Fabrication of thermo-sensitive complex micelles for reversible cell targeting

  • Yukun Wu
  • Chengling Yang
  • Quanyong Lai
  • Qian Zhang
  • Wei Wang
  • Zhi Yuan
Biomaterials Synthesis and Characterization Original Research
Part of the following topical collections:
  1. Biomaterials Synthesis and Characterization


To ideally solve the contradiction between enhanced cellular uptake and prolonged blood circulation, reversible targeting polymeric micelles based on the expanding and shrinking behavior of a temperature-responsive polymer were developed. The micelle contained a hydrophobic PCL core and a mixed shell consisting of poly(N-isopropylacrylamide) (PNIPAAm) and biotin-terminated poly(ethylene glycol) (Biotin-PEG), and its targeting ability could be switched on/off by temperature. The cellular uptake of the complex polymeric micelles was studied. The results from a quantitative enzyme-linked immunosorbent assay (ELISA) indicated that the surface biotin content increased by as much as 11.6-fold when the temperature increased above the lower critical solution temperature (LCST). More importantly, the ELISA confirmed that biotin-mediated targeting on the surface was reversibly switched on and off for at least five cycles. In addition, the results from quantitative flow cytometry and confocal spectroscopy indicated that the cellular uptake of the targeted micelles at temperatures above the LCST was much higher than that at temperatures below the LCST. This complex polymeric micelle with reversible targeting property could be a promising alternative for drug delivery.


HepG2 Cell Block Copolymer Cellular Uptake Atom Transfer Radical Polymerization Atom Transfer Radical Polymerization 



This work was supported by the National Natural Science Foundation of China (51433004), Natural Science Foundation of Tianjin (13JCYBJC25100), and PCSIRT (IRT1257). We thank Prof. Deling Kong, Nankai University, for his help with the whole cell experiments.


  1. 1.
    Maeda H, Bharate GYY, Daruwalla J. Polymeric drugs for efficient tumor-targeted drug delivery based on EPR-effect. Eur J Pharm Biopharm. 2009;71:409–19. doi: 10.1016/j.ejpb.2008.11.010.CrossRefGoogle Scholar
  2. 2.
    Kwon IK, Lee SC, Han B, Park K. Analysis on the current status of targeted drug delivery to tumors. J Control Release. 2012;164:108–14. doi: 10.1016/j.jconrel.2012.07.010.CrossRefGoogle Scholar
  3. 3.
    Fang J, Nakamura H, Maeda H. The EPR effect: unique features of tumor blood vessels for drug delivery, factors involved, and limitations and augmentation of the effect. Adv Drug Deliv Rev. 2011;63:136–51. doi: 10.1016/j.addr.2010.04.009.CrossRefGoogle Scholar
  4. 4.
    Manuscript A, Targeting D, Heterogeneity T, Bae YH. Drug targeting and tumor heterogeneity. J Control Release. 2009;133:2–3. doi: 10.1016/j.jconrel.2008.09.074.CrossRefGoogle Scholar
  5. 5.
    Chen CC, Borden MA. The role of poly(ethylene glycol) brush architecture in complement activation on targeted microbubble surfaces. Biomaterials. 2011;32:6579–87. doi: 10.1016/j.biomaterials.2011.05.027.CrossRefGoogle Scholar
  6. 6.
    McNeeley KM, Karathanasis E, Annapragada AV, Bellamkonda RV. Masking and triggered unmasking of targeting ligands on nanocarriers to improve drug delivery to brain tumors. Biomaterials. 2009;30:3986–95. doi: 10.1016/j.biomaterials.2009.04.012.CrossRefGoogle Scholar
  7. 7.
    Gullotti E, Yeo Y. Extracellularly activated nanocarriers: a new paradigm of tumor targeted drug delivery. Mol Pharm. 2009;6:1041–51. doi: 10.1021/mp900090z.CrossRefGoogle Scholar
  8. 8.
    Low PS. The optimal strategy for drug targeting. Mol Pharm. 2007;4:629–30. doi: 10.1021/mp700111w.CrossRefGoogle Scholar
  9. 9.
    Fan N-C, Cheng F-Y, Ho JA, Yeh C-S. Photocontrolled targeted drug delivery: photocaged biologically active folic acid as a light-responsive tumor-targeting molecule. Angew Chem Int Ed Engl. 2012;51:8806–10. doi: 10.1002/anie.201203339.CrossRefGoogle Scholar
  10. 10.
