Inhibition Of Mitochondrial Metabolism Research Articles

Nutrient-sensitizing drug repurposing screen identifies lomerizine as a mitochondrial metabolism inhibitor of chronic myeloid leukemia.

In chronic myeloid leukemia (CML), the persistence of leukemic stem cells (LSCs) after treatment with tyrosine kinase inhibitors (TKIs), such as imatinib, can lead to disease relapse. It is known that therapy-resistant LSCs rely on oxidative phosphorylation (OXPHOS) for their survival and that targeting mitochondrial respiration sensitizes CML LSCs to imatinib treatment. However, current OXPHOS inhibitors have demonstrated limited efficacy or have shown adverse effects in clinical trials, highlighting that identification of clinically safe oxidative pathway inhibitors is warranted. We performed a high-throughput drug repurposing screen designed to identify mitochondrial metabolism inhibitors in myeloid leukemia cells. This identified lomerizine, a US Food and Drug Administration (FDA)-approved voltage-gated Ca2+ channel blocker now used for the treatment of migraines, as one of the top hits. Transcriptome analysis revealed increased expression of voltage-gated CACNA1D and receptor-activated TRPC6 Ca2+ channels in CML LSCs (CD34+CD38-) compared with normal counterparts. This correlated with increased endoplasmic reticulum (ER) mass and increased ER and mitochondrial Ca2+ content in CML stem/progenitor cells. We demonstrate that lomerizine-mediated inhibition of Ca2+ uptake leads to ER and mitochondrial Ca2+ depletion, with similar effects seen after CACNA1D and TRPC6 knockdown. Through stable isotope-assisted metabolomics and functional assays, we observe that lomerizine treatment inhibits mitochondrial isocitrate dehydrogenase activity and mitochondrial oxidative metabolism and selectively sensitizes CML LSCs to imatinib treatment. In addition, combination treatment with imatinib and lomerizine reduced CML tumor burden, targeted CML LSCs, and extended survival in xenotransplantation model of human CML, suggesting this as a potential therapeutic strategy to prevent disease relapse in patients.

Deoxycytidine kinase inactivation enhances gemcitabine resistance and sensitizes mitochondrial metabolism interference in pancreatic cancer

Pancreatic ductal adenocarcinoma (PDAC) is considered one of the most lethal forms of cancer. Although in the last decade, an increase in 5-year patient survival has been observed, the mortality rate remains high. As a first-line treatment for PDAC, gemcitabine alone or in combination (gemcitabine plus paclitaxel) has been used; however, drug resistance to this regimen is a growing issue. In our previous study, we reported MYC/glutamine dependency as a therapeutic target in gemcitabine-resistant PDAC secondary to deoxycytidine kinase (DCK) inactivation. Moreover, enrichment of oxidative phosphorylation (OXPHOS)-associated genes was a common property shared by PDAC cell lines, and patient clinical samples coupled with low DCK expression was also demonstrated, which implicates DCK in cancer metabolism. In this article, we reveal that the expression of most genes encoding mitochondrial complexes is remarkably upregulated in PDAC patients with low DCK expression. The DCK-knockout (DCK KO) CFPAC-1 PDAC cell line model reiterated this observation. Particularly, OXPHOS was functionally enhanced in DCK KO cells as shown by a higher oxygen consumption rate and mitochondrial ATP production. Electron microscopic observations revealed abnormal mitochondrial morphology in DCK KO cells. Furthermore, DCK inactivation exhibited reactive oxygen species (ROS) reduction accompanied with ROS-scavenging gene activation, such as SOD1 and SOD2. SOD2 inhibition in DCK KO cells clearly induced cell growth suppression. In combination with increased anti-apoptotic gene BCL2 expression in DCK KO cells, we finally reveal that venetoclax and a mitochondrial complex I inhibitor are therapeutically efficacious for DCK-inactivated CFPAC-1 cells in in vitro and xenograft models. Hence, our work provides insight into inhibition of mitochondrial metabolism as a novel therapeutic approach to overcome DCK inactivation-mediated gemcitabine resistance in PDAC patient treatment.

