Nicole Wong's SOD1 aptamer project (2014)
Nucleic Acid Aptamer Selection Against Superoxide Dismutase-1 for Amyotrophic Lateral Sclerosis Drug Delivery
An aggressive neurodegenerative disease with few effective treatment options, amyotrophic lateral sclerosis (ALS) is characterized by gradual motor neuron breakdown, muscle atrophy, and eventually, muscle paralysis. Several studies have demonstrated evidence that the characteristic motor neuron degeneration may be caused by an aggregation of mutated forms of Cu, Zn-superoxide dismutase 1 (SOD1). This accumulation has been implicated in creating a toxic environment for motor neurons, contributing to their degeneration, decreased neural stimulation, and resulting muscle atrophy.
Localized delivery of drugs offers many therapeutic possibilities and can be achieved through a nucleic aptamer selection against superoxide dismutase-1. Aptamers are oligonucleotides that tightly bind their targets; since most mutated forms of SOD1 differ from the wild-type SOD1 by only one amino acid, an aptamer capable of tightly binding to the wild-type SOD1 enzyme could potentially bind to most of the mutated forms as well. Then, the aptamer could be functionalized to deliver drugs directly to the aggregations of superoxide dismutase 1. The advantage of targeted disease therapy is that it prevents unnecessary alterations of other proteins, thereby minimizing the treatment’s unwarranted side effects.
The aptamer for SOD1 would be isolated through a process of in vitro bead-based selection in which SOD1 would be immobilized on nickel beads, incubated with a pool of RNA sequences, and washed with Tris buffer to remove the unbound species. After many rounds of selection, the target’s most tightly binding aptamer would remain. The aptamer could then be loaded with an inhibitory drug molecule similar to a transthyretin ligand that could stabilize the superoxide dismutase 1 enzymes so that they do not aggregate and create a toxic environment for motor neurons (Baures, Peterson, and Kelly, 1998). With a reduction in the toxicity, the main symptoms of amyotrophic lateral sclerosis, initially caused by motor neuron degeneration, could be alleviated. Identification of an aptamer capable of accurately delivering drugs to SOD1 enzymes would significantly further our progress in generating treatment options for ALS and the 30,000 people affected by its paralyzing effects.
Figure 1. Schematic of an aptamer functionalized for targeted disease therapy (Zhou and Rossi, 2014).
Figure 2. X-ray crystal structure of superoxide dismutase 1 (Parge, Hallewell, and Tainer, 1992).
Budget
● Human superoxide dismutase protein (His Tag) from Sino Biological Inc.
● Available in -80° freezer in Painter 2.14
● Catalog Number is 11727-H07E-50.
● Sino Biological Inc.’s telephone number is +86-400-890-9989.
● Sino Biological Inc.’s website is http://www.sinobiological.com/SOD1-Protein-g-3370.html
● Molecular Weight: 16,800 g/mol
● Costs $138 for 50 µg
● Costs $9.85 per round of selection (using 200 pmol/round) [2]
References
1. "Diagnosing ALS." The ALS Association. N.p., n.d. Web. 17 Sept. 2014.
2. "Human SOD1 / Superoxide Dismutase Protein (His Tag)." Sino Biological Inc.. Sino Biological Inc., n.d. Web. 2 Sept. 2014.
3. "Protein Calculator v3.4." Protein Calculator. N.p., n.d. Web. 17 Sept. 2014.
4. "What is ALS?" The ALS Association. N.p., n.d. Web. 17 Sept. 2014. <http://www.als-ny.org/index.php?page=about_als&sub=what_is>.
5. Baures, P.W., Peterson, S.A., Kelly, J.W. (1998). Discovering transthyretin amyloid fibril inhibitors by limited screening. Science Direct. 6 (8), 1389–1401.
6. Broering, T.J., Wang, H., Boatright, N.K., Wang, Y., Baptista, K., Shayan, G., Garrity, K.A., Kayatekin, C., Bosco, D.A., Matthews, R., et.al. (2013). Identification of Human Monoclonal Antibodies Specific for Human SOD1 Recognizing Distinct Epitopes and Forms of SOD1. PlosOne.
7. Desmarais, W.T., Bienvenue, D.L., Bzymek, K.P., Holz, R.C., Petsko, G.A., Ringe, D. (2002).The 1.20 Å Resolution Crystal Structure of the Aminopeptidase from Aeromonas proteolytica Complexed with Tris: A Tale of Buffer Inhibition. Science Direct. 10 (8), 1063–1072.
8. Fessenden, J., "Gene connection extends possible treatments to more ALS patients." UMass Medical School. University of Massachusetts Medical School, n.d. Web. 2 Sept. 2014.
9. Goodsell, D. (2007). Superoxide Dismutase. RCSB Protein Data Bank. 10.
10. Kretschmer, B.D., Kratzer, U., Schmidt, W.J. (1998). Riluzole, a glutamate release inhibitor, and motor behavior. PubMed. 358 (2), 181-190.
11. Rowland, L.P., Shneider, N.A. (2001). Amytrophic Lateral Sclerosis. The New England Journal of Medicine. 344, 1688-1700.
12. Zhou, J., Rossi, J.J. (2014). Cell-type-specific, Aptamer-functionalized Agents for Targeted Disease Therapy. PubMed. 17 (3), 169.
13. Overview Figure 1. Schematic of an aptamer functionalized for targeted disease therapy. Adapted from “Cell-type-specific, Aptamer-functionalized Agents for Targeted Disease Therapy” by Zhou, J. and Rossi, J.J., 2014, Molecular Therapy-Nucleic Acids, 3, e169.
14. Overview Figure 2. X-ray crystal structure of superoxide dismutase 1. Adapted from “Atomic Structures of wild-type and thermostable mutant recombinant human Cu, Zn Superoxide Dismutase” by Parge, H.E., Hallewell, R.A., and Tainer, J.A., 1992, RCSB Protein Data Bank, 89: 6109-6113.
15. Figure 1. Motor neuron degeneration leading to muscle atrophy. Adapted from “What is ALS?” by The ALS Association, 2014.
16. Figure 2. Superoxide dismutase active site. Adapted from “Superoxide dismutase” by the Goodsell, D., 2007, RCSB Protein Data Bank.
17. Figure 3. Amino acid sequence of human SOD1. Adapted from “Superoxide dismutase [Cu-Zn] [Homo sapiens]” by the National Center for Biotechnology, 2014. U.S. National Library of Medicine.