Browse
Submit Data
Databases
API
Help

Dataset Information

10 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Selection and Characterization of Vimentin-Binding Aptamer Motifs for Ovarian Cancer

ABSTRACT: The application of aptamers in biomedicine is emerging as an essential technology in the field of cancer research. As small single-stranded DNA or RNA ligands with high specificity and low immunogenicity for their targets, aptamers provide many advantages in cancer therapeutics over protein-based molecules, such as antibodies. Vimentin is an intermediate filament protein that is overexpressed in endothelial cells of cancerous tissue. High expression levels of vimentin have been associated with increased capacity for migration and invasion of the tumor cells. We have selected and identified thioated aptamers with high specificity for vimentin using human ovarian cancer tissues. Tentative binding motifs were chosen for two vimentin aptamers based on predicted secondary structures. Each of these shorter, tentative binding motifs was synthesized, purified, and characterized via cell binding assays. Two vimentin binding motifs with high fidelity binding were selected and further characterized via cell and tissue binding assays, as well as flow cytometric analysis. The equilibrium binding constants of these small thioated aptamer constructs were also determined. Future applications for the vimentin binding aptamer motifs include conjugation of the aptamers to synthetic dyes for use in targeted imaging and therapy, and ultimately more detailed and precise monitoring of treatment response and tumor progression in ovarian pathology.

SUBMITTER: Costello A

PROVIDER: S-EPMC8588432 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Similar Datasets

Morph-X-Select: Morphology-based tissue aptamer selection for ovarian cancer biomarker discovery.

Project description:High affinity aptamer-based biomarker discovery has the advantage of simultaneously discovering an aptamer affinity reagent and its target biomarker protein. Here, we demonstrate a morphology-based tissue aptamer selection method that enables us to use tissue sections from individual patients and identify high-affinity aptamers and their associated target proteins in a systematic and accurate way. We created a combinatorial DNA aptamer library that has been modified with thiophosphate substitutions of the phosphate ester backbone at selected 5´dA positions for enhanced nuclease resistance and targeting. Based on morphological assessment, we used image-directed laser microdissection (LMD) to dissect regions of interest bound with the thioaptamer (TA) library and further identified target proteins for the selected TAs. We have successfully identified and characterized the lead candidate TA, V5, as a vimentin-specific sequence that has shown specific binding to tumor vasculature of human ovarian tissue and human microvascular endothelial cells. This new Morph-X-Select method allows us to select high-affinity aptamers and their associated target proteins in a specific and accurate way, and could be used for personalized biomarker discovery to improve medical decision-making and to facilitate the development of targeted therapies to achieve more favorable outcomes.

| S-EPMC5136366 | biostudies-literature

Selection, Characterization, and Application of ssDNA Aptamer against Furaneol.

Project description:Furaneol is an aroma compound which occurs naturally in foods and is used as an artificial flavor. Detection of furaneol is required in food science and food processing industry. Capture- Systematic Evolution of Ligands by EXponential enrichment (SELEX) protocol was applied for the isolation of an aptamer binding to furaneol, a small volatile organic substance contributing to the flavor of various products. Thirteen cycles of selection were performed. The resulting DNA pool was cloned, using blunt-end cloning, and ninety-six plasmids were sequenced and analyzed. Eight oligonucleotides were selected as aptamer candidates and screened for the ability to bind to furaneol, using three different methods-magnetic-beads associated elution assay, SYBR Green I assay, and exonuclease protection assay. One of the candidates was further characterized as an aptamer. The apparent equilibrium constant was determined to be (1.1 ± 0.4) µM, by the fluorescent method. The reported aptamer was applied for development of the ion-sensitive field-effect transistor (ISFET)-based biosensor, for the analysis of furaneol, in the concentration range of 0.1⁻10 µM.

| S-EPMC6320952 | biostudies-literature

Selection and Characterization of an α6β4 Integrin blocking DNA Aptamer.

Project description:The heterodimeric laminin receptor α6β4 integrin plays a central role in the promotion of tumor cell growth, invasion, and organotropic metastasis. As an overproduction of the integrin is often linked to a poor prognosis, the inhibition of integrin α6β4 binding to laminin is of high therapeutical interest. Here, we report on the combination of a cell-systematic evolution of ligands by exponential enrichment and a bead-based selection resulting in the first aptamer inhibiting the interaction between α6β4 integrin and laminin-332. This Integrin α6β4-specific DNA Aptamer (IDA) inhibits the adhesion of prostate cancer cells (PC-3) to laminin-332 with an IC50 value of 149 nmol/l. The Kd value concerning the aptamer's interaction with PC-3 cells amounts to 137 nmol/l. Further characterization showed specificity to α6 integrins and a half-life in murine blood plasma of 6 hours. Two truncated versions of the aptamer retained their binding capacity, but lost their ability to inhibit the interaction between laminin-332 and PC-3 cells. Confocal laser scanning microscope studies revealed that the aptamer was internalized into PC-3-cells. Therefore, in addition to the adhesion-blocking function of this aptamer, IDA could also be applied for the delivery of siRNA, microRNA or toxins to cancer cells presenting the integrin α6β4.

| S-EPMC5014452 | biostudies-literature

A Novel ssDNA Aptamer Targeting Carcinoembryonic Antigen: Selection and Characterization.

