MicroRNA‑425 inhibits proliferation of chronic lymphocytic leukaemia cells through regulation of the Bruton's tyrosine kinase/phospholipase Cγ2 signalling pathway

Chen,Jianying; Li,Yuhua; Xie,Xiaoling

doi:10.3892/etm.2020.8771

MicroRNA‑425 inhibits proliferation of chronic lymphocytic leukaemia cells through regulation of the Bruton's tyrosine kinase/phospholipase Cγ2 signalling pathway

Authors:
- Jianying Chen
- Yuhua Li
- Xiaoling Xie
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Affiliations: Department of Rheumatology, Hunan Provincial People's Hospital, Changsha, Hunan 410012, P.R. China, Department of Hematology, Zhujiang Hospital, Southern Medical University, Guangzhou, Guangdong 510006, P.R. China
Published online on: May 19, 2020 https://doi.org/10.3892/etm.2020.8771
Pages: 1169-1175

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Abstract

The present study aimed to investigate the effects of microRNA (miR)-425 on the proliferation of chronic lymphocytic leukaemia (CLL) cells and the possible underlying mechanisms. The expression of miR‑425 was determined in the B lymphocytes of CLL patients and in normal B lymphocytes by reverse transcription‑quantitative polymerase chain reaction (RT‑qPCR). In addition, MEC‑1 cells transfected with miR‑425 negative control (NC) or miR‑425 mimic were examined. The cell proliferation of different groups was evaluated using an MTT assay, and cell cycle distribution was evaluated using flow cytometry analysis. A dual‑luciferase reporter assay was used to verify whether Bruton's tyrosine kinase (BTK) was a target of miR‑425. Furthermore, the expression levels of BTK, phospholipase Cγ2 (PLCγ2), Ki‑67 and proliferating cell nuclear antigen (PCNA) were determined by RT‑qPCR and western blotting. The results revealed that the expression of miR‑425 was significantly downregulated in B lymphocytes obtained from CLL patients as compared with that in normal B lymphocytes. When cells were transfected with miR‑425 mimic, the proliferation of MEC‑1 cells was significantly inhibited at 24, 48 and 72 h compared with the proliferation of control cells. Additionally, the ratio of G0/G1 cells was significantly increased and the ratio of G2/M cells was significantly decreased in miR‑425‑overexpressing cells compared with that in control cells. The luciferase reporter assay revealed that miR‑425 binds to the 3'‑untranslated region of BTK mRNA. Finally, BTK, PLCγ2, Ki‑67 and PCNA expression was significantly inhibited at the mRNA and protein level in cells where miR‑425 was upregulated. In conclusion, miR‑425 inhibits the proliferation of MEC‑1 cells, potentially by inhibiting BTK/PLCγ2 signalling, and Ki‑67 and PCNA expression levels. These results provide a deeper insight for understanding the development of CLL and suggest a potential novel target for the treatment of CLL patients.

