Question: What type of linkers are used for antibody-siRNA conjugation?
Can you share the type of linker that BioSynthesis uses to conjugate antibodies (Abs) to siRNAs?
A variety of linkers is available for antibody-siRNA conjugates. Linker types are often categorized by their cleavage mechanism.
[1] Cleavable Linkers
Cleavable linkers are designed to be stable in circulation but break down under specific intracellular conditions, releasing the siRNA payload at the target site. This controlled release is vital for therapeutic efficacy and minimizing off-target effects.
Peptide-based Linkers (Enzyme-cleavable) are very common and are designed to be cleaved by specific lysosomal proteases such as Cathepsin B found within target cells.
Valine-Citrulline (Val-Cit or VC) linkers are widely used dipeptide linkers, often combined with a self-immolative spacer like p-aminobenzylcarbamate (PABC) to ensure efficient release of the unmodified payload.
Others are Phenylalanine-Lysine (Phe-Lys) or Valine-Alanine (Val-Ala) linkers.
Multi-functional peptides: Some more complex linkers incorporate cell-penetrating peptides (CPPs) and enzyme-cleavable substrate peptides to enhance intracellular delivery and release specifically at tumor sites.
Disulfide Linkers (Glutathione-sensitive) linkers contain a disulfide bond (-SS-) that is relatively stable in the extracellular environment but is cleaved by the high concentration of intracellular glutathione (a reducing agent) allowing the release of the siRNA once it's inside the cell. SPDP (N-Succinimidyl 3-(2-pyridyldithio)propionate) is a common reagent used to introduce disulfide linkages.
pH-Sensitive Linkers (Acid-cleavable) linkers are stable at neutral pH (bloodstream) but become unstable and hydrolyze in acidic environments, such as the low pH of endosomes (pH 5-6) and lysosomes (pH 4.8).
Hydrazone linkers are a well-known example that exploits acidic conditions for cleavage.
Carbonate linkers are hydrolyzed in acidic environments.
β-Glucuronide Linkers are cleaved by lysosomal β-glucuronidases, which are hydrolytic enzymes found in lysosomes.
[2] Non-Cleavable Linkers
Non-cleavable linkers form stable, permanent bonds between the antibody and the siRNA. The siRNA is released in the cell only after the entire conjugate is internalized and degraded by lysosomal enzymes within the cell.
Thioether Linkers are formed through maleimide conjugation reaction to a thiol group. SMCC - Succinimidyl-4-(N-maleimidomethyl)cyclohexane-1-carboxylate is an example that forms a robust and stable linkage.
Other non-cleavable linkers contain amide bonds, oxime bonds (C=N-O-), triazole rings, and aromatic heterocycles formed via click chemistry.
Enzymatically-Cleavable-Linker-Sequence-Motifs
Oligonucleotide-linkers-for-conjugation
Photocleavable linkers
Spacer-18-heg-oligonucleotide-modification
SMCC-and-Sulfo-SMCC-for-Cross-Linking-and-Conjugation
Peptide-Linkers-and-Linker-Peptides-for-Antibody-Drug-Conjugates-(ADCs)
Amino-Linker-Oligonucleotide-Modification
Thiol-c6-modified-oligonucleotide
Comparison-of-long-tethers
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