Targeting immune checkpoint co-inhibitory pathways is a promising novel therapeutic approach for several types of cancer. Therefore, we aimed to determine which co-inhibitory pathways can be targeted to enhance the functionality of intra-tumoral T cells in hepatocellular carcinoma (HCC).
We measured the expression of co-inhibitory receptors and ligands on leukocytes isolated from paired tumors, tumor-free liver tissues (TFL), and peripheral blood of HCC patients, and studied the effects of blocking co-inhibitory pathways on functional responses of CD4+ and CD8+ tumor-infiltrating lymphocytes (TIL) in ex vivo assays.
Expression of PD-1, TIM-3, LAG-3 and CTLA-4 on CD8+ T cells and/or CD4+Foxp3- T helper cells was significantly higher in tumors than in TFL or blood. Dendritic cells, monocytes and B cells in tumors expressed their ligands. Using MHC class I multimers loaded with immunogenic Glypican-3 or MAGE-C2 peptides, we found that PD-1, TIM-3 and LAG-3 expressions were selectively increased on tumor-associated antigen-specific CD8+ TIL. Compared to the TIL without co-inhibitory receptor expression, CD8+ and CD4+ TIL expressing those co-inhibitory receptors displayed a more activated status but similar or decreased granzyme B expression and effector cytokine production. Blocking PD-L1, TIM-3 or LAG-3 with neutralizing antibodies enhanced proliferative and cytokine responses of CD8+ and CD4+ TIL to polyclonal CD3/D28-stimulation as well as to Glypican-3 or MAGE-C2 presented by mRNA-transfected autologous antigen-presenting cells. Importantly, combining PD-L1 blockade with TIM-3, LAG-3 or CTLA-4 blockade further enhanced ex vivo TIL responses.
Conclusions: PD-1, TIM-3 and LAG-3 may be promising immunotherapeutic targets for the most prevalent primary liver cancer.