Abstract C129: Development of new increased potency Ref-1/APE1 targeted inhibitors that show promise for clinical applications against solid tumors

Abstract AP endonuclease-1/Redox factor-1 (APE1/Ref-1 or Ref-1) is a multifunctional protein that is over-expressed in most aggressive cancers and impacts various cancer cell signaling pathways. Among its primary functions, Ref-1’s redox activity plays a significant role in activating transcription factors (TFs) such as NFkB, HIF1a, and AP-1, among others, which are crucial contributors to the development of tumors and metastatic growth. Therefore, development of potent, selective inhibitors to target Ref-1 redox function is an appealing approach for therapeutic intervention. A first-generation compound, APX3330 successfully completed phase I clinical trial in adults with progressing solid tumors with favorable response rate, pharmacokinetics (PK), and minimal toxicity. These positive results prompted us to develop even more potent analogs of APX3330 to effectively target Ref-1 in solid tumors. In this study, we present structure-activity relationship (SAR) identification and validation of more potent lead compounds that exhibit a similar/better safety profile to APX3330. Pharmacokinetics, mouse and human S9 fraction metabolic stability, in silico ADMET properties, ligand-based WaterLOGSY NMR measurements, TF-driven luciferase activity, and two 3-dimensional (3D) cell killing assays (interstitial Tumor-Microenvironment (TME) on a Chip and 3D spheroid) were used for inhibitor characterization. These efforts have elucidated five compounds with improved PK, target engagement, and increased efficacy for cell killing compared to parent compound. Ligand-based WaterLOGSY NMR measurements demonstrate direct interaction of these inhibitors with Ref-1. Target inhibition was further confirmed following Ref-1 treatment via reporter assay with significant and dose-dependent decreases in TF-driven luciferase activity (3-6-fold compared to APX3330). In two 3D co-culture models, second-generation Ref-1 redox analogs suppressed tumor survival significantly with significantly increased potency (~10-fold) compared to APX3330. APX2014, 44, and 51 also inhibited tumor cell proliferation more dramatically (1.5-3.5-fold) than cancer-associated fibroblasts from the TME. Two compounds, APX2044 and 51 outperformed the other analogs as well as the parent compound as assessed by inhibition of downstream TFs regulated by Ref-1 activity, inhibition of mitochondrial metabolism, greater potency against tumors than CAFs in monolayer and in 3D co-culture experiments. PK studies also demonstrated that efficacious doses are clinically achievable for these new inhibitors. Additionally, these findings have allowed us to develop additional analogs for further refinement. Thus, our preclinical findings identified more potent inhibitors of Ref-1 redox function (5-10-fold more than APX3330) that can be translated to the clinic. This study provides invaluable information regarding new Ref-1 analogs for advancement to in vivo and eventual IND-enabling studies and demonstrates the effectiveness of these compounds in sophisticated in vitro models in cancers that are in need of therapeutic options. Citation Format: Silpa Gampala, Hye-Ran Moon, Randall Wireman, Jacqueline Peil, Sonia Kiran, Andi Masters, Christine Bach, Whitney Smith-Kinnamen, Emma Doud, Ratan Rai, Amber L Mosley, Millie M Georgiadis, Bumsoo Han, Chi Zhang, Chafiq Hamdouchi, James Wikel, Mark R Kelley, Melissa L Fishel. Development of new increased potency Ref-1/APE1 targeted inhibitors that show promise for clinical applications against solid tumors [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr C129.

Staufen1 controls mitochondrial metabolism via HIF2α in embryonal rhabdomyosarcoma and promotes tumorigenesis.

Elevated mitochondrial metabolism promotes tumorigenesis of Embryonal Rhabdomyosarcomas (ERMS). Accordingly, targeting oxidative phosphorylation (OXPHOS) could represent a therapeutic strategy for ERMS. We previously demonstrated that genetic reduction of Staufen1 (STAU1) levels results in the inhibition of ERMS tumorigenicity. Here, we examined STAU1-mediated mechanisms in ERMS and focused on its potential involvement in regulating OXPHOS. We report the novel and differential role of STAU1 in mitochondrial metabolism in cancerous versus non-malignant skeletal muscle cells (NMSkMCs). Specifically, our data show that STAU1 depletion reduces OXPHOS and inhibits proliferation of ERMS cells. Our findings further reveal the binding of STAU1 to several OXPHOS mRNAs which affects their stability. Indeed, STAU1 depletion reduced the stability of OXPHOS mRNAs, causing inhibition of mitochondrial metabolism. In parallel, STAU1 depletion impacted negatively the HIF2α pathway which further modulates mitochondrial metabolism. Exogenous expression of HIF2α in STAU1-depleted cells reversed the mitochondrial inhibition and induced cell proliferation. However, opposite effects were observed in NMSkMCs. Altogether, these findings revealed the impact of STAU1 in the regulation of mitochondrial OXPHOS in cancer cells as well as its differential role in NMSkMCs. Overall, our results highlight the therapeutic potential of targeting STAU1 as a novel approach for inhibiting mitochondrial metabolism in ERMS.

