Activatable Antibodies Using Cleavable Masking Peptides
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Solution Overview
Problem
The development of activatable binding polypeptides, such as activatable antibodies, is a slow, labor-intensive, and costly process, necessitating improved methods for identifying self-blocking peptides that can effectively target antigens like CTLA4 or CD137 only in specific contexts, such as the tumor microenvironment.
Innovation Solution
The use of polynucleotide libraries encoding polypeptides with a specific amino acid sequence structure, comprising a first peptide, a cleavable moiety, and a target binding moiety, where the first peptide masks the antibody activity until cleavage by specific proteases, allowing for precise activation and enhanced therapeutic indexes and safety profiles.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If traditional methods are used to develop activatable binding polypeptides, then the process can identify functional antibodies, but the development process becomes slow, labor-intensive, and costly
Solution Approach 1:
The invention segments the antibody structure by introducing a cleavable moiety that divides the antibody into distinct regions: a target-binding region and a masking region. This segmentation allows the antibody to be screened in inactive form and then activated by proteolytic cleavage, dramatically accelerating the development process by enabling parallel screening of multiple candidates without requiring functional activation during the screening phase.
Solution Approach 2:
The invention applies preliminary action by pre-installing the cleavable moiety and masking peptide structure onto the antibody candidates before screening. This preliminary structural modification allows non-functional or weakly functional antibodies to be identified and selected based on their structural properties, which can then be activated later. This eliminates the need for time-consuming functional activation and testing during the screening process itself.
2Reliability
If activatable binding polypeptides are designed with masking peptides, then therapeutic index and safety are improved, but the design and screening process becomes more complex
Solution Approach 1:
The invention extracts the masking function from the target-binding function by using a cleavable moiety that separates these two activities. The masking peptide can be independently optimized for blocking activity, while the target-binding region can be independently optimized for affinity and specificity. This extraction simplifies the design process by allowing separate optimization of each function rather than trying to balance both simultaneously in a single continuous structure.
Solution Approach 2:
The invention applies parameter changes by modifying the molecular structure to include specific cleavage sites with defined protease recognition sequences. This introduces a new parameter (protease specificity) that can be controlled and optimized independently. The cleavable moiety contains specific amino acid sequences that determine which proteases can activate the antibody, allowing precise control over activation conditions while maintaining overall structural simplicity.
3Object-affected harmful factors
If self-blocking peptides are used to mask antibody activity, then cytotoxicity is reduced, but the masking efficiency must be precisely controlled to maintain therapeutic efficacy
Solution Approach 1:
The invention introduces the cleavable moiety as an intermediary between the masking peptide and the target-binding region. This intermediary component serves as a controlled release mechanism: it maintains the masking peptide in place during circulation (reducing cytotoxicity) but allows for precise removal upon encountering specific proteases in the tumor microenvironment. The cleavable moiety's specific protease recognition sequence acts as a molecular key that triggers activation only under the correct conditions, providing precise control over masking efficiency.
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
These polynucleotide libraries enable the identification of activatable binding polypeptides that efficiently treat multiple cancer types with reduced cytotoxicity, maintaining an activatable phenotype and binding affinity post-cleavage, while minimizing activity before activation, thus improving therapeutic efficacy and safety.
Implementation Method 1
capable of binding their target (when in active form) only after cleavage of the cleavable moiety (CM) to remove the first peptide (FP) (i.e., a masking moiety (MM) or self-blocking peptide)
Data Source
AI summary
Provided herein are libraries containing synthetic polynucleotides that encode activatable binding polypeptides. Further provided herein are activatable binding polypeptides and polypeptide libraries containing such activatable binding polypeptides. Also provided herein are vectors, vector libraries, cells, kits, and methods of making and using activatable polypeptide libraries.