    Chen Y, Gao D-Y, Huang L. In vivo delivery of miRNAs for cancer therapy: challenges and strategies. Adv Drug Deliv Rev. 2014;. doi: 10.1016/j.addr.2014.05.009.Google Scholar
  11. 11.
    Niu Y, Yu M, Hartono SB, Yang J, Xu H, Zhang H, et al. Nanoparticles mimicking viral surface topography for enhanced cellular delivery. Adv Mater. 2013;25:6232–3. doi: 10.1002/adma.201302737.CrossRefGoogle Scholar
  12. 12.
    Yao X, Peng R, Ding J. Cell-material interactions revealed via material techniques of surface patterning. Adv Mater. 2013;25(37):5257–86. doi: 10.1002/adma.201301762.CrossRefGoogle Scholar
  13. 13.
    Poon Z, Chen S, Engler AC, Lee HI, Atas E, von Maltzahn G, et al. Ligand-clustered “patchy” nanoparticles for modulated cellular uptake and in vivo tumor targeting. Angew Chemie Int Ed. 2010;49:7266–70. doi: 10.1002/anie.201003445.CrossRefGoogle Scholar
  14. 14.
    Ashley CE, Carnes EC, Phillips GK, Padilla D, Durfee PN, Brown PA, et al. The targeted delivery of multicomponent cargos to cancer cells by nanoporous particle-supported lipid bilayers. Nat Mater. 2011;10:389–97. doi: 10.1038/NMAT2992.CrossRefGoogle Scholar
  15. 15.
    Kim C, Lee Y, Kim JS, Jeong JH, Park TG. Thermally triggered cellular uptake of quantum dots immobilized with poly(N-isopropylacrylamide) and cell penetrating peptide. Langmuir. 2010;26:14965–9. doi: 10.1021/la102632m.CrossRefGoogle Scholar
  16. 16.
    Moradi E, Vllasaliu D, Garnett M, Falcone F, Stolnik S. Ligand density and clustering effects on endocytosis of folate modified nanoparticles. RSC Adv. 2012;2:3025. doi: 10.1039/c2ra01168a.CrossRefGoogle Scholar
  17. 17.
    Sansone F, Casnati A. Multivalent glycocalixarenes for recognition of biological macromolecules: glycocalyx mimics capable of multitasking. Chem Soc Rev. 2013;42:4623–39. doi: 10.1039/c2cs35437c.CrossRefGoogle Scholar
  18. 18.
    Zhang Z, Ali MM, Eckert MA, Kang D-KD-K, Chen YY, Sender LS, et al. A polyvalent aptamer system for targeted drug delivery. Biomaterials. 2013;34:9728–35. doi: 10.1016/j.biomaterials.2013.08.079.CrossRefGoogle Scholar
  19. 19.
    Mastrotto F, Caliceti P, Amendola V, Bersani S, Magnusson JP, Meneghetti M, et al. Polymer control of ligand display on gold nanoparticles for multimodal switchable cell targeting. Chem Commun. 2011;47:9846–8. doi: 10.1039/c1cc12654g.CrossRefGoogle Scholar
  20. 20.
    Macewan SR, Chilkoti A. Digital switching of local arginine density in a genetically encoded self-assembled polypeptide nanoparticle controls cellular uptake. Nano Lett. 2012;12:3322–8. doi: 10.1021/nl301529p.CrossRefGoogle Scholar
  21. 21.
    Lin Q, Bao C, Yang Y, Liang Q, Zhang D, Cheng S, et al. Highly discriminating photorelease of anticancer drugs based on hypoxia activatable phototrigger conjugated chitosan nanoparticles. Adv Mater. 2013;25:1981–6. doi: 10.1002/adma.201204455.CrossRefGoogle Scholar
  22. 22.
    Borden MA, Zhang H, Gillies RJ, Dayton PA, Ferrara KW. A stimulus-responsive contrast agent for ultrasound molecular imaging. Biomaterials. 2008;29:597–606. doi: 10.1016/j.biomaterials.2007.10.011.CrossRefGoogle Scholar
  23. 23.
    Lee ES, Gao Z, Bae YH, Manuscript A. Recent progress in tumor pH targeting nanotechnology. J Control Release. 2008;132:164–70. doi: 10.1016/j.jconrel.2008.05.003.CrossRefGoogle Scholar
  24. 24.
    Sethuraman VA, Bae YH. TAT peptide-based micelle system for potential active targeting of anti-cancer agents to acidic solid tumors. J Control Release. 2007;118:216–24. doi: 10.1016/j.jconrel.2006.12.008.CrossRefGoogle Scholar
  25. 25.
