Biochemical Regulation of the Tim-3-Galectin-9 Immunosuppressive Pathway under Normal Conditions and upon Cell Malignant Transformation

Cancer is a group of diseases involving abnormal cell proliferation with a high potential to spread or invade into other parts of the body and is currently considered as one of the leading causes of death worldwide. Cancer cells can be recognised and killed by cytotoxic immune cells of lymphoid lineage. However, malignant cells are capable of suppressing the host's cytotoxic immune responses using a variety of biochemical mechanisms. One of the critical and poorly understood mechanisms of immune evasion is the Tim-3 (T cell immunoglobulin and mucin domain containing protein 3)-galectin-9 immunosuppressive pathway. Galectin-9, a protein highly conserved through evolution, is used to suppress anti-cancer activities of cytotoxic lymphoid cells. It has recently been reported that acute myeloid leukaemia (AML) cells are able to promote galectin-9 and Tim-3 translation and secretion through stimulation induced by fibronectin leucine-rich transmembrane protein 3 (FLRT3), which activates transmembrane receptor latrophilin-1 (LPHN1). LPHN1 mediates activation of Tim-3 and galectin-9 translation and facilitates their exocytosis. However, the molecular mechanisms underlying biochemical regulation of galectin-9 expression and control of the pathway activity remain unknown. Thus, uncovering this crucial biochemical machinery was the aim of this PhD programme.

First of all, our group discovered that the Tim-3-galectin-9 pathway is active in a variety of human solid tumours and is also used to suppress the activities of cytotoxic lymphoid cells. We found that transforming growth factor beta 1 (TGF-β) is crucial for upregulation of galectin-9 expression in human cancer but not in non-malignant human cells. The transcription factors hypoxia-inducible factor 1 (HIF-1) and activator protein 1 (AP-1) were found to upregulate TGF-β expression, leading to activation of the transcription factor Smad3 through autocrine action. Smad3 was found to be responsible for TGF-β-mediated upregulation of galectin-9 expression. As a follow-up, we have discovered that high-mobility group box 1 (HMGB1, a nuclear protein secreted by stressed, damaged or dying cells and thus highly present in the tumour microenvironment) upregulates TGF- β production through the innate immune receptor called Toll-like receptor 4 (TLR4). Thus, HMGB1 can lead to induction of galectin-9 expression in cancer cells directly (if they express functional TLR4) or indirectly through TLR4 expressing myeloid cells present in the tumour microenvironment. Collectively, our results suggest that Tim-3-galectin-9 and TGF-β-dependent signalling pathways are potential targets for the immunotherapy of a large number of human cancers.