Ex vivo enrichment of circulating anti-tumor T cells from both cutaneous and ocular melanoma patients: clinical implications for adoptive cell transfer therapy

Tumor-infiltrating lymphocytes (TILs) have been successfully used for adoptive
cell transfer (ACT) immunotherapy; however, due to their scarce availability,
this therapy is possible for a limited fraction of cutaneous melanoma patients.
We assessed whether an effective protocol for ex vivo T-cell expansion from
peripheral blood mononuclear cells (PBMCs), suitable for ACT of both cutaneous
and ocular melanoma patients, could be identified. PBMCs from both cutaneous and
ocular melanoma patients were stimulated in vitro with autologous, irradiated
melanoma cells (mixed lymphocyte tumor cell culture; MLTCs) in the presence of
IL-2 and IL-15 followed by the rapid expansion protocol (REP). The functional
activity of these T lymphocytes was characterized and compared with that of TILs.
In addition, the immune infiltration in vivo of ocular melanoma lesions was
analyzed. An efficient in vitro MLTC expansion of melanoma reactive T cells was
achieved from all PBMC's samples obtained in 7 cutaneous and ocular metastatic
melanoma patients. Large numbers of melanoma-specific T cells could be obtained
when the REP protocol was applied to these MLTCs. Most MLTCs were enriched in
non-terminally differentiated T(EM) cells homogeneously expressing co-stimulatory
molecules (e.g., NKG2D, CD28, CD134, CD137). A similar pattern of anti-tumor
activity, in association with a more variable expression of co-stimulatory
molecules, was detected on short-term in vitro cultured TILs isolated from the
same patients. In these ocular melanoma patients, we observed an immune
infiltrate with suppressive characteristics and a low rate of ex vivo growing
TILs (28.5% of our cases). Our MLTC protocol overcomes this limitation, allowing
the isolation of T lymphocytes with effector functions even in these patients.
Thus, anti-tumor circulating PBMC-derived T cells could be efficiently isolated
from melanoma patients by our novel ex vivo enrichment protocol. This protocol
appears suitable for ACT studies of cutaneous and ocular melanoma patients.