Skip to main content
SpringerLink
Log in

Star-Shaped Tetraspermine Enhances Cellular Uptake and Cytotoxicity of T-Oligo in Prostate Cancer Cells

  • Research Article
  • Published:
Pharmaceutical Research Aims and scope Submit manuscript

ABSTRACT

Purpose

An oligonucleotide termed ‘T-oligo’ having sequence homology with telomere overhang has shown cytotoxicity in multiple cancers. We have demonstrated that T-oligo can induce apoptosis in androgen independent prostate cancer cell line DU-145. In this report, we evaluate the use of star-shaped tetraspermine (SSTS) for delivery of T-oligo.

Methods

SSTS was synthesized from spermine and its intrinsic cytotoxicity towards DU-145 cells was compared with spermine and branched polyethyleneimine (bPEI). Atomistic molecular dynamic (MD) simulations were conducted to understand binding and complexation of spermine and SSTS with T-oligo. Complexation was also determined using gel electrophoresis and SYBR gold assay. Complexes were characterized for size, cellular uptake and antiproliferative effect.

Results

SSTS exhibited significantly lower toxicity than spermine and bPEI. Its affinity towards T-oligo was significantly higher than spermine as determined by experimental studies and confirmed by MD simulations and it formed stable complexes (TONPs) with T-oligo. TONPs facilitated cellular uptake and nuclear accumulation of T-oligo and their cytotoxic potential was observed at concentration several folds lower than that required for T-oligo alone.

Conclusion

SSTS significantly enhanced therapeutic benefits associated with the use of T-oligo and can be developed as a delivery vehicle for its in-vivo therapeutic applications.

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

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Scheme 1
Scheme 2
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7
Fig. 8
Fig. 9

Similar content being viewed by others

Abbreviations

6-FAM:

6-carboxyfluorescein

BCA:

Bicinchoninic acid

bPEI:

Polyethylenimine, branched

CRPC:

Castration resistant prostate cancer

DAPI:

4′,6-Diamidino-2-Phenylindole, Dihydrochloride

EDTA(ONP)4 :

Ethylenediaminetetrakis-(p-nitrophenyl) ester

SSTS:

Star-shaped tetraspermine

TONPs:

T-oligo nanoplexes

REFERENCES

  1. Siegel R, Ma J, Zou Z, Jemal A. Cancer statistics. CA Cancer J Clin. 2014;64:9–29.

    Article  PubMed  Google Scholar 

  2. Cookson MS, Roth BJ, Dahm P, Engstrom C, Freedland SJ, Hussain M, et al.. Castration-Resistant Prostate Cancer: AUA Guideline. The Journal of urology. 2013.

  3. Martinezand P, Blasco MA. Telomeric and extra-telomeric roles for telomerase and the telomere-binding proteins. Nat Rev Cancer. 2011;11:161–76.

    Article  Google Scholar 

  4. Aoki H, Iwado E, Eller MS, Kondo Y, Fujiwara K, Li GZ, et al. Telomere 3′ overhang-specific DNA oligonucleotides induce autophagy in malignant glioma cells. FASEB J: Off Publ Fed Am Soc Exp Biol. 2007;21:2918–30.

    Article  Google Scholar 

  5. Longe HO, Romesser PB, Rankin AM, Faller DV, Eller MS, Gilchrest BA, et al. Telomere homolog oligonucleotides induce apoptosis in malignant but not in normal lymphoid cells: mechanism and therapeutic potential. Int J Cancer J Int du Cancer. 2009;124:473–82.

    Article  CAS  Google Scholar 

  6. Li GZ, Eller MS, Firoozabadi R, Gilchrest BA. Evidence that exposure of the telomere 3′ overhang sequence induces senescence. Proc Natl Acad Sci U S A. 2003;100:527–31.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  7. Sarkarand S, Faller DV. T-oligos inhibit growth and induce apoptosis in human ovarian cancer cells. Oligonucleotides. 2011;21:47–53.

    Article  Google Scholar 

  8. Cerone MA, Londoño-Vallejo JA, Autexier C. Telomerase inhibition enhances the response to anticancer drug treatment in human breast cancer cells. Mol Cancer Ther. 2006;5:1669–75.

    Article  CAS  PubMed  Google Scholar 

  9. Puri N, Eller MS, Byers HR, Dykstra S, Kubera J, Gilchrest BA. Telomere-based DNA damage responses: a new approach to melanoma. FASEB J: Off Publ Fed Am Soc Exp Biol. 2004;18:1373–81.

