SP2509 is a specific histone demethylase LSD1 inhibitor(IC50 =13 nM).

LSD1:13 nM

In OCI-AML3, SP250 inhibited colony formation and induced apoptosis. In AML cells, SP2509 inhibited the interaction of LSD1 with CoREST, increased promoter-specific H3K4Me3, and induced p53, p21 and C/EBPα. In primary AML cells, SP2509 induced cell proliferation.

In OCI-AML3, SP250 inhibited colony formation and induced apoptosis. In AML cells, SP2509 inhibited the interaction of LSD1 with CoREST, increased promoter-specific H3K4Me3, and induced p53, p21 and C/EBPα. In primary AML cells, SP2509 induced cell proliferation.

SP2509 activity assays: Test compounds are diluted to 20 × the desired test concentration in 100% DMSO and 2.5?μL of the diluted drug sampleis added to a black 384-well plate. The LSD1 enzyme stock is diluted 17-fold with assay buffer and 40?μL of the diluted LSD1 enzyme is added to the appropriate wells. Substrate, consisting of horseradish peroxidase, dimethyl K4 peptide corresponding to the first 21 amino acids of the N-terminal tail of histone H3, and 10-acetyl-3,7-dihydroxyphenoxazine is then added to wells. Resorufin is analyzed on an Envision plate reader with an excitation wavelength of 530?nm and an emission wavelength of 595?nm. The activity of SP2509 on the other oxidases is determined by using commercially available kits. The glucose oxidase activity (which also noncovalently binds FAD in an elongate conformation), is determined using the glucose oxidase kit. The MAO assays are performed using the MAO-glo kit with MAO-A and MAO-B.

Cultured AML cells are treated with SP2509 and/or PS for 96?h. At the end of treatment, cells are washed free of the drugs and 500 cells per condition are plated in methylcellulose and incubated at 37?°C. Colony formation is measured 7–10 days after plating.(Only for Reference)