The protease interpain A of Prevotella intermedia promotes human oral squamous cell carcinoma cells proliferation and migration

Abstract

Prevotella intermedia has been increasingly recognized as a potential contributor to oral squamous cell carcinoma (OSCC), yet the underlying mechanisms remain poorly defined. In this study, we identified interpain A (InpA), a cysteine protease secreted by P. intermedia, as a key virulence factor that promotes oral epithelial cell proliferation and OSCC cell migration. Conditioned medium (CM) derived from P. intermedia strain ATCC 25611 enhanced proliferation in both normal oral keratinocytes (SG cells) and OSCC cell lines (SCC-15, SAS). The pro-proliferative effect was abolished upon heat inactivation of the CM and inhibited by E64, a cysteine protease inhibitor, and FSLLRY-NH2, a PAR-2 antagonist, suggesting the involvement of protease-activated receptor-2 (PAR-2) signaling. InpA was highly secreted by strain ATCC 25611 but not by clinical isolates lacking proliferative effects, and RT-qPCR confirmed higher inpA expression in OSCC-derived strains compared to non-cancer controls. Recombinant InpA (rInpA) mimicked the effect of CM, inducing proliferation and migration, which were blocked by PAR-2 inhibition. Transcriptomic and protein-level screening in SG cells revealed activation of EGFR and downstream RAS-RAF-MEK-ERK signaling. Furthermore, in a colorectal cancer (CRC) mouse model, oral administration of P. intermedia led to increased tumor formation, suggesting a broader oncogenic potential. These findings highlight InpA as a PAR-2-activating protease that may contribute to OSCC and potentially other cancers associated with oral microbiota dysbiosis.