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Background: Head and neck cancers (HNSCC) are highly immunosuppressive. Plasma-derived exosomes of HNSCC patients carry immunomodulatory molecules, and their cargo correlates with clinical parameters. Here, we evaluated the exosomal molecular profile for early detection of treatment failure in locally advanced HNSCC patients treated with conventional therapy.Methods: Plasma from 17 HNSCC patients was collected before, during, and after treatment by surgery with adjuvant (chemo)radiation and at recurrence. Exosomes were isolated by size-exclusion chromatography. Total exosomal protein (TEP) was used to estimate exosome load and on-bead flow cytometry to evaluate relative fluorescence intensity (RFI) of tumour-associated and immunoregulatory proteins on exosomes. Exosomal effects on the activity of and adenosine production by T cells was assessed by flow cytometry and mass spectrometry.Results: TEP and the ratio of tumour-/immune-cell-derived exosomes varied during and after therapy with an overall decrease in the tumour-free follow-up but an increase at recurrence. RFI values of immunoregulatory proteins on exosomes, their ability for T cell inhibition and adenosine production changed during and after therapy. PD-L1 was the earliest discriminator for treatment failure and disease-free survival.Conclusions: Monitoring of plasma exosomes during therapy represents a promising opportunity for early detection of treatment failure and risk stratification to delay/avoid recurrence. [ABSTRACT FROM AUTHOR]

Summary: Exosomes in plasma of head and neck squamous cell carcinoma (HNSCC) patients comprise subsets of vesicles derived from various cells. Recently, we separated CD3(+) from CD3(–) exosomes by immune capture. CD3(–) exosomes were largely tumour‐derived (CD44v3+). Both subsets carried immunosuppressive proteins and inhibited functions of human immune cells. The role of these subsets in immune cell reprogramming by the tumour was investigated by focusing on the adenosine pathway components. Spontaneous adenosine production by CD3(+) or CD3(–) exosomes was measured by mass spectrometry, as was the production of adenosine by CD4+CD39+ regulatory T cells (Treg) co‐incubated with these exosomes. The highest level of CD39/CD73 ectoenzymes and of adenosine production was found in CD3(–) exosomes in patients with the stages III/IV HNSCCs). Also, the production of 5′‐AMP and purines was significantly higher in Treg co‐incubated with CD3(–) than CD3(+) exosomes. Consistently, CD26 and adenosine deaminase (ADA) levels were higher in CD3(+) than CD3(–) exosomes. ADA and CD26 levels in CD3(+) exosomes were significantly higher in patients with early (stages I/II) than advanced (stages III/IV) disease. HNSCC patients receiving and responding to photodynamic therapy had increased ADA levels in CD3(+) exosomes with no increase in CD3(–) exosomes. The opposite roles of CD3(+) ADA+CD26+ and CD3(–)CD44v3+ adenosine‐producing exosomes in early versus advanced HNSCC suggest that, like their parent cells, these exosomes serve as surrogates of immune suppression in cancer. [ABSTRACT FROM AUTHOR]