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Nucleic acid monomers and miniTAR±aptamer kissing complex. ( A ) Schematic representation of the sugar chemical structure adopted by a DNA, RNA and LNA monomer. ( B ) Loop±loop complex formed between the R06 16 aptamer (bold) and miniTAR RNA used in this study. MiniTAR is an imperfect hairpin corresponding to the top part of the retroviral TAR
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One of the major limitations of the use of phosphodiester oligonucleotides in cells is their rapid degradation by nucleases. To date, several chemical modifications have been employed to overcome this issue but insufficient efficacy and/or specificity have limited their in vivo usefulness. In this work conformationally restricted nucleotides, locke...
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Context 1
... furanose ring of LNA monomers is locked and adopts a C3¢-endo conformation similar to the RNA one (Fig. 1A) that marks them, a priori, as good structural analogues of RNA (24). We took advantage of this to generate oligonucleotides with LNA modi®cations, derived from a shortened version of the RNA aptamer of highest af®nity identi®ed by in vitro selection against TAR. The results show that a fully modi®ed LNA version of this aptamer is not ...
Context 2
... and RNA molecules including the biotinylated target, miniTAR, were synthesized on an Expedite 8908 synthesizer. MiniTAR (Fig. 1B) is an imperfect RNA hairpin correspond- ing to the top part of the retroviral TAR element, which maintains biological responsiveness. Compounds containing LNA units were synthesized on an automated DNA synthe- sizer as previously described (34). All oligonucleotides were puri®ed by electrophoresis on denaturing 20% polyacryl- amide/7 M ...
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... 1, the stem can be shortened to 3 bp without signi®cant loss of af®nity for miniTAR (K d = 9.1 T 1.1 nM). The octameric 5¢- GUCCCAGA-3¢ consensus sequence, R06 8 , characteristic of the selected kissing aptamers, is not a good TAR ligand in agreement with previous results (20,35). We chose to design the LNA derivatives on the basis of R06 16 (Fig. 1B), a truncated oligomer eight bases shorter than R06 24 ...
Similar publications
Locked nucleic acid (LNA), a recently introduced nucleic acid
analogue with a bicyclic 2′-O,4′-C-methylene
linked furanose sugar, exhibits enhanced affinities for DNA and RNA
relative to the corresponding oligodeoxyribonucleotides and
oligoribonucleotides; we report the first crystal structure of an LNA unit
incorporated in an oligonucleotide...
Citations
... 29,30 Several protective chemical modifications can be introduced to improve the stability of DNA oligonucleotides, and this can affect their functionality, especially when using aptamers. 31 As an alternative, the use of resistant oligonucleotide analogs, for self-assembly into nanoparticles, has become increasingly important. 17 Peptide nucleic acids (PNAs), 32 polyamide analogs of DNA, are more stable and easier to produce, and their derivatives can be particularly useful in nanotechnology. ...
Organosilica nanoparticles that contain responsive organic building blocks as constitutive components of the silica network offer promising opportunities for the development of innovative drug formulations, biomolecule delivery, and diagnostic tools. However, the synthetic challenges required to introduce dynamic and multifunctional building blocks have hindered the realization of biomimicking nanoparticles. In this study, capitalizing on our previous research on responsive nucleic acid-based organosilica nanoparticles, we combine the supramolecular programmability of nucleic acid (NA) interactions with sol–gel chemistry. This approach allows us to create dynamic supramolecular bridging units of nucleic acids in a silica-based scaffold. Two peptide nucleic acid-based monoalkoxysilane derivatives, which self-assemble into a supramolecular bis-alkoxysilane through direct base pairing, were chosen as the noncovalent units inserted into the silica network. In addition, a bridging functional NA aptamer leads to the specific recognition of ATP molecules. In a one-step bottom-up approach, the resulting supramolecular building blocks can be used to prepare responsive organosilica nanoparticles. The supramolecular Watson–Crick–Franklin interactions of the organosilica nanoparticles result in a programmable response to external physical (i.e., temperature) and biological (i.e., DNA and ATP) inputs and thus pave the way for the rational design of multifunctional silica materials with application from drug delivery to theranostics.
... First of all, modifications are made to nucleic acid-based aptamers (DNA, RNA) to overcome nuclease-mediated cleavage [100]. Some examples of modifications on DNA aptamers are done on linkage (addition of an extra guanine base to the 3 -end) [101], and also on the sugar ring and bases (addition of a methylene linkage between the 2oxygen and the 4 -carbon of the sugar ring) [102]. For RNA aptamers, the most common modification is on the 2 -position of the sugar ring, for example, the addition of a 2 -OH group [103], and substitution with 2 -F on pyrimidine [104]. ...
