title: Laccase purified fromCerrena unicolorexerts antitumor activity against leukemic cells.
abstract: Chronic lymphocytic leukemia (CLL) is the most commonly observed adult hematological malignancy in Western countries. Despite the fact that recent improvements in CLL treatment have led to an increased percentage of complete remissions, CLL remains an incurable disease. Cerrena unicolor is a novel fungal source of highly active extracellular laccase (ex-LAC) that is currently used in industry. However, to the best of our knowledge, no reports regarding its anti-leukemic activity have been published thus far. In the present study, it was hypothesized that C. unicolor ex-LAC may possess cytotoxic activity against leukemic cell lines and CLL primary cells. C. unicolor ex-LAC was separated using anion exchange chromatography on diethylaminoethyl cellulose-Sepharose and Sephadex G-50 columns. The cytotoxic effects of ex-LAC upon 24- and 48-h treatment on HL-60, Jurkat, RPMI 8226 and K562 cell lines, as well as CLL primary cells of nine patients with CLL, were evaluated using 2,3-bis-(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide (XTT) assay. Annexin V/propidium iodide staining of Jurkat cells treated with ex-LAC was used to investigate apoptosis via flow cytometry. Ex-LAC induced changes in Jurkat and RPMI 8226 cells, as visualized by fluorescence and scanning electron microscopy (SEM). The XTT assay revealed high cytotoxic rates following treatment with various concentrations of ex-LAC on all the cell lines and CLL primary cells analyzed, with a half maximal inhibitory concentration ranging from 0.4 to 1.1 µg/ml. Fluorescence microscopy and SEM observations additionally revealed apoptotic changes in Jurkat and RPMI 8226 cells treated with ex-LAC, compared with control cells. These results were in agreement with the apoptosis analysis of Jurkat cells on flow cytometry. In conclusion, C. unicolor ex-LAC was able to significantly induce cell apoptosis, and may represent a novel therapeutic agent for the treatment of various hematological neoplasms.
authors.affiliation: Department of Biochemistry, Maria Curie-Skłodowska University, Lublin 20-033, Poland.
authors.forename: Anna
authors.initials: A
authors.lastname: Matuszewska
authors.affiliation: Department of Experimental Hemato-Oncology, Medical University of Lublin, Lublin 20-093, Poland; Postgraduate School of Molecular Medicine, Medical University of Warsaw, Warsaw 02-091, Poland.
authors.forename: Marta
authors.initials: M
authors.lastname: Karp
authors.affiliation: Department of Biochemistry, Maria Curie-Skłodowska University, Lublin 20-033, Poland.
authors.forename: Magdalena
authors.initials: M
authors.lastname: Jaszek
authors.affiliation: Department of Biochemistry, Maria Curie-Skłodowska University, Lublin 20-033, Poland.
authors.forename: Grzegorz
authors.initials: G
authors.lastname: Janusz
authors.affiliation: Department of Biochemistry, Maria Curie-Skłodowska University, Lublin 20-033, Poland.
authors.forename: Monika
authors.initials: M
authors.lastname: Osińska-Jaroszuk
authors.affiliation: Department of Biochemistry, Maria Curie-Skłodowska University, Lublin 20-033, Poland.
authors.forename: Justyna
authors.initials: J
authors.lastname: Sulej
authors.affiliation: Department of Biochemistry, Maria Curie-Skłodowska University, Lublin 20-033, Poland.
authors.forename: Dawid
authors.initials: D
authors.lastname: Stefaniuk
authors.affiliation: Department of Hemato-Oncology and Bone Marrow Transplantation, Medical University of Lublin, Lublin 20-081, Poland.
authors.forename: Waldemar
authors.initials: W
authors.lastname: Tomczak
authors.affiliation: Department of Experimental Hemato-Oncology, Medical University of Lublin, Lublin 20-093, Poland.
authors.forename: Krzysztof
authors.initials: K
authors.lastname: Giannopoulos
keywords: Cerrena unicolor; HL-60 human acute promyelocytic leukemia cells; Jurkat human T cell leukemia cells; K562 human chronic myeloid leukemia in blast crisis cells; RPMI 8226 human multiple myeloma cells; chronic lymphocytic leukemia; laccase