Meth0xyethyl Chimeras Secondgeneration Antisense Drugs

The 2'-0-(2-methoxyethyl) (Fig. 5.11) substitution represents a significant advance in an-

Figure 5.10. PNA peptide nucleic acid without (A) and with Lys end (B).

tisense therapeutics and the culmination of more than a decade of progress in antisense technology. Although oligonucleotides in which every nucleotide contains a 2'-0-(2-methoxyethyl) modification have proved of great value when used for occupancy-only-mediated mechanisms (e.g., induction of alternative splicing; for review, see Ref. 361). They

Figure 5.10. PNA peptide nucleic acid without (A) and with Lys end (B).

thoxyethyl) RNA.

thoxyethyl) RNA.

have been used most broadly in chimeric structures designed to serve as substrates for RNase H. Numerous chimeric 2'-0-(2-me-thoxyethyl) oligonucleotides have not been studied extensively. Although such oligonu-cleotides typically display affinity for target RNA that is several orders of magnitude greater than that for phosphorothioate oli-godeoxynucleotides, they are less attractive substrates for RNase H (for review, see ref. 362). Consequently, they typically display only 5- to 15-fold increases in potency both in vitro and in vivo. For example, a direct comparison of two antisense inhibitors of C-raf kinase with the same sequence, one of which is a phosphorothioate oligodeoxynucleotide, the other a 2'-O-(2-methoxyethyl) chimera, showed that in vitro the 2'-0-(2-methoxyethyl) chimera was approximately fivefold more potent. Similarly, after i.v. administration in a rat heart allograph model, the modified oligonucleotide was at least fivefold more potent (363). Similar data were reported from a direct comparison of antisense inhibitors of survivin both in vitro and in vivo in a human tumor xenograft model (364). Pharmacological studies of antisense oligomers modified with 2'-0-(2-methoxyethyl) chemistry have recently been reviewed (365).

Perhaps more important, 2'-0-(2-methoxy-ethyl) chimeras are substantially more stable than phosphorothioate oligonucleotides. In mice, rats, and monkeys, the elimination halflife is nearly 30 days in plasma and several tissues (314, 366-368). Furthermore, in the liver, elegant correlations between pharmacokinetic and pharmacodynamic effects have been reported to show a duration of pharmacological action of nearly 20 days in the mouse (369).

Studies in animals have recently been extended to humans. ISIS 104838, a TNFa antisense inhibitor, a 2'-0-(2-methoxyethyl) chimera, was shown to reduce TNFa secretion at doses at least 10 times lower than would be expected in humans by first-generation antisense drugs (Dorr, unpublished observations, 2001) and to have an elimination half-life in plasma of approximately 30 days (Geary, unpublished observations, 2001). These studies demonstrate the feasibility of once-a-month dosing.

Although significant progress in achieving acceptable oral bioavailability of antisense inhibitors has been reported, much remains to be accomplished and key clinical studies in progress will help determine how feasible oral delivery is (for review, see Ref. 215). Based on the progress, two key barriers to oral bioavail-ability have been identified and partially overcome: presystemic metabolism and penetration across the gastrointestinal (GI) mucosa.

Because the gut has a very high level of nucleases contributed by both the host and bacteria resident in the GI tract, metabolism of phosphorothioate oligodeoxynucleotides in the gut occurs much too rapidly to support adequate oral bioavailability (370). Chimeric modifications of the oligonucleotides have been shown to enhance stability and oral bioavailability (328, 371). With the development of2'-0-MOE phosphorothioates, oral bioavailability is potentially feasible (215).

In rodents, dogs, and monkeys, the permeability of the GI tract to antisense inhibitors has been significantly enhanced by using formulations containing penetration enhancers such as bile acid salts and fatty acids. After intrajejunal administration of several 2'-0-MOE-modified oligonucleotides in the presence of penetration enhancer-containing formulations, systemic bioavailability in excess of 20% was observed in all three species (215). Initial studies with solid dose forms containing penetration enhancers resulted in significantly less systemic bioavailability (5.5%); clearly therefore more progress is required before solid dose forms with adequate (10-20%) oral bioavailability are available (215, 366, 367). Initial clinical trials in which a variety of solid dose forms containing ISIS 104803, a 2'-O-MOE chimeric antisense inhibitor of TNFa, and various penetration enhancers are in progress, so within the next few years we should have some sense of the near-term potential for oral delivery of antisense inhibitors in humans.

Finally, 2'-0-(2-methoxyethyl) chimeras have been shown to be less proinflammatory (169, 372). After repeated subcutaneous dosing in humans, ISIS 104838 produced dramatically lower local inflammation than that of first-generation antisense drugs. Thus, more convenient and better locally tolerated subcutaneous administration may be feasible.

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  • poppy
    Are phosphorothioate oligonucleotides better than chimera nucleotides?
    8 years ago

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