SCIENCE CHINA Materials, Volume 58, Issue 8: 640-648(2015) https://doi.org/10.1007/s40843-015-0073-y

The formation of intracellular nanoparticles correlates with cisplatin resistance

Meng Cao1,2,†, Fangzhou Liu1,3,4,†, Xiquan Zhang1,5,†, Ming Zheng5, Ziqi Ye1,6, Weiwei Chang1,5, Min Ji1, Xi Zhan3,*, Ning Gu1,*
More info
  • ReceivedJun 18, 2015
  • AcceptedJul 15, 2015
  • PublishedAug 11, 2015


Patients treated with the cisplatin often develop strong resistance to the drug after prolonged treatments, ultimately resulting in limited clinical efficacy. One of the possible mechanisms is that the internalized compound may be inactivated before getting access to the nucleus where cisplatin forms a complex with the genomic DNA and triggers a cell death program. However, the nature and intracellular fate of inactivated cisplatin is poorly illustrated. In the present study, we reported for the first time the presence of platinum nanoparticles (Pt-NPs) in the cytoplasm of cells treated with cisplatin. Further analysis also evidenced a correlation of the increased intracellular Pt-NPs formation with cisplatin resistance, and confirmed the process was glutathione S-transferase relevant. Our data suggest that tumor cells may develop cisplatin resistance by converting the drug into less toxic intracellular Pt-NPs, thereby impeding the drug from targeting its substrates.


This work was Funded by the National Key Basic Research Program of China (2011CB933503), the National Natural Science Foundation of China (NSFC) for Key Project of International Cooperation (61420106012), the Special Funds of National Natural Science Foundation of China For Basic Research Projects of Scientific Instruments (61127002), the Special Project on the Development of National Key Scientific Instruments and Equipment of China (2011YQ03013403), and China Postdoctoral Science Foundation funded project (2013M541592).

Interest statement

The authors declare that they have no conflict of interest.

Contributions statement

Gu N, Zhan X, Ji M and Cao M conceived and designed the reported research. Liu F and Zhang X prepared the samples and performed the TEM experiments, assisted by Zheng M. Cao M, Ye Z and Chang W maintained the four cell lines and performed cell viability assays as well as western blot. Cao M, Chang W and Zheng M designed the particle validation experiments and performed spectrometry. Cao M, Liu F and Zhang X analyzed the data. Gu N, Zhan X, Ji M and Cao M wrote the manuscript. All authors discussed the results and commented on the manuscript.

Author information

Meng Cao was born in 1983. He received his PhD degree in biomedical engineering from the School of Biological Science & Medical Engineering, Southeast University, Nanjing, China, in 2013. Currently he is a postdoctoral fellow in Professor Ning Gu’s group. His research interests mainly focus on biochemistry and anti-tumor therapeutics.

Ning Gu was born in 1964. He received his PhD degree in biomedical engineering from the Department of Biomedical Engineering, Southeast University, Nanjing, China, in 1996. Currently he is a Changjiang Scholar Professor and NSFC Outstanding Young Investigator Fund Winner at the School of Biological Science and Medical Engineering, Southeast University. He also serves as the president of Jiangsu Society of Biomedical Engineering, the director of the Research Center for Nanoscale Science and Technology of Southeast University. His research interests include biomaterials, nanobiology, medical imaging, and advanced instrument development.


Supplementary information

Supplementary information including the SEM examination of particles generated from cisplatin is available in the online version of the paper.


[1] Prestayko AW, Daoust JC, Issell BF, et al. Cisplatin (cis-diamminedichloroplatinum-II). Cancer Treat Rev, 1979, 6: 17–39. Google Scholar

[2] Chen X, Wu Y, Dong H, et al. Platinum-based agents for individualized cancer treatment. Curr Mol Med, 2013, 13: 1603–1612. Google Scholar

[3] Maccio A, Madeddu C. Cisplatin: an old drug with a newfound efficacy-from mechanisms of action to cytotoxicity. Expert Opin Pharmaco, 2013, 14: 1839–1857. Google Scholar

[4] Stathopoulos GP. Cisplatin: process and future. J Buon, 2013, 18: 564–569. Google Scholar