    Article  Google Scholar 

  10. Yaar M, Eller MS, Panova I, Kubera J, Wee LH, Cowan KH, et al. Telomeric DNA induces apoptosis and senescence of human breast carcinoma cells. Breast Cancer Res. 2007;9:R13.

    Article  PubMed Central  PubMed  Google Scholar 

  11. Gnanasekar M, Thirugnanam S, Zheng G, Chen A, Ramaswamy K. T-oligo induces apoptosis in advanced prostate cancer cells. Oligonucleotides. 2009;19:287–92.

    Article  CAS  PubMed  Google Scholar 

  12. Rankin AM, Forman L, Sarkar S, Faller DV. Enhanced cytotoxicity from deoxyguanosine-enriched T-oligo in prostate cancer cells. Nucleic Acid Ther. 2013;23:311–21.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  13. Juliano RL, Ming X, Carver K, Laing B. Cellular Uptake and Intracellular Trafficking of Oligonucleotides: Implications for Oligonucleotide Pharmacology. Nucleic acid therapeutics 2014.

  14. Juliano R, Bauman J, Kang H, Ming X. Biological barriers to therapy with antisense and siRNA oligonucleotides. Mol Pharm. 2009;6:686–95.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  15. Lee H, Lytton-Jean AK, Chen Y, Love KT, Park AI, Karagiannis ED, et al. Molecularly self-assembled nucleic acid nanoparticles for targeted in vivo siRNA delivery. Nat Nanotechnol. 2012;7:389–93.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  16. Dong Y, Love KT, Dorkin JR, Sirirungruang S, Zhang Y, Chen D, et al. Lipopeptide nanoparticles for potent and selective siRNA delivery in rodents and nonhuman primates. Proceedings of the National Academy of Sciences of the United States of America 2014.

  17. Zheng M, Pavan GM, Neeb M, Schaper AK, Danani A, Klebe G, et al. Targeting the blind spot of polycationic nanocarrier-based siRNA delivery. ACS Nano. 2012;6:9447–54.

    Article  CAS  PubMed  Google Scholar 

  18. Scholz C, Kos P, Wagner E. Comb-Like Oligoaminoethane Carriers: Change in Topology Improves pDNA Delivery. Bioconjugate chemistry 2014.

  19. Behr JP. Synthetic gene transfer vectors II: back to the future. Acc Chem Res. 2012;45:980–4.

    Article  CAS  PubMed  Google Scholar 

  20. Tang MX, Redemann CT, Szoka Jr FC. In vitro gene delivery by degraded polyamidoamine dendrimers. Bioconjug Chem. 1996;7:703–14.

    Article  CAS  PubMed  Google Scholar 

  21. Mintzerand MA, Simanek EE. Nonviral vectors for gene delivery. Chem Rev. 2009;109:259–302.

    Article  Google Scholar 

  22. Scholzand C, Wagner E. Therapeutic plasmid DNA versus siRNA delivery: common and different tasks for synthetic carriers. J Control Release: Off J Control Release Soc. 2012;161:554–65.

    Article  Google Scholar 

  23. Alabi C, Vegas A, Anderson D. Attacking the genome: emerging siRNA nanocarriers from concept to clinic. Curr Opin Pharmacol. 2012;12:427–33.

    Article  CAS  PubMed  Google Scholar 

  24. Kang HC, Huh KM, Bae YH. Polymeric nucleic acid carriers: current issues and novel design approaches. J Control Release: Off J Control Release Soc. 2012;164:256–64.

    Article  CAS  Google Scholar 

  25. Merkeland OM, Kissel T. Nonviral pulmonary delivery of siRNA. Acc Chem Res. 2012;45:961–70.

    Article  Google Scholar 

  26. Kolhatkar RB, Kitchens KM, Swaan PW, Ghandehari H. Surface acetylation of polyamidoamine (PAMAM) dendrimers decreases cytotoxicity while maintaining membrane permeability. Bioconjug Chem. 2007;18:2054–60.

    Article  CAS  PubMed  Google Scholar 

  27. Kunath K, von Harpe A, Fischer D, Petersen H, Bickel U, Voigt K, et al. Low-molecular-weight polyethylenimine as a non-viral vector for DNA delivery: comparison of physicochemical properties, transfection efficiency and in vivo distribution with high-molecular-weight polyethylenimine. J Control Release: Off J Control Release Soc. 2003;89:113–25.

    Article  CAS  Google Scholar 

  28. Fischer D, Li Y, Ahlemeyer B, Krieglstein J, Kissel T. In vitro cytotoxicity testing of polycations: influence of polymer structure on cell viability and hemolysis. Biomaterials. 2003;24:1121–31.

