title: Detection and knockdown of microRNA-34a using thioacetamido nucleic acid.
abstract: Thioacetamido nucleic acids (TANA) contain a backbone modification of dinucleotides consisting of a 5-atom amide linker N3'-COCH2-S-CH2 at thymidine or thymidine-cytidine dimer blocks. Here, the chemical synthesis of the TANA linked 5-methyl-cytidine-cytidine ((Me)cc) dimer block and its incorporation into the DNA sequence, complementary to human microRNA 34 (miR-34) is described. Further, for the first time, we demonstrate the biological applications of TANA modified oligonucleotides in detection and intracellular knockdown of a cancer related microRNA in comparison with DNA containing locked nucleic acid (LNA) and 2'-O-methyl modifications. The human microRNA miR-34 is a pro-apoptotic microRNA under the transcriptional control of protein 53 (p53). It gets expressed in response to DNA damage and regulates several cell cycle and apoptosis related targets. Here, we show that the TANA modified antisense oligonucleotide binds specifically to miR-34a, allowing its detection using primer extension. We also show that, using the TANA modified antisense oligonucleotide against miR-34a, intracellular levels of miR-34 can be reduced, and consequently, the expression of its target oncogene V-myc myelocytomatosis viral related oncogene, neuroblastoma derived (MYCN) is enhanced. Further, we assessed the toxicity and serum stability of the oligonucleotide to conclude that it is suitable for detection and modulation of the vital biomarker and tumor suppressor microRNA.
authors: Council of Scientific and Industrial Research (CSIR)-Institute of Genomics and Integrative Biology, Delhi, India.
authors: Kartik
authors: K
authors: Soni
authors: Shivani
authors: S
authors: Gupta
authors: Sachin S
authors: SS
authors: Gokhale
authors: Rakesh
authors: R
authors: Dey
authors: Anita D
authors: AD
authors: Gunjal
authors: Vaijayanti A
authors: VA
authors: Kumar
authors: Beena
authors: B
authors: Pillai
mesh_terms: D017209:Apoptosis / Q000235:genetics*; D045744:Cell Line, Tumor; D015972:Gene Expression Regulation, Neoplastic; D055785:Gene Knockdown Techniques; D016259:Genes, myc; D057809:HEK293 Cells; D006801:Humans; D035683:MicroRNAs / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D009841:Oligonucleotides / Q000235:genetics; D013936:Thymidine / Q000031:analogs & derivatives* / Q000138:chemical synthesis / Q000235:genetics; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism
chemical_list: C522425:MIRN34 microRNA, human; D035683:MicroRNAs; D009841:Oligonucleotides; D016159:Tumor Suppressor Protein p53; C477371:locked nucleic acid; C525861:thioacetamido nucleic acid analog; D013936:Thymidine