[5] Perry DJ, Weltz MD, Brown AW, et al. Vinblastine, bleomycin and cisplatin for recurrent or metastatic squamous-cell carcinoma of the head and neck. Cancer, 1982, 50: 2257–2260. Google Scholar

[6] Leipzig B. Cisplatin sensitization to radiotherapy of squamous-cell carcinomas of the head and neck. Am J Surg, 1983, 146: 462–465. Google Scholar

[7] Brizel DM, Albers ME, Fisher SR, et al. Hyperfractionated irradiation with or without concurrent chemotherapy for locally advanced head and neck cancer. New Engl J Med, 1998, 338: 1798–1804. Google Scholar

[8] Bernier J, Domenge C, Ozsahin M, et al. Postoperative irradiation with or without concomitant chemotherapy for locally advanced head and neck cancer. New Engl J Med, 2004, 350: 1945–1952. Google Scholar

[9] Khuri FR, Nemunaitis J, Ganly I, et al. A controlled trial of intratumoral ONYX-015, a selectively-replicating adenovirus, in combination with cisplatin and 5-fluorouracil in patients with recurrent head and neck cancer. Nat Med, 2000, 6: 879–885. Google Scholar

[10] Posner MR, Hershock DM, Blajman CR, et al. Cisplatin and fluorouracil alone or with docetaxel in head and neck cancer. New Engl J Med, 2007, 357: 1705–1715. Google Scholar

[11] Jamieson ER, Lippard SJ. Structure, recognition, and processing of cisplatin-DNA adducts. Chem Rev, 1999, 99: 2467–2498. Google Scholar

[12] Jordan P, Carmo-Fonseca M. Molecular mechanisms involved in cisplatin cytotoxicity. Cell Mol Life Sci, 2000, 57: 1229–1235. Google Scholar

[13] Cepeda V, Fuertes MA, Castilla J, et al. Biochemical mechanisms of cisplatin cytotoxicity. Anticancer Agents Med Chem, 2007, 7: 3–18. Google Scholar

[14] Dasari S, Tchounwou PB. Cisplatin in cancer therapy: molecular mechanisms of action. Eur J Pharmacol, 2014, 740: 364–378. Google Scholar

[15] Siddik ZH. Cisplatin: mode of cytotoxic action and molecular basis of resistance. Oncogene, 2003, 22: 7265–7279. Google Scholar

[16] Shen DW, Pouliot LM, Hall MD, et al. Cisplatin resistance: a cellular self-defense mechanism resulting from multiple epigenetic and genetic changes. Pharmacol Rev, 2012, 64: 706–721. Google Scholar

[17] Galluzzi L, Senovilla L, Vitale I, et al. Molecular mechanisms of cisplatin resistance. Oncogene, 2012, 31: 1869–1883. Google Scholar

[18] Galluzzi L, Vitale I, Michels J, et al. Systems biology of cisplatin resistance: past, present and future. Cell Death Dis, 2014, 5: e1257. Google Scholar

[19] Arnesano F, Losacco M, Natile G. An updated view of cisplatin transport. Eur J Inorg Chem, 2013, 2701–2711. Google Scholar

[20] Katano K, Kondo A, Safaei R, et al. Acquisition of resistance to cisplatin is accompanied by changes in the cellular pharmacology of copper. Cancer Res, 2002, 62: 6559–6565. Google Scholar

[21] Tsai CY, Larson CA, Safaei R, et al. Molecular modulation of the copper and cisplatin transport function of CTR1 and its interaction with IRS-4. Biochem Pharmacol, 2014, 90: 379–387. Google Scholar

[22] Nishimura T, Newkirk K, Sessions RB, et al. Immunohistochemical staining for glutathione S-transferase predicts response to platinum-based chemotherapy in head and neck cancer. Clin Cancer Res, 1996, 2: 1859–1865. Google Scholar

[23] Cullen KJ, Newkirk KA, Schumaker LM, et al. Glutathione S-transferase π amplification is associated with cisplatin resistance in head and neck squamous cell carcinoma cell lines and primary tumors. Cancer Res, 2003, 63: 8097–8102. Google Scholar

