Iranian Journal of Basic Medical Sciences

Iranian Journal of Basic Medical Sciences

ADAMTS9-AS2 suppresses hypoxia-driven glycolytic reprogramming through EZH2-dependent regulation of ENO1 alternative splicing in bladder cancer

Articles in Press

Document Type : Original Article

Authors
Gang Liu 1, 2
Xiaoying Wang 1, 2
Dingwen Gui 1, 2
Tingting Zhang 2, 3
1 Department of Urology, Huangshi Central Hospital, Affiliated Hospital of Hubei Polytechnic University, Huangshi, Hubei, China
2 Hubei Key Laboratory of Kidney Disease Pathogenesis and Intervention, Huangshi, Hubei, China
3 Department of Clinical Laboratory, Huangshi Central Hospital, Affiliated Hospital of Hubei Polytechnic University, Huangshi, Hubei, China
10.22038/ijbms.2026.96791.20827
Abstract
Objective(s): Tumor hypoxia is a key driver of metabolic reprogramming and aggressive progression in bladder cancer; however, the upstream long non-coding RNA (lncRNA)-mediated epigenetic mechanisms that regulate these processes remain incompletely understood. This study investigated the role of ADAMTS9-AS2 in hypoxia-induced metabolic remodeling and its interaction with EZH2 and ENO1 alternative splicing.
Materials and Methods: Bladder cancer cell lines (T24 and UM-UC-3) were cultured under hypoxic or normoxic conditions. Stable knockdown and overexpression models of ADAMTS9-AS2, EZH2, and ENO1 were generated using lentiviral vectors. Functional assays included CCK-8, colony formation, wound-healing, Transwell invasion, Seahorse XF metabolic analysis, glucose uptake, lactate/ATP quantification, ROS detection, JC-1 staining, and transmission electron microscopy. Molecular interactions were assessed by RNA-FISH, RNA immunoprecipitation, biotinylated RNA pull-down, ChIP-qPCR, and RNA sequencing with alternative splicing analysis (rMATS/SUPPA2). In vivo effects were evaluated using subcutaneous xenograft models with bioluminescence imaging and immunohistochemistry. Rescue experiments established pathway causality.
Results: Hypoxia significantly reduced ADAMTS9-AS2 expression, which correlated with advanced clinicopathological stage and poor survival. ADAMTS9-AS2 directly interacted with nuclear EZH2 and regulated chromatin remodeling at the ENO1 locus, thereby altering ENO1 alternative splicing. Loss of ADAMTS9-AS2 promoted glycolysis, mitochondrial dysfunction, ROS accumulation, proliferation, migration, invasion, and xenograft growth. Conversely, ENO1 silencing or EZH2 signaling restoration attenuated these malignant phenotypes.
Conclusion: ADAMTS9-AS2 acts as a hypoxia-responsive tumor suppressor that restrains bladder cancer progression by regulating the EZH2/ENO1 signaling axis. These findings identify a novel lncRNA-mediated epigenetic mechanism linking hypoxia, alternative splicing, and metabolic reprogramming, highlighting ADAMTS9-AS2 and the EZH2/ENO1 pathway as promising therapeutic targets for bladder cancer.
Keywords
Urinary bladder neoplasms
Chromatin remodeling
Glycolysis
Tumor hypoxia
Enolase
Subjects
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Iranian Journal of Basic Medical Sciences

Articles in Press, Accepted Manuscript
Available Online from 11 October 2026