Argininosuccinate synthase 1 suppresses tumor progression through activation of PERK/eIF2α/ATF4/CHOP axis in hepatocellular carcinoma

Background Hepatocellular carcinoma (HCC) is one of the most common malignant cancers worldwide, and liver cancer has increased in mortality due to liver cancer because it was detected at an advanced stages in patients with liver dysfunction, making HCC a lethal cancer. Accordingly, we aim to new targets for HCC drug discovery using HCC tumor spheroids. Methods Our comparative proteomic analysis of HCC cells grown in culture as monolayers (2D) and spheroids (3D) revealed that argininosuccinate synthase 1 (ASS1) expression was higher in 3D cells than in 2D cells due to upregulated endoplasmic reticulum (ER) stress responses. We investigated the clinical value of ASS1 in Korean patients with HCC. The mechanism underlying ASS1-mediated tumor suppression was investigated in HCC spheroids. ASS1-mediated improvement of chemotherapy efficiency was observed using high content screening in an HCC xenograft mouse model. Results Studies of tumor tissue from Korean HCC patients showed that, although ASS1 expression was low in most samples, high levels of ASS1 were associated with favorable overall survival of patients. Here, we found that bidirectional interactions between ASS1 ER stress responses in HCC-derived multicellular tumor spheroids can limit HCC progression. ASS1 overexpression effectively inhibited tumor growth and enhanced the efficacy of in vitro and in vivo anti-HCC combination chemotherapy via activation of the PERK/eIF2 alpha/ATF4/CHOP axis, but was not dependent on the status of p53 and arginine metabolism. Conclusions These results demonstrate the critical functional roles for the arginine metabolism-independent tumor suppressor activity of ASS1 in HCC and suggest that upregulating ASS1 in these tumors is a potential strategy in HCC cells with low ASS1 expression.

Automated summary

The expression of ASS1 was higher in HCC spheroids than in monolayer cells, and high levels of ASS1 were associated with favorable overall survival in Korean HCC patients. Bidirectional interactions between ASS1 ER stress responses in HCC-derived spheroids limit HCC progression. Overexpression of ASS1 inhibited tumor growth and enhanced the efficacy of anti-HCC chemotherapy via activation of the PERK/eIF2 alpha/ATF4/CHOP axis, independent of p53 and arginine metabolism.