Technology

Antisense oligonucleotides (ASO) are short polymers of linked nucleotides (combinations of one of the nucleobases adenine, guanine, cytosine or guanine, with a ribose sugar and a phosphate group) that are complementary in sequence to (the mirror image of) a messenger ribonucleic acid (mRNA) of interest (Figure 1). mRNAs are long stretches of nucleotides that carry the instructions for the synthesis of proteins. ASO recognize and bind to mRNA following the principles of Watson-Crick hybridization first described for the formation of the double-stranded deoxyribonucleic acid (DNA) helix and essential for in situ hybridization and diagnostic probe technologies. Upon binding mRNA, ASO mediate the destruction of the mRNA or inactivate its function(s). Therefore, the recognition of mRNA by ASO is nucleotide sequence-dependent, producing a specific, rational method for altering the production of disease-related proteins and/or changing biological processes within cells

There are many ways that ASO can achieve the desired effects in cells and in tissues. However, the only marketed ASO product (Vitravene, CibaVision, Isis Pharmaceuticals/Novartis) and the most clinically advanced ASO compounds cleave mRNA by invoking the activation of a ubiquitously present enzyme, RNase H. Altair’s ASO programs exploit the RNase H mechanism to block the production of proteins encoded by the targeted mRNA.

Mechanism of Action of Antisense Oligonucleotides
Figure 1 Mechanism of Action of Antisense Oligonucleotides
Antisense oligonucleotides selectively inhibit the synthesis of proteins by hybridizing with a target mRNA
and preventing translation to protein.

Altair has licensed a proprietary chemical modification of ASO from Isis Pharmaceuticals, based on over 15 years of medicinal chemistry technology advances. Addition of a methoxyethyl (MOE) group to the 2’-position on the ribose sugar ring in combination with replacement of a non-bridging oxygen atom in the phosphate group with sulfur (phosphorothioate backbone modification) yields an ASO (Figure 2) with greatly improved potency, stability, safety and tolerability in vivo. Because the 2’-MOE modification does not allow recruitment of RNase H when bound to mRNA, a hybrid ASO molecular design has been adopted (termed “MOE gapmer”) in which the terminal residues of the ASO sequence are 2’-MOE-modified, leaving the central region with only the phosphorothioate backbone modification (Figure 3). This design couples the resistance to degradation in plasma and tissues with improved potency and safety while supporting the activation of RNase H.

Chemical Structure First-Generation
Figure 2 Chemical Structure of First-Generation (Phosphorothioate Oligodeoxynucleotide) and Second-Generation [Phosphorothioate 2′-O-(2-Methoxyethyl) Oligonucleotides (2′-MOE)].

Design of Chimeric
Figure 3 Design of Chimeric 2′-MOE Phosphorothioate Oligonucleotide (MOE-Gapmer). N can be any nucleotide (A, G, C or T).

Published References

Altmann, K.H., Dean, N.M., Fabbro, D., Freier, S.M., Geiger, T., Haner, R., Husken, D., Martin, P., Monia, B.P., Muller, M., Natt, F., Nicklin, P., Phillips, J., Pieles, U., Sasmor, H. and Moser, H.E. Second-generation of antisense oligonucleotides: from nuclease resistance to biological efficacy in animals. Chimia 1996; 50: 168-176.

Bennett, C.F. Pharmacological properties of 2’-O-methoxyethyl-modified oligonucleotides, in Antisense Drug Technology: Principles, Strategies, and Applications, Second Edition, Crooke, S.T., ed., Marcel Dekker, Inc. New York, 2007.

Capaldi, D.C., and Scozzari, A.N. Manufacturing and analytical processes for 2’-O-methoxyethyl-modified oligonucleotides, in in Antisense Drug Technology: Principles, Strategies, and Applications, Second Edition, Crooke, S.T., ed., Marcel Dekker, Inc. New York, 2007.

Chi, K.N., Eisenhauer, E., Fazli, L., Jones, E.C., Goldenberg, S.L., Powers, J., Tu, D. and Gleave, M.E. A phase I pharmacokinetic and pharmacodynamic study of OGX-011, a 2’-methoxyethyl antisense oligonucleotide to clusterin, in patients with localized prostate cancer. J Natl Cancer Inst 2005; 97: 1287-96.

Geary, R.S. Yu, R.Z., Siwkowski, A. and Levin, A.A. Pharmacokinetic/pharmacodynamic properties of phosphorothioate 2’-O-methoxyethyl-modified antisense oligonucleotides in animals and man, in Antisense Drug Technology: Principles, Strategies, and Applications, Second Edition, Crooke, S.T., ed., Marcel Dekker, Inc. New York, 2007.

Henry, S.P., Geary, R.S., Yu, R.Z., Templin, M.V., Pallman, J. and Levin, A.A. Comparison of toxicity and tissue concentrations for 1st and 2nd generation antisense oligonucleotides. The Toxicologist 2001; 60: 325-34.

Henry, S.P., Kim T-W., Kramer-Stickland, K., Zanardi, T.A., Fey, R.A. and Levin, A.A. Toxicologic properties of 2’-O-methoxyethyl chimeric antisense inhibitors in animals and man, in Antisense Drug Technology: Principles, Strategies, and Applications, Second Edition, Crooke, S.T., ed., Marcel Dekker, Inc. New York, 2007.

Kastelein, J.J.P., Wedel, M.K., Baker, B.F., Su, J., Bradley, J.D., Yu, R.Z., Chuang, E., Graham, M.J. and Crooke, R.M. Potent reduction of apolipoprotein B and low-density lipoprotein cholesterol by short-term administration of an antisense inhibitor of apolipoprotein B. Circulation 2006; 114: 1729-35.

Kwoh, T.J. An overview of the clinical safety experience of first- and second-generation antisense oligonucleotides, in Antisense Drug Technology: Principles, Strategies, and Applications, Second Edition, Crooke, S.T., ed., Marcel Dekker, Inc. New York, 2007.

Monia, B.P., Lesnik, E.A., Gonzalez, C., Lima, W.F., McGee, D., Guinosso, C.J., Kawasaki, A.M., Cook, P.D. and Freier, S.M. Evaluation of 2'-modified oligonucleotides containing 2'-deoxy gaps as antisense inhibitors of gene expression. J Biol Chem 1993; 268(19): 14514-22.