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. 2023 Jul 25;12:e83659. doi: 10.7554/eLife.83659

Figure 5. The in vitro model predicted tumor evolution in time and space.

(a) Scheme of cell detachment and serial engagement in the in vitro model. Immunological synapses (IS) duration is set to 150 min. Pe, encounter probability, Pa, adhesion probability; (b) Long-term simulation (72 hr) of target cell depletion across drug concentrations; (c–d) The effects of drug concentration (c) and incubation time (d) on CD19 expression. Dashed line, a pre-defined threshold value of CD19 expression for 15% target cell depletion within 72 hr (initial setup: 2 X, E:T(1), CD3(L), 0.65 ng/mL, 72 h). Ctrl, the initial distribution of CD19 expression in the target cell population. (e–f) the effects of effector and target cell density on target cell depletion (%). Dots indicated the effect and target cell densities in healthy human organs. White color, 15% target cell depletion. BM, bone marrow; LN, lymph nodes; SP, spleen; Remainder, all the rest of the non-lymphoid organs. Initial setup: CD3(L), CD19(M) for (e), CD19 (M/20) for (f), 0.65 ng/mL, 72 h.

Figure 5.

Figure 5—figure supplement 1. The effects of ET ratio (a), cell density (b), and binding affinity (c) on CD19 evolution.

Figure 5—figure supplement 1.

The in vitro model was applied to perform simulations. Dashed line, the threshold value of CD19 expression for 15% target cell depletion within 72 h (initial setup: 2 X, E:T(1), CD3(L), 0.65 ng/mL, 72 h). Ctrl, initial CD19 distribution in the target cell population. Kd, dissociation constant; 2 X, 2 × 106 total cells/mL; E:T(1), E:T ratio = 1; CD3(L), CD3 expression (Low); CD19(M), CD19 expression (medium); 0.65, 20 ng/mL, blinatumomab concentration; 72 h, incubation duration.
Figure 5—figure supplement 2. The effects of effector and target cell density on target cell depletion (%) at 72 h.

Figure 5—figure supplement 2.

Simulations were performed by the in vitro model. Dots indicated the effect and target cell densities in healthy human organs. White color, 15% target cell depletion. BM, bone marrow; LN, lymph nodes; SP, spleen; Remainder, all the rest of non-lymphoid organs. Initial setup: CD3(L), CD19(M) for a, CD19 (M×4) for b, CD19 (M/4) for c, CD19 (M/20) for d, 0.65 ng/mL, 72 h.
Figure 5—figure supplement 3. Different effects of CD19 on cellular and molecular processes.

Figure 5—figure supplement 3.

(a–b) Different effects of target cell density and CD19 expression on target cell depletion amount (a) and fraction (b). The in vitro model was used in the simulation. Initial setup: CD3(L), effector cell (1 X), target cell (X/20 to 500 X), CD19 expression (M/20 to 500 M), 0.65 ng/mL, 72 h. Total CD19 in the system was jointly influenced by CD19 expression per target cell and target cell density. CD19 expression influenced cell lysis to a similar extent as target cell density when both factors were low (e.g. CD19 expression <M, target cell density <X in (a)). However, further increase of CD19 expression on cell membrane did not further improve cell lysis (e.g. from point A to point C), indicating maximum ternary complexes at each interface have been reached. In contrast, the increase of target cell density continuously promotes cell lysis, e.g., from point A to B, due to enhanced probability of cell-cell encounter. When the target cell density reaches extremely high (>50 X), cell lysis started to decrease, resulting from fewer cell-cell adhesion events due to insufficient bispecific T cell engager (BiTE) concentration. As shown, although total CD19 in the system at point B and C are identical (10 × M × X), different cell lysis level is yielded, supporting different effects of CD19 on cellular and molecular processes. (c) Effects of B cell density and CD19 expression on B cell depletion in vivo. In each group, the change of total CD19 density from the reference (1 X, 200 B cells/μL × 30,000 CD19/cell) was achieved through changing B cell density (red bar) or CD19 expression (gray bar). Their effects on B cell depletion amount were simulated by the in vivo model. Initial setup for reference (1 X): T cell (200 /μL), B cell (200 /μL), CD3 (50, 000 /cell), CD19 (30, 000 /cell), 0.73 ng/mL, 72 h. Different effects of CD19 on cellular and molecular processes have been confirmed by the in vivo model. Similarly, the increase of CD19 expression within low-level range (3 × 103–3 × 105 CD19/cell from 0.1X to 10X) constantly improved cell depletion, as ternary complexes formation at each interface increased with CD19 expression. By contrast, a bidirectional effect was shown by increasing B cell density, owing to enhanced probability of cell-cell encounter at the population level and then insufficient BiTE concentration. Herein, B cell density at 10 X (2000 /μL) in the blood indicates extremely high organ B cell density in the model, e.g., spleen (~9 × 108 /mL) and lymph nodes (~3 × 108 /mL).