Fullerenol/iron nanocomposite diminishes doxorubicin-induced toxicity

  • Mariana SekeEmail author
  • Danijela PetrovicEmail author
  • Milica Labudovic Borovic
  • Ivana Borisev
  • Mirjana Novakovic
  • Zlatko Rakocevic
  • Aleksandar Djordjevic
Research Paper


Fullerenol C60(OH)24 with its spherical shape, symmetrical structure, 1 nm size and the ability to form polyionic nanoparticles in water solution, was used to synthesise a novel nanocomposite made of fullerenol nanoparticles (FNP) and iron ions (Fe2+). The FNP/Fe2+ nanocomposite was characterised by DLS and TEM-EDS analyses which have shown that the size distribution of FNP/Fe2+ stayed in the same scope as the size distribution of FNP, ranging from 11 to 60 nm. However, Fe2+ did affect the change of FNP’s zeta potential (− 49.2 mV), shifting it to more positive values (− 30.8 mV). In this study, it was assumed that FNP/Fe2+ could reduce the toxic effects of doxorubicin (Dox). Male Wistar rats were treated i.p. with FNP/Fe2+ nanocomposite 1 h prior to Dox treatment. At the subcellular level, the ultrastructural analysis revealed minor alterations sporadically displayed within the heart and liver tissues. Moreover, at the molecular level, the gene expressions analysis of mRNAs for catalase (heart and liver) and MnSOD (only liver) were significantly downregulated, indicating reduction in oxidative stress. Overall, the pretreatment with FNP/Fe2+ nanocomposite, followed by Dox application, significantly diminished harmful effects of the applied drug on the heart and liver, suggesting the potential protective effect of the nanocomposite on the healthy tissues.


Fullerenol Iron Doxorubicin Nanocomposite qRTR-PCR Ultrastructural analysis Nanomedicine Health effects 



This work has received a financial support from the Ministry of Education, Science and Technological Development, Republic of Serbia, Grant No. III 45005.We thank Professor Vladimir Srdic (Faculty of Technology, University of Novi Sad, Serbia) for DLS measurements and MSc Ivana Andjelkovic for technical support.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.


  1. Aggarwal P, Hall JB, McLeland CB, Dobrovolskaia MA, McNeil SE (2009) Nanoparticle interaction with plasma proteins as it relates to particle biodistribution, biocompatibility and therapeutic efficacy. Adv Drug Deliv Rev 61:428–437CrossRefGoogle Scholar
  2. Alcolado R, Arthur MJP, Iredale JP (1997) Pathogenesis of liver fibrosis. Clin Sci 92:103–112CrossRefGoogle Scholar
  3. Anderson R, Barron AR (2005) Reaction of hydroxyfullerene with metal salts: a route to remediation and immobilization. J Am Chem Soc 127:10458–10459CrossRefGoogle Scholar
  4. Aryal B, Jeong J, Ashutosh Rao V (2014) Doxorubicin-induced carbonylation and degradation of cardiac myosin binding protein C promote cardiotoxicity. Proc Natl Acad Sci 111:2011–2016CrossRefGoogle Scholar
  5. Baimanov D, Cai R, Chen C (2019) Understanding the chemical nature of nanoparticle-protein interactions. Bioconjug ChemGoogle Scholar
