Functional Analysis of microRNA in Multiple Myeloma

  • Maria Teresa Di MartinoEmail author
  • Nicola Amodio
  • Pierfrancesco Tassone
  • Pierosandro Tagliaferri
Part of the Methods in Molecular Biology book series (MIMB, volume 1375)


MicroRNAs (miRNAs) are short non coding RNAs that regulate the gene expression and play a relevant role in physiopathological mechanisms such as development, proliferation, death, and differentiation of normal and cancer cells. Recently, abnormal expression of miRNAs has been reported in most of solid or hematopoietic malignancies, including multiple myeloma (MM), where miRNAs have been found deeply dysregulated and act as oncogenes or tumor suppressors. Presently, the most recognized approach for definition of miRNA portraits is based on microarray profiling analysis. We here describe a workflow based on the identification of dysregulated miRNAs in plasma cells from MM patients based on Affymetrix technology. We describe how it is possible to search miRNA putative targets performing whole gene expression profile on MM cell lines transfected with miRNA mimics or inhibitors followed by luciferase reporter assay to analyze the specific targeting of the 3′untranslated region (UTR) sequence of a mRNA by selected miRNAs. These technological approaches are suitable strategies for the identification of relevant druggable targets in MM.


microRNA miRNA Microarray profiling miRNA replacement miRNA inhibition Transfection 


