Long-term adaptation of lymphoma cells to hypoxia is mediated by diverse molecular mechanisms that are targetable with specific inhibitors

In this study we analyzed impact of long-term hypoxia on a panel of lymphoma cell lines. Only 2 out of 8 tested lymphoma cell lines (Ramos, and HBL2) survived > 4 weeks under deep hypoxia (1% O2). The hypoxia-adapted (HA) Ramos and HBL2 cells had severely decreased proliferation rate accompanied by complex changes of the transcriptome, proteome, and metabolome. Seahorse analysis demonstrated near complete blockage of both oxidative phosphorylation and glycolytic pathways. Both transcriptome and proteome analyses showed significant downregulation of mitochondrial respiration complexes I and IV, many mitochondrial ribosomal proteins, and increase of proteins regulating glycolysis and mitophagy. Both HA cell lines had an increased total mitochondrial mass, but a decreased mitochondrial DNA content compared to normoxic controls. Sensitivity of HA cells to 2-deoxyglucose, an inhibitor of glycolysis, and to A1155463, a BCL-XL inhibitor were, was markedly increased. Indeed, co-culture on CD40 ligand expressing fibroblasts (that induce BCL-XL expression) significantly increased survival of lymphoma cells under hypoxia. Of note, prolyl hydroxylase P4HA1 involved in the stabilization of hypoxia-induced factor (HIF) 1 alpha was increased in both HA cell lines on both mRNA and protein levels. Transgenic (over)expression of P4HA1 in hypoxia-sensitive MINO cells was associated with increased survival under hypoxia. Our data suggest that under long-term hypoxia, lymphoma cells try to compensate for the decrease in ATP production from the oxidative phosphorylation process by boosting structural machinery of glycolysis, as well as amino-acid recycling by mitophagy, on which they become vitally dependent for survival, and which can be targeted by specific inhibitors. The data also suggest increased dependence of lymphoma cells on BCL-XL under hypoxia. Finally, P4HA1 plausibly represents a novel druggable target for more effective elimination of hypoxia-adapted lymphoma cells.