Analysis of metalothionein in patients with malignant liver tumors

Czech version

Authors: L. Melich ¹; J. Werle ¹; M. Fořtová ¹; L. Vysloužilová ²; K. Burešová ¹; K. Kotaška ¹; E. Klapková ¹; O. Štěpánková ²; J. Čepová ¹; R. Průša ¹; R. Kizek ¹
Authors‘ workplace: Ústav lékařské chemie a klinické bio chemie 2. LF UK a FN Motol, Praha ¹; Český institut informatiky, robotiky a kybernetiky, ČVUT v Praze, Praha ²
Published in: Klin Onkol 2024; 38(5): 345-353
Category: Original Articles
doi: https://doi.org/10.48095/ccko2024345

Overview

Background: Malignant liver tumors are highly aggressive with a poor prognosis. Metalothionein (MT) is a low-molecular intracellular protein, whose primary function is to regulate the homeostasis of heavy metals in many organisms. There are only few studies focusing on the molecular mechanisms of MT expression. Recent studies show its significant relations to carcinogenesis, spontaneous mutagenesis and efficiency of antitumor medicine. In previous studies, the increase of MT levels in cancer patients was proven. The aim of this work is to study MT as well as to increase the efficiency of malignant liver tumor diagnosis. Methods: In our pilot study (2022–2023) we observed a group of 15 patients with hepatocellular carcinoma (diagnosis C220) and a group of 15 patients with hepatoblastoma (diagnosis C222). The control group included 20 healthy probands. We developed our own modified method for the analysis. Blood serum samples of the probands were denaturated (99 ˚C, 20 min). MT was determined by an electrochemical method. Obtained data were stored and processed in the laboratory information system QINSLAB. Results: In denaturated blood serum samples, we obtained voltametric curves of MT. We determined concentrations of MT by evaluating the area under the curve (AUC). To differentiate normal and abnormal concentrations of MT, blood samples of healthy probands were used (N = 20), with the average MT levels of 2.0 ± 1.3 µg/L and median 1.9 µg/L. In patients diagnosed with HCC, the average MT levels were 9.1 ± 6.5 µg/L and median 9.0 µg/L. The receiver operating characteristic (ROC) analysis showed AUC 0.864 (95% CI 0.736–0.992), sensitivity 0.74 and specificity 0.75. In patients diagnosed with hepatoblastoma, the average MT concentrations measured were 11.5 ± 7.5 µg/L and the median was 10.9 µg/L. The ROC analysis displayed AUC 0.868 (95% CI 0.751–0.993), sensitivity 0.84 and specificity 0.86. The correlation analysis showed correlation between MT and carcinoembryonic antigen (CEA) (r = 0.99), uric acid (r = −0.86) and potassium ions (r = −0.94). Conclusion: In this pilot study, we observed the association of MT levels in healthy probands and malignant liver tumor patients. Many previous studies show that MT concentrations are increasing as the illness progresses. We assume that this increase is connected to the high metabolic activity of cancer cells. This study will continue with collecting a larger number of samples.

Keywords:

nanomedicine – cancer – targeted therapy – malignant tumor – Brdička’s reaction

Sources

1. Cossiga V, Guarino M, Capasso M et al. Relevance of bile acids in cholangiocarcinoma pathogenesis: critical revision and future directions. Cells 2023; 12 (12): 1576. doi: 10.3390/cells12121576.

2. Ilyas SI, Affo S, Goyal L et al. Cholangiocarcinoma – novel biological insights and therapeutic strategies. Nat Rev Clin Oncol 2023; 20 (7): 470–486. doi: 10.1038/s41571-023-00770-1.

3. Calvisi DF, Boulter L, Vaquero J et al. Criteria for preclinical models of cholangiocarcinoma: scientific and medical relevance. Nat Rev Gastroenterol Hepatol 2023; 20 (7): 462–480. doi: 10.1038/s41575-022-00739-y.