Mystery of bisphenol F-induced nonalcoholic fatty liver disease-like changes: Roles of Drp1-mediated abnormal mitochondrial fission in lipid droplet deposition

As one of the major substitutes for bisphenol A (BPA), bisphenol F (BPF) has been widely used. Our previous study demonstrated that BPF exposure facilitates lipid droplet deposition in hepatic cells, contributing to nonalcoholic fatty liver disease (NAFLD)-like changes. However, the underlying mechanisms remain poorly understood. Here, with a metabolic cage system, we observed the perturbation of energy metabolism in mice treated with BPF. BPF obviously suppressed metabolic capacity, which manifested as decreased energy expenditure, low O2 consumption and CO2 levels in mice. Consistent with the in vivo results, a Seahorse XF Cell Mito Stress Test showed significant reductions in mitochondrial ATP production capacity, maximum respiratory capacity, and residual respiratory capacity after BPF treatment in an in vitro study. Electron microscopy revealed a striking increase in mitochondrial fission that was synchronous with excessive expression and activation of dynamin-related protein 1 (Drp1). Intriguingly, chemical inhibition of Drp1 by Mdivi-1 and/or silencing of Drp1 dramatically hampered mitochondrial fission and ameliorated BPF-induced lipid droplet deposition both in mouse liver and human hepatic cells. Mechanistically, mitochondrial dynamics imbalance played prominent roles in these processes, since suppression of Drp1 by chemical inhibition or knockdown substantially reversed BPF-induced mitochondrial fission and ameliorated the suppression of mitochondrial metabolism as well as excessive mitochondrial ROS, which was verified to be key to lipid droplet deposition. Collectively, the findings of the current study reveal previously unrecognized effects involving Drp1-mediated mitochondrial injury in BPF-induced lipid droplet deposition. Therefore, targeted intervention against mitochondrial dysfunction may be a promising therapeutic strategy for BPF-induced NAFLD-like changes.

Preclinical evaluation of Mito-LND, a targeting mitochondrial metabolism inhibitor, for glioblastoma treatment

BackgroundGlioblastoma (GBM) is a brain tumor with the highest level of malignancy and the worst prognosis in the central nervous system. Mitochondrial metabolism plays a vital role in the occurrence and development of cancer, which provides critical substances to support tumor anabolism. Mito-LND is a novel small-molecule inhibitor that can selectively inhibit the energy metabolism of tumor cells. However, the therapeutic effect of Mito-LND on GBM remains unclear.MethodsThe present study evaluated the inhibitory effect of Mito-LND on the growth of GBM cells and elucidated its potential mechanism.ResultsThe results showed that Mito-LND could inhibit the survival, proliferation and colony formation of GBM cells. Moreover, Mito-LND induced cell cycle arrest and apoptosis. Mechanistically, Mito-LND inhibited the activity of mitochondrial respiratory chain complex I and reduced mitochondrial membrane potential, thus promoting ROS generation. Importantly, Mito-LND could inhibit the malignant proliferation of GBM by blocking the Raf/MEK/ERK signaling pathway. In vivo experiments showed that Mito-LND inhibited the growth of GBM xenografts in mice and significantly prolonged the survival time of tumor-bearing mice.ConclusionTaken together, the current findings support that targeting mitochondrial metabolism may be as a potential and promising strategy for GBM therapy, which will lay the theoretical foundation for further clinical trials on Mito-LND in the future.

Downregulation of cyclic adenosine monophosphate levels in leukocytes of hibernating captive black bears is similar to reported cyclic adenosine monophosphate findings in major depressive disorder.

