RESUMO
The investigation of long noncoding RNAs (lncRNAs) and RNA binding proteins (RBPs) interactions in living cell holds great significance for elucidating their critical roles in a variety of biological activities, but limited techniques are available to profile the temporal-spatial dynamic heterogeneity. Here, we introduced a molecular beacon-functionalized nanoneedle array designed for spatially resolved profiling of lncRNA-RBP interactions (Nano-SpatiaLR). A nanoneedle array modified with a molecular beacon is employed to selectively isolate specific intracellular lncRNAs and their associated RBPs without affecting cell viability. The RBPs are then in situ analyzed with a fluorescent labeled antibody and colocalized with lncRNA signals to get a quantitative measurement of their dynamic interactions. Additionally, leveraging the spatial distribution and nanoscale modality of the nanoneedle array, this technique provides the spatial heterogeneity information on cellular lncRNA-RBPs interaction at single cell resolution. In this study, we tracked the temporal-spatial interactive heterogeneity dynamics of lncRNA-RBPs interaction within living cells across different biological progresses. Our findings demonstrated that the interactions between lncRNA HOTAIR and RBPs EZH2 and LSD1 undergo significant changes in response to drug treatments, particularly in tumor cells. Moreover, these interactions become more intensified as tumor cells aggregate during the proliferation process.
Assuntos
RNA Longo não Codificante , Proteínas de Ligação a RNA , Análise de Célula Única , RNA Longo não Codificante/metabolismo , RNA Longo não Codificante/genética , Humanos , Proteínas de Ligação a RNA/metabolismo , Proteínas de Ligação a RNA/química , Proteína Potenciadora do Homólogo 2 de Zeste/metabolismoRESUMO
Sonodynamic therapy (SDT) has attracted intensive attention, but is still hindered by low sonosensitization and non-biodegradability of the traditional sonosensitizers. Herein, perovskite-type manganese vanadate (MnVO3 ) sonosensitizers integrating high reactive oxide species (ROS) production efficiency and appropriate bio-degradability are developed for enhanced SDT. Taking advantage of the intrinsic properties of perovskites such as narrow bandgap and substantial oxygen vacancies, MnVO3 shows a facile ultrasound (US)-triggered electrons-holes separation and restrained recombination, thus enhancing the ROS quantum yield in SDT. Furthermore, MnVO3 exhibits a considerable chemodynamic therapy (CDT) effect under the acidic condition probably owing to the presence of manganese and vanadium ions. Due to the presence of high-valent vanadium, MnVO3 can also eliminate glutathione (GSH) within the tumor microenvironment, which synergistically amplifies the efficacy of SDT and CDT. Importantly, the perovskite structure bestows MnVO3 with superior biodegradability, which alleviates the long-term presence of residues in metabolic organs after therapeutic actions. Based on these characteristics, US-assisted MnVO3 achieves an excellent antitumor outcome along with low systemic toxicity. Overall, perovskite-type MnVO3 may be promising sonosensitizers for highly efficient and safe treatment of cancer. The work attempts to explore the potential utility of perovskites in the design of degradable sonosensitizers.
Assuntos
Neoplasias , Terapia por Ultrassom , Humanos , Vanadatos , Vanádio , Manganês , Espécies Reativas de Oxigênio , Neoplasias/terapia , Glutationa , Óxidos , Linhagem Celular Tumoral , Microambiente TumoralRESUMO
Aptamers, which are called chemical antibodies for their high affinity and specificity to targets, have great potential as analytical tools to detect pesticides. In this work, a DNA aptamer for thiamethoxam was isolated by an improved SELEX (systematic evolution of ligands by exponential enrichment) strategy, in which the ssDNA library was fixed on streptavidin-agarose beads through a short biotin labeled complementary strand. After 13 rounds of selection, the random ssDNA pool was successfully enriched. Three sequences were chosen as aptamer candidates through sequencing and analysis and were transformed into fluorescent probes to evaluate their interactions with thiamethoxam. A fluorescent turn-on aptasensor for thiamethoxam based on the best aptamer (FAM-Thi13) and a short quenching strand were further designed and showed a quantitative linear range from 10 to 1000 nM with a detection limit of 1.23 nM for thiamethoxam. Molecular docking and molecular dynamics were used to investigate the binding site of the main probe of the aptasensor (FAM-Thi13) and thiamethoxam. Satisfactory results were also obtained in quantifying thiamethoxam in environmental water samples by the developed fluorescent aptasensor.
