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The mitochondrial electron transport chain (ETC) is necessary for tumour growth1–6 and its inhibition has demonstrated anti-tumour efficacy in combination with targeted therapies7–9. Furthermore, human brain and lung tumours display robust glucose oxidation by mitochondria10,11. However, it is unclear why a functional ETC is necessary for tumour growth in vivo. ETC function is coupled to the generation of ATP—that is, oxidative phosphorylation and the production of metabolites by the tricarboxylic acid (TCA) cycle. Mitochondrial complexes I and II donate electrons to ubiquinone, resulting in the generation of ubiquinol and the regeneration of the NAD+ and FAD cofactors, and complex III oxidizes ubiquinol back to ubiquinone, which also serves as an electron acceptor for dihydroorotate dehydrogenase (DHODH)—an enzyme necessary for de novo pyrimidine synthesis. Here we show impaired tumour growth in cancer cells that lack mitochondrial complex III. This phenotype was rescued by ectopic expression of Ciona intestinalis alternative oxidase (AOX)12, which also oxidizes ubiquinol to ubiquinone. Loss of mitochondrial complex I, II or DHODH diminished the tumour growth of AOX-expressing cancer cells deficient in mitochondrial complex III, which highlights the necessity of ubiquinone as an electron acceptor for tumour growth. Cancer cells that lack mitochondrial complex III but can regenerate NAD+ by expression of the NADH oxidase from Lactobacillus brevis (LbNOX)13 targeted to the mitochondria or cytosol were still unable to grow tumours. This suggests that regeneration of NAD+ is not sufficient to drive tumour growth in vivo. Collectively, our findings indicate that tumour growth requires the ETC to oxidize ubiquinol, which is essential to drive the oxidative TCA cycle and DHODH activity. Oxidation of ubiquinol by the mitochondrial electron transfer chain drives tumour growth by maintaining the function of the oxidative Krebs cycle and de novo pyrimidine synthesis.

Mitochondrial complex I inhibition in the presence of a mito-antioxidant induces reductive stress, ISR, and cancer cell death., Mitochondria-derived reactive oxygen species (mROS) are required for the survival, proliferation, and metastasis of cancer cells. The mechanism by which mitochondrial metabolism regulates mROS levels to support cancer cells is not fully understood. To address this, we conducted a metabolism-focused CRISPR-Cas9 genetic screen and uncovered that loss of genes encoding subunits of mitochondrial complex I was deleterious in the presence of the mitochondria-targeted antioxidant mito-vitamin E (MVE). Genetic or pharmacologic inhibition of mitochondrial complex I in combination with the mitochondria-targeted antioxidants, MVE or MitoTEMPO, induced a robust integrated stress response (ISR) and markedly diminished cell survival and proliferation in vitro. This was not observed following inhibition of mitochondrial complex III. Administration of MitoTEMPO in combination with the mitochondrial complex I inhibitor phenformin decreased the leukemic burden in a mouse model of T cell acute lymphoblastic leukemia. Thus, mitochondrial complex I is a dominant metabolic determinant of mROS-dependent cellular fitness.

