The World Health Organization has declared the outbreak of a novel coronavirus (SARS-CoV-2 or 2019-nCoV) as a global pandemic. However, the mechanisms behind the coronavirus infection are not yet ...fully understood, nor are there any targeted treatments or vaccines. In this study, we identified high-binding-affinity aptamers targeting SARS-CoV-2 RBD, using an ACE2 competition-based aptamer selection strategy and a machine learning screening algorithm. The K d values of the optimized CoV2-RBD-1C and CoV2-RBD-4C aptamers against RBD were 5.8 nM and 19.9 nM, respectively. Simulated interaction modeling, along with competitive experiments, suggests that two aptamers may have partially identical binding sites at ACE2 on SARS-CoV-2 RBD. These aptamers present an opportunity for generating new probes for recognition of SARS-CoV-2 and could provide assistance in the diagnosis and treatment of SARS-CoV-2 while providing a new tool for in-depth study of the mechanisms behind the coronavirus infection.
Circulating tumor cell (CTC)‐enrichment by using aptamers has a number of advantages, but the issue of compromised binding affinities and stabilities in real samples hinders its wide applications. ...Inspired by the high efficiency of the prey mechanism of the octopus, we engineered a deterministic lateral displacement (DLD)‐patterned microfluidic chip modified with multivalent aptamer‐functionalized nanospheres (AP‐Octopus‐Chip) to enhance capture efficiency. The multivalent aptamer–antigen binding efficiency improves 100‐fold and the capture efficiency is enhanced more than 300 % compared with a monovalent aptamer‐modified chip. Moreover, the captured cancer cells can be released through a thiol exchange reaction with up to 80 % efficiency and 96 % viability, which is fully compatible with downstream mutation detection and CTC culture. Using the chip, we were able to find CTCs in all cancer samples analyzed.
Octopus chip cell capture: Inspired by the high efficiency of the prey mechanism of the octopus, an aptamer‐tailed octopus chip (AP‐Octopus‐Chip) for CTC enrichment was developed. The design of the chip and the high binding affinity of the multivalent structures against the target cells significantly improved the CTC capture efficiency and enrichment. The enriched cancer cells can be released through a thiol exchange reaction, which is fully compatible with downstream mutation detection and CTC culture.
Tumor‐derived exosomal proteins have emerged as promising biomarkers for cancer diagnosis, but the quantitation accuracy is hindered by large numbers of normal cell‐derived exosomes. Herein, we ...developed a dual‐target‐specific aptamer recognition activated in situ connection system on exosome membrane combined with droplet digital PCR (ddPCR) (TRACER) for quantitation of tumor‐derived exosomal PD‐L1 (Exo‐PD‐L1). Leveraging the high binding affinity of aptamers, excellent selectivity of dual‐aptamer recognition, and the high sensitivity of ddPCR, this method exhibits significant sensitivity and selectivity for tracing tumor‐derived Exo‐PD‐L1 in a wash‐free manner. Due to the excellent sensitivity, the level of tumor‐derived Exo‐PD‐L1 detected by TRACER can distinguish cancer patients from healthy donors, and for the first time was identified as a more reliable tumor diagnostic marker than total Exo‐PD‐L1. The TRACER strategy holds great potential for converting exosomes into reliable clinical indicators and exploring the biological functions of exosomes.
We developed a dual‐target‐specific aptamer recognition system combined with droplet digital PCR for precise quantitative analysis of exosomal PD‐L1 (Exo‐PD‐L1). This method can distinguish tumor‐derived from non‐tumor‐derived Exo‐PD‐L1, holding great potential for the analysis of exosome subtypes and offering unprecedented opportunities for the study of the biological functions of exosomes and their conversion into reliable clinical indicators.
