Next generation APOBEC3 inhibitors: Optimally designed for potency and nuclease stability

Area of Science:

• Biochemistry
• Molecular Biology
• Drug Discovery

Background:

• APOBEC3 (A3) enzymes contribute to viral and cancer heterogeneity, often causing drug resistance.
• Current inhibition strategies using phosphodiester (PO) DNA-based inhibitors face limitations in stability and potency.

Purpose of the Study:

• To enhance the potency and nuclease stability of 2'-deoxy-zebularine (dZ) oligonucleotide inhibitors against A3A and A3G enzymes.
• To develop novel therapeutic strategies for inhibiting A3 enzymes.

Main Methods:

• Modification of oligonucleotide backbones with phosphorothioate (PS) linkages and 2'-fluoro sugar modifications.
• Design of mixed PO/PS backbones and hairpin-structured inhibitors with LNA sugar modifications.
• Assessment of inhibitor potency and nuclease resistance through enzymatic assays and serum stability tests.

Main Results:

• Mixed PO/PS backbones significantly enhanced inhibitor potency (2.3-9.2 fold) and nuclease resistance.
• A novel nanomolar inhibitor for A3G-CTD2 was developed using 2'-fluoro modifications.
• Single-digit nanomolar inhibitors targeting A3A were achieved with hairpin structures, optimized PS patterns, and LNA modifications, demonstrating high serum stability.

Conclusions:

• Optimized oligonucleotide inhibitors exhibit superior potency and stability compared to previous approaches.
• These findings present a promising therapeutic avenue for A3 enzyme inhibition in viral infections and cancer treatment.