Chimeric Polypeptide Conversion via Site-Specific Recombination
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Solution Overview
Problem
Existing methods for converting polypeptides of one type into another, such as converting a single-chain variable fragment (scFv) into a chimeric polypeptide, are inefficient and lack effective solutions for generating specific antibody forms like IgG or chimeric antigen receptors (CARs).
Innovation Solution
The method involves using two vectors with specific site-specific recombination motifs, a recombinase enzyme, and regulatory elements to fuse portions of the scFv with constant regions or other polypeptides, enabling the conversion of scFv into chimeric polypeptides like IgG or CARs, allowing for the creation of antibodies and antigen receptors with desired functionalities.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If existing methods are used to convert polypeptides of one type into another, then the conversion process can be performed, but the efficiency is low and the process is cumbersome
Solution Approach 1:
The patent applies preliminary action by pre-designing vectors that contain predetermined recombination motifs (such as attP and attB sites) and polypeptide sequences ready for integration. The system is prepared in advance with all necessary components (vectors, motifs, and polypeptide sequences) so that when conversion is needed, the recombination can proceed directly without time-consuming setup or design steps.
Solution Approach 2:
The patent uses recombination motifs as intermediaries to facilitate the conversion process. These motifs act as mediators between the polypeptide sequences and the vector systems, enabling efficient and specific recombination. The motifs serve as recognition sites that guide the recombination process, making the conversion of polypeptides between different types (e.g., from scFv to IgG or CAR) more efficient and less time-consuming.
2Manufacturing precision
If existing conversion methods are used, then polypeptide type conversion is possible, but the methods lack precision and flexibility in generating specific antibody forms
Solution Approach 1:
The patent applies segmentation by dividing the antibody polypeptide into distinct modular segments (such as variable regions, constant regions, and recombination motifs). Each segment can be independently designed, selected, and recombined. This modular segmentation enables precise control over which parts of the polypeptide are retained or modified, allowing accurate generation of specific antibody forms like IgG or CAR while maintaining flexibility to create different configurations.
Solution Approach 2:
The patent applies local quality by enabling different regions of the polypeptide to have different properties and functions. Specific segments can be optimized for particular purposes (e.g., binding affinity in variable regions, stability in constant regions) while maintaining overall functionality. This allows precise tailoring of each antibody form to meet specific requirements for different applications.
3Ease of manufacture
If conventional polypeptide conversion methods are used, then basic conversion is achieved, but the process is inefficient and lacks effective solutions for specific applications
Solution Approach 1:
The patent applies universality by creating a multi-functional vector system that can handle multiple types of polypeptide conversions through a unified approach. The same basic vector architecture and recombination mechanism can be used to convert scFv to IgG, to CAR, or to other antibody forms, making the process easy to manufacture while maintaining high efficiency. The universal system eliminates the need for separate specialized methods for each conversion type.
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
This approach enables efficient and precise conversion of scFv into chimeric polypeptides, such as IgG antibodies and CARs, enhancing their functionality and application in immunotherapy and diagnostics.
Implementation Method 1
The vectors may further include complementary site-specific recombination motifs, such that site-specific recombination between the two vectors results in the generation of a chimeric polypeptide
Implementation Method 2
contacting the first vector and the second vector in the presence of a recombinase enzyme, in which the recombinase enzyme combines the first vector and the second vector in a site-specific manner to form an integrant vector
Data Source
AI summary
The present invention provides methods and compositions for converting a first polypeptide into a chimeric polypeptide. The invention includes two vectors: a first vector including the sequence of the first polypeptide and a second vector including a second polypeptide. The vectors include complementary site-specific recombination motifs such that site-specific recombination between the two vectors results in the generation of a chimeric polypeptide including at least a portion of the first polypeptide and at least a portion of the second polypeptide. A site-specific recombination motif may be positioned within an intron or within a coding sequence on the first or second vector.