Metformin Inhibits Mitochondrial Complex I To Promote Health

Abstract The major function of mitochondria in cellular homeostasis has been the generation of ATP through oxidative phosphorylation. However, we have previously demonstrated that mitochondria can serve as signaling organelles by releasing low levels of reactive oxygen species (ROS) and TCA cycle metabolites that are essential for hypoxic activation of HIF, antigen activation of T cells, cellular differentiation and proliferation of cancer cells. The anti-diabetic drug metformin has been proposed to inhibit mitochondrial complex I. We will present data indicating that metformin inhibits mitochondrial complex I to exert it’s biological effects through controlling ROS, ATP, and NAD+.

Download Full-text

Glutaminase Inhibition Overcomes Acquired Resistance to Mitochondrial Complex I in NOTCH1-Driven T-Cell Acute Lymphoblastic Leukemias (T-ALL) Via Block of Glutamine Driven Reductive Metabolism

Blood ◽

10.1182/blood-2019-128930 ◽

2019 ◽

Vol 134 (Supplement_1) ◽

pp. 806-806

Author(s):

Natalia Baran ◽

Alessia Lodi ◽

Shannon Renee Sweeney ◽

Vinitha Mary Kuruvilla ◽

Antonio Cavazos ◽

...

Keyword(s):

Cell Proliferation ◽

Dna Damage ◽

Complex I ◽

Research Funding ◽

Tca Cycle ◽

Mitochondrial Complex I ◽

Mitochondrial Complex ◽

The Tca Cycle ◽

Dual Blockade ◽

Reductive Metabolism

Notch1-mutated T-ALL is an aggressive hematologic malignancy lacking targeted therapeutic options. Genomic alterations in Notch1-gene and its activated downstream pathways are associated with metabolic stress response and heightened glutamine (Gln) utilization to fuel oxidative phosphorylation (OxPhos) (Kishton at al., Cell Metabolism 2016, 23:649, Herranz at al., Nat Med, 2015, 21(10): 1182-1189). Hence, targeting NOTCH1-associated OxPhos and/or Gln dependency could constitute a plausible therapeutic strategy for T-ALL. In this study we examined metabolic vulnerabilities of NOTCH1-driven T-ALL and tested pre-clinical efficacy of novel mitochondrial complex I (OxPhosi) IACS-010759 and of glutaminase inhibitor CB-839 (GLSi) in T-ALL models including Notch1-mutated T-ALL cell lines, patient-derived xenograft (PDX) and primary T-ALL cells. We have previously reported and confirmed in this expanded study the anti-leukemia efficacy of IACS-010759 (EC50s 0.1-15 nM) (Molina at al., Nat Med, 2018, 24: 1036; Baran at al., Blood, 2018, 132:4020). Metabolic characterization demonstrated that OxPhosi caused striking dose-dependent decrease in basal and maximal oxygen consumption rate (OCR), ATP and NADH generation in T-ALL cell lines and primary T-ALL samples (p<0.001). OxPhosi, similar to knockout of complex I subunit NDUFS4 using CRISPR-CAS9, induced profound changes in T-ALL mitochondria, with induction of mitochondrial reactive oxygen species (ROS), DNA damage, activation of AMPK and inhibition of mTOR pathway. OxPhosi altered cellular energy homeostasis by reduction of TCA cycle intermediates, glutathione and reduction of intracellular nucleotides ATP, CTP, GTP, and UTP, translating into inhibition of DNA and RNA synthesis (p<0.0001) (by UPLC-MS/MS). IACS-010759 significantly reduced the glucose flux through the TCA cycle, redirected it towards lactate production and triggered increased utilization