TIM-3 blockade enhances ex vivo stimulated allogeneic NK cell therapy for relapsed murine neuroblastoma after hematopoietic cell transplant. J Immunother Cancer 2025 Dec 12;13(12)
Date
12/13/2025Pubmed ID
41386972Pubmed Central ID
PMC12699621DOI
10.1136/jitc-2024-010239Scopus ID
2-s2.0-105024833850 (requires institutional sign-in at Scopus site) 2 CitationsAbstract
BACKGROUND: High-risk neuroblastoma (HR-NBL) is an aggressive tumor of the sympathetic nervous system with high risk of relapse and poor overall survival. Allogeneic hematopoietic cell transplant (allo-HCT) has been used previously in patients with HR-NBL; however, graft-versus-host disease (GVHD) and disease progression have limited clinical application. Ex vivo stimulated allogeneic natural killer (NK) cells represent an approach to enhance the graft-versus-tumor (GVT) effect without exacerbation of GVHD but have not shown efficacy in NBL.
METHODS: Ex vivo stimulated NK cells from C57BL/6NCr (B6) mice were expanded with soluble interleukin-15 (IL-15) and IL-15 receptor alpha (IL-15Rα) alone or with irradiated CD137L/CD54+ aggressive variant of the Neuro-2a murine neuroblastoma cell line (15-4P) at a 1:1 ratio for 10-12 days. Allogeneic NK cells were then analyzed for activation, proliferation, cytokine production, and cytotoxicity against two murine NBL cell lines, Neuro2a and NXS2, in the absence or presence of anti-T-cell immunoglobulin and mucin-domain containing-3 (TIM-3). Lethally irradiated B6AJF1 mice received allo-HCT from B6 donors followed by NBL challenge after 7 days to mimic tumor relapse. Select groups received anti-TIM-3 starting on day 9 for every 4 days with/without infusions of 15-4P B6 NK cells on days 14, 21, and 28. In select experiments, T cell and NK cells were selectively depleted to establish contribution to the GVT effect. All groups were analyzed for tumor growth, GVHD and survival.
RESULTS: Co-culturing NK cells with 15-4P results in 78-fold expansion with increased expression of Kiel-67 (Ki-67) and Natural Killer Group 2, Member D (NKG2D), NKp46, TNF-Related Apoptosis-Inducing Ligand (TRAIL) and TIM-3. 15-4P stimulated allogeneic NK cells showed enhanced cytotoxicity against NBL compared with IL-15 NK cells alone but was limited in part due to high expression of TIM-3 ligands on Neuro-2a compared with NXS2. The addition of TIM-3 blockade further enhanced NK cytotoxicity versus Neuro-2a, with enhanced 15-4P NK cell degranulation, Eomesodermin, TRAIL and Fas Ligand expression observed. In vivo, the combination of 15-4P stimulated allogeneic NK cells and TIM-3 blockade after allo-HCT resulted in prolonged survival against NBL with decreased tumor burden compared with NK cells or anti-TIM-3 alone. Depletion of NK cells, but not T cells, abrogated the GVT effect.
CONCLUSION: Allo-HCT can be a platform for treating NBL using combination ex vivo stimulated allogeneic NK cell therapy with TIM-3 blockade to enhance the GVT effect without inducing GVHD.
Author List
Quamine AE, Dray EL, Mohrdieck NR, King CA, Griggs AA, Kline JM, Cho MM, Rinella SP, Tippins KE, Bates PD, Shi L, Song L, Hess NJ, Kearl TJ, Johnson BD, Capitini CMAuthors
Bryon D. Johnson PhD Adjunct Professor in the Medicine department at Medical College of WisconsinTyce J. Kearl PhD, MD Assistant Professor in the Medicine department at Medical College of Wisconsin
MESH terms used to index this publication - Major topics in bold
AnimalsCell Line, Tumor
Graft vs Tumor Effect
Hematopoietic Stem Cell Transplantation
Hepatitis A Virus Cellular Receptor 2
Humans
Killer Cells, Natural
Mice
Mice, Inbred C57BL
Neuroblastoma
Transplantation, Homologous
