HK2 knockdown is associated with context-dependent inflammatory and angiogenesis-related transcriptional programmes...

Deregulation of cellular metabolism is a hallmark of cancer, in which tumour cells upregulate glycolysis and preferentially ferment glucose to lactate in the 'Warburg effect'. Tumour-specific upregulation of hexokinase 2 (HK2), which catalyses the first rate-limiting step of glycolysis, is a key driver of this phenotype and represents a promising therapeutic target in glioblastoma. Here, we sought to identify HK2-associated tumour processes that could serve as therapeutic co-targets by profiling the transcriptomic consequences of HK2 knockdown (HK2KD) in model established glioblastoma cell lines and patient-derived short-term cell cultures.

Glioblastoma cell lines and patient-derived short-term cell cultures were subjected to HK2KD using HK2-targeted siRNA. Glycolytic activity was assessed using the Seahorse Glycolytic Stress Test, and transcriptomic changes were analysed by RNA sequencing.

HK2KD was associated with reduced glycolytic activity in high-HK2-expressing models as well as extensive transcriptional remodelling. Integrative analysis using RNA sequencing, gene ontology and pathway enrichment, protein-protein interaction networks, and transcriptome-based perturbation matching indicated coordinated modulation of inflammatory, immune-associated and angiogenesis-linked gene programmes, including cytokine, chemokine and pro-angiogenic signalling axes. Notably, inflammatory pathway responses diverged markedly between independent glioblastoma models, representing a central finding of this study and highlighting context-dependent immunometabolic transcriptional changes following HK2KD.

These findings suggest that HK2KD is associated with transcriptional changes extending beyond glycolytic pathways, linking metabolic activity to immunological and angiogenic transcriptional networks in a model-dependent manner. Although functional effects on the tumour microenvironment were not directly assessed, this study provides a transcriptomic framework for understanding how HK2 targeting may be associated with changes in tumour-associated signalling pathways, with potential implications for stratified metabolic and combination therapeutic strategies in glioblastoma.