In the early1990s, a new class of molecules called “aptamer” has been developed. Aptamers are single-stranded oligonucleotides (either DNA or RNA) that can bind to target molecules with affinity and specificity comparable to antibodies. Aptamers can be developed using in vitro selection or SELEX techniques (Systematic Evolution of Ligands by EXponential enrichment) as summarized in the diagram below. The techniques are very powerful in allowing the isolation of rare RNA or DNA molecules, or “aptamers”, from a pool of random RNAs or DNAs of more than 1016 different sequences. The selection process starts with a library of RNAs with randomized sequences. The library is generated from ss-DNA containing a randomized region flanked by constant sequences at both 5′ and 3′ ends, which are used as primer binding sites during RT-PCR amplification. In addition, the promoter of T7 RNA polymerase is incorporated into the 5′ end of ss-DNAs. This random ss-DNA molecules are converted into double-stranded DNAs (ds-DNA) by PCR, and the ds-DNAs are subsequently used as templates for in vitro transcription using T7 RNA polymerase to generate a pool of random RNAs. The complexity of the library depends on the number of nucleotides present in the randomized regions (varied from 25-80 nucleotides long). The commonly-used libraries have the complexity of ~ 10^15-10^16 different sequences. The aptamers are partitioned from the noninteracting RNA ligands based on their ability to bind the target ligands. The pool of RNAs recovered after partitioning is used to generate ds-DNA by RT-PCR amplification. The ds-DNA templates are then used for in vitro transcription to produce a new pool of RNA for use in the next round of selection. The selection process is repeated until the aptamers with desired properties are highly enriched. Because the random RNA library with a large complexity is used, it is likely that some of the molecules could fold into the conformations that are able to recognize and bind to the selected target molecules even though they are not natural ligands for RNAs. So far, aptamers to a wide range of targets have been discovered, ranging from simple ions, small molecules, peptides, single proteins, organelles, viruses and even entire cells.

Since the invention of the aptamer technology, aptamers have now become an emerging class of molecules that can rival antibodies in both therapeutic and diagnostic applications. In addition to the comparable affinity and specificity of aptamers and antibodies to target molecules, there are several desirable properties of aptamers over those of antibodies. For example, the production of aptamers is faster and cheaper because the selection processes are done in vitro and no animals involved. Aptamers can also be raised against some antigenic epitopes on the target protein that are important for activity or functions but have poor immunogenicity because the selection of aptamers is not based on the immunological response. In addition, because aptamers are nucleic acids, which are not good immunogens and have a shorter half-life in the body, they could be used more safely as a therapeutic agent compared with antibodies, particular those produced from other species.

We are currently developing RNA aptamers to the dengue virus envelope protein with neutralizing activity against dengue virus infection. The aptamers might be useful for diagnostics or therapeutics of dengue infection. In addition, we are interested in generating RNA aptamers with specificity to various types of hemoglobin in order to use them for diagnosis of hemoglobinopathies or thalassemia, which is one of the most common hematologic diseases in Thailand. In the future, we plan to develop RNA aptamers against target molecules that could be used in diagnostics or therapeutics of the diseases with medical importance in Thailand.

Besides the aptamer project, we are also studying the suppression of collagen production in keloid fibroblasts using a pharmacologic agent, curcumin, and RNA interference against collagen genes, i.e. COL1A1 and COL1A2. The results from this study might be useful for treatment of keloids in the future.