Indian Journal of Clinical Biochemistry

, Volume 34, Issue 1, pp 26–38 | Cite as

Interrelation of the Circulating and Tissue MicroRNA-21 with Tissue PDCD4 Expression and the Invasiveness of Iraqi Female Breast Tumors

  • Meena M. Abdulhussain
  • Najat A. HasanEmail author
  • Alaa G. Hussain
Original Research Article


The changes in the translational repression and variation in mRNA degradation induced by micro RNA are important aspects of tumorigenesis. The association of microRNA-21 with clinicopathologic features and expression of programed cell death 4 (PDCD4) in Iraqi female’s with breast tumors has not been studied. MicroRNAs were extracted from a set of 60 breast tumor tissues and blood samples of females with breast cancer and benign breast lesions obtained after breast-reductive surgery, and only blood samples from 30 normal volunteers. These extracts were evaluated for miR-21 expression by quantitative RT-PCR. Analysis of PDCD4 protein expression was carried out as miR-21 target gene by immunohistochemical tests and correlating the results with patients’ clinicopathological features. Significant overexpression of miRNA-21 was found in breast cancer group. The fold increase in the miR-21 gene expression was significantly higher in circulating exosomes and breast tissues of breast cancer patients as compared to other groups (P < 0.001). Overexpression of miR-21 was also significantly associated with the advanced tumor stage and histological grade. In breast cancer patients, PDCD4 protein expression was decreased to about 70% of the level in the control group. The delta Ct of exosomal and breast tissue miRNA-21 was negatively associated with PDCD4 expression. In conclusion, the translational repression of the PDCD4 induced by the high expression of miR-21 promotes breast cell transformation and development of breast tumor, and circulating miR-21 level could be applied to the screening panels for early detection of women breast cancer.


Breast cancer Benign breast lesion Programed death cell 4 MicroRNA-21 



Authors would express their sincere appreciation to Professor Robert C. Benjamin, University of North Texas, USA for revising and editing this article prior to submission to the journal.

Author Contribution

Professor Dr. NAH has conceived, designed the experiments and share in writing the paper. Dr. MMA has collected the samples, performed the molecular experiments under the supervision of Professor Dr. NAH, contributed reagents/materials/analysis tools and analysed the data. Professor Dr. AGH has performed the immunohistochemical analyses and interpretation of the results.

Compliance with Ethical Standards

Conflict of interest

All authors declare that they have no conflict of interest.

Supplementary material

12291_2017_710_MOESM1_ESM.docx (2.6 mb)
Supplementary material 1 (DOCX 2652 kb)


