Rapid Complexation of Aptamers by Their Specific Antidotes

Area of Science:

• Biochemistry
• Molecular Biology
• Pharmacology

Background:

• Aptamers are nucleic acid ligands with high specificity and affinity for various targets.
• Aptamers offer advantages like chemical modifiability for bioavailability and antidote (AD)-mediated controllability.
• Controllability is crucial for safe and effective in vivo applications of aptamers.

Purpose of the Study:

• To determine the AD-mediated complexation and neutralization of two aptamers: NU172 (thrombin binding) and R10-60 (TLR9 binding).
• To analyze the kinetics of aptamer/AD complex formation in human serum at 37°C.
• To evaluate the blockade of aptamer functions by ADs in relevant biological environments.

Main Methods:

• Gel electrophoresis was used to analyze the time course of aptamer/AD complex formation in human serum.
• Activated clotting time (ACT) assay measured the neutralization of NU172's anticoagulant activity in whole blood.
• Gene expression analysis (IFN-1β, IL-6, CXCL-10, IL-1β) assessed the inhibition of R10-60's TLR9-mediated immune activation in PMDC05 cells.

Main Results:

• Rapid complexation of both NU172 and R10-60 aptamers by their respective ADs was observed within 2 minutes in human serum.
• Anticoagulant activity of NU172 was neutralized within 5 minutes of AD addition in fresh human whole blood.
• TLR9-mediated activation of PMDC05 cells by R10-60 was effectively interrupted upon addition of the R10-60 AD.

Conclusions:

• Nucleic acid aptamers demonstrate rapid antagonizability by complementary ADs across diverse biological matrices (serum, whole blood, cell culture media).
• ADs provide a rapid and effective mechanism for controlling aptamer activity in vitro and potentially in vivo.
• Nucleic acid ADs are promising tools for applications including aptamer-based drug antagonism, immobilization, and targeted delivery of oligonucleotides.