4. Banales JM, Marin JJG, Lamarca A et al. Cholangiocarcinoma 2020: the next horizon in mechanisms and management. Nat Rev Gastroenterol Hepatol 2020; 17 (9): 557–588. doi: 10.1038/s41575-020-0310-z.

5. Befeler AS, Di Bisceglie AM. Hepatocellular carcinoma: diagnosis and treatment. Gastroenterology 2002; 122 (6): 1609–1619. doi: 10.1053/gast.2002.33411.

6. Wei LY, Zhai YZ, Feng GH. New advances in the use of serum prealbumin as an index of liver function. World Chin J Digest 2013; 21 (15): 1387–1393. doi: 10.1017/S0007114518002854.

7. Zavadil J, Juráček J, Čechová B et al. Dynamic changes in circulating microRNA levels in liver cancer patients undergoing thermal ablation and transarterial chemoembolization. Klin Onkol 2019; 32 (Suppl 1): S164–S166.

8. Zelená J, Potěšil D, Vacek J et al. Metallothionein as a prognostic marker of tumor disease. Klin Onkol 2004; 17 (6): 190–195.

9. Fabrik I, Adam V, Křížková S et al. Level of heat-stable thiols in patients with a malignant tumor. Klin Onkol 2007; 20 (6): 384–389.

10. Horakova Z, Starek I. Serum metallothionein – a potential oncomarker? Bratisl Lek Listy 2021; 122 (8): 577–581. doi: 10.4149/BLL_2021_092.

11. Miaou E, Tissot FLH. Copper isotope ratios in serum do not track cancerous tumor evolution, but organ failure. Metallomics 2023; 15 (11): mfad060. doi: 10.1093/mtomcs/mfad060.

12. Abelev G. Alpha-fetoprotein in ontogenesis and its association with malignant tumors. Adv Cancer Res 1971; 14: 295–358. doi: 10.1016/s0065-230x (08) 60523-0.

13. Kithier K, Houstek J, Masopust J et al. Occurrence of a specific foetal protein in a primary liver carcinoma. Nature 1966; 212 (5060): 414. doi: 10.1038/212414a0.

14. Zizkovsky V, Masopust J, Kordac V et al. Alpha1-fetoprotein in carcinoid. New England Journal of Medicine 1972; 287 (21): 1102–1103.

15. Masopust J, Kithier K, Radl J et al. Occurrence of fetoprotein in patients with neoplasms and non-neoplastics diseases. Int J Cancer 1968; 3 (3): 364–373. doi: 10.1002/ijc.2910030306.

16. Fabrik I, Krizkova S, Huska D et al. Employment of electrochemical techniques for metallothionein determination in tumor cell lines and patients with a tumor disease. Electroanalysis 2008; 20 (14): 1521–1532.

17. Zhang XS, Ma YX, Liu KJ et al. Risk prediction for dermatomyositis-associated hepatocellular carcinoma. [online]. Available from: https: //bmcbioinformatics.biomedcentral.com/articles/10.1186/s12859-023-05353-6BMC.

18. Vij M, Menon J, Subbiah K et al. Pathologic and immunophenotypic characterization of syncytial giant cell variant of pediatric hepatocellular carcinoma. A distinct subtype. Fetal Ped Pathol 2023 Jul; 42 (4): 709–718. doi: 10.1080/15513815.2023.2201318.

19. Nishikawa H, Asai A, Fukunishi S. The significance of zinc in patients with chronic liver disease. Nutrients 2022; 14 (22): 4855. doi: 10.3390/nu14224855.

20. Petrlová J, Blaštík O, Průša R et al. Determination of metallothionein content in patients with breast cancer, colon cancer, and malignant melanoma. Klin Onkol 2006; 19 (2): 138–142.

21. Melich L, Fortova M, Hosnedlova B et al. Studium hladiny metalothionienu v séru pacientů se zhoubným nádorem. Chem Listy 2023; 117 (9): 573–580.

22. Wang XL, Schnoor M, Yin LM. Metallothionein-2: an emerging target in inflammatory diseases and cancers. Pharmacol Therap 2023; 244: 108374.

