Background
A-1210477 is an effective and specific MCL-1 inhibitor with an EC50 value below 5 μmol/L [1]. Selectively, it binds to MCL-1 with an affinity of 0.45 nM [2].
MCL-1, an anti-apoptotic Bcl-2 family member, is an anti-apoptotic protein. It is a key regulator of cancer cell survival [3, 4].
In MCL-1-dependent SVEC cells, treatment with A-1210477 at varying doses, induced cell death in a dose-dependent manner. SYTOX Green exclusion and live-cell imaging were used to determine cell viability. In line with increased potency, cell death was more rapidly induced by A-1210477. To examine the selectivity of A-1210477 for targeting Bcl-2 family members, BcL-xL-, BcL-2-, and MCL-1-dependent SVEC cells were treated with A-1210477. A-1210477 only killed MCL-1-dependent cells. Compared with UMI-77, A-1210477 showed greater potency and specificity as an MCL-1 inhibitor, the EC50 value of UMI-77 is 10 μmol/L [1]. In living cells, A-1210477 disrupted BIM/MCL-1 complexes. In MCL-1-dependent cancer cells, A-1210477 induced the hallmarks of mitochondrial apoptosis. In various malignant cell lines, A-1210477 induced apoptosis, synergizing with navitoclax. Data also demonstrate that A-1210477 acted through an on-target mechanism. It appeared as the first BH3 mimetic targeting MCL-1 [2].
The pharmacokinetics of A-1210477 are not favorable for in vivo use [5].
参考文献:
[1].? Lopez J, Bessou M, Riley JS, et al. Mito-priming as a method to engineer Bcl-2 addiction. Nature communications, 2016, 7:10538.
[2].? Besbes S, Mirshahi M, Pocard M, et al. New dimension in therapeutic targeting of BCL-2 family proteins. Oncotarget, 2015, 6(15): 12862.
[3].? Leverson JD, Zhang H, Chen J, et al. Potent and selective small-molecule MCL-1 inhibitors demonstrate on-target cancer cell killing activity as single agents and in combination with ABT-263 (navitoclax). Cell death & disease, 2015, 6(1): e1590.
[4].? Mott JL, Kobayashi S, Bronk SF, et al. mir-29 regulates Mcl-1 protein expression and apoptosis. Oncogene, 2007, 26(42): 6133-6140.
[5].? Opferman JT. Attacking cancer's Achilles heel: antagonism of anti-apoptotic BCL-2 family members. FEBS Journal, 2015.
Protocol
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Kinase experiment [1]:
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Binding affinity assays
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TR-FRET-binding affinity assays were performed for BCL-2, BCL-XL and MCL-1 in 4.52 mM monobasic potassium phosphate, 15.48 mM dibasic potassium phosphate, 1 mM sodium EDTA, 0.05% Pluronic F-68 detergent, 50 mM sodium chloride and 1 mM DTT (pH 7.5). For MCL-1 assays, GST-tagged MCL-1 (1 nM) was mixed with 100 nM f-Bak, 1 nM Tb-labeled anti-GST antibody and compound at room temperature for 60 mins. Fluorescence was measured on an Envision plate reader using a 340/35 nm excitation filter and 520/525 (f-Bak) and 495/510 nm (Tb-labeled anti-GST antibody) emission filters.
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Cell experiment [1]:
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Cell lines
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H929 cells
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Preparation method
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This compound is soluble in DMSO. General tips for obtaining a higher concentration: Please warm the tube at 37 °C for 10 minutes and/or shake it in the ultrasonic bath for a while. Stock solution can be stored below - 20 °C for several months.
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Reacting condition
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0 ~ 30 μM; 4 hrs
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Applications
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In H929 cells, A-1210477 bound selectively and strongly to MCL-1, and reduced the amount of BIM co-immunoprecipitated with MCL-1 in a dose-dependent manner with an IC50 value in the low-μM range.
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参考文献:
[1]. Leverson JD, Zhang H, Chen J, et al. Potent and selective small-molecule MCL-1 inhibitors demonstrate on-target cancer cell killing activity as single agents and in combination with ABT-263 (navitoclax). Cell death & disease, 2015, 6(1): e1590.
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