[24] Shiga H, Heath EI, Rasmussen AA, et al. Prognostic value of p53, glutathione S-transferase π, and thymidylate synthase for neoadjuvant cisplatin-based chemotherapy in head and neck cancer. Clin Cancer Res, 1999, 5: 4097–4104. Google Scholar

[25] Chiu CY, Ruan L, Huang Y. Biomolecular specificity controlled nanomaterial synthesis. Chem Soc Rev, 2013, 42: 2512–2527. Google Scholar

[26] Faramarzi MA, Sadighi A. Insights into biogenic and chemical production of inorganic nanomaterials and nanostructures. Adv Colloid Interface Sci, 2013, 189-190: 1–20. Google Scholar

[27] Konishi Y, Ohno K, Saitoh N, et al. Bioreductive deposition of platinum nanoparticles on the bacterium shewanella algae. J Biotechnol, 2007, 128: 648–653. Google Scholar

[28] Song JY, Kwon EY, Kim BS. Biological synthesis of platinum nanoparticles using Diopyros kaki leaf extract. Bioprocess Biosyst Eng, 2010, 33: 159–164. Google Scholar

[29] Venu R, Ramulu TS, Anandakumar S, et al. Bio-directed synthesis of platinum nanoparticles using aqueous honey solutions and their catalytic applications. Colloids and Surfaces A: Physicochem Eng Aspects, 2011, 384: 733–738. Google Scholar

[30] Govender Y, Riddin TL, Gericke M, et al. On the enzymatic formation of platinum nanoparticles. J Nanopart Res, 2010, 12: 261–271. Google Scholar

[31] Aika K, Ban LL, Okura I, et al. Chemisorption and catalytic activity of a set of platinum catalysts. J Res Inst Catalysis Hokkaido Univ, 1976, 24: 54–64. Google Scholar

[32] Li M, Yang DP, Wang XS, et al. Mixed protein-templated luminescent metal clusters (Au and Pt) for H2O2 sensing. Nanoscale Res Lett, 2013, 8: 182. Google Scholar

[33] Li YJ, Whyburn GP, Huang Y. Specific peptide regulated synthesis of ultrasmall platinum nanocrystals. J Am Chem Soc, 2009, 131: 15998–15999. Google Scholar

[34] Konieczny P, Goralczyk AG, Szmyd R, et al. Effects triggered by platinum nanoparticles on primary keratinocytes. Int J Nanomed, 2013, 8: 3963–3975. Google Scholar

[35] Yamagishi Y, Watari A, Hayata Y, et al. Acute and chronic nephrotoxicity of platinum nanoparticles in mice. Nanoscale Res Lett, 2013, 8: 395. Google Scholar

[36] Elder A, Yang H, Gwiazda R, et al. Testing nanomaterials of unknown toxicity: an example based on platinum nanoparticles of different shapes. Adv Mater, 2007, 19: 3124–3129. Google Scholar

[37] Asharani PV, Xinyi N, Hande MP, et al. DNA damage and p53-mediated growth arrest in human cells treated with platinum nanoparticles. Nanomedicine, 2010, 5: 51–64. Google Scholar

[38] Asharani PV, Yi LW, Gong ZY, et al. Comparison of the toxicity of silver, gold and platinum nanoparticles in developing zebrafish embryos. Nanotechnology, 2011, 5: 43–54. Google Scholar

[39] Gehrke H, Pelka J, Hartinger CG, et al. Platinum nanoparticles and their cellular uptake and DNA platination at non-cytotoxic concentrations. Arch Toxicol, 2011, 85: 799–812. Google Scholar

[40] Horie M, Kato H, Endoh S, et al. Evaluation of cellular influences of platinum nanoparticles by stable medium dispersion. Metallomics, 2011, 3: 1244–1252. Google Scholar

[41] Tsuchida S, Sato K. Glutathione transferases and cancer. Crit Rev Biochem Mol Biol, 1992, 27: 337–384. Google Scholar

