Ubiquitin carboxyl‐terminal hydrolase L1 promotes hypoxia‐inducible factor 1‐dependent tumor cell malignancy in spheroid models

Hypoxia‐inducible factor 1 (HIF‐1) is a critical heterodimeric transcription factor for tumor malignancy. Recently, ubiquitin carboxyl‐terminal hydrolase L1 (UCHL1) has been reported to function as a deubiquitinating enzyme for the stabilization of its α subunit (HIF‐1α). In the present study, we showed that UCHL1 inhibition can be an effective therapeutic strategy against HIF‐1‐dependent tumor malignancy. In 2D monolayer culture, a UCHL1 inhibitor suppressed HIF activity and decreased the transcription of HIF downstream genes by inhibiting the UCHL1‐mediated accumulation of HIF‐1α. Phenotypically, UCHL1 inhibition remarkably blocked cell migration. In 3D spheroid culture models, ectopic expression of UCHL1 significantly upregulated malignancy‐related factors such as solidity, volume, as well as viable cell number in an HIF‐1α‐dependent manner. Conversely, inhibition of the UCHL1‐HIF‐1 pathway downregulated these malignancy‐related factors and also abolished UCHL1‐mediated cell proliferation and invasiveness. Finally, inhibition of UCHL1 promoted HIF‐1α degradation and lowered the expression of HIF‐1 target genes in the 3D model, as also observed in 2D monolayer culture. Our research indicates that the UCHL1‐HIF‐1 pathway plays a crucial role in tumor malignancy, making it a promising therapeutic target for cancer chemotherapy. We revealed that depletion of UCHL1 by siRNA or blockade of its deubiquitinating activity with a specific inhibitor caused a remarkable decrease in HIF‐1 protein levels in the spheroids of UCHL1‐expressing MDA‐MB‐436 cells. The resulting reduction in the expression of HIF‐1 targeted genes in the spheroids, which are closely related to tumor malignancy including metastasis, cell proliferation and angiogenesis, was observed. These findings suggest that the UHCL1‐HIF‐1α pathway is a promising therapeutic target in anticancer therapy.