Purpose/Objective(s) Primary tumor failure is common in patients treated with chemoradiation (CRT) for unresectable LA-NSCLC. Stereotactic body radiation therapy (SBRT) yields high rates of primary tumor control in Stage I NSCLC. This trial tested an SBRT boost to the primary tumor prior to the start of CRT to improve primary tumor control (PTC). Materials/Methods Patients with LA-NSCLC with primary tumor ≤10 cm were enrolled on a prospective phase II trial testing an SBRT boost in 2 fractions (central 6 Gy x 2, peripheral 8 Gy x 2) immediately followed by standard concurrent CRT (60 Gy in 30 fractions). Patients were staged with PET-CT, brain MRI, and underwent 4D-CT simulation for radiation planning using a customized immobilization device that enabled radiation planning for the sequential delivery of the SBRT boost and conventionally-fractionated radiation from the same CT dataset. Chemotherapy was carboplatin/paclitaxel (C/P) or cisplatin/etoposide. For patients receiving C/P, consolidation C/P for 2 cycles was given at the discretion of the medical oncologist. The trial was later amended to allow for consolidation durvalumab. The primary objective was to estimate PTC rate at 1-year with a hypothesis that the 1-year PTC rate is ≥90%. Secondary objectives were to establish the safety, feasibility, objective response rate (ORR; RECISTv1.1), and rates of regional & distant control, disease-free survival (DFS), and overall survival (OS). Exploratory functional MRI (fMRI) scans before and within the first 30 hrs of the first SBRT fraction were performed in 11 patients. Results The study enrolled 21 patients (10 male, 11 female), with median age 62 years (range 52-78). 16 patients received 6 Gy x 2 SBRT boost, while 5 patients received 8 Gy x 2 SBRT boost. 18 patients received C/P chemotherapy, of whom 9 patients received consolidation C/P. Six patients received consolidation durvalumab. Median pre-treatment primary tumor size was 5.0 cm (range 1.0-8.3). Median and mean follow-up were17.9 and 20.2 months, respectively. The most common toxicities were fatigue, neutropenia, leukopenia, lymphopenia, anemia, pneumonitis, fibrosis, dyspnea and esophagitis. Five grade 4 toxicities related to treatment occurred [lymphopenia (3), neutropenia (1), and leukopenia (1)], but no grade 5 toxicities related to treatment occurred. ORR at 3 and 6 months was 72.7% and 80.0%. The PTC rate was 100% and 92.3% at 1 and 2 years, respectively. The 2-year regional and distant control were 81.6% and 70.3%, respectively. Disease-free survival and overall survival at 2 yrs were 46.1% and 50.3%, respectively. Median survival was 37.8 months. fMRI detected a mean relative decrease in BOLD signal of -87.1% (P = 0.05). Conclusion Dose escalation to the primary tumor utilizing upfront SBRT appears feasible and safe. PTC was high and other oncologic endpoints compared favorably with the literature. Through the use of 1 CT simulation dataset, accurate calculation of the planned dose through the 2 sequentially-delivered plans is achievable.

There are numerous mechanisms by which mammals respond to hypoxia. These include acute changes in pulmonary arterial tone due to smooth muscle cell contraction, acute increases in respiration triggered by the carotid body chemosensory cells, and chronic changes such as induction of red blood cell proliferation and angiogenesis by hypoxia inducible factor targets erythropoietin and vascular endothelial growth factor, respectively. Mitochondria account for the majority of oxygen consumption in the cell and have recently been appreciated to serve as signaling organelles required for the initiation or propagation of numerous homeostatic mechanisms. Mitochondria can influence cell signaling by production of reactive oxygen species and metabolites. Here we review recent evidence that mitochondrial signals can imitate acute and chronic hypoxia responses.

Thiocyanate (SCN) is used by the innate immune system, but less is known about its impact on inflammation and oxidative stress. Granulocytes oxidize SCN to evolve the bactericidal hypothiocyanous acid, which we previously demonstrated is metabolized by mammalian, but not bacterial, thioredoxin reductase (TrxR). There is also evidence that SCN is dysregulated in cystic fibrosis (CF), a disease marked by chronic infection and airway inflammation. To investigate antiinflammatory effects of SCN, we administered nebulized SCN or saline to β epithelial sodium channel (βENaC) mice, a phenotypic CF model. SCN significantly decreased airway neutrophil infiltrate and restored the redox ratio of glutathione in lung tissue and airway epithelial lining fluid to levels comparable to wild type. Furthermore, in Pseudomonas aeruginosa–infected βENaC and wild-type mice, SCN decreased inflammation, proinflammatory cytokines, and bacterial load. SCN also decreased airway neutrophil chemokine keratinocyte chemoattractant (also known as C-X-C motif chemokine ligand 1) and glutathione sulfonamide, a biomarker of granulocyte oxidative activity, in uninfected βENaC mice. Lung tissue TrxR activity and expression increased in inflamed lung tissue, providing in vivo evidence for the link between hypothiocyanous acid metabolism by TrxR and the promotion of selective biocide of pathogens. SCN treatment both suppressed inflammation and improved host defense, suggesting that nebulized SCN may have important therapeutic utility in diseases of both chronic airway inflammation and persistent bacterial infection, such as CF.