    Quan C-Y, Chen J-X, Wang H-Y, Li C, Chang C, Zhang X, et al. Core-shell nanosized assemblies mediated by the alpha-beta cyclodextrin dimer with a tumor-triggered targeting property. ACS Nano. 2010;4:4211–9. doi: 10.1021/nn100534q.CrossRefGoogle Scholar
  26. 26.
    Gao H, Cheng T, Liu J, Liu J, Yang C, Chu L, et al. Self-regulated multifunctional collaboration of targeted nanocarriers for enhanced tumor therapy. Biomacromolecules. 2014;15:3634–42. doi: 10.1021/bm5009348.CrossRefGoogle Scholar
  27. 27.
    Tian Z, Yang C, Wang W, Yuan Z. Shieldable tumor targeting based on pH responsive self-assembly/disassembly of gold nanoparticles. ACS Appl Mater Interfaces. 2014;. doi: 10.1021/am5045339.Google Scholar
  28. 28.
    Mok H, Bae KH, Ahn C-H, Park TG. PEGylated and MMP-2 specifically dePEGylated quantum dots: comparative evaluation of cellular uptake. Langmuir. 2009;25:1645–50. doi: 10.1021/la803542v.CrossRefGoogle Scholar
  29. 29.
    Deng C, Jiang Y, Cheng R, Meng F, Zhong Z. Biodegradable polymeric micelles for targeted and controlled anticancer drug delivery: promises, progress and prospects. Nano Today. 2012;7:467–80. doi: 10.1016/j.nantod.2012.08.005.CrossRefGoogle Scholar
  30. 30.
    Mura S, Nicolas J, Couvreur P. Stimuli-responsive nanocarriers for drug delivery. Nat Mater. 2013;12:991–1003. doi: 10.1038/nmat3776.CrossRefGoogle Scholar
  31. 31.
    Hildebrandt B, Wust P, Ahlers O, Dieing A, Sreenivasa G, Kerner T, et al. The cellular and molecular basis of hyperthermia. Crit Rev Oncol Hematol. 2002;43:33–56. doi: 10.1016/s1040-8428(01)00179-2.CrossRefGoogle Scholar
  32. 32.
    Mcdaniel JR, Macewan SR, Dewhirst M, Chilkoti A. Doxorubicin-conjugated chimeric polypeptide nanoparticles that respond to mild hyperthermia. J Control Release. 2012;159:362–7. doi: 10.1016/j.jconrel.2012.02.030.CrossRefGoogle Scholar
  33. 33.
    McDaniel JR, Dewhirst MW, Chilkoti A. Actively targeting solid tumours with thermoresponsive drug delivery systems that respond to mild hyperthermia. Int J Hyperth. 2013;29:501–10. doi: 10.3109/02656736.2013.819999.CrossRefGoogle Scholar
  34. 34.
    Miyata K, Christie RJ, Kataoka K. Polymeric micelles for nano-scale drug delivery. React Funct Polym. 2011;71:227–34. doi: 10.1016/j.reactfunctpolym.2010.10.009.CrossRefGoogle Scholar
  35. 35.
    Li C, Ge Z, Fang J, Liu S. Synthesis and self-assembly of coil–rod double hydrophilic diblock copolymer with dually responsive asymmetric centipede-shaped polymer brush as the rod segment. Macromolecules. 2009;42:2916–24. doi: 10.1021/ma900165z.CrossRefGoogle Scholar
  36. 36.
    Li W, Li J, Gao J, Li B, Xia Y, Meng Y, et al. The fine-tuning of thermosensitive and degradable polymer micelles for enhancing intracellular uptake and drug release in tumors. Biomaterials. 2011;32:3832–44. doi: 10.1016/j.biomaterials.2011.01.075.CrossRefGoogle Scholar
  37. 37.
    Akimoto J, Nakayama M, Sakai K, Okano T. Temperature-induced intracellular uptake of thermoresponsive polymeric micelles. Biomacromolecules. 2009;10:1331–6. doi: 10.1021/bm900032r.CrossRefGoogle Scholar
  38. 38.
    Sawant RM, Hurley JP, Salmaso S, Kale A, Tolcheva E, Levchenko TS, et al. “SMART” drug delivery systems: double-targeted pH-responsive pharmaceutical nanocarriers. Bioconjug. Chem. 2006;17:943–9. doi: 10.1021/bc060080h.CrossRefGoogle Scholar
  39. 39.
