Regulated Biocircuit Systems for Tunable Protein Expression
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Solution Overview
Problem
Current gene therapy technologies lack the ability to titrate the timing and levels of target protein induction, making it difficult to safely and effectively deploy many potential gene therapy applications, especially for proteins with narrow therapeutic windows or those requiring transient expression.
Innovation Solution
The development of regulatable and tunable biocircuit systems, specifically using stimulus response elements (SREs) such as FKBP or E.coli DHFR destabilizing domains, to modulate protein levels in response to specific stimuli, allowing for precise control of gene expression and protein function.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Manufacturing precision
If current gene therapy technologies are used to induce target protein expression, then protein expression is achieved, but the timing and levels of protein induction cannot be titrated
Solution Approach 1:
The patent applies dynamics by making the protein expression system adjustable and responsive to external stimuli. Destabilizing domains are designed to respond dynamically to small molecule ligands, allowing the system to transition between stable and unstable states, thereby enabling temporal and quantitative control of protein expression levels.
Solution Approach 2:
The patent implements parameter changes by modifying protein stability parameters through the use of destabilizing domains. By changing the stability parameter in response to ligand binding, the system can precisely control the timing and levels of protein induction, transforming a static expression system into a tunable one.
2Duration of action of moving object
If destabilizing domains are used to control protein stability, then temporal control of protein levels is achieved, but the system complexity increases
Solution Approach 1:
The patent applies segmentation by dividing the protein of interest into separate functional domains: a stable core domain and a destabilizing domain. This segmentation allows the destabilizing domain to be independently controlled by small molecule ligands, enabling temporal control of protein expression without requiring complex regulatory machinery.
Solution Approach 2:
The patent uses small molecule ligands as intermediaries to control protein stability. These small molecules bind to destabilizing domains, acting as mediators that translate external signals into intracellular protein stability changes, thereby simplifying the control mechanism while achieving precise temporal regulation.
3Ease of operation
If top level transcriptional regulation via inducible promoters is used, then protein expression regulation is achieved, but the response time is delayed
Solution Approach 1:
The patent extracts the regulatory control from the transcriptional level and relocates it to the post-translational level. By removing the need for transcriptional regulation and instead controlling protein stability directly, the system eliminates the time delay associated with transcription and translation initiation, achieving rapid response to regulatory signals.
Solution Approach 2:
The patent substitutes the mechanical transcriptional regulation system with a chemical stabilization system. Instead of relying on complex promoter-regulator interactions that require time for transcriptional activation, the system uses small molecule-ligand binding to directly stabilize or destabilize proteins, replacing a slow mechanical process with a faster chemical interaction.
Data Source
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AI summary
The present invention provides regulatable biocircuit systems. Such systems provide modular and tunable protein expression systems in support of the discovery and development of therapeutic modalities.