Assuntos
Aptâmeros de Nucleotídeos , Praguicidas , DNA de Cadeia Simples/genética , Simulação de Acoplamento Molecular , Técnica de Seleção de Aptâmeros , TiametoxamRESUMO
G-Quadruplexes (G4s) assembled from tandem G-rich repeat sequences exhibit significant biological functions and applications, which may well depend on their structural features, such as the planar arrangement of G-tetrads and flexibility of loop regions. It has been found that cytosine-intercalated G-repeat sequences also assemble to be quadruplex structures, involving the formation of nonplanar GCGC-tetrads. Herein, to investigate the effect of GCGC-tetrads on structural properties of G4s, some previously studied quadruplexes with or without GCGC-tetrads were selected, and were used to interact with various developed G4 ligands. Our data show that stacked G-tetrads in quadruplexes are important for the π-π stacking interactions, thus promoting the combination with end-stacking ligands, such as porphyrins or planar small molecules. This is confirmed by the observation that the quadruplex formed by d(GGGCT4 GGGC) with two internal G-tetrads and two external GCGC-tetrads can bind to planar ligands in the presence of specific G4-stabilizing cations, including K+ and Pb2+ , and can realize the sensitive detection of Pb2+ . However, the quadruplex composed of two external G-tetrads and two internal GCGC-tetrads formed by d(GCGGT3 GCGG) facilitates the binding of nonplanar ligands, such as triphenylmethane (TPM) dyes, owing to the structural flexibility induced by internal GCGC-tetrads. This work provides new insights into the interaction between DNA quadruplexes and specific ligands, which is beneficial to the development of quadruplex-based biosensors and the design of anticancer drugs.
Assuntos
DNA/química , Quadruplex G , Porfirinas , Cátions/química , LigantesRESUMO
Numerous studies have shown compelling evidence that incorporation of an inversion of polarity site (IPS) in G-rich sequences can affect the topological and structural characteristics of G-quadruplexes (G4s). Herein, the influence of IPS on the formation of a previously studied intramolecular parallel G4 of d(G3 TG3 TG3 TG3 ) (TTT) and its stacked higher-order structures is explored. Insertion of 3'-3' or 5'-5' IPS did not change the parallel folding pattern of TTT. However, both the species and position of the IPS in TTT have a significant impact on the G4 stability and end-stacking through the alteration of G4-G4 interfaces properties. The data demonstrate that one base flip in each terminal G-tetrad can stabilize parallel G4s and facilitate intermolecular packing of monomeric G4s. Such modifications can also enhance the fluorescence and enzymatic performances by promoting interactions between parallel G4s with N-methyl mesoporphyrin IX (NMM) and hemin, respectively.
Assuntos
DNA Catalítico/química , Guanosina/química , Hemina/química , Mesoporfirinas/química , Quadruplex G , Estrutura MolecularRESUMO
A quadruplex-integrated assembly method is proposed for the organization and regulation of various nanoscale architectures. In this method, two types of one-dimensional DNA nanostructures formed by two well-designed GC-rich single strands assemble into two-dimensional (2D) DNA nanostructures based on the self-assembly of dimeric G-quadruplex and I-motif structures. Subsequently, a C-rich strand and two biotin-modified G-rich strands primordially form a notched double helix in LiCl solution (pH 8). However, a linear "DNA-protein" nanostructure linked by I-motif structures and biotin-streptavidin interaction can be formed when hydrogen ions and streptavidin are sequentially titrated. Furthermore, the linear "DNA-protein" nanostructure is assembled into 2D nanomaterials connected by K+-stabilized G-quadruplexes formed from terminal G-rich repeats of the two G-rich strands. Interestingly, the 2D nanohybrids form two-lined "DNA-protein" nanostructures if the terminal G-rich repeats in one of the biotin-modified G-rich strands are removed. Our results indicate that quadruplex DNAs are promising building blocks in the fabrication of nanomaterials and that the assembly of quadruplex DNAs has potential applications in the directional arrangement of macromolecules.