    Article  CAS  PubMed  Google Scholar 

  29. Elsayed M, Corrand V, Kolhatkar V, Xie Y, Kim NH, Kolhatkar R, et al. The influence of oligospermine architecture on their suitability for siRNA delivery. Biomacromolecules 2014.

  30. Shankar R, Samykutty A, Riggin C, Kannan S, Wenzel U, Kolhatkar R. Cathepsin B degradable star-shaped peptidic macromolecules for delivery of 2-methoxyestradiol. Mol Pharm. 2013;10:3776–88.

    CAS  PubMed  Google Scholar 

  31. Merkel OM, Mintzer MA, Librizzi D, Samsonova O, Dicke T, Sproat B, et al. Triazine dendrimers as nonviral vectors for in vitro and in vivo RNAi: the effects of peripheral groups and core structure on biological activity. Mol Pharm. 2010;7:969–83.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  32. Foloppeand N, MacKerell AD. All-atom empirical force field for nucleic acids: I. Parameter optimization based on small molecule and condensed phase macromolecular target data. J Comput Chem. 2000;21:86–104.

    Article  Google Scholar 

  33. MacKerelland AD, Banavali NK. All-atom empirical force field for nucleic acids: II. Application to molecular dynamics simulations of DNA and RNA in solution. J Comput Chem. 2000;21:105–20.

    Article  Google Scholar 

  34. Vanommeslaeghe K, Hatcher E, Acharya C, Kundu S, Zhong S, Shim J, et al. CHARMM general force field: a force field for drug-like molecules compatible with the CHARMM All-atom additive biological force fields. J Comput Chem. 2010;31:671–90.

    CAS  PubMed Central  PubMed  Google Scholar 

  35. Vanommeslaeghe K, Raman EP, MacKerell AD. Automation of the CHARMM general force field (CGenFF) II: assignment of bonded parameters and partial atomic charges. J Chem Inf Model. 2012;52:3155–68.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  36. Vanommeslaegheand K, MacKerell AD. Automation of the CHARMM general force field (CGenFF) I: bond perception and atom typing. J Chem Inf Model. 2012;52:3144–54.

    Article  Google Scholar 

  37. Darden T, York D, Pedersen L. Particle mesh ewald - an N.Log(N) method for ewald sums in large systems. J Chem Phys. 1993;98:10089–92.

    Article  CAS  Google Scholar 

  38. Borgman MP, Ray A, Kolhatkar RB, Sausville EA, Burger AM, Ghandehari H. Targetable HPMA copolymer-aminohexylgeldanamycin conjugates for prostate cancer therapy. Pharm Res. 2009;26:1407–18.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  39. Rejman J, Bragonzi A, Conese M. Role of clathrin- and caveolae-mediated endocytosis in gene transfer mediated by lipo- and polyplexes. Mol Ther:J Am Soc Gene Ther. 2005;12:468–74.

    Article  CAS  Google Scholar 

  40. Shi J, Schellinger JG, Johnson RN, Choi JL, Chou B, Anghel EL, et al. Influence of histidine incorporation on buffer capacity and gene transfection efficiency of HPMA-co-oligolysine brush polymers. Biomacromolecules. 2013;14:1961–70.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  41. Vercauteren D, Vandenbroucke RE, Jones AT, Rejman J, Demeester J, De Smedt SC, et al. The use of inhibitors to study endocytic pathways of gene carriers: optimization and pitfalls. Mol Ther:J Am Soc Gene Ther. 2010;18:561–9.

    Article  CAS  Google Scholar 

  42. Shankar R, Samykutty A, Riggin C, Kannan S, Wenzel U, Kolhatkar R. Cathepsin B Degradable Star Shaped Peptidic Macromolecules for Delivery of 2-methoxyestradiol. Molecular pharmaceutics 2013.

  43. Humphrey W, Dalke A, Schulten K. VMD: Visual molecular dynamics. J Mol Graph Model. 1996;14:33–8.

    Article  CAS  Google Scholar 

  44. Weisell J, Hyvonen MT, Hakkinen MR, Grigorenko NA, Pietila M, Lampinen A, et al. Synthesis and biological characterization of novel charge-deficient spermine analogues. J Med Chem. 2010;53:5738–48.

    Article  CAS  PubMed  Google Scholar 

  45. Feuerstein BG, Pattabiraman N, Marton LJ. Spermine-DNA interactions: a theoretical study. Proc Natl Acad Sci U S A. 1986;83:5948–52.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  46. Feuerstein BG, Pattabiraman N, Marton LJ. Theoretical models of spermine/DNA interactions. Ann N Y Acad Sci. 1986;482:251–4.

