Synthesis of Flavone-DNA Conjugates Using Click Chemistry
Poster Number
23B
Faculty Mentor Name
Liang Xue
Format
Poster Presentation
Research or Creativity Area
Natural Sciences
Abstract
The DNA double helix can be bound by sequence-specific triplex-forming oligonucleotides (TFOs) to form a DNA triplex. These TFOs can be synthesized to be complementary to potential sequences of interest in duplex DNA, where, once bound to form the triplex, they will regulate the transcription of the target sequence and the subsequent production of its encoded proteins. TFOs’ high specificity, as well as the limited off-target effects they produce, make them a promising anti-gene therapeutic strategy. However, an inherent instability under physiological conditions limits triplex formation. Recent studies from our laboratory have shown that quercetin, a flavonoid found in many plants, and several of its derivatives stabilize the DNA triplex while leaving duplex DNA unaffected. To further advance this approach, we are now interested in improving the delivery efficiency of both TFOs and ligands to their target sequences. To this end, a covalent conjugate between the TFO and ligand is an avenue we are currently pursuing. This conjugate has been synthesized using copper-catalyzed click chemistry, which required an alkyne-modified TFO and an azide handle on our ligand derived from 5-hydroxyflavone. Synthesis and purification of the former can be an expensive and delicate affair; as such, testing highly efficient purification methods, such as gel purification, on standard synthesized oligonucleotides was conducted first to establish robust methods for the synthesis of the modified TFO. Initial trials in the synthesis have been successful, and we are now optimizing our protocol to achieve sufficient concentrations for future biophysical studies.
Purpose
The purpose of this research is to improve the delivery efficiency of triplex-forming oligonucleotides and small molecules to DNA sequences of interest. To do so, a covalent conjugate between the oligonucleotide and small molecule is synthesized using copper-catalyzed click chemistry, which will then be tested for stabilizing effects when bound to duplex DNA.
Results
Synthesis of azide-containing 5-hydroxyflavone derivatives and alkyne-modified oligonucleotides has proven successful and readily scalable to sufficient quantities for our click protocol. The covalent conjugate has also been synthesized at this time with sufficient time and heating, and optimizations to the protocol are currently underway.
Significance
Triplex-forming oligonucleotides are a promising anti-gene therapy, as they can regulate transcription of a target DNA sequence by forming a TFO strand complementary to it, producing a therapy with high specificity and limited side effects. This therapy is hindered by inherent instability under physiological conditions, a drawback that can be mitigated by adding small molecules that bind to and stabilize the structure. By conjugating our TFO to the small molecule, we can increase the efficiency of its delivery to a target sequence while ideally maintaining the small molecule’s stabilizing effect.
Location
University of the Pacific, DeRosa University Center
Start Date
24-4-2026 11:00 AM
End Date
24-4-2026 2:00 PM
Synthesis of Flavone-DNA Conjugates Using Click Chemistry
University of the Pacific, DeRosa University Center
The DNA double helix can be bound by sequence-specific triplex-forming oligonucleotides (TFOs) to form a DNA triplex. These TFOs can be synthesized to be complementary to potential sequences of interest in duplex DNA, where, once bound to form the triplex, they will regulate the transcription of the target sequence and the subsequent production of its encoded proteins. TFOs’ high specificity, as well as the limited off-target effects they produce, make them a promising anti-gene therapeutic strategy. However, an inherent instability under physiological conditions limits triplex formation. Recent studies from our laboratory have shown that quercetin, a flavonoid found in many plants, and several of its derivatives stabilize the DNA triplex while leaving duplex DNA unaffected. To further advance this approach, we are now interested in improving the delivery efficiency of both TFOs and ligands to their target sequences. To this end, a covalent conjugate between the TFO and ligand is an avenue we are currently pursuing. This conjugate has been synthesized using copper-catalyzed click chemistry, which required an alkyne-modified TFO and an azide handle on our ligand derived from 5-hydroxyflavone. Synthesis and purification of the former can be an expensive and delicate affair; as such, testing highly efficient purification methods, such as gel purification, on standard synthesized oligonucleotides was conducted first to establish robust methods for the synthesis of the modified TFO. Initial trials in the synthesis have been successful, and we are now optimizing our protocol to achieve sufficient concentrations for future biophysical studies.
