Cancer cells possess aberrant proteomes that can arise by the disruption of genes involved in physiological protein degradation. Here we demonstrate the presence of promoter CpG island hypermethylation-linked inactivation of DERL3 (Derlin-3), a key gene in the endoplasmic reticulum-associated protein degradation pathway, in human tumours. The restoration of in vitro and in vivo DERL3 activity highlights the tumour suppressor features of the gene. Using the stable isotopic labelling of amino acids in cell culture workflow for differential proteome analysis, we identify SLC2A1 (glucose transporter 1, GLUT1) as a downstream target of DERL3. Most importantly, SLC2A1 overexpression mediated by DERL3 epigenetic loss contributes to the Warburg effect in the studied cells and pinpoints a subset of human tumours with greater vulnerability to drugs targeting glycolysis.