    Article  CAS  PubMed  Google Scholar 

  47. Jiang HL, Lim HT, Kim YK, Arote R, Shin JY, Kwon JT, et al. Chitosan-graft-spermine as a gene carrier in vitro and in vivo. Eur J Pharmaceutics and Biopharmaceutics: Off J Arbeitsgemeinschaft fur Pharmazeutische Verfahrenstechnik eV. 2011;77:36–42.

    Article  CAS  Google Scholar 

  48. Abdullah S, Wendy-Yeo WY, Hosseinkhani H, Hosseinkhani M, Masrawa E, Ramasamy R, et al. Gene transfer into the lung by nanoparticle dextran-spermine/plasmid DNA complexes. J Biomed Biotechnol. 2010;2010:284840.

    Article  PubMed Central  PubMed  Google Scholar 

  49. Pack DW, Hoffman AS, Pun S, Stayton PS. Design and development of polymers for gene delivery. Nat Rev Drug Discov. 2005;4:581–93.

    Article  CAS  PubMed  Google Scholar 

  50. Eliyahu H, Makovitzki A, Azzam T, Zlotkin A, Joseph A, Gazit D, et al. Novel dextran-spermine conjugates as transfecting agents: comparing water-soluble and micellar polymers. Gene Ther. 2005;12:494–503.

    Article  CAS  PubMed  Google Scholar 

  51. Amini R, Jalilian FA, Abdullah S, Veerakumarasivam A, Hosseinkhani H, Abdulamir AS, et al. Dynamics of PEGylated-dextran-spermine nanoparticles for gene delivery to leukemic cells. Appl Biochem Biotechnol. 2013;170:841–53.

    Article  CAS  PubMed  Google Scholar 

  52. Jere D, Kim JE, Arote R, Jiang HL, Kim YK, Choi YJ, et al. Akt1 silencing efficiencies in lung cancer cells by sh/si/ssiRNA transfection using a reductable polyspermine carrier. Biomaterials. 2009;30:1635–47.

    Article  CAS  PubMed  Google Scholar 

  53. Duan SY, Ge XM, Lu N, Wu F, Yuan W, Jin T. Synthetic polyspermine imidazole-4, 5-amide as an efficient and cytotoxicity-free gene delivery system. Int J Nanomedicine. 2012;7:3813–22.

    CAS  PubMed Central  PubMed  Google Scholar 

  54. Duan S, Yuan W, Wu F, Jin T. Polyspermine Imidazole-4,5-imine, a Chemically Dynamic and Biologically Responsive Carrier System for Intracellular Delivery of siRNA. Angew Chem Int Ed Engl 2012.

  55. Barnard A, Posocco P, Pricl S, Calderon M, Haag R, Hwang ME, et al. Degradable self-assembling dendrons for gene delivery: experimental and theoretical insights into the barriers to cellular uptake. J Am Chem Soc. 2011;133:20288–300.

    Article  CAS  PubMed  Google Scholar 

  56. Jones SP, Gabrielson NP, Wong CH, Chow HF, Pack DW, Posocco P, et al. Hydrophobically modified dendrons: developing structure-activity relationships for DNA binding and gene transfection. Mol Pharm. 2011;8:416–29.

    Article  CAS  PubMed  Google Scholar 

  57. Pavan GM, Danani A, Pricl S, Smith DK. Modeling the multivalent recognition between dendritic molecules and DNA: understanding How ligand “sacrifice” and screening Can enhance binding. J Am Chem Soc. 2009;131:9686–94.

    Article  CAS  PubMed  Google Scholar 

  58. Jones SP, Pavan GM, Danani A, Pricl S, Smith DK. Quantifying the effect of surface ligands on dendron-DNA interactions: insights into multivalency through a combined experimental and theoretical approach. Chemistry. 2010;16:4519–32.

    Article  CAS  PubMed  Google Scholar 

  59. Hardy JG, Kostiainen MA, Smith DK, Gabrielson NP, Pack DW. Dendrons with spermine surface groups as potential building blocks for nonviral vectors in gene therapy. Bioconjug Chem. 2006;17:172–8.

    Article  CAS  PubMed  Google Scholar 

  60. Rejman J, Conese M, Hoekstra D. Gene transfer by means of lipo- and polyplexes: role of clathrin and caveolae-mediated endocytosis. J Liposome Res. 2006;16:237–47.

