Publikationsserver der Universitätsbibliothek Marburg

Titel:Functional analysis of non-viral miRNA replacement therapy in colon carcinoma in vitro and in vivo
Autor:Ibrahim, Ahmed Fawzy Moustafa
Weitere Beteiligte: Aigner, Achim (Prof. Dr.)
Veröffentlicht:2011
URI:https://archiv.ub.uni-marburg.de/diss/z2011/0273
URN: urn:nbn:de:hebis:04-z2011-02735
DOI: https://doi.org/10.17192/z2011.0273
DDC: Medizin
Titel (trans.):Funktionelle Analyse von Micro-RNAs Einschleusungs-Therapie in Kolonkarzinomen in vitro und in vivo
Publikationsdatum:2011-03-18
Lizenz:https://rightsstatements.org/vocab/InC-NC/1.0/

Dokument

Schlagwörter:
Small RNA, Small RNA, colon cancer, Colonkrebs

Summary:
MicroRNAs (miRNAs) are small RNA molecules with perfect or imperfect homology to their target mRNA. They have been shown to specifically interfere with the expression of their target protein(s). Moreover, microRNAs play an important role in the pathogenesis of cancer, where some microRNAs act as tumor suppressors by targeting lethal oncogenes, and others act as oncogenes by targeting suppressor genes. Thus, microRNAs can be involved directly or indirectly in important cellular processes such as proliferation, differentiation and apoptosis. Understanding the functions and importance of microRNAs could lead to the discovery of new strategies for controlling of cancer. This thesis was aiming at the possibility of using non-viral delivery systems for microRNAs into mammalian cell lines, and examined their efficacies. To this end, three different transfection reagents, linear Jet-PEI, branched PEI F25-LMW and INTERFERin, were used in vitro, either commercially available or prepared in our lab. The work was concentrating on colon cancer as a tumor model, and two representative cell lines, LS174t and HCT-116, were selected as a platform to conduct the experiments. The studies focused on miR-143 and miR-145, which had been implicated previously with cancer development and progression. It was found in miRNA transfection experiments that miR-143 and miR-145, which are downregulated in colon cancer, have the ability to inhibit the progression of the cancer through different regulatory mechanisms, proliferation and apoptosis. Some important genes, which play important roles in tumorigenesis and tumor progression, were found to be negatively regulated by both miR-143 and miR-145. More specifically, ERK5 (Mitogen Activated Protein Kinase) which plays an essential role in the proliferation of colon cancer was suppressed by miR-143 and miR-145. Likewise, c-Myc was negatively regulated by miR-145, but these results were only at the protein level, i.e., no significant effects were determined on c-Myc transcription. Furthermore, it was found that the^inhibitory effects were depending on several factors such as cell line, transfection reagent, time of incubation and concentration of miRNA complex. Thus, the cellular delivery of microRNAs miR-143 and miR-145 showed profound inhibitory effects on the proliferation and soft agar colony formation of LS174t and HCT- 116 cells, and induced apoptosis in LS174t cells. Moreover, miR-145 was examined for its tumor inhibitory effect in vivo. To this end, athymic nude mice bearing s.c. colon carcinoma xenografts were treated with PEIcomplexed miR-145 by systemic or local injection. Profound anti-tumor effects upon PEImediated miR-145 delivery were found which were again dependent on the downregulation of specific target genes. Taken together, microRNA replacement therapy may represent a promising approach in tumor treatment, and this thesis establishes the PEI-mediated therapeutic delivery of miR- 145 in colon carcinoma.

