Reversible Aptamer Cell Selection for High-Purity, High-Yield Isolation
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Solution Overview
Problem
Current cell therapy methods, such as CAR T cell therapies, face challenges with low cell yield and high costs due to the need for successive positive selections, particularly in label-free protein-based isolation technologies.
Innovation Solution
Aptamer-based reversible cell selection technology for production-scale isolation of cells, utilizing aptamers that specifically bind cell surface markers, followed by separation and recovery of target cells using solid supports or phase-changing agents, and reversal agents to disrupt binding.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Manufacturing precision
If label-free protein-based isolation technologies are used to isolate specific cell subsets, then cell purity is improved, but cell yield decreases and costs increase due to the number of successive positive selections required
Solution Approach 1:
The invention changes the binding parameter from protein-antigen interaction to nucleic acid aptamer-target interaction. Aptamers are selected through SELEX to have high affinity and specificity for target cells, enabling single-step isolation that achieves both high purity and high yield without requiring multiple successive selections
Solution Approach 2:
The invention replaces the mechanical/process-intensive approach of successive positive selections with a molecular recognition system based on aptamer binding. This substitution allows for more efficient cell isolation in a single step, improving both productivity and reducing operational complexity
2Manufacturing precision
If label-free protein-based isolation technologies are used to isolate specific cell subsets, then cell purity is improved, but costs increase due to the number of successive positive selections required
Solution Approach 1:
The invention changes the isolation mechanism from protein-based to aptamer-based, where aptamers can be synthesized chemically at lower cost compared to protein production. This parameter change reduces the cost burden while maintaining the ability to achieve high cell purity through specific binding
Solution Approach 2:
Aptamers are chemically synthesized nucleic acid molecules that can be produced more cheaply and stable than protein-based reagents. They can be used as disposable binding agents in the isolation process, reducing overall manufacturing costs while achieving the desired cell purity
3Manufacturing precision
If successive positive selections are performed to isolate specific cell subsets, then cell purity is improved, but the number of steps and time required increases
Solution Approach 1:
The invention changes the selection approach from multiple iterative protein-based selections to a single aptamer-based selection step. Aptamers are engineered to have high specificity and affinity, allowing one-step isolation that achieves the same or better purity in significantly less time
Solution Approach 2:
The aptamers are pre-selected and optimized through SELEX before use in the isolation process. This preliminary action creates ready-to-use binding agents that can directly isolate target cells in a single step, eliminating the need for multiple successive selections and reducing overall isolation time
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
Enables efficient and cost-effective isolation of viable cells, such as CD8+ T cells, with high purity and yield, suitable for large-scale applications.
Implementation Method 1
contacting the biological sample with an aptamer that specifically binds a cell surface marker that is specific for the cell of interest under conditions that permit forming aptamer-bound cells
Implementation Method 2
the step of separating aptamer-bound cells from cells not bound to the aptamer comprises the use of a first solid support or a phase-changing agent
Implementation Method 3
the aptamer is conjugated to a phase-changing agent. In one embodiment, contacting step (i) is performed under conditions in which the phase-changing agent is in the solution phase and the separating step comprises (ii) inducing the phase-changing agent to precipitate from solution
Implementation Method 4
the first and/or second solid support comprises a magnetoresponsive bead. the separating step comprises subjecting the sample to a magnetic field, whereby magnetoresponsive beads or solid support comprising aptamer-bound cells are separated from the biological sample
Implementation Method 5
recovering a cell of interest by disrupting binding of the aptamer to the cell surface marker
Data Source
AI summary
Described herein is a reversible aptamer selection technology for production-scale isolation of label-free cells (e.g., CD8+ T cells). Provided herein are methods for isolating a cell of interest from a biological sample by contacting the biological sample with an aptamer that specifically binds a cell surface marker specific for the cell of interest; separating the aptamer-bound cells from cells not bound to the aptamer; and recovering a cell of interest by disrupting binding of the aptamer to the cell surface marker.