Assuntos
DNA/química , Quadruplex G , Nanoestruturas/químicaRESUMO
G-quadruplex (G4)/hemin DNAzymes have been extensively applied in bioanalysis and molecular devices. However, their catalytic activity is still much lower than that of proteinous enzymes. The G4/hemin DNAzyme activity is correlated with the G4 conformations and the solution conditions. However, little is known about the effect of the flanking sequences on the activity, though they are important parts of G4s. Here, we report sequences containing d(CCC), flanked on both ends of the G4-core sequences remarkably enhance their DNAzyme activity. By using circular dichroism and UV-visible spectroscopy, the d(CCC) flanking sequences were demonstrated to improve the hemin binding affinity to G4s instead of increasing the parallel G4 formation, which might explain the enhanced DNAzyme activity. Meanwhile, the increased hemin binding ability promoted the degradation of hemin within the DNAzyme by H2O2. Furthermore, the DNAzyme with d(CCC) flanking sequences showed strong tolerance to pH value changes, which makes it more suitable for applications requiring wide pH conditions. The results highlight the influence of the flanking sequences on the DNAzyme activity and provide insightful information for the design of highly active DNAzymes.
Assuntos
DNA Catalítico/química , Quadruplex G , Oligodesoxirribonucleotídeos/química , Dicroísmo Circular , Hemina/química , Peróxido de Hidrogênio/metabolismoRESUMO
Circulating tumor cells (CTCs) are valuable circulating biomarkers of cancer, which carry primary tumor information and may provide real-time assessment of tumor status as well as treatment response in cancer patients. Herein, we developed a novel assay for accurate diagnosis and dynamic monitoring of epithelial ovarian cancer (EOC) using CTC RNA analysis. Multiantibody-modified magnetic nanoparticles were prepared for purification of EOC CTCs from whole blood samples of clinical patients. Subsequently, nine EOC-specific mRNAs of purified CTCs were quantified using droplet digital PCR. The EOC CTC Score was generated using a multivariate logistic regression model for each sample based on the transcripts of the nine genes. This assay exhibited a distinguishing diagnostic performance for the detection of EOC (n = 17) from benign ovarian tumors (n = 30), with an area under the receiver operating characteristic curve (AUC) of 0.96 (95% CI = 0.91-1.00). Moreover, dynamic changes of the EOC CTC Score were observed in patients undergoing treatment, demonstrating the potential of the assay for monitoring EOC. In conclusion, we present an accurate assay for the diagnosis and monitoring of EOC via CTC RNA analysis, and the results suggest that it may provide a promising solution for the detection and treatment response assessment of EOC.
Assuntos
Nanopartículas de Magnetita , Células Neoplásicas Circulantes , Neoplasias Ovarianas , Humanos , Feminino , Carcinoma Epitelial do Ovário/diagnóstico , Células Neoplásicas Circulantes/patologia , Biomarcadores Tumorais/genética , Neoplasias Ovarianas/diagnóstico , Neoplasias Ovarianas/tratamento farmacológico , Neoplasias Ovarianas/patologia , RNARESUMO
Circulating tumor cells (CTCs) have been well-established as promising biomarkers that can be leveraged to gauge the prognosis of patients with cancers and to guide patient treatment efforts. Although the scarcity of CTCs within peripheral circulation and the associated phenotypic changes that they exhibit owing to the epithelial-mesenchymal transition (EMT) process make the reliable isolation of these cells very challenging. Recently, several studies have discussed platforms capable of mediating the efficient and sensitive isolation of CTCs, but these approaches are nonetheless subject to certain limitations that preclude their clinical application. For example, these platforms are poorly-suited to minimizing damage in the context of cellular capture and release or the in vitro culture of captured cells for subsequent molecular analyses, which would better enable clinicians to select appropriate precision treatments on an individualized basis. In this study, we report the layer-by-layer assembly approach to synthesize a novel composite nanomaterial consisting of modified zirconium-based metal-organic-frameworks (MOFs) on the surface of magnetic beads with dual antibody surface modifications capable of capturing CTCs without being hampered by the state of cellular EMT process. Our analyses indicated that these dual antibody-modified nanomaterials exhibited greater capture efficiency than that observed for single antibody. Importantly, captured cells can be gradually released following capture and undergo subsequent in vitro proliferation following water molecule-induced MOF structural collapse. This release mechanism, which does not require operator intervention, may be effective as a means of minimizing damage and preserving cellular viability such that cells can be more reliably utilized for downstream molecular analyses and associated treatment planning. To further confirm the potential clinical applicability of the developed nanomaterial, it was successfully utilized for capturing CTCs from peripheral blood samples collected from cases diagnosed with gastrointestinal tumors.