Cyclic adenosine monophosphate (cAMP) levels in the lymphoblasts and leukocytes of patients with major depressive disorder (MDD) have been reported to be downregulated compared to in controls. cAMP is a derivative of adenosine triphosphate (ATP), and low ATP turnover has been reported in the state of hypometabolism associated with human MDD and with mammalian hibernation due to suppression of mitochondrial metabolism. Similarities have been noted between many state-dependent neurobiological changes associated with MDD in humans and with mammalian hibernation. To compare cAMP levels between human MDD and mammalian hibernation and to investigate whether cAMP downregulation is another state-dependent neurobiological finding, we measured cAMP concentrations in lysed leukocytes, plasma, and serum in serial blood specimens from nine female captive black bears (Ursus americanus; CBBs), and cortisol levels in serum from 10 CBBs. Cortisol levels were significantly higher during hibernation in CBBs, confirming previous findings in hibernating black bears and similar to findings in humans with MDD. cAMP levels were significantly lower during hibernation versus active states (pre-hibernation and exit from hibernation) and were similar to the cAMP downregulation reported in MDD patients versus euthymic patients or controls. cAMP level changes during the different states (hibernation, pre-hibernation, active) confirm their state-dependent status. These findings are similar to the neurobiological findings associated with the hypometabolism (metabolic depression) observed during mammalian hibernation and reported during MDD. A sudden increase in cAMP levels was observed before entrance into pre-hibernation and during exit from hibernation. Further investigation is suggested into the possible role of elevated cAMP levels in initiation of the chain reaction of changes in gene expression, proteins, and enzymes leading to the suppression of mitochondrial metabolism and to low ATP turnover. This process leads to hypometabolism, the old adaptive mechanism that is used by organisms for energy preservation and is associated with both mammalian hibernation and human MDD.

Inhibition of mitochondrial metabolism by (-)-jerantinine A: synthesis and biological studies in triple-negative breast cancer cells.

A concise semi-synthesis of the Aspidosperma alkaloids, (-)-jerantinine A and (-)-melodinine P, and derivatives thereof, is reported. The novel compounds were shown to have potent activity against MDA-MB-231 triple-negative breast cancer cells. Furthermore, unbiased metabolomics and live cell reporter assays reveal (-)-jerantinine A alters cellular redox metabolism and induces oxidative stress that coincides with cell cycle arrest.

Abstract B012: Differential stress responses drive asparagine exchange in pancreatic cancer

Abstract Pancreatic ductal adenocarcinoma (PDAC) is the third leading cause of cancer-related death in the United States, contrasting a relatively low incidence rate. A principal barrier to the treatment of pancreatic cancer is the densely fibrotic tumor microenvironment and the high interstitial pressure which acts to collapse blood vessels, impairing the delivery of chemotherapy. This lack of functional vasculature limits nutrient availability within the tumor, causing cancer cells to develop numerous metabolic adaptations to allow for proliferation in hypoxic, austere conditions. Recent studies have demonstrated several clonal behaviors of cancer cells within individual pancreatic tumors, although it is unclear if this is true of metabolism. Exploring this, we found the metabolic profiles and sensitivities of a series of clonal cell lines derived from a single pancreatic tumor clustered into two distinct groups. These groups demonstrated differential sensitivities to inhibitors of mitochondrial metabolism; whereby the growth of the sensitive clones during impaired respiration can be rescued through co-culture with the insensitive cells. Profiling metabolites exchanged by the clonal populations, we identified this is mediated by asparagine released by insensitive cells. Mechanistically, the production of asparagine is driven by differential stress response pathway activation between clones. We further determined that exogenous asparagine supports the growth of both cancer and non-cancerous cells during limited respiration. Conversely, we found that the use of asparaginase to degrade asparagine sensitizes PDA tumors to mitochondrial inhibition. Collectively these data demonstrate distinct metabolic classes of cancer cells within single tumors and identify asparagine as a potential target to sensitize tumors to mitochondrial inhibition. Citation Format: Cecily Anaraki, Galloway Thurston, Nina G. Steele, Samuel A. Kerk, Hanna Hong, Peter Sajjakulnukit, Anthony Andren, Catherine Felton, Alica Beutel, Rima Singh, Heather R. Christofk, Andrea Viale, Howard Crawford, Marina Pasca di Magliano, Claus Jorgensen, Costas A. Lyssiotis, Christopher J. Halbrook. Differential stress responses drive asparagine exchange in pancreatic cancer [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer; 2022 Sep 13-16; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2022;82(22 Suppl):Abstract nr B012.

