Cytokine working group study of lymphodepleting chemotherapy, interleukin-2, and granulocyte-macrophage colony-stimulating factor in patients with metastatic melanoma: clinical outcomes and peripheral-blood cell recovery

Abstract

Purpose: Recovery of lymphocyte populations after lymphocyte depletion is implicated in therapeutic immune pathways in animal models and in patients with cancer. We sought to evaluate the effects of chemotherapy-induced lymphodepletion followed by granulocyte-macrophage colony-stimulating factor (GM-CSF) and high-dose interleukin-2 (IL-2) therapy on clinical response and the recovery of lymphocyte subcompartments in patients with metastatic melanoma.

Patients and methods: This was a two-stage phase II trial design. Patients with measurable metastatic melanoma were treated with intravenous cyclophosphamide (60 mg/kg, days 1 and 2) and fludarabine (25 mg/m(2), day 3 through 7) followed by two 5-day courses of intravenous high-dose bolus IL-2 (600,000 U/kg; days 8 through 12 and 21 through 25). GM-CSF (250 microg/m(2)/d beginning day 8) was given until granulocyte recovery. Lymphocyte recovery profiles were determined by flow cytometric phenotyping at regular intervals, and clinical outcome was assessed by Response Evaluation Criteria in Solid Tumors (RECIST).

Results: The trial was stopped at the end of stage 1 with four of 18 objective responses noted. Twelve patients had detailed lymphocyte subcompartments evaluated. After lymphodepletion, we observed an induction of regulatory cells (CD4+ T regulatory cells; CD8+ T suppressor cells) and of T memory cells (CD8+ T central memory cells; T effector memory RA+ cells). Expansion of circulating melanoma-specific CD8(+) cells was observed in one of four HLA-A2-positive patients.

Conclusion: Chemotherapy-induced lymphodepletion modulates the homeostatic repopulation of the lymphocyte compartment and influences recovering lymphocyte subpopulations. Clinical activity seems similar to standard high-dose aldesleukin alone.

Fig 1.

Peripheral-blood cell count recovery in 12 patients. (A) Leukocyte recovery. (B) Peripheral-blood platelet count recovery. LYMPH, lymphocytes; GRAN, granulocytes; PLT, platelets.

Fig 2.

Lymphocyte recovery: x-axis represents day of treatment from day 0. Days 8 and 11 were excluded because of insufficient cells to do subset phenotyping for most patients. y-axis represents cell numbers for each graph. The solid blue circles in each graph represent the number of cells for each patient, and the solid blue vertical lines represent the 95% CIs. The symbols represent statistically significant difference from baseline value: (*) .001 < P ≤ .01; (†) .0001 < P ≤ .001; (‡) P ≤ .0001. The panels represent absolute lymphocytes, absolute CD4⁺ cells, absolute CD8⁺ cells, CD4/CD8 ratio, number of T_Reg cells, T Eff cells; number of T effector memory CD8⁺/CD45RO⁺/CCR7⁺ cells, and T_Reg/CD8 T Eff cells ratio.

Fig 3.

Dot plots from flow cytometric analysis of tetramer (Tet) -positive (A, C, E) pretreatment and (B, D, F) post-treatment CD8 cells. One of four HLA-A2-positive patients had a treatment related shift in tetramer-positive cells. CD8⁺ cells are on the y-axis; tetramer-positive cells are on the x axis. (A, B) Mart 1; (C, D) gp100; (E, F) tyrosinase.

Fig 4.

Kaplan-Meier estimates for progression-free and overall survival. (Survival with 95% CI and progression-free survival with 95% CI.)