23. Petrlova J, Potesil D, Mikelova R et al. Attomole voltammetric determination of metallothionein. Electrochim Acta 2006; 51 (24): 5112–5119.

24. Skotak V, Sehnal K, Banas D et al. Využití Brdičkovy reakce pro detekci metalothioneinu v moči. Prakt Lék 2023; 103 (6): 299–303.

25. Hubaux A, Vos G. Decision and detection limits for linerar calibration curves. Anal Chem 1970; 42 (8): 849–851.

26. Melich L, Fortova M, Vyslouzilova L et al. Analýza metalothioneinu u pacientů se zhoubnými nádory jater. Klin Onkol 2023; 36 (Suppl 1): S74–S74.

27. Brdicka R. Polarographic investigation in serological cancer diagnosis. Nature 1937; 139: 1020–1021.

28. Brdicka R. Application of the polarographic effect of proteins in cancer diagnosis. Nature 1937; 139: 330–330.

29. Dabrio M, Rodríguez AR, Bordin G et al. Recent developments in quantification methods for metallothionein. J Inorg Biochem 2002; 88 (2): 123–134. doi: 10.1016/s0162-0134 (01) 00374-9.

30. Babula P, Masarik M, Adam V et al. Mammalian metallothioneins: properties and functions. Metallomics 2012; 4 (8): 739–750. doi: 10.1039/c2mt20081c.

31. Eckschlager T, Adam V, Hrabeta J et al. Metallothioneins and cancer. Current Prot Pept Sci 2009; 10 (4): 360–375. doi: 10.2174/138920309788922243.

32. Krizkova S, Fabrik I, Adam V et al. Metallothionein – a promising tool for cancer diagnostics. Bratisl Lek Listy 2009; 110 (2): 93–97.

33. Adam V, Fabrik I, Eckschlager T et al. Vertebrate metallothioneins as target molecules for analytical techniques. TrAC Trends in Analytical Chemistry 2010; 29 (5): 409–418.

34. Vyslouzilova L, Adam V, Szaboova A et al. Brdicka curve – a new source of biomarkers. IEEE International Conference on Bioinformatics and Biomedicine Workshops (BIBMW); 2011 Nov 12–15; Atlanta, GA.

35. Vyslouzilova L, Krizkova S, Anyz J et al. Use of brightness wavelet transformation for automated analysis of serum metallothioneins- and zinc-containing proteins by Western blots to subclassify childhood solid tumours. Electrophoresis 2013; 34 (11): 1637–1648. doi: 10.1002/elps.201200561.

36. Tamai Y, Iwasa M, Eguchi A et al. Serum copper, zinc and metallothionein serve as potential biomarkers for hepatocellular carcinoma. Plos One 2020; 15 (8): e0237370. doi: 10.1371/journal.pone.0237370.

37. Kruseova J, Hynek D, Adam V et al. Serum metallothioneins in childhood tumours – a potential prognostic marker. Int J Mol Sci 2013; 14 (6): 12170–12185. doi: 10.3390/ijms140612170.

38. Krizkova S, Masarik M, Majzlik P et al. Serum metallothionein in newly diagnosed patients with childhood solid tumours. Acta Biochim Pol 2010; 57 (4): 561–566.

39. Tomášek J, Prášek J, Kiss I et al. Liver function assessment in oncology practice. Klin Onkol 2012; 25 (6): 427–433.

40. Sawyers CL. The cancer biomarker problem. Nature 2008; 452 (7187): 548–552. doi: 10.1038/nature06913.

41. Berndt N, Bulik S, Wallach I et al. Hepatokin1 is a biochemistry-based model of liver metabolism for applications in medicine and pharmacology. Nat Commun 201; 9 (1): 2386. doi: 10.1038/s41467-018-04720-9.

42. Sharon A. Antioxidants in liver disease: a focus on thiol supplementation. [online]. Available from: https: //www.vin.com/apputil/content/defaultadv1.aspx?pId=11223&meta=generic&catId=31461&id=3859034&ind=148&objTypeID=17.