Hybrid Antisense Oligonucleotide Delivery via Protein Binding
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Solution Overview
Problem
Current methods for delivering antisense oligonucleotides into cells are limited by their inability to penetrate cell membranes, leading to low bioavailability and ineffective cytosolic access, with existing approaches being unsafe or unreliable.
Innovation Solution
The development of hybrid antisense oligonucleotides (ASO hybrids) that form a double-stranded protein-binding sequence and a single-stranded antisense sequence, non-covalently conjugated with a shuttle protein like LFn-GAL4, allowing for membrane translocation without the need for polycationic affinity handles or toxic condensing agents.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If traditional antisense oligonucleotide delivery methods are used, then membrane penetration is achieved through mechanical or chemical cell damage, but safety and reliability are compromised
Solution Approach 1:
The patent uses a cell-penetrating peptide (CPP) as an intermediary carrier that facilitates the transport of antisense oligonucleotides across the cell membrane without causing mechanical or chemical damage. The CPP-ASO conjugate system allows the oligonucleotide to penetrate the membrane through the CPP's inherent ability to traverse membranes, eliminating the need for harmful delivery methods while maintaining safety and reliability
Solution Approach 2:
The invention creates a composite structure by chemically conjugating the cell-penetrating peptide with the antisense oligonucleotide to form a CPP-ASO conjugate. This composite material combines the membrane-penetrating capability of the CPP with the gene-silencing function of the ASO, enabling effective and safe intracellular delivery
2Reliability
If polycationic affinity handles or toxic condensing agents are used for delivery, then oligonucleotide complexation is achieved, but toxicity increases
Solution Approach 1:
The patent extracts and eliminates the toxic components (polycationic affinity handles and condensing agents) from the delivery system by using a cell-penetrating peptide instead. The CPP performs the complexation and delivery function without requiring toxic chemicals, thereby maintaining delivery effectiveness while removing the harmful elements that caused toxicity in traditional methods
Solution Approach 2:
The cell-penetrating peptide acts as a temporary, biodegradable carrier that fulfills its delivery function and is then naturally degraded by cellular processes. This disposable approach eliminates the need for persistent toxic materials while achieving reliable oligonucleotide delivery, as the CPP is broken down after completing its transport mission
Data Source
AI summary
The present invention relates to antisense oligonucleotide (ASO) compositions and particularly to compositions and methods for the cytosolic delivery of antisense oligonucleotides (ASOs). Hybrid ASOs, part single-stranded and part double-stranded, are provided, hybridizing to form a double-stranded region that can non-covalently bond to nucleic-acid-binding protein regions. In this way, ASO::protein complexes may be produced that facilitate delivery of antisense DNA into target cells. Such complexes may be used to down-regulate gene expression in cells.