High-dimensional single-cell definition of CLL T cells identifies Galectin-9 as novel immunotherapy target

Failure of cancer immunotherapy is linked to T cell exhaustion. To decipher the underlying mechanisms, we explored the T cell landscape in blood, bone marrow and lymph node samples of patients with chronic lymphocytic leukemia (CLL), and spleen samples of a CLL mouse model. By single-cell RNA-sequencing, mass cytometry (CyTOF), and multiplex image analysis of tissue microarrays, we identified a disease-specific accumulation of distinct regulatory T cell subsets and T cell exhaustion stages and their trajectories in CLL lymph nodes. Integration of T cell receptor sequencing data revealed a clonal expansion of CD8+ precursor exhausted T cells (TPEX), suggesting their CLL reactivity. Interactome analyses identified the TIM3 ligand Galectin-9 as a novel immunoregulatory molecule in CLL. Blocking of Galectin-9 in CLL-bearing mice slowed down disease development and reduced the number of TIM3-expressing T cells. Galectin-9 expression correlated with shorter survival of patients with CLL, renal cell carcinoma or glioma.Statement of significance Our findings for the first time define the T cell landscape in CLL lymph nodes and reshape the current understanding of T cell exhaustion in this malignancy. They further introduce Galectin-9 as novel immune checkpoint with a high potential to overcome resistance to PD1 targeting drugs in CLL and beyond.Competing Interest StatementThe authors declare no competing interests directly related to this work. The authors however disclose unrelated funding and honorariums as follows: D.S. reports funding from GSK and receives honorariums from immuneai and Alpenglow. M.S. reports research funding from Bayer AG.