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1	Byrd JC, Furman RR, Coutre SE, Flinn IW, Burger JA, Blum KA, Grant B, Sharman JP, Coleman M, Wierda WG, et al: Targeting BTK with ibrutinib in relapsed chronic lymphocytic leukemia. N Engl J Med. 369:32–42. 2013.PubMed/NCBI View Article : Google Scholar
2	Siegel R, DeSantis C, Virgo K, Stein K, Mariotto A, Smith T, Cooper D, Gansler T, Lerro C, Fedewa S, et al: Cancer treatment and survivorship statistics, 2012. CA Cancer J Clin. 62:220–241. 2012.PubMed/NCBI View Article : Google Scholar
3	Singh SP, Pillai SY, de Bruijn MJW, Stadhouders R, Corneth OBJ, van den Ham HJ, Muggen A, van IJcken W, Slinger E, Kuil A, et al: Cell lines generated from a chronic lymphocytic leukemia mouse model exhibit constitutive Btk and Akt signaling. Oncotarget. 8:71981–71995. 2017.PubMed/NCBI View Article : Google Scholar
4	Gribben JG and O'Brien S: Update on therapy of chronic lymphocytic leukemia. J Clin Oncol. 29:544–550. 2011.PubMed/NCBI View Article : Google Scholar
5	Seda V and Mraz M: B-cell receptor signalling and its crosstalk with other pathways in normal and malignant cells. Eur J Haematol. 94:193–205. 2015.PubMed/NCBI View Article : Google Scholar
6	Liu Y, Dong Y, Jiang QL, Zhang B and Hu AM: Bruton's tyrosine kinase: Potential target in human multiple myeloma. Leuk Lymphoma. 55:177–181. 2014.PubMed/NCBI View Article : Google Scholar
7	Yaktapour N, Meiss F, Mastroianni J, Zenz T, Andrlova H, Mathew NR, Claus R, Hutter B, Fröhling S, Brors B, et al: BRAF inhibitor-associated ERK activation drives development of chronic lymphocytic leukemia. J Clin Invest. 124:5074–5084. 2014.PubMed/NCBI View Article : Google Scholar
8	Rushworth SA, Bowles KM, Barrera LN, Murray MY, Zaitseva L and MacEwan DJ: BTK inhibitor ibrutinib is cytotoxic to myeloma and potently enhances bortezomib and lenalidomide activities through NF-κB. Cell Signal. 25:106–112. 2013.PubMed/NCBI View Article : Google Scholar
9	Barrientos J and Rai K: Ibrutinib: A novel Bruton's tyrosine kinase inhibitor with outstanding responses in patients with chronic lymphocytic leukemia. Leuk Lymphoma. 54:1817–1820. 2013.PubMed/NCBI View Article : Google Scholar
10	Hwang HW and Mendell JT: MicroRNAs in cell proliferation, cell death, and tumorigenesis. Br J Cancer. 94:776–780. 2006.PubMed/NCBI View Article : Google Scholar
11	Chang HM, Martinez NJ, Thornton JE, Hagan JP, Nguyen KD and Gregory RI: Trim71 cooperates with microRNAs to repress Cdkn1a expression and promote embryonic stem cell proliferation. Nat Commun. 3(923)2012.PubMed/NCBI View Article : Google Scholar
12	Kovaleva V, Mora R, Park YJ, Plass C, Chiramel AI, Bartenschlager R, Döhner H, Stilgenbauer S, Pscherer A, Lichter P and Seiffert M: miRNA-130a targets ATG2B and DICER1 to inhibit autophagy and trigger killing of chronic lymphocytic leukemia cells. Cancer Res. 72:1763–1772. 2012.PubMed/NCBI View Article : Google Scholar
13	Audrito V, Vaisitti T, Rossi D, Gottardi D, D'Arena G, Laurenti L, Gaidano G, Malavasi F and Deaglio S: Nicotinamide blocks proliferation and induces apoptosis of chronic lymphocytic leukemia cells through activation of the p53/miR-34a/SIRT1 tumor suppressor network. Cancer Res. 71:4473–4483. 2011.PubMed/NCBI View Article : Google Scholar
14	Gao Y, Yin Y, Xing X, Zhao Z, Lu Y, Sun Y, Zhuang Z, Wang M, Ji W and He Y: Arsenic-induced anti-angiogenesis via miR-425-5p-regulated CCM3. Toxicol Lett. 254:22–31. 2016.PubMed/NCBI View Article : Google Scholar
