Antibody-Drug Conjugate Linker Design for Species-Specific Stability
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Solution Overview
Problem
ADCs with linkers comprising dipeptides like Val-Cit are unstable in mouse plasma due to Ces1c, leading to differences in pharmacokinetics between mice and humans, making it difficult to evaluate drug efficacy in humans.
Innovation Solution
A conjugate with a linker that allows drug release by cleavage of a cleavable site and π electron resonance system interlocking, linked to a thiol group generated by disulfide bond cleavage in antibodies, maintaining hydrophilicity before and allowing hydrophobicity post-cleavage for enhanced cell permeability.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If a linker comprising Val-Cit dipeptide is used in ADC, then the ADC can release drug in human cancer cells via cathepsin B cleavage, but the ADC becomes unstable in mouse plasma due to Ces1c cleavage
Solution Approach 1:
The patent introduces a species-specific recognition element (the dipeptide sequence) at a specific location within the linker, making the linker's cleavage properties location-dependent and species-specific. The Val-Cit dipeptide is positioned between the drug and the antibody, creating a localized cleavage site that is recognized by human cathepsin B but not by mouse Ces1c in the same manner, thus achieving differential stability and activity between species.
Solution Approach 2:
The patent changes the chemical structure parameter of the linker by incorporating the Val-Cit dipeptide sequence, which fundamentally alters the cleavage characteristics. This structural parameter change enables the linker to be cleaved by human cathepsin B at a specific pH and ionic environment while resisting cleavage by mouse Ces1c, thereby achieving species-specific pharmacokinetic properties.
2Quantity of substance
If all four interchain disulfide bonds in IgG are cleaved to generate eight thiol groups, then more drug conjugation sites are available, but the antibody structure may become less stable
Solution Approach 1:
The patent performs preliminary reduction of disulfide bonds to generate free thiol groups before drug conjugation. By pre-reducing the disulfide bonds using a reducing agent, the antibody is prepared in advance with available thiol groups that can then be efficiently conjugated with the drug, avoiding the need for simultaneous reduction and conjugation steps.
Solution Approach 2:
The patent uses a reducing agent as an intermediary substance to cleave the disulfide bonds. The reducing agent temporarily interacts with the antibody's disulfide bonds, converting them to thiol groups, and then dissociates, allowing the reduced antibody to subsequently undergo drug conjugation without the reducing agent present during the conjugation step.
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 conjugate achieves a long residence time in the body with low aggregation and high monomer ratio, facilitating drug efficacy by suppressing hydrophobicity before and enabling hydrophobicity post-cleavage, thus enhancing cell permeability.
Implementation Method 1
Such a disulfide bond can be cleaved by a reducing agent. For example, when all the four disulfide bonds in IgG are cleaved, an IgG antibody having eight thiol groups is generated.
Implementation Method 2
cathepsin B in lysosomes in human cancer cells recognizes a VC structure and cleaves an amide bond present on a carboxy terminal side of citrulline. Therefore, an ADC having a linker comprising such a dipeptide can release a drug and exhibit a drug efficacy in human cancer cells.
Data Source
AI summary
A conjugate of an antibody and a functional substance, comprising a structural unit represented by the following Formula (1):whereinIg represents an immunoglobulin unit comprising two heavy chains and two light chains, and is bonded to LA adjacent to Ig via a thiol group in side chains of a plurality of cysteine residues in the two heavy chains and the two light chains,HG represents a hydrophilic group or a monovalent group comprising a hydrophilic group,CS represents a divalent group comprising a cleavable site,ring A represents a divalent aromatic ring group optionally having a substituent wherein the divalent aromatic ring group constitutes a π electron conjugated system with the cleavable site,V represents an oxygen atom, a sulfur atom, or an amino group (NH),LA and LB each independently represent a divalent group,D represents a functional substance, andan average number n of the bonds per the immunoglobulin unit is 1.5 or more, ora salt thereof, anda substance associated therewith,is excellent in desired properties.