Figure 2.

TIL cultures had variable proliferation and terminal differentiation.

1 × 10⁶ T cells were activated with CD3/28 Dynabeads and IL-2 (600 IU/ml). (a) TIL proliferation rates are demonstrated for individual patients designated by patient #number. (b) TIL proliferation rates are compared with allogeneic donor PBMCs (n = 4 PBMC cultures, 6 TIL cultures. p < 0.0001 by one-way analysis of variance). (c) TIL#11, #20 and allogeneic PBMCs were thawed, activated using CD3/28 beads, labeled with CTV and cultured in complete media with IL-2 (600 IU/ml). Cells were then analyzed for CTV fluorescence daily until day 7 for TIL or day 6 for PBMC. TIL#11 and #20 were cultured with IL-2 and IL-7 (10 ng/ml) for comparison. Cell proliferation, as seen from CTV dilution over time, was similar irrespective of whether IL-2 alone or IL-2+IL-7 was used in culture (results of a single experiment). (d) CD4⁺, and (e) CD8⁺, demonstrate TIL subtypes for TIL#11 with and without the addition of IL-7 to media. Cells were gated on morphology, viability, CD3⁺, CD4⁺ or CD8⁺. Naïve T cells: CD45RA⁺ CCR7⁺; T_EMRA: CD45RA⁺ CCR7⁻; T_CM: CD45RA⁻ CCR7⁺; T_EM: CD45RA⁻ CCR7⁻. Similar proportions of cell subsets were observed in IL-2- and IL-2/IL-7-supplemented cultures (results from a single experiment).

CTV, cell trace violet; PBMC, peripheral blood mononuclear cell; T_CM, central memory T cell; T_EM, effector memory T cell; T_EMRA, effector memory T cell expressing CD45RA; TIL, tumor-infiltrating lymphocyte.