Project description:One of the major causes of a drastically shorter life expectancy and one of the most prevalent diseases in the world today is cancer. Given the data on the rise in cancer cases throughout the world, it is obvious that, despite the diagnostic techniques currently being used, there is a pressing need to develop precise and sensitive techniques for early diagnosis of the disease. A high degree of affinity and specificity towards particular targets is maintained by the short nucleic acid molecules known as aptamers. Aptamers outperform antibodies due to their unique benefits, such as their simplicity in synthesis and modification, lack of toxicity, and long-term stability. Utilizing an accurate recognition element and a robust signal transduction mechanism, molecular diagnostics can be extremely sensitive and specific. In this study, development of new single-stranded DNA aptamers against CEA for use in cancer diagnostics was accomplished using SELEX and NGS methods. As a result of 12 iterative SELEX rounds, nine aptamer candidates against CEA were developed. NGS comparative analysis revealed that round twelve had an enriched number of aptamers that were specifically bound, as opposed to round eight. Among the selected nine sequences characterized by bioinformatics analysis and ELONA, an aptamer sequence with the highest specificity and affinity for the target protein was identified and further examined. Aptamer sequence (6) was screened in a concentration-dependent assay, specificity analysis was performed, and its potential secondary and tertiary structures were predicted, which enabled us to test one of the possible putative interactions with CEA. Finally, aptamer sequence (6) labelled with a Cy5 fluorescent tag was used in confocal microscopy to observe its binding towards the CEA expressed in HT-29 human colon adenocarcinoma cell line.

| S-EPMC9598387 | biostudies-literature

DNA-aptamer targeting vimentin for tumor therapy in vivo.

Project description:In recent years, new prospects for the use of nucleic acids as anticancer drugs have been discovered. Aptamers for intracellular targets can regulate cellular functions and cause cell death or proliferation. However, intracellular aptamers have limited use for therapeutic applications due to their low bioavailability. In this work, we selected DNA aptamers to cell organelles and nucleus of cancer cells, and showed that an aptamer NAS-24 binds to vimentin and causes apoptosis of mouse ascites adenocarcinoma cells in vitro and in vivo. To deliver the aptamer NAS-24 inside cells, natural polysaccharide arabinogalactan was used as a carrier reagent. The mixture of arabinogalactan and NAS-24 was injected intraperitonealy for 5 days into mice with adenocarcinoma and inhibited adenocarcinoma growth more effectively than free arabinogalactan or the aptamer alone. The use of aptamers to intracellular targets together with arabinogalactan becomes a promising approach for anticancer therapy.

| S-EPMC3962644 | biostudies-literature

Downregulation of vimentin expression increased drug resistance in ovarian cancer cells.

Project description:Cisplatin and other platinum-based drugs have been widely used in the treatment of ovarian cancer, but most patients acquire the drug resistance that greatly compromises the efficacy of drugs. Understanding the mechanism of drug resistance is important for finding new therapeutic approaches. In the present study, we found that the expression of vimentin was downregulated in drug-resistant ovarian cancer cell lines A2780-DR and HO-8910 as compared to their respective control cells. Overexpression of vimentin in A2780-DR cells markedly increased their sensitivity to cisplatin, whereas knockdown of vimentin in A2780, HO-8910-PM and HO-8910 cells increased the resistance to cisplatin, demonstrating that vimentin silencing enhanced cisplatin resistance in ovarian cancer cells. Quantitative proteomic analysis identified 95 differentially expressed proteins between the vimentin silenced A2780 cells (A2780-VIM-KN) and the control cells, in which downregulation of endocytic proteins and the upregulation of exocytotic proteins CHMP2B and PDZK1 were proposed to contribute the decreased cisplatin accumulation in vimentin knockdown cells. Silencing of vimentin induced upregulation of cancer stem cell markers and both A2780-DR and A2780-VIM-KN cells were more facile to form spheroids than control cells under serum-free culture condition. Our results also revealed that vimentin knockdown increased the 14-3-3 mediated retention of Cdc25C in the cytoplasm, leading to inactivation of Cdk1 and the prolonged G2 phase arrest that allowed the longer period of time for cells to repair cisplatin-damaged DNA. Taken together, we demonstrated that vimentin silencing enhanced cells' resistance to cisplatin via prolonged G2 arrest and increased exocytosis, suggesting that vimentin is a potential target for treatment of drug resistant ovarian cancer.

| S-EPMC5216767 | biostudies-literature

Evolution and characterization of a benzylguanine-binding RNA aptamer.

