Biological Trace Element Research

, Volume 186, Issue 2, pp 413–429 | Cite as

Interaction of Zn with Losartan. Activation of Intrinsic Apoptotic Signaling Pathway in Lung Cancer Cells and Effects on Alkaline and Acid Phosphatases

  • Valeria R. Martínez
  • María V. Aguirre
  • Juan S. Todaro
  • Oscar E. Piro
  • Gustavo A. Echeverría
  • Luciana G. Naso
  • Evelina G. Ferrer
  • Patricia A. M. WilliamsEmail author


A new losartan [2-butyl-5-chloro-3-[[4-[2-(2H-tetrazol-5-yl)phenyl]phenyl]methyl]imidazol-4-yl]methanol zinc(II) complex [Zn(Los)Cl], was synthesized and characterized. The crystal structure was determined by x-ray diffraction methods. When aqueous solutions of the ligand and the metal were mixed, the known and more soluble powder [Zn(Los)2].3H2O (ZnLos) complex has been obtained. The interactions with phosphatases showed a concerted mechanism displayed by the Zn ions and ZnLos up to 500 μM concentration: a decrease of the acid phosphatase (AcP) associated with an increase in the alkaline phosphatase (ALP) activities. The complex and ZnSO4 showed a cytotoxic behavior on human lung A549 cancer cell line at concentrations higher than 75 μM with reactive oxygen species (ROS) generation and GSH (and GSH/GSSG ratio) depletion. Apoptotic cells were observed using terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) method, a mechanism accompanied by upregulation of BAX protein, downregulation of Bcl-XL and release of caspase-3. The BAX/Bcl-XL ratio was found to be significantly higher in cells exposure to ZnLos than cells treated with ZnSO4, in agreement with the higher apoptotic percentage of cells found for the complex. Cell death was found to be produced by apoptosis and no necrosis has been observed. On the contrary, losartan exerted low effects on phosphatases, produced some reduction of cancer cell viability (concentrations > 250 μM, number of apoptotic cells similar to the basal) with low ROS depletion, without alteration of the GSH/GSSG and low BAX/Bcl-XL ratios. In the MRC-5, normal lung fibroblasts cell line only ZnSO4 at concentrations higher than 200 μM displays cytotoxic effects.

Graphical abstract

Interaction of Zn with losartan. Activation of intrinsic apoptotic signaling pathway in lung cancer cells and effects on alkaline and acid phosphatases


Losartan Zinc coordination Enzymatic inhibition Anticancer mechanism 



This work was supported by UNLP, CONICET (PIP 0611), CICPBA (PICyT 813/13), and ANPCyT (PICT-2016-1814, PME06 2804 and PICT06 2315), Argentina. VRM is a fellowship holder from ANPCyT. EGF, LGN, GAE, and OEP are research fellows of CONICET. PAMW is a research fellow of CICPBA, Argentina.

Supplementary material

12011_2018_1334_MOESM1_ESM.docx (622 kb)
ESM 1 (DOCX 622 kb)


