IsoPlexis’ Functional Phenotyping Reveals Insights That Could Help Create More Robust CAR-T Therapies

Recent advances in immuno-oncology therapeutics have helped improve clinical outcomes for patients with hard-to-treat tumors. However, since not all patients respond to these treatments, there is an urgent need for additional novel strategies to enhance the efficacy of immunotherapies.

In a recent study2, researchers hypothesized that bispecific anti-CD20 and anti-CD19 (LV20.19) CAR-T cells improve outcomes in relapsed, refractory (R/R) non-Hodgkin lymphoma (NHL) by limiting relapse from single antigen downregulation.

Previously reported results from a Phase I trial of LV20.19 CAR-T cells expanded in IL-2 in R/R NHL & CLL showed high response rates. In addition, preclinical models have demonstrated that, compared to IL-2, CAR-T cells expanded in IL-7 & IL-15 (IL7+15) improve expansion, in vivo persistence, and anti-tumor efficacy.2

In the current study, the team assessed a Phase I/II trial of LV20.19 (bispecific anti-CD20, anti-CD19 CAR-T cells) expanded in IL-7+15 by varying the lengths of CAR-T manufacturing time (8 vs 12 days) in R/R NHL. They hypothesized that expansion in IL-7+15 and manufacturing for shorter durations would generate a CAR-T product with superior functionality.

IsoPlexis’ Functional Phenotyping Provides Correlative Clinical and Preclinical Immune Biomarkers

To accomplish this, CAR-T cells were manufactured and thawed. CD4 and CD8 cells were sorted via immunomagnetic separation and stimulated with CD19+ K562 cells, which were then loaded onto IsoPlexis’ Single-Cell Adaptive Immune IsoCode chips and read in IsoPlexis’ IsoLight instrument. The single-cell production of 32 cytokines was measured. Polyfunctionality and polyfunctional strength index (PSI) were calculated for each CAR-T product.2

The team found that PFA was higher for cells expanded in IL-7+15 compared to IL-2. The PSI was also significantly higher for IL-7+15 versus IL-2 expanded cells. CD4 cells expanded in IL-7+15 also had substantially higher levels of IFN-y, Gzmb, MIP-1a, IP-10 and IL-2 and CD8 cells had drastically higher levels of Gzmb, IFN-y, TNF-a, MIP-1a, GM-CSF, IP-10, and IL-2 when expanded in IL-7+15.

In the IL-7+15 trial, eight patients were included in the 8-day manufacturing arm and six patients in the 12-day arm. PSI was similar between the two groups. The 8-day CD4 T cells had higher levels of perforin, while 8-day CD8 T cells had higher levels of MCP-1 and lower IL-4.

According to the paper’s authors, these results suggest that bispecific LV20.19 CAR-T cells expanded in IL-7+15 have greater PSI than IL-2 expanded cells and thus may generate more robust CAR-T products which could improve persistence, cytotoxicity, and patient outcomes.


Results from this study indicate that bispecific LV20.19 CAR-T cells expanded in IL-7+15 have superior polyfunctionality and polyfunctional strength index compared to IL-2 expanded cells. Because highly polyfunctional CAR-T cells and higher PSI have been linked to clinical response in single antigen targeted CAR-T cells, the authors conclude that the expansion of LV20.19 CAR-T cells in IL-7+15 may produce a more robust CAR-T product than expansion in IL-2, which could improve persistence, cytotoxicity, and ultimately patient outcomes.2

This is yet another example of the power of IsoPlexis’ unique functional phenotyping technology, which provides correlative clinical and preclinical immune biomarkers for advancing the manufacturing intricacies of  the competitive clinical world of immuno-oncology.

Want to learn how IsoPlexis technology can help advance your next-gen therapies? Get in touch.




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