Assessment of miR-21-5p, miR-451a, and miR-144-3p level in urine in differential diagnosis of localized prostate cancer

Background. Limited sensitivity and specificity of existing prostate cancer (PCa) diagnosis methods drive the search for new markers. A number of studies has demonstrated the potential for measuring expression of certain microRNAs in urine.

Aim. To evaluate the diagnostic potential of measuring microRNA expression in urine in PCa.

Materials and methods. A collection of urine sediment samples from 19 patients with benign prostatic hyperplasia and 44 patients with PCa was analyzed. RNA was isolated using the miRNEasy Serum/Plasma Kit. 16 µL of RNA isolated from each sample were converted into cDNA, which served as a template for real-time polymerase chain reaction. For sequencing, microRNA libraries were prepared using MGIEasy Small RNA Library Prep Kit v.2.0. The formed DNA nanoballs were placed into an MGI DNBSEQ-G400 sequencer. Sequencing results were processed using IsoMiRmap. Differences in microRNA abundance were analyzed using DESeq2. For miRNA-21, high-throughput sequencing data were corroborated by the results of quantitative real-time polymerase chain reaction.

Results. 1154 types of microRNA were identified, 11 were differentially represented in all comparison groups. The most significant differences in cell sediment between benign prostatic hyperplasia and PCa patients were recorded for miR-451a (area under the curve (AUC) 0.98). Additionally, the abundance levels of two microRNA isoforms were significantly different: hsa-miR-144-3p|-1 (AUC 0.96) and hsa-miR-21-5p|+4 (AUC 0.68).

Сonclusion. This study confirms that altered expression of microRNAs miR-21, miR-451a and miR-144-3p is associated  with PCa, can be detected in urine samples, and can also be a potential non-invasive diagnostic criterion.

1. Rawla P. Epidemiology of prostate cancer. World J Oncol 2019;10(2):63–89. DOI: 10.14740/wjon1191

2. Sung H., Ferlay J., Siegel R.L. et al. Global Cancer Statistics 2020: GLOBOCAN Estimates of Incidence and Mortality Worldwide for 36 Cancers in 185 Countries. CA Cancer J Clin 2021;71(3):209–49. DOI: 10.3322/caac.21660

3. Archer M., Dogra N., Kyprianou N. Inflammation as a driver of prostate cancer metastasis and therapeutic resistance. Cancers 2020;12:2984. DOI: 10.3390/cancers12102984

4. Prensner J.R., Rubin M.A., Wei J.T., Chinnaiyan A.M. Beyond PSA: the next generation of prostate cancer biomarkers. Sci Transl Med 2012;4(127):127rv123. DOI: 10.1126/scitranslmed.3003180

5. Draisma G., Etzioni R., Tsodikov A. et al. Lead time and overdiagnosis in prostate-specific antigen screening: importance of methods and context. J Natl Cancer Inst 2009;101(6):374–83. DOI: 10.1093/jnci/djp001

6. Carroll P.R., Parsons J.K., Andriole G. et al. NCCN guidelines insights: prostate cancer early detection, version 2.2016. J Natl Compr Cancer Netw 2016;14(5):509–19. DOI: 10.6004/jnccn.2016.0060

7. Wang W., Wang M., Wang L. et al. Diagnostic ability of %p2PSA and prostate health index for aggressive prostate cancer: a meta-analysis. Sci Rep 2014;4:5012. DOI: 10.1038/srep05012

8. Fradet Y., Saad F., Aprikian A. et al. Upm3, a new molecular urine test for the detection of prostate cancer. Urology 2004;64(2):311–5. DOI: 10.1016/j.urology.2004.03.052

9. Hessels D., Klein Gunnewiek J.M., van Oort I. et al. Dd3(pca3)-based molecular urine analysis for the diagnosis of prostate cancer. Eur Urol 2003;44(1):8–15. DOI: 10.1016/S0302-2838(03)00201-X

10. Nakanishi H., Groskopf J., Fritsche H.A. et al. PCA3 molecular urine assay correlates with prostate cancer tumor volume: Implication in selecting candidates for active surveillance. J Urol 2008;179(5):1804–9. DOI: 10.1016/j.juro.2008.01.013

11. Van Gils M.P., Cornel E.B., Hessels D. et al. Molecular PCA3 diagnostics on prostatic fluid. Prostate 2007;67(8):881–7. DOI: 10.1002/pros.20564

12. Cui Y., Cao W., Li Q. et al. Evaluation of prostate cancer antigen 3 for detecting prostate cancer: a systematic review and meta-analysis. Sci Rep 2016;6:25776. DOI: 10.1038/srep25776

13. He L., Hannon G.J. MicroRNAs: small RNAs with a big role in gene regulation. Nat Rev Genet 2004;5(7):522–31. DOI: 10.1038/nrg1379