of Gln for fueling of the TCA cycle and reductive metabolism (Fig.1, Flux metabolic analysis SIRM). In concert with these findings, supplementation with Gln partially rescued growth-inhibitory effects of OxPhos inhibition. These results uncover metabolic gap used by T-ALL to escape OxPhos block, and identifying reliance on glutaminolysis as a critical therapeutic target. To confirm that blockade of Gln entry into TCA cycle with GLSi synergistically reduced viable ALL cell numbers, we studied potential synergy of OxPhosi and GLSi. The key role of Gln in maintaining energy production and cell proliferation via OxPhos in Notch1-mutated T-ALL cells was confirmed by the findings that Gln starvation or pharmacological GLS inhibition by CB-839 reduced ATP production and OCR and decreased cell proliferation by more than 50% in vitro (Fig.2, Fig.3). Dual blockade of OxPhos together with GLS induced DNA damage response via accumulation of ROS upon glutathione deprivation, induced AMPK signaling through profound reduction of all adenosine related intermediates and inhibited mTOR signaling. This translated into significant reduction of leukemia burden and extension of overall survival in vivo (p<0.0001) in Notch1-mutated T-ALL PDX models (n=2) with IACS-010759/CB-839 co-treatment and in the Notch1-mutated GLS fl/fl murine model upon tamoxifen-induced GLS knockout (p<0.0001) (Fig.4, Fig.5). In summary, our findings indicate that dual blockade of metabolic processes by inhibiting complex I of mitochondria and restricting Gln utilization results in metabolic catastrophe in Notch1-mutated T-ALL associated with energy depletion and oxidative stress, which combined severely inhibit T-ALL growth and survival. We postulate that targeting this unique metabolic vulnerability of Notch1-mutated T-ALL cells constitutes a novel therapeutic modality in this aggressive malignancy. Disclosures Kuruvilla: The University of Texas M.D.Anderson Cancer Center: Employment. Jabbour:AbbVie: Consultancy, Research Funding; Pfizer: Consultancy, Research Funding; Cyclacel LTD: Research Funding; Takeda: Consultancy, Research Funding; BMS: Consultancy, Research Funding; Adaptive: Consultancy, Research Funding; Amgen: Consultancy, Research Funding. Konopleva:Agios: Research Funding; Stemline Therapeutics: Consultancy, Honoraria, Research Funding; Calithera: Research Funding; Astra Zeneca: Research Funding; Kisoji: Consultancy, Honoraria; Ascentage: Research Funding; Genentech: Honoraria, Research Funding; F. Hoffman La-Roche: Consultancy, Honoraria, Research Funding; Amgen: Consultancy, Honoraria; Eli Lilly: Research Funding; Cellectis: Research Funding; Forty-Seven: Consultancy, Honoraria; Ablynx: Research Funding; Reata Pharmaceuticals: Equity Ownership, Patents & Royalties; AbbVie: Consultancy, Honoraria, Research Funding.

Download Full-text

Mitochondrial Complex I Inhibitor Iacs-010759 Reverses the NOTCH1-Driven Metabolic Reprogramming in T-ALL Via Blockade of Oxidative Phosphorylation: Synergy with Chemotherapy and Glutaminase Inhibition

Blood ◽

10.1182/blood-2018-99-117310 ◽

2018 ◽

Vol 132 (Supplement 1) ◽

pp. 4020-4020 ◽

Cited By ~ 1

Author(s):

Natalia Baran ◽

Alessia Lodi ◽

Shannon Renee Sweeney ◽

Pandey Renu ◽

Vinitha Mary Kuruvilla ◽

...

Keyword(s):

