Fluorescent Probes for Peptide Nucleic Acid (PNA) based Diagnostics

Shankar Naik, Institute for Drug Research, School of Pharmacy, The Hebrew University of Jerusalem, Jerusalem, Israel
Eylon Yavin, Institute for Drug Research, School of Pharmacy, The Hebrew University of Jerusalem, Jerusalem, Israel

Fluorescent nucleobase surrogates provide nucleic acids with unique and interesting properties. The introduction of a fluorophore as a base surrogate into peptide nucleic acid (PNA) oligomers has been well documented. Such PNA probes signal hybridization by enhancement of fluorescence and also allow the measurements of local structure and dynamics of nucleic acids. The increased fluorescence of such a dye when bound to nucleic acids has been attributed to the restricted rotation of monomethine bond that connects the two aromatic systems. Previous studies in our lab has demonstrated the ability of thiazole orange (TO) embedded PNA-molecular beacons (MBs) in discriminating complementary sequence from mismatch mRNA sequences at a single base resolution in living cells. Realizing the potential of such fluorophores, a quest for molecules that have the ability to absorb light at a longer wavelengths relative to TO, began with the synthesis of acridine and indanone based fluorescent dyes. Acridine conjugated to benzothiazole derivative was introduced into the Fmoc-2-aminoethylglycine backbone yielding a PNA building block as a light-up probe. Similarly, indanone modified as its dicyano- and tetracyano derivatives acted as precursors for PNA monomer synthesis. These monomers were incorporated into the PNA sequence by Fmoc-solid phase peptide synthesis. The fluorescent properties of these probes with respect to single PNA strands and pairing DNA sequences are currently being studied and may pave the way to real-time in-vivo imaging of RNA biomarkers for a variety of disease-related indications.