Backbone Hydrocarbon-Constrained Nucleic Acids Modulate Hybridization Kinetics for RNA

Area of Science:

• Medicinal Chemistry
• Oligonucleotide Therapeutics
• Molecular Biophysics

Background:

• Therapeutic oligonucleotide (ON) binding affinity relies on association (ka) and dissociation (kd) rates.
• Single-stranded ONs exhibit flexibility, potentially hindering hybridization with target RNA.
• Modulating ON backbone conformation is crucial for optimizing hybridization kinetics.

Purpose of the Study:

• To investigate if restricting rotation around the sugar-phosphate backbone of ONs can modulate their hybridization kinetics for complementary RNA.
• To employ molecular dynamic simulations to optimize hydrocarbon bridge design for backbone-constrained ON analogues.
• To synthesize and evaluate modified ONs with backbone constraints to improve hybridization properties.

Main Methods:

• Utilized molecular dynamic simulations to guide the design of hydrocarbon bridges for backbone constraint.
• Synthesized backbone-constrained nucleotide trimers using ring-closing metathesis and incorporated them into oligonucleotides.
• Incorporated phosphoramidites via in situ synthesis and coupling to solid supports for oligonucleotide assembly.

Main Results:

• 15-membered macrocyclic-constrained ON analogues exhibited similar/improved on-rates but significantly increased off-rates, reducing duplex stability compared to DNA ONs.
• Locked nucleic acid (LNA) ONs demonstrated similar on-rates to DNA ONs but very slow off-rates, enhancing duplex stability.
• Experimental data generally supported molecular dynamics simulation predictions for constrained backbone ON analogues.

Conclusions:

• Backbone constraint strategies can modulate oligonucleotide hybridization kinetics.
• Molecular dynamics simulations serve as a predictive tool for designing next-generation constrained backbone ON analogues.
• Optimized backbone constraints, like those in LNA, can lead to improved duplex stability and therapeutic potential.