14. Lewis H., Lance R., Troyer D. et al. miR-888 is an expressed prostatic secretions-derived microRNA that promotes prostate cell growth and migration. Cell Cycle 2014;13(2):227–39. DOI: 10.4161/cc.26984

15. Haj-Ahmad T.A., Abdalla M.A., Haj-Ahmad Y. Potential urinary miRNA biomarker candidates for the accurate detection of prostate cancer among benign prostatic hyperplasia patients. J Cancer 2014;5(3):182–91. DOI: 10.7150/jca.6799

16. Stuopelyte K., Daniunaite K., Bakavicius A. et al. The utility of urine-circulating miRNAs for detection of prostate cancer. Br J Cancer 2016;115(6):707–15. DOI: 10.1038/bjc.2016.233

17. Fredsøe J., Rasmussen A.K.I., Thomsen A.R. et al. Diagnostic and prognostic microRNA biomarkers for prostate cancer in cell-free urine. Eur Urol Focus 2018;4(6):825–33. DOI: 10.1016/j.euf.2017.02.018

18. Loher P., Karathanasis N., Londin E. et al. IsoMiRmap: fast, deterministic and exhaustive mining of isomiRs from short RNA-seq datasets. Bioinformatics 2021;37(13):1828–38. DOI: 10.1093/bioinformatics/btab016

19. Love M.I., Huber W., Anders S. Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2. Genome Biol 2014;15(12):550. DOI: 10.1186/s13059-014-0550-8

20. Pan X., Wang R., Wang Z.X. The potential role of miR-451 in cancer diagnosis, prognosis, and therapy. Mol Cancer Ther 2013;12(7):1153–62. DOI: 10.1158/1535-7163.MCT-12-0802

21. Kretov D.A., Walawalkar I.A., Mora-Martin A. et al. Ago2-dependent processing allows miR-451 to evade the global microRNA turnover elicited during erythropoiesis. Mol Cell 2020;78(2):317–28.e6. DOI: 10.1016/j.molcel.2020.02.020

22. Kohrs N., Kolodziej S., Kuvardina O.N. et al. MiR144/451 expression is repressed by RUNX1 during megakaryopoiesis and disturbed by RUNX1/ETO. PLoS Genet 2016;12(3):e1005946. DOI: 10.1371/journal.pgen.1005946

23. Azzouzi I., Moest H., Wollscheid B. et al. Deep sequencing and proteomic analysis of the microRNA-induced silencing complex in human red blood cells. Exp Hematol 2015;43(5):382–92. DOI: 10.1016/j.exphem.2015.01.007

24. Foley S.J., Soloman L.Z., Wedderburn A.W. et al. A prospective study of the natural history of hematuria associated with benign prostatic hyperplasia and the effect of finasteride. J Urol 2000;163(2):496–8. DOI: 10.1016/S0022-5347(05)67910-4

25. Ghandi M., Huang F.W., Jané-Valbuena J. et al. Next-generation characterization of the Cancer Cell Line Encyclopedia. Nature 2019;569(7757):503–8. DOI: 10.1038/s41586-019-1186-3

26. Liu Y., Yang H.Z., Jiang Y.J., Xu L.Q. MiR-451a is downregulated and targets PSMB8 in prostate cancer. Kaohsiung J Med Sci 2020;36(7):494–500. DOI: 10.1002/kjm2.12196

27. Sun X.B., Chen Y.W., Yao Q.S. et al. MicroRNA-144 suppresses prostate cancer growth and metastasis by targeting EZH2. Technol Cancer Res Treat 2021;20:1533033821989817. DOI: 10.1177/1533033821989817

28. Bai M., Lei Y., Wang M. et al. Long non-coding RNA SNHG17 promotes cell proliferation and invasion in castration-resistant prostate cancer by targeting the miR-144/CD51 axis. Front Genet 2020;11:274. DOI: 10.3389/fgene.2020.00274

29. Wang G., Yao L., Yang T. et al. MiR-451 suppresses the growth, migration, and invasion of prostate cancer cells by targeting macrophage migration inhibitory factor. Transl Cancer Res 2019;8(2):647–54. DOI: 10.21037/tcr.2019.03.28

30. Fan B., Jin X., Ding Q. et al. Expression of miR-451a in prostate cancer and its effect on prognosis. Iran J Public Health 2021;50(4):772–9. DOI: 10.18502/ijph.v50i4.6002

31. Cappellesso R., Tinazzi A., Giurici T. et al. Programmed cell death 4 and microRNA 21 inverse expression is maintained in cells and exosomes from ovarian serous carcinoma effusions. Cancer Cytopathol 2014;122(9):685–93. DOI: 10.1002/cncy.21442

