OV16 Improves Radiation-Induced Intestinal Injury by Targeting Transglutaminase 2

Irradiation (IR) can cause intestinal epithelial cell death, damage to crypt stem cells, and mucosal barrier dysfunction, which are the features of radiation-induced intestinal injury (RIII). Our study first discovered a natural small-molecule alkaloid Orychophragine D (OV16) with an obvious radiation protection effect. This study aims to investigate the radiation protection effect of OV16 on RIII and its potential molecular mechanism. The results showed that in vitro OV16 exhibited a significant protective effect on an irradiated human small intestinal epithelial cell-6 (HIEC-6) model. Then, transglutaminase 2 (TGM2), which is the key protein for OV16 to exert its anti-RIII protective effect, was identified as a crucial cellular target of OV16 using drug affinity responsive target stability (DARTS), molecular docking, molecular dynamics simulation, cell thermal shift assay (CETSA), and microscale thermophoresis (MST). Moreover, OV16 can upregulate the expression level of TGM2 in the nucleus of HIEC-6. TGM2 can reduce radiation-induced damage by enhancing the proliferation ability and migration ability of HIEC-6 and reducing the generation of γ-H2AX. Collectively, our study first identified TGM2 as a previously unreported therapeutic target for RIII, and provided a future drug design direction for TGM2 allosteric activators using OV16 as a novel molecular template.