Melanoma cell lines A375 (BRAFV600E mutation), MeWo (BRAF/NRAS wild-type), SK-MEL-28 (BRAFV600E mutation) and SK-MEL-2 (NRASQ61R mutation) were acquired from the American Type Culture Collection (ATCC, Manassas, VA, USA). Cells were cultured as monolayers according to the supplier’s instructions in RPMI-1640 medium containing 11 mM glucose (PAN Biotech #17500, Aidenbach, Germany) with 3 mM L-glutamine (ThermoFisher Scientific #25030024, Waltham, MA, USA) and 10% FBS for a maximum of 50 passages. Sodium dichloroacetate (DCA) was purchased from Sigma-Aldrich (#347795, St. Louis, MO, USA) and AZD7545 from SelleckChem (#S7517, Houston, TX, USA). DCA was dissolved in either PBS or medium (1M stock concentration) and used in concentrations between 0–50 mM. AZD7545 was dissolved in DMSO (either 10 or 100 mM stock concentration), keeping the DMSO concentration at a maximum of 0.1%, and used in concentrations between 0–100 μM. The following antibodies were used: PDH (Cell Signaling, cat.nr. 3205, Danvers, MA, USA), pPDHSer293 (Calbiochem, AP1062, San Diego, CA, USA), pDPHSer232 (Calbiochem, AP1063, San Diego, CA, USA), pPDHSer300 (Calbiochem, AP1064, San Diego, CA, USA), PDK1 (Cell Signaling #3820, Danvers, MA, USA), PDK2 (Novus Biological, NBP2-75611, Englewood, CO, USA) and HSP90 (Santa Cruz, sc-13119, Dallas, TX, USA).
Sodium dichloroacetate dca
Manufactured by Merck Group
Sourced in United States
Sodium dichloroacetate (DCA) is a chemical compound used in various laboratory applications. It functions as a water-soluble sodium salt. DCA is often utilized in research and analytical settings, but its specific applications should be determined by the user based on their research needs and objectives.
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Lab products found in correlation
Oligomycin, by Santa Cruz Biotechnology (2 mentions)
Antimycin, by Santa Cruz Biotechnology (2 mentions)
Adenosine diphosphate adp, by Merck Group (2 mentions)
Rotenone, by Santa Cruz Biotechnology (2 mentions)
6 diazo 5 oxo l norleucine don, by Merck Group (2 mentions)
Doxycycline dox, by Merck Group (2 mentions)
Fluoro carbonyl cyanide phenylhydrazone (fccp), by Santa Cruz Biotechnology (2 mentions)
Sodium l lactate, by Merck Group (2 mentions)
Rpmi 1640, by Thermo Fisher Scientific (2 mentions)
Penicillin streptomycin, by Thermo Fisher Scientific (2 mentions)
Hsp90, by Santa Cruz Biotechnology (1 mentions)
Azd7545, by Selleck Chemicals (1 mentions)
Sk mel 28, by American Type Culture Collection (1 mentions)
Sk mel 2, by American Type Culture Collection (1 mentions)
Phgdh inactive, by Cayman Chemical (1 mentions)
Cb 839, by Focus Biomolecules (1 mentions)
Hct116 ccl 247, by American Type Culture Collection (1 mentions)
Fetal bovine serum (fbs), by Harvard Bioscience (1 mentions)
5 6 dichlorobenzimidazole 1 β d ribofuranoside, by Merck Group (1 mentions)
Mc coy s, by Merck Group (1 mentions)
12 protocols using sodium dichloroacetate dca
1
Investigating Metabolic Regulation in Melanoma
2
Metabolic Modulation in Cancer Research
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The following drugs were used: 6‐Diazo‐5‐oxo‐l ‐norleucine (Don, D2141, Sigma‐Aldrich); Sodium dichloroacetate (DCA, 347795, Sigma‐Aldrich); NCT‐502 and PHGDH inactive (19716 and 19717, Cayman); Doxycycline (Dox, D9891, Sigma‐Aldrich), Oligomycin (sc‐203342, Santa Cruz Biotechnology); FCCP (sc‐203578, Santa Cruz Biotechnology); Antimycin (sc‐202467, Santa Cruz Biotechnology); Rotenone (sc‐203242, Santa Cruz Biotechnology); Cyt.C (C2037, Sigma‐Aldrich) CB‐839 (10‐4556, Focus Biomolecules); Adenosine diphosphate (ADP, A2754, Sigma‐Aldrich).
3
Colon Carcinoma Cell Line Cultures
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Cell culture material was from Becton Dickinson Labware (Basel, Switzerland) and Corning (Bodenheim, Germany). Dulbecco's Modified Eagle's Medium (DMEM) and Mc Coy's were purchased from Sigma Chemicals (Buchs, Switzerland). Fetal bovine serum (FBS) was from Biochrom AG (Berlin, Germany). Insulin, cycloheximide (CHX), PD098059, Sodium Dichloroacetate (DCA), Sodium L-lactate and 5,6-dichlorobenzimidazole 1β-D-ribofuranoside (DRB) were purchased from Sigma-Aldrich (Fluka AG, Buchs, Switzerland). The following cell lines were purchased from ATCC (Manassas, VA): human colon carcinoma cell line HT-29 (accession number: HTB-38), SW-620 (CLL-227), HCT116 (CCL-247) and JEG-3 (HTB-36). Akt Inhibitor VIII was from Calbiochem (Merck, Zug, Switzerland). Adenosine 5′-triphosphate [gamma-32P] ([gamma32P] ATP) was from Perkin Elmer (Maanstraat, The Netherlands).
