Tumor hypoxia is a hallmark of head and neck squamous cell carcinoma (HNSCC) and promotes tumor prgression. Tumor-derived small extracellular vesicles (sEVs) are mediators of hypoxia-driven signaling and contribute to immune suppression within the tumor microenvironment. ADAM17, a membrane-associated sheddase regulating inflammatory and immune pathways, is sensitive to membrane lipid composition, particularly phosphatidylserine (PS). We investigated how hypoxia-induced lipidomic remodeling of tumor-derived sEVs affects ADAM17 activity and CD8+ T-cell apoptosis.
HNSCC cells (PCI-30) and normal keratinocytes (HaCaT) were cultured under normoxia (21% O₂) or hypoxia (1% O₂). sEVs were isolated by size-exclusion chromatography and characterized by nanoparticle tracking analysis, transmission electron microscopy, immunoblotting, and high-resolution untargeted lipidomics. Surface PS and ADAM17 expression were assessed by bead-based flow cytometry, and ADAM17 activity was measured using fluorogenic assays. Human CD8+ T cells were incubated with hypoxic or normoxic tumor-derived sEVs. PS-mediated interactions were blocked using Annexin V, and CD8+ T-cell apoptosis was quantified by flow cytometry.
sEVs (125—135 nm) expressed Alix, CD63 and CD9 but not Grp94. Hypoxia selectively enriched PS species with mono-, polyunsaturated and very long acyl chains, reducing surface-exposed PS. Despite decreased PS exposure, hypoxic TEX displayed significantly increased ADAM17 expression and enzymatic activity, resulting in an elevated ADAM17/PS ratio. Functionally, hypoxic TEX induced higher levels of CD8+ T-cell apoptosis compared with normoxic TEX. Blockade of PS with annexin V significanlty reduced ADAM17 activity and attenuated TEX-induced CD8+ T-cell apoptosis.
Hypoxia alters phosphatidylserine organization in tumor-derived sEVs, enhancing ADAM17 activation, promoting PS-dependent CD8+ T-cell apoptosis, and facilitating immune evasion in HNSCC.
n/a
Tumor Immunology: Cellular Responses and Tumor Microevironment (TIME)
Alicja Głuszko, Ahmad Aljohmani, Andrzej Ciechanowicz et al.· Journal of Immunology· 0 citations
Pancreatic ductal adenocarcinoma (PDAC) is an aggressive malignancy with limited therapeutic options, driven in part by its immunosuppressive tumor microenvironment (TME). Tumor-associated macrophages (TAMs) and neutrophils (TANs) contribute to tumor progression and immune evasion. A Disintegrin and Metalloproteinase 8 (ADAM8), a zinc-dependent protease, is strongly upregulated in PDAC and correlates with poor clinical outcomes, suggesting a regulatory role in tumor progression. Wild-type (WT) and Adam8 knockout (A8KO) PDAC cell lines were generated using the CRISPR-Cas9 technique, and PDAC mouse models with or without Adam8 expression were established to investigate the role of ADAM8 in tumor and immune cells. In vitro assays, including Western blotting, qPCR, migration and invasion assays, proliferation assays, ELISA, cytokine and proteome analyses, as well as co-culture experiments with PDAC cells and either macrophages or neutrophils, were employed to assess the effects of ADAM8 on tumor–immune cell crosstalk. In parallel, in vivo WT and A8KO KPC models were generated, genotyped, and monitored to evaluate the impact of ADAM8 on survival, tumor growth, and immune cell recruitment within the PDAC TME. ADAM8 deletion reduced tumor cell proliferation and migration, associated with reduced activation of FAK/Src/STAT3 signaling and altered secretion of cytokines including GM-CSF, M-CSF, ICAM-1, and TNF-α. Co-culture assays demonstrated that ADAM8 enhanced reciprocal signaling between tumor cells and TAMs/TANs, promoting pro-oncogenic activation. Migration assays and in vivo analyses revealed that ADAM8 facilitated recruitment of macrophages and neutrophils in PDAC TME, while Adam8KO tumors exhibited reduced immune infiltration and altered macrophage polarization. Our findings demonstrate that ADAM8 promotes PDAC aggressiveness by enhancing tumor cell proliferation and migration, activating FAK/Src/STAT3 signaling, and driving macrophage and neutrophil recruitment through cytokine regulation. By orchestrating both tumor-intrinsic pathways and tumor–immune interactions, ADAM8 emerges as a key determinant of PDAC progression and a systemic target for therapeutic intervention.
Kimia Zandieh, L. Cook, Kai Zhao et al.· Cellular Oncology· 0 citations
We use cookies to run the site and, with your consent, for analytics and to show ads.
See our Cookie Policy.
