title: Identification of an aptamer targeting hnRNP A1 by tissue slide-based SELEX.
abstract: We report a new in situ tissue slide-based SELEX strategy targeting neoplastic tissues from breast cancer patients. The methodology, using the molecular differences between clinical specimens, can evolve aptamers to all fractions of tissue. The aptamers may be used as new molecular probes for pathological diagnosis and tumour imaging, and also to reveal the molecular differences that are responsible for the diseases. The specific aptamers were enriched by unequal length strand PCR employing a structured (-) strand primer. After 12 rounds of selection, using the paraffin tissue sections from infiltrating ductal carcinomas as targets, and using the adjacent normal tissue from the same case as controls, one of the enriched ssDNA aptamers, BC15, was selected from a nucleic acid library and characterized as recognizing breast cancer cells either within the tissue sections or from the culture medium, but only weakly binding to adjacent normal cells or immortalized breast cell line MCF10A. The calculated equilibrium dissociation constants (K(d)) of BC15 bound to MCF7 cells was 111.0 +/- 36.9 nM. Through streptavidin magnetic beads mediated affinity purification assay followed by mass spectrometry identification and western blot confirmation, the target of BC15 was characterized to be hnRNP A1, which was further verified to be specifically and highly expressed in cancerous tissues of breast by hnRNP A1 antibody immunostaining as well as western blot. BC15 aptamer was also used to probe cancer cells in tissues from other pathological types of breast cancers including lobular carcinoma, ductal carcinoma complicated with lobular carcinoma, comedo carcinoma, and lymph node metastasis of breast ductal carcinoma origin or breast lobular carcinoma origin. Therefore, tissue slide-based SELEX holds promise in identifying tumour markers and developing specific molecular probes for cancer diagnosis.
authors.affiliation: Department of Biochemistry and Molecular Biology, Beijing Institute of Basic Medical Sciences, China.
authors.forename: Shaohua
authors.initials: S
authors.lastname: Li
authors.forename: Hua
authors.initials: H
authors.lastname: Xu
authors.forename: Hongmei
authors.initials: H
authors.lastname: Ding
authors.forename: Yanping
authors.initials: Y
authors.lastname: Huang
authors.forename: Xiaoxiao
authors.initials: X
authors.lastname: Cao
authors.forename: Guang
authors.initials: G
authors.lastname: Yang
authors.forename: Jie
authors.initials: J
authors.lastname: Li
authors.forename: Zhigang
authors.initials: Z
authors.lastname: Xie
authors.forename: Yuhong
authors.initials: Y
authors.lastname: Meng
authors.forename: Xiaobing
authors.initials: X
authors.lastname: Li
authors.forename: Qiang
authors.initials: Q
authors.lastname: Zhao
authors.forename: Beifen
authors.initials: B
authors.lastname: Shen
authors.forename: Ningsheng
authors.initials: N
authors.lastname: Shao
mesh_terms: D000595:Amino Acid Sequence; D052157:Aptamers, Nucleotide / Q000235:genetics*; D014408:Biomarkers, Tumor / Q000235:genetics* / Q000378:metabolism; D001706:Biopsy; D001943:Breast Neoplasms / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D018270:Carcinoma, Ductal, Breast / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D004273:DNA, Neoplasm / Q000235:genetics; D005260:Female; D015723:Gene Library; D000076245:Heterogeneous Nuclear Ribonucleoprotein A1; D034481:Heterogeneous-Nuclear Ribonucleoprotein Group A-B / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D018613:Microscopy, Confocal; D008969:Molecular Sequence Data; D009363:Neoplasm Proteins / Q000235:genetics / Q000378:metabolism; D052156:SELEX Aptamer Technique / Q000379:methods; D014407:Tumor Cells, Cultured
chemical_list: D052157:Aptamers, Nucleotide; D014408:Biomarkers, Tumor; D004273:DNA, Neoplasm; D000076245:Heterogeneous Nuclear Ribonucleoprotein A1; D034481:Heterogeneous-Nuclear Ribonucleoprotein Group A-B; D009363:Neoplasm Proteins