Abstract 12401: Short-term Pdl-1 and Ctla-4 Blocking Agent Therapy Reduces Myocardial Pampk and Il-10 Levels, Increases Vascular Nf-kb Expression and Serum Il-1 / Il-6 Levels

Introduction: Several strategies based on immune checkpoint inhibitors (ICIs) have been developed or are under investigation for cancer therapy, opening to advantages in cancer outcomes. However, several ICIs-induced side effects emerged in these patients, especially a rare but clinically significant cardiotoxicity with high rate of mortality. Hypothesis: We analyzed the differential vasculo and cardiotoxicity of Pembrolizumab, Nivolumab and Ipilimumab in preclinical models highlighting on the molecular pathways involved. Methods: C57 female mice were treated with Ipilimumab, Pembrolizumab or Nivolumab (15 mg/kg) through intraperitoneal injection for 10 days. pAMPK expression and vascular p65/NF-kB was analyzed through Immunohistochemistry. Cardiac and systemic levels of IL-1α, IL-1β, IL-2, IL-6 and IL-10 were quantified through ELISA method Results: In preclinical models, all ICIs increased p65/NF-kB expression in vascular endothelial cells of mice compared to saline group. IHC analysis indicates that ICIs reduced pAMPK expression in myocardial cells. Notably, tissue levels of IL-1α, IL-1β, IL-2, IL-6 were strongly increased in ICIs group vs saline (p<0.05 vs saline). On the other hand, levels of the anti-inflammatory cytokine IL-10 were strongly reduced ( overall reduction of 43.3-54.5 % vs saline) Conclusions: In preclinical models, a short treatment with ICIs is sufficient to confer a pro-inflammatory phenotype in the heart of mice. Furthermore, reductions in pAMPK indicate the inhibition of mitochondrial metabolism after ICIs. Furthermore ICIs induce chemokines involved in fibrosis, myocarditis and vascultitis. These results could indicate new therapeutic candidates aimed to prevent myocardial and vascular damages in cancer patients treated with ICIs

Pembrolizumab, Ipilimumab and Nivolumab reduces cardiac pAMPK and IL-10, increases vascular NF-kB expression and levels of IL-1b, IL-2 and IL-6 in myocardial tissue

Abstract Background Several strategies based on immune checkpoint inhibitors (ICIs) have been developed or are under investigation for cancer therapy, opening to advantages in cancer outcomes. However, several ICIs-induced side effects emerged in these patients, especially a rare but clinically significant cardiotoxicity with high rate of mortality. Purpose We analyzed the differential vasculo and cardiotoxicity of Pembrolizumab, Nivolumab and Ipilimumab in preclinical models highlighting on the molecular pathways involved. Methods C57 female mice were treated with Ipilimumab, Pembrolizumab or Nivolumab (15 mg/kg) through intraperitoneal injection for 10 days. pAMPK expression and vascular p65/NF-kB was analyzed through Immunohistochemistry. Cardiac levels of IL-1α, IL-1β, IL-2, IL-6 and IL-10 were quantified through ELISA method Results In preclinical models, all ICIs increased p65/NF-kB expression in vascular endothelial cells of mice compared to saline group. IHC analysis indicates that ICIs reduced pAMPK expression in myocardial cells. Notably, tissue levels of IL-1α, IL-1β, IL-2, IL-6 were strongly increased in ICIs group vs saline (p<0.05 vs saline). On the other hand, levels of the anti-inflammatory cytokine IL-10 were strongly reduced (overall reduction of 43.3–54.5% vs saline) Conclusion In preclinical models, a short treatment with ICIs is sufficient to confer a pro-inflammatory phenotype in the heart of mice. Furthermore, reductions in pAMPK indicate the inhibition of mitochondrial metabolism after ICIs. Furthermore ICIs induce chemokines involved in fibrosis, myocarditis and vascultitis. These results could indicate new therapeutic candidates aimed to prevent myocardial and vascular damages in cancer patients treated with ICIs. Funding Acknowledgement Type of funding sources: Public Institution(s). Main funding source(s): Ricerca Corrente, Ministero della Salute

Small molecules targeting the NADH-binding pocket of VDAC modulate mitochondrial metabolism in hepatocarcinoma cells.