  1. 1.
    International Agency for Research on Cancer: Early Detection and Prevention. Globocan 2012. Available from: Accessed 13 June 2017.
  2. 2.
    Alsaraj M, Alsaed SJ. Iraqi cancer registry 2011. Baghdad: Iraqi Cancer Board, Ministry of Health; 2011. p. 25–38.Google Scholar
  3. 3.
    Majid RA, Hassan HA, Muhealdeen DN, Mohammed HA, Hughson MD. Breast cancer in Iraq is associated with a unimodally distributed predominance of luminal type B over luminal type A, surrogates from young to old age. BMC Womens Health. 2017;17:27. Scholar
  4. 4.
    AL-Janabi AA, Naseer ZH, Hamody TA. Epidemiological study of cancers in Iraq-Karbala from 2008 to 2015. Intern J Med Res Health Sci. 2017;6(1):79-86Google Scholar
  5. 5.
    The Cancer Genome Atlas Network. Comprehensive molecular portraits of human breast tumors. Nature. 2012;490(7418):61–70. Scholar
  6. 6.
    Lu J, Getz G, Miska EA, Alvarez-Saavedra E, Lamb J, Peck D, et al. MicroRNA expression profiles classify human cancers. Nature. 2005;435(7043):834–8.CrossRefGoogle Scholar
  7. 7.
    Iorio MV, Croce CM. MicroRNA involvement in human cancer. Carcinogen. 2012;33(6):1126–33.CrossRefGoogle Scholar
  8. 8.
    Yanaihara N, Caplen N, Bowman E, Seike M, Kumamoto K, Yi M, et al. Unique microRNA molecular profiles in lung cancer diagnosis and prognosis. Cancer Cell. 2006;9(3):189–98.CrossRefGoogle Scholar
  9. 9.
    Ambros V. The functions of animal microRNAs. Nature. 2004;431:350–5.CrossRefGoogle Scholar
  10. 10.
    Qian B, Katsaros D, Lu L, Preti M, Durando A, Arisio R, et al. High miR-expression in breast cancer associated with poor disease-free survival in early stage disease and high TGF-beta1. Breast Cancer Res Treat. 2009;117:13–40.CrossRefGoogle Scholar
  11. 11.
    Sioud M, Cekaite L. Profiling of miRNA expression and prediction of target genes. Methods Mol Biol. 2010;629:257–71.PubMedGoogle Scholar
  12. 12.
    Si ML, Zhu S, Wu H, Lu Z, Wu F, Mo YY. MiR-21-mediated tumor growth. Oncogene. 2007;26:2799–803.CrossRefGoogle Scholar
  13. 13.
    Zhu S, Si ML, Wu H, Mo YY. MicroRNA-21 targets the tumor suppressor gene tropomyosin 1 (TPM1). J Biol Chem. 2007;282:14328–36.CrossRefGoogle Scholar
  14. 14.
    Frankel LB, Christoffersen NR, Jacobsen A, Lindow M, Krogh A, Lund AH. Programmed cell death 4 (PDCD4) is an important functional target of the microRNA miR-21 in breast cancer cells. J Biol Chem. 2008;283:1026–33.CrossRefGoogle Scholar
  15. 15.
    Yan LX, Huang XF, Shao Q, Huang MY, Deng L, Wu QL, et al. MicroRNA miR-21 overexpression in human breast cancer is associated with advanced clinical stage, lymph node metastasis and patient poor prognosis. RNA. 2008;14:2348–60.CrossRefGoogle Scholar
  16. 16.
    Asangani IA, Rasheed SA, Nikolova DA, Leupold JH, Colburn NH, Post S, et al. MicroRNA-21(miR-21) post-transcriptionally downregulates tumor suppressor Pdcd4 and stimulates invasion, intravasation and metastasis in colorectal cancer. Oncogene. 2008;27:2128–36.CrossRefGoogle Scholar
  17. 17.
    Lu Z, Liu M, Stribinskis V, Klinge CM, Ramos KS, Colburn NH, et al. MicroRNA-21 promotes cell transformation by targeting the programmed cell death 4 gene. Oncogene. 2008;27:4373–9.CrossRefGoogle Scholar
  18. 18.
    Zhu S, Wu H, Wu F, Nie D, Sheng S, Mo YY. MicroRNA-21 targets tumor suppressor genes in invasion and metastasis. Cell Res. 2008;18:350–9.CrossRefGoogle Scholar
  19. 19.
    Schwarzenbach H, Hoon DS, Pantel K. Cell-free nucleic acids as biomarkers in cancer patients. Nat Rev Cancer. 2011;11:426–37.CrossRefGoogle Scholar
  20. 20.
    Simpson RJ, Lim JW, Moritz RL, Mathivanan S. Exosomes: proteomic insights and diagnostic potential. Expert Rev Proteomics. 2009;6:267–83.CrossRefGoogle Scholar