RESUMO
Colorectal cancer (CRC) is one of the main causes of cancer-related morbidity and mortality across the globe. Although serum biomarkers such as carcinoembryonic antigen (CEA) and carbohydrate antigen 19-9 (CA-199) have been prevalently used as biomarkers in various cancers, they are neither very sensitive nor highly specific. Repeated tissue biopsies at different times of the disease can be uncomfortable for cancer patients. Additionally, the existence of tumor heterogeneity and the results of local biopsy provide limited information about the overall tumor biology. Against this backdrop, it is necessary to look for reliable and noninvasive biomarkers of CRC. Circulating tumor cells (CTCs), which depart from a primary tumor, enter the bloodstream, and imitate metastasis, have a great potential for precision medicine in patients with CRC. Various efficient CTC isolation platforms have been developed to capture and identify CTCs. The count of CTCs, as well as their biological characteristics and genomic heterogeneity, can be used for the early diagnosis, prognosis, and prediction of treatment response in CRC. This study reviewed the existing CTC isolation techniques and their applications in the clinical diagnosis and treatment of CRC. The study also presented their limitations and provided future research directions.
Assuntos
Neoplasias Colorretais , Células Neoplásicas Circulantes , Animais , DNA Tumoral Circulante , Neoplasias Colorretais/diagnóstico , Neoplasias Colorretais/genética , Neoplasias Colorretais/terapia , Humanos , Medicina de PrecisãoRESUMO
Ovarian cancer (OC) is a lethal disease occurring in women worldwide. Due to the lack of obvious clinical symptoms and sensitivity biomarkers, OC patients are often diagnosed in advanced stages and suffer a poor prognosis. Circulating tumor cells (CTCs), released from tumor sites into the peripheral blood, have been recognized as promising biomarkers in cancer prognosis, treatment monitoring, and metastasis diagnosis. However, the number of CTCs in peripheral blood is low, and it is a technical challenge to isolate, enrich, and identify CTCs from the blood samples of patients. This work develops a simple, effective, and inexpensive strategy to capture and identify CTCs from OC blood samples using the folic acid (FA) and antifouling-hydrogel-modified fluorescent-magnetic nanoparticles. The hydrogel showed a good antifouling property against peripheral blood mononuclear cells (PBMCs). The FA was coupled to the hydrogel surface as the targeting molecule for the CTC isolation, held a good capture efficiency for SK-OV-3 cells (95.58%), and successfully isolated 2-12 CTCs from 10 OC patients' blood samples. The FA-modified fluorescent-magnetic nanoparticles were successfully used for the capture and direct identification of CTCs from the blood samples of OC patients.
Assuntos
Nanopartículas de Magnetita , Células Neoplásicas Circulantes , Neoplasias Ovarianas , Linhagem Celular Tumoral , Feminino , Ácido Fólico , Humanos , Leucócitos Mononucleares , Neoplasias Ovarianas/diagnósticoRESUMO
Tympanic membrane (TM) perforation leads to persistent otitis media, conductive deafness, and affects life quality. Ointment medication may not be sufficient to treat TM perforation (TMP) due to the lack of an underlying tissue matrix and thus requiring a scaffold-based application. The engineering of scaffold biointerface close to the matrix via tissue-specific decellularized extracellular matrix (dECM) is crucial in instructing cell behaviour and regulating cell-material interaction in the bioengineering domain. Herein, polycaprolactone (PCL) and TM-dECM (from Sprague-Dawley rats) were combined in a different ratio in nanofibrous form using an electrospinning process and crosslinked via tannic acid. The histological and biochemical assays demonstrated that chemical and enzymatic decellularization steps removed cellular/immunogenic contents while retaining collagen and glycosaminoglycan. The morphological, physicochemical, thermomechanical, contact angle, and surface chemical studies demonstrated that the tannin crosslinked PCL/dECM nanofibers fine-tune biophysical and biochemical properties. The multifaceted crosslinked nanofibers hold the tunable distribution of dECM moieties, assembled into a spool-shaped membrane, and could easily insert into perforated sites. The dECM decorated fibers provide a preferable biomimetic matrix for L929 fibroblast adhesion, proliferation, matrix adsorption, and f-actin saturation, which could be crucial for bioengineering. Overall, dECM patterning, surface hydrophilicity, interconnected microporosities, and multifaceted nanofibrous biosystem modulate cell-scaffold performance and could open opportunities to reconstruct TMP in a biomimetic fashion.