Voltage dependent anion channels (VDAC) control the flux of most anionic respiratory substrates, ATP, ADP, and small cations, crossing the outer mitochondrial membrane. VDAC closure contributes to the partial suppression of mitochondrial metabolism that favors the Warburg phenotype of cancer cells. Recently, it has been shown that NADH binds to a specific pocket in the inner surface of VDAC1, also conserved in VDAC2 and 3, closing the channel. We hypothesized that binding of small molecules to the NADH pocket, maintain VDAC in an open configuration by preventing closure induced by NADH and possible other endogenous regulators. We screened in silico, the South Carolina Compound Collection SC3 (~ 100,000 proprietary molecules), using shape-based queries of the NADH binding region of VDAC. After molecular docking of selected compounds, we physically screened candidates using mitochondrial membrane potential (ΔΨm), as an overall readout of mitochondrial metabolism. We identified SC18, as the most potent compound. SC18 bound to VDAC1, as assessed by a thermal shift assay. Short-term treatment with SC18 decreased ΔΨm in SNU-449 and HepG2 human hepatocarcinoma cells. Mitochondrial depolarization was similar in wild type, VDAC1/2, 1/3, and 2/3 double KO HepG2 cells indicating that the effect of SC18 was not VDAC isoform-dependent. In addition, SC18 decreased mitochondrial NADH and cellular ATP production; and increased basal respiration. Long-term exposure to SC18, decreased cell proliferation as determined by wound-healing and cell viability assays. In summary, SC18 is a novel VDAC-targeting small molecule that induces mitochondrial dysfunction and inhibits cell proliferation.

Arousal from Torpor Increases Oxidative Damage in the Hibernating Thirteen-Lined Ground Squirrel (Ictidomys tridecemlineatus).

AbstractDuring hibernation, especially during arousal from torpor to interbout euthermia (IBE), blood flow changes drastically. In nonhibernating mammals, similar changes during ischemia/reperfusion lead to oxidative damage. We hypothesized that suppression of mitochondrial metabolism during hibernation protects against such damage. We compared markers of oxidative damage and total antioxidant capacity in eight tissues among summer, torpid, and IBE thirteen-lined ground squirrels. Overall, summer tissue had less lipid and protein oxidative damage than tissue from the hibernation season, but DNA damage (in four tissues) and total antioxidant capacity (in all eight tissues) were similar among all groups. During torpor, when mitochondrial metabolism is suppressed, lipid damage in heart, brown adipose tissue, and small intestine was lower than IBE by as much as fivefold. By contrast, oxidative damage to protein was at least twofold higher in liver and skeletal muscle in torpor compared with IBE. Our findings suggest that arousal from torpor creates oxidative damage similar to ischemia/reperfusion injury but that this damage is repaired during IBE. These differences cannot be explained by changes in antioxidant capacity, so they are likely due to differences is reactive oxygen species production among hibernation states that may relate to the well-characterized reversible suppression of mitochondrial metabolism during torpor.

CRISPR-Cas9 screen identifies oxidative phosphorylation as essential for cancer cell survival at low extracellular pH.

SummaryUnlike most cell types, many cancer cells survive at low extracellular pH (pHe), a chemical signature of tumors. Genes that facilitate survival under acid stress are therefore potential targets for cancer therapies. We performed a genome-wide CRISPR-Cas9 cell viability screen at physiological and acidic conditions to systematically identify gene knockouts associated with pH-related fitness defects in colorectal cancer cells. Knockouts of genes involved in oxidative phosphorylation (NDUFS1) and iron-sulfur cluster biogenesis (IBA57, NFU1) grew well at physiological pHe, but underwent profound cell death under acidic conditions. We identified several small-molecule inhibitors of mitochondrial metabolism that can kill cancer cells at low pHe only. Xenografts established from NDUFS1−/− cells grew considerably slower than their wild-type controls, but growth could be stimulated with systemic bicarbonate therapy that lessens the tumoral acid stress. These findings raise the possibility of therapeutically targeting mitochondrial metabolism in combination with acid stress as a cancer treatment option.

Small molecules targeting the VDAC-NADH binding pocket modulate mitochondrial metabolism in cancer cells

Enhanced glycolysis and partial suppression of mitochondrial metabolism characterize the Warburg phenotype of cancer cells (CC) and cancer stem cells (CSC). Voltage dependent anion channels (VDAC 1/2/3) in the outer mitochondrial membrane, regulate the flux of most metabolites entering and exiting mitochondria, including ATP. VDAC closure partially suppress mitochondrial metabolism in cancer cells. VDAC isoform specificity and lack of identified druggable sites, are major obstacles to develop VDAC-dependent “metabolic” cancer treatments.