Bibliographie / References

  1. MICHAEL ET AL. (2003). Reduced accumulation of specific microRNAs in colorectal neoplasia. Mol Cancer Res 1, 882-891.
  2. MONTERO, J.C., OCANA, A., ABAD, M., ORTIZ-RUIZ, M.J., PANDIELLA, A., and ESPARIS-OGANDO, A. (2009). Expression of Erk5 in early stage breast cancer and association with disease free survival identifies this kinase as a potential therapeutic target. PLoS One 4, e5565.
  3. SOHN, S.J., SARVIS, B.K., CADO, D., and WINOTO, A. (2002). ERK5 MAPK regulates embryonic angiogenesis and acts as a hypoxia-sensitive repressor of vascular endothelial growth factor expression. J Biol Chem 277, 43344-43351.
  4. VOORHOEVE, P.M., LE SAGE, C., SCHRIER, M., GILLIS, A.J., STOOP, H., NAGEL, R., LIU, Y.P., VAN DUIJSE, J., DROST, J., GRIEKSPOOR, A., ZLOTORYNSKI, E., YABUTA, N., DE VITA, G., NOJIMA, H., LOOIJENGA, L.H., and AGAMI, R. (2006). A genetic screen implicates miRNA-372 and miRNA-373 as oncogenes in testicular germ cell tumors. Cell 124, 1169-1181.
  5. MILNE, A.N., SITARZ, R., CARVALHO, R., CARNEIRO, F., and OFFERHAUS, G.J. (2007). Early onset gastric cancer: on the road to unraveling gastric carcinogenesis.
  6. MOORE, M.D., MCGARVEY, M.J., RUSSELL, R.A., CULLEN, B.R., and MCCLURE, M.O. (2005). Stable inhibition of hepatitis B virus proteins by small interfering RNA expressed from viral vectors. J Gene Med 7, 918-925.
  7. Many roads to maturity: microRNA biogenesis pathways and their regulation. Nat Cell Biol 11, 228-234.
  8. WANG, X., TANG, S., LE, S.Y., LU, R., RADER, J.S., MEYERS, C., and ZHENG, Z.M. (2008). Aberrant expression of oncogenic and tumor-suppressive microRNAs in cervical cancer is required for cancer cell growth. PLoS One 3, e2557.
  9. SHI, B., SEPP-LORENZINO, L., PRISCO, M., LINSLEY, P., DEANGELIS, T., and BASERGA, R. (2007). Micro RNA 145 targets the insulin receptor substrate-1 and inhibits the growth of colon cancer cells. J Biol Chem 282, 32582-32590.
  10. A.1-Prof.Dr. Frank Czubayko czubayko@staff.uni-marburg.de, Tel.: 06421 / 2865003 Fax: 06421 / 2865600
  11. A.2-Prof.Dr. Achim Aigner aigner@staff.uni-marburg.de, Tel.: 06421 / 2862262 Fax: 06421 / 2865600
  12. MULLOY, R., SALINAS, S., PHILIPS, A., and HIPSKIND, R.A. (2003). Activation of cyclin D1 expression by the ERK5 cascade. Oncogene 22, 5387-5398.
  13. SLABY, O., SVOBODA, M., FABIAN, P., SMERDOVA, T., KNOFLICKOVA, D., BEDNARIKOVA, M., NENUTIL, R., and VYZULA, R. (2007). Altered expression of miR-21, miR-31, miR-143 and miR-145 is related to clinicopathologic features of colorectal cancer. Oncology 72, 397-402.
  14. NAGATA, S. (1997). Apoptosis by death factor. Cell 88, 355-365.
  15. MOURRAIN, P., BECLIN, C., ELMAYAN, T., FEUERBACH, F., GODON, C., MOREL, J.B., JOUETTE, D., LACOMBE, A.M., NIKIC, S., PICAULT, N., REMOUE, K., SANIAL, M., VO, T.A., and VAUCHERET, H. (2000). Arabidopsis SGS2 and SGS3 genes are required for posttranscriptional gene silencing and natural virus resistance. Cell 101, 533-542.
  16. SU, L.K., VOGELSTEIN, B., and KINZLER, K.W. (1993). Association of the APC tumor suppressor protein with catenins. Science 262, 1734-1737.
  17. WIDLAK, P., LI, P., WANG, X., and GARRARD, W.T. (2000). Cleavage preferences of the apoptotic endonuclease DFF40 (caspase-activated DNase or nuclease) on naked DNA and chromatin substrates. J Biol Chem 275, 8226-8232.
