Phototriggered Drug Release from Functionalized Oligonucleotides by a Molecular Beacon Strategy

Abstract

DNA hybridization biosensors offer considerable promise for obtaining sequence information of genes in a fast and simple manner. Various DNA probes that give signals in a sequencespecific fashion, as represented by molecular beacons, have been widely used. [1][2][3] However, there are very few DNA probes that release functional molecules on recognition of the target sequence. [4,5] The sequence-specific molecule-releasing system that is triggered by external stimulation, such as irradiation would be a very powerful tool for gene analysis.Herein we report for the first time a molecule-releasing system controllable by an intramolecular quenching based on a molecular-beacon strategy by using photoactive probe oligodeoxynucleotides (ODNs). The photoreaction of the probe ODN containing a photoactive group and a triplet quencher at both ends of the strand was very inefficient when it was in the closed-form ODN (a stem-and-loop structure), whereas irradiation of the open-form ODN hybridized with the complementary DNA resulted in a rapid release of the functional molecule from the ODN (see Figure 1).ODN sequences for the molecule-releasing system were designed according to the molecular-beacon strategy.[1] Phenacyl ester [6][7][8] as a photocleavable group via a triplet excited state, and substituted naphthalene [9,10] as a triplet quencher were incorporated into the 3'-and 5'-ends of the ODN, respectively. The stem of the closed-form ODN keeps a phenacyl ester and a naphthalene ring in close proximity to each other, causing the efficient triplet quenching, as in the case of a molecular beacon. In contrast, when the probe ODN is transformed to the open form by hybridization with target DNA, [1] photocleavage of the phenacyl ester proceeds very smoothly under irradiation, because the photoactive phenacyl ester is far away from the naphthalene quencher.The synthetic route for the functionalized ODN is shown in Scheme 1. The photocleavable phenacyl ester 3, which has both a maleimide functional group to be connected to thiolmodified DNA and a biotin unit as a leaving functional molecule, was prepared from 4-hydroxyacetophenone (1). Naphthalene phosphoramidite, 6, was prepared from 1-naphthylacetic acid (4) in four steps. ODNs containing both a naphthalene unit at the 5' end and a disulfide unit at the 3' end were synthesized by using a DNA synthesizer by a conventional phosphoramidite method. The disulfide group of the given ODNs 7 was reduced to thiol by dithiothreitol, and then coupled with 3 to give ODNs 9 a and 9 b that were equipped with a photoactive group and a quencher at each end.We initially investigated the efficiency of the photoreaction of ODN 9 a by irradiation at 312 nm before and after hybridization with the complementary target DNA 5'-d(CAT-AGGTCTTAACTT)-3'. The irradiation of a solution of 9 a in 15 mm sodium cacodylate (pH 7.0) and 2-propanol (2:1) was Figure 1. Outline for the phototriggered drug release from functionalized oligonucleotides by a molecular beacon strategy. Scheme 1. Reagents and condition...

Nucleic Acid Templated Reactions for Chemical Biology

“…Upon interaction with aD NA target, the stem opened up, and the para-hydroxyphenacyl ester could be cleaved via at riplet excited state by UV irradiation (312 nm), thus releasing biotin. [28] Tanabe et al improved this strategy by developing as imilar system,b ut with irradiation at 365 nm. [29] Rçthlingshçfer et al [30] reported the templated photocleavage of an itrobenzyl-type linker occurring at 405 nm.…”

Development and Applications of Fluorescent Oligonucleotides

“…A moleculereleasing system controllable by an intramolecular quenching based on a MB strategy by using photoactive probe ONs has been reported [295]. Phenacyl ester as a photocleavable group via a triplet excited state, and substituted naphthalene as a triplet Q was incorporated into the 3'-and the 5'-ends of the ON, respectively.…”

Synthesis of Nitrogen-containing 4-alkoxybenzaldehyde Analogues

“…Indeed, our research group had spent much time synthesizing Afatinib derivatives to find compounds that show more potency. During the development process of these kinds of new compounds, nitrogen-containing 4-alkoxybenzaldehyde analogues play an important role since they have wide usefulness in the pharmaceutical and chemical fields [5][6][7]. These groups are frequently focused of interest in medicinal chemistry because of their broad spectrum of activities: anticancer, antimicrobial, antiviral, analgesic, anti-inflammatory, antimicrobial, antihistaminic, antiangiogenic etc.…”