Targeting Mitochondrial Metabolism and RNA Polymerase POLRMT to Overcome Multidrug Resistance in Cancer.

Clinically, the prognosis of tumor therapy is fundamentally affected by multidrug resistance (MDR), which is primarily a result of enhanced drug efflux mediated by channels in the membrane that reduce drug accumulation in tumor cells. How to restore the sensitivity of tumor cells to chemotherapy is an ongoing and pressing clinical issue. There is a prevailing view that tumor cells turn to glycolysis for energy supply due to hypoxia. However, studies have shown that mitochondria also play crucial roles, such as providing intermediates for biosynthesis through the tricarboxylic acid (TCA) cycle and a plenty of ATP to fuel cells through the complete breakdown of organic matter by oxidative phosphorylation (OXPHOS). High OXPHOS have been found in some tumors, particularly in cancer stem cells (CSCs), which possess increased mitochondria mass and may be depends on OXPHOS for energy supply. Therefore, they are sensitive to inhibitors of mitochondrial metabolism. In view of this, we should consider mitochondrial metabolism when developing drugs to overcome MDR, where mitochondrial RNA polymerase (POLRMT) would be the focus, as it is responsible for mitochondrial gene expression. Inhibition of POLRMT could disrupt mitochondrial metabolism at its source, causing an energy crisis and ultimately eradicating tumor cells. In addition, it may restore the energy supply of MDR cells to glycolysis and re-sensitize them to conventional chemotherapy. Furthermore, we discuss the rationale and strategies for designing new therapeutic molecules for MDR cancers by targeting POLRMT.

VDAC Modulation of Cancer Metabolism: Advances and Therapeutic Challenges.

Most anionic metabolites including respiratory substrates, glycolytic adenosine triphosphate (ATP), and small cations that enter mitochondria, and mitochondrial ATP moving to the cytosol, cross the outer mitochondrial membrane (OMM) through voltage dependent anion channels (VDAC). The closed states of VDAC block the passage of anionic metabolites, and increase the flux of small cations, including calcium. Consequently, physiological or pharmacological regulation of VDAC opening, by conditioning the magnitude of both anion and cation fluxes, is a major contributor to mitochondrial metabolism. Tumor cells display a pro-proliferative Warburg phenotype characterized by enhanced aerobic glycolysis in the presence of partial suppression of mitochondrial metabolism. The heterogeneous and flexible metabolic traits of most human tumors render cells able to adapt to the constantly changing energetic and biosynthetic demands by switching between predominantly glycolytic or oxidative phenotypes. Here, we describe the biological consequences of changes in the conformational state of VDAC for cancer metabolism, the mechanisms by which VDAC-openers promote cancer cell death, and the advantages of VDAC opening as a valuable pharmacological target. Particular emphasis is given to the endogenous regulation of VDAC by free tubulin and the effects of VDAC-tubulin antagonists in cancer cells. Because of its function and location, VDAC operates as a switch to turn-off mitochondrial metabolism (closed state) and increase aerobic glycolysis (pro-Warburg), or to turn-on mitochondrial metabolism (open state) and decrease glycolysis (anti-Warburg). A better understanding of the role of VDAC regulation in tumor progression is relevant both for cancer biology and for developing novel cancer chemotherapies.

Mitochondrial Metabolism Targeted Nanoplatform for Efficient Triple-Negative Breast Cancer Combination Therapy.

