J Cancer 2019; 10(27):6767-6778. doi:10.7150/jca.32167

Research Paper

Fenretinide-induced Apoptosis of Acute Myeloid Leukemia Cells via NR4A1 Translocation into Mitochondria and Bcl-2 Transformation

Jie Xiong1,2, Xingyi Kuang2, Tingting Lu2, Xu Liu3, Bingqing Cheng2, Weili Wang2, Danna Wei2, Xinyao Li2, Zhaoyuan Zhang2, Qin Fang4, Depei Wu1✉, Jishi Wang2✉

1. Jiangsu Institute of Hematology, The First Affiliated Hospital of Soochow University, Key Laboratory of Thrombosis and Hemostasis under Ministry of Health, Collaborative Innovation Center of Hematology, Suzhou Institute of Blood and Marrow Transplantation,188 Shizi Street, Suzhou 215006, Jiangsu, China
2. Department of Hematology, The Affiliated Hospital of Guizhou Medical University. Hematopoietic Stem Cell Transplantation Center of Guizhou Province, Key Laboratory of Hematological Disease Diagnostic & Treat Centre of Guizhou Province. Guizhou Medical University, Guiyang 550001, China.
3. Department of Critical Care Medicine, Affiliated Hospital of Guizhou Medical University, Guiyang 550001, China
4. Department of Pharmacy, Affiliated Hospital of Guizhou Medical University, Guiyang 550001, China.

This is an open access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/). See http://ivyspring.com/terms for full terms and conditions.
Citation:
Xiong J, Kuang X, Lu T, Liu X, Cheng B, Wang W, Wei D, Li X, Zhang Z, Fang Q, Wu D, Wang J. Fenretinide-induced Apoptosis of Acute Myeloid Leukemia Cells via NR4A1 Translocation into Mitochondria and Bcl-2 Transformation. J Cancer 2019; 10(27):6767-6778. doi:10.7150/jca.32167. Available from http://www.jcancer.org/v10p6767.htm

File import instruction

Abstract

OBJECTIVE: Fenretinide is reported to induce NR4A1-associated apoptosis in several types of cancer cells. However, it remains unclear about its specific role and the underlying mechanism in acute myeloid leukemia (AML). Therefore, this study aimed to explore the role and mechanism of fenretinide-induced apoptosis in AML.

METHOD: Firstly, the NR4A1 mRNA level in the newly diagnosed AML patients was measured, then AML cells were treated with fenretinide at various time points and doses, and cell viability was investigated by using the cell-counting kit-8 (CCK-8) assay. Additionally, apoptosis and cell cycles were analyzed by using flow cytometry. Moreover, siNR4A1 was utilized to knockdown NR4A1 expression, and leptomycin B (LMB) was adopted to inhibit the nuclear export; afterwards, the apoptosis rate and expression of apoptotic proteins in AML cells were detected. In addition, the expression levels of NR4A1 in the nuclei and mitochondria of fenretinide-treated AML cells were also measured. Meanwhile, the interaction between NR4A1 and Bcl-2, as well as the Bcl-2 transformation, was also examined. The anti-leukemic effect of fenretinide on NOD/SCID mice was also determined through subcutaneous injection of HL-60 cells.

RESULTS: NR4A1 expression in AML patients was markedly down-regulated compared with that in normal donors. Fenretinide induced the expression of NR4A1 and mitochondria-mediated apoptotic pathway-associated proteins in a time- and concentration-dependent manner. Importantly, both siNR4A1 alone or the combination of fenretinide with LMB could attenuate the fenretinide-induced apoptosis and expression of apoptotic proteins. Under the action of fenretinide, the NR4A1 protein expression was down-regulated in nuclear extracts whereas up-regulated in mitochondrial extracts. At the same time, fenretinide promoted NR4A1 translocation from nuclei into mitochondria, and enhanced the interaction between NR4A1 and Bcl-2, thereby exposing the BH3 domain of Bcl-2 to exert the anti-apoptotic effect. Moreover, fenretinide also exhibited an anti-leukemic effect and induced NR4A1 expression in the AML mouse model.

CONCLUSIONS: Fenretinide exerts an obvious effect on AML cells both in vitro and in vivo. Besides, the NR4A1-mediated signaling pathway is highly involved in the fenretinide-induced apoptosis of AML cells.

Keywords: NR4A1, Fenretinide, Acute myeloid leukemia, Apoptosis, Nuclear export