Circular Prodrug Nucleic Acids for Stable Low-Immunogenic RNA Delivery
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Solution Overview
Problem
Existing antisense oligonucleotides face challenges with nuclease stability, inflammatory responses, and immune activation due to interactions with Pattern Recognition Receptors, limiting their therapeutic efficacy.
Innovation Solution
Development of circular prodrug nucleic acids (CPNs) with a functional domain and a circularizing domain, forming an intramolecular duplex that masks the 5′- and 3′-ends, reducing interactions with PRRs and exonucleases, and allowing conditional release of the functional domain within cells.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Stability of the object's composition
If 3′-end modifications (capping, hairpin loop) are applied to antisense oligonucleotides, then nuclease stability is improved, but inflammatory responses increase and therapeutic index is limited
Solution Approach 1:
The antisense oligonucleotide is divided into distinct functional segments: a 5′-end domain with specific modifications for stability, a central functional domain for target RNA binding, and a 3′-end domain with different modifications. This segmentation allows each region to be optimized independently for its specific function while avoiding the harmful effects of uniform modification.
Solution Approach 2:
Different modification chemistries are applied to different regions of the oligonucleotide. The 5′-end receives modifications optimized for nuclease protection and stability, while the 3′-end receives modifications optimized for reducing inflammatory responses. This local differentiation of properties resolves the contradiction between stability and reduced inflammation.
2Reliability
If DNA phosphorothioate antisense is used, then RNase H activation is improved, but immune activation increases
Solution Approach 1:
The oligonucleotide employs a hybrid structure where the 5′-end contains DNA phosphorothioate modifications that activate RNase H, while the 3′-end contains RNA or 2′-substituted RNA modifications that reduce immune activation. This local differentiation allows simultaneous achievement of reliable RNase H activation and reduced immune responses.
Solution Approach 2:
The antisense oligonucleotide is designed as a composite structure combining DNA and RNA elements with different properties. The DNA phosphorothioate portion provides RNase H activation capability, while the RNA portion provides reduced immune activation, creating a hybrid molecule that integrates the benefits of both chemistries.
3Strength
If RNA or 2′-substituted RNA antisense is used, then affinity binding to RNA is improved, but RNase H activation is lost
Solution Approach 1:
The oligonucleotide is segmented into a 5′-end domain with DNA phosphorothioate that provides RNase H activation, and a 3′-end domain with RNA or 2′-substituted RNA that provides high affinity binding. This segmentation allows each domain to specialize in its respective function.
Solution Approach 2:
A hybrid antisense structure combines DNA phosphorothioate elements (for RNase H activation) with RNA or 2′-substituted RNA elements (for high affinity binding), creating a composite molecule that achieves both RNase H activation and strong target binding simultaneously.
4Stability of the object's composition
If gapmer antisense with modified RNA segments at both ends is used, then nuclease stability is improved, but immune activation increases
Solution Approach 1:
Within the gapmer structure, the 5′-end modified RNA segment is optimized for nuclease stability, while the 3′-end modified RNA segment is optimized for reducing immune activation. This local quality differentiation allows the molecule to achieve both stability and reduced immunogenicity that cannot be obtained with uniform modifications.
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
CPNs enhance nuclease stability, reduce immune activation, and improve delivery to the cytoplasm or nucleus, minimizing off-target effects and enhancing therapeutic efficacy.
Implementation Method 1
the first nucleic acid molecule and the second nucleic acid molecule are complementary to each other and of opposite polarity to each other and hybridize to form a double-stranded section
Data Source
AI summary
The present invention provides oligonucleotides referred to as circular prodrug nucleic acid (“CPN”) as described herein, compositions comprising same, and methods of using same. This design of circular prodrug nucleic acids maintains a circular form until the circularizing domain is cleaved in situ by RNase H or Dicer or other intracellular factors.