  6. Borović ML, Ičević I, Kanački Z, Žikić D, Seke M, Injac R, Djordjević A (2014) Effects of fullerenol C60(OH)24 nanoparticles on a single-dose doxorubicin-induced cardiotoxicity in pigs: an ultrastructural study. Ultrastruct Pathol 38:150–163CrossRefGoogle Scholar
  7. Cai X, Jia H, Liu Z, Hou B, Luo C, Feng Z, Li W, Liu J (2008) Polyhydroxylated fullerene derivative C60(OH)24 prevents mitochondrial dysfunction and oxidative damage in an MPP+-induced cellular model of Parkinson’s disease. J Neurosci Res 86:3622–3634CrossRefGoogle Scholar
  8. Carvalho C, Santos RX, Cardoso S, Correia S, Oliveira PJ, Santos MS, Moreira PI (2009) Doxorubicin: the good, the bad and the ugly effect. Curr Med Chem 16:3267–3285CrossRefGoogle Scholar
  9. Chen M, von Mikecz A (2005) Formation of nucleoplasmic protein aggregates impairs nuclear function in response to SiO2 nanoparticles. Exp Cell Res 305:51–62CrossRefGoogle Scholar
  10. Chen J, Rider DA, Ruan R (2006) Identification of valid housekeeping genes and antioxidant enzyme gene expression change in the aging rat liver. J Gerontol Ser A Biol Med Sci 61(2006):20–27CrossRefGoogle Scholar
  11. Childs AC, Phaneuf SL, Dirks AJ, Phillips T, Leeuwenburgh C (2002) Doxorubicin treatment in vivo causes cytochrome C release and cardiomyocyte apoptosis, as well as increased mitochondrial efficiency, superoxide dismutase activity, and Bcl-2: Bax ratio. Cancer Res 62:4592–4598Google Scholar
  12. Deavall DG, Martin EA, Horner JM, Roberts R (2012) Drug-induced oxidative stress and toxicity. J Toxicol 2012:645460CrossRefGoogle Scholar
  13. Deng ZJ, Mortimer G, Schiller T, Musumeci A, Martin D, Minchin RF (2009) Differential plasma protein binding to metal oxide nanoparticles. Nanotechnology 20:455101CrossRefGoogle Scholar
  14. Dixon SJ, Lemberg KM, Lamprecht MR, Skouta R, Zaitsev EM, Gleason CE, Patel DN et al (2012) Ferroptosis: an iron-dependent form of nonapoptotic cell death. Cell 149:1060–1072CrossRefGoogle Scholar
  15. Djordjević A, Vojinović-Miloradov M, Petranović N, Devečerski A, Lazar D, Ribar B (1998) Catalytic preparation and characterization of C60Br24. Fuller Sci Technol 6:689–694CrossRefGoogle Scholar
  16. Djordjevic A, Canadanovic-Brunet J, Vojinovic-Miloradov M, Bogdanovic G (2005) Antioxidant properties and hypothetical radical mechanism of fullerol C60(OH)24. Oxid Commun 27:806–812Google Scholar
  17. Djordjevic A, Ajdinović B, Dopudja M, Trajkovic S, Milovanovic Z, Maksin T, Neskovic O, Bogdanović VB, Trpkov Djordje T, Cveticanin J (2011) Scintigraphy of the domestic dog using [Tc-99m(CO)(3)(H2O)(3)]-C-60(OH)(22-24). Dig J Nanomater Biostructures 6(2011):99–106Google Scholar
  18. Djordjevic A, Srdjenovic B, Seke M, Petrovic D, Injac R, Mrdjanovic J (2015) Review of synthesis and antioxidant potential of fullerenol nanoparticles. J Nanomater 16:280Google Scholar
  19. Đorđević AN, Ičević ID, Bogdanović VV (2009) Complex with fullerenol and copper (II). Hemijska Industrija 63:171–175CrossRefGoogle Scholar
  20. Doroshow JH, Locker GY, Myers CE (1980) Enzymatic defenses of the mouse heart against reactive oxygen metabolites: alterations produced by doxorubicin. J Clin Invest 65:128–135CrossRefGoogle Scholar