  1. 1.
    Ferlay J, Shin HR, Bray F, Forman D, Mathers C, Parkin DM (2010) Estimates of worldwide burden of cancer in 2008: GLOBOCAN 2008. Int J Cancer 127:2893–2917. CrossRefPubMedPubMedCentralGoogle Scholar
  2. 2.
    Timmermans PB, Duncia JV, Carini DJ, Chiu AT, Wong PC, Wexler RR, Smith RD (1995) Discovery of losartan, the first angiotensin II receptor antagonist. J Hum Hypertens 5:S3–S18Google Scholar
  3. 3.
    Li J, Chen L, Yu P, Liu B, Zhu J, Yang Y (2014) Telmisartan exerts anti-tumor effects by activating peroxisome proliferator-activated receptor-γ in human lung adenocarcinoma A549 cells. Molecules 19:2862–2876. CrossRefPubMedGoogle Scholar
  4. 4.
    Gallagher PE, Tallant EA (2004) Inhibition of human lung cancer cell growth by angiotensin-(1-7). Carcinogenesis 25:2045–2052. CrossRefPubMedPubMedCentralGoogle Scholar
  5. 5.
    Islas MS, Luengo A, Franca CA, Griera Merino M, Calleros L, Rodríguez-Puyol M, Lezama L, Ferrer EG, Williams PAM (2016) Experimental and DFT characterization, antioxidant and anticancer activities of a cu(II)-irbesartan complex: structure-antihypertensive activity relationships in cu(II)-sartan complexes. J Biol Inorg Chem 21:851–863. CrossRefPubMedGoogle Scholar
  6. 6.
    Ho E (2004) Zinc deficiency, DNA damage and cancer risk. J Nutr Biochem 10:572–578CrossRefGoogle Scholar
  7. 7.
    Singh KP, Zaidi SI, Raisuddin S, Saxena AK, Murthy RC, Ray PK (1992) Effect of zinc on immune functions and host resistance against infection and tumor challenge. Immunopharmacol Immunotoxicol 14:813–840CrossRefGoogle Scholar
  8. 8.
    Prasad AS, Kucuk O (2002) Zinc in cancer prevention. Cancer Metastasis Rev 21:291–295. CrossRefPubMedGoogle Scholar
  9. 9.
    Kocdor H, Ates H, Aydin S, Cehreli R, Soyarat F, Kemanli P, Harmanci D, Cengiz H, Kocdor MA (2015) Zinc supplementation induces apoptosis and enhances antitumor efficacy of docetaxel in non-small-cell lung cancer. Dovepress 9:3899–3909. CrossRefGoogle Scholar
  10. 10.
    Braun LA, Rosenfeldt F (2013) Pharmaco-nutrient interactions—a systematic review of zinc and antihypertensive therapy. Int J Clin Pract 67:717–725. CrossRefPubMedGoogle Scholar
  11. 11.
    Martínez VR, Aguirre MV, Todaro JS, Piro OE, Echeverría GA, Ferrer EG, Williams PAM (2018) Azilsartan and its Zn(II) complex. Synthesis, anticancer mechanisms of action and binding to bovine serum albumin. Toxicol In Vitro 48:205–220. CrossRefGoogle Scholar
  12. 12.
    Teixeira JA, Siqueira AB (2016) Thermal and spectroscopic characterization, antioxidant evaluation and pyrolysis of losartan with some bivalent metals. J Anal Appl Pyrol 117:17–24. CrossRefGoogle Scholar
  13. 13.
    Lachowicz JI, Nurchi VM, Crisponi G, Jaraquemada-Pelaez MG, Caltagirone C, Peana M, Zoroddu MA, Szewczuk Z, Cooper GJS (2017) Complex formation equilibria of Cu(2+) and Zn(2+) with irbesartan and losartan. Eur J Pharm Sci 97:158–169. CrossRefPubMedGoogle Scholar
  14. 14.
    Fishman WH (1987) Alkaline phosphatase: an overview. Clin Biochem 20:387–392. CrossRefPubMedGoogle Scholar
  15. 15.
    Bozzo GG, Raghothama KG, Plaxton WC (2004) Structural and kinetic properties of a novel purple acid phosphatase from phosphate-starved tomato (Lycopersicon esculentum) cell cultures. Biochem J 377:419–428. CrossRefPubMedPubMedCentralGoogle Scholar
  16. 16.
    CrysAlisPro, Oxford Diffraction Ltd., version (release 15–09-2009 CrysAlis171.NET)Google Scholar
  17. 17.
    Sheldrick GM (2008) A short history of SHELX. Acta Crystallogr A64:112–122. CrossRefGoogle Scholar
  18. 18.
    Ferrer EG, Salinas MV, Correa MJ, Naso L, Barrio DA, Etcheverry SB, Lezama L, Rojo T, Williams PAM (2006) Synthesis, characterization, antitumoral and osteogenic activities of quercetin vanadyl (IV) complexes. J Biol Inorg Chem 11:791–801. CrossRefPubMedGoogle Scholar
