Background:
HNSCCs with high expression levels of PD-L1 have especially poor outcome. However, many patients with PD-L1+ HNSCC don't benefit from checkpoint inhibitor therapy. Tumor derived exosomes carry numerous immunosuppressive molecules and deliver them concentrated to recipient immune cells. We have demonstrated that elevated levels of circulating immunosuppressive exosomes in HNSCC patients play a key role in immune suppression and disease progression. Here, we show that surface PD-L1 on exosomes is responsible for these effects.
Material and Methods:
Exosomes were isolated from plasma of 40 HNSCC patients by mini size exclusion chromatography, captured on beads using anti-CD63 Abs, stained for PD-1 and PD-L1 and analyzed by flow cytometry. Exosomes, which were either PD-L1 high or PD-L1 low were incubated with activated human CD8 T-cells ± PD-1 inhibitor, and CD69 expression levels on T-cells were measured. Patients' plasma was also tested for soluble PD-L1.
Results:
The PD-L1 surface expression on exosomes correlated with patient's disease activity and UICC stage. In contrast, plasma PD-L1 levels and exosomal PD-1 levels were not informative. T-cell activation was inhibited by co-incubation with PD-L1high but not by PD-L1low exosomes. This inhibition could be reversed by adding a PD-1 inhibitor to T-cells prior to their co-incubation with exosomes.
Conclusions:
We show that PD-L1 levels on exosomes, but not plasma levels of soluble PD-L1, correlated with clinicopathological data in HNSCC patients. Blocking of PD-L1+ exosomes signaling to PD-1+ T-cells with anti-PD-1 Ab attenuated immune suppression. Altogether, PD-L1+ exosomes serve as useful metrics of disease and immune activity in HNSCC patients.
