Optimizing T Cell Functionality : Tips to Prevent Exhaustion

In our previous article, we explored the morphological changes that T cells undergo upon activation and the underlying reasons for these transformations.However, we left certain questions unanswered: Can exhausted T cells ever regain their full potential? Or are there ways to prevent exhaustion in the first place?

Research indicates that exhausted T cells have difficulty fully recovering once they become exhausted. While they might regain some ability to proliferate, they struggle to produce key cytokines like IL-2 and TNF-α. Although there are strategies to help exhausted T cells regain some functionality, such as PD-1 blockade or targeting stem-like T cell subsets like precursor of exhausted T cells (TPEX), these approaches only enhance the proliferative potential of specific T cell populations, but do not fully restore the overall functionality of exhausted T cells (Tsui et al., 2022). Fortunately, there are ways to prevent T cell exhaustion from occurring in the first place, and one of the crucial factors is the timing of restimulation.

What is restimulation?

Restimulation is the process of reactivating T cells through their T cell receptor after an initial activation. This can happen when T cells encounter the same antigen again, either naturally during an ongoing immune response, where antigen presenting cells such as dendritic cells present the antigen to T cells, or artificially in experimental settings.

And why is it important?

The importance of restimulation lies in its ability to sustain T cell growth and function. T cells typically grow more slowly two weeks after initial activation, and eventually stop growing altogether. Restimulation helps T cells keep proliferating and allows researchers and clinicians to generate sufficient cells for treatments. However, prolonged exposure of T cells to their specific antigen leads to ongoing antigen signals through T cells’ receptors, which eventually contribute to T cell exhaustion.

So how can we prevent exhaustion from improper restimulation?

Managing the timing of restimulations is crucial.

Waiting, is the first thing we need to do before restimulating T cells. This is to avoid restimulation-induced cell death (RICD). RICD is a natural mechanism that regulates immune responses by controlling effector T cell expansion and preventing unnecessary damage to the body. It is triggered by the re-engagement of the TCR on a cycling effector T cell, resulting in apoptosis. (Pohida et al., 2022) If restimulation occurs prematurely, it can lead to the death of a significant portion of T cells.

So, how long should we wait?

Based on current research, it's recommended to restimulate T cells 8 to 10 days after their initial activation. This timing aligns with the reduction in their suppressive effect on naive T cell growth, which starts to decline around day 7.

Besides, a treatment free interval (TRI) of 7 days before restimulating T cells can transcriptionally reprogram T cells and boost their performance. When T cells are allowed to rest, they show greater ability to kill target cells and produce more granzyme B compared to those that are constantly stimulated. Also, rested T cells release more IFN-γ and TNF-α when they are stimulated again. (Philipp et al., 2022) Therefore, restimulating T cells between day 8-10 after their initial activation appears to provide the most effective outcomes.

When you give T cells a longer break between activations, they can keep multiplying without limits. T cell growth reaches its peak when there’s a gap of 60 days or more between immunization boosts. This break allows the T cells to expand and thrive even after several rounds of stimulation, while maintaining their ability to grow (Soerens et al., 2023).

Apart from the timing of restimulation, another crucial factor in avoiding T cell exhaustion is the nature of the signals presented during T cell activation. There are four types of signals: Strong-Transient, Strong-Persistent, Weak-Transient, and Weak-Persistent. Each type impacts T cell activation efficiency and the likelihood of exhaustion. The Weak-Persistent signal is generally the best.

The graph shows that AimGel enhances T cell growth, while anti-CD3/28 beads inhibit it due to their strong, concentrated signals, which increase the risk of RICD. A key difference lies in signal density: anti-CD3/28 beads typically feature an IgG density of approximately 30,000 per μm², essentially saturating the bead surface. This high density is designed to maximize interactions with T cells as the signals remain steady. AimGel mitigates this with lower signal density, providing milder yet persistent stimulation that effectively activates T cells. Specifically, AimGel beads present signals on a mobile membrane surface, enabling adaptive protein-protein interactions with T cells. This dynamic presentation allows for a significant reduction in protein density to approximately 1,000 - 3,000 per μm² or even lower. Its complex stimulation with additional signals closely mimics natural T cell activation by antigen-presenting cells. This sustained signalling is crucial, as it avoids the rapid on/off stimulation which can lead to cell stress and death. The accompanying graphics illustrate these signal patterns and their effects on T cell functionality.

In summary, by giving T cells adequate intervals between restimulations—ideally 8 to 10 days, or even longer for the optimal results—and using the right activation signals, we can promote sustained T cell proliferation while minimizing exhaustion. Also, for restoration of their capacity to produce key cytokines like IL-2, IL-10 and IL-15, which are important in T cell proliferation and overall immune response.

Interested in learning more about the significance of soluble cytokine in T cell function? Stay tuned for our next blog.

Read our previous article for more on the Morphological changes in T cells

Chloe Chan: Graphics

Kiki Choi: Write-up

Reference list:

Tsui, C., Kretschmer, L., Rapelius, S., Gabriel, S. S., Chisanga, D., Knöpper, K., Utzschneider, D. T., Nüssing, S., Liao, Y., Mason, T., Valle Torres, S., Wilcox, S. A., Kanev, K., Jarosch, S., Leube, J., Nutt, S. L., Zehn, D., Parish, I. A., Kastenmüller, W., Shi, W., Buchholz, V. R., & Kallies, A. (2022). MYB orchestrates T cell exhaustion and response to checkpoint inhibition. Nature, 609(7927), 354–360. https://doi.org/10.1038/s41586-022-05105-1

Pohida, K., Lake, C. M., Yee, D., & Snow, A. L. (2022). Restimulation-induced cell death (RICD): Methods for modeling, investigating, and quantifying RICD sensitivity in primary human T cells via flow cytometric analysis. Journal of Immunological Methods, 505, Article 113203. https://doi.org/10.1016/j.jim.2022.113203

Philipp, N., Kazerani, M., Nicholls, A., Vick, B., Wulf, J., Straub, T., Scheurer, M., Muth, A., Hänel, G., Nixdorf, D., Sponheimer, M., Ohlmeyer, M., Lacher, S. M., Brauchle, B., Marcinek, A., Rohrbacher, L., Leutbecher, A., Rejeski, K., Weigert, O., von Bergwelt-Baildon, M., Theurich, S., Kischel, R., Jeremias, I., Bücklein, V., & Subklewe, M. (2022). T-cell exhaustion induced by continuous bispecific molecule exposure is ameliorated by treatment-free intervals. Blood, 140(10), 1104–1118. https://doi.org/10.1182/blood.2022015956

Soerens, A. G., Künzli, M., Quarnstrom, C. F., Scott, M. C., Swanson, L., Locquiao, J. J., Ghoneim, H. E., Zehn, D., Youngblood, B., Vezys, V., & Masopust, D. (2023). Functional T cells are capable of supernumerary cell division and longevity. Nature, 614(7949), 762–766. https://doi.org/10.1038/s41586-023-05846-4