*Article* **Trichosanthin Promotes Anti-Tumor Immunity through Mediating Chemokines and Granzyme B Secretion in Hepatocellular Carcinoma**

**Kaifang Wang 1,2,†, Xiaona Wang 3,†, Minghuan Zhang 3, Zhenguang Ying 1, Zeyao Zhu 4, Kin Yip Tam 5, Chunman Li 6, Guowei Zhou 3, Feng Gao 1, Meiqi Zeng 1, Stephen Cho Wing Sze 2, Xia Wang 3,\* and Ou Sha 1,\***


**Abstract:** Trichosanthin (TCS) is a type I ribosome-inactivating protein extracted from the tuberous root of the plant *Trichosanthes*. TCS shows promising potential in clinical drug abortion, anti-tumor and immunological regulation. However, the molecular mechanisms of its anti-tumor and immune regulation properties are still not well discovered. In the present study, we investigated the anti-tumor activity of TCS in hepatocellular carcinoma (HCC), both in vitro and in vivo. Both HCC cell lines and xenograft tumor tissues showed considerable growth inhibition after they were treated with TCS. TCS provoked caspase-mediated apoptosis in HCC cells and xenograft tumor tissues. The recruitment of CD8+ T cells to HCC tissues and the expression of chemokines, CCL2 and CCL22, were promoted upon TCS treatment. In addition, TCS induced an upregulation of Granzyme B (GrzB), TNF-α and IFN-γ in HCC tissues, which are the major cytotoxic mediators produced by T cells. Furthermore, TCS also resulted in an increase of mannose-6-phosphate receptor (M6PR), the major receptor of GrzB, in HCC tissues. In summary, these results suggest that TCS perhaps increases T-cell immunity via promoting the secretion of chemokines and accelerating the entry of GrzB to HCC cells, which highlights the potential role of TCS in anti-tumor immunotherapy.

**Keywords:** Trichosanthin (TCS); hepatocellular carcinoma (HCC); T cell; chemokine; Granzyme B (GrzB); apoptosis
