MYC Inhibition and TP53 Activation Synergistically Suppress Immune Checkpoint Ligand PD-L1 Expression in AML Cells

• Published in: Blood 2016-12, Vol.128 (22), p.1667-1667
• Main Authors: Bland, Joshua B., Tse, William T.
• Format: Article
• Language: English
• ISSN: 0006-4971; 1528-0020
• DOI: 10.1182/blood.V128.22.1667.1667

Expression of immune checkpoint ligands is a mechanism that many tumors use to escape attack by host immune cells. PD-L1, the ligand for checkpoint receptor PD-1 on T cells, is often expressed on tumor cells. Engagement of PD-1 on T cells by PD-L1 on tumor cells attenuates T-cell receptor signaling and suppresses anti-tumor response. PD-1 and PD-L1 blocking antibodies have been implemented clinically as treatment for many cancers, but the pattern of PD-L1 expression on AML is not well characterized. To answer this question, we studied how PD-L1 expression on AML is regulated under in vitro conditions that simulate the leukemia-host microenvironment. We examined surface expression of PD-L1 by flow cytometry on 4 AML lines, THP-1, KG1, KG1a, HL60, and a CML line, K562. Under basal conditions, these lines expressed no or low levels of PDL1. The AML cells were then subjected to conditions that mimic the leukemia-host microenvironment. AML cells were stained with the green fluorescent dye CFSE and co-cultured with Ficoll-separated PMNCs from healthy donors. After a day of co-culture, expression of PD-L1 was analyzed on AML cells and PD-1, CD25 and CD69 activation markers on PMNCs. Only a small increase of PD-L1, up to 2-4 fold, was seen on AML cells under this condition. To simulate the pro-inflammatory milieu in the tumor microenvironment, anti-CD3/CD28 microbeads were then added in culture to activate T cells. We observed a marked up-regulation of PD-L1 on AML cells, up to 5-60 fold, plus prominent expression of PD-1, CD25 and CD69 on T cells. These findings were confirmed by an alternative method of T cell activation in which AML cells were first coated with an anti-CD123 antibody, linked to anti-CD3/CD28 antibodies via a biotin-streptavidin bridge, and then cultured with PMNCs. To test whether pro-inflammatory cytokines were the sole inducers of PD-L1 expression, AML cells were treated with IFN-γ or TNF-α alone. IFN-γ treatment enhanced PD-L1 expression by 2-10 fold, while TNF-α showed a