The local accumulation of Tregs, highly expressing the costimulatory receptor OX40 and displaying a stable and activated phenotype, contributes to maintain immunosuppression in hepatic cirrhosis and hepatocellular carcinoma microenvironments. In these contexts, our preliminary data highlight a connection between OX40 signal and specific metabolic routes, namely glycolysis, cholesterol/lipid biosynthesis, and iron/transferrin capture.
We hypothesize that OX40 orchestrates a metabolic program of Treg expansion and suppression, based on the effects of extracellular iron sequestration from effector cells. Our main aim will be to demonstrate that iron scavenging mediates OX40+ Treg suppressive function thus promoting tumor progression.
The impact of iron/transferrin uptake on Treg proliferation and suppression will be assessed, in relation to OX40 expression and engagement, by multiple techniques (multicolor flow cytometry, microscopy, biochemical and functional assays), in three settings: in vitro, using stimulated human Tregs; ex vivo, analyzing hepatic Tregs from hepatocellular carcinoma patients with variable iron homeostasis perturbations; in vivo, monitoring Tregs and tumor growth in mouse models of high fat choline-deficient diet-induced, or transplanted, hepatocellular carcinoma, combined with the Treg-restricted genetic deletion of transferrin receptor.
We expect to delineate a previously unrecognized mechanism of Treg suppression, based on iron sequestration from the microenvironment: this event may become especially relevant in tumor-infiltrating OX40+ Tregs and may thus represent new targets for more focused immunotherapies.
Therapies based on OX40 agonists are currently under development for cancer, even though it is unclear whether Treg depletion, rather than engagement, contributes to a potential benefit. Many efforts have been made to identify Treg-selective targets, and it is increasingly clear that immunotherapy can be achieved through T cell metabolism modulation. Our proposal points at demonstrating that iron metabolism plays non-redundant roles in Treg-mediated immune suppression and thus can be exploited to improve anti-cancer response, besides blocking tumor cell growth. Many pieces of evidence link iron overload with cancer risk and progression, and interrupting this pathway (by iron-chelating therapies, or transferrin-receptor targeted drug delivery to cancer cells) is a promising target for cancer therapy [8]. Our results will not only provide novel pieces of knowledge about the interplay between immune regulation and metabolism, but also lay the foundation to the development of innovative immunotherapies for cancer, based on the concomitant targeting of tumor cells and Tregs sharing key metabolic pathways. Indeed, tumor-infiltrating Tregs may be "poisoned" by exploiting iron endocytic pathway as a Trojan horse.
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