Carotenoids serve antioxidant functions in plant photosynthetic processes, as well as in actions of disease reduction in mammalian systems. Lutein and zeaxanthin are important dietary carotenoids in suppressing aging eye diseases. Age-related macular degeneration now affects more than 1.75 million individuals in the US. Sweet corn (Zea mays var. rugosa) is one of only a few vegetable sources high in zeaxanthin. Mesotrione herbicide is currently labeled for selective pre- and post-emergence weed control in sweet corn production. Mesotrione competitively inhibits phytoene desaturase, a critical enzyme in carotenoid biosynthesis, which results in bleaching of leaf tissues in susceptible species and eventual plant death. Sweet corn is tolerant to mesotrione applications; however, differing sensitivity exists among genotypes. What remains unclear is the impact of mesotrione on carotenoid concentrations in mature sweet corn kernels following post-emergent applications to young corn plants. Our research objective was to measure mature kernel carotenoid concentrations in response to post-emergence applications of mesotrione to genotypes of different sensitivities. Post-emergence treatments included mesotrione applied alone, or in mixtures with the photosystem II inhibitor atrazine applied to corn plants of either 5-10 or 15-20 cm in height. Kernels were harvested from mature plants and measured for carotenoid concentrations. Mesotrione applied alone, or in mixtures with atrazine acted to increase concentrations of kernel antheraxanthin, lutein, and zeaxanthin in several sweet corn genotypes. Mesotrione applications resulted in greater pools of kernel carotenoids once the sweet corn genotypes overcame initial herbicidal photo-oxidative stress. This is the first report of herbicides directly up regulating a key biochemical pathway linked to the nutritional quality of a vegetable crop.

Sulfur mustard (bis 2-chloroethyl ethyl sulfide, SM) is a powerful bi-functional vesicating chemical warfare agent. SM tissue injury is partially mediated by the overproduction of reactive oxygen species resulting in oxidative stress. We hypothesized that using a catalytic antioxidant (AEOL 10150) to alleviate oxidative stress and secondary inflammation following exposure to SM would attenuate the toxic effects of SM inhalation. Adult male rats were intubated and exposed to SM (1.4 mg/kg), a dose that produces an LD50 at approximately 24 h. Rats were randomized and treated via subcutaneous injection with either sterile PBS or AEOL 10150 (5 mg/kg, sc, every 4 h) beginning 1 h post-SM exposure. Rats were euthanized between 6 and 48 h after exposure to SM and survival and markers of injury were determined. Catalytic antioxidant treatment improved survival after SM inhalation in a dose-dependent manner, up to 52% over SM PBS at 48 h post-exposure. This improvement was sustained for at least 72 h after SM exposure when treatments were stopped after 48 h. Non-invasive monitoring throughout the duration of the studies also revealed blood oxygen saturations were improved by 10% and clinical scores were reduced by 57% after SM exposure in the catalytic antioxidant treatment group. Tissue analysis showed catalytic antioxidant therapy was able to decrease airway cast formation by 69% at 48 h post-exposure. To investigate antioxidant induced changes at the peak of injury, several biomarkers of oxidative stress and inflammation were evaluated at 24 h post-exposure. AEOL 10150 attenuated SM-mediated lung lipid oxidation, nitrosative stress and many proinflammatory cytokines. The findings indicate that catalytic antioxidants may be useful medical countermeasure against inhaled SM exposure.