  18. WIGHTMAN, L., KIRCHEIS, R., ROSSLER, V., CAROTTA, S., RUZICKA, R., KURSA, M., and WAGNER, E. (2001). Different behavior of branched and linear polyethylenimine for gene delivery in vitro and in vivo. J Gene Med 3, 362-372.
  19. MORIGUCHI, T., KAWACHI, K., KAMAKURA, S., MASUYAMA, N., YAMANAKA, H., MATSUMOTO, K., KIKUCHI, A., and NISHIDA, E. (1999). Distinct domains of mouse dishevelled are responsible for the c-Jun N-terminal kinase/stress- activated protein kinase activation and the axis formation in vertebrates. J Biol Chem 274, 30957-30962.
  20. ـــــــــــــ ــــــــ ـــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــــ ـــ 115 METZLER, M., WILDA, M., BUSCH, K., VIEHMANN, S., and BORKHARDT, A. (2004). High expression of precursor microRNA-155/BIC RNA in children with Burkitt lymphoma. Genes Chromosomes Cancer 39, 167-169.
  21. WINGARD. (1928). Hosts and symptoms of ring spot, a virus disease of plants. J Agric Res 37, 127-153.
  22. VANDENBOOM LI, T.G.L., Y.PHILIP, P. A.SARKAR, F. H. (2008). MicroRNA and Cancer: Tiny Molecules with Major Implications. Curr Genomics 9, 97-109.
  23. NAM, E.J., YOON, H., KIM, S.W., KIM, H., KIM, Y.T., KIM, J.H., KIM, J.W., and KIM, S. (2008). MicroRNA expression profiles in serous ovarian carcinoma. Clin Cancer Res 14, 2690-2695.
  24. SOREIDE, K., SLEWA, A., STOKKELAND, P.J., VAN DIERMEN, B., JANSSEN, E.A., SOREIDE, J.A., BAAK, J.P., and KORNER, H. (2009). Microsatellite instability and DNA ploidy in colorectal cancer: potential implications for patients undergoing systematic surveillance after resection. Cancer 115, 271-282.
  25. Mutational analysis of the APC/beta-catenin/Tcf pathway in colorectal cancer. Cancer Res 58, 1130-1134.
  26. SIJEN, T., FLEENOR, J., SIMMER, F., THIJSSEN, K.L., PARRISH, S., TIMMONS, L., PLASTERK, R.H., and FIRE, A. (2001). On the role of RNA amplification in dsRNA-triggered gene silencing. Cell 107, 465-476.
  27. A.3-Prof.Dr. Timothy D. Plant plant@staff.uni-marburg.de, Tel.: 06421 / 2865038 Fax: 06421 / 2865600 B. National Research Center, Division of Pharmaceutical Sciences, Department of Chemistry Natural and Microbial products, Cairo, Egypt B.1-Prof.Dr. Ahmed I. Eldewany aieldewany_1@yahoo.com Tel : 0020101220046 Fax : 0020233370931
  28. STURGILL, T.W., and WU, J. (1991). Recent progress in characterization of protein kinase cascades for phosphorylation of ribosomal protein S6. Biochim Biophys Acta 1092, 350-357.
  29. REFERENCES A- Institute of Pharmacology and Toxicology, Philipps-Universität Marburg, Karl-von-Frisch- Straße 1, 35032 Marburg, Germany.
  30. URBAN-KLEIN, B., WERTH, S., ABUHARBEID, S., CZUBAYKO, F., and AIGNER, A. (2005). RNAi-mediated gene-targeting through systemic application of polyethylenimine (PEI)-complexed siRNA in vivo. Gene Ther 12, 461-466.
  31. SMARDON, A., SPOERKE, J.M., STACEY, S.C., KLEIN, M.E., MACKIN, N., and MAINE, E.M. (2000). EGO-1 is related to RNA-directed RNA polymerase and functions in germ-line development and RNA interference in C. elegans. Curr Biol 10, 169-178.
  32. VOGELSTEIN, B., FEARON, E.R., HAMILTON, S.R., KERN, S.E., PREISINGER, A.C., LEPPERT, M., NAKAMURA, Y., WHITE, R., SMITS, A.M., and BOS, J.L. (1988). Genetic alterations during colorectal-tumor development. N Engl J Med 319, 525-532.


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