MicroRNA‑34b‑5p inhibits proliferation, stemness, migration and invasion of retinoblastoma cells via Notch signaling

Zhang,Shurong; Cui,Zhe

doi:10.3892/etm.2021.9686

MicroRNA‑34b‑5p inhibits proliferation, stemness, migration and invasion of retinoblastoma cells via Notch signaling

Authors:
- Shurong Zhang
- Zhe Cui
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Affiliations: Department of Ophthalmology, The Third Affiliated Hospital of Qiqihar Medical University, Qiqihar, Heilongjiang 161000, P.R. China
Published online on: January 25, 2021 https://doi.org/10.3892/etm.2021.9686
Article Number: 255
Copyright: © Zhang et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

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Abstract

Retinoblastoma (RB) is one of the most common forms of childhood intraocular cancer. While the occurrence of RB is traditionally associated with dysregulation of the RB1 gene, efforts have been made to assess the role of several other pathways that may result in RB. The Notch signaling pathway has been identified as one of the sentinel pathways in retinal development and has been indicated to serve as a tumor suppressor. However, epigenetic modifications of the Notch signaling pathway, and their consequences on tumor establishment and progression, have received little attention. The present study attempted to elucidate the microRNA (miR)‑mediated dysregulation of the Notch signaling pathway and its implications on tumor initiation. Upon recruitment of patients with RB (age, 4‑25 months), the levels of miR‑34b‑5p were determined in tumor and adjacent healthy tissues. Simultaneously, the serum levels of miR‑34b‑5p were measured in tumor and healthy samples using reverse transcriptase‑quantitative PCR (RT‑qPCR). Binding of miR‑34b‑5p to Notch1 and Notch2 were confirmed bioinformatically. In vitro studies were performed in Y79 and Weri‑Rb‑1 RB cell lines. The cell lines were transfected with miR‑34b‑5p constructs and miR‑34b‑5p overexpression was confirmed using RT‑qPCR. The impact of miR‑34b‑5p overexpression on cell growth and cancer stemness markers (Sox‑2, Nanog, and CD133) was examined. The expression levels of Notch1 and Notch2 were evaluated in the presence of miR‑34b‑5p. The rescue of cell growth and cancer stemness phenotypes was evaluated by co‑transfection of miR‑34b‑5p with Notch1 or Notch2. The results of the present study indicated that the expression levels of miR‑34b‑5p were reduced in patient tissues and serum samples compared with those in healthy tissues and samples. Notch1 and Notch2 expression level was negatively correlated with the expression level of miR‑34b‑5p. Overexpression of miR‑34b‑5p resulted in reduced cell proliferation, migration, invasion and cancer stemness compared with the control group. Further in vivo experiments confirmed the inhibitory effects of miR‑34b‑5p on RB cell proliferation. Upon co‑transfection of miR‑34b‑5p with Notch1 or Notch2, these phenotypes were rescued with reversal of cell growth and tumor sphere formation. Collectively, the results indicated that miR‑34b‑5p functions as a tumor suppressor in RB via regulating the Notch signaling pathway. Therefore, miR‑34b‑5p may be explored for its utility as a therapeutic target in RB.

