Chimeric Polypeptides for Exosome Secretion and Antigen Loading
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Solution Overview
Problem
Current methods lack an effective tool to target antigenic proteins for immunization using exosomes, as previous studies have not described a molecular tool for efficiently loading antigenic proteins onto exosomes for immunization, and existing chimeric constructs are limited in their ability to produce exosomes across various cell types.
Innovation Solution
Development of novel chimeric polypeptides comprising an infra-membrane targeting domain, a peptide or polypeptide of interest, and a cytoplasmic domain, which can efficiently address and secrete the peptide or polypeptide of interest onto exosomes produced by various cell types, including HEK293 cells, thereby amplifying exosome production and enabling their use in immunization, diagnostics, and research.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If previous chimeric constructs are used, then some exosome production is achieved, but the production level is insufficient (not amplified 10 to 100 times) and is limited to specific cell types
Solution Approach 1:
The chimeric polypeptide construct is designed with a cytoplasmic domain from a widely expressed protein (such as GFP, RFP, or β-actin) that functions across multiple cell types. This universal domain enables the construct to be expressed and secreted in various cell types including HEK293, 293F, and other eukaryotic cells, achieving both high productivity and broad adaptability simultaneously
Solution Approach 2:
The invention modifies the cytoplasmic domain parameters by selecting specific protein sequences with optimized expression and secretion characteristics. The domain is engineered to interact with exosome formation machinery, changing the biological parameters of the construct to achieve 10 to 100 times amplification of exosome production while maintaining versatility across cell types
2Ease of manufacture
If no effective molecular tool is used, then existing methods are simple, but they lack the ability to efficiently target and load antigenic proteins onto exosomes for immunization
Solution Approach 1:
The chimeric polypeptide acts as an intermediary molecule that bridges the antigenic protein and the exosome. It contains a peptide of interest (antigen) fused to a cytoplasmic domain that mediates interaction with exosome formation machinery. This intermediary construct enables efficient targeting and loading of antigens onto exosomes while maintaining ease of manufacture through standard recombinant DNA techniques
Solution Approach 2:
The molecular tool is segmented into functional domains: a peptide of interest (antigen) and a cytoplasmic domain. This segmentation allows independent optimization of each domain - the antigen domain for immunogenicity and the cytoplasmic domain for exosome targeting - while maintaining overall simplicity in the cloning and expression process
Data Source
AI summary
The present invention provides a chimeric polypeptide comprising a plurality of polypeptide domains that are capable of being secreted in combination with membrane vesicles and in particular exosomes.The invention also concerns the use of polypeptides of the invention and polynucleotides coding for these polypeptides, for the production of immunogenic compositions based on exosomes or DNA, to screen protein interactions. The present invention also concerns exploiting the properties of exosomes comprising a polypeptide of the invention and immunogenic compositions of the invention in immunology.The present invention concerns the use of exosomes comprising a polypeptide of the invention as a diagnostic tool.The present invention also concerns exploiting the properties of membrane vesicles and protein compositions of the invention for the prophylaxis and/or treatment of a disease due to a functional deficit, in particular to transport a protein or a nucleic acid, in particular to compensate for or make up for an enzymatic deficit, or in particular to induce a transcriptional or translational modification in the target cells or organs.