Advertisement

AAPS PharmSciTech

, 20:111 | Cite as

Design, Synthesis, and In Vitro Evaluation of Low Molecular Weight Protamine (LMWP)-Based Amphiphilic Conjugates as Gene Delivery Carriers

  • Sepideh Arabzadeh
  • Zeinab Amiri Tehranizadeh
  • Hamideh Moalemzadeh Haghighi
  • Fahimeh Charbgoo
  • Mohammad RamezaniEmail author
  • Fatemeh SoltaniEmail author
Research Article

Abstract

Development of efficient non-viral carriers is one of the major challenges of gene delivery. In the current study, we designed, synthesized, and evaluated the in vitro gene delivery efficiency of novel amphiphilic constructs composed of cholesterol and low molecular weight protamine (LMWP: VSRRRRRRGGRRRR) peptide. Vectors having both hydrophobic and hydrophilic moieties were evaluated in terms of particle size and charge, DNA condensation ability, cytotoxicity, and gene transfection efficiency. The prepared vectors spontaneity self-assembled into the liposome-like particles with a high local positive density. The nano-vehicle A (H5-LMWP-Cholestrol) and nano-vehicle B (LMWP-Cholesterol) could form micelles at concentrations above 50 μg/mL and 65 μg/mL, respectively. The gel retardation assay showed that nano-vehicles A and B could condense pDNA more efficiently than the corresponding unconjugated peptides. The mean of size and zeta potential of complexed nano-vehicle A at N/P ratios of 5, 15, and 30 were 151 nm and 23 mv, and those of nano-vehicle B were 224 nm and 19 mv, respectively. In terms of transfection efficiency, the designed nano-vehicles showed almost two-fold higher gene expression level compared to PEI 25 kDa at optimal N/P ratios, and also exhibited negligible cytotoxicity on a model cancer cell, Neuro 2a. The findings of the present study revealed that these cationic micelles can be promising candidates as non-viral gene delivery vehicles.

KEY WORDS

gene delivery vectors micelles low molecular weight protamine cholesterol 

Notes

Acknowledgments

This work was supported by Mashhad University of Medical Sciences.

Compliance with Ethical Standards

Conflict of Interest

The authors declare that they have no conflict of interest.

Supplementary material

12249_2018_1235_MOESM1_ESM.jpg (379 kb)
ESM 1 (JPG 378 kb)
12249_2018_1235_MOESM2_ESM.jpg (406 kb)
ESM 2 (JPG 406 kb)

