Aptamer‐mediated complement system killing of MCF‐7 and MEAR cancer cells

Abstract

Aptamer MUSC1-5TR-1 has high affinity for Mucin 1 (MUC1) which is over expressed in the adenocarcinoma cell line MCF-7. Aptamer TLS-11a recognizes BNL 1ME A.7R.1 (MEAR) hepatocarncinoma cells (Shangguan, D, et al., Anal Chem 80, no. 3: 721–728). Biotinylation of the aptamers' 5′ end allowed conjugation to streptavidin that was in turn conjugated to C1q, the first protein in the classical complement immune response pathway. Killing of MCF-7 and MEAR cells in vitro by complement-mediated lysis resulting from exposure to the biotinylated aptamers-streptavidin-C1q, in the presence of human complement protein serum, was visualized using SYTOX Green nucleic acid stain. SYTOX Green (Invitrogen) brightly labels nucleic acid in cells with compromised membrane integrity. MCF-7 and MEAR cells exposed to the full treatment both showed increased nuclear staining within a few hours, whereas cells not exposed to the aptamer, streptavidin-C1q conjugate, or complement protein serum showed little to no nuclear staining. Future experiments will incorporate FRET detection of membrane potential changes resulting from MAC formation, and will utilize immuno-fluorescence and immuno-gold to localize components of the MAC by laser scanning confocal and electron microscopy. Supported by funding from OTC Biotechnologies, LLC and NSF DBI-0821252 to JK.