  21. Dugan LL, Gabrielsen JK, Yu Shan P, Lin T-S, Choi DW (1996) Buckminsterfullerenol free radical scavengers reduce excitotoxic and apoptotic death of cultured cortical neurons. Neurobiol Dis 3:129–135CrossRefGoogle Scholar
  22. Engin AB, Nikitovic D, Neagu M, Henrich-Noack P, Docea AO, Shtilman MI, Golokhvast K, Tsatsakis AM (2017) Mechanistic understanding of nanoparticles’ interactions with extracellular matrix: the cell and immune system. Part Fibre Toxicol 14:22CrossRefGoogle Scholar
  23. Fonseca-Nunes, Ana, Paula Jakszyn, and Antonio Agudo. "Iron cancer risk-a systematic review and meta-analysis of the epidemiological evidence." Cancer epidemiology and prevention biomarkers 23 (2014): 12-31Google Scholar
  24. Fröhlich E (2012) The role of surface charge in cellular uptake and cytotoxicity of medical nanoparticles. Int J Nanomedicine 7:5577CrossRefGoogle Scholar
  25. Goormaghtigh E, Chatelain P, Caspers J, Ruysschaert JM (1980) Evidence of a specific complex between adriamycin and negatively-charged phospholipids. Biochim Biophys Acta Biomembr 597:1–14CrossRefGoogle Scholar
  26. Guenancia C, Li N, Hachet O, Rigal E, Cottin Y, Dutartre P, Rochette L, Vergely C (2015) Paradoxically, iron overload does not potentiate doxorubicin-induced cardiotoxicity in vitro in cardiomyocytes and in vivo in mice. Toxicol Appl Pharmacol 284:152–162CrossRefGoogle Scholar
  27. Gutteridge JM (1984) Lipid peroxidation and possible hydroxyl radical formation stimulated by the self-reduction of a doxorubicin-iron (III) complex. Biochem Pharmacol 33:1725–1728CrossRefGoogle Scholar
  28. Haber F, Weiss J (1932) Über die katalyse des hydroperoxydes. Naturwissenschaften 20:948–950CrossRefGoogle Scholar
  29. Heimann J, Morrow L, Anderson RE, Barron AR (2015) Understanding the relative binding ability of hydroxyfullerene to divalent and trivalent metals. Dalton Trans 44:4380–4388CrossRefGoogle Scholar
  30. Ichikawa Y, Ghanefar M, Bayeva M, Wu R, Khechaduri A, Prasad SV, Mutharasan RK, Naik TJ, Ardehali H (2014) Cardiotoxicity of doxorubicin is mediated through mitochondrial iron accumulation. J Clin Invest 124:617CrossRefGoogle Scholar
  31. Injac R, Perse M, Obermajer N, Djordjevic-Milic V, Prijatelj M, Djordjevic A, Cerar A, Strukelj B (2008a) Potential hepatoprotective effects of fullerenol C60(OH)24 in doxorubicin-induced hepatotoxicity in rats with mammary carcinomas. Biomaterials 29:3451–3460CrossRefGoogle Scholar
  32. Injac R, Perse M, Boskovic M, Djordjevic-Milic V, Djordjevic A, Hvala A, Cerar A, Strukelj B (2008b) Cardioprotective effects of fullerenol C60(OH)24 on a single dose doxorubicin-induced cardiotoxicity in rats with malignant neoplasm. Technol Cancer Res Treat 7:15–25CrossRefGoogle Scholar
  33. Injac R, Boskovic M, Perse M, Koprivec-Furlan E, Cerar A, Djordjevic A, Strukelj B (2008c) Acute doxorubicin nephrotoxicity in rats with malignant neoplasm can be successfully treated with fullerenolC60(OH)24 via suppression of oxidative stress. Pharmacol Rep 60:742Google Scholar
  34. Injac R, Perse M, Cerne M, Potocnik N, Radic N, Govedarica B, Djordjevic A, Cerar A, Strukelj B (2009a) Protective effects of fullerenol C60(OH)24 against doxorubicin-induced cardiotoxicity and hepatotoxicity in rats with colorectal cancer. Biomaterials 30:1184–1196CrossRefGoogle Scholar
