospitals, Bournemouth, UK
M. Z. Michel
Affiliation:
Department Anaesthesia, Poole and Royal Bournemouth Hospitals, Bournemouth, UK
*
Correspondence to: Brian Peter Sweeney, Department of Anaesthesia, Poole and Royal Bournemouth Hospitals, Royal Bournemouth Hospital, Castle Lane East, Bournemouth, Dorset BH7 7DW, UK. E-mail: [email protected]; Tel: +1 202 442443; Fax: +1 202 704196/442672

Abstract

Type
Editorial
Copyright
Copyright © European Society of Anaesthesiology 2008

In 2006, Andrew Fire and Craig Mello were awarded the Nobel Prize for medicine for their pioneering work in developing techniques that allow a better understanding of gene function. Using segments of double-stranded RNA (dsRNA) injected into cells, they were able to show that the messenger RNA (mRNA) of a specific target gene could be effectively blocked or switched off. Using the tiny nematode Caenorhabditis elegans, they found that they could block the expression of specific genes whose nucleotide sequence was complementary to the sequence of the injected RNA [Reference Fire1]. This revolutionary method of gene silencing is now referred to as RNA interference (RNAi) [Reference Elbashir, Harborth and Lendeckel2].

The injected dsRNA molecule is broken down by the elegantly named enzyme, Dicer ribonuclease into a number of shorter sequences, 21–23 nucleotides in length. Each oligonucleotide, now called a ‘short interfering RNA’ (siRNA), is taken up into a protein complex where it is unwound and cleaved by means of an RNA helicase enzyme. The single-stranded siRNA now base pairs or hybridizes to the target mRNA, which it inactivates. Longer sections of dsRNA, although able to mediate gene silencing, can also trigger in mammalian cells an unwanted interferon response, which results in the widespread shut-down of protein synthesis. By designing short sequences of dsRNA against specific genes, researchers can exploit this novel concept and use siRNA as a genetic tool for inhibiting gene expression on a gene-by-gene basis without triggering unpredictable, off-target side-effects [Reference Elbashir, Harborth and Weber3]. Using siRNA to disrupt genes that are specifically involved in disease represents a promising new method both to learn more about the role of the gene in the disease process and to quickly screen potential drug targets – a costly and time consuming aspect of drug development [Reference Ruvkun4]. Furthermore, using siRNA therapeutically has become a distinct possibility and may help in the treatment of some intractable conditions where conventional therapy is lacking or remains ineffective [Reference Ryther, Flynt and Phillips5]. An important example of such a condition is neuropathic pain.

Neuropathic pain is a debilitating, complex, chronic painful state that is usually accompanied by tissue injury and in particular, injury to the nervous system itself. The aetiology includes diverse conditions such as post-herpetic neuralgia, trigeminal neuralgia, diabetic neuropathy, spinal cord injury, cancer, stroke and degenerative neurological diseases. The pathophysiology of neuropathic pain is complex involving several pathways and receptors. Of pivotal importance, however, is the role of the N-methyl-d-aspartate (NMDA) receptor and its activation in the posterior horn of the spinal cord [Reference Hudspith, Siddall and Munglani6]. Prolonged or repeated activation of this receptor results in a cascade of neurochemical events, which manifest themselves as hyperalgaesia, allodynia and an increase in transmission for a given afferent input, which is referred to as ‘wind-up’. In addition, and perhaps more significantly, there are thought to be changes in the genetic machinery leading to alteration in synaptic morphology. Recent work has suggested that there are electrophysiological changes in the NMDA receptor in peripheral nerve-injured neuropathic mice together with changes in the NMDA receptor subunits, which may underlie the hyperalgaesia [Reference Iwata, Takasusuki and Yamaguchi7].

