In silico Selection Approach to Develop DNA Aptamers for a Stem–like Cell Subpopulation of Non-Small Lung Cancer Adenocarcinoma Cell Line A549
Background. Detection of circulating lung cancer cells with cancer-stem like characteristics would represent an improved tool for disease prognosis. However, current antibodies based methods have some disadvantages and therefore cell SELEX (Systematic Evolution of Ligands by Exponential Enrichment) was used to develop DNA aptamers, recognizing cell surface markers of non-small lung cancer (NSLC) cells.
Materials and methods. The human adenocarcinoma cell line A549 was used for selection in seven cell SELEX cycles. We used human blood leukocytes as negative selection, and lung cancer membrane lung stem cell protein marker CD90 antibody A549 cell binding as positive selection step.
Results. The obtained oligonucleotide sequences after the seventh SELEX cycle were subjected to in silico selection analysis based on three independent types of bioinformatics approaches, selecting two closely related aptamer candidates in terms of consensus sequences, structural motifs, affinity (Kd) and binding stability (∆G). We selected and identified the aptamer A155_18 with very good binding characteristics to A459 cells slected for CD90 antibody binding. The calculated phylogenetic tree showed that aptamers A155_18 and the known A549 cell aptamer S6 have a close structural relationship. MEME sequence analysis showed that they share two unique motifs, not present in other sequences.Conclusions. The novel aptamer A155_18 has strong binding affinity for A549 lung cancer cell line subpopulation that is expressing stem cell marker CD90, indicating a possible stemness characteristics of A459 line, or a subpopulation present within this cell line. This aptamer can be applied as diagnostic tool, identifying NSLC circulating cells.