T-cell Avidity

What is avidity, why is it important, and how can we measure it

Avidity is the combined strength of multiple affinities, such as between cell receptors and their corresponding ligands.

For example, while the affinity between a T cell receptor (TCR) and an antigen on the cell surface of a tumor cell contributes to the strength of the interaction, the sum of other membrane-associated factors will ultimately define the total intercellular forces. Examples of these factors include co-receptor binding, TCR clustering, or even the orientation and valency of each binding partner.

So, while affinity describes the strength of a single interaction, avidity defines the overall binding strength and determines how strongly a cell interacts with its target.

T Cell binding schemcatic

Physiologically, cells communicate with each other, by establishing molecular bonds: a cell expresses multiple TCR-pMHC interactions and interacts with various co-stimulatory molecules and co-receptors.

Why T-cell avidity?

As shown on the figure at the left, cells can establish multiple interactions, for example between TCRs and pMHCs or various co-stimulatory molecules and co-receptors. Assessing the overall strength of all of these simultaneous interactions provides information about avidity which has been shown to correlate with the outcomes of cell-cell interactions. This information can be essential for immuno-oncology and of unprecedented value for future immune therapies, as the ability of T cells to recognize and encounter cancer cells is often dictated by their avidity.

So far, researchers have mostly been using surface plasmon resonance (SPR) to assess affinity. However recent studies have shown that affinity is a poor predictor of T-cell activation or functionality and cannot discriminate functional potency (Leah V. Sibener et al., 2018. Cell). On the other hand, techniques for avidity readouts, such as micropipette adhesion assay is technically challenging to use and provides low throughput. By contrast, the tetramer staining is a high-throughput fluorescence technique. However, recent findings have shown fluorescence determined by tetramer staining lead to false negative readouts of T-cell functionality (Zhang et al., 2016. Science Translational Medicine). Also,  the established readouts are based on bulk populations, offering only averaged functionality readouts of the mixed population of T-cells.

Here, we present z-Movi®: a high throughput single-cell resolution, label-free, lab-on-a-chip technology that enables reliable high throughput measurements of T-cell avidity.

z-Movi™ Avidity Analyzer: Principle and Workflow

The z-Movi technology is a lab-on-a-chip device that uses acoustic waves to apply controllable and non-invasive forces on cells.

In the figures below we present a case study showing the typical workflow of z-Movi technology to compare the avidity between non-transduced (control) T cells and transduced CAR-T cells with low or high avidity. In short:

  • You can plate target cancer cells of your choice on the surface of the z-Movi chip (step 1);
  • Next, you can apply the effector T cells you want to assess to the chip and let them sediment onto the target cells (step 2);
  • An applied electric tension (voltage) drives a piezo element within the z-Movi Chip to generate a standing acoustic wave that forces the hundreds of effector cells to move vertically to the nodal plane (step 3);
  • Because of the heterogenous avidities within the cell populations, you would need to apply varying forces to force different T cells towards the nodal plane (step 4).

The graph at the right shows the data we typically retrieve from these types of avidity experiments, that is, real-time data revealing effector cell populations that interact strongly with the target cancer cells.

The working principle of z-Movi, which is based on a lab-on-a-chip with microfluidics channels that enables the introduction of target and effector cells in a controlled environment. Brightfield microscopy with low-power LED illumination enables the imaging of cells without damaging them.

Real-time avidity curves on applying force ramp showing three distinct populations of effector CAR-T cells exhibiting different levels of acidity to the target cancer cells.

T-cell avidity - z-Movi - target cells

Step 1: Culture monolayer of target cancer cells in the z-Movi Chip

T-cell avidity - z-Movi - flushed CAR-T cells

Step 2: Flow in the effector T cells

Step 3: Region of Interest (ROI) selection of CAR-T cells on top of tumor cells monolayer

T-cell avidity - z-Movi - force ramp on CAR-T cells and tumor cells

Step 4: Force ramp applied to differentiate between the types of effector CAR-T cells based on their avidity to the cancer cells.



T Cell Avidity Analyzer

The z-Movi is the only instrument that can directly measure the avidity, or overall strength of interaction, between cells  (e.g., CAR-T and target tumor cells) or between cells and ligands (e.g., TCR-ligand interactions). Introduced in 2018, this new technology based on acoustic waves paves the way for the study of yet unexplored avenues in basic and translational research, impacting applications where cell-cell interactions are key, including immunotherapy, antigen presentation, therapeutic antibodies, vaccination, immunological synapse, and cellular adhesion. … Read More

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