The role of IL-32β in anti-tumor T cell responses

Tumor development and progression are decisively affected by the immune system, especially by T cells. Tumors develop strategies to evade T cell rejection and concomitantly promote suppression of T cell responses. Despite growing knowledge, the underlying mechanisms are not completely understood. Unraveling these phenomena is important for the development of efficient immunotherapies against cancer. Previous work by Dr. Xydia in our group suggested a pro-tumorigenic role of the cytokine Interleukin-32 (IL-32) in breast cancer patients. Single-cell RNA-Sequencing revealed that IL-32 is highly upregulated in intratumoral immunosuppressive regulatory T cells (Treg) and in a subset of circulating tumor antigen (TA)-specific effector T cells (Teff) with tumor-promoting properties. IL-32 has isoform-specific and context-dependent functions in viral infections, autoimmune diseases but also in cancer. However, the role of IL-32 in anti-tumor T cell immunity has been poorly investigated. Here, it was hypothesized that T cells upregulate specific IL-32 isoforms after their activation through tumor cell recognition, leading to the inhibition of Teff and the enhancement of Treg functions, and thereby a direct and indirect regulation of anti-tumor T cell responses. In this thesis, I could unravel that the IL-32β isoform is upregulated by a broad variety of T cells after T cell receptor (TCR) activation and IL-2 exposure. In tumor patients and healthy donors, CD4+ cells, in particular Th1 and Treg, were the major T cell source of IL-32β production. The secretion of IL-32β proteins by T cells was initiated after TCR activation and occurred through unconventional secretion pathways including secretory vesicles. The extracellular presence of IL-32β did not considerably affect the Th1 and Treg phenotype or their function. In contrast, IL-32β exerted an inhibitory impact on the effector function of CD8+ T cells. Specifically, IL-32β impaired the cytotoxic capacity of antigen-specific CD8+ Teff cells by diminishing their secretion of pro-inflammatory molecules, thereby inhibiting T cell-mediated tumor cell eradication. The immunosuppressive role of IL-32β was confirmed through a CRISPR/Cas9-mediated knock-out (KO) of IL-32 in antigen-specific CD8+ Teff cells, which resulted in elevated secretion levels of cytotoxic molecules and improved tumor cell eradication. The superior effector responses of IL-32KO CD8+ Teff cells were associated with higher glycolytic metabolism and enhanced TCR downstream signaling. Thus, IL-32β displays a potential target for future immunotherapeutic approaches to treat cancer patients.