title: Reduced Warburg effect in cancer cells undergoing autophagy: steady- state 1H-MRS and real-time hyperpolarized 13C-MRS studies.
abstract: Autophagy is a highly regulated, energy dependent cellular process where proteins, organelles and cytoplasm are sequestered in autophagosomes and digested to sustain cellular homeostasis. We hypothesized that during autophagy induced in cancer cells by i) starvation through serum and amino acid deprivation or ii) treatment with PI-103, a class I PI3K/mTOR inhibitor, glycolytic metabolism would be affected, reducing flux to lactate, and that this effect may be reversible. We probed metabolism during autophagy in colorectal HT29 and HCT116 Bax knock-out cells using hyperpolarized (13)C-magnetic resonance spectroscopy (MRS) and steady-state (1)H-MRS. 24 hr PI103-treatment or starvation caused significant reduction in the apparent forward rate constant (k(PL)) for pyruvate to lactate exchange compared with controls in HT29 (100 μM PI-103: 82%, p = 0.05) and HCT116 Bax-ko cells (10 μM PI-103: 53%, p = 0.05; 20 μM PI-103: 42%, p<0.0001; starvation: 52%, p<0.001), associated with reduced lactate excretion and intracellular lactate in all cases, and unchanged lactate dehydrogenase (LDH) activity and increased NAD+/NADH ratio following PI103 treatment or decreased LDH activity and unchanged NAD+/NADH ratio following starvation. After 48 hr recovery from PI103 treatment, k(PL) remained below control levels in HT29 cells (74%, p = 0.02), and increased above treated values, but remained below 24 hr vehicle-treated control levels in HCT116 Bax-ko cells (65%, p = 0.004) both were accompanied by sustained reduction in lactate excretion, recovery of NAD+/NADH ratio and intracellular lactate. Following recovery from starvation, k(PL) was significantly higher than 24 hr vehicle-treated controls (140%, p = 0.05), associated with increased LDH activity and total cellular NAD(H). Changes in k(PL) and cellular and excreted lactate provided measureable indicators of the major metabolic processes accompanying starvation- and drug-induced autophagy. The changes are reversible, returning towards and exceeding control values on cellular recovery, which potentially identifies resistance. k(PL) (hyperpolarized (13)C-MRS) and lactate ((1)H-MRS) provide useful biomarkers for the autophagic process, enabling non-invasive monitoring of the Warburg effect.
authors.affiliation: Cancer Research UK and EPSRC Cancer Imaging Centre, Division of Radiotherapy and Imaging, The Institute of Cancer Research and Royal Marsden Hospital, Sutton, Surrey, United Kingdom; Department of Medical Imaging and Intervention, Chang Gung Memorial Hospital at Linkou, College of Medicine, Chang Gung University, Taoyuan, Taiwan.
authors.forename: Gigin
authors.initials: G
authors.lastname: Lin
authors.affiliation: Cancer Research UK and EPSRC Cancer Imaging Centre, Division of Radiotherapy and Imaging, The Institute of Cancer Research and Royal Marsden Hospital, Sutton, Surrey, United Kingdom.
authors.forename: Gabriela
authors.initials: G
authors.lastname: Andrejeva
authors.affiliation: Cancer Research UK and EPSRC Cancer Imaging Centre, Division of Radiotherapy and Imaging, The Institute of Cancer Research and Royal Marsden Hospital, Sutton, Surrey, United Kingdom.
authors.forename: Anne-Christine
authors.initials: AC
authors.lastname: Wong Te Fong
authors.affiliation: Cancer Research UK and EPSRC Cancer Imaging Centre, Division of Radiotherapy and Imaging, The Institute of Cancer Research and Royal Marsden Hospital, Sutton, Surrey, United Kingdom.
authors.forename: Deborah K
authors.initials: DK
authors.lastname: Hill
authors.affiliation: Cancer Research UK and EPSRC Cancer Imaging Centre, Division of Radiotherapy and Imaging, The Institute of Cancer Research and Royal Marsden Hospital, Sutton, Surrey, United Kingdom.
authors.forename: Matthew R
authors.initials: MR
authors.lastname: Orton
authors.affiliation: Cancer Research UK and EPSRC Cancer Imaging Centre, Division of Radiotherapy and Imaging, The Institute of Cancer Research and Royal Marsden Hospital, Sutton, Surrey, United Kingdom.
authors.forename: Harry G
authors.initials: HG
authors.lastname: Parkes
authors.affiliation: Cancer Research UK and EPSRC Cancer Imaging Centre, Division of Radiotherapy and Imaging, The Institute of Cancer Research and Royal Marsden Hospital, Sutton, Surrey, United Kingdom.
authors.forename: Dow-Mu
authors.initials: DM
authors.lastname: Koh
authors.affiliation: Cancer Research UK and EPSRC Cancer Imaging Centre, Division of Radiotherapy and Imaging, The Institute of Cancer Research and Royal Marsden Hospital, Sutton, Surrey, United Kingdom.
authors.forename: Simon P
authors.initials: SP
authors.lastname: Robinson
authors.affiliation: Cancer Research UK and EPSRC Cancer Imaging Centre, Division of Radiotherapy and Imaging, The Institute of Cancer Research and Royal Marsden Hospital, Sutton, Surrey, United Kingdom.
authors.forename: Martin O
authors.initials: MO
authors.lastname: Leach
authors.affiliation: Cancer Research UK and EPSRC Cancer Imaging Centre, Division of Radiotherapy and Imaging, The Institute of Cancer Research and Royal Marsden Hospital, Sutton, Surrey, United Kingdom; Division of Imaging Sciences and Biomedical Engineering, King's College London, The Rayne Institute, St Thomas Hospital, London, United Kingdom.
authors.forename: Thomas R
authors.initials: TR
authors.lastname: Eykyn
authors.affiliation: Cancer Research UK and EPSRC Cancer Imaging Centre, Division of Radiotherapy and Imaging, The Institute of Cancer Research and Royal Marsden Hospital, Sutton, Surrey, United Kingdom.
authors.forename: Yuen-Li
authors.initials: YL
authors.lastname: Chung
mesh_terms: D001343:Autophagy; D002247:Carbon Isotopes; D045744:Cell Line, Tumor; D005663:Furans / Q000494:pharmacology; D055785:Gene Knockdown Techniques; D006801:Humans; D007553:Isotope Labeling; D007770:L-Lactate Dehydrogenase / Q000235:genetics / Q000378:metabolism; D019344:Lactic Acid / Q000378:metabolism*; D009243:NAD / Q000235:genetics / Q000378:metabolism; D009369:Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D011725:Pyridines / Q000494:pharmacology; D011743:Pyrimidines / Q000494:pharmacology; D051028:bcl-2-Associated X Protein / Q000235:genetics / Q000378:metabolism
chemical_list: C494826:BAX protein, human; D002247:Carbon Isotopes; D005663:Furans; C522973:PI103; D011725:Pyridines; D011743:Pyrimidines; D051028:bcl-2-Associated X Protein; D009243:NAD; D019344:Lactic Acid; D007770:L-Lactate Dehydrogenase