  21. 21.
    Valadi H, Ekstrom K, Bossios A, Sjostrand M, Lee JJ, Lotvall JO. Exosome-mediated transfer of mRNAs and microRNAs is a novel mechanism of genetic exchange between cells. Nat Cell Biol. 2007;9:654–9.CrossRefGoogle Scholar
  22. 22.
    Edge SB, Compton CC. The American Joint Committee on Cancer: the 7th edition of the AJCC cancer staging manual and the future of TNM. Ann Surg Oncol. 2010;17(6):1471–4. Scholar
  23. 23.
    Yang M, Chen J, Su F, Yu B, Lin L, Liu Y, et al. Microvesicles secreted by macrophages shuttle invasion-potentiating microRNAs into breast cancer cells. Mol Cancer. 2011;10:117. Scholar
  24. 24.
    Alwan, N. Iraqi initiative of a regional comparative breast cancer research project in the Middle East. J Cancer Biol Res. 2014. Available from: Accessed 13 June 2017.
  25. 25.
    AL-wasiti EA, Hasan NA, AL-Salhi AR. Evaluation of markers of oxidative DNA damage in females with breast tumors. Iraqi J Med Sci. 2010;8(1):51–65.Google Scholar
  26. 26.
    Abed Oun MA, El-Yssin HD, Al-Alwan NA. Prevalence of soluble fas protein in breast cancer patients: correlation with the clinico-pathological parameter. Iraqi Postgr Med J. 2016;15:107–17.Google Scholar
  27. 27.
    DeSantis C, Siegel R, Bandi P, Jemal A. breast cancer statistics, 2011. CA Cancer J Clin. 2011;61(6):409–18.CrossRefGoogle Scholar
  28. 28.
    Ravdin PM, Cronin KA, Howlader N, Berg CD, Chlebowski RT, Feuer EJ, et al. The decrease in breast-cancer incidence in 2003 in the United States. N Engl J Med. 2007;356(16):1670–4.CrossRefGoogle Scholar
  29. 29.
    Leong SP, Shen ZZ, Liu TJ, Agarwal G, Tajima T, Paik NS, et al. Is breast cancer the same disease in Asian and Western countries? World J Surg. 2010;34(10):2308–24.CrossRefGoogle Scholar
  30. 30.
    Asaga S, Kuo C, Nguyen T, Terpenning M, Giuliano AE, Hoon DS. Direct serum assay for microRNA-21 concentrations in early and advanced breast cancer. Clin Chem. 2011;57(1):84–91.CrossRefGoogle Scholar
  31. 31.
    Mar-Aguilar F, Mendoza-Ramíreza JA, Malagón-Santiago I, Espino-Silva PK, Santuario-Facio SK, Ruiz-Flores P, et al. Serum circulating microRNA profiling for identification of potential breast cancer biomarkers. Dis Markers. 2013;34:163–9.CrossRefGoogle Scholar
  32. 32.
    Han JG, Jiang YD, Zhang CH, Yang YM, Pang D, Song YN, et al. A novel panel of serum miR-21/miR-155/miR-365 as a potential diagnostic biomarker for breast cancer. Ann Surg Treat Res. 2017;92(2):55–66.CrossRefGoogle Scholar
  33. 33.
    Chen H, Liu H, Zou H, Chen R, Dou Y, Sheng S, et al. Evaluation of plasma miR-21 and miR-152 as diagnostic biomarkers for common types of human cancers. J Cancer. 2016;7(5):490–9.CrossRefGoogle Scholar
  34. 34.
    Abdel-Hamid NR, Mohammed EA, Abbas AH, Badr FM. MicroRNA-21 expression in primary breast cancer tissue among Egyptian female patients and its correlation with chromosome 17 aneusomy. Mol Diagn Ther. 2015;19(6):365–73.CrossRefGoogle Scholar
  35. 35.
    De Mattos-Arruda L, Bottai G, Nuciforo PG, Di Tommaso L, Giovannetti E, Peg V, et al. MicroRNA-21 links epithelial-to-mesenchymal transition and inflammatory signals to confer resistance to neoadjuvant trastuzumab and chemotherapy in HER2-positive breast cancer patients. Oncotarget. 2015;6(35):37269–80.PubMedPubMedCentralGoogle Scholar
  36. 36.
    Yan LX, Liu YH, Xiang JW, Wu QN, Xu LB, Luo XL, et al. PIK3R1 targeting by miR-21 suppresses tumor cell migration and invasion by reducing PI3 K/AKT signaling and reversing EMT, and predicts clinical outcome of breast cancer. Int J Oncol. 2016;48(2):471–84.CrossRefGoogle Scholar
  37. 37.
