Protease-Activatable Multispecific Binding Proproteins for Tumor Selectivity
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Solution Overview
Problem
Existing bispecific antibody therapeutics face challenges in selectively activating T-cells against tumor cells while minimizing toxicity to normal cells due to the expression of tumor-associated antigens on both tumor and normal tissues.
Innovation Solution
Development of protease-activatable antigen-binding molecule proproteins (ABMs) with protease-cleavable linkers that mask the T-cell engaging antigen-binding site (TCE ABS) until activated in the tumor environment, reducing toxicity by minimizing binding to normal tissues.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If bisspecific antibodies are designed to bind to both TAA and TCR complex components to activate cytotoxic T cells, then immune response against tumor cells is enhanced, but toxicity to normal cells increases due to TAA expression on healthy tissues
Solution Approach 1:
The antibody is administered in an inactive proprotein form where the TCE ABS is pre-masked by a masking moiety. This preliminary masking prevents premature activation and off-target effects. The antibody reaches the tumor site first, then activates only when protease-cleavable linkers are cleaved by tumor-associated proteases, ensuring selective activation at the target site rather than systemic activation.
Solution Approach 2:
A masking moiety is introduced as an intermediary element that temporarily blocks the TCE ABS from binding to the TCR complex. This masking moiety is connected via protease-cleavable linkers, creating a conditional blocking mechanism. The intermediary allows the antibody to circulate safely without activating T cells, then enables controlled activation when the masking is removed by tumor-specific proteases.
2Productivity
If the T-cell engaging antigen-binding site is kept active to ensure rapid T-cell activation, then treatment efficacy is improved, but binding to normal tissues increases causing adverse side effects
Solution Approach 1:
The antibody is designed to rapidly bind to TAA upon reaching the tumor site, but the TCE ABS is preliminarily masked to prevent immediate activation. This ensures rapid accumulation at the target while delaying activation until the antibody is positioned correctly, preventing premature systemic activation and associated side effects.
Solution Approach 2:
The masking moiety performs a preliminary anti-action by blocking the TCE ABS from binding to the TCR complex. This preliminary blocking prevents harmful activation in normal tissues, while the protease-cleavable linkers allow the blocking to be removed at the tumor site, enabling selective activation only where needed.
3Reliability
If a masking moiety is introduced to block the TCE ABS to reduce toxicity, then selectivity is improved, but molecule complexity increases
Solution Approach 1:
The masking moiety, protease-cleavable linkers, and the antibody itself are merged into a single integrated proprotein molecule. This combining reduces the need for separate components and simplifies manufacturing, as the entire construct is produced as one molecule that self-activates through proteolytic cleavage in the tumor microenvironment.
Solution Approach 2:
The proprotein is designed to self-activate through proteolytic cleavage by tumor-associated proteases. The masking moiety and linkers are configured so that the molecule automatically transitions from inactive to active state in response to the tumor microenvironment, eliminating the need for external activation mechanisms or complex delivery systems.
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 ABMs provide reduced toxicity and adverse side effects by selectively activating T-cells against tumor cells, enhancing targeted cancer treatment efficacy while minimizing impact on healthy tissues.
Implementation Method 1
the masking moiety is cleaved and a binding molecule is produced having enhanced target binding. This is achieved through the inclusion of one or more protease-cleavable linkers ("PCLs") that comprise one or more protease substrate sequences
Data Source
AI summary
The present disclosure provides proproteins of antigen-binding molecules, including multispecific binding molecules (MBMs). The antigen-binding molecule proproteins comprise an antigen-binding site, one or more components that inhibit the binding of the antigen-binding site to its target, and a protease-cleavable linker that can be cleaved by a protease and whose cleavage removes the inhibition of binding of the antigen-binding site to its target. The MBM proproteins and MBMs resulting from cleavage of a protease-cleavable linker. e.g., tandem Fab MBMs, comprise a T-cell engaging antigen-binding site and a tumor-associated antigen antigen-binding site. The MBM proproteins further comprise a component that inhibits the binding of the T-cell engaging antigen-binding site to its target and a protease-cleavable linker that can be cleaved by a protease in the tumor environment and whose cleavage removes the inhibition of binding of the T-cell engaging antigen-binding site to its target. The disclosure further provides pharmaceutical compositions comprising the antigen-binding molecule proproteins, and methods of use of the antigen-binding molecule proproteins in therapy, as well as nucleic acids encoding the antigen-binding molecule proproteins, recombinant cells that express the antigen-binding molecule proproteins, and methods of producing the antigen-binding molecule proproteins.