Engineered Microvesicles for Direct Protein Delivery
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Solution Overview
Problem
Current methods for introducing proteins into target cells, such as transfection and transduction, are inefficient, slow, and often associated with toxicity and delayed protein expression, particularly in non-dividing cells like neurons, and require complex and time-consuming viral vector production.
Innovation Solution
Engineering eukaryotic cells to overexpress viral membrane fusion proteins, which leads to the secretion of microvesicles containing the protein of interest, enabling rapid and efficient delivery to target cells without altering the protein's function.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Speed
If transfection techniques (calcium phosphate, electroporation, lipofection) are used to introduce genetic material into target cells, then protein expression can be achieved, but the process is slow (requiring transcription and translation) and causes toxicity and immune responses
Solution Approach 1:
The patent applies preliminary action by pre-synthesizing the protein of interest in producer cells and packaging it into viral particles beforehand. This eliminates the need for transcription and translation in target cells, achieving immediate protein expression upon particle entry while avoiding transfection reagent toxicity and immune responses associated with genetic material delivery.
Solution Approach 2:
The patent extracts the protein expression step from the target cell by producing the protein in advance in separate producer cells. The extracted protein is then delivered directly to target cells via viral particles, separating the protein synthesis function from the target cell's transcriptional and translational machinery, thereby avoiding associated toxicities and delays.
2Reliability
If retroviral transduction is used to transfer genetic material, then stable long-term protein expression is achieved, but the process is tedious and time-consuming due to production of multiple plasmids and viral particles
Solution Approach 1:
The patent extracts the protein synthesis function from the viral transduction process by producing the protein of interest in advance in producer cells. This separates protein production from viral particle production, eliminating the need to produce and manipulate multiple plasmids and viral particles, thereby significantly reducing preparation time while maintaining the ability to achieve stable expression through viral delivery.
Solution Approach 2:
The patent applies preliminary action by pre-producing and pre-purifying the protein of interest in producer cells before packaging it into viral particles. This preliminary protein synthesis step eliminates the time-consuming processes of co-producing multiple plasmids and viral particles simultaneously, streamlining the overall procedure while ensuring reliable protein delivery.
3Ease of manufacture
If retroviral vectors are used for gene transfer, then genetic material can be delivered to target cells, but there is a sanitary risk even when vectors are inactivated
Solution Approach 1:
The patent extracts the harmful viral genetic material from the delivery system by producing only the necessary viral envelope proteins in producer cells without incorporating functional viral genomes into the particles. This creates a sanitized delivery system that retains viral particle functionality for cell entry while eliminating the sanitary risks associated with retroviral genetic material.
Solution Approach 2:
The patent uses a disposable, non-integrating viral particle system that delivers protein without integrating genetic material into the target cell genome. These particles are essentially 'single-use' delivery vehicles that enter cells, release their protein cargo, and do not establish persistent viral infections, thereby eliminating long-term sanitary risks while maintaining delivery effectiveness.
4Speed
If microinjection is used to deliver protein into target cells, then direct protein delivery is achieved, but the method is costly and poorly amenable to large-scale uses
Solution Approach 1:
The patent applies self-service by engineering producer cells to autonomously produce and package the protein of interest into viral particles. This self-organizing system eliminates the need for manual microinjection operations, enabling automated, high-throughput protein delivery that is scalable to large numbers of cells while maintaining the speed and directness of protein delivery.
Solution Approach 2:
The patent merges protein production, viral particle assembly, and delivery functions into a single integrated system using engineered producer cells. This consolidation combines the advantages of direct protein delivery with the scalability of cell-based production systems, eliminating the labor-intensive microinjection step while maintaining efficient protein transfer to target cells.
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 method allows for transient and rapid introduction of functional proteins into target cells, avoiding the need for transcription and translation in the target cell, reducing toxicity and activation of immune responses, and is applicable to various cell types, including non-dividing cells.
Implementation Method 1
contacting said target cell with a microvesicle according to the disclosure comprising said protein of interest, wherein the microvesicle is formed by a eukaryotic cell overexpressing a viral membrane fusion protein
Data Source
Figure 1A~1D
Figure 2A~2D
Figure 3A~3B
AI summary
The present invention relates to direct protein delivery with engineered micro vesicles.