Assuntos
Nanofibras , Perfuração da Membrana Timpânica , Animais , Bioengenharia , Matriz Extracelular/metabolismo , Nanofibras/química , Ratos , Ratos Sprague-Dawley , Taninos , Engenharia Tecidual , Alicerces Teciduais/química , Perfuração da Membrana Timpânica/metabolismo , Perfuração da Membrana Timpânica/terapiaRESUMO
The majority of current methods of isolating circulating tumor cells (CTCs) rely on a biomarker. However, the isolation efficiency may be compromised due to the heterogeneity of CTCs. In this work, a simple and broad-spectrum method is established to efficiently isolate the heterogeneous CTCs from patient blood samples using tannic acid (TA)-functionalized magnetic nanoparticles (MNPs). The TA-functionalized MNPs (MNPs-TA) inhibit the nonspecific adhesion of peripheral blood mononuclear cell (PBMC) and enhance cancer cell capture, resulting from the unique interaction between TA and glycocalyx on cancer cells. The MNPs-TA was demonstrated to effectively capture seven kinds of cancer cells (HeLa, PC-3, T24, MAD-MB-231, MCF-7, HT1080, A549) from artificial samples (62.3-93.7%). Moreover, this epithelial cell adhesion molecule (EpCAM)-independent CTC isolation method was also tested using clinical blood samples from patients with different cancers (21 patients), which may provide a universal tool to detect CTCs in the clinic.
Assuntos
Separação Celular/métodos , Imãs/química , Nanopartículas/química , Neoplasias/patologia , Células Neoplásicas Circulantes/patologia , Taninos/química , Adulto , Idoso , Linhagem Celular Tumoral , Molécula de Adesão da Célula Epitelial/análise , Feminino , Humanos , Masculino , Pessoa de Meia-IdadeRESUMO
The success of complex tissue and internal organ reconstruction relies principally on the fabrication of a 3D vascular network, which guarantees the delivery of oxygen and nutrients in addition to the disposal of waste. In this study, a rapidly forming cell-encapsulated double network (DN) hydrogel is constructed by an ultrasonically activated silk fibroin network and bioorthogonal-mediated polyethylene glycol network. This DN hydrogel can be solidified within 10 s, and its mechanical property gradually increases to â¼20 kPa after 30 min. This work also demonstrates that coencapsulation of human umbilical vein endothelial cells (HUVECs) and umbilical cord-derived mesenchymal stem cells (UCMSCs) into the DN hydrogel can facilitate the formation of more mature vessels and complete the capillary network in comparison with the hydrogels encapsulated with a single cell type both in vitro and in vivo. Taking together, the DN hydrogel, combined with coencapsulation of HUVECs and UCMSCs, represents a strategy for the construction of a functional vascular network.
Assuntos
Fibroínas/química , Hidrogéis/química , Polietilenoglicóis/química , Materiais Biocompatíveis/química , Materiais Biocompatíveis/farmacologia , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Técnicas de Cocultura , Módulo de Elasticidade , Células Endoteliais da Veia Umbilical Humana , Humanos , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/metabolismo , Neovascularização Fisiológica/efeitos dos fármacos , Molécula-1 de Adesão Celular Endotelial a Plaquetas/genética , Molécula-1 de Adesão Celular Endotelial a Plaquetas/metabolismo , Cordão Umbilical/citologiaRESUMO
The isolation of specific and sensitive circulating tumor cells (CTCs) is significant for applying them in cancer diagnosis and monitoring. In this work, dual aptamer-modified poly(lactic-co-glycolic acid) (PLGA) nanofiber-based microfluidic devices were fabricated to achieve the highly efficient capture and specific release of epithelial and mesenchymal CTCs of ovarian cancer. Dual aptamer targeting epithelial cell adhesion molecules (EpCAM) and N-cadherin proteins to improve the capture sensitivity, bovine serum albumin (BSA) to guarantee the capture purity and the nanofibers to increase the capture efficiency via synchronously and effectively capturing the epithelial and mesenchymal CTCs with good capture specificity and sensitivity from blood samples were used. We used the target cells including the ovarian cancer A2780 cells (N-cadherin-high, EpCAM-low) and OVCAR-3 cells (EpCAM-high, N-cadherin-low) to test the devices, which exhibited good capture efficiency (91% for A2780 cells, 89% for OVCAR-3 cells), release efficiency (95% for A2780 cells, 88% for OVCAR-3 cells), and sensitivity for rare cells (92% for A2780 cells, 88% for OVCAR-3 cells). Finally, the clinical blood samples of ovarian cancer patients were detected by the PLGA nanofiber-based microfluidic device, and 1 to 13 CTCs were successfully confirmed to be captured with the help of immunofluorescence staining identification. The results exhibited that the dual aptamer-modified PLGA nanofiber-based microfluidic device used as a tool for CTC capture has the potential for clinical application to guide the diagnosis, treatment, and prognosis of ovarian cancer patients.
