Metabolic targeting of multiple myeloma using the Warburg Trap

Background The Warburg effect, characterized by enhanced glycolysis and lactate production, is a universal feature of cancer cells, even under normoxic conditions. The resulting extracellular acidification and intracellular alkalization create a selective vulnerability that is absent in normal tissues. We previously developed the Warburg Trap (WT) approach, which exploits this vulnerability by combining inhibitors of mitochondrial complex I with H + -ionophores to collapse pH homeostasis and energy balance, thereby inducing cancer-specific cell death. Given that the bone marrow microenvironment in multiple myeloma (MM) is markedly acidic (pH 6.7–6.8 compared to ~ 7.4 in normal bone marrow), we hypothesized that MM cells would be highly sensitive to WT treatment. Methods The effects of individual WT drugs and drug combinations on three human myeloma cell lines (hMCLs), primary MM cells and one non-neoplastic human osteoblast-like cell line (hOBs) were analyzed in vitro. To evaluate the cytotoxic effects, the IC 50 values of the drugs were calculated, and the Zero Interaction Potency (ZIP) model was used to identify possible synergies. Cell viability, intracellular pH values (pHi), the invasive potential, apoptosis induction, glucose uptake, ATP levels, mitochondrial depolarization and glucose transporter expression were analyzed using fluorescence microscopy, flow cytometry, RT-qPCR analysis and various in vitro assays. Results Combinations of the mitochondrial complex I inhibitors metformin (Me) and fenofibrate (Fe) with one of the H + -ionophores drotaverine (Dro) or monensin (Mo) significantly induced apoptosis in hMCLs and primary MM cells, while hOBs remained unaffected. The triple medication regimen exhibited a clear synergistic effect compared to monotherapy. Mechanistic analyses revealed a pronounced drop in pH i , disruption of glucose metabolism, depletion of ATP and loss of mitochondrial potential in response to these drugs. Furthermore, the WT drugs suppressed the invasive potential of MM cells and induced the upregulation of the stress-associated gene DDIT3 and the encoded CHOP protein. RT-qPCR profiling of solute carrier family 2 A ( SLC2A ) genes, encoding glucose transporters (GLUTs) revealed that SLC2A1 , SLC2A8 and SLC2A9 were the most abundant transcripts in MM cells, with SLC2A8 showing the most pronounced difference between MM cells and non-neoplastic hOBs. Additionally, SLC2A8 expression associated with the observed WT-drug sensitivity of the analyzed cell lines.