References

  1. 1.
    Verma IM, Somia N. Gene therapy-promises, problems and prospects. Nature. 1997;389:239–42.CrossRefGoogle Scholar
  2. 2.
    Niidome T, Huang L. Gene therapy progress and prospects: nonviral vectors. Gene Ther. 2002;9:1647–52.CrossRefGoogle Scholar
  3. 3.
    Belting M, Sandgren S, Wittrup A. Nuclear delivery of macromolecules: barriers and carriers. Adv Drug Deliv Rev. 2005;57:505–27.CrossRefGoogle Scholar
  4. 4.
    Liu F, Huang L. Development of non-viral vectors for systemic gene delivery. J Control Release. 2002;78:259–66.CrossRefGoogle Scholar
  5. 5.
    Soltani F, Parhiz H, Mokhtarzadeh A, Ramezani M. Synthetic and biological vesicular nano-carriers designed for gene delivery. Curr Pharm Des. 2015;21:6214–35.CrossRefGoogle Scholar
  6. 6.
    Mitchell P. Vector problems still thwart gene-therapy promise. Lancet. 1998;351:346.CrossRefGoogle Scholar
  7. 7.
    Akinc A, Anderson DG, Lynn DM, Langer R. Synthesis of poly(β-amino ester)s optimized for highly effective gene delivery. Bioconjug Chem. 2003;14:979–88.CrossRefGoogle Scholar
  8. 8.
    Mintzer MA, Simanek EE. Nonviral vectors for gene delivery. Chem Rev. 2008;109:259–302.CrossRefGoogle Scholar
  9. 9.
    Nguyen DN, Green JJ, Chan JM, Langer R, Anderson DG. Polymeric materials for gene delivery and DNA vaccination. Adv Mater. 2009;21:847–67.CrossRefGoogle Scholar
  10. 10.
    Pathak A, Patnaik S, Gupta KC. Recent trends in non-viral vector-mediated gene delivery. Biotechnol J. 2009;4:1559–72.CrossRefGoogle Scholar
  11. 11.
    Jafari M, Soltani M, Naahidi S, Karunaratne DN, Chen P. Nonviral approach for targeted nucleic acid delivery. Curr Med Chem. 2012;19:197–208.CrossRefGoogle Scholar
  12. 12.
    Milletti F. Cell-penetrating peptides: classes, origin, and current landscape. Drug Discov Today. 2012;17:850–60.CrossRefGoogle Scholar
  13. 13.
    Zou L-L, Ma J-L, Wang T, Yang T-B, Liu C-B. Cell-penetrating peptide-mediated therapeutic molecule delivery into the central nervous system. Curr Neuropharmacol. 2013;11:197–208.CrossRefGoogle Scholar
  14. 14.
    Hoyer J, Neundorf I. Peptide vectors for the nonviral delivery of nucleic acids. Acc Chem Res. 2012;45:1048–56.CrossRefGoogle Scholar
  15. 15.
    Kaouass M, Beaulieu R, Balicki D. Histonefection: novel and potent non-viral gene delivery. J Control Release. 2006;113:245–54.CrossRefGoogle Scholar
  16. 16.
    Soltani F, Sankian M, Hatefi A, Ramezani M. Development of a novel histone H1-based recombinant fusion peptide for targeted non-viral gene delivery. Int J Pharm. 2013;441:307–15.CrossRefGoogle Scholar
  17. 17.
    Ma K, Wang DD, Lin Y, Wang J, Petrenko V, Mao C. Synergetic targeted delivery of sleeping-beauty transposon system to mesenchymal stem cells using LPD nanoparticles modified with a phage-displayed targeting peptide. Adv Funct Mater. 2013;23:1172–81.CrossRefGoogle Scholar
  18. 18.
    Choi YS, Lee JY, Suh JS, Kwon YM, Lee SJ, Chung JK, et al. The systemic delivery of siRNAs by a cell penetrating peptide, low molecular weight protamine. Biomaterials. 2010;31:1429–43.CrossRefGoogle Scholar
  19. 19.
    Park YJ, Liang JF, Ko KS, Kim SW, Yang VC. Low molecular weight protamine as an efficient and nontoxic gene carrier: in vitro study. J Gene Med. 2003;5:700–11.CrossRefGoogle Scholar
  20. 20.
    Chen M, Liu Y, Yang W, Li X, Liu L, Zhou Z, et al. Preparation and characterization of self-assembled nanoparticles of 6-O-cholesterol-modified chitosan for drug delivery. Carbohydr Polym. 2011;84:1244–51.CrossRefGoogle Scholar
  21. 21.
    Guo XD, Tandiono F, Wiradharma N, Khor D, Tan CG, Khan M, et al. Cationic micelles self-assembled from cholesterol-conjugated oligopeptides as an efficient gene delivery vector. Biomaterials. 2008;29:4838–46.CrossRefGoogle Scholar
  22. 22.
    Tang Q, Cao B, Wu H, Cheng G. Cholesterol-peptide hybrids to form liposome-like vesicles for gene delivery. PLoS One. 2013;8:e54460.CrossRefGoogle Scholar
  23. 23.
    Pichon C, Gonçalves C, Midoux P. Histidine-rich peptides and polymers for nucleic acids delivery. Adv Drug Deliv Rev. 2001;53:75–94.CrossRefGoogle Scholar
  24. 24.
    Midoux P, Pichon C, Yaouanc JJ, Jaffrès PA. Chemical vectors for gene delivery: a current review on polymers, peptides and lipids containing histidine or imidazole as nucleic acids carriers. Br J Pharmacol. 2009;157:166–78.CrossRefGoogle Scholar
  25. 25.
    Patil SN, Liu F. Regioselective synthesis and structural studies of substituted γ-hydroxybutenolides with use of a tandem Baylis–Hillman/singlet oxygenation reaction. J Org Chem. 2008;73:4476–83.CrossRefGoogle Scholar
  26. 26.
    Domínguez A, Fernández A, González N, Iglesias E, Montenegro L. Determination of critical micelle concentration of some surfactants by three techniques. J Chem Educ. 1997;74:1227.CrossRefGoogle Scholar
  27. 27.
    Dehshahri A, Oskuee RK, Shier WT, Hatefi A, Ramezani M. Gene transfer efficiency of high primary amine content, hydrophobic, alkyl-oligoamine derivatives of polyethylenimine. Biomaterials. 2009;30:4187–94.CrossRefGoogle Scholar
  28. 28.
    Salmasi Z, Shier WT, Hashemi M, Mahdipour E, Parhiz H, Abnous K, , Ramezani M. Heterocyclic amine-modified polyethylenimine as gene carriers for transfection of mammalian cells Eur J Pharm Biopharm 2015;96:76–88.CrossRefGoogle Scholar
  29. 29.
    Balhorn R, Brewer L, Corzett M. DNA condensation by protamine and arginine-rich peptides: analysis of toroid stability using single DNA molecules. Mol Reprod Dev. 2000;56:230–4.CrossRefGoogle Scholar
  30. 30.
    Sabouri-Rad S, Oskuee RK, Mahmoodi A, Gholami L, Malaekeh-Nikouei B. The effect of cell penetrating peptides on transfection activity and cytotoxicity of polyallylamine. Bioimpacts. 2017;7:139–45.CrossRefGoogle Scholar
  31. 31.
    Ayatollahi S, Salmasi Z, Hashemi M, Askarian S, Oskuee RK, Abnous K, et al. Aptamer-targeted delivery of Bcl-xL shRNA using alkyl modified PAMAM dendrimers into lung cancer cells. Int J Biochem Cell Biol. 2017;92:210–7.CrossRefGoogle Scholar
  32. 32.
    Ter-Avetisyan G, Tünnemann G, Nowak D, Nitschke M, Herrmann A, Drab M, et al. Cell entry of arginine-rich peptides is independent of endocytosis. J Biol Chem. 2009;284:3370–8.CrossRefGoogle Scholar
  33. 33.
    Erazo-Oliveras A, Muthukrishnan N, Baker R, Wang TY, Pellois JP. Improving the endosomal escape of cell-penetrating peptides and their cargos: strategies and challenges. Pharmaceuticals. 2012;5:1177–209.CrossRefGoogle Scholar

Copyright information

© American Association of Pharmaceutical Scientists 2019

Authors and Affiliations

  1. 1.Biotechnology Research Center, Pharmaceutical Technology InstituteMashhad University of Medical SciencesMashhadIran
  2. 2.Department of Medicinal Chemistry, School of PharmacyMashhad University of Medical SciencesMashhadIran
  3. 3.Pharmaceutical Research Center, Pharmaceutical Technology InstituteMashhad University of Medical SciencesMashhadIran

Personalised recommendations