G-CSF Upregulates the Expression of Aquaporin-9 through C/EBP-Beta to Enhance the Cytotoxic Activity of Arsenic Trioxide to Acute Myeloid Leukemia Cells

Abstract Arsenic trioxide (ATO) is the first-line drug for the treatment of acute promyelocytic leukemia (APL). Aquaporin-9 (AQP9), a transmembrane transporter, is required for leukemia cells to uptake ATO. APL cells express high levels of AQP9, while other types of acute myeloid leukemia (AML) cells express much lower levels of AQP9, which limits the transportation and the cytotoxic activity of ATO in those types of leukemia. Recently, we found that granulocyte colony stimulating factor (G-CSF) can significantly upregulate the expression of AQP9 in AML cell lines (THP-1 and HL-60), and can significantly enhance the intracellular concentrations of ATO, when compared with the treatment with ATO alone. We also found that the combination of ATO and G-CSF inhibited the cell proliferation and induced cell apoptosis more significantly than the treatment with ATO or G-CSF alone. The overexpression of AQP9 with lentivirus in HL-60 or THP-1 cells significantly enhanced the cytotoxic activity of ATO, while the knock-down of AQP9 gene with small interfering RNA (siRNA) significantly attenuated the combination effect of G-CSF and ATO. Moreover, we found that the upregulation of AQP9 by G-CSF depends on the upregulation of C/EBP-beta, the transcription factor downstream of G-CSF. In conclusion, our study suggests that the pretreatment of G-CSF could significantly enhance the cytotoxic activity of ATO in AML cells through the upregulation of AQP9, and that the combination of G-CSF and ATO would be a potential therapeutic strategy for AML patients. Disclosures No relevant conflicts of interest to declare.

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RANKL Expressed by Acute Myeloid Leukemia Cells Impairs NK Cell-Mediated Immune Surveillance

Blood ◽

10.1182/blood.v116.21.2164.2164 ◽

2010 ◽

Vol 116 (21) ◽

pp. 2164-2164

Author(s):

Benjamin J Schmiedel ◽

Constantin M Wende ◽

Tina Baessler ◽

Carolin Scheible ◽

Stefan Wirths ◽

...

Keyword(s):

Acute Myeloid Leukemia ◽

Nk Cells ◽

Myeloid Leukemia ◽

Nk Cell ◽

Conflicts Of Interest ◽

Immune Surveillance ◽

Surface Protein ◽

Leukemia Cells ◽

Acute Myeloid Leukemia Cells ◽

Acute Myeloid

Abstract Abstract 2164 NK cells play an important role in tumor immunosurveillance, especially of leukemia. Their reactivity is governed by various activating and inhibitory molecules expressed by their targets including multiple members of the TNF family. The TNF family member Receptor Activator of NF-κB ligand (RANKL) and its receptors RANK and osteoprotegerin (OPG) are key regulators of bone remodelling, but recently have also been shown to influence progression of hematopoetic malignancies. Here we studied the yet unkown role of the RANK/RANKL molecule system in NK cells and their reactivity against acute myeloid leukemia (AML). Primary leukemia cells from AML patients were found to substantially express RANKL mRNA and surface protein in 75% of the investigated cases (n=40). Reverse signaling via surface-expressed RANKL into AML blasts induced the release of soluble factors including the immunoregulatory cytokines TNF and IL-10, which impaired NK cell anti-tumor reactivity. Moreover, we observed upregulation of RANK on NK cells among PBMC of healthy donors upon exposure to IL-10. This was not caused by direct effects on NK cells, but was rather due to yet unidentified factors released by monocytes among the PBMC upon IL-10 exposure and could be prevented by the activating cytokine IL-2. Furthermore, functional experiments with NK cells and RANKL transfectants or RANKL-negative controls revealed that forward signaling into RANK-expressing NK cells by tumor-expressed RANKL also directly impaired NK cytotoxicity and IFN-γ production. In line, blocking RANK-RANKL interaction using anti-RANKL antibodies or RANK-Fc fusion protein increased cytotoxicity and cytokine production of allogenic NK cells in cultures with RANKL-positive primary AML cells. Our data indicate that RANKL expression enables immune evasion of leukemia cells both by directly inhibiting reactivity of RANK-expressing NK cells and by orchestrating a reciprocal interplay between AML cells, monocytes and NK cells resulting in an immunosuppressive cytokine milieu. Thus, therapeutic modulation of the RANK/RANKL system, e.g. with Denosumab/AMG162, which is presently being evaluated for treatment of both non-malignant and malignant osteolysis, holds promise to reinforce NK reactivity against hematopoietic malignancies. Disclosures: No relevant conflicts of interest to declare.

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