Currently, the outcome of treatment of neuropathic pain is disappointing. Although there are a number of pharmacotherapeutic options, which include tricyclic antidepressants, antiepileptics and weak NMDA-receptor antagonists such as dextromethorphan and ketamine, the overall results from therapy provide patients with poor relief from symptoms, which may be both unrelenting and debilitating. In an attempt to evaluate more precisely the role of the NMDA receptor, researchers examined in vivo the effect of experimentally blocking the receptor using specific NMDA-receptor antagonists. They found that such a strategy was highly effective in increasing the efficacy of opiates in the experimental animal [Reference Fisher, Carrigan and Dykstra8]. Furthermore, in animals there is clear evidence that when the NMDA gene was blocked by silencing the NR1 subunit, there was significantly less pain compared with controls [Reference Quintero, Erzurumlu and Vaccarino9] and that there was markedly reduced associated neuropathic features such as hyperalgaesia and morphine tolerance [Reference Shimoyama, Shimoyama and Davis10].

The introduction of novel NMDA antagonists has unfortunately been limited until now by their unacceptable side-effect profile [Reference Fisher, Coderre and Hagen11]. However, one new and exciting therapeutic option that will shortly be available is the targeting of the NMDA receptor using siRNA. Recently Tan and colleagues [Reference Tan, Yang and Shih12] targeted the gene, which encrypts the NR2 subunit of the NMDA receptor with specific siRNA injected intrathecally in mice. The result was potent, long-lasting silencing of the gene with abolition of formalin-induced pain behaviour. Similar results were obtained by Garaway [Reference Garraway, Xu and Inturrisi13] who demonstrated, also in mice, that injection of siRNA into the spinal cord dorsal horn, aimed at the NR1 subunit of the NMDA receptor, effectively achieved 75% knockdown of the receptor, which lasted for up to 6 months. This was accompanied by alleviation of symptoms of experimentally induced allodynia [Reference Garraway, Xu and Inturrisi13].

At present, the focus of attention has mainly been on the NMDA receptor as a conduit for the transmission of neuropathic pain. One other family of proteins currently under investigation are the transient receptor potential (TRP) group, which are expressed in the nervous system and which is involved in sensory physiology. One member of this family is the vanilloid type 1 receptor, which is encrypted by the TRPV1 gene. Using similar strategies of injecting siRNA intrathecally, Christoph and colleagues [Reference Christoph, Grünweller and Mika14] were similarly able to demonstrate significant reductions in the levels of allodynia in a neuropathic rat model. Viscerally induced pain levels were also attenuated [Reference Christoph, Grünweller and Mika14].

Only a few years after their discovery, RNAi has become an invaluable tool for researchers working in the field of functional genomics and target validation and has become the gold standard for studying loss of function phenotype, i.e. gene knockout models. This exciting new technology will shortly be within the reach of clinicians and already the number of possible applications, apart from the treatment of chronic pain syndromes, is rapidly growing. The potential therapeutic applications include the treatment of a variety of virus-induced diseases such as HIV, hepatitis, polio and influenza [Reference Lieberman, Song and Lee15,Reference Saulnier, Pelletier and Labadie16]. siRNA can also specifically inhibit oncogenes associated with leukaemia and has been found to have other potential roles in the treatment of neurodegenerative disease such as amyotrophic lateral sclerosis [Reference Koutsilieri, Rethwilm and Scheller17]. At present considerable effort is being made to develop appropriate vehicles to optimize the delivery and stability for siRNA. Both lipid-based delivery systems as well as vectors such as adenoviruses, retroviruses and plasmids have been examined. One promising avenue is the complexing of siRNA with polyethyleneimine (PEI), which both stabilizes the siRNA and releases it intact when injected systemically [Reference Aigner18].

The discovery of RNAi has opened a window on the control mechanisms of intracellular protein production. Already pharmaceutical companies have begun to formulate strategies for harnessing this valuable therapeutic tool; indeed the first siRNA therapeutic substance Bevasiranib Sodium (Acuity Pharmaceuticals Inc., Philadelphia, PA, USA), intended for use in a common form of macular degeneration, has already successfully passed phase 2 trials. Despite the recent introduction of new classes of pain killers such as the N-type calcium channel blockers pregabalin and ziconotide, clinicians continue to be frustrated by neuropathic pain in their attempts to provide effective and reliable therapy. It is hoped that for the many patients whose lives are blighted by this unfortunate debilitating condition, siRNA will provide the much longed-for panacea.