Tumor reprogram pathway of mitochondrial metabolism is an emerging approach for malignant tumor treatment, such as triple-negative breast cancer. In this study, a tumor/mitochondria cascaded targeting, adenosine-triphosphate (ATP) responsive nanocarrier of zeolitic imidazolate framework-90 (ZIF-90) for breast cancer combination therapy is reported. Atovaquone (AVO) and hemin are loaded into ZIF-90, then a peptide iRGD with tumor-targeting ability is modified on the ZIF-90 nanoplatform. Hemin can specifically degrade BTB and CNC homology1 (BACH1), resulting in the changes of mitochondrial metabolism, and AVO acts as the inhibitor of the electron transport chain (ETC). The degradation of BACH1 using hemin can effectively improve the anti-tumor efficiency of mitochondrial metabolism inhibitor AVO, by increasing dependency on mitochondrial respiration. This nanoplatform displays both tumor-targeting and mitochondria-targeting capacity with high level of ATP responsive drug release behavior. The specific characteristic of mitochondria-targeting ability of this nanoplatform can increase the accumulation of AVO in the mitochondria, and in turn, can effectively improve the inhibition of the ETC. Both in vitro and in vivo results reveal that this composite nanocarrier has excellent tumor inhibition ability with limited side effects. Accordingly, this study provides an attractive strategy in the mitochondrial metabolism for cancer targeted therapy.

Prostate cancer cells survive anti-androgen and mitochondrial metabolic inhibitors by modulating glycolysis and mitochondrial metabolic activities.

Most cancer cells are more glycolytic even under aerobic conditions compared with their normal counterparts. Recent evidence of tumor cell metabolism, however, shows that some tumors also increase mitochondrial oxidative phosphorylation (ox-phos) at some disease states during progression and/or development of drug resistance. Our data show that anti-androgen enzalutamide (ENZA) resistant prostate cancer (PCa) cells use more mitochondrial metabolism leading to higher ox-phos as compared to the ENZA-sensitive cells and can become vulnerable to mitochondrial metabolism targeted therapies. Seahorse assay, mass spectrometry and high resolution fluorescence confocal microscopy coupled with image analysis has been used to compare mitochondrial metabolism in ENZA-treated and -untreated anti-androgen-sensitive LNCaP and -resistant C4-2, CWR22ν1, and PCa2b cells. Ex vivo fluorescence microscopy and image analysis has been standardized to monitor mitochondrial electron transport (ETS) activity that likely increases ox-phos in circulating tumor cells (CTCs) isolated fom patients undergoing AR-targeted therapies. Our data show that PCa cells that are resistant to anti-androgen ENZA switch from glycolysis to ox-phos leading to an increased ETS activity. ENZA pretreated cells are more vulnerable to ETS component complex I inhibitor IACS-010759 (IACS) and mitochondrial glutaminase inhibitor CB-839 that reduces glutamate supply to tricarboxylic acid cycle. CTCs isolated from 6 of 20 patient blood samples showed relatively higher ETS activity than the rest of the patients. All six patients have developed ENZA resistance within less than 6 months of the sample collection. The enhanced growth inhibitory effects of mitochondrial metabolic inhibitors IACS and CB-839 in ENZA pretreated PCa cells provides a rationale for designing a drug combination trial. Patients can be selected for such trials by monitoring the mitochondrial ETS activities in their CTCs to maximize success.

Inhibition of oxidative metabolism by nitric oxide restricts EMCV replication selectively in pancreatic beta-cells

Environmental factors, such as viral infection, are proposed to play a role in the initiation of autoimmune diabetes. In response to encephalomyocarditis virus (EMCV) infection, resident islet macrophages release the pro-inflammatory cytokine IL-1β, to levels that are sufficient to stimulate inducible nitric oxide synthase (iNOS) expression and production of micromolar levels of the free radical nitric oxide in neighboring β-cells. We have recently shown that nitric oxide inhibits EMCV replication and EMCV-mediated β-cell lysis and that this protection is associated with an inhibition of mitochondrial oxidative metabolism. Here we show that the protective actions of nitric oxide against EMCV infection are selective for β-cells and associated with the metabolic coupling of glycolysis and mitochondrial oxidation that is necessary for insulin secretion. Inhibitors of mitochondrial respiration attenuate EMCV replication in β-cells, and this inhibition is associated with a decrease in ATP levels. In mouse embryonic fibroblasts (MEFs), inhibition of mitochondrial metabolism does not modify EMCV replication or decrease ATP levels. Like most cell types, MEFs have the capacity to uncouple the glycolytic utilization of glucose from mitochondrial respiration, allowing for the maintenance of ATP levels under conditions of impaired mitochondrial respiration. It is only when MEFs are forced to use mitochondrial oxidative metabolism for ATP generation that mitochondrial inhibitors attenuate viral replication. In a β-cell selective manner, these findings indicate that nitric oxide targets the same metabolic pathways necessary for glucose stimulated insulin secretion for protection from viral lysis.