The Mitochondria-Independent Cytotoxic Effect of Leflunomide on RPMI-8226 Multiple Myeloma Cell Line

• Published in: Molecules (Basel, Switzerland) Switzerland), 2021-09, Vol.26 (18), p.5653
• Main Authors: Adamczuk, Grzegorz, Humeniuk, Ewelina, Iwan, Magdalena, Natorska-Chomicka, Dorota, Adamczuk, Kamila, Korga-Plewko, Agnieszka
• Format: Article
• Language: English
• Subjects: Anti-inflammatory agents; Antineoplastic Agents - pharmacology; Apoptosis; Apoptosis - drug effects; Arthritis; Biosynthesis; Cancer; Cell cycle; Cell Cycle - drug effects; Cell division; Cell Line, Tumor; Cell Proliferation - drug effects; Cell Survival - drug effects; Clinical trials; Crotonates - pharmacology; Cytotoxicity; dehydrogenase dihydroorotate; Dihydroorotate Dehydrogenase; Disease; Enzymes; Gene expression; High-performance liquid chromatography; Humans; Hydroxybutyrates - pharmacology; Inflammation; Kinases; Leflunomide; Leflunomide - pharmacology; Liquid chromatography; Localization; Membrane potential; Membrane Potential, Mitochondrial - drug effects; Membrane proteins; Metabolites; Mitochondria; Mitochondria - drug effects; Mitochondria - metabolism; Multiple myeloma; Multiple Myeloma - drug therapy; Multiple Myeloma - metabolism; Multiple Myeloma - pathology; Nitriles - pharmacology; Oxidative stress; Oxidative Stress - drug effects; Oxidoreductases Acting on CH-CH Group Donors - antagonists & inhibitors; Oxidoreductases Acting on CH-CH Group Donors - metabolism; protein tyrosine kinases; Protein-tyrosine kinase; teriflunomide; Thiols; Toluidines - pharmacology; Toxicity; Tyrosine; Uridine
• ISSN: 1420-3049; 1420-3049
• DOI: 10.3390/molecules26185653

Leflunomide, an anti-inflammatory agent, has been shown to be effective in multiple myeloma (MM) treatment; however, the mechanism of this phenomenon has not been fully elucidated. The aim of the study was to assess the role of mitochondria and dihydroorotate dehydrogenase (DHODH) inhibition in the cytotoxicity of leflunomide in relation to the MM cell line RPMI 8226. The cytotoxic effect of teriflunomide-an active metabolite of leflunomide-was determined using MTT assay, apoptosis detection, and cell cycle analysis. To evaluate DHODH-dependent toxicity, the cultures treated with teriflunomide were supplemented with uridine. Additionally, the level of cellular thiols as oxidative stress symptom was measured as well as mitochondrial membrane potential and protein tyrosine kinases (PTK) activity. The localization of the compound in cell compartments was examined using HPLC method. Teriflunomide cytotoxicity was not abolished in uridine presence. Observed apoptosis occurred in a mitochondria-independent manner, there was also no decrease in cellular thiols level. Teriflunomide arrested cell cycle in the G2/M phase which is not typical for DHODH deficiency. PTK activity was decreased only at the highest drug concentration. Interestingly, teriflunomide was not detected in the mitochondria. The aforementioned results indicate DHODH- and mitochondria-independent mechanism of leflunomide toxicity against RPMI 8226 cell line.