Tempol

Prkaa1^fl and Prkaa2^fl mice were reported before (Nakada et al., 2010 (link)). C57BL/Ka-Thy-1.2 (CD45.1), C57BL/Ka-Thy-1.1 (CD45.2), and NOD-SCID mice (8-12 weeks of age) were used for AML transplantation assays. Poly(I:C) (Amersham, Piscataway, NJ) was injected intraperitoneally with 0.5 μg/gram of body mass every other days for 6 days. 5 mg of tamoxifen (Sigma, St. Louis, MO) was administered daily by oral gavage for 5 days. 1 mM Tempol (4-hydroxy-2,2,6,6-tetramethylpiperidinyloxy) (Enzo Life Science, Farmingdale, NY) was given to mice in the drinking water. 4 mg/kg of compound C (Cayman Chemical, Ann Arbor, MI) diluted in DMSO was intraperitoneally injected for 4 weeks. Metformin (1.5 mg/ml) was given to mice in the drinking water. All mice were fed Harlan Rodent Diet 2920X, either ad libitum (AL) or with DR regimen (70% of the normal food intake). Mice were housed in AAALAC-accredited, specific-pathogen-free animal care facilities at Baylor College of Medicine (BCM). All procedures were approved by BCM Institutional Animal Care and Use Committees.

Nitroxide radicals TEMPOL and mitoTEMPO were purchased from Enzo Life Sciences (San Diego, CA), bromomethyl acetate was purchased from Sigma-Aldrich (St. Louis, MO). The 3,3-dicarboxy-2,2,5,5-tetra-methylpyrrolidine-1-oxyl (DCP) was prepared in analogy to original method [14 (link),20 ] with some modifications described in [21 (link)]. The synthesis of nitroxides DCP-AM, DCP-AM2 [14 (link)] and DCAP-AM2 and of corresponding hydroxylamine DCP-AM-H is shown in Schemes 1 and 2.

DIPPMPO (5-diisopropoxyphosphoryl-5-methyl-1-pyrroline N-oxide) was purchased from Radical Vision (Marseille, France) and from Enzo Life Science (Antwerpen, Belgium). CMH (1-hydroxy-3-methoxycarbonyl-2,2,5,5-tetramethylpyrrolidine), TEMPOL (4-hydroxy-2,2,6,6-tetramethylpiperidine-1-oxyl), mitoTEMPO ((2-(2,2,6,6-tetramethylpiperidin-1-oxyl-4-ylamino)-2-oxoethyl)triphenylphosphonium chloride) and mitoTEMPO-H (1-hydroxy-4-[2-(triphenylphosphonio)-acetamido]-2,2,6,6-tetramethylpiperidine) were from Enzo Life Science. CT-03 (Finland radical) was synthesized as previously described [28 (link),29 ]. Catalase conjugated with polyethylene glycol (PEG-CAT), superoxide dismutase conjugated with polyethylene glycol (PEG-SOD), superoxide dismutase (SOD), catalase (CAT), xanthine oxidase (XO), hypoxanthine (HX), hydrogen peroxide, ferric sulfate, diethylene triamine pentaacetic acid (DTPA), cytochrome c oxidase and phorbol 12-myristate 13-acetate (PMA) were from Sigma-Aldrich (Overijse, Belgium). Dulbecco’s modified Eagle medium (DMEM), (catalogue number: 41966029) and phosphate buffered saline solution (PBS), (catalogue number: 10010023) were from ThermoFisher.

SOD mimics and non-SOD mimics were analyzed. Mn-, Fe porphyrins, Mn Salen and EUK-8 were synthesized as reported [45 –50 (link)]. Mn(II) cyclic polyamines, M40403 and M40404 were synthesized based on the literature procedures [51 (link)]. Nitrone NXY-059 was purchased from Selleckchem, Cat No. S6002, >99 % and nitroxide, Tempol, was obtained from Enzo, Cat No. ALX-430-081-G001, Lot No. L27338C. Catalase enzyme from bovine liver was bought from Sigma, Cat No. C1345-1G, Lot No. 010M7011V. Ebselen was purchased Cayman Chemical Company, Cat No. 70530, Lot No. 0406449-57. Figure 1 shows the structures of Mn porphyrins and Figure 2 the structures of other redox-active drugs studied here.