幽门螺杆菌ssDNA适配子的筛选及其应用

目的通过富集的配基系统进化技术(SELEX)筛选出与幽门螺杆菌(HP)菌体具有高亲和性适配子,并且利用其检测组织和活体内HP的感染。方法培养HP细菌,利用SELEX筛选获得HP菌体的适配子。在HP感染患者活检组织验证其结合。结果从第6轮筛选所得ssDNA中筛选出一条高结合力的适配子命名为HP1,将适配子HP1区别于其他筛选出的适配子,并检测其与幽门螺旋杆菌的亲和力。结论初步获得了针对HP菌体筛选出高亲和性适配子,为开发HP诊断试剂奠定了基础。     

新型HER2适配体的筛选及鉴定

目的研发能特异性结合人表皮生长因子受体2(HER2)的DNA适配体,为开发针对HER2的新型肿瘤靶向诊疗技术提供依据。方法体外合成全长86个碱基并含有40个随机寡核苷酸的单链DNA文库,以HER2表位肽为靶标,利用指数富集的配体系统进化技术(SELEX),从单链DNA文库中筛选能够选择性结合HER2多肽的核酸适配体;流式细胞术检测富集进度、适配体与HER2蛋白及HER2阳性细胞的结合特性;MFold软件预测二级结构。结果经多轮筛选获得了能够识别HER2多肽的DNA适配体HA5,其能够选择性地结合HER2蛋白及HER2阳性的乳腺癌细胞,而不结合胰蛋白酶和HER2阴性细胞。结论 DNA适配体HA5能选择性地识别HER2阳性的乳腺癌细胞,在研发针对HER2的新型肿瘤靶向诊疗技术方面具有应用潜能。     

Metastasis‐focused cell‐based SELEX generates aptamers inhibiting cell migration and invasion

Metastasis, the capacity of tumour cells to disseminate and grow at distant sites, is the main factor in cancer mortality. Compounds inhibiting migration and invasion of cancer cells are promising candidates for anticancer therapy strategies. We have generated nuclease‐resistant RNA ligands (aptamers) recognizing highly metastatic cells with high affinity and specificity, and inhibiting their migratory and invasive potentials. Aptamers were generated by a cell‐based subtractive SELEX technology using isogenic cell lines with similar tumorigenic potentials but opposite metastatic aggressiveness. Two aptamers, E37 and E10, bound specifically to the metastatically aggressive cell line and altered the phosphorylation of several tyrosine kinases. Fluorescent microscopy showed intracellular uptake of E37, in contrast to membrane binding of E10. Both aptamers inhibited migration of tumour cells in culture (50 and 85% inhibition with respect to control pool for E10 and E37, respectively) while only E10 inhibited cell invasion (?75% with respect to control pool). This proof‐of‐concept study demonstrates the potential of cell‐based SELEX to yield ligands that selectively recognize aggressive metastatic cells and inhibit phenotypes linked to metastatic potential.     

ATP-Responsive Drug Delivery Systems

The combination of targeted drug delivery and controlled-release technology may pave the road for more effective yet safer chemotherapeutic options for cancer therapy. Drug-encapsulated polymeric nanoparticle–aptamer bioconjugates represent an emerging technology that can facilitate the delivery of chemotherapeutics to primary and metastatic tumours. Aptamers are short nucleic acid molecules with binding properties and biochemical characteristics that may make them suitable for use as targeting molecules. The goal of this review is to summarise the key components that are required for creating effective cancer targeting nanoparticle–aptamer bioconjugates. The field of controlled release and the structure and properties of aptamers, as well as the criteria for constructing effective conjugates, will be discussed.     

B淋巴细胞活化因子特异的单链DNA适体的人工进化

目的: 进化筛选与B淋巴细胞活化因子（B cell activating factor belonging to the TNF family，BAFF）特异结合的单链DNA适体，鉴定其结合力。方法:应用指数方式富集的配体系统进化（systematic evolution of ligands by exponential enrichment, SELEX）技术，将随机序列寡核     

SELEX技术及其应用的研究进展

SELEX技术近年来不断完善与发展，应用领域不断扩宽，筛选的适配子不断优化，使该技术在疾病的诊断、治疗及药物的开发等方面更具潜在价值。     

Aptamers in Diagnostics and Treatment of Viral Infections

Aptamers are in vitro selected DNA or RNA molecules that are capable of binding a wide range of nucleic and non-nucleic acid molecules with high affinity and specificity. They have been conducted through the process known as SELEX (Systematic Evolution of Ligands by Exponential Enrichment). It serves to reach specificity and considerable affinity to target molecules, including those of viral origin, both proteins and nucleic acids. Properties of aptamers allow detecting virus infected cells or viruses themselves and make them competitive to monoclonal antibodies. Specific aptamers can be used to interfere in each stage of the viral replication cycle and also inhibit its penetration into cells. Many current studies have reported possible application of aptamers as a treatment or diagnostic tool in viral infections, e.g., HIV (Human Immunodeficiency Virus), HBV (Hepatitis B Virus), HCV (Hepatitis C Virus), SARS (Severe Acute Respiratory Syndrome), H5N1 avian influenza and recently spread Ebola. This review presents current developments of using aptamers in the diagnostics and treatment of viral diseases.     

针对于白蛋白的新型核酸适配体的筛选与鉴定

目的研发能特异结合白蛋白的DNA核酸适配体,为去除实验样品中的白蛋白提供新型技术手段。方法体外合成全长为59个碱基并含有21个随机寡核苷酸的单链DNA文库,以白蛋白为靶标,利用指数富集的配体系统进化技术(SELEX),从单链DNA文库中筛选能特异结合白蛋白的核酸适配体;用流式细胞计量术来检测核酸适配体的富集程度、适配体与白蛋白的结合特性;用MFold软件预测其二级结构。结果经过多轮筛选得到了能特异识别白蛋白的DNA核酸适配体A6,其Kd值为77.4 nmol/L,且基本不结合卵清蛋白、免疫球蛋白Ig G等对照蛋白。结论针对于白蛋白的DNA核酸适配体A6能选择性识别白蛋白,在去除实验样品中的白蛋白方面具有应用潜能。