Finding the Cells That Matter

Directly link T cell phenotype and function to genotype

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T cell-based cancer immunotherapies show great promise but are also associated with severe side effects. To date, it has been challenging for scientists to improve efficacy and reduce toxicity because T cell-mediated tumor cell death relies on complicated and heterogeneous cell-cell interactions that are difficult to measure using traditional tools. The T Cell Analysis Suite on the Lightning™ optofluidic platform changes this by providing a direct link between T cell phenotype, function and genotype for thousands of
individual cells so you can find the cells that matter the first time you look.

Many T cell therapy side effects are related to cytokine release intensity and target cell killing kinetics.This could be addressed by selectively treating patients with only tumor antigen-specific T cells that quickly kill multiple target cells in succession and avoid high levels of cytokine release. These cells, however, represent a tiny fraction of an entire T cell population and are incredibly difficult to identify and isolate for further analysis. Single-cell approaches have allowed scientists to characterize this T cell diversity, but they only enable correlative conclusions based on data collected on multiple platforms and from different T cell populations. Additionally, they do not directly link a single T cell’s phenotype to its target cell killing kinetics or genotype. For example, it is simply not possible to determine whether a specific cytokine-secreting cell can kill tumor targets or to link this behavior directly to gene expression or genotype. Standard assays also make it challenging to discriminate between multiplexed and serial killing or to assess key killing characteristics such as length of T cell-tumor cell conjugation. In short, traditional methods make it nearly impossible to find the subset of cells that matter—those that are able to rapidly kill tumor cells with minimal toxicity to healthy cells.

The T Cell Analysis Suite on the Lightning platform can find and isolate these cells in just days. By taking advantage of this expanding suite of software capabilities, reagents and protocols scientists can define and test the function of individual cells (Figure 1). Individual T cell-target cell interactions are precisely assembled in thousands of nano-liter sized NanoPen™ chambers across a microfluidic chip. T cells can be selectively penned based on phenotype and co-cultured with target cells and cytokine capture beads. Function (cytokine secretion) and killing kinetics (caspase-3 activation in target cells) can be assessed using time-lapse brightfield and fluorescence microscopy. Cells with secretion profiles and killing kinetics of interests can be exported for downstream analysis such as sequencing.

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Figure 1. Assembly, visualization, and characterization of individual cell–cell interactions and recovery of live cells of interest

This enables previously impossible experiments, such as the analysis of variation in antigen-specific killing kinetics, recovery of TCR sequences directly associated with desired killing behavior, and discrimination between multiplexed and serial killing. With the T Cell Analysis Suite on the Lightning platform, scientists can now sift through thousands of individual T cells in parallel in a single day and on a single platform, which allows them to draw fast, actionable, causative conclusions about T cell function and biology. Such rapid and deep characterization has the potential to not only provide novel insights into T cell biology but also to accelerate the development of more efficacious cell therapies.

 

Visit berkeleylights.com/AnalysisSuite to download an application note exploring CAR-T and TCR use cases.

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