    Toraih EA, Mohammed EA, Farrag S, Ramsis N, Hosny S. Pilot study of serum MicroRNA-21 as a diagnostic and prognostic biomarker in Egyptian breast cancer patients. Mol Diagn Ther. 2015;19:179–90.CrossRefGoogle Scholar
  38. 38.
    Li X, Xin S, Yang D, Li X, He Z, Che X, et al. Down-regulation of PDCD4 expression is an independent predictor of poor prognosis in human renal cell carcinoma patients. J Cancer Res Clin Oncol. 2012;138(3):529–35.CrossRefGoogle Scholar
  39. 39.
    Hornick N, Huan J, Goloviznina NA, Potter A, Kurre P. Hypoxia regulates exosomal microrna content, trafficking and function of key elements in the AML microenvironment. Blood. 2013. Accessed 13 July 2017.
  40. 40.
    Niu Z, Goodyear SM, Rao S, Wu X, Tobias JW, Avarbock MR, et al. MicroRNA-21 regulates the self-renewal of mouse spermatogonial stem cells. Proc Natl Acad Sci USA. 2011;108:12740–5.CrossRefGoogle Scholar
  41. 41.
    Yang W, Lee DY, Ben-David Y. The roles of microRNAs in tumorigenesis and angiogenesis. Int J Physiol Pathophysiol Pharmacol. 2011;3(2):140–55.PubMedGoogle Scholar
  42. 42.
    Gao Y, Cai Q, Huang Y, Li S, Yang H, Sun L, et al. MicroRNA-21 as a potential diagnostic biomarker for breast cancer patients: a pooled analysis of individual studies. Oncotarget. 2016;7(23):34498–506.PubMedPubMedCentralGoogle Scholar
  43. 43.
    Gao J, Zhang Q, Xu J, Guo L, Li X. Clinical significance of serum miR-21 in breast cancer compared with CA153 and CEA. Chin J Cancer Res. 2013;25(6):743–8.PubMedPubMedCentralGoogle Scholar
  44. 44.
    Badr FM. Potential role of miR-21 in breast cancer diagnosis and therapy. JSM Biotechnol Bioeng. 2016;3(5):1068.Google Scholar
  45. 45.
    Leupold JH, Yang HS, Colburn NH, Asangani I, Post S, Allgayer H. Tumor suppressor Pdcd4 inhibits invasion/intravasation and regulates urokinase receptor (u-PAR) gene expression via Sp-transcription factors. Oncogene. 2007;26:4550–62.CrossRefGoogle Scholar
  46. 46.
    Wen YH, Shi X, Chiriboga L, Matsahashi S, Yee H, Afonja O. Alterations in the expression of PDCD4 in ductal carcinoma of the breast. Oncol Rep. 2007;18(6):1387–93.PubMedGoogle Scholar
  47. 47.
    Kumar N, Wethkamp N, Waters LC, Carr MD, Klempnauer K-H. Tumor suppressor protein Pdcd4 interacts with Daxx and modulates the stability of Daxx and the Hipk2-dependent phosphorylation of p53 at serine 46. Oncogenesis. 2013;. Scholar
  48. 48.
    Modelska A, Turro E, Russell R, Beaton J, Sbarrato T, Spriggs K, et al. The malignant phenotype in breast cancer is driven by eIF4A1-mediated changes in the translational landscape. Cell Death Dis. 2015;. Scholar
  49. 49.
    Böhm M, Sawicka K, Siebrasse JP, Brehmer-Fastnacht A, Peters R, Klempnauer K-H. The transformation suppressor protein Pdcd4 shuttles between nucleus and cytoplasm and binds RNA. Oncogene. 2003;22:4905–10.CrossRefGoogle Scholar
  50. 50.
    Feng YH, Tsao CJ. Emerging role of microRNA-21 in cancer. Biomed Rep. 2016;5(4):395–402.CrossRefGoogle Scholar
  51. 51.
    Qi L, Bart J, Tan LP, Platteel I, Sluis TVD, Huitema S, et al. Expression of miR-21 and its targets (PTEN, PDCD4, TM1) in flat epithelial atypia of the breast in relation to ductal carcinoma in situ and invasive carcinoma. BMC Cancer. 2009;9:163. Scholar

Copyright information

© Association of Clinical Biochemists of India 2017

Authors and Affiliations

  • Meena M. Abdulhussain
    • 1
  • Najat A. Hasan
    • 1
    Email author
  • Alaa G. Hussain
    • 2
  1. 1.Department of Chemistry and Biochemistry, College of MedicineAlnahrain UniversityBaghdadIraq
  2. 2.Department of Clinical Pathology, College of MedicineAlnahrain UniversityBaghdadIraq

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