Skip to main content
SpringerLink
Log in
Book cover

Therapeutic Oligonucleotides pp 317–335Cite as

Inhibition of Human Papillomavirus Expression Using DNAzymes

  • Protocol
  • First Online:

Part of the book series: Methods in Molecular Biology ((MIMB,volume 764))

Abstract

Deoxyribozymes (DXZs) are catalytic oligodeoxynucleotides capable of performing diverse functions including the specific cleavage of a target RNA. These molecules represent a new type of therapeutic oligonucleotides combining the efficiency of ribozymes and the intracellular endurance and simplicity of modified antisense oligonucleotides. Commonly used DXZs include the 8–17 and 10–23 motifs, which have been engineered to destroy disease-associated genes with remarkable efficiency. Targeting DXZs to disease-associated transcripts requires extensive biochemical testing to establish target RNA accessibility, catalytic efficiency, and nuclease sensibility. The usage of modified nucleotides to render nuclease-resistance DXZs must be counterweighted against deleterious consequences on catalytic activity. Further intracellular testing is required to establish the effect of microenvironmental conditions on DXZ activity and off-target issues. Application of modified DXZs to cervical cancer results in specific growth inhibition, cell death, and apoptosis. Thus, DXZs represent a highly effective antisense moiety with minimal secondary effects.

This is a preview of subscription content, log in via an institution.

Protocol
USD   49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD   89.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD   119.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD   169.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Learn about institutional subscriptions

Springer Nature is developing a new tool to find and evaluate Protocols. Learn more

References

  1. Baum, D. A., and Silverman, S. K. (2008) Deoxyribozymes: useful DNA catalysts in vitro and in vivo. Cell Mol. Life Sci. 65, 2156–2174.

    Article  PubMed  CAS  Google Scholar 

  2. Santoro, S. W., and Joyce, G. F. (1997) A general purpose RNA-cleaving DNA enzyme. Proc. Natl. Acad. Sci. USA 94, 4262–4266.

    Article  PubMed  CAS  Google Scholar 

  3. Santoro, S. W., and Joyce, G. F. (1998) Mechanism and utility of an RNA-cleaving DNA enzyme. Biochemistry 37, 13330–13342.

    Article  PubMed  CAS  Google Scholar 

  4. Bhindi, R., Fahmy, R. G., Lowe, H. C., Chesterman, C. N., Dass, C. R., Cairns, M. J., Saravolac, E. G., Sun, L. Q., and Khachigian, L. M. (2007) Brothers in arms. DNA enzymes, short interfering RNA, and the emerging wave of small-molecule nucleic acid-based gene-silencing strategies. Am. J. Pathol. 171, 1079–1088.

    Article  PubMed  CAS  Google Scholar 

  5. Dass, C. R., Choong, P. F., and Khachigian, L. M. (2008) DNAzyme technology and cancer therapy: cleave and let die. Mol. Cancer Ther. 7, 243–251.

    Article  PubMed  CAS  Google Scholar 

  6. Alvarez-Salas, L. M., and DiPaolo, J. A. (2007) Molecular approaches to cervical cancer therapy. Curr. Drug Discov. Technol. 4, 208–219.

    Article  PubMed  CAS  Google Scholar 

  7. Faulhammer, D., and Famulok, M. (1997) Characterization and divalent metal-ion dependence of in vitro selected deoxyribozymes which cleave DNA/RNA chimeric oligonucleotides. J. Mol. Biol. 269, 188–202.

    Article  PubMed  CAS  Google Scholar 

  8. Cairns, M. J., King, A., and Sun, L. Q. (2000) Nucleic acid mutation analysis using catalytic DNA. Nucleic Acids Res. 28, E9.

    Article  PubMed  CAS  Google Scholar 

  9. Yuan, B. F., Xue, Y., Luo, M., Hao, Y. H., and Tan, Z. (2007) Two DNAzymes targeting the telomerase mRNA with large difference in Mg2+ concentration for maximal catalytic activity. Int. J. Biochem. Cell Biol. 39, 1119–1129.

    Article  PubMed  CAS  Google Scholar 

  10. Takamori, K., Kubo, T., Zhelev, Z., Rumiana, B., Ohba, H., Doi, K., and Fujii, M. (2005) Suppression of bcr/abl chimeric gene by conjugate DNA enzymes in human cells. Nucl. Acids Symp. Ser. (Oxf.) 49, 333–334.

    Article  Google Scholar 

  11. Reyes-Gutierrez, P., and Alvarez-Salas, L. M. (2009) Cleavage of HPV-16 E6/E7 mRNA mediated by modified 10-23 deoxyribozymes. Oligonucleotides 19, 233–242.