Immunotherapy has revolutionized cancer treatment, but its efficacy is severely hindered by the lack of effective predictors. Herein, we developed a homogeneous, low‐volume, efficient, and sensitive ...exosomal programmed death‐ligand 1 (PD‐L1, a type of transmembrane protein) quantitation method for cancer diagnosis and immunotherapy response prediction (HOLMES‐ExoPD‐L1). The method combines a newly evolved aptamer that efficiently binds to PD‐L1 with less hindrance by antigen glycosylation than antibody, and homogeneous thermophoresis with a rapid binding kinetic. As a result, HOLMES‐ExoPD‐L1 is higher in sensitivity, more rapid in reaction time, and easier to operate than existing enzyme‐linked immunosorbent assay (ELISA)‐based methods. As a consequence of an outstanding improvement of sensitivity, the level of circulating exosomal PD‐L1 detected by HOLMES‐ExoPD‐L1 can effectively distinguish cancer patients from healthy volunteers, and for the first time was found to correlate positively with the metastasis of adenocarcinoma. Overall, HOLMES‐ExoPD‐L1 brings a fresh approach to exosomal PD‐L1 quantitation, offering unprecedented potential for early cancer diagnosis and immunotherapy response prediction.
An aptamer‐induced thermophoresis quantitation of exosomal programmed death‐ligand 1 (PD‐L1, a transmembrane protein) was developed, which integrates effective recognition of aptamer and homogeneous thermophoresis. The facile technique is more sensitive and efficient than the current enzyme‐linked immunosorbent assay (ELISA)‐based methods. Translation of the method into standard clinical practice for immunotherapy prediction and monitoring is anticipated.
The COVID‐19 pandemic caused by SARS‐CoV‐2 is threating global health. Inhibiting interaction of the receptor‐binding domain of SARS‐CoV‐2 S protein (SRBD) and human ACE2 receptor is a promising ...treatment strategy. However, SARS‐CoV‐2 neutralizing antibodies are compromised by their risk of antibody‐dependent enhancement (ADE) and unfavorably large size for intranasal delivery. To avoid these limitations, we demonstrated an aptamer blocking strategy by engineering aptamers’ binding to the region on SRBD that directly mediates ACE2 receptor engagement, leading to block SARS‐CoV‐2 infection. With aptamer selection against SRBD and molecular docking, aptamer CoV2‐6 was identified and applied to prevent, compete with, and substitute ACE2 from binding to SRBD. CoV2‐6 was further shortened and engineered as a circular bivalent aptamer CoV2‐6C3 (cb‐CoV2‐6C3) to improve the stability, affinity, and inhibition efficacy. cb‐CoV2‐6C3 is stable in serum for more than 12 h and can be stored at room temperature for more than 14 days. Furthermore, cb‐CoV2‐6C3 binds to SRBD with high affinity (Kd=0.13 nM) and blocks authentic SARS‐CoV‐2 virus with an IC50 of 0.42 nM.
We propose an aptamer blocking strategy to inhibit SARS‐CoV‐2 infection. With the advantages of small size, rapid kinetics, high stability, sophisticated programmability and high security, our aptamers have great potential as prophylactic and therapeutic agents, which could greatly assist in the intervention of prevailing and emerging infectious diseases other than COVID‐19.
The gamma coronavirus infectious bronchitis virus (IBV) is known to cause an acute and highly contagious infectious disease in poultry. Here, this study aimed to investigate the impact of virulent or ...avirulent IBV infection on the avian host by conducting proteomics with data-independent acquisition mass spectrometry (DIA-MS) in the kidneys of IBV-infected chickens. The results revealed 267, 489, and 510 differentially expressed proteins (DEPs) in the chicken kidneys at 3, 5, and 7 days postinfection (dpi), respectively, when infected with the GD17/04 strain, which is a highly nephrogenic strain and belongs to the 4/91 genotype. In contrast, the attenuated 4/91 vaccine resulted in the identification of 144, 175, and 258 DEPs at 3, 5, and 7 dpi, respectively. Functional enrichment analyses indicated distinct expression profiles between the 2 IBV strains. Upon GD17/04 infection, metabolic pathways respond initially in the early stage (3 dpi) and immune-related signaling pathways respond in the middle and late stages (5 and 7 dpi). The 4/91 vaccine elicited a completely opposite response compared to the GD17/04 infection. Among all DEPs, 62 immune-related DEPs were focused on and found to be mainly enriched in the type I interferon (IFN-I) signaling pathway and involved in humoral and cellular immunity. Notably, key molecules in the IFN-I signaling pathway including MDA5, LGP2, and TBK1 may serve as regulatory targets of IBV. Overall, this study highlights similarities and discrepancies in the patterns of protein expression at different stages of infection with virulent and avirulent IBV strains, with the IFN-I signaling pathway emerging as a critical response to IBV infection.