32. Chan J.A., Krichevsky A.M., Kosik K.S. MicroRNA-21 is an antiapoptotic factor in human glioblastoma cells. Cancer Res 2005;65(14):6029–33. DOI: 10.1158/0008-5472.CAN-05-0137

33. Meng F., Henson R., Wehbe-Janek H. et al. MicroRNA-21 regulates expression of the PTEN tumor suppressor gene in human hepatocellular cancer. Gastroenterology 2007;133(2):647–58. DOI: 10.1053/j.gastro.2007.05.022

34. Venturutti L., Romero L.V., Urtreger A.J. et al. Stat3 regulates ErbB-2 expression and co-opts ErbB-2 nuclear function to induce miR-21 expression, PDCD4 downregulation and breast cancer metastasis. Oncogene 2016;35(17):2208–22. DOI: 10.1038/onc.2015.28

35. Chan J.K., Blansit K., Kiet T. et al. The inhibition of mir-21 promotes apoptosis and chemosensitivity in ovarian cancer. Gynecol Oncol 2014;132(3):739–44. DOI: 10.1016/j.ygyno.2014.01.034

36. Xue X., Liu Y., Wang Y. et al. Mir-21 and mir-155 promote non-small cell lung cancer progression by downregulating socs1, socs6, and pten. Oncotarget 2016;7(51):84508–19. DOI: 10.18632/oncotarget.13022

37. Wu Y., Song Y., Xiong Y. et al. MicroRNA-21 (miR-21) promotes cell growth and invasion by repressing tumor suppressor PTEN in colorectal cancer. Cell Physiol Biochem 2017;43(3):945–58. DOI: 10.1159/000481648

38. Asangani I.A., Rasheed S.A.K., Nikolova D.A. et al. MicroRNA-21 (miR-21) post-transcriptionally downregulates tumor suppressor Pdcd4 and stimulates invasion, intravasation and metastasis in colorectal cancer. Oncogene 2008;27(15):2128–36. DOI: 10.1038/sj.onc.1210856

39. Wang C., Peng R., Zeng M. et al. An autoregulatory feedback loop of miR-21/VMP1 is responsible for the abnormal expression of miR-21 in colorectal cancer cells. Cell Death Dis 2020;11(12):1067. DOI: 10.1038/s41419-020-03265-4

40. Melbø-Jørgensen C., Ness N., Andersen S. et al. Stromal expression of miR-21 predicts biochemical failure in prostate cancer patients with Gleason score 6. PLoS One 2014;9:e113039. DOI: 10.1371/journal.pone.0113039

41. Ribas J., Ni X., Haffner M. et al. MiR-21: an androgen receptor-regulated microRNA that promotes hormone-dependent and hormone-independent prostate cancer growth. Cancer Res 2009;69(18):7165–9. DOI: 10.1158/0008-5472.CAN-09-1448

42. Li T., Li R.S., Li Y.H. et al. MiR-21 as an independent biochemical recurrence predictor and potential therapeutic target for prostate cancer. J Urol 2012;187(4):1466–72. DOI: 10.1016/j.juro.2011.11.082

43. Porzycki P., Ciszkowicz E., Semik M., Tyrka M. Combination of three miRNA (miR-141, miR-21, and miR-375) as potential diagnostic tool for prostate cancer recognition. Int Urol Nephrol 2018;50(9):1619–26. DOI: 10.1007/s11255-018-1938-2

44. Yang B., Liu Z., Ning H. et al. MicroRNA-21 in peripheral blood mononuclear cells as a novel biomarker in the diagnosis and prognosis of prostate cancer. Cancer Biomark 2016;17(2):223–30. DOI: 10.3233/CBM-160634

45. Seputra K.P., Purnomo B.B., Susianti H. et al. miRNA-21 as reliable serum diagnostic biomarker candidate for metastatic progressive prostate cancer: meta-analysis approach. Med Arch 2021;75(5):347–50. DOI: 10.5455/medarh.2021.75.347-350

46. Ghorbanmehr N., Gharbi S., Korsching E. et al. miR-21-5p, miR-141-3p, and miR-205-5p levels in urine-promising biomarkers for the identification of prostate and bladder cancer. Prostate 2019;79(1):88–95. DOI: 10.1002/pros.23714

47. Danarto R., Astuti I., Umbas R., Haryana S.M. Urine miR-21-5p and miR-200c-3p as potential non-invasive biomarkers in patients with prostate cancer. Turk J Urol 2019;46(1):26–30. DOI: 10.5152/tud.2019.19163

48. Gunawan R.R., Astuti I., Danarto H.R. miRNA-21 as high potential prostate cancer biomarker in prostate cancer patients in Indonesia. Asian Pac J Cancer Prev 2023;24(3):1095–9. DOI: 10.31557/APJCP.2023.24.3.1095