LSD1 demethylation assay

Human recombinant LSD1 protein is incubated with dimethylated H3K4 peptide as the substrate in the presence of various concentrations of lead compound inhibitors (0 to 75 μM ) or control tranylcypromine. The demethylase activity is measured by the release of H2O2 produced during the catalytic process, using the Amplex red peroxide/peroxidase-coupled assay kit. Each reaction is done in triplicate. The maximum LSD1 demethylase activity is obtained in the absence of inhibitor and corrected for background fluorescence. The Ki (IC50) of OG-L002 is calculated as half-maximal activity.

Cell lines

HeLa, HFF cells, MRC-5 cells

Preparation method

The solubility of this compound in DMSO is >10 mM. General tips for obtaining a higher concentration: Please warm the tube at 37℃ for 10 minutes and/or shake it in the ultrasonic bath for a while. Stock solution can be stored below -20℃ for several months.

Reacting condition

50 μM for 12 hr;

Applications

OG-L002 inhibited HSV IE gene expression and progeny virus production in both HeLa and HFF cells.

Animal models

Mouse ganglion explant model

Dosage form

2 to 40 mg/kg /day; intraperitoneal administration for 15-17 days

Applications

OG-L002 repressed herpes simplex virus (HSV) primary infection in vivo.

Other notes

Please test the solubility of all compounds indoor, and the actual solubility may slightly differ with the theoretical value. This is caused by an experimental system error and it is normal.

OG-L002 is a specific and potent inhibitor of lysine (K)-specific demethylase 1A (LSD1) with IC50 value of 20 nM [1].
LSD1 is a flavin-dependent monoamine oxidase, which can demethylate lysines. LSD1 plays critical roles in oocyte growth, embryogenesis and tissue-specific differentiation [2].
OG-L002 potently inhibited HSV IE gene expression in both HFF and HeLa cells with IC50 of ~3 μM and ~10 μM, respectively. OG-L002 treatment can potently reduce production of progeny virus (~100-fold) with no significant toxicity in HeLa or HFF cells. In chromatin immunoprecipitation assays, OG-L002 increased the levels of total histone H3K9-me2 and H3 (20- to 30-fold) associated with viral IE promoters, which resulted in decreased viral IE gene expression. In addition, OG-L002 also repressed the expression of adenovirus E1A gene and hCMV IE genes [1].
In a mouse model, OG-L002 repressed primary HSV infection in a dose-dependent manner. Moreover, OG-L002 plays an important role in the viral latency-reactivation cycle in a mouse ganglion explant model [1].
References:
[1]. Liang Y, Quenelle D, Kristie TM, et al. A Novel Selective LSD1/KDM1A Inhibitor epigenetically blocks herpes simplex virus lytic replication and reactivation from latency. mBio, 2013, 4(1), e00558-12.
[2]. Pedersen MT, Helin K. Histone demethylases in development and disease. Trends in Cell Biology, 2010, 20 (11): 662-71.