    Lee F, Chung JE, Kurisawa M. An injectable hyaluronic acid-tyramine hydrogel system for protein delivery. J Control Release. 2009;134:186–93. doi: 10.1016/j.jconrel.2008.11.028.CrossRefGoogle Scholar
  40. 40.
    Huang W, Wang W, Wang P, Tian Q, Zhang C, Wang C, et al. Glycyrrhetinic acid-modified poly(ethylene glycol)-b-poly(gamma-benzyl L-glutamate) micelles for liver targeting therapy. Acta Biomater. 2010;6:3927–35. doi: 10.1016/j.actbio.2010.04.021.CrossRefGoogle Scholar
  41. 41.
    Ho K, Lapitsky Y, Shi M, Shoichet MS. Tunable immunonanoparticle binding to cancer cells: thermodynamic analysis of targeted drug delivery vehicles. Soft Matter. 2009;5:1074. doi: 10.1039/b814204a.CrossRefGoogle Scholar
  42. 42.
    Glavas L, Olsén P, Odelius K, Albertsson AC. Achieving micelle control through core crystallinity. Biomacromolecules. 2013;14:4150–6. doi: 10.1021/bm401312j.CrossRefGoogle Scholar
  43. 43.
    Pulkkinen M, Pikkarainen J, Wirth T, Tarvainen T, Haapa-aho V, Korhonen H, et al. Three-step tumor targeting of paclitaxel using biotinylated PLA-PEG nanoparticles and avidin-biotin technology: formulation development and in vitro anticancer activity. Eur J Pharm Biopharm. 2008;70:66–74. doi: 10.1016/j.ejpb.2008.04.018.CrossRefGoogle Scholar
  44. 44.
    Liu X, Ma R, Shen J, Xu Y, An Y, Shi L. Controlled release of ionic drugs from complex micelles with charged channels. Biomacromolecules. 2012;13:1307–14. doi: 10.1021/bm2018382.CrossRefGoogle Scholar
  45. 45.
    Copolymers D, Li G, Shi L, Ma R, An Y, Huang N. Formation of complex micelles with double-responsive channels from self-assembly of two diblock copolymers. Angew Chemie. 2006;118:5081–4. doi: 10.1002/ange.200600172.CrossRefGoogle Scholar
  46. 46.
    Kataoka K, Harada A, Nagasaki Y. Block copolymer micelles for drug delivery: design, characterization and biological significance. Adv Drug Deliv Rev. 2001;47:113–31. doi: 10.1016/s0169-409x(00)00124-1.CrossRefGoogle Scholar
  47. 47.
    Simnick AJ, Amiram M, Liu W, Hanna G, Dewhirst MW, Kontos CD, et al. In vivo tumor targeting by a NGR-decorated micelle of a recombinant diblock copolypeptide. J Control Release. 2011;155:144–51. doi: 10.1016/j.jconrel.2011.06.044.CrossRefGoogle Scholar
  48. 48.
    Li G, Guo L, Ma S, Liu J. Complex micelles formed from two diblock copolymers for applications in controlled drug release. J Polym Sci A. 2009;47:1804–10. doi: 10.1002/pola.23274.CrossRefGoogle Scholar
  49. 49.
    Gao H, Xiong J, Cheng T, Liu JJ, Chu L, Ma R, et al. In vivo biodistribution of mixed shell micelles with tunable hydrophilic/hydrophobic surface. Biomacromolecules. 2013;14:460–7. doi: 10.1021/bm301694t.CrossRefGoogle Scholar
  50. 50.
    Huang Y, Jiang Y, Wang H, Wang J, Shin MC, Byun Y, et al. Curb challenges of the “Trojan Horse” approach: smart strategies in achieving effective yet safe cell-penetrating peptide-based drug delivery. Adv Drug Deliv Rev. 2013;65:1299–315. doi: 10.1016/j.addr.2012.11.007.CrossRefGoogle Scholar
  51. 51.
    Taghdisi SM, Lavaee P, Ramezani M, Abnous K. Reversible Targeting and controlled release delivery of daunorubicin to cancer cells by aptamer-wrapped carbon nanotubes. Eur J Pharm Biopharm. 2011;77:200–6. doi: 10.1016/j.ejpb.2010.12.005.CrossRefGoogle Scholar
  52. 52.
    Yellepeddi VK, Kumar A, Palakurthi S. Biotinylated poly(amido)amine (PAMAM) dendrimers as carriers for drug delivery to ovarian cancer cells in vitro. Anticancer Res 2009;29:2933–2943. http://www.ncbi.nlm.nih.gov/pubmed/19661298. Accessed 18 Apr 2014.
  53. 53.
