ANNA ZAMAY^1,3, GALINA ZAMAY¹, YURY GLAZYRIN¹, TATIANA ZAMAY¹, ALEXEY KRAT², ANDREY MODESTOV², OLGA ZUBKOVA,¹ EKATERINA SPIVAK,¹ MARIIA SUKHOVOLSKAIA¹, SVETLANA KUZNETSOVA³, ALLA SALMINA¹, ANA GARGAUN⁴, MAXIM V. BEREZOVSKI⁴, OLGA KOLOVSKAYA¹*
*Corresponding author
1. Research Institute of Molecular Medicine and Pathobiochemistry, Krasnoyarsk State Medical University, Krasnoyarsk, Russia
2. Krasnoyarsk Regional Clinical Cancer Center named after A.I. Kryzhanovsky, Krasnoyarsk, Russia
3. Institute of Chemistry and Chemical Technology, Siberian Branch Russian Academy of Sciences, Krasnoyarsk, Russia
4. Department of Chemistry, University of Ottawa, Ottawa, Ontario, Canada

Abstract

Aptamers are oligonucleotides that can bind with high affinity and specificity to their targets due to their unique three dimensional structure. Moreover, they are becoming increasingly attractive as therapeutic agents as they have demonstrated other virtues, such as antitumor effects. Here we present selection of DNA-aptamers specific to human lung adenocarcinoma tissues obtained immediately after surgery. Flow cytometry was used to compare relative affinities of aptamers to the cancer cells, and aptamers with the highest binding were selected, identified and then evaluated in a cell-based assay. Our findings showed that one of the aptamers has the potential to inhibit growth of primary cancer cell cultures without influencing growth of healthy lung cells. Following cell-based assays, we identified a protein target for this aptamer using high performance liquid chromatography and high resolution tandem mass spectrometry. Protein cathepsin D was found to be the most likely target for this aptamer; it is associated with cancer growth, invasion and metastasis, and more recently has been considered as a potential target for anticancer therapy. These results are promising and may lead the way toward the development of novel aptamer-based anticancer therapeutics.

INTRODUCTION

Aptamers are single-stranded DNA or RNA oligonucleotides selected from large random-sequence nucleic acid libraries for their selective affinity to different molecular targets (1-6). They fold into well-defined three-dimensional structures (1, 2) and show high affinity and specificity for their targets. Aptamers are often viewed as synthetic affinity probes, with applications ranging from target detection to drug discovery and delivery. These molecules are non-immunogenic and non-toxic; therefore, they have the potential to become good drug candidates (7). Furthermore, aptamers to various cancer cell lines have been found to exhibit anticancer activity themselves as well as in combination with toxins (8, 9). When interacting with molecular targets, aptamers can also change the functional state of a cell (10-12).

RESULTS AND DISCUSSION
Adenocarcinoma is one of the most abundant histological types of human lung cancer, which is why it was chosen as a target for the aptamer selection. Human cancer lung tissues were aseptically taken right after surgery and utilized for aptamer selection on the basis of cell-SELE ...