Aptamers are a group of synthetic single-stranded nucleic acids. They are generated from a random library of single-stranded DNA or RNA by a technology named systematic evolution of ligands by exponential enrichment (SELEX). SELEX is a repetitive process to select and identify suitable aptamers that show high affinity and specificity towards target cells. Great strides have been achieved in the design, construction, and use of aptamers up to this point. However, only a small number of aptamer-based applications have achieved widespread commercial and clinical acceptance. Additionally, finding more effective ways to acquire aptamers with high affinity remains a challenge. Therefore, it is crucial to thoroughly examine the existing dearth and advancement in aptamer-related technologies. This review focuses on aptamers that are generated by SELEX to detect pathogenic microorganisms and mammalian cells, as well as in cell-internalizing SELEX for diagnostic and therapeutic purposes. The development of novel aptamer-based biosensors using optical and electrical methods for microbial detection is reported. The applications and limitations of aptamers are also discussed.
... However, Darfeuille et al. studied LNA nucleotides as nuclease-resistant aptamers that are targeted against the HIV-1 trans-activation response RNA, which indicates that LNA units alternated with DNA could increase their stability and impart the capacity of resistance to nuclease action without reducing their affinity for the target. 28 The reports on nucleic acid sensing using XNA probes at the nanoscale, especially at the single or few molecules level, are limited in number. In our laboratory, we have developed simple methods of generating self-assembled and ordered LNA monolayers 18 that are capable of producing a stronger DNA detection signal and improved single nucleobase mismatch recognition, compared to the case of DNA monolayers, as detected by the molecularly resolved SMFS measurements 14,19,23 (Table 3). ...
The strategies for nucleic acid sensing based on nucleic acid hybridization between the target sequence and the capture probe sequence are considered to be largely successful as far as detection of a specific target of known sequence is concerned. However, when compared with other complementary methods, like direct sequencing, a number of results are still found to be either "false positives" or "false negatives". This suggests that modifications in these strategies are necessary to make them more accurate. In this minireview, we propose that one way toward improvement could be replacement of the DNA capture probes with the xeno nucleic acid or XNA capture probes. This is because the XNAs, especially the locked nucleic acid, the peptide nucleic acid, and the morpholino, have shown better single nucleobase mismatch discrimination capacity than the DNA capture probes, indicating their capacity for more precise detection of nucleic acid sequences, which is beneficial for detection of gene stretches having point mutations. Keeping the current trend in mind, this minireview will include the recent developments in nanoscale, fluorescent label-free applications, and present the cases where the XNA probes show clear advantages over the DNA probes.
... LNAs increase the thermodynamic stability, binding affinity, and enable the oligonucleotides to prevent serum degradation. [50][51][52] These modifications enable the aptamers for biological applications. ...
Aptamers are a short polymer of oligonucleotides (natural or modified) that can bind to its cognate target (small molecules to large macromolecules like proteins, cells, microorganisms etc.) with high affinity and selectivity. They can fold into unique secondary and tertiary conformation in solution (pH, ionic concentration) and bind to their targets in a specific manner (binding constants in sub-nano to pico molar range). They rival the monoclonal antibodies and other specific biological ligands with respect to affinity, stability, robustness, non-immunogenicity and facile to synthesis. Nucleic acid aptamers are selected from an oligonucleotide library by an iterative process called SELEX (Systematic Evolution of Ligands by Exponential Enrichment Analysis). These aptamers are compatible to any kind of chemical modification , conjugation and functionalization. Briefly, this chapter discusses about the diagnostic and therapeutic application of aptamers.
... 33 It was reported that a chimeric LNA/DNA aptamer targeting HIV-1 trans-activating response protein could remain intact for more than 20 h in serum. 75 Replacement of the phosphodiester bonds of aptamers with phosphorothioate or methylphosphonate analogues is also a conventional strategy. 76 To enhance the stability of the phosphodiester backbone in the anti-thrombin aptamer, replacements with thiophosphoryl at different nucleotide sites were evaluated. ...
Aptamers are oligonucleotide sequences with a length of about 25-80 bases which have abilities to bind to specific target molecules that rival those of monoclonal antibodies. They are attracting great attention in diverse clinical translations on account of their various advantages, including prolonged storage life, little batch-to-batch differences, very low immunogenicity, and feasibility of chemical modifications for enhancing stability, prolonging the half-life in serum, and targeted delivery. In this Review, we demonstrate the emerging aptamer discovery technologies in developing advanced techniques for producing aptamers with high performance consistently and efficiently as well as requiring less cost and resources but offering a great chance of success. Further, the diverse modifications of aptamers for therapeutic applications including therapeutic agents, aptamer-drug conjugates, and targeted delivery materials are comprehensively summarized.
