Optimizing Human T Cell Expansion With IL-2

Adoptive T cell therapy capitalizes on the body’s immune system to combat cancer by infusing ex vivo expanded T cells engineered to target cancerous cells. The process of large-scale expansion, particularly for therapies like CAR-T, is intricate and requires meticulous optimization to maximize T cell yield while preserving their desired function.

Optimal growth and viability are achieved through culturing immune cells with a high-quality activator supplemented with growth factors and immunomodulators in the growth medium, with a notable combination being Interleukin-7 (IL-7) and IL-15. These cytokines play pivotal roles as growth factors in ex vivo T cell expansion protocols, promoting rapid T cell multiplication and enhancing cytotoxicity, resulting in more potent cancer treatments. 

Beyond their growth-promoting functions, IL-7 and IL-15 also fortify the survival and functionality of transferred T cells within the patient, ensuring a sustained assault on malignant cells.

Results With IL-2 Supplemented T Cell Expansion

Stem cell memory T cells (TSCM) are preferred for adoptive T cell therapy due to their capacity to self-renew and differentiate into various T cell subsets, ensuring long-term persistence and sustained anti-cancer activity. Their unique properties enhance the durability and efficacy of the therapy by maintaining a potent and adaptable immune response against cancerous cells.

When supplemented with IL-2, T cells cultured using Nanotein’s NanoSpark STEM-T Soluble T Cell Activator demonstrated 100-fold expansion by day 10 with industry-leading enrichment of TSCM at a final concentration of 40% and 90% for CD4+ and CD8+ subpopulations, respectively (Figure 1).

IL-7/IL-15 Data from Datasheet

Recommended Protocol

The NanoSparkTM STEM-T Soluble T Cell Activator is intended for ex vivo activation of fresh or cryopreserved CD3+ T cells or human resting T lymphocytes from peripheral blood mononuclear cells (PBMCs). Xeno-free T cell expansion media can be used and supplemented with high-quality recombinant human IL-2. Fluorophore-conjugated antibodies may be used for flow cytometry characterization as needed. 

Fresh Cells

Day 0 – Activation: Exchange CD3+ T cells or PBMCs into xeno-free culture media. Cells should be seeded at approximately 1×106 cells/mL. To activate cells, add 10 μL of NanoSparkTM STEM-T Soluble T Cell Activator for every mL of cell suspension (e.g., 80 μL of the activator for 8 mL of cell suspension). Add IL-2 cytokine supplement to culture media at 20 ng/mL final concentration. Incubate cells at 37 °C and 5% CO2 overnight in a humidified incubator.

(OPTIONAL) Day 1 –  Transduction: 24 hours after adding the NanoSparkTM Activator, apply

viral vector for ~24-48 hours.

Cell Expansion & Maintenance: It is important to ensure the activator is in culture media for at least 72 hours (up to six days). Continue to incubate cells at 37 °C and 5% CO2 in a humidified incubator. Every two to three days, monitor and count the cells for viability and adjust confluency as needed. 

Add fresh culture medium every three days supplemented with 20 ng/mL IL-2. For optimization in your specific setup, consider using the NanoSparkTM Activator at concentrations of 5-10 μL per mL for each culture media refresh. Repeat these maintenance steps until the desired cell number is reached or up to 14 days after initial activation.

Cryopreserved Cells

Day -1 – Thaw Cells: Remove cryovial containing frozen CD3+ T cells or PBMCs from storage and quickly dip them into a 37 °C bath. Holding the top of the vial, gently swirl by hand in the water bath until almost completely thawed, about one minute. There should be a small bit of ice remaining in the vial. Sterilize the vial and transfer it to a laminar flow hood. Work quickly to count and seed cells at 1×106 cells/mL into culture media to maintain viability. 

Day 0 – Follow the remaining “Fresh Cells” protocol steps, starting with the addition of NanoSparkTM STEM-T Soluble T Cell Activator. 


Discussion and the Benefits of NanoSpark™ STEM-T

The NanoSpark™ STEM-T Soluble T Cell Activator is a cutting-edge product developed by Nanotein Technologies that has revolutionized adoptive cancer therapy. The STEM-T Activator enriches TSCM cells with all protein-based nanoparticle technology that complements well with IL-2 cytokine support. 

This activator addresses a critical challenge in adoptive T cell therapy: the efficient and selective expansion of TSCM cells. These remarkable cells possess the unique ability to self-renew and differentiate into diverse T cell subsets, ensuring their long-term persistence within the patient’s body. The NanoSpark™ STEM-T Soluble T Cell Activator’s precision-engineered formulation amplifies this inherent potential, resulting in a substantial increase in TSCM cell numbers available for infusion.

The NanoSpark™ platform synergizes seamlessly with IL-2 cytokine support, further enhancing the effectiveness of the adoptive therapy. By providing a tailored microenvironment that sustains T cell viability and function, IL-2 ensures the prolonged presence of these potent immune effectors in the fight against cancer. 

This innovative platform empowers future developments in CAR-T cancer therapy by offering a streamlined, highly efficient, and scientifically validated method to bolster CAR-T and CAR-NK cells, making it an invaluable asset for researchers and patients alike. 

NanoSpark™ STEM-T Soluble T Cell Activator marks a pivotal step forward in the quest to harness adoptive T cell therapy’s full potential and represents a promising avenue for advancing the treatment of cancer. Contact us today to learn more.

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