  1. 1.
    Anderson KC (2014) Multiple myeloma. Hematol Oncol Clin North Am 28:xi–xii. doi: 10.1016/j.hoc.2014.08.001 CrossRefPubMedGoogle Scholar
  2. 2.
    Tagliaferri P et al (2012) Promises and challenges of microRNA-based treatment of multiple myeloma. Current Cancer Drug Targets 12:838–846CrossRefPubMedPubMedCentralGoogle Scholar
  3. 3.
    Tassone P, Tagliaferri P (2012) Editorial: new approaches in the treatment of multiple myeloma: from target-based agents to the new era of microRNAs (dedicated to the memory of Prof. Salvatore Venuta). Curr Cancer Drug Targets 12:741–742CrossRefPubMedGoogle Scholar
  4. 4.
    Rossi M et al (2013) From target therapy to miRNA therapeutics of human multiple myeloma: theoretical and technological issues in the evolving scenario. Curr Drug Targets 14:1144–1149CrossRefPubMedGoogle Scholar
  5. 5.
    Rossi M et al (2014) MicroRNA and multiple myeloma: from laboratory findings to translational therapeutic approaches. Curr Pharm Biotechnol 15:459–467CrossRefPubMedGoogle Scholar
  6. 6.
    Misso G et al (2013) Emerging pathways as individualized therapeutic target of multiple myeloma. Expert Opin Biol Ther 13(Suppl 1):S95–S109. doi: 10.1517/14712598.2013.807338 CrossRefPubMedGoogle Scholar
  7. 7.
    Misso G et al (2014) Mir-34: a new weapon against cancer? Mol Ther Nucleic Acids 3:e194. doi: 10.1038/mtna.2014.47 CrossRefPubMedGoogle Scholar
  8. 8.
    Lionetti M et al (2013) Biological and clinical relevance of miRNA expression signatures in primary plasma cell leukemia. Clin Cancer Res 19:3130–3142. doi: 10.1158/1078-0432.CCR-12-2043 CrossRefPubMedGoogle Scholar
  9. 9.
    Lionetti M, Agnelli L, Lombardi L, Tassone P, Neri A (2012) MicroRNAs in the pathobiology of multiple myeloma. Curr Cancer Drug Targets 12:823–837CrossRefPubMedGoogle Scholar
  10. 10.
    Amodio N et al (2013) miR-29b induces SOCS-1 expression by promoter demethylation and negatively regulates migration of multiple myeloma and endothelial cells. Cell Cycle 12:3650–3662. doi: 10.4161/cc.26585 CrossRefPubMedPubMedCentralGoogle Scholar
  11. 11.
    Amodio N, Di Martino MT, Neri A, Tagliaferri P, Tassone P (2013) Non-coding RNA: a novel opportunity for the personalized treatment of multiple myeloma. Expert Opin Biol Ther 13(Suppl 1):S125–S137. doi: 10.1517/14712598.2013.796356 CrossRefPubMedGoogle Scholar
  12. 12.
    Amodio N et al (2012) DNA-demethylating and anti-tumor activity of synthetic miR-29b mimics in multiple myeloma. Oncotarget 3:1246–1258CrossRefPubMedPubMedCentralGoogle Scholar
  13. 13.
    Di Martino MT et al (2014) In vivo activity of miR-34a mimics delivered by stable nucleic acid lipid particles (SNALPs) against multiple myeloma. PloS One 9:e90005. doi: 10.1371/journal.pone.0090005 CrossRefPubMedPubMedCentralGoogle Scholar
  14. 14.
    Di Martino MT et al (2014) In vitro and in vivo activity of a novel locked nucleic acid (LNA)-inhibitor-miR-221 against multiple myeloma cells. PloS One 9:e89659. doi: 10.1371/journal.pone.0089659 CrossRefPubMedPubMedCentralGoogle Scholar
  15. 15.
    Di Martino MT et al (2012) Synthetic miR-34a mimics as a novel therapeutic agent for multiple myeloma: in vitro and in vivo evidence. Clin Cancer Res 18:6260–6270. doi: 10.1158/1078-0432.CCR-12-1708 CrossRefPubMedPubMedCentralGoogle Scholar
  16. 16.
    Leone E et al (2013) Targeting miR-21 inhibits in vitro and in vivo multiple myeloma cell growth. Clin Cancer Res 19:2096–2106. doi: 10.1158/1078-0432.CCR-12-3325 CrossRefPubMedPubMedCentralGoogle Scholar
  17. 17.
    Leotta M et al (2014) A p53-dependent tumor suppressor network is induced by selective miR-125a-5p inhibition in multiple myeloma cells. J Cell Physiol 229:2106–2116. doi: 10.1002/jcp.24669 CrossRefPubMedGoogle Scholar
  18. 18.
    Rossi M et al (2013) miR-29b negatively regulates human osteoclastic cell differentiation and function: implications for the treatment of multiple myeloma-related bone disease. J Cell Physiol 228:1506–1515. doi: 10.1002/jcp.24306 CrossRefPubMedGoogle Scholar
  19. 19.
    Scognamiglio I et al (2014) Transferrin-conjugated SNALPs encapsulating 2′-O-methylated miR-34a for the treatment of multiple myeloma. Biomed Res Int 2014:217365. doi: 10.1155/2014/217365 CrossRefPubMedPubMedCentralGoogle Scholar
  20. 20.
    Monroig PD, Chen L, Zhang S, Calin GA (2014) Small molecule compounds targeting miRNAs for cancer therapy. Adv Drug Deliv Rev. doi: 10.1016/j.addr.2014.09.002 Google Scholar
  21. 21.
    Amodio N et al (2012) miR-29b sensitizes multiple myeloma cells to bortezomib-induced apoptosis through the activation of a feedback loop with the transcription factor Sp1. Cell Death Dis 3:e436. doi: 10.1038/cddis.2012.175 CrossRefPubMedPubMedCentralGoogle Scholar
  22. 22.
    Di Martino MT et al (2013) In vitro and in vivo anti-tumor activity of miR-221/222 inhibitors in multiple myeloma. Oncotarget 4:242–255CrossRefPubMedPubMedCentralGoogle Scholar
  23. 23.
    Lionetti M et al (2009) Identification of microRNA expression patterns and definition of a microRNA/mRNA regulatory network in distinct molecular groups of multiple myeloma. Blood 114:e20–e26. doi: 10.1182/blood-2009-08-237495 CrossRefPubMedGoogle Scholar
  24. 24.
    Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) method. Methods 25:402–408. doi: 10.1006/meth.2001.1262 CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2015

Authors and Affiliations

  • Maria Teresa Di Martino
    • 1
    Email author
  • Nicola Amodio
    • 1
  • Pierfrancesco Tassone
    • 1
    • 2
  • Pierosandro Tagliaferri
    • 1
  1. 1.Department of Experimental and Clinical Medicine, T. Campanella Cancer CenterMagna Graecia University and Medical Oncology UnitCatanzaroItaly
  2. 2.Sbarro Institute for Cancer Research and Molecular Medicine, Center for Biotechnology, College of Science and TechnologyTemple UniversityPhiladelphiaUSA

Personalised recommendations