... LNA/DNA chimera LNA5, forming a stable complex against HIV-1 trans-activating response (TAR) RNA, was synthesized from a shortened and stable version of the hairpin RNA aptamer identified by in vitro selection against TAR. The results indicated that these modifications provide good protection towards nuclease digestion in bovine serum and keep the same binding affinity of the unmodified RNA aptamer [89]. Shi et al. developed another LNA/DNA chimeric aptamer probe through proper LNA incorporation and 3′-3′-thymidine (3′-3′-T) capping ( Figure 8). ...
Soon after they were first described in 1990, aptamers were largely recognized as a new class of biological ligands that can rival antibodies in various analytical, diagnostic, and therapeutic applications. Aptamers are short single-stranded RNA or DNA oligonucleotides capable of folding into complex 3D structures, enabling them to bind to a large variety of targets ranging from small ions to an entire organism. Their high binding specificity and affinity make them comparable to antibodies, but they are superior regarding a longer shelf life, simple production and chemical modification, in addition to low toxicity and immunogenicity. In the past three decades, aptamers have been used in a plethora of therapeutics and drug delivery systems that involve innovative delivery mechanisms and carrying various types of drug cargos. However, the successful translation of aptamer research from bench to bedside has been challenged by several limitations that slow down the realization of promising aptamer applications as therapeutics at the clinical level. The main limitations include the susceptibility to degradation by nucleases, fast renal clearance, low thermal stability, and the limited functional group diversity. The solution to overcome such limitations lies in the chemistry of aptamers. The current review will focus on the recent arts of aptamer chemistry that have been evolved to refine the pharmacological properties of aptamers. Moreover, this review will analyze the advantages and disadvantages of such chemical modifications and how they impact the pharmacological properties of aptamers. Finally, this review will summarize the conjugation strategies of aptamers to nanocarriers for developing targeted drug delivery systems.
... Otras modificaciones, como la inversión del 3' , 2'-fluoro, 2'-O-metil y la sustitución del 2' mediante aminoácidos captados por ácido nucleico bloqueado, mejoran la estabilidad metabólica, la afinidad de la interacción y la filtración renal (44,45). ...
Los aptámeros son secuencias de ADN o ARN de cadena sencilla que adoptan la forma de estructuras tridimensionales únicas, lo cual les permite reconocer un blanco específico con gran afinidad. Sus usos potenciales abarcan, entre otros, el diagnóstico de enfermedades, el desarrollo de nuevos agentes terapéuticos, la detección de riesgos alimentarios, la producción de biosensores, la detección de toxinas, el transporte de fármacos en el organismo y la señalización de nanopartículas.
El pegaptanib es el único aptámero aprobado para uso comercial por la Food and Drug Administration (FDA).
En parasitología, se destacan los estudios que se vienen realizando en Leishmania spp., con la obtención de aptámeros que reconocen la proteína de unión a poliA (LiPABP) y que pueden tener potencial utilidad en la investigación, el diagnóstico y el tratamiento de la leishmaniasis. En cuanto a la malaria, se han obtenido aptámeros que permiten identificar eritrocitos infectados e inhiben la formación de rosetas, y otros que prometen ser alternativas para el diagnóstico al detectar de forma específica la proteína lactato deshidrogenasa (PfLDH). Para Cryptosporidium parvuum se han seleccionado aptámeros que detectan ooquistes a partir de alimentos o aguas contaminadas. Para Entamoeba histolytica se han aislado dos aptámeros llamados C4 y C5, que inhiben la proliferación in vitro de los trofozoítos y tienen potencial terapéutico. Los aptámeros contra Trypanosoma cruzi inhiben la invasión de células LLC-MK2 (de riñón de mono) en un 50 a 70 % y aquellos contra T. brucei transportan moléculas tóxicas al lisosoma parasitario como una novedosa estrategia terapéutica.
... 71 The effect of the incorporation of LNA nucleotides into an RNA aptamer specific to the HIV-1 TAR RNA element has also been reported. 72 Fang et al. reported LNA microarrays for the detection of LNA-captured miRNAs by first adding poly(A) tail to the surface-bound miRNA via the poly(A) polymerase reaction and then hybridizing with T 30 DNA-modified AuNPs for AuNP-amplified surface plasmon resonance imaging (SPRI) detection. 69 LNA has been incorporated into molecular beacons (MB), resulting in the combined advantage of sensitivity that MB-based detection imparts and the high binding affinity and stability that the LNA-containing nucleic acid sequences exhibit. ...