    Article  PubMed  CAS  Google Scholar 

  12. Pirisi, L., Yasumoto, S., Feller, M., Doniger, J., and DiPaolo, J. A. (1987) Transformation of human fibroblasts and keratinocytes with human papillomavirus type 16 DNA. J. Virol. 61, 1061–1066.

    PubMed  CAS  Google Scholar 

  13. Durst, M., Dzarlieva-Petrusevska, R. T., Boukamp, P., Fusenig, N. E., and Gissmann, L. (1987) Molecular and cytogenetic analysis of immortalized human primary keratinocytes obtained after transfection with human papillomavirus type 16 DNA. Oncogene 1, 251–256.

    PubMed  CAS  Google Scholar 

  14. Alvarez-Salas, L. M., Benitez-Hess, M. L., and DiPaolo, J. A. (2003) Advances in the development of ribozymes and antisense oligodeoxynucleotides as antiviral agents for human papillomaviruses. Antivir. Ther. 8, 265–278.

    PubMed  CAS  Google Scholar 

  15. Broker, T. R. (1987) Structure and genetic expression of papillomaviruses. Obstet. Gynecol. Clin. North Am. 14, 329–348.

    PubMed  CAS  Google Scholar 

  16. Smotkin, D., Prokoph, H., and Wettstein, F. O. (1989) Oncogenic and nononcogenic human genital papillomaviruses generate the E7 mRNA by different mechanisms. J. Virol. 63, 1441–1447.

    PubMed  CAS  Google Scholar 

  17. Schneider-Gadicke, A., and Schwarz, E. (1986) Different human cervical carcinoma cell lines show similar transcription patterns of human papillomavirus type 18 early genes. EMBO J. 5, 2285–2292.

    PubMed  CAS  Google Scholar 

  18. DiPaolo, J. A., Popescu, N. C., Alvarez, L., and Woodworth, C. D. (1993) Cellular and molecular alterations in human epithelial cells transformed by recombinant human papillomavirus DNA. Crit. Rev. Oncog. 4, 337–360.

    PubMed  CAS  Google Scholar 

  19. Duensing, S., and Munger, K. (2002) The human papillomavirus type 16 E6 and E7 oncoproteins independently induce numerical and structural chromosome instability. Cancer Res. 62, 7075–7082.

    PubMed  CAS  Google Scholar 

  20. Alvarez-Salas, L. M., Cullinan, A. E., Siwkowski, A., Hampel, A., and DiPaolo, J. A. (1998) Inhibition of HPV-16 E6/E7 immortalization of normal keratinocytes by hairpin ribozymes. Proc. Natl. Acad. Sci. USA 95, 1189–1194.

    Article  PubMed  CAS  Google Scholar 

  21. Alvarez-Salas, L. M., Arpawong, T. E., and DiPaolo, J. A. (1999) Growth inhibition of cervical tumor cells by antisense oligodeoxynucleotides directed to the human papillomavirus type 16 E6 gene. Antisense Nucl. Acid Drug Dev. 9, 441–450.

    Article  CAS  Google Scholar 

  22. Butz, K., Denk, C., Ullmann, A., Scheffner, M., and Hoppe-Seyler, F. (2000) Induction of apoptosis in human papillomaviruspositive cancer cells by peptide aptamers targeting the viral E6 oncoprotein. Proc. Natl. Acad. Sci. USA 97, 6693–6697.

    Article  PubMed  CAS  Google Scholar 

  23. Jiang, M., and Milner, J. (2002) Selective silencing of viral gene expression in HPV-positive human cervical carcinoma cells treated with siRNA, a primer of RNA interference. Oncogene 21, 6041–6048.

    Article  PubMed  CAS  Google Scholar 

  24. Romani, A. M., and Scarpa, A. (2000) Regulation of cellular magnesium. Front Biosci. 5, D720–D734.

    Article  PubMed  CAS  Google Scholar 

  25. Alvarez-Salas, L. M. (2008) Nucleic acids as therapeutic agents. Curr. Top Med. Chem. 8, 1379–1404.

    Article  PubMed  CAS  Google Scholar 

  26. Santoro, S. W., Joyce, G. F., Sakthivel, K., Gramatikova, S., and Barbas, C. F. (2000) RNA cleavage by a DNA enzyme with extended chemical functionality. J. Am. Chem. Soc. 122, 2433–2439.

