Folate-IRNA Conjugates for Cellular Uptake
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Solution Overview
Problem
Current methods for delivering oligonucleotides face limitations in transmembrane transport, particularly due to poor solubility of folic acid and its derivatives, leading to variable conjugate transport and susceptibility to exonucleases, which hinders effective pharmacologic properties for therapeutic applications.
Innovation Solution
Development of oligonucleotide-folate conjugates with a folic acid moiety attached to modified subunits, such as ribose sugar replacements, optimized for improved cellular uptake through receptor-mediated endocytosis, utilizing a folate receptor binding ligand and a tethering mechanism for enhanced affinity and stability.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Ease of operation
If folic acid is conjugated to oligonucleotides to improve cellular uptake, then cellular penetration is enhanced, but solubility deteriorates leading to variable conjugate transport
Solution Approach 1:
The patent modifies the chemical structure of folic acid by replacing the pterin ring system with various heterocyclic alternatives (e.g., triazolo-pyridine, pyrimido-pyrimidine, triazolo-pyrimidine structures). These structural parameter changes maintain the folate receptor binding capability while improving solubility properties and reducing aggregation, thereby enabling more consistent conjugate transport without sacrificing cellular uptake efficiency.
Solution Approach 2:
The invention creates composite structures by combining oligonucleotide sequences with modified folic acid moieties through linker structures. These composite conjugates integrate the gene-silencing function of oligonucleotides with the targeted cellular delivery capability of folate analogs, while the modified folate structure ensures adequate solubility. The composite nature allows simultaneous optimization of multiple properties: cellular targeting, solubility, and nuclease resistance.
2Reliability
If folic acid is conjugated to oligonucleotides to enhance stability, then resistance to exonucleases improves, but solubility deteriorates leading to variable transport
Solution Approach 1:
The patent employs parameter changes in the folate structure (replacing pterin with heterocyclic variants) to decouple the stability function from solubility limitations. The modified folate structures maintain the protective effect against exonuclease degradation while exhibiting improved aqueous solubility compared to conventional folic acid, thereby eliminating the trade-off between stability and solubility.
Solution Approach 2:
The invention introduces modified folate analogs as intermediary structures that mediate between the oligonucleotide and the cellular environment. These intermediaries provide both the stability function (protecting oligonucleotides from degradation) and the solubility function (ensuring proper dissolution and transport), acting as a bridge that reconciles the otherwise conflicting requirements.
3Productivity
If conventional folic acid conjugates are used to target cells, then receptor-mediated endocytosis is enhanced, but variable transport occurs due to poor solubility
Solution Approach 1:
The patent systematically varies parameters of the folate structure (heterocyclic ring systems, linker configurations, substitution patterns) to optimize the balance between receptor binding affinity and solubility. The modified structures maintain high-affinity binding to folate receptors (ensuring efficient endocytosis) while their improved solubility characteristics ensure consistent dissolution and uniform distribution in biological fluids, leading to reliable and reproducible transport.
Solution Approach 2:
The invention introduces dynamic flexibility through variable linker structures and flexible heterocyclic frameworks that can adapt to different biological environments. These dynamic structures maintain optimal conformation for receptor binding while accommodating variations in solubility conditions, enabling consistent transport performance across different physiological contexts.
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
The conjugation of folic acid to oligonucleotides improves their cellular penetration and stability, optimizing gene silencing activity and tissue targeting, thereby enhancing the therapeutic potential of oligonucleotides.
Implementation Method 1
optimized for improved cellular uptake through receptor-mediated endocytosis, utilizing a folate receptor binding ligand
Data Source
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AI summary
The present invention provides iRNA agent including at least one monomer having the structure shown in formula (I') wherein: A and B are each independently for each occurrence O, N(R ) or S; X is H, a protecting group, a phosphate group, a phosphodiester group, an activated phosphate group, an activated phosphite group, a phosphoramidite, a solid support, - P(Z')(Z")O-nucleoside, -P(Z')(Z")O-oligonucleotide, a lipid, a PEG, a steroid, a polymer, -P(Z' )(Z")O-L6-Q'-L7-OP(Z'")(Z"")O-oligonucleotide, a nucleotide, or an oligonucleotide; Y is H, a protecting group, a phosphate group, a phosphodiester group, an activated phosphate group, an activated phosphite group, a phosphoramidite, a solid support, - P(Z')(Z")O-nucleoside, -P(Z')(Z")O-oligonucleotide, a lipid, a PEG, a steroid, a lipophile, a polymer, -P(Z')(Z")O-L6-Q'-L7-OP(Z'")(Z"")O-oligonucleotide, a nucleotide, or an oligonucleotide; R is folate, a folate analog a folate mimic or a folate receptor binding ligand; L6 and L7 are each independently for each occurrence -(CH2)I1-, -C(R')(R")(CH2)n-, - (CH2)nC(R')(R")-, -(CH2CH2O)mCH2CH2-, or -(CH2CH2O)mCH2CH2NH-; Q' is NH, O, S, CH2, C(O)O, C(O)NH, -NH-CH(Ra)-C(0)-, -C(0)-CH(Ra)-NH-, CO, formula (A) where Ra is H or amino acid side;;chain. R' and R" are each independently H, CH3, OH, SH, NH2, NH(Alkyl = Me, Et, Pr, isoPr, Bu, Bn) or N(diAlkyl = Me2, Et2, Bn2); Z', Z", Z' " and Z"" are independently 0 or S; n represent independently for each occurrence 1-20; and m represent independently for each occurrence 0-50.