[42] Hayes JD, Pulford DJ. The glutathione S-transferase supergene family: regulation of gst and the contribution of the isoenzymes to cancer chemoprotection and drug resistance. Crit Rev Biochem Mol Biol, 1995, 30: 445–600. Google Scholar

[43] Townsend DM, Tew KD. The role of glutathione-s-transferase in anti-cancer drug resistance. Oncogene, 2003, 22: 7369–7375. Google Scholar

[44] Henglein A, Ershov BG, Malow M. Absorption-spectrum and some chemical-reactions of colloidal platinum in aqueous-solution. J Phys Chem, 1995, 99: 14129–14136. Google Scholar

  • Figure 1

    Cisplatin-treated cells containing particle-like structures. HEp2 cells were grown in RPMI 1640 medium supplemented with 10% (v/v) FBS and treated with cisplatin at 0.5 µg/mL. After 24 h, cells were fixed and analyzed by TEM. (a) Particles as pointed by arrow heads were frequently found in the cytoplasm and intracellular vesicles of a treated cell. (b) However, these particles were absent in the untreated cell.

  • Figure 2

    Formation of nanoparticles in the lysate of cisplatin-treated cells. (a) Cells were treated with 0.8 µg/mL cisplatin for 12 h and lysed subsequently by sonication. The lysate was then examined by TEM, which revealed many particles with sizes around 2–5 nm. (b) The lysate of HEp2 cells were prepared and incubated with 0.8 µg/mL cisplatin for 12 h. The cisplatin treated lysate was examined by TEM, which demonstrated the presence of numerous amounts of particles with sizes similar to those as shown in (a). (c) Synthesized nanoparticles were prepared by incubation of sodium citrate with H2PtCl6 and were examined by TEM. (d) Nanoparticles were also formed in cell lysates that were incubated with 1 µg/mL H2PtCl6 for 12 h.

  • Figure 3

    Spectral property and sizes of cisplatin-derived particles. (a) HEp2 cells were treated by cisplatin from 0 to 12 h and subsequently examined by fluorescent microscopy (top panels) or phase-contrast microscopy (bottom panels). (b) Synthesized Pt-NPs were mixed with different doses of cell lysates and subsequently examined for absorbance at UV spectra from 200 to 300 nm. (c) The emission spectrum determination. (d) Cell lysates of different amounts of proteins were incubated with 0.25 mg/mL cisplatin for 2 h at 37°C under different conditions as indicated. The treated lysates were examined by DLS for the hydrodynamic sizes of nanoparticles. NA means not detectable.

  • Figure 4

    Characterization of lattice spacing of cisplatin-derived nanoparticles formed under different conditions. (a) The lattice spacing was estimated by TEM of the particles formed in the lysates of HEp2 cells treated with cisplatin at 0.8 µg/mL for 12 h; (b) cell lysates incubated with 0.8 µg/mL cisplatin for 12 h; (c) cisplatin molecules that were incubated with NADPH for 2 h at 37°C; (d) cisplatin incubated with GSH for 2 h at 37°C; (e) cisplatin incubated with ascorbic acid for 2 h at 37°C; (f) cisplatin incubated with nucleotides for 2 h at 37°C. A presentative lattice spacing for the particles in each sample was indicated.

  • Figure 5

    Pt-NPs have poor cytotoxic activity towards tumor cells. Cells were treated with either cisplatin or Pt-NPs at concentrations as indicated. After 24 h, cell viability was analyzed by CC8 assay. (a) HEp2; (b) FaDu.

  • Figure 6

    The formation of intracellular nanoparticles correlates with GST and cisplatin resistance. (a) HEp2 and FaDu cells were treated with 0.8 µg/mL cisplatin for 12 h and lysed. The lysates were analyzed for the UV absorbance. (b) Cells grown in the absence of cisplatin were lysed and the cell lysates were analyzed for the expression of GST by Western blot. As the loading control, the samples were also analyzed for α-tubulin expression.

  • Figure 7

    Schematic illustration of the possible relevance between intracellular Pt-nanoparticles formation and cisplatin resistance.

Copyright 2020 Science China Press Co., Ltd. 《中国科学》杂志社有限责任公司 版权所有

京ICP备18024590号-1       京公网安备11010102003388号