  35. Injac R, Radic N, Govedarica B, Perse M, Cerar A, Djordjevic A, Strukelj B (2009b) Acute doxorubicin pulmotoxicity in rats with malignant neoplasm is effectively treated with fullerenolC60(OH)24 through inhibition of oxidative stress. Pharmacol Rep 61:335–342CrossRefGoogle Scholar
  36. Isakovic A, Markovic Z, Todorovic-Markovic B, Nikolic N, Vranjes-Djuric S, Mirkovic M, Dramicanin M et al (2006) Distinct cytotoxic mechanisms of pristine versus hydroxylated fullerene. Toxicol Sci 91:173–183CrossRefGoogle Scholar
  37. Ji ZQ, Sun H, Wang H, Xie Q, Liu Y, Wang Z (2006) Biodistribution and tumor uptake of C60(OH)x in mice. J Nanopart Res 8:53–63CrossRefGoogle Scholar
  38. Jović DS, Seke MN, Djordjevic AN, Mrđanović JZ, Aleksić LD, Bogdanović GM, Pavić AB, Plavec J (2016) Fullerenol nanoparticles as a new delivery system for doxorubicin. RSC Adv 6:38563–38578CrossRefGoogle Scholar
  39. Koppenol WH (1993) The centennial of the Fenton reaction. Free Radic Biol Med 15:645–651CrossRefGoogle Scholar
  40. Li T, Danelisen I, Belló-Klein A, Singal PK (2000) Effects of probucol on changes of antioxidant enzymes in adriamycin-induced cardiomyopathy in rats. Cardiovasc Res 46:523–530CrossRefGoogle Scholar
  41. Li Y, Luo HB, Zhang HY, Guo Q, Yao HC, Li JQ, Chang Q et al (2016) Potential hepatoprotective effects of fullerenol nanoparticles on alcohol-induced oxidative stress by ROS. RSC Adv 6:31122–31130CrossRefGoogle Scholar
  42. Link G, Tirosh R, Pinson A, Hershko C (1996) Role of iron in the potentiation of anthracyclinecardiotoxicity: identification of heart cell mitochondria as a major site of iron-anthracycline interaction. J Lab Clin Med 127:272–278CrossRefGoogle Scholar
  43. Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2− ΔΔCT method. Methods 25:402–408CrossRefGoogle Scholar
  44. Manoli I, Alesci S, Blackman MR, Su YA, Rennert OM, Chrousos GP (2007) Mitochondria as key components of the stress response. Trends Endocrinol Metab 18:190–198CrossRefGoogle Scholar
  45. Marzenell P, Hagen H, Sellner L, Zenz T, Grinyte R, Pavlov V, Daum S, Mokhir A (2013) Aminoferrocene-based prodrugs and their effects on human normal and cancer cells as well as bacterial cells. J Med Chem 56:6935–6944CrossRefGoogle Scholar
  46. Minotti G, Recalcati S, Mordente A, Liberi G, Calafiore AM, Mancuso C, Preziosi P, Cairo G (1998) The secondary alcohol metabolite of doxorubicin irreversibly inactivates aconitase/iron regulatory protein-1 in cytosolic fractions from human myocardium. FASEB J 12:541–552CrossRefGoogle Scholar
  47. Minotti G, Ronchi R, Salvatorelli E, Menna P, Cairo G (2001) Doxorubicin irreversibly inactivates iron regulatory proteins 1 and 2 in cardiomyocytes: evidence for distinct metabolic pathways and implications for iron-mediated cardiotoxicity of antitumor therapy. Cancer Res 61:8422–8428Google Scholar
  48. Mirkov SM, Djordjevic AN, Andric NL, Andric SA, Kostic TS, Bogdanovic GM, Vojinovic-Miloradov MB, Kovacevic RZ (2004) Nitric oxide-scavenging activity of polyhydroxylatedfullerenol, C60(OH)24. Nitric Oxide 11:201–207CrossRefGoogle Scholar
