Aspartame (asparagynal)-β-hydroxylase (ASPH) leads to hepatocellular carcinoma (HCC) progression, but the potential molecular mechanism of ASPH on HCC progression to metastasis is still obscure. Our aim was to explore the correlation between ASPH and HIF1α-mediated mitophagy fueling via glutamine during tumor progression to metastasis in HCC.
Bioinformatics analysis constructed a composite gene-gene functional interaction network for ASPH regulatory genes to predict potential cellular mechanisms and then verified in vivo and vitro. ASPH+/-expressing stable cell line of HepG2 and Huh7 cell lines were structured by lentivirus encoding the wild-type ASPH open reading frame and ASPH-CRISPR/Cas9 system. 140 cases of HCC patients and a sbcutaneous xenograft HCC tumor model were used to evaluate HCC tumor growth and progression in vivo. The effect of ASPH on mitophagy, glutamine metabolism, angiogenesis and metastasis in HCC were tested via western blot, RT-PCR, immunohistochemical staining, electron microscopy, hematoxylin-eosin (HE) and immunocytofluorescent staining. The HIF1α inhibitor 2-Methoxyestradiol (2-MeOE2) and the serum free cultivation were used to conduct the related rescue experiments in vitro.
In this study, ASPH improved the expression of LC3-II/LC3-I,Atg7, Atg5 and Beclin1 and down-regulated the expression of TOMM20 and P62. Meanwhile, ASPH also significantly increased the expression of GLS1, which reflects the glutamine metabolism level of tumor cells. In addition, ASPH also significantly inhibited the expressions of HIF1α, BNIP3 and NIX proteins in HCC. Furthermore, a significant association between ASPH and CD34 expression was observed in HCC patients (P = 0.024).
Our finding suggests that ASPH fuels glutamine via HIF1α-mediated mitophagy to drive cancer angiogenesis and metastasis in HCC. ASPH may act as a promising therapeutic target in HCC patients.
Clinical trial identification
Legal entity responsible for the study
National Natural Science Foundation of China (No. 81470877) Beijing Natural Science Foundation (No.7192085).
All authors have declared no conflicts of interest.