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Complement factor H protects tumor cell-derived exosomes from complement-dependent lysis and phagocytosis.

ABSTRACT: Exosomes are a class of extracellular vesicles (EVs) that are mediators of normal intercellular communication, but exosomes are also used by tumor cells to promote oncogenesis and metastasis. Complement factor H (CFH) protects host cells from attack and destruction by the alternative pathway of complement-dependent cytotoxicity (CDC). Here we show that CFH can protect exosomes from complement-mediated lysis and phagocytosis. CFH was found to be associated with EVs from a variety of tumor cell lines as well as EVs isolated from the plasma of patients with metastatic non-small cell lung cancer. Higher levels of CFH-containing EVs correlated with higher metastatic potential of cell lines. GT103, a previously described antibody to CFH that preferentially causes CDC of tumor cells, was used to probe the susceptibility of tumor cell-derived exosomes to destruction. Exosomes were purified from EVs using CD63 beads. Incubation of GT103 with tumor cell-derived exosomes triggered exosome lysis primarily by the classical complement pathway as well as antibody-dependent exosome phagocytosis by macrophages. These results imply that GT103-mediated exosome destruction can be triggered by antibody Fc-C1q interaction (in the case of lysis), and antibody-Fc receptor interactions (in the case of phagocytosis). Thus, this work demonstrates CFH is expressed on tumor cell derived exosomes, can protect them from complement lysis and phagocytosis, and that an anti-CFH antibody can be used to target tumor-derived exosomes for exosome destruction via innate immune mechanisms. These findings suggest that a therapeutic CFH antibody has the potential to inhibit tumor progression and reduce metastasis promoted by exosomes.

SUBMITTER: Bushey RT

PROVIDER: S-EPMC8208531 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:The suppressive nature of immune cells in the tumor microenvironment plays a major role in regulating anti-tumor immune responses. Our previous work demonstrated that a soluble factor from tumor cells is able to induce a suppressor phenotype (SP) in human CD8+ T cells typified by loss of CD27/CD28 expression and acquisition of a potent suppressor function. The present study hypothesized that the soluble mechanism that is inducing the SP in CD8+ T cells are tumor-derived exosomes (TDEs).Membrane vesicles and TDEs from multiple head and neck cancer cell line's conditioned growth media were isolated by ultracentrifugation and precipitation, respectively. Human purified CD3+CD8+ T cells were assessed for their induction of the T cell SP by flow cytometry identifying loss of CD27/CD28 expression and in vitro suppression assays. Furthermore, the T cell SP was characterized for the attenuation of IFN-γ production. To delineate exosomal proteins contributing to T cell SP, mass spectrometry was used to identify unique proteins that were present in TDEs. CRISPR/Cas9 knockout constructs were used to examine the role of one of these proteins, galectin-1. To assess the role of exosomal RNA, RNA purified from TDEs was nucleofected into CD8+ T cells followed by suppression analysis.Using fractionated conditioned growth media, factors >200 kDa induced CD8+ T cell SP, which was determined to be an exosome by mass spectrometry analysis. Multiple head and neck cancer-derived cell lines were found to secrete T cell SP-inducing exosomes. Mass spectrometry analysis revealed that an immunoregulatory protein, galectin-1 (Gal-1), was expressed in those exosomes, but not in TDEs unable to induce T cell SP. Galectin-1 knockout cells were found to be less able to induce T cell SP. Furthermore, RNA purified from the T cell SP-inducing exosomes were found to partially induce the SP when transfected into normal CD8+ T cells.For the first-time, TDEs have been identified to induce a SP in CD8+ T cells and their mode of action may be synergistic effects from exosomal proteins and RNA. One protein in particular, galectin-1, appears to play a significant role in inducing T cell SP. Therefore, tumor-derived immunosuppressive exosomes are a potential therapeutic target to prevent T cell dysfunction and enhance anti-tumor immune responses.

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Complement factor H protects tumor cell-derived exosomes from complement-dependent lysis and phagocytosis.

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