Shangguan Dihua, Cao Zehui Charles, Li Ying, Tan Weihong
Department of Chemistry, University of Florida, Gainesville, FL 32611, USA.
Clin Chem. 2007 Jun;53(6):1153-5. doi: 10.1373/clinchem.2006.083246. Epub 2007 Apr 26.
Molecular-level differentiation of neoplastic cells is essential for accurate and early diagnosis, but effective molecular probes for molecular analysis and profiling of neoplastic cells are not yet available. We recently developed a cell-based SELEX (systematic evolution of ligands by exponential enrichment) strategy to generate aptamers (designer DNA/RNA probes) as molecular probes to recognize neoplastic cells.
We tested 6 cell-SELEX-generated aptamers with equilibrium dissociation constants in the nanomolar to subnanomolar range: sgd5, selected from Toledo cells, a human diffuse large-cell lymphoma cell line (B-cell), and sgc8, sgc3, sgc4, sgd2, and sgd3 from CCRF-CEM cells, a human precursor T cell acute lymphoblastic leukemia (T-ALL) cell line. Aptamers were labeled with fluorescein isothiocyanate fluorophores and then used to recognize, by flow cytometric analysis, neoplastic cells in cultured hematopoietic cell lines and clinical samples.
Aptamer sgd5 recognized only its target cells. Aptamers sgc3, sgd2, sgd3, sgc4, and sgc8, selected from a T-cell leukemia cell line, identified all of the cultured T-cell leukemia cell lines with relatively high fluorescence intensity. Aptamers sgc8, sgc3, and sgd3 showed good selectivity toward T-ALL cells and almost no binding to normal hematopoietic cells or lymphoma and myeloma cells. Selected aptamers also detected targets on the cell membranes of neoplastic cells in patient samples.
Aptamers selected against cultured neoplastic cells can effectively be used as molecular probes for recognition of neoplastic cells in patient samples. Cell-based aptamer selection can be used to generate aptamer probes to obtain molecular signatures of neoplastic cells in patient samples.
肿瘤细胞的分子水平分化对于准确和早期诊断至关重要，但目前尚无用于肿瘤细胞分子分析和谱分析的有效分子探针。我们最近开发了一种基于细胞的SELEX（指数富集配体系统进化）策略，以生成适体（定制的DNA/RNA探针）作为识别肿瘤细胞的分子探针。
我们测试了6种通过细胞SELEX生成的适体，其平衡解离常数在纳摩尔至亚纳摩尔范围内：sgd5，从托莱多细胞（一种人弥漫性大细胞淋巴瘤细胞系（B细胞））中筛选得到；sgc8、sgc3、sgc4、sgd2和sgd3，从CCRF-CEM细胞（一种人T细胞前体急性淋巴细胞白血病（T-ALL）细胞系）中筛选得到。适体用异硫氰酸荧光素荧光团标记，然后通过流式细胞术分析用于识别培养的造血细胞系和临床样本中的肿瘤细胞。
适体sgd5仅识别其靶细胞。从T细胞白血病细胞系中筛选得到的适体sgc3、sgd2、sgd3、sgc4和sgc8，以相对较高的荧光强度识别所有培养的T细胞白血病细胞系。适体sgc8、sgc3和sgd3对T-ALL细胞表现出良好的选择性，几乎不与正常造血细胞、淋巴瘤细胞和骨髓瘤细胞结合。所选适体还检测到患者样本中肿瘤细胞膜上的靶标。
针对培养的肿瘤细胞筛选得到的适体可有效用作识别患者样本中肿瘤细胞的分子探针。基于细胞的适体筛选可用于生成适体探针，以获得患者样本中肿瘤细胞的分子特征。