    Yang W, Cheng Y, Xu T, Wang X, Wen L-P. Targeting cancer cells with biotin-dendrimer conjugates. Eur J Med Chem. 2009;44:862–8. doi: 10.1016/j.ejmech.2008.04.021.CrossRefGoogle Scholar
  54. 54.
    Quan C-Y, Wu D-Q, Chang C, Zhang G, Cheng S, Zhang X, et al. Synthesis of thermo-sensitive micellar aggregates self-assembled from biotinylated PNAS- b -PNIPAAm- b -PCL triblock copolymers for tumor targeting. J Phys Chem C. 2009;113:11262–7. doi: 10.1021/jp902637n.CrossRefGoogle Scholar
  55. 55.
    Cheng C, Wei H, Shi B-X, Cheng H, Li C, Gu Z-W, et al. Biotinylated thermoresponsive micelle self-assembled from double-hydrophilic block copolymer for drug delivery and tumor target. Biomaterials. 2008;29:497–505. doi: 10.1016/j.biomaterials.2007.10.004.CrossRefGoogle Scholar
  56. 56.
    Cheng C, Wei H, Zhang X-Z, Cheng S-X, Zhuo R-X. Thermo-triggered and biotinylated biotin-P(NIPAAm-co-HMAAm)-b-PMMA micelles for controlled drug release. J Biomed Mater Res A. 2009;88:814–22. doi: 10.1002/jbm.a.31770.CrossRefGoogle Scholar
  57. 57.
    Salmaso S, Caliceti P, Amendola V, Meneghetti M, Magnusson JP, Pasparakis G, et al. Cell up-take control of gold nanoparticles functionalized with a thermoresponsive polymer. J Mater Chem. 2009;19:1608. doi: 10.1039/b816603j.CrossRefGoogle Scholar
  58. 58.
    Abulateefeh SR, Spain SG, Thurecht KJ, Aylott JW, Chan WC, Garnett MC, et al. Enhanced uptake of nanoparticle drug carriers via a thermoresponsive shell enhances cytotoxicity in a cancer cell line. Biomater Sci. 2013;1:434. doi: 10.1039/c2bm00184e.CrossRefGoogle Scholar
  59. 59.
    Meyer DE, Shin BC, Kong GA, Dewhirst MW, Chilkoti A. Drug targeting using thermally responsive polymers and local hyperthermia. J Control Release. 2001;74:213–24. doi: 10.1016/S0168-3659(01)00319-4.CrossRefGoogle Scholar
  60. 60.
    Wu C, Han D, Chen T, Peng L, Zhu G, You M, et al. Building a multifunctional aptamer-based DNA nanoassembly for targeted cancer therapy. J Am Chem Soc. 2013;. doi: 10.1021/ja4094617.Google Scholar
  61. 61.
    Tian Q, Zhang C-N, Wang X-H, Wang W, Huang W, Cha R-T, et al. chitosan/poly(ethylene glycol) nanoparticles for liver-targeted delivery. Biomaterials. 2010;31:4748–56. doi: 10.1016/j.biomaterials.2010.02.042.CrossRefGoogle Scholar
  62. 62.
    Zhang C, Wang W, Liu T, Wu Y, Guo H, Wang P, et al. Doxorubicin-loaded glycyrrhetinic acid-modified alginate nanoparticles for liver tumor chemotherapy. Biomaterials. 2012;33:2187–96. doi: 10.1016/j.biomaterials.2011.11.045.CrossRefGoogle Scholar
  63. 63.
    Macewan SR, Chilkoti A. Controlled apoptosis by a thermally toggled nanoscale amplifier of cellular uptake. Nano Lett. 2014;14:2058–64. doi: 10.1021/nl5002313.CrossRefGoogle Scholar
  64. 64.
    McDaniel JR, MacEwan SR, Li X, Radford DC, Landon CD, Dewhirst M, et al. Rational design of “heat seeking” drug loaded polypeptide nanoparticles that thermally target solid tumors. Nano Lett. 2014;14:2890–5. doi: 10.1021/nl5009376.CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2015

Authors and Affiliations

  • Yukun Wu
    • 1
  • Chengling Yang
    • 1
  • Quanyong Lai
    • 1
  • Qian Zhang
    • 1
  • Wei Wang
    • 1
  • Zhi Yuan
    • 1
  1. 1.Key Laboratory of Functional Polymer Materials of Ministry of Education and Institute of Polymer ChemistryNankai University, and Collaborative Innovation Center of Chemical Science and Engineering (Tianjin)TianjinChina

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