Glycyrrhizic acid reduces neutrophil extracellular trap formation to ameliorate colitis-associated colorectal cancer by inhibiting peptidylarginine deiminase 4.

ETHNOPHARMACOLOGICAL RELEVANCE: In traditional Chinese medicine, the radices of Glycyrrhiza uralensis Fisch., known as liquorice, have been used for relieving cough, alleviating pain and harmonizing the actions of all medicinals in a formula. Glycyrrhizic acid (GA), a natural compound derived from licorice, exhibits notable anti-inflammatory properties.

AIM: Neutrophil extracellular trap (NET) generated by peptidylarginine deiminase 4 (PAD4) has been implicated in the progression of colitis to colitis-associated colorectal cancer (CAC). This study aims to investigate whether GA can ameliorate CAC through the inhibition of PAD4 activity and reduction of NET formation.

METHODS

We investigated the correlation between PAD4 expression levels and immune cell infiltration in colorectal cancer utilizing the TIMER database, while also assessing PAD4 levels and activity in human CAC biopsy samples. To evaluate the therapeutic potential of licorice acid on CAC in vivo, we employed the AOM/DSS model and confirmed its inhibitory effects on NET formation in vitro. Furthermore, we explored whether licorice acid can restore immune cell cytotoxicity by diminishing NET formation through fluorescence transfection of CT26 cell lines and subsequent sorting of CD8+ T cells. Additionally, we elucidated the detrimental role of PAD4 in CAC progression using PAD4-/- mice.

RESULTS

We observed that GA ameliorated colonic inflammation, reduced tumorigenicity, and decreased NET formation, as evidenced by decreased levels of PAD4, citH3, MPO and MMP-9. In vitro experiments demonstrated that GA effectively bound to PAD4 and inhibited its enzyme activity. Furthermore, GA prevented epithelial cell destruction while enhancing CD8+ T-cell-mediated tumor killing through the suppression of NET formation in a coculture system.

CONCLUSIONS

We demonstrate that GA inhibits CAC occurrence by suppressing PAD4 activity and reducing NET formation.