Combination of downregulating FEN1 and PD-1 blockade enhances antitumor activity of CD8+ T cells against HNSCC cells in vitro

Programmed cell death ligand 1 (PD-L1) and human leukocyte antigen/major histocompatibility complex (HLA/MHC) are two main kinds of immunophenotypes affecting the susceptibility to anti-PD therapy. Our previous study found that down-regulation of flap endonuclease-1 (FEN1) could not only inhibit PD-L1 expression, but also upregulate HLA expression in head and neck squamous cell carcinoma (HNSCC). We aimed to clarify whether downregulating FEN1 cloud enhance the response to PD-1 blockade, and possible mechanisms in HNSCC in vitro. Differential expression of FEN1 in HNSCC tumor and normal tissues were explored in the TIMER and TISIDB datasets. A HNSCC cells/CD8+ T cells co-culture model was established. HNSCC cell cycle and apoptosis were recorded by flow cytometry. Immune activity markers of granzyme A, granzyme B, and PRF1 expressed in the CD8+ T cells, and IFN-γ, IL-2, and TNF-α secreted in the supernatants were detected by western blot, ELISA, respectively. FEN1 was highly expressed in HNSCC and associated with low immune infiltration. Downregulating FEN1 could induce HLA class I expression, and inhibit PD-L1 expression in HNSCC cells. Functionally, FEN1 knockdown enhanced the response to αPD-1 mAb by mediating G2/M phase arrest, apoptosis of HNSCC cells. Mechanistically, targeting FEN1 synergized with αPD-1 mAb could reinforce the antitumor response of CD8+ T cells against HNSCC cells, as indicated by increasing granzyme A, granzyme B, and PRF1 expressions, and promoting IFN-γ, IL-2, and TNF-α secretions. These findings might offer a potential combined strategy for patients resistant to anti-PD therapy via combining FEN1 knockdown and PD-1 blockade.