Amicarbazone is a photosystem II (PSII)-inhibiting herbicide in the triazolinone family, which is similar in mode of action to the triazines. Annual bluegrass is a cool-season weed and has shown resistance to some PSII-inhibiting herbicides. The objective was to evaluate triazine-resistant and -susceptible annual bluegrass populations for potential cross-resistance to amicarbazone. Two triazine-resistant (MS-01, MS-02) and triazine-susceptible (AL-01, COM-01) annual bluegrass populations were treated with amicarbazone, atrazine, and simazine at 0.26, 1.7, and 1.7 kg ai ha−1, respectively. All herbicide treatments controlled the susceptible populations greater than 94% 2 wk after treatment (WAT). No visual injury of MS-01 and MS-02 was observed at any time following herbicide treatment. Quantum yield (ΦPSII) of annual bluegrass was measured 0 to 72 h after application (HAA) to determine the photochemical effects of amicarbazone compared to other PSII inhibitors. ΦPSIIof triazine-susceptible populations was reduced at all measurement times by all three herbicides. However, amicarbazone decreased ΦPSIIof susceptible populations faster and greater than atrazine and simazine at most measurement times. Amicarbazone did not reduce ΦPSIIof the MS-01 population. Amicarbazone significantly reduced ΦPSIIof the MS-02 population during several measurement timings; however, these reductions were short-lived compared to the susceptible populations and no trend in ΦPSIIreduction was observed. Sequencing of thepsbAgene revealed a Ser to Gly substitution at amino acid position 264 known to confer resistance to triazine herbicides. These data indicate amicarbazone efficiently inhibited PSII of susceptible annual bluegrass populations; however, triazine-resistant annual bluegrass populations with Ser264to Gly mutations are cross-resistant to amicarbazone.

Amicarbazone is a photosystem II (PSII) inhibiting herbicide of the triazolinone herbicide family. Greenhouse experiments were conducted to compare the effects of amicarbazone and atrazine on annual bluegrass control and quantum yield (ΦPSII) when applied at three treatment placements (soil-only, foliage-only, and foliage + soil). Herbicide rates for amicarbazone and atrazine were 0.53 and 2.25 kg ha−1, respectively. Amicarbazone applied soil-only and foliage + soil controlled annual bluegrass 57 and 59%, respectively, 1 wk after treatment (WAT). Atrazine applied to foliage + soil controlled annual bluegrass 48% at 1 WAT. All soil-only and foliage + soil treatments were similar 2 WAT. Foliage-only application of amicarbazone provided significantly less control than other amicarbazone treatments throughout the study. Amicarbazone applied soil-only and foliage + soil controlled annual bluegrass 100% at 3 WAT. Soil-only and foliage + soil applications of atrazine and amicarbazone had similar reductions in quantum yield (ΦPSII) at 1 to 3 WAT. Foliar-applied amicarbazone reduced ΦPSII 78, 84, and 86% at 1, 2, and 3 WAT, respectively. The rapid reduction in annual bluegrass ΦPSII and the increase in control resulting from soil contact of amicarbazone indicate root exposure of amicarbazone is beneficial for annual bluegrass control.

Carotenoids are important photoprotectant and light-harvesting pigments within the photosynthetic apparatus. Little information is available regarding carotenoid physiology in creeping bentgrass (Agrostis stolonifera L.). Research was conducted to investigate relative high and low irradiance adaptation of creeping bentgrass with respect to β-carotene and xanthophyll composition. 'Crenshaw' creeping bentgrass plants were acclimated for 7 d to relative high [47.9 mol m -2 d -1 photosynthetically active radation (PAR)] or low irradiance (4.7 mol m 2 d -1 PAR). After the acclimation period, plants were transferred from high to low (low irradiance treatment) and low to high (high irradiance treatment) irradiance. Clippings were harvested at 0, 24, 72, and 168 h after the acclimation period. Zeaxanthin and antheraxanthin decreased from 5.1 and 3.4 to 0.9 and 0.6 mg 100 g -1 fresh weight (FW), respectively, over 168 h in low irradiance. As the turf adapted to low irradiance, violaxanthin, lutein, and lutein-5,6-epoxide (epoxylutein) increased at 24 h, but levels decreased from 24 to 168 h. Zeaxanthin and antheraxanthin increased in high irradiance, while violaxanthin and β-carotene decreased. Lutein was the predominant carotenoid quantified regardless of irradiance treatment. Cumulative zeaxanthin, antheraxanthin, and violaxanthin increased as a percentage of the total carotenoids as the turfgrass adapted to high irradiance, but decreased in low irradiance. Conversely, neoxanthin and β-carotene decreased in high irradiance and increased in low irradiance. Creeping bentgrass produces carotenoid amounts comparable to other plant species potentially attributable to selection efforts of more stress-tolerant varieties.