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ML-265 Catalog No.GC11741

tumor-specific PKM2 activator

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5mg
$72.00
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10mg
$128.00
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25mg
$233.00
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Sample solution is provided at 25 µL, 10mM.

Quality Control

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Protocol

Kinase experiment:

Pyruvate kinase activity is measured by monitoring pyruvate-dependent conversion of NADH to NAD+ by lactate dehydrogenase (LDH) as described previously. Briefly, for cell line experiments, the medium is replaced with fresh medium 1 hour prior to the start of treatment with DMSO or activator. Also, where indicates, 100 μM pervanadate is added 10 minutes prior to cell lysis. Cells are lysed on ice with NP-40 buffer containing 2 mM DTT and protease inhibitors and clarified by centrifugation at 21,000 x g. 5 μL of the supernatant is used to assess pyruvate kinase activity. Pyruvate kinase activity is subsequently normalized for total protein content.

Cell experiment:

2,000 cells are seeded in 96-well plates 24 h prior to treatment start. CellTiter96® AQueous is used to assess cell viability following oxidant and PKM2 activator combination treatments. MTS: (3-(4,5-dimethylthiazol-2-yl)-5- (3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium).

Animal experiment:

H1299 parental and H1299 cells with constitutive expression of a mouse PKM1 cDNA (H1299-PKM1 cells) are propagated in RPMI supplemented with 10% fetal bovine serum, penicillin/streptomycin, 2 mM glutamine, and hygromycin for transgene selection. Cells are harvested, resuspended in sterile PBS, and 5×105 cells are injected subcutaneously into nu/nu mice. Tumor growth is monitored by caliper measurement, the mice are sacrificed and tumors harvested after the time indicated. Tumors are weighed, divided and either flash-frozen in liquid nitrogen or fixed in formalin for later analysis.

References:

[1]. Anastasiou D, et al. Pyruvate kinase M2 activators promote tetramer formation and suppress tumorigenesis. Nat Chem Biol. 2012 Oct;8(10):839-847.
[2]. Palsson-McDermott EM, et al. Pyruvate kinase M2 regulates Hif-1α activity and IL-1β induction and is a critical determinant of the warburg effect in LPS-activated macrophages. Cell Metab. 2015 Jan 6;21(1):65-80.

Chemical Properties

Cas No. 1221186-53-3 SDF
Synonyms CID-44246499,NCGC00186528,TEPP-46
Chemical Name 6-[(3-aminophenyl)methyl]-4,6-dihydro-4-methyl-2-(methylsulfinyl)-5H-thieno[2’,3’:4,5]pyrrolo[2,3-d]pyridazin-5-one
Canonical SMILES CS(C1=CC(N2C)=C(S1)C3=C2C(N(CC4=CC(N)=CC=C4)N=C3)=O)=O
Formula C17H16N4O2S2 M.Wt 372.5
Solubility ≥37.3mg/mL in DMSO Storage Store at -20°C
General tips For obtaining a higher solubility , please warm the tube at 37 ℃ and shake it in the ultrasonic bath for a while.Stock solution can be stored below -20℃ for several months.
Shipping Condition Evaluation sample solution : ship with blue ice
All other available size: ship with RT , or blue ice upon request
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Background

ML-265 (TEPP-46) is a potent and selective PKM2 activator [1][2].

Pyruvate kinase catalyzes the final step in glycolysis and transfers the phosphate group from phosphoenolpyruvate (PEP) to adenosine diphosphate (ADP) to yield adenosine triphosphate (ATP) and pyruvate. Pyruvate kinase M2 (PKM2) is considerably less active and expressed in highly proliferative cells including all cancer cell lines and tumors, which require high amounts of glucose for proliferation [1][2].

ML-265 is a potent and selective PKM2 activator with AC50 value of 92 nM. ML-265 exhibits high selectivity over PKM1, PKR and PKL. ML-265 decreased the Km of PKM2 for PEP in a way similar to the endogenous activator FBP. ML-265 also increased PKM2 activity by promoting the tetrameric state [1].

In mice with A549 xenograft tumors, ML-265 exhibited good oral bioavailability with relatively low clearance, long half-life. ML-265 (150 mg/kg) readily achieved maximal PKM2 activation. In mice bearing H1299 xenograft tumors, ML-265 inhibited tumor growth, suggesting that increased pyruvate kinase activity can impair tumorigenesis [1][2].

References:
[1].  Anastasiou D, Yu Y, Israelsen WJ, et al. Pyruvate kinase M2 activators promote tetramer formation and suppress tumorigenesis. Nat Chem Biol. 2012 Oct;8(10):839-47.
[2].  Walsh MJ, Brimacombe KR, Anastasiou D, et al. ML265: A potent PKM2 activator induces tetramerization and reduces tumor formation and size in a mouse xenograft model. Probe Reports from the NIH Molecular Libraries Program [Internet].