  19. 19.
    Blum U, Schwedt G (1998) Inhibition behavior of acid phosphatase, phosphodiesterase I and adenosine deaminase as tools for trace metal analysis and speciation. Anal Chim Acta 360:101–108. CrossRefGoogle Scholar
  20. 20.
    Glaysher S, Cree I (2011) Isolation and culture of colon cancer cells and cell lines. Meth Mol Biol 731:135–140. CrossRefGoogle Scholar
  21. 21.
    Qin Y, Lu M, Gong X (2008) Dihydrorhodamine 123 superior to 2,7-dichlorodihydrofluorescein diacetate and dihydrorhodamine 6G in detecting intracellular hydrogen peroxide tumor cells. Cell Biol Int 32:224–228. CrossRefPubMedGoogle Scholar
  22. 22.
    Bradford M (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:249–254. CrossRefGoogle Scholar
  23. 23.
    Hissin PJ, Hilf R (1976) A fluorometric method for determination of oxidized and reduced glutathione in tissues. Anal Biochem 74:214–226. CrossRefPubMedGoogle Scholar
  24. 24.
    Negoescu A, Guillermet C, Lorimier P, Brambilla E, Labat-Moleur F (1998) Importance of DNA fragmentation in apoptosis with regard to TUNEL specificity. Biomed Pharmacother 52:252–258. CrossRefPubMedGoogle Scholar
  25. 25.
    Liu QY, Stein C (1997) Taxol and estramustine-induced modulation of human prostate cancer cell apoptosis via alteration in bcl-xL and bak expression. Clin Cancer Res 3(11):2039–2046PubMedGoogle Scholar
  26. 26.
    Aguirre MV, Juaristi JA, Alvarez MA, Brandan NC (2005) Characteristics of in vivo murine erythropoietic response to sodium orthovanadate. Chem Biol Interact 156:55–68. CrossRefPubMedGoogle Scholar
  27. 27.
    Ribble D, Goldstein NB, Norris DA, Shellman YG (2005) A simple technique for quantifying apoptosis in 96-well plates. BMC Biotechnol 5:1–7. CrossRefGoogle Scholar
  28. 28.
    Farrugia LJ (1997) ORTEP3 for windows. J Appl Crystallogr 30:565CrossRefGoogle Scholar
  29. 29.
    Franca CA, Etcheverry SB, Pis Diez R, Williams PAM (2009) Irbesartan: FTIR and Raman spectra. Density functional study on vibrational and NMR spectra. J Raman Spectrosc 40:1296–1300. CrossRefGoogle Scholar
  30. 30.
    Islas MS, Franca CA, Etcheverry SB, Ferrer EG, Williams PAM (2012) Computational study and spectroscopic investigations of antihypertensive drugs. Vibrat Spectrosc 62:143–151. CrossRefGoogle Scholar
  31. 31.
    Könczöl M, Goldenberg E, Ebeling S, Schäfer B, Garcia-Käufer M, Gminski R, Grobéty B, Rothen-Rutishauser B, Merfort I, Gieré R, Mersch-Sundermann V (2012) Cytotoxicity and genotoxicity of size-fractionated iron oxide (magnetite) in A549 human lung epithelial cells: role of ROS, JNK, and NF-κB. Chem Res Toxicol 25:2687–2703. CrossRefPubMedGoogle Scholar
  32. 32.
    Vink H (1994) Electrolytic conductivity of mixed electrolyte solutions. Ber Bunsenges Phys Chem 98:1039–1045CrossRefGoogle Scholar
  33. 33.
    de Oliveira RP, Felix FS, Angnes L (2012) A simple and precise conductometric method for the determination of losartan in pharmaceutical products. Cent Eur J Chem 10:1842–1849. CrossRefGoogle Scholar
  34. 34.
    Kellet KAB, Williams J, Vardy ERLC, Smith AD, Hooper NM (2011) Plasma alkaline phosphatase is elevated in Alzheimer's disease and inversely correlates with cognitive function. Int J Mol Epidemiol Genet 2:114–121Google Scholar
  35. 35.
    Thompson KH, McNeill JH, Orvig C (1999) Vanadium compounds as insulin mimics. Chem Rev 99:2561–2571. CrossRefPubMedGoogle Scholar
  36. 36.
    Dean RL (2002) Kinetic studies with alkaline phosphatase in the presence and absence of inhibitors and divalent cations. Biochem. Mol Biol Educ 30:401–407. CrossRefGoogle Scholar
