Background Upregulation of inhibitory checkpoint molecules (ICM) on T-Cells and their ligands on AML blasts may be a mechanism of AML relapse after allogeneic stem cell transplantation (SCT). Better understanding of relapse biology may improve treatment efficacy.
Materials and Methods We examined peripheral blood (PB) and bone marrow (BM) samples of 5 AML patients (PTs) relapsing after SCT and PB from 5 healthy individuals, including 2 stem cells donors. ICM (PD-1, CTLA-4)/ligand (CD86, PD-L1, PD-L2) expression on T-cells and blasts was assessed by flow cytometry. PTs’ PB was cultivated with ‘KitM’ (GM-CSF+PGE-1) and without (Ctr) to generate leukemia-derived dendritic cells (DCleu), followed by MLC enriched with PTs’/donors’ T-Cells. After MLC, immune activation and functionality (degranulation, intracellular cytokine production, blast lysis) was assessed.
Results All PTs showed high expression of PD-1 on T-Cells, additional overexpression of CTLA-4 correlated negatively with responses to relapse treatment. Expression of ICM was low on T-Cells of 4/5 healthy individuals.
Influence of KitM on ICM/ligand expression:
a. ICM/ligand expression on uncultured T-Cells/blasts : Contrary to H, PTs presented high expressions of CTLA-4 and PD-1 on PB T-Cells. PTs also showed high frequencies of PB/BM blasts expressing CD86.
b. DC/DCleu in PB : Generation of DCleu in AML and generation of DC in H was successful with KitM pretreated PB vs. Ctr.
c. ICM expression on T-Cells after MLC : MLC of KitM treated PB enriched with unstimulated PTs’ T-Cells resulted in reduced frequencies of ICM-positive T-Cells in 3/5 PTs and increased frequencies of activated (leukemia specific) T-Cells in 3/5 PTs. Blast lysis was improved in 4/5 samples treated with KitM vs. Ctr.
Possible impact of ICM on clinical outcome (case study): PT1suffered early relapses after 2 SCTs from her healthy father. A role of ICM in AML relapse was suggested by CTLA-4/PD-1 expression on her T-Cells and CD86 expression on her blasts. Also, >90% of the father’s T-Cells expressed CTLA-4/PD-1, which might have contributed to treatment failure. In contrast, T-Cells from PT1’s mother presented with low ICM levels, suggesting that she may have been a better donor. Stimulation of PT1’s PB cells with KitM generated DCleu, decreased ICM expression and increased T-Cell activity. KitM pretreated samples showed improved blast lysis after MLC.
Conclusions Concisely, T-cells and blasts of AML PTs relapsing after SCT uniformly expressed ICM and their ligands, possibly leading to inferior immune responses.High aberrant ICM expression on donor T-Cells (particularly CTLA-4) may be a reason for relapse after SCT by inhibiting antileukemic immune reactions. Further, generation of DCleu through KitM triggers immune responses in MLC along reduced ICM expression on T-Cells, possibly reducing their inhibitory effects and thereby improving antileukemic responses.
Disclosure Information S. Bohlscheid: None. G. Filipini Velazquez: None. H. Aslan: None. T. Baudrexler: None. H. Schmetzer: E. Ownership Interest (stock, stock options, patent or other intellectual property); Significant; Modiblast Pharma GmbH. C. Schmid: None.
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