15	Liu L, Zhao Z, Zhou W, Fan X, Zhan Q and Song Y: Enhanced expression of miR-425 promotes esophageal squamous cell carcinoma tumorigenesis by targeting SMAD2. J Genet Genomics. 42:601–611. 2015.PubMed/NCBI View Article : Google Scholar
16	Nakagawa R, Muroyama R, Koike K, Saeki C, Ito S, Morimoto S, Goto K, Matsubara Y, Kato N and Zeniya M: Decreased mir-425 induced inflammatory cytokine production via N-RAS upregulation in CD4+ T cells of primary biliar cholangitis. J Hepatol. 64 (Suppl):S643–S644. 2016.
17	He B, Li T, Guan L, Liu FE, Chen XM, Zhao J, Lin S, Liu ZZ and Zhang HQ: CTNNA3 is a tumor suppressor in hepatocellular carcinomas and is inhibited by miR-425. Oncotarget. 7:8078–8089. 2016.PubMed/NCBI View Article : Google Scholar
18	Peng WZ, Ma R, Wang F, Yu J and Liu ZB: Role of miR-191/425 cluster in tumorigenesis and diagnosis of gastric cancer. Int J Mol Sci. 15:4031–4048. 2014.PubMed/NCBI View Article : Google Scholar
19	Fu Y, Xu X, Xue J, Duan W and Yi Z: Deregulated lncRNAs in B cells from patients with active tuberculosis. PLoS One. 12(e0170712)2017.PubMed/NCBI View Article : Google Scholar
20	Cai BL, Xu XF, Fu SM, Shen LL, Zhang J, Guan SM and Wu JZ: Nuclear translocation of MRP1 contributes to multidrug resistance of mucoepidermoid carcinoma. Oral Oncol. 47:1134–1140. 2011.PubMed/NCBI View Article : Google Scholar
21	Cai BL, Li Y, Shen LL, Zhao JL, Liu Y, Wu JZ, Liu YP and Yu B: Nuclear multidrug resistance-related protein 1 is highly associated with better prognosis of human mucoepidermoid carcinoma through the suppression of cell proliferation, migration and invasion. PLoS One. 11(e0148223)2016.PubMed/NCBI View Article : Google Scholar
22	Livak KJ and Schmittgen TD: Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) method. Methods. 25:402–408. 2001.PubMed/NCBI View Article : Google Scholar
23	Bottoni A, Rizzotto L, Lai TH, Liu C, Smith LL, Mantel R, Reiff S, El-Gamal D, Larkin K, Johnson AJ, et al: Targeting BTK through microRNA in chronic lymphocytic leukemia. Blood. 128:3101–3112. 2016.PubMed/NCBI View Article : Google Scholar
24	Kaderi MA, Kanduri M, Buhl AM, Sevov M, Cahill N, Gunnarsson R, Jansson M, Smedby KE, Hjalgrim H, Jurlander J, et al: LPL is the strongest prognostic factor in a comparative analysis of RNA-based markers in early chronic lymphocytic leukemia. Haematologica. 96:1153–1160. 2011.PubMed/NCBI View Article : Google Scholar
25	Ma J, Liu J, Wang Z, Gu X, Fan Y, Zhang W, Xu L, Zhang J and Cai D: NF-kappaB-dependent microRNA-425 upregulation promotes gastric cancer cell growth by targeting PTEN upon IL-1β induction. Mol Cancer. 13(40)2014.PubMed/NCBI View Article : Google Scholar
26	Feng F, Song T, Zhang T, Cui Y, Zhang G and Xiong Q: MiR-425-5p promotes invasion and metastasis of hepatocellular carcinoma cells through SCAI-mediated dysregulation of multiple signaling pathways. Oncotarget. 8:31745–31757. 2017.PubMed/NCBI View Article : Google Scholar
27	Di Leva G, Piovan C, Gasparini P, Ngankeu A, Taccioli C, Briskin D, Cheung DG, Bolon B, Anderlucci L, Alder H, et al: Estrogen mediated-activation of miR-191/425 cluster modulates tumorigenicity of breast cancer cells depending on estrogen receptor status. PLoS Genet. 9(e1003311)2013.PubMed/NCBI View Article : Google Scholar
28	Chen FF, Xiong Y, Peng Y, Gao Y, Qin J, Chu GY, Pang WJ and Yang GS: miR-425-5p inhibits differentiation and proliferation in porcine intramuscular preadipocytes. Int J Mol Sci. 18(pii: E2101)2017.PubMed/NCBI View Article : Google Scholar