The analysis of circulating tumor cells (CTCs) holds great significance for cancer diagnosis, prognosis, and personalized therapy. However, the rarity, vulnerability and heterogeneity of CTCs bring ...daunting technical challenges to their isolation, release and analysis. Recent exciting advances in microfluidics have greatly promoted CTC isolation and analysis due to its merits of precise control of fluid behavior, integration, and automation. Especially, aptamer-based microfluidic chip is considered as promising platform, because aptamers as recognition ligands have inherent superiority in CTC isolation and release for their convenient modification and controllable recognition ability. This review focuses on recent progresses in aptamer-based microfluidics for isolation, release and analysis of CTCs. First, existing CTC-related aptamers and their selection methods are briefly introduced. Strategies for conjugating aptamers onto microfluidic chips are also reviewed. Then, aptamer-based microfluidic chips for CTC isolation, release and analysis are summarized. Finally, future research directions and challenges in this field are discussed.
•CTC analysis is important for cancer diagnosis, prognosis, and personalized therapy.•Microfluidic chip is emerged as powerful CTC analysis platform.•Aptamers are considered as ideal recognition tools for CTC isolation and release.•Aptamer-based microfluidics for CTC isolation, release, and analysis is summarized.
In this work, we report a reducing agent-free strategy for the synthesis of Fe3O4 nanoparticle/nitrogen-doped graphene quantum dot (Fe3O4/N-GQD) hybrids, and constructed a sensing platform based on ...Fe3O4/N-GQDs for the visual discrimination of phenylenediamine isomers. Fe3O4/N-GQDs were facilely prepared by hydrothermal treatment of Fe(3+)/N-GQD solutions under alkaline conditions without other reagents. The prepared Fe3O4/N-GQDs exhibited outstanding peroxidase-like activity and were stable under a wide range of pH values and temperatures. The phenylenediamine isomers (o-phenylenediamine, m-phenylenediamine, and p-phenylenediamine) were discriminated through the H2O2-mediated oxidation reaction using Fe3O4/N-GQDs as novel peroxidase mimics, which resulted in appreciable color changes. The proposed method is simple, economical, and effective for discrimination of isomers, and can be used for sensitive and selective quantitative analysis of o-phenylenediamine and p-phenylenediamine. A good linear relationship from 1 to 90 μM and a detection limit of 230 nM for o-phenylenediamine were achieved, and the linear relationship for p-phenylenediamine was from 2 to 70 μM with a detection limit of 530 nM. The proposed method may open new applications of Fe3O4/N-GQDs in biomedicine and environmental chemistry.
Molecular recognition ligands are of great significance in many fields, but our ability to develop new recognition molecules remains to be expanded. Here, we developed a Sequential Multidimensional ...Analysis algoRiThm for aptamer discovery (SMART-Aptamer) from high-throughput sequencing (HTS) data of SELEX libraries based on multilevel structure analysis and unsupervised machine learning to discover nucleic acid recognition ligands with high accuracy and efficiency. We validated SMART-Aptamer with three sets of HTS data from screening pools against hESCs, EpCAM, and CSV. High affinity aptamers for all three targets were successfully obtained, and the results revealed that SMART-Aptamer is able to pick out high affinity aptamers with low false positive and negative rates. With the advantages of accuracy, efficiency, and robustness, SMART-Aptamer represents a paradigm-shift strategy for the discovery of binding ligands for a variety of biomedical applications.
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