Challenges in reliable nucleic acid detection are manifold. The major ones are related to false positive or negative signals due to a lack of target specificity in detection, and to low sensitivity, especially when a plethora of background sequences are present that can mask the specific recognition signal. Utilizing designed synthetic nucleic acids that are commonly called xeno nucleic acids could offer potential routes to meet such challenges. In this article, we have presented the general framework of nucleic acid detection, especially keeping nanoscale applications in mind, and discussed how and why the xeno nucleic acids could be truly an alternative to the DNA probes. Two specific cases of locked nucleic acid (LNA) and peptide nucleic acid (PNA), which are nuclease-resistant and can form thermally stable duplexes with DNA, have been addressed. It is shown that the relative ease of the conformationally rigid LNA probe to be oriented upright on substrate surface, and of the non-ionic PNA probe to result into high probe density assist in their use in nanoscale nucleic acid recognition. It is anticipated that success with these probes may lead to important developments such as PCR-independent approaches where the major aim is to detect low number of target sequences in the analyte medium.
... Two groups first used SELEX to select highly avid and specific RNA aptamers for particular targets, including organic dyes [10] and bacteriophage T4 DNA polymerase [12,87]. Since then, aptamers have been selected for a variety of targets, including metal ions [88], organic molecules [89], amino acids [90][91][92], viral nucleic acid components [93][94][95][96], peptides [97], proteins [5,6,98], drugs [99][100][101], macromolecules [102][103][104][105], cells [106,107], and pathogens [108][109][110][111][112][113]. ...
Aptamers are versatile oligonucleotide ligands used for molecular recognition of diverse targets. However, application of aptamers to the field of amyloid β-protein (Aβ) has been limited so far. Aβ is an intrinsically disordered protein that exists in a dynamic conformational equilibrium, presenting time-dependent ensembles of short-lived, metastable structures and assemblies that have been generally difficult to isolate and characterize. Moreover, despite understanding of potential physiological roles of Aβ, this peptide has been linked to the pathogenesis of Alzheimer disease, and its pathogenic roles remain controversial. Accumulated scientific evidence thus far highlights undesirable or nonspecific interactions between selected aptamers and different Aβ assemblies likely due to the metastable nature of Aβ or inherent affinity of RNA oligonucleotides to β-sheet-rich fibrillar structures of amyloidogenic proteins. Accordingly, lessons drawn from Aβ–aptamer studies emphasize that purity and uniformity of the protein target and rigorous characterization of aptamers’ specificity are important for realizing and garnering the full potential of aptamers selected for recognizing Aβ or other intrinsically disordered proteins. This review summarizes studies of aptamers selected for recognizing different Aβ assemblies and highlights controversies, difficulties, and limitations of such studies.
... This modification showed great resistance to nucleases and increased thermostability thus could be used to generate the most stable pairs [50,51]. Darfeuille et al. also found that the LNA/DNA chimera LNA5, a stable complex that against HIV-1 trans-activating response (TAR) RNA, was able to maintain the intact structure within 20 h in bovine serum [52]. ...
... LNA, UNA, 2′-F ANA Locked nucleic acid (LNA)(Figure 7)is an analog of ribonucleotide with a methylene linkage between 2′-O and 4′-C of the sugar ring. This modification showed great resistance to nucleases and increased thermostability thus could be used to generate the most stable pairs[50,51].Darfeuille et al. also found that the LNA/DNA chimera LNA5, a stable complex that against HIV-1 trans-activating response (TAR) RNA, was able to maintain the intact structure within 20 h in bovine serum[52]. Shi et al. developed a new LNA/DNA chimeric aptamer probe through proper LNA incorporation and ...
... Chemical modifications of nucleic acid aptamers for different purposes. 2′-fluoro, 2′-O-methyl and 2′-amino-substitutions1[39,48,49] locked nucleic acid (LNA)[52,53] unlocked nucleic acid (UNA)[54,55] 2′-deoxy-2′-fluoro-D-arabinonucleic acid (2′-F ANA) ...
Nucleic acid aptamers have minimal immunogenicity, high chemical synthesis production, low cost and high chemical stability when compared with antibodies. However, the susceptibility to nuclease degradation, rapid excretion through renal filtration and insufficient binding affinity hindered their development as drug candidates for therapeutic applications. In this review, we will discuss methods to conquer these challenges and highlight recent developments of chemical modifications and technological advances that may enable early aptamers to be translated into clinical therapeutics.