  49. Mobaraki M, Faraji A, Zare M, Dolati P, Ataei M, Dehghan Manshadi HR (2017) Molecular mechanisms of cardiotoxicity: a review on major side-effect of doxorubicin. Indian J Pharm Sci 79:335–344CrossRefGoogle Scholar
  50. Moulin M, Piquereau J, Mateo P, Fortin D, Rucker-Martin C, Gressette M, Lefebvre F et al (2015) Sexual dimorphism of doxorubicin-mediated cardiotoxicity: potential role of energy metabolism remodeling. Circ Heart Fail 8:98–108CrossRefGoogle Scholar
  51. Myers CE, Gianni L, Simone CB, Klecker R, Greene R (1982) Oxidative destruction of erythrocyte ghost membranes catalyzed by the doxorubicin-iron complex. Biochemistry 21:1707–1713CrossRefGoogle Scholar
  52. Olson RD, Mushlin PS, Brenner DE, Fleischer S, Cusack BJ, Chang BK, Boucek RJ (1988) Doxorubicin cardiotoxicity may be caused by its metabolite, doxorubicinol. Proc Natl Acad Sci 85:3585–3589CrossRefGoogle Scholar
  53. Parker MA, King V, Howard KP (2001) Nuclear magnetic resonance study of doxorubicin binding to cardiolipin containing magnetically oriented phospholipid bilayers. Biochim Biophys Acta Biomembr 1514:206–216CrossRefGoogle Scholar
  54. Petrovic D, Seke M, Srdjenovic B, Djordjevic A (2015) Applications of anti/prooxidant fullerenes in nanomedicine along with fullerenes influence on the immune system. J Nanomater 16:279Google Scholar
  55. Petrovic D, Seke M, Labudovic Borovic M, Jovic D, Borisev I, Srdjenovic B, Rakocevic Z, Pavlovic V, Djordjevic A (2018) Hepatoprotective effect of fullerenol/doxorubicin nanocomposite in acute treatment of healthy rats. Exp Mol Pathol 104:199–211CrossRefGoogle Scholar
  56. Pradhan A, Pinheiro JP, Seena S, Pascoal C, Cássio F (2014) Polyhydroxyfullerene binds cadmium ions and alleviates metal-induced oxidative stress in Saccharomyces cerevisiae. Appl Environ Microbiol 80:5874–5881CrossRefGoogle Scholar
  57. Seke M, Petrovic D, Djordjevic A, Jovic D, Labudovic Borovic M, Kanacki Z, Jankovic M (2016) Fullerenol/doxorubicin nanocomposite mitigates acute oxidative stress and modulates apoptosis in myocardial tissue. Nanotechnology 27:485101CrossRefGoogle Scholar
  58. Siddiqi S, Sussman MA (2014) The heart: mostly postmitotic or mostly premitotic? Myocyte cell cycle, senescence, and quiescence. Can J Cardiol 30:1270–1278CrossRefGoogle Scholar
  59. Slavic M, Djordjevic A, Radojicic R, Milovanovic S, Orescanin-Dusic Z, Rakocevic Z, Spasic MB, Blagojevic D (2013) Fullerenol C60(OH)24 nanoparticles decrease relaxing effects of dimethyl sulfoxide on rat uterus spontaneous contraction. J Nanopart Res 15:1650CrossRefGoogle Scholar
  60. Srdjenovic B, Milic-Torres V, Grujic N, Stankov K, Djordjevic A, Vasovic V (2010) Antioxidant properties of fullerenol C60(OH)24 in rat kidneys, testes, and lungs treated with doxorubicin. Toxicol Mech Methods 20:298–305CrossRefGoogle Scholar
  61. Sugino N, Telleria CM, Gibori G (1998) Differential regulation of copper-zinc superoxide dismutase and manganese superoxide dismutase in the rat corpus luteum: induction of manganese superoxide dismutase messenger ribonucleic acid by inflammatory cytokines. Biol Reprod 59:208–215CrossRefGoogle Scholar