    Article  CAS  PubMed  Google Scholar 

  61. Sahay G, Alakhova DY, Kabanov AV. Endocytosis of nanomedicines. J Control Release: Off J Control Release Soc. 2010;145:182–95.

    Article  CAS  Google Scholar 

  62. Dong Y, Li J, Wu C, Oupicky D. Bisethylnorspermine lipopolyamine as potential delivery vector for combination drug/gene anticancer therapies. Pharm Res. 2010;27:1927–38.

    Article  CAS  PubMed  Google Scholar 

  63. Chen Y, Sen J, Bathula SR, Yang Q, Fittipaldi R, Huang L. Novel cationic lipid that delivers siRNA and enhances therapeutic effect in lung cancer cells. Mol Pharm. 2009;6:696–705.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  64. Martinand ME, Rice KG. A novel class of intrinsic proteasome inhibitory gene transfer peptides. Bioconjug Chem. 2008;19:370–6.

    Article  Google Scholar 

  65. Li J, Zhu Y, Hazeldine ST, Li C, Oupicky D. Dual-function CXCR4 antagonist polyplexes to deliver gene therapy and inhibit cancer cell invasion. Angew Chem Int Ed Engl. 2012;51:8740–3.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  66. Sun C, Tang T, Uludag H. Molecular dynamics simulations for complexation of DNA with 2 kDa PEI reveal profound effect of PEI architecture on complexation. J Phys Chem B. 2012;116:2405–13.

    Article  CAS  PubMed  Google Scholar 

  67. Pavan GM, Mintzer MA, Simanek EE, Merkel OM, Kissel T, Danani A. Computational insights into the interactions between DNA and siRNA with “rigid” and “flexible” triazine dendrimers. Biomacromolecules. 2010;11:721–30.

    Article  CAS  PubMed  Google Scholar 

  68. Pavan GM, Albertazzi L, Danani A. Ability to adapt: different generations of PAMAM dendrimers show different behaviors in binding siRNA. J Phys Chem B. 2010;114:2667–75.

    Article  CAS  PubMed  Google Scholar 

  69. Meneksedag-Erol D, Tang T, Uludag H. Molecular modeling of polynucleotide complexes. Biomaterials. 2014;35:7068–76.

    Article  CAS  PubMed  Google Scholar 

  70. Elder RM, Emrick T, Jayaraman A. Understanding the effect of polylysine architecture on DNA binding using molecular dynamics simulations. Biomacromolecules. 2011;12:3870–9.

    Article  CAS  PubMed  Google Scholar 

Download references

ACKNOWLEDGMENTS AND DISCLOSURES

The work was supported by Department of Biopharmaceutical Sciences, College of Pharmacy, University of Illinois at Chicago. Peter Shanine received Riback Fellowship from College of Pharmacy, University of Illinois at Chicago. We also thank Dr. Onyuksel for allowing us to use NICOMP light scattering instrument. Petr Král’s work was supported by the NSF-DMR grant No. 1309765 and ACS PRF grant #53062-ND6.

Author information

Authors and Affiliations

  1. Department of Biopharmaceutical Sciences, University of Illinois Chicago, 1601 Parkview Ave, Rm N302, Rockford, Illinois, 61107, USA

    Vidula Kolhatkar, Hiren Khambati, Asawari Lote, Peter Shanine & Rohit Kolhatkar

  2. Department of Chemistry, University of Illinois at Chicago, Chicago, Illinois, 60607, USA

    Thomas Insley, Soumyo Sen & Petr Král

  3. Department of Biomedical Sciences, College of Medicine, University of Illinois Chicago, Rockford, Illinois, 61107, USA

    Gnanasekar Munirathinam & Rohit Kolhatkar

  4. Department of Physics, University of Illinois at Chicago, Chicago, Illinois, 60607, USA

    Petr Král

Authors
  1. Vidula Kolhatkar
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Hiren Khambati
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Asawari Lote
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Peter Shanine
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Thomas Insley
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Soumyo Sen
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Gnanasekar Munirathinam
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Petr Král
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Rohit Kolhatkar
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Rohit Kolhatkar.

Electronic Supplementary Material

Below is the link to the electronic supplementary material.

ESM 1

(DOCX 360 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Kolhatkar, V., Khambati, H., Lote, A. et al. Star-Shaped Tetraspermine Enhances Cellular Uptake and Cytotoxicity of T-Oligo in Prostate Cancer Cells. Pharm Res 32, 196–210 (2015). https://doi.org/10.1007/s11095-014-1455-7

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11095-014-1455-7

KEY WORDS

Search

Navigation