  37. 37.
    Gellatly KS, Moorhead GBC, Duff SMC, Lefebvre DD, Plaxton WC (1994) Purification and characterization of a potato tuber. Acid phosphatase having significant phosphotyrosine phosphatase activity. Plant Physiol 106:223–232. CrossRefPubMedPubMedCentralGoogle Scholar
  38. 38.
    Schenk G, Miti N, Hanson GR, Comba P (2013) Purple acid phosphatase: a journey into the function and mechanism of a colorful enzyme. Coord Chem Rev 257:473–482. CrossRefGoogle Scholar
  39. 39.
    Rane SY, Badave KD, Ahmed K (2009) Insight into bio-physiological functions of acid phosphatase from potatoes (Solanum tuberosum): a bioanalytical approach. Indian J Chem 48A:15–24Google Scholar
  40. 40.
    Kuftinec MM, Miller SA, Kuftinec MM, Miller SA (1972) Alkaline and acid phosphatase activities during growth of long bones and mandibles. Calcif Tissue Res 9:173–178. CrossRefPubMedGoogle Scholar
  41. 41.
    De la Iglesia IS, López-Jorge CE, Gómez-Casares MT, Lemes Castellano A, Cabrera PM, López Brito J, Suárez Cabrera A, Molero Labarta T (2009) Induction of apoptosis in leukemic cell lines treated with captopril, trandolapril and losartan: a new role in the treatment of leukaemia for these agents. Leuk Res 33:810–816. CrossRefGoogle Scholar
  42. 42.
    Godugu C, Patel AR, Doddapaneni R, Marepally S, Jackson T, Singh M (2013) Inhalation delivery of Telmisartan enhances intratumoral distribution of nanoparticles in lung cancer models. J Control Release 172:86–95. CrossRefPubMedPubMedCentralGoogle Scholar
  43. 43.
    Zhao W, Song Q, Zhang Z, Mao L, Zheng W, Hu X, Lian H (2015) The kinetic response of the proteome in A549 cells exposed to ZnSO4 stress. PLoS One 10:e0133451. CrossRefPubMedPubMedCentralGoogle Scholar
  44. 44.
    Jiang D, Sullivan PG, Sensi SL, Steward O, Weiss JH (2001) Zn(2+) induces permeability transition pore opening and release of pro-apoptotic peptides from neuronal mitochondria. J Biol Chem 276:47524–47529CrossRefGoogle Scholar
  45. 45.
    Donadelli M, Dalla Pozza E, Scupoli MT, Costanzo C, Scarpa A, Palmieri M (2009) Intracellular zinc increase inhibits p53−/− pancreatic adenocarcinoma cell growth by ROS/AIF-mediated apoptosis. Biochim Biophys Acta 1793:273–280. CrossRefPubMedGoogle Scholar
  46. 46.
    Yuan N, Wang YH, Ll KJ, Zhao Y, Hu X, Mao L, Zhao WJ, Lian HZ, Zheng WJ (2012) Effects of exogenous zinc on the cellular zinc distribution and cell cycle of A549 cells. Biosci Biotechnol Biochem 76:2014–2020. CrossRefPubMedGoogle Scholar
  47. 47.
    Goodsell DS (2002) The molecular perspective: Bcl-2 and apoptosis. Stem Cells 20:355–356. CrossRefPubMedGoogle Scholar
  48. 48.
    Ku JH, Seo SY, Kwak C, Kim HH (2012) The role of survivin and Bcl-2 in zinc-induced apoptosis in prostate cancer cells. Urol Oncol 30:562–568. CrossRefPubMedGoogle Scholar
  49. 49.
    Etcheverry SB, Ferrer EG, Naso L, Barrio DA, Lezama L, Rojo T, Williams PAM (2007) Losartan and its interaction with copper(II): biological effects. Bioorg Med Chem 15:6418–6424CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2018

Authors and Affiliations

  • Valeria R. Martínez
    • 1
  • María V. Aguirre
    • 2
  • Juan S. Todaro
    • 2
  • Oscar E. Piro
    • 3
  • Gustavo A. Echeverría
    • 3
  • Luciana G. Naso
    • 1
  • Evelina G. Ferrer
    • 1
  • Patricia A. M. Williams
    • 1
    Email author
  1. 1.Centro de Química Inorgánica (CEQUINOR-CONICET-CICPBA-UNLP)La PlataArgentina
  2. 2.Laboratorio de Investigaciones Bioquímicas, Facultad de Medicina, UNNECorrientesArgentina
  3. 3.Departamento de Física, Facultad de Ciencias ExactasUniversidad Nacional de La Plata y IFLP (CONICET, CCT La Plata)La PlataArgentina

Personalised recommendations