    Article  PubMed  CAS  Google Scholar 

  27. Cieslak, M., Niewiarowska, J., Nawrot, M., Koziolkiewicz, M., Stec, W. J., and Cierniewski, C. S. (2002) DNAzymes to beta 1 and beta 3 mRNA down-regulate expression of the targeted integrins and inhibit endothelial cell capillary tube formation in fibrin and matrigel. J. Biol. Chem. 277, 6779–6787.

    Article  PubMed  CAS  Google Scholar 

  28. Fokina, A., Novopashina, D., Meschaninova, M., Vorobjeva, M., Zenkova, M., Francois, J. C., and Venyaminova, A. (2008) Effective cleavage of structured RNAs by tandems of 10-23 DNAzymes with 3'-modified oligo(2'-O-methylribonucleotide)-effectors. Nucl. Acids Symp. Ser. (Oxf.) 52, 525–526.

    Article  CAS  Google Scholar 

  29. Vester, B., Hansen, L. H., Lundberg, L. B., Babu, B. R., Sorensen, M. D., Wengel, J., and Douthwaite, S. (2006) Locked nucleoside analogues expand the potential of DNAzymes to cleave structured RNA targets. BMC Mol. Biol. 7, 19.

    Article  PubMed  Google Scholar 

  30. Guapillo, M. R., Marquez-Gutiérrez, M. A., Benitez-Hess, M. L., and Alvarez-Salas, L. M. (2006) A bacterial reporter system for the evaluation of antisense oligodeoxynucleotides directed against human papillomavirus type 16 (HPV-16). Arch. Med. Res. 37, 584–592.

    Article  PubMed  CAS  Google Scholar 

  31. Marquez-Gutierrez, M. A., Benitez-Hess, M. L., DiPaolo, J. A., and Alvarez-Salas, L. M. (2007) Effect of combined antisense oligodeoxynucleotides directed against the human papillomavirus type 16 on cervical carcinoma cells. Arch. Med. Res. 38, 730–738.

    Article  PubMed  CAS  Google Scholar 

  32. Aquino-Jarquin, G., Benitez-Hess, M. L., DiPaolo, J. A., and Alvarez-Salas, L. M. (2008) A triplex ribozyme expression system based on a single hairpin ribozyme. Oligonucleotides 18, 213–224.

    Article  PubMed  CAS  Google Scholar 

  33. DeYoung, M. B., Siwkowski, A., and Hampel, A. (1997) Determination of catalytic parameters for hairpin ribozymes. Methods Mol. Biol. 74, 209–220.

    PubMed  CAS  Google Scholar 

  34. Baker, C. C., Phelps, W. C., Lindgren, V., Braun, M. J., Gonda, M. A., and Howley, P. M. (1987) Structural and transcriptional analysis of human papillomavirus type 16 sequences in cervical carcinoma cell lines. J. Virol. 61, 962–971.

    PubMed  CAS  Google Scholar 

  35. Yee, C., Krishnan-Hewlett, I., Baker, C. C., Schlegel, R., and Howley, P. M. (1985) Presence and expression of human papillomavirus sequences in human cervical carcinoma cell lines. Am. J. Pathol. 119, 361–366.

    PubMed  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Laboratorio de Terapia Génica, Departamento de Genética y Biología Molecular, CINVESTAV, Av. IPN 2508, México D.F., 07360, México

    María Luisa Benítez-Hess, Pablo Reyes-Gutiérrez & Luis Marat Alvarez-Salas

Authors
  1. María Luisa Benítez-Hess
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Pablo Reyes-Gutiérrez
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Luis Marat Alvarez-Salas
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Luis Marat Alvarez-Salas .

Editor information

Editors and Affiliations

  1. Dept. Chemistry, Worcester State College, Chandler St. 486, Worcester, 01602, Massachusetts, USA

    John Goodchild

Rights and permissions

Reprints and permissions

Copyright information

© 2011 Springer Science+Business Media, LLC

About this protocol

Cite this protocol

Benítez-Hess, M.L., Reyes-Gutiérrez, P., Alvarez-Salas, L.M. (2011). Inhibition of Human Papillomavirus Expression Using DNAzymes. In: Goodchild, J. (eds) Therapeutic Oligonucleotides. Methods in Molecular Biology, vol 764. Humana Press. https://doi.org/10.1007/978-1-61779-188-8_21

Download citation

  • DOI: https://doi.org/10.1007/978-1-61779-188-8_21

  • Published:

  • Publisher Name: Humana Press

  • Print ISBN: 978-1-61779-187-1

  • Online ISBN: 978-1-61779-188-8

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation