Aptamer-based approaches to detect nucleolin in prostate cancer

Abstract

We have investigated the expression of nucleolin (NCL) in liquid biopsies of prostate cancer (PCa) patients and healthy controls to determine its correlation with tumor prognosis. To detect NCL we used a modified AS1411 aptamer designated by AS1411-N5. In presence of NCL, AS1411-N5 increases the fluorescence by assuming a G-quadruplex (G4) structure, while in the absence of NCL the fluorescence signal remains quenched. The structural characterization of AS1411-N5 was performed by biophysical studies, which demonstrated the formation of G4 parallel conformation in the presence of 100 mM K⁺ and the ability to recognize NCL with high affinity (K_D = 138.1 ± 5.5 nM). Furthermore, the clinical relevance of NCL in PCa liquid biopsies was assessed by using an NCL-based ELISA assay. The protein was measured in the peripheral blood mononuclear cells (PBMCs) cell lysate of 158 individuals, including PCa patients and healthy individuals. The results depicted a remarkable increase of NCL levels in the PBMC's lysate of PCa patients (mean of 626.1 pg/mL whole blood) when compared to healthy individuals (mean of 198.5 pg/mL whole blood). The ELISA results also provided evidence for the usefulness of determining NCL levels in advanced PCa stages. Furthermore, a microfluidic assay showed the ability of AS1411-N5 in recognizing NCL in spiked human plasma samples.

Keywords: AS1411-N5; G-quadruplex; Microfluidic; Nucleolin; Prostate cancer.