Engineering a Facile Aptamer “Molecule-Doctor” with Hairpin-Contained I-Motif Enables Accurate Imaging and Killing of Cancer Cells

Abstract

Herein, we subtly engineered a pH and membrane receptor dual-activatable aptamer therapeutic for bispecific tumor cell imaging and in situ drug release by utilizing a hairpin-contained i-motif as the acid-responsive element to be complementary with a tumor-targeted aptamer, named as an aptamer “molecule-doctor” (pH-Apt-MD). Specifically, the pH-Apt-MD consisted of two DNA strands, where the Apt-sgc8c was labeled with AF488 and Cy3 at its 5′- and 3′-end, respectively. The I-strand, a hairpin-contained i-motif, was complementary to the Apt-sgc8c strand partially, labeled with a BHQ2 in the middle, thus generating Cy3 with quenched fluorescence and only AF488-emitted fluorescence. The double-helix region of pH-Apt-MD was designed rich in GC bases, providing sites for doxorubicin (Dox) intercalation. Once target cells were encountered, the pH-Apt-MD disassembled due to the specific recognition of the aptamer and conformation change of the i-motif, with activated fluorescence resonance energy transfer (FRET) signals between AF488 and Cy3, accompanied by Dox release in situ. Benefiting from the design of the hairpin-contained i-motif, the pH-Apt-MD presented a narrow pH response range (pH 6.0–6.8) with a transition midpoint (pHT) of 6.50 ± 0.04. Furthermore, living cell studies revealed that the stimuli-responsive FRET signal activation of pH-Apt-MD was successfully achieved on the HCT116 cell surface with ultralow background and enhanced imaging contrast. Then, the cytotoxicity experiments proved that accurate drug release and cell killing were realized to target cells in an acidic microenvironment. As a facile double stimuli-responsive strategy, the pH-Apt-MD may hold great promise for application in precise diagnosis and therapy of cancer cells.

Self-assembly of DNA nanostructure containing cell-specific aptamer as a precise drug delivery system for cancer therapy in non-small cell lung cancer

“…It is well known that DNA nanostructure can be loaded with anthracyclines owing to the rich of GC base pair [ 45 ]. DOX, a widely used anthracycline anticancer agent, can insert into double-stranded DNAs, inducing the quenching of drug fluorescence due to fluorescence resonance energy transfer (FRET) [ 46 – 48 ]. Thus, DOX loading and release of DNA nanostructure can be evaluated via the changes of fluorescence.…”

Framework Nucleic Acid-Based Multifunctional Tumor Theranostic Nanosystem for miRNA Fluorescence Imaging and Chemo/Gene Therapy

“…Afterward, 10 μL of CCK-8 reagent mixed with 90 μL of culture medium without serum was added and incubated for another 1 h. The absorbance ( A ) at 450 nm of the solution for each well was measured by a microplate reader. The relative cell viability (%) was calculated as follows: relative .25em cell .25em viability ( % ) = ( A sample − A blank ) / ( A control − A blank ) × 100 % …”

Engineered aptamers for molecular imaging

“…When pH-Apt-MD encountered a target cell, it underwent disassembly due to the specic recognition of the aptamer and a conformational change in the i-motif structure, which led to the activation of FRET for signal output. 86 Another approach to inducing conformational changes in aptamer sequences involves the incorporation of complementary sequences that bind to the aptamer region (Fig. 4C and D).…”