Assuntos
Separação Celular/instrumentação , Dispositivos Lab-On-A-Chip , Nanofibras , Células Neoplásicas Circulantes/patologia , Fenótipo , Copolímero de Ácido Poliláctico e Ácido Poliglicólico/química , Caderinas/metabolismo , Linhagem Celular Tumoral , Molécula de Adesão da Célula Epitelial/metabolismo , Humanos , Limite de Detecção , Células Neoplásicas Circulantes/metabolismoRESUMO
The detection and analysis of circulating tumor cells (CTCs) from cancer patients' blood samples present a powerful means to monitor cancer progression. In this work, an antifouling nanostructure substrate made of hydrogel nanoparticles was fabricated for an effective capture of CTCs from the blood samples. The hydrogel nanoparticles were synthesized by zwitterionic sulfobetaine methacrylate (SBMA), methacrylic acid (MAA) and N, N'-methylene bisacrylamide (MBA) through a simple polymerization. SBMA could provide an effective antifouling layer for the substrate to prevent nonspecific cell adhesion, MAA could afford active carboxyl groups for the immobilization of antibody to achieve specific CTC capture, and the nanostructured surface could improve the interaction of the target cells with the antibody modified substrate surface to enhance the capture efficiency of CTCs. Moreover, it was not necessary to further modify the antifouling molecules on the hydrogel nanoparticle substrate's surface, reducing the complexity and difficulty of the substrate preparation. The results showed that about 87 % of target cells (MCF-7 cells) were captured on the antibody modified hydrogel nanoparticle substrate. In contrast, the substrate showed little adhesive capacity for the nonspecific cells (K562 cells), and only 0.15 % of cells were captured. And 98 % of the captured cells kept good cell viability. Finally, 1-32 CTCs/mL were detected from the blood samples of five cancer patients, while no CTC was found in five healthy samples. It is envisaged that the new hydrogel nanostructure substrate is capable of capturing CTCs efficiently and specifically from patient blood samples to be used in cancer treatment.
Assuntos
Incrustação Biológica , Nanopartículas , Nanoestruturas , Células Neoplásicas Circulantes , Incrustação Biológica/prevenção & controle , Linhagem Celular Tumoral , Separação Celular , Humanos , Hidrogéis , Células MCF-7RESUMO
Four types of quadruplex supramolecules containing G-quadruplex (G4)-I-motif interfaces were assembled under slightly acidic conditions, which can interact with hemin to form I-motif-linked G4/hemin DNAzymes. Our data demonstrated that some I-motif-linked DNAzymes are highly acid-dependent due to the stabilization of hemiprotonated cytosine-cytosine (CËCH+) pairs for the G4 units.
