Modular DNA Tetrahedron Nanomachine-Guided Dual-Responsive Hybridization Chain Reactions for Discernible Bivariate Assay and Cell Imaging.

Abstract

Engineering of multivariate biosensing and imaging platforms involved in disease plays a vital role in effectively discerning cancer cells from normal cells and facilitating reliable targeted therapy. Multiple biomarkers such as mucin 1 (MUC1) and nucleolin are typically overexpressed in breast cancer cells compared to normal human breast epithelium cells. Motivated by this knowledge, a dual-responsive DNA tetrahedron nanomachine (drDT-NM) is constructed through immobilizing two recognition modules, MUC1 aptamer (MA) and a hairpin H1* encoding nucleolin-specific G-rich AS1411 aptamer, in two separate vertexes of a functional DT architecture tethering two localized pendants (PM and PN). When drDT-NM identifiably binds bivariate MUC1 and nucleolin, two independent hybridization chain reactions (HCRM and HCRN) as amplification modules are initiated with two sets of four functional hairpin reactants. Among them, one hairpin for HCRM is dually ended by fluorescein and quencher BHQ1 to sense MUC1. The responsiveness of nucleolin is executed by operating HCRN utilizing another two hairpins programmed with two pairs of AS1411 splits. In the shared HCRN duplex products, the parent AS1411 aptamers are cooperatively merged and folded into G-quadruplex concatemers to embed Zn-protoporphyrin IX (ZnPPIX/G4) for fluorescence signaling readout, thereby achieving a highly sensitive intracellular assay and discernible cell imaging. The tandem ZnPPIX/G4 unities also act as imaging agents and therapeutic cargos for efficient photodynamic therapy of cancer cells. Based on drDT-NM to guide bispecific HCR amplifiers for adaptive bivariate detection, we present a paradigm of exquisitely integrating modular DNA nanostructures with nonenzymatic nucleic acid amplification, thus creating a versatile biosensing platform as a promising candidate for accurate assay, discernible cell imaging, and targeted therapy.