Uses advantages and disadvantages of oligos

Antisense oligonucleotides (oligos) are being assessed in preclinical and clinical studies as therapeutic agents in the treatment of cancer, as well as a variety of viral diseases (e.g. HIV, hepatitis B, herpes and papillomavirus infections). They also have potential application in treating other disease states for which blocking of gene expression would likely have a beneficial effect. Such medical conditions include restenosis, rheumatoid arthritis and allergic disorders. Cancer, however, remains the most common target indication. The favoured approach is to target genes whose expression/up-regulation triggers or fuels tumorigenesis. These include products of the BCL-2, survivin and clusterin genes. The BCL-2 oncogene products drive neoplastic progression by enhancing cell survival via inhibition of apoptosis. Survivin is generally not expressed in healthy tissue, but expressed at high levels in a range of common cancer types, including lung, colon, breast and prostate cancers. It plays an important role in both promoting cell division and inhibiting apoptosis. The clusterin gene codes for a cytoprotective 'chaperone' protein whose up-regulation is associated with various human cancers.

As potential drugs, antisense oligos display a number of desirable characteristics, the most significant of which is their likely specificity. Statistical analysis reveals that any specific base sequence of 17 or more bases is extremely unlikely to occur more than once in a human cell's nucleic acid complement. It thus follows that an oligonucleotide of 17 or more nucleotide units in length, which is designed to duplex successfully with a specific mRNA species, is unlikely to form a duplex with any other (unintended) mRNA species. Most synthetic oligos, therefore, are in the region of 17 nucleotide units long. These will display virtually an absolute specificity for the target sequence. Additional advantages of the oligonucleotide antisense approach include:

• Relatively low toxicity: thus far, most trials report relatively few significant side effects. This is likely due to the highly specific nature of oligo duplexing, and the fact that they are 'natural' biomolecules. Some toxicity may, however, by triggered by non-specific binding to proteins, and most antisense agents appear to promote pro-inflammatory effects at high dosage levels.

• The requirement for only low levels of the oligo to be present inside the cell, as target mRNA is, itself, usually present only in nanomolar concentrations.

• The ability to manufacture oligos of specified nucleotide sequence is relatively straightforward using automated synthesizers.

However, native antisense oligonucleotides also suffer from a number of disadvantages, which are ultimately responsible for numerous disappointing trial results, and the fact that, after almost two decades of clinical investigation, only a single product has gained approval to date. Disadvantages include:

• sensitivity to nucleases;

• very low serum half lives;

• poor rate of cellular uptake;

• orally inactive.

Linkage name

Substituent (R)

BASE

BASE

O v CH2 Phosphorothioate

Linkage

Linkage

Methylphosphonate

Methylphosphotriester

Ethylphosphotriester

-CH3

-O-CH2-CH3

Alkylphosphoramidate

-NH-CH3

Figure 14.15 Major types of modification potentially made to an oligo's phosphodiester linkage in order to increase their stability or enhance some other functional characteristic. The native phosphodiester link is shown to the left

Some progress has been made to overcome such difficulties, and continued progress in the area is expected to render the next generation of oligos more therapeutically effective.

Native oligonucleotides display a 3'-5' phosphodiester linkage in their backbone (Figure 14.15). These are sensitive to a range of nucleases naturally present in most extracellular fluids and intracellular compartments. The half-life of native oligonucleotides in serum is only of the order of 15 min, and oligoribonucleotides are less stable than oligodeoxynucleotides. Selective modification of the native phosphodiester bond can render the product resistant to nuclease degradation.

Modification usually entails replacement of one of the free (non-bridging) oxygen atoms of the phosphodiester linkage with an alternative atom or chemical group (Figure 14.15). Most commonly, the oxygen has been replaced with a sulfur atom and the resultant phosphorothioates display greatest clinical promise. Phosphorothioate-based oligos, 'S-oligos', display increased resistance to nuclease attack while remaining water-soluble. They are also easy to synthesize chemically and they display a biological half-life of several hours. Most antisense oligos currently assessed in clinical trials are S-oligos, as is the sole antisense agent approved for general medical use to date (Vitravene, Box 14.3). Further modified oligos may well improve product pharmacokinetic and pharmacodynamic properties, as alluded to towards the end of the next section.

Was this article helpful?

0 0
Natural Arthritis Relief

Natural Arthritis Relief

Natural Arthritis Relief details a unique method of reversing Rheumatoid Arthritis Symptoms by removing numerous arthritis triggers as well as toxins using a simple 5 step natural process.

Get My Free Ebook


Post a comment