A universal strategy designed for selecting bench-to-bedside aptamers to serum and validated in hepatocellular carcinoma diagnosis

Aptamers are single-stranded oligonucleotide ligands selected in vitro, with high specificity and affinity to their targets and great potential of diagnostic application. Serum is the most widely-used specimen in clinical diagnosis, but few aptamers to serum have been selected due to the serum complexity. Here we designed a universal strategy to generate serum aptamers for bench-to-bedside diagnosis and illustrated it in hepatocellular carcinoma (HCC). The strategy encompassed design of a special library, use of pooled serum as the selection target, elimination of oligonucleotides to normal serum molecules, enrichment of oligonucleotides specific to HCC serum, verification and validation of aptamer specificity, and diagnostic evaluation of aptamers in clinical setting. A total of 181 aptamers were selected, and all of them were specific to HCC to various extents. Seven aptamers chosen for specificity validation with serum specimens exhibited areas under the receiver operating characteristic curve (AUROCs) of 0.694-0.937 for HCC diagnosis, all except for one better than alpha-fetoprotein (AFP) (0.792). Two aptamers further evaluated in an enlarged serum sample showed AUROCs of 0.875 and 0.840, superior to AFP (0.766). The AUROC increased to 0.964 when the two aptamers combined with AFP, with sensitivity and specificity of over 90%. The aptamers alone and in combination yielded high true-positive rates (86.4%-90.9%) for AFP-negative HCC and high net benefits for clinical decision-making in HCC diagnosis. Conclusively, we developed and validated a universal method to select serum aptamers for bench-to-bedside diagnosis, and generated a group of serum aptamers valuable for HCC diagnosis and with translational potential.