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1	Dimaras H, Kimani K, Dimba EA, Gronsdahl P, White A, Chan HS and Gallie BL: Retinoblastoma. Lancet. 379:1436–1446. 2012.PubMed/NCBI View Article : Google Scholar
2	Fabian ID and Sagoo MS: Understanding retinoblastoma: Epidemiology and genetics. Community Eye Health. 31(7)2018.PubMed/NCBI
3	Knudson AG Jr, Meadows AT, Nichols WW and Hill R: Chromosomal deletion and retinoblastoma. N Engl J Med. 295:1120–1123. 1976.PubMed/NCBI View Article : Google Scholar
4	Chinnam M and Goodrich DW: RB1, development, and cancer. Curr Top Dev Biol. 94:129–169. 2011.PubMed/NCBI View Article : Google Scholar
5	Ishak CA, Coschi CH, Roes MV and Dick FA: Disruption of CDK-resistant chromatin association by pRB causes DNA damage, mitotic errors, and reduces Condensin II recruitment. Cell Cycle. 16:1430–1439. 2017.PubMed/NCBI View Article : Google Scholar
6	Mastrangelo D, De Francesco S, Di Leonardo A, Lentini L and Hadjistilianou T: Retinoblastoma epidemiology: Does the evidence matter? Eur J Cancer. 43:1596–1603. 2007.PubMed/NCBI View Article : Google Scholar
7	Dimaras H, Khetan V, Halliday W, Orlic M, Prigoda NL, Piovesan B, Marrano P, Corson TW, Eagle RC Jr, Squire JA and Gallie BL: Loss of RB1 induces non-proliferative retinoma: Increasing genomic instability correlates with progression to retinoblastoma. Hum Mol Genet. 17:1363–1372. 2008.PubMed/NCBI View Article : Google Scholar
8	Dimaras H and Gallie BL: The p75 NTR neurotrophin receptor is a tumor suppressor in human and murine retinoblastoma development. Int J Cancer. 122:2023–2029. 2008.PubMed/NCBI View Article : Google Scholar
9	Dimaras H, Coburn B, Pajovic S and Gallie BL: Loss of p75 neurotrophin receptor expression accompanies malignant progression to human and murine retinoblastoma. Mol Carcinog. 45:333–343. 2006.PubMed/NCBI View Article : Google Scholar
10	Kandalam M, Mitra M, Subramanian K and Biswas J: Molecular pathology of retinoblastoma. Middle East Afr J Ophthalmol. 17:217–223. 2010.PubMed/NCBI View Article : Google Scholar
11	Castro-Magdonel BE, Orjuela M, Camacho J, García-Chéquer AJ, Cabrera-Muñoz L, Sadowinski-Pine S, Durán-Figueroa N, Orozco-Romero MJ, Velázquez-Wong AC, Hernández-Ángeles A, et al: miRNome landscape analysis reveals a 30 miRNA core in retinoblastoma. BMC Cancer. 17(458)2017.PubMed/NCBI View Article : Google Scholar
12	Xiao W, Gao Z, Duan Y, Yuan W and Ke Y: Notch signaling plays a crucial role in cancer stem-like cells maintaining stemness and mediating chemotaxis in renal cell carcinoma. J Exp Clin Cancer Res. 36(41)2017.PubMed/NCBI View Article : Google Scholar
13	Xiao W, Chen X and He M: Inhibition of the Jagged/Notch pathway inhibits retinoblastoma cell proliferation via suppressing the PI3K/Akt, Src, p38MAPK and Wnt/β-catenin signaling pathways. Mol Med Rep. 10:453–458. 2014.PubMed/NCBI View Article : Google Scholar
14	Chen CY, Chen YY, Hsieh MS, Ho CC, Chen KY, Shih JY and Yu CJ: Expression of Notch gene and its impact on survival of patients with resectable non-small cell lung cancer. J Cancer. 8:1292–1300. 2017.PubMed/NCBI View Article : Google Scholar
15	Mollen EWJ, Ient J, Tjan-Heijnen VCG, Boersma LJ, Miele L, Smidt ML and Vooijs MAGG: Moving Breast Cancer Therapy up a Notch. Front Onco. 8(518)2018.PubMed/NCBI View Article : Google Scholar
16	Baker A, Wyatt D, Bocchetta M, Li J, Filipovic A, Green A, Peiffer DS, Fuqua S, Miele L, Albain KS and Osipo C: Notch-1-PTEN-ERK1/2 signaling axis promotes HER2+ breast cancer cell proliferation and stem cell survival. Oncogene. 37:4489–4504. 2018.PubMed/NCBI View Article : Google Scholar
17	Sun Q, Wang R, Wang Y, Luo J, Wang P and Cheng B: Notch1 is a potential therapeutic target for the treatment of human hepatitis B virus X protein-associated hepatocellular carcinoma. Oncol Rep. 31:933–939. 2014.PubMed/NCBI View Article : Google Scholar
18	Venkatesh V, Nataraj R, Thangaraj GS, Karthikeyan M, Gnanasekaran A, Kaginelli SB, Kuppanna G, Kallappa CG and Basalingappa KM: Targeting Notch signalling pathway of cancer stem cells. Stem Cell Investig. 5(5)2018.PubMed/NCBI View Article : Google Scholar
19	Takebe N, Nguyen D and Yang SX: Targeting notch signaling pathway in cancer: Clinical development advances and challenges. Pharmacol Ther. 141:140–149. 2014.PubMed/NCBI View Article : Google Scholar