Overall Survival ◽

Oxidative Phosphorylation ◽

Complex I ◽

Cell Metabolism ◽

Tca Cycle ◽

Tumor Burden ◽

Mitochondrial Complex I ◽

Mitochondrial Complex ◽

The Tca Cycle

Abstract Adult T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive hematologic malignancy characterized by limited therapeutic options and a high rate of treatment failure due to chemoresistance. T-ALL is largely driven by activating NOTCH1 mutations, where oncogenic NOTCH1 facilitates glutamine oxidation, induces metabolic stress, and facilitates reliance on oxidative phosphorylation (OXPHOS)1. In other malignancies, the shift toward OXPHOS-dependent high-energy status is associated with acquired chemoresistance. In this study, we found that the novel inhibitor of mitochondrial complex I (OXPHOSi) IACS-0107592 has preclinical activity in NOTCH1-mutated T-ALL; we also characterize the cellular and metabolic responses to OXPHOS inhibition and propose that an OXPHOSi be incorporated into standard-of-care therapy to improve outcomes in patients harboring NOTCH1-mutated T-ALL. Exposure to IACS-010759 (0-370 nM) in vitro drastically reduced T-ALL viability, with EC50 ranging from 0.1-10 nM for cell lines (n=7) and from 13-60 nM for patient-derived xenograft (PDX)-derived and primary T-ALL cells (n=10) (Fig.1). Oral administration of IACS-010759 (7.5 mg/kg/day) significantly reduced leukemia burden and extended overall survival (p<0.0001) in two aggressive NOTCH1-mutated T-ALL PDX models and in a murine NOTCH1-driven T-ALL model (Fig.4). Addition of OXPHOS inhibitor to dexamethasone (X), vincristine (V), asparaginase (L), or a combination (VXL) led to additive/synergistic inhibition of cell proliferation in vitro and to doubling of overall survival in vivo (p<0.0001) (Fig.4). Metabolic characterization confirmed that IACS-010759 caused striking dose-dependent decreases in basal and maximal oxygen consumption rates (OCR) and ATP and NADH production in T-ALL cell lines and primary T-ALL samples (p<0.001; Fig.2). Further, pretreatment with V, X, or L shifted T-ALL cell metabolism toward OXPHOS, increasing significantly the OCR that was effectively inhibited by IACS-010759. Pharmacological inhibition of complex I with IACS-010759, similar to knockout of complex I subunit NDUFS4 using CRISPR-CAS9, induced catastrophic changes in mitochondria, with induction of mitochondrial reactive oxygen species (ROS), DNA damage, and activation of the compensatory mTOR pathway. OXPHOS inhibition altered cellular energy homeostasis through reduction of TCA cycle intermediates; decreased glutathione level (by UPLC-MS/MS; p<0.0001) with ROS induction (Fig.3); and depleted the pool of intracellular nucleotides, affecting DNA and RNA synthesis (Fig.2C). Stable isotope-resolved metabolomics (SIRM) flux analysis showed that IACS-010759 (30 nM at 24 h) significantly decreased the flux of glucose through the TCA cycle and redirected it toward lactate production and increased utilization of glutamine for fueling the TCA cycle, in particular through reductive metabolism, uncovering reliance on glutaminolysis as an additional therapeutic target. Consistent with this was the finding that combined OXPHOSi with glutaminase inhibitor CB-839 caused additive reduction of viability of T-ALL cells lines and primary T-ALL cells in vitro (Fig.2), decreased tumor burden (p<0.02), and increased survival in a T-ALL PDX model (p<0.01). This was supported by IACS-induced reduction of tumor burden in a NOTCH1-mutated GLS fl/fl murine model upon tamoxifen-induced GLS knockout (p<0.01). In summary, our findings indicate that OXPHOSi, alone and particularly in combination with standard chemotherapy and GLS inhibition, constitutes a novel therapeutic modality that targets a unique metabolic vulnerability of NOTCH1-mutated T-ALL cells. References:Kishton RJ, Barnes CE, Nichols AG at al., AMPK Is Essential to Balance Glycolysis and Mitochondrial Metabolism to Control T-ALL Cell Stress and Survival, Cell Metabolism, 2016, 23(4):649-62Molina JR, Sun Y, Protopopova M et al., An inhibitor of oxidative phosphorylation exploits cancer vulnerability, Nat Med, 2018, 24: 1036-1046 Disclosures Lorenzi: NIH: Patents & Royalties; Erytech Pharma: Consultancy. Konopleva:Stemline Therapeutics: Research Funding.

Download Full-text