American Society of Hematology

Figure 6.

$Tregs suppress the proliferation and killing capacity of CD4+CD25-effector Tcells. (A) Tregs and CD4+CD25− effector T cells were isolated from healthy donor-derived buffy coats by using an immune-magnetic cell isolation kit. Baseline immune cell frequencies and purity of isolated fractions were determined by flow cytometry, representative density plots are depicted. (B) Effector cells were labeled with violet tracer and incubated with or without Tregs in the presence of anti-CD3/CD28-coated beads. Proliferation was measured by flow cytometry after 5 days. (C) RPMI-8226 cells were incubated with Tregs and/or effector cells for 48 hours at an E:T ratio of 1:1 and 3:1, with solvent control or talquetamab (4.0 µg/mL). In combination experiments, Tregs and effector T cells were combined in a 1:1 ratio. Lysis of MM cells was determined using a BLI-based cytotoxicity assay. (D) Cytokine and granzyme B concentrations in cell-culture supernatants were measured using flow cytometry, and concentrations in talquetamab-treated conditions are depicted as fold change compared with solvent control. Data represent mean ± SEM of 3 independent experiments, performed in triplicate. Groups were compared using ANOVA with Tukey’s posttest. *P < .05; **P < .01; ***P < .001; ****P < .0001.$

Tregs suppress the proliferation and killing capacity of CD4⁺CD25^-effector Tcells. (A) Tregs and CD4⁺CD25⁻ effector T cells were isolated from healthy donor-derived buffy coats by using an immune-magnetic cell isolation kit. Baseline immune cell frequencies and purity of isolated fractions were determined by flow cytometry, representative density plots are depicted. (B) Effector cells were labeled with violet tracer and incubated with or without Tregs in the presence of anti-CD3/CD28-coated beads. Proliferation was measured by flow cytometry after 5 days. (C) RPMI-8226 cells were incubated with Tregs and/or effector cells for 48 hours at an E:T ratio of 1:1 and 3:1, with solvent control or talquetamab (4.0 µg/mL). In combination experiments, Tregs and effector T cells were combined in a 1:1 ratio. Lysis of MM cells was determined using a BLI-based cytotoxicity assay. (D) Cytokine and granzyme B concentrations in cell-culture supernatants were measured using flow cytometry, and concentrations in talquetamab-treated conditions are depicted as fold change compared with solvent control. Data represent mean ± SEM of 3 independent experiments, performed in triplicate. Groups were compared using ANOVA with Tukey’s posttest. *P < .05; **P < .01; ***P < .001; ****P < .0001.

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