References

1.Fire, AZ. Gene silencing by double stranded RNA. Cell Death Differ 2007; 14: 19982012.CrossRefGoogle ScholarPubMed
2.Elbashir, SM, Harborth, J, Lendeckel, W et al. Duplexes of 21-nucleotide RNAs mediate RNA interference in cultured mammalian cells. Nature 2001; 411: 494498.CrossRefGoogle ScholarPubMed
3.Elbashir, SM, Harborth, J, Weber, K et al. Analysis of gene function in somatic mammalian cells using small interfering RNAs. Methods 2002; 26: 199213.CrossRefGoogle ScholarPubMed
4.Ruvkun, G. Molecular biology. Glimpses of a tiny RNA world. Science 2001; 294: 797799.CrossRefGoogle ScholarPubMed
5.Ryther, RC, Flynt, AS, Phillips, JA. siRNA therapeutics: big potential from small RNAs. Gene Ther 2005; 12: 511.CrossRefGoogle ScholarPubMed
6.Hudspith, MJ, Siddall, PJ, Munglani, R. Physiology of pain. In Foundations of Anaesthesia, Chapter 23. Philadelphia: Mosby Elsevier, 2006: 267286.CrossRefGoogle Scholar
7.Iwata, H, Takasusuki, T, Yamaguchi, S et al. NMDA receptor 2B subunit-mediated synaptic transmission in the superficial dorsal horn of peripheral nerve-injured neuropathic mice. Brain Res 2007; 1135: 92101.CrossRefGoogle ScholarPubMed
8.Fisher, BD, Carrigan, KA, Dykstra, LA. Effects of NMDA receptor antagonists on acute morphine-induced and l-methadone-induced antinociception in mice. J Pain 2005; 6: 425433.CrossRefGoogle Scholar
9.Quintero, GC, Erzurumlu, RS, Vaccarino, AL. Decreased pain response following cortex-specific knockout of the N-methyl-D aspartate NR1 subunit. Neurosci Lett 2007; 425: 8993.CrossRefGoogle ScholarPubMed
10.Shimoyama, N, Shimoyama, M, Davis, AM et al. An antisense oligonucleotide to the N-methyl-D-aspartate (NMDA) subunit NMDAR1 attenuates NMDA-induced nociception, hyperalgesia, and morphine tolerance. J Pharmacol Exp Ther 2005; 312: 834840.CrossRefGoogle Scholar
11.Fisher, K, Coderre, TJ, Hagen, NA. Targeting the NMDA receptor for chronic pain management. Preclinical animal studies, recent clinical experience and future research directions. J Pain Symptom Manage 2000; 20: 358373.CrossRefGoogle ScholarPubMed
12.Tan, PH, Yang, LC, Shih, HC. Gene knockdown with intrathecal siRNA of NMDA nociceptor NR2B subunit reduces formalin-induced nociception in the rat. Gene Ther 2005; 12: 5966.Google Scholar
13.Garraway, SM, Xu, Q, Inturrisi, CE. Design and evaluation of small interfering RNAs that target expression of the N-methyl-D-aspartate receptor NR1 subunit gene in the spinal cord dorsal horn. J Pharmacol Exp Ther 2007; 322: 982988.CrossRefGoogle ScholarPubMed
14.Christoph, T, Grünweller, A, Mika, J. Silencing of vanilloid receptor TRPV1 by RNAi reduces neuropathic and visceral pain in vivo. Biochem Biophys Res Commun 2006; 350: 238243.CrossRefGoogle ScholarPubMed
15.Lieberman, J, Song, E, Lee, SK et al. Interfering with disease: opportunities and roadblocks to harnessing RNA interference. Trends Mol Med 2003; 9: 397403.CrossRefGoogle ScholarPubMed
16.Saulnier, A, Pelletier, I, Labadie, K. Complete cure of persistent virus infections by antiviral siRNAs. Mol Ther 2006; 13: 142150.CrossRefGoogle ScholarPubMed
17.Koutsilieri, E, Rethwilm, A, Scheller, C. The therapeutic potential of siRNA in gene therapy of neurodegenerative disorders. J Neural Transm Suppl 2007; 72: 4349.CrossRefGoogle Scholar
18.Aigner, A. Delivery systems for the direct application of siRNA to induce RNA interference (RNAi) in vivo. J Biomed Biotechnol 2006; 2006: 461466.CrossRefGoogle ScholarPubMed