Assuntos
Ácidos/química , DNA Catalítico/química , Quadruplex G , Hemina/química , Dicroísmo Circular , Citosina/química , DNA Catalítico/metabolismo , Corantes Fluorescentes/química , Concentração de Íons de Hidrogênio , Oligonucleotídeos/químicaRESUMO
Analysis of circulating tumor cells (CTCs) can provide significant clinical information for tumors, which has proven to be helpful for cancer diagnosis, prognosis monitoring, treatment efficacy, and personalized therapy. However, CTCs are an extremely rare cell population, which challenges the isolation of CTCs from patient blood. Over the last few decades, many strategies for CTC detection have been developed based on the physical and biological properties of CTCs. Among them, nanostructured interfaces have been widely applied as CTC detection platforms to overcome the current limitations associated with CTC capture. Furthermore, aptamers have attracted significant attention in the detection of CTCs due to their advantages, including good affinity, low cost, easy modification, excellent stability, and low immunogenicity. In addition, effective and nondestructive release of CTCs can be achieved by aptamer-mediated methods that are used under mild conditions. Herein, we review some progress in the detection and release of CTCs through aptamer-functionalized nanostructured interfaces.
Assuntos
Aptâmeros de Nucleotídeos/química , Nanoestruturas/química , Células Neoplásicas Circulantes/patologia , Humanos , Tamanho da Partícula , Propriedades de SuperfícieRESUMO
Circulating tumor cells (CTCs) are an important part of liquid biopsy as they represent a potentially rich source of information for cancer diagnosis, monitoring, prognosis, and treatment guidance. It has been proved that the nanotopography interaction between cells and the surface of CTC detection platforms can significantly improve the capture efficiency of CTCs, whereas many mature nanostructure substrates have been developed based on chemistry materials. In this work, a natural biointerface with unique biological properties is fabricated for efficient isolation and nondestructive release of CTCs from blood samples using the cancer cell membranes. The cell membrane interfaces are proved to have a good antiadhesion property for nonspecific cells because of their own electronegativity. A natural surface nanostructure is provided by the cancer cell membrane to nicely match with the surface nanotopography of CTCs. Bovine serum albumin (BSA) as a linker and DNA aptamer against the epithelial cell adhesion molecule (EpCAM) as a specific affinity molecule are then introduced onto the cell membrane interfaces to achieve the highly efficient and specific capture of CTCs. Finally, the captured target cells can be intactly released from the substrate using the complementary DNA sequence with controlling the incubation time. This study provides a smart strategy in the development of a natural biological interface for the isolation and release of CTCs with high purity.
Assuntos
Membrana Celular/metabolismo , Separação Celular/métodos , Células Neoplásicas Circulantes/metabolismo , Adulto , Idoso , Animais , Aptâmeros de Nucleotídeos/química , Neoplasias da Mama/sangue , Neoplasias da Mama/patologia , Bovinos , Linhagem Celular Tumoral , Membrana Celular/química , Molécula de Adesão da Célula Epitelial/química , Molécula de Adesão da Célula Epitelial/metabolismo , Feminino , Células HEK293 , Humanos , Pessoa de Meia-Idade , Soroalbumina Bovina/químicaRESUMO
Circulating tumor cells (CTCs) acquire mesenchymal markers (e.g., N-cadherin) and lose epithelial markers (e.g., epithelial cell adhesion molecule, EpCAM) during the epithelial-mesenchymal transition (EMT) and are therefore ideal biomarkers of tumor metastasis. However, it is still a challenge to efficiently capture and detect circulating tumor cells with different phenotypes simultaneously. In this work, to obtain aptamers targeting N-cadherin in the native conformation on live cells, we established stable N-cadherin overexpressing cells (N-cadherin cells) and used these cells to identify a panel of N-cadherin-specific aptamers through the cell-SELEX approach. Two aptamer candidates obtained after 12 rounds of selection showed a low equilibrium dissociation constant in the nanomolar range, indicating high binding affinity. The truncated aptamer candidate NC3S showed the highest binding affinity to N-cadherin cells with a low Kd value of 20.08 nM. The SYL3C aptamer was reported to target cancer cell surface biomarker EpCAM. Then, we synthesized two kinds of aptamer-modified magnetic nanoparticles (SYL3C-MNPs and NC3S-MNPs). Both SYL3C and NC3S aptamers possess excellent capture specificity and efficiency for the target cells. The aptamer-MNP cocktail exhibits a considerable capture efficiency and sensitivity for rare cancer cells of epithelial and mesenchymal phenotypes. Furthermore, no CTCs were found in blood samples from healthy donors, while CTCs were successfully isolated by using the aptamer-MNP cocktail for 15 out of 16 samples collected from patients. In summary, the two kinds of aptamer-modified MNPs could be utilized as a promising tool for capturing CTCs from clinical samples.