Project description:Repurposing the "protein-labeling toolkit" for RNA research could be a pragmatic approach for developing new RNA-labeling methods. We have evolved an RNA aptamer that tightly binds benzylguanine (bG), the key ligand for the protein SNAP-tag. The aptamer tightly binds bG fluorophores and can be purified from cellular RNA with bG agarose under native conditions.

| S-EPMC5020705 | biostudies-literature

Computable structured aptamer for targeted treatment of ovarian cancer.

Project description:Nucleolin protein expression is higher on the ovarian cancer cell surface. AS1411, a DNA aptamer, can bind with nucleolin protein specifically. In this study, we developed HA and ST DNA tiles to assemble six AS1411 aptamers to deliver doxorubicin. In addition, to superior serum stability and drug loading, HA-6AS and ST-6AS outperformed TDN-AS in cellular uptake. HA-6AS and ST-6AS exhibited satisfactory targeted cytotoxicity and achieved resounding lysosomal escape. Moreover, when injected into nude mice subcutaneous xenograft models, HA-6AS reached the peak in tumor more quickly than ST-6AS, and better expressed the active targeting ability of AS1411. Our study suggests that designing appropriate DNA tiles to assemble different aptamers to deliver different chemotherapeutic drugs is a promising treatment for ovarian cancer.

| S-EPMC10189780 | biostudies-literature

Selection and characterization of DNA aptamer against glucagon receptor by cell-SELEX.

Project description:Excessive secretion of glucagon, a functional insulin antagonist, significantly contributes to hyperglycemia. Glucagon exerts its physiological functions through activation of the glucagon receptor (GCGR). Inhibition of GCGR activity represents a potential therapeutic approach for reducing excess glucose production in diabetes mellitus. Aptamers are short DNA or RNA oligonucleotides evolved from systematic evolution of ligands by exponential enrichment (SELEX). Here, we have successfully selected a DNA aptamer against GCGR by cell-SELEX, which can specifically bind membrane protein of CHO-GCGR cells with a K d of 52.7 ± 5.1 nM. Aptamer-mediated pull-down and gcgr knockdown assay verified that GCGR was the target of aptamer GR-3. Binding analysis revealed that GR-3 could recognize other cells with different affinity according to the level of GCGR protein expressed in these cells. Hepatic tissue imaging suggested that GR-3 could bind the cell membrane of hepatic tissues. With the advantages of small size, high binding affinity, good stability, lack of immunogenicity, and easy synthesis, aptamer GR-3 against GCGR can be a promising tool with the potential to attenuate hyperglycemia in diabetes mellitus.

| S-EPMC5543139 | biostudies-literature

Native Top-Down Mass Spectrometry Uncovers Two Distinct Binding Motifs of a Functional Neomycin-Sensing Riboswitch Aptamer.

Project description:Understanding how ligands bind to ribonucleic acids (RNA) is important for understanding RNA recognition in biological processes and drug development. Here, we have studied neomycin B binding to neomycin-sensing riboswitch aptamer constructs by native top-down mass spectrometry (MS) using electrospray ionization (ESI) and collisionally activated dissociation (CAD). Our MS data for a 27 nt aptamer construct reveal the binding site and ligand interactions, in excellent agreement with the structure derived from nuclear magnetic resonance (NMR) studies. Strikingly, for an extended 40 nt aptamer construct, which represents the sequence with the highest regulatory factor for riboswitch function, we identified two binding motifs for neomycin B binding, one corresponding to the bulge-loop motif of the 27 nt construct and the other one in the minor groove of the lower stem, which according to the MS data are equally populated. By replacing a noncanonical with a canonical base pair in the lower stem of the 40 nt aptamer, we can reduce binding to the minor groove motif from ∼50 to ∼30%. Conversely, the introduction of a CUG/CUG motif in the lower stem shifts the binding equilibrium in favor of minor groove binding. The MS data reveal site-specific and stoichiometry-resolved information on aminoglycoside binding to RNA that is not directly accessible by other methods and underscore the role of noncanonical base pairs in RNA recognition by aminoglycosides.

| S-EPMC10360057 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data