  62. Swain K, Yusuf MY, Kalender S (2005) Doxorubicin hepatotoxicity and hepatic free radical metabolism in rats: the effects of vitamin E and catechin. Toxicology 209:39–45CrossRefGoogle Scholar
  63. Ta HT, Li Z, Hagemeyer CE, Cowin G, Zhang S, Palasubramaniam J, Alt K, Wang X, Peter K, Whittaker AK (2017) Molecular imaging of activated platelets via antibody-targeted ultra-small iron oxide nanoparticles displaying unique dual MRI contrast. Biomaterials 134:31–42CrossRefGoogle Scholar
  64. Theil, Elisabeth C.,Gunther L.Eichhorn, and Luigi G. Morzilli. "Iron bindingproteins without cofactors or sulfur clusters."" Advances in inorganic biochemistry 5 (1983): 1-38Google Scholar
  65. Thorn CF, Oshiro C, Marsh S, Hernandez-Boussard T, McLeod H, Klein TE, Altman RB (2011) Doxorubicin pathways: pharmacodynamics and adverse effects. Pharmacogenet Genomics 21:440CrossRefGoogle Scholar
  66. Torres VM, Srdjenovic B, Jacevic V, Dragojevic Simic V, Djordjevic A, Simplício AL (2010) Fullerenol C60(OH)24 prevents doxorubicin-induced acute cardiotoxicity in rats. Pharmacol Rep 62:707–718CrossRefGoogle Scholar
  67. Torti SV, Torti FM (2013) Iron and cancer: more ore to be mined. Nat Rev Cancer 13:342CrossRefGoogle Scholar
  68. Von Hoff DD, Layard MW, Basa P, Davis HL Jr, Von Hoff AL, Rozencweig M, Muggia FM (1979) Risk factors for doxorubicin-induced congestive heart failure. Ann Intern Med 91:710–717CrossRefGoogle Scholar
  69. Wang Z, Gao X, Zhao Y (2018) Mechanisms of Antioxidant Activities of Fullerenols from First-Principles Calculation. J Phys Chem A 122:8183–8190CrossRefGoogle Scholar
  70. Xia T, Kovochich M, Brant J, Hotze M, Sempf J, Oberley T, Sioutas C, Yeh JI, Wiesner MR, Nel AE (2006) Comparison of the abilities of ambient and manufactured nanoparticles to induce cellular toxicity according to an oxidative stress paradigm. Nano Lett 6:1794–1807CrossRefGoogle Scholar
  71. Yagmurca M, Bas O, Mollaoglu H, Sahin O, Nacar A, Karaman O, Songur A (2007) Protective effects of erdosteine on doxorubicin-induced hepatotoxicity in rats. Arch Med Res 38:380–385CrossRefGoogle Scholar
  72. Yang F, Teves SS, Kemp CJ, Henikoff S (2014) Doxorubicin, DNA torsion, and chromatin dynamics. Biochim Biophys Acta Rev Cancer 1845:84–89CrossRefGoogle Scholar
  73. Yang L-Y, Gao J-L, Gao T, Dong P, Ma L, Jiang F-L, Liu Y (2016) Toxicity of polyhydroxylated fullerene to mitochondria. J Hazard Mater 301:119–126CrossRefGoogle Scholar

Copyright information

© Springer Nature B.V. 2019

Authors and Affiliations

  1. 1.Institute of Nuclear Sciences “Vinca”University of BelgradeBelgradeSerbia
  2. 2.Department of Natural Sciences and Management in Education, Faculty of Education SomborUniversity of Novi SadNovi SadSerbia
  3. 3.Institute of Histology and Embryology “AleksandarDj. Kostic”, Faculty of MedicineUniversity of BelgradeBelgradeSerbia
  4. 4.Department of Chemistry, Biochemistry and Environmental Protection, Faculty of SciencesUniversity of Novi SadNovi SadSerbia

Personalised recommendations