en Immunology Immunology پژوهشي Original Article <div style="text-align: justify;"><span style="font-family:Times New Roman;"><span style="line-height:2;"><span style="font-size:14px;"><strong>Background:</strong> In adoptive cell therapy, cytokine-induced killer (CIK) cells have been used as a promising modality due to their potent antitumor activity and MHC-independent function. IL-15 has been proposed as a promising alternative to IL-2 because it enhances survival, maintains a memory-like phenotype, and improves cytotoxic function; however, its utility in optimizing CIK cells for the treatment of hepatocellular carcinoma (HCC) remains to be investigated.<br> <strong>Methods:</strong> CIK cells were generated from human peripheral blood mononuclear cells (hPBMCs) using a 12-day step-wise expansion protocol including treatment with IFN-&gamma;, anti-CD3 antibody, IL-2, and IL-15 in culture medium containing human platelet lysate. The phenotypic characteristics of the cells were examined by flow cytometry, and the expression of CD3⁺, CD8⁺, and CD56⁺ was assessed. The cytotoxic activity of CIK cells against HepG2 cells was evaluated by flow cytometry using a CFSE/PI assay at different effector-to-target ratios (E: T), and ELISA measured the secretion of IFN-&gamma; and TNF-&alpha;.<br> <strong>Results: </strong>The expanded CIK cells exhibited a characteristic CD3⁺CD8⁺ and CD3⁺CD56⁺ phenotype. These cells demonstrated significant cytotoxic activity against HepG2 cells across different E: T ratio and showed enhanced production of IFN-&gamma; and TNF-&alpha;, indicating robust antitumor activity.<br> <strong>Conclusion: </strong>The short-term IL-15-enriched expansion protocol generated stable CIK cells with an NK-like phenotype and significant cytotoxic activity against HCC cells. Increased secretion of inflammatory cytokines also indicated effective antitumor activation of these cells. &nbsp;These findings support further development and preclinical evaluation of IL-15-CIK-based cell therapy for the treatment of HCC.</span></span></span></div> Immunotherapy, Adoptive cell therapy, Cytokine-induced killer cells, Hepatocellular carcinoma 88 95 http://ijbc.ir/browse.php?a_code=A-10-2459-1&slc_lang=en&sid=1 Neda Minaei 100319475328460013501 100319475328460013501 No Department of Applied Cell Sciences, Faculty of Basic Sciences and Advanced Medical Technologies, Royan Institute, Academic Center for Education, Culture and Research (ACECR), Tehran, Iran. Behnoosh Tayebi 100319475328460013502 100319475328460013502 No Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran. Shukoofeh Torabi shekoofe_torabi@yahoo.com 100319475328460013503 100319475328460013503 No Department of Regenerative Medicine, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran. Marzieh Ebrahimi marzieh.ebrahimi@gmail.com 100319475328460013504 100319475328460013504 No Department of Regenerative Medicine, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran. Abbas Piryaei abbaspiryaei@gmail.com 100319475328460013505 100319475328460013505 No Department of Biology and Anatomical Sciences, School of Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran. Massoud Vosough masvos@yahoo.com 100319475328460013506 100319475328460013506 Yes Department of Applied Cell Sciences, Faculty of Basic Sciences and Advanced Medical Technologies, Royan Institute, Academic Center for Education, Culture and Research (ACECR), Tehran, Iran & Department of Regenerative Medicine, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran.