Abstract
New drugs are needed for glioblastoma, an aggressive brain tumor with a dismal prognosis. We recently reported that gallium maltolate (GaM) retards the growth of glioblastoma in a rat orthotopic brain tumor model by inhibiting mitochondrial function and iron-dependent ribonucleotide reductase (RR). However, GaM’s mechanism of action at the mitochondrial level is not known. Given the interaction between gallium and iron metabolism, we hypothesized that gallium might target iron-sulfur (Fe-S) cluster-containing mitochondrial proteins. Using Extracellular Flux Analyzer technology, we confirmed that after a 24-h incubation, GaM 50 μmol/L inhibited glioblastoma cell growth by <10% but inhibited cellular oxygen consumption rate by 44% and abrogated mitochondrial reserve capacity. GaM blocked mitochondrial complex I activity and produced a 2.9-fold increase in cellular ROS. NMR spectroscopy revealed that gallium binds to IscU, the bacterial scaffold protein for Fe-S cluster assembly and stabilizes its folded state. Gallium inhibited the rate of in vitro cluster assembly catalyzed by bacterial cysteine desulfurase in a reaction mixture containing IscU, Fe (II), DTT, and L-cysteine. Metformin, a complex I inhibitor, enhanced GaM’s inhibition of complex I, further increased cellular ROS levels, and synergistically enhanced GaM’s cytotoxicity in glioblastoma cells in 2-D and 3-D cultures. Metformin did not affect GaM action on cellular iron uptake or transferrin receptor1 expression nor did it enhance the cytotoxicity of the RR inhibitor Didox. Our results show that GaM inhibits complex I by disrupting iron-sulfur cluster assembly and that its cytotoxicity can be synergistically enhanced by metformin through combined action on complex I.
Highlights
Glioblastoma accounts for approximately 15% of all central nervous system (CNS) tumors and approximately 47% of all malignant CNS tumors [1]
Cells exposed to 50 μmol/L gallium maltolate (GaM) displayed less than a 10% decrease in their growth at 24 h compared to control cells, their basal cellular oxygen consumption rate (OCR, a measure of mitochondrial function) at this time-point was decreased by approximately 44% (Figure 1B)
Using XF Analyzer technology, we show for the first time that GaM blocks the activity of mitochondrial complex I in glioblastoma cells and that it does so at a time-point that precedes a decrease in cell proliferation
Summary
Glioblastoma accounts for approximately 15% of all central nervous system (CNS) tumors and approximately 47% of all malignant CNS tumors [1]. In 2018, an estimated 12,760 new cases of glioblastoma were diagnosed in the US [1] Current treatment for this malignancy has not changed in several years and focuses primarily on maximal surgical resection followed by radiation and temozolomide chemotherapy [2]. Delivery of alternating tumor-treating electric field was shown to prolong patient survival by a few months and FDA-approved as an additional glioblastoma therapy [3]. Outcomes in this disease are dismal; the median survival is reported to be approximately 14 months from diagnosis [4], while the one-year and five-year survival is approximately 40% and 5.5%, respectively [1]. There is a great need to develop additional efficacious therapies
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