4
Establishing GBM and Non-Cancerous Cell Lines
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GBM cell lines (U87, U251, LN229, DBTRG) and non-cancerous human lung fibroblast cell line (MRC-5) were purchased from ATCC. U87, U251 and MRC-5 were cultured in MEM (Gibco) with 10% fetal bovine serum (FBS) and 2 mM L-glutamine. LN229 was cultured in DMEM (Gibco) with 10% FBS and 2 mM L-glutamine. DBTRG was cultured in RPMI1640 (Gibco) supplemented with 10% FBS, 2 mM L-glutamine, 25 mM HEPES and 1 mM sodium pyruvate. Patient-derived GBM cell line BAH1 was kindly provided by our collaborative researchers at Queensland Institute of Medical Research and was cultured in Advanced DMEM/F12 (Gibco) mixed with NeurobasalTM-A medium (Gibco) (1:1) supplemented with B-27 (1×), FGF (20 ng/mL) and EGF (20 ng/mL). Normal human astrocytes were purchased from Lonza and cultured in Astrocyte Growth Medium with Astrocyte Medium Bullet Kit (Lonza). PENAO was synthesized as previously published [25 (link)]. Sodium dichloroacetate (DCA), N-acetyl-L-cysteine (NAC), glutathione reduced ethyl ester (GSH-MEE) and buthionine sulphoximine (BSO) were purchased from Sigma-Aldrich.
5
Immortalized Head and Neck PGL Cell Lines
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We have previously established and immortalized head and neck PGL cell cultures PTJ64i and PTJ86i from two patients carrying the SDHC c.43 C > T (p.Arg15*) and SDHD c.27delC (p.Val10Phefs*5) mutations, respectively9 (link). PTJ64i and PTJ86i cell lines were cultured in DMEM-F12 (Sigma), supplemented with 10% FBS, at 37 °C, 5% CO2. Sodium dichloroacetate (DCA) and metformin were purchased from Sigma Aldrich. GW6471 was obtained from Tocris Bioscience (Bristol, UK).
6
Metabolic Regulation in Cell Culture
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Flasks and plates were obtained from Corning Inc. High-glucose D-MEM and fetal bovine serum (FBS) were from Lonza, and High-glucose D-MEM without phenol red, geneticin (G418 sulfate), pyruvate, l -glutamine and penicillin/streptomycin were from GIBCO, Invitrogen. [U-13C6]glucose and [U-13C5]glutamine were from Cambridge Isotope Laboratories (Andover, MA, USA). Sodium dichloroacetate (DCA) was from Sigma-Aldrich.
The primary antibodies were hexokinase II, glyceraldehyde 3-phosphate dehydrogenase, pyruvate kinase isozyme M2, pyruvate dehydrogenase α subunit (Glycolysis Antibody Sampler Kit), lactate dehydrogenase A (#2012), all from Cell Signaling; pyruvate dehydrogenase kinase 1 (KAP-PK112) from Stressgen; hypoxia-inducible factor 1α from Novus Biologicals (NB100-479); peroxisome proliferator-activated receptor γ co-activator-1α from Santa Cruz Biotechnology (sc-13067); glutamine synthetase from Abcam (ab49873); monocarboxylate transporter 1 from Millipore (#AB1286-I); actin from Millipore (MAB 1501).
The primary antibodies were hexokinase II, glyceraldehyde 3-phosphate dehydrogenase, pyruvate kinase isozyme M2, pyruvate dehydrogenase α subunit (Glycolysis Antibody Sampler Kit), lactate dehydrogenase A (#2012), all from Cell Signaling; pyruvate dehydrogenase kinase 1 (KAP-PK112) from Stressgen; hypoxia-inducible factor 1α from Novus Biologicals (NB100-479); peroxisome proliferator-activated receptor γ co-activator-1α from Santa Cruz Biotechnology (sc-13067); glutamine synthetase from Abcam (ab49873); monocarboxylate transporter 1 from Millipore (#AB1286-I); actin from Millipore (MAB 1501).
7
Metabolic Modulation in Disease Models
8
Sodium Dichloroacetate and Trichloroacetate Synthesis
9
Culturing and Conditioning CRC Cell Lines
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Human CRC cell lines SW620, HCT116, SW480, Colo320, Caco2, SW48, and HIEC-6 were obtained from the American Type Culture Collection (ATCC). Cells were incubated at 37 °C in a 5% CO2-humidified incubator and cultured in DMEM medium (Gibco, NY, USA) or RPMI 1640 (Gibco, NY, USA) with 10% fetal bovine serum (Gibco, NY, USA) and 1% penicillin-streptomycin (Gibco, CA, USA). After growing to 80% confluence, cells were washed with PBS, and fresh serum-free media was added. Conditioned medium (CM) was harvested 48 h later, filtered through a 0.22-μm filter to remove cell debris, and then stored at −80 °C until use. To block the production of lactate, the indicated cells were treated with 2 mM sodium dichloroacetate (DCA, Sigma) for 72 h.
10
Investigating PDK4 and PKA Modulators
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The PDK4 inhibitor sodium dichloroacetate (DCA) and the PKA activator dibutyryl cyclic-AMP (dbcAMP) were from Sigma-Aldrich/Merck, USA.
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