64Cu and fluorescein labeled anti-miRNA peptide nucleic acids for the detection of miRNA expression in living cells.

MiRNAs are single stranded RNAs of 18-22 nucleotides. They are promising diagnostic and prognostic markers for several pathologies including tumors, neurodegenerative, cardiovascular and autoimmune diseases. In the present work the development and characterization of anti-miRNA radiolabeled probes based on peptide nucleic acids (PNAs) for potential non-invasive molecular imaging in vivo of giant cell arteritis are described. MiR-146a and miR-146b-5p were selected as targets because they have been found up-regulated in this disease. Anti-miR and scramble PNAs were synthesized and linked to carboxyfluorescein or DOTA. DOTA-anti-miR PNAs were then labelled with copper-64 (⁶⁴Cu) to function as non-invasive molecular imaging tools. The affinity of the probes for the targets was assessed in vitro by circular dichroism and melting temperature. Differential uptake of fluorescein and ⁶⁴Cu labeled anti-miRNA probes was tested on BCPAP and A549 cell lines, expressing different levels of miR-146a and -146b-5p. The experiments showed that the anti-miR-146a PNAs were more effective than the anti-miR-146b-5p PNAs. Anti-miR-146a PNAs could bind both miR-146a and miR-146b-5p. The uptake of fluorescein and ⁶⁴Cu labeled anti-miR-146a PNAs was higher than that of the negative control scramble PNAs in miRNA expressing cells in vitro. ⁶⁴Cu-anti-miR-146a PNAs might be further investigated for non-invasive PET imaging of miR-146 overexpressing diseases. [ABSTRACT FROM AUTHOR]