FIG. 1.

Selection of CCKBR-binding DNA aptamers. (A) Peptide sequences of the two regions of the CCKBR N-terminus (amino acids 5–21 and 40–57) used for initial aptamer selection and the sequence of the corresponding regions from the CCKAR. Residue in bold show amino acids which differ between human and mouse CCKBR. (B) Dendrogram comparison of the DNA sequence for selected CCKBR aptamers. Aptamers (boxed) were identified for further individual characterization. (C) Proliferation of PANC-1 human pancreatic cancer cells in the presence of CCKBR-selected aptamers or human recombinant EGF, a proproliferative control. Light cross-hatched bars represent 72-h time points, and darker cross-hatched bars represent 96-h treatment. Bars represent the standard error of the mean of three experimental replicates. None of the selected aptamers significantly stimulated pancreatic cancer cell proliferation compared to vehicle-treated cells. Instead, several APs produced significant growth inhibition by 96 h. *P < 0.05, **P < 0.01 compared to control (1152 and 1153 were significant at P < 0.001). (D) Western blots of vehicle (V), gastrin-17 (G), or AP1153 (A)-treated PANC-1 cells demonstrated that although gastrin-16 stimulates Akt phosphorylation, AP1153 did not. Adjacent panel shows densitometric quantification of the blot normalized for β-actin. AP, aptamer; CCKAR, cholecystokinin A receptor; CCKBR, cholecystokinin B receptor; EGF, epidermal growth factor.