20	Fabbri G, Holmes AB, Viganotti M, Scuoppo C, Belver L, Herranz D, Yan XJ, Kieso Y, Rossi D, Gaidano G, et al: Common nonmutational NOTCH1 activation in chronic lymphocytic leukemia. Proc Natl Acad Sci USA. 114:E2911–E2919. 2017.PubMed/NCBI View Article : Google Scholar
21	Lee H, Kim KR, Cho NH, Hong SR, Jeong H, Kwon SY, Park KH, An HJ, Kim TH, Kim I, et al: MicroRNA expression profiling and Notch1 and Notch2 expression in minimal deviation adenocarcinoma of uterine cervix. World J Surg Oncol. 12(334)2014.PubMed/NCBI View Article : Google Scholar
22	Maroof H, Islam F, Dong L, Ajjikuttira P, Gopalan V, McMillan NAJ and Lam AK: Liposomal delivery of miR-34b-5p induced cancer cell death in thyroid carcinoma. Cells. 7(265)2018.PubMed/NCBI View Article : Google Scholar
23	Singh L and Kashyap S: Update on pathology of retinoblastoma. Int J Ophthalmol. 11:2011–2016. 2018.PubMed/NCBI View Article : Google Scholar
24	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
25	Phillips PC, Levow C, Catterall M, Colvin OM, Pastan I and Brem H: Transforming growth factor-alpha-Pseudomonas exotoxin fusion protein (TGF-alpha-PE38) treatment of subcutaneous and intracranial human glioma and medulloblastoma xenografts in athymic mice. Cancer Res. 54:1008–1015. 1994.PubMed/NCBI
26	Zhao D and Cui Z: MicroRNA-361-3p regulates retinoblastoma cell proliferation and stemness by targeting hedgehog signaling. Expe Ther Med. 17:1154–1162. 2019.PubMed/NCBI View Article : Google Scholar
27	Jadhav AP, Mason HA and Cepko CL: Notch 1 inhibits photoreceptor production in the developing mammalian retina. Development. 133:913–923. 2006.PubMed/NCBI View Article : Google Scholar
28	Zhang L, Liao Y and Tang L: MicroRNA-34 family: A potential tumor suppressor and therapeutic candidate in cancer. J Exp Clin Cancer Res. 38(53)2019.PubMed/NCBI View Article : Google Scholar
29	Misso G, Di Martino MT, De Rosa G, Farooqi AA, Lombardi A, Campani V, Zarone MR, Gullà A, Tagliaferri P, Tassone P and Caraglia M: Mir-34: A new weapon against cancer? Mol Ther Nucleic Acids. 3(e194)2014.PubMed/NCBI View Article : Google Scholar
30	Engkvist ME, Stratford EW, Lorenz S, Meza-Zepeda LA, Myklebost O and Munthe E: Analysis of the miR-34 family functions in breast cancer reveals annotation error of miR-34b. Sci Rep. 7(9655)2017.PubMed/NCBI View Article : Google Scholar
31	Jiang L and Hermeking H: miR-34a and miR-34b/c suppress intestinal tumorigenesis. Cancer Res. 77:2746–2758. 2017.PubMed/NCBI View Article : Google Scholar
32	Majid S, Dar AA, Saini S, Shahryari V, Arora S, Zaman MS, Chang I, Yamamura S, Tanaka Y, Chiyomaru T, et al: miRNA-34b inhibits prostate cancer through demethylation, active chromatin modifications, and AKT pathways. Clin Cancer Res. 19:73–84. 2013.PubMed/NCBI View Article : Google Scholar
33	Li Z, Luo Q, Xu H, Zheng M, Abdalla BA, Feng M, Cai B and Zhang X, Nie Q and Zhang X: MiR-34b-5p suppresses melanoma differentiation-associated gene 5 (MDA5) signaling pathway to promote avian leukosis virus subgroup J (ALV-J)-infected cells proliferaction and ALV-J replication. Front Cell Infect Microbiol. 7(17)2017.PubMed/NCBI View Article : Google Scholar
34	Agarwal V, Bell GW, Nam JW and Bartel DP: Predicting effective microRNA target sites in mammalian mRNAs. Elife. 4(e05005)2015.PubMed/NCBI View Article : Google Scholar
35	Kakuda S, LoPilato RK, Ito A and Haltiwanger RS: Canonical Notch ligands and Fringes have distinct effects on NOTCH1 and NOTCH2. J Biol Chem. 295:14710–14722. 2020.PubMed/NCBI View Article : Google Scholar
36	Joshi I, Minter LM, Telfer J, Demarest RM, Capobianco AJ, Aster JC, Sicinski P, Fauq A, Golde TE and Osborne BA: Notch signaling mediates G1/S cell-cycle progression in T cells via cyclin D3 and its dependent kinases. Blood. 113:1689–1698. 2009.PubMed/NCBI View Article : Google Scholar
37	Dai MY, Fang F, Zou Y, Yi X, Ding YJ, Chen C, Tao ZZ and Chen SM: Downregulation of Notch1 induces apoptosis and inhibits cell proliferation and metastasis in laryngeal squamous cell carcinoma. Oncol Rep. 34:3111–3119. 2015.PubMed/NCBI View Article : Google Scholar
38	Abel EV, Kim EJ, Wu J, Hynes M, Bednar F, Proctor E, Wang L, Dziubinski ML and Simeone DM: The Notch pathway is important in maintaining the cancer stem cell population in pancreatic cancer. PLoS One. 9(e91983)2014.PubMed/NCBI View Article : Google Scholar