Background Cancer is a condition in which abnormal cells in a particular part of the body can proliferate in an unchecked way and affect its surrounding healthy tissues. Inflammatory pathways are some of the most important factors involved in cancer and tumor formation (1). These pathways are activated by various agents, which are responsible for 95% of cancers, including smoking, stress, food, obesity, alcohol, infectious agents, environmental triggers, and exposure to radiation (2). Inflammation is an immune response to cellular or tissue injury or infection caused by pathogens. Clinically, inflammation is determined by features such as redness, warmth, swelling, and pain (3). The mechanism of inflammatory pathways can be divided into the arachidonic acid (AA)-dependent pathway and the AA-independent pathway (4). Cyclooxygenase (COX), lipoxygenase (LOX), and phospholipase A2 (PLA2) are considered as AA-dependent pathways. On the other hand, nitric oxide synthase (NOS) and NF-кB (i.e., nuclear factor kappa-light-chain-enhancer of activated B cells) are AA-independent pathways (5). The products of both pathways play an important role in the inflammation process (6). In the past two decades, several molecules involved in the inflammation have been identified, including tumor necrosis factor (TNF), interleukin 1 (IL- 1), interleukin 6 (IL-6), COX -2, matrix metalloproteinase (MMP) and vascular endothelial growth factor (VEGF). The common feature of all these molecules is the fact that they are regulated by the transcription factor NF-кB (7). The NF-кB which is known as a transcription activator is a heterodimer protein composed of two subunits called p50 and RelA (p65). In the absence of stimulus, the NF-кB is inhibited in the cytoplasm by an inhibitor agent called IкBα; but in the presence of the stimulus, the signaling pathway is activated. The protein IкB creates a heterodimer nuclear translocation signal that prevents the transferring of NF-кB into the nucleus. In these conditions, the NF-кB is activated and enters into the nucleus which will affect the expression of target genes (8). The NF- кB connects cells that are involved in inflammation and cancer (9). The expression of NF-кB is increased in the majority of cancer tissues and is known as an inducer of prostate cancer progression and tumor metastasis in lymph nodes (10). Moreover, lung cancer progression (11), breast cancer (12) and its response to chemotherapy (13) are other NF-кB activities. Due to the large number of cellular processes affected by NF-кB, modulators of this pathway have received a considerable attention. For Avicenna Journal of Medical Biochemistry © 2021 The Author(s); Published by Hamadan University of Medical Sciences. This is an open-access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. 2021 December;9(2):48-53 doi:10.34172/ajmb.2021.09 Inhibition of NF-кB Expression in LPS-Induced RAW264.7 Macrophage Cells by a Thiazolidinone Derivative (TZD- OCH 2 CH 3 ) Farahnaz Hasanzadeh 1 , Hossein Ghafouri 2,3* ID , Salman Ahmadi 2 , Sevda Zarei 2 ID , Mahmoud Reza Aghamaali 2 , Asadollah Mohammadi 4 1 Department of Biology, Faculty of Basic Sciences, University of Guilan, University Campus 2, Rasht, Iran 2 Department of Biology, Faculty of Basic Sciences, University of Guilan, Rasht, Iran 3 Department of Marine Sciences, The Caspian Sea Basin Research Center, University of Guilan, Rasht, Iran 4 Department of Chemistry, Faculty of Sciences, University of Guilan, Rasht, Iran http://ajmb.umsha.ac.ir Research Article Abstract To date, various derivatives of thiazolidinone in a variety of cell lines have been investigated. The present study aimed to evaluate the toxicity and inhibitory effects of a thiazolidinone derivative called 5-(2,4-bis- 4-ethoxy-phenyl azo)-3-hydroxy-benzylidine)-2,4-thiazolidinone (TZD-OCH 2 CH 3 ) on the expression of NF-кB in LPS-induced RAW264.7 macrophage cell lines. Different concentrations of the MTT assay (0-120 μg/mL) were performed to estimate the biological rate of the cells. The half-maximal inhibitory concentration (IC 50 ) of TZD-OCH 2 CH 3 -treated RAW264.7 cells was found to be 115 μg/mL. To determine the inhibitory effect of the synthesized compound on the expression changes of NF-кB, the RAW264.7 cells were initially induced with LPS and then treated by 15, 30 and 60 µg/mL of TZD-OCH 2 CH 3 . Real- time PCR results confirmed a strong inhibitory effect of TZD-OCH 2 CH 3 on the expression of NF-кB in LPS-induced RAW264.7 cells (IC 50 = 48 μg/mL). Overall, these findings suggested that the derivative TZD- OCH 2 CH 3 had a significant anti-inflammatory effect. Keywords: Thiazolidinone, NF-кB, RAW264.7 cells, LPS- induced, Inflammatory pathways Received: 13 July 2021 Revised: 22 August 2021 Accepted: 22 August 2021 ePublished: 29 December 2021 *Corresponding author: Hossein Ghafouri, Associate Professor in Biochemistry, Department of Biology, Faculty of Basic Sciences, University of Guilan, Rasht, Iran. Tel: +981333333647; Email: h.ghafoori@guilan.ac.ir