Differences in Expansion Potential of Naive Chimeric Antigen Receptor T Cells from Healthy Donors and Untreated Chronic Lymphocytic Leukemia Patients

Abstract

IntroductionTherapy with chimeric antigen receptor T (CART) cells for hematological malignancies has shown promising results. Effectiveness of CART cells may depend on the ratio of naive (TN) vs. effector (TE) T cells, TN cells being responsible for an enduring antitumor activity through maturation. Therefore, we investigated factors influencing the TN/TE ratio of CART cells.Materials and methodsCART cells were generated upon transduction of peripheral blood mononuclear cells with a CD19.CAR-CD28-CD137zeta third generation retroviral vector under two different stimulating culture conditions: anti-CD3/anti-CD28 antibodies adding either interleukin (IL)-7/IL-15 or IL-2. CART cells were maintained in culture for 20 days. We evaluated 24 healthy donors (HDs) and 11 patients with chronic lymphocytic leukemia (CLL) for the composition of cell subsets and produced CART cells. Phenotype and functionality were tested using flow cytometry and chromium release assays.ResultsIL-7/IL-15 preferentially induced differentiation into TN, stem cell memory (TSCM: naive CD27+ CD95+), CD4+ and CXCR3+ CART cells, while IL-2 increased effector memory (TEM), CD56+ and CD4+ T regulatory (TReg) CART cells. The net amplification of different CART subpopulations derived from HDs and untreated CLL patients was compared. Particularly the expansion of CD4+ CARTN cells differed significantly between the two groups. For HDs, this subtype expanded >60-fold, whereas CD4+ CARTN cells of untreated CLL patients expanded less than 10-fold. Expression of exhaustion marker programmed cell death 1 on CARTN cells on day 10 of culture was significantly higher in patient samples compared to HD samples. As the percentage of malignant B cells was expectedly higher within patient samples, an excessive amount of B cells during culture could account for the reduced expansion potential of CARTN cells in untreated CLL patients. Final TN/TE ratio stayed <0.3 despite stimulation condition for patients, whereas this ratio was >2 in samples from HDs stimulated with IL-7/IL-15, thus demonstrating efficient CARTN expansion.ConclusionUntreated CLL patients might constitute a challenge for long-lasting CART effects in vivo since only a low number of TN among the CART product could be generated. Depletion of malignant B cells before starting CART production might be considered to increase the TN/TE ratio within the CART product.

Idelalisib for optimized CD19‐specific chimeric antigen receptor T cells in chronic lymphocytic leukemia patients

“…30 In regard to stimulation, it was shown that addition of IL-7/IL-15 is superior to cultivation of CART cells in the presence of IL-2 in terms of maintenance and survival of lessdifferentiated T cells. [11][12][13] In this study, addition of idelalisib could, even in the presence of IL-7/IL-15, further increase T N and T SCM cell subpopulations within the CART cell product. Another cytokine associated with enhancement of lessdifferentiated T cells is IL-21.…”

“…An enrichment of CD8+ T cells and less‐differentiated T cells can be reached by retronectin‐based T cell activation . In regard to stimulation, it was shown that addition of IL‐7/IL‐15 is superior to cultivation of CART cells in the presence of IL‐2 in terms of maintenance and survival of less‐differentiated T cells . In this study, addition of idelalisib could, even in the presence of IL‐7/IL‐15, further increase T N and T SCM cell subpopulations within the CART cell product.…”

“…In recent years, CART cell therapy evolved with regards to CAR vector design, but a vast pursuit is to further expand less‐differentiated T cells. Cytokines such as interleukin (IL)‐2, IL‐7 and IL‐15 can influence the cellular composition during the expansion stage of CART cell production . In addition, inhibition of specific signaling pathways during ex vivo T cell expansion can influence T cell differentiation .…”

“…Cytokines such as interleukin (IL)-2, IL-7 and IL-15 can influence the cellular composition during the expansion stage of CART cell production. [10][11][12][13] In addition, inhibition of specific signaling pathways during ex vivo T cell expansion can influence T cell differentiation. 14 Besides their antitumor activity, the approved B cell receptor (BCR) inhibitors ibrutinib and idelalisib can modulate critical pathways in T cells.…”

Dawn of chimeric antigen receptor T cell therapy in non-Hodgkin Lymphoma

“…Conditioning chemotherapy is given 2‐7 days prior to T‐cell infusion. The theoretical benefits of conditioning include direct cytotoxic effect against malignancy, depletion of regulatory T cell and other suppressive immune cells and decreased competition for homeostatic cytokines . The optimal conditioning regimen has not been rigorously examined, but clinical experiences from the NCI and FHCRC are instructive.…”