Home Ubiquitin DUB inhibitor ML323 For research use only.

ML323

ML323 displays reversible, nanomolar inhibitory activity and excellent selectivity toward USP1/UAF1 with IC50 of 76 nM.

ML323 Chemical Structure

ML323 Chemical Structure

CAS No. 1572414-83-5

Purity & Quality Control

ML323 Related Products

Signaling Pathway

DUB Signaling Pathways

DUB Signaling Pathway

Biological Activity

Description ML323 displays reversible, nanomolar inhibitory activity and excellent selectivity toward USP1/UAF1 with IC50 of 76 nM.
Features USP1-UAF1-selective inhibitor.
Targets
USP1-UAF1 [1]
(Cell-free assay)
76 nM
In vitro
In vitro

ML323 inhibits the deubiquitination of PCNA and FANCD2 by inhibiting USP1–UAF1 activity in H596 cells. Moreover, ML323 potentiates cytotoxicity in NSCLC H596 cells and U2OS osteosarcoma cells by targeting two major DNA damage response pathways (TLS and FA). [1]
Kinase Assay High-throughput screening
For HTS, USP1-UAF1 activity is monitored using ubiquitin-rhodamine 110 as a substrate. The assay is miniaturized to a 4 μL volume in a 1,536-well format and is used to screen approximately 402,701 compounds in quantitative HTS mode, with each compound tested over a range of four to five concentrations. The assay shows robust performance with an average Z’factor of 0.8 throughout the screen.
Cell Research Cell lines H596 cells
Concentrations ~30 μM
Incubation Time 7-12 days
Method

For the colony-forming assay, cells are seeded at a density of 300–500 cells per well in six-well plates and grown overnight. Cells are then treated with ML323 alone at the indicated concentrations. Cells treated with an equal volume of DMSO and saline are used as control. After 48 h of treatment, fresh growth medium is added, and cells are incubated for an additional 5-10 d to allow for colony formation. For UV combination treatment, the cells are treated with ML323 at the indicated concentrations or an equal volume of DMSO. After 48 h, the medium is removed, and cells are irradiated at 254 nm at the indicated dosage. Fresh growth medium is added, and the cells are incubated for an additional 5-10 d to allow for colony formation. The cells without UV irradiation but treated with ML323 or an equal volume of DMSO are used as controls and designated as 100%. After the formation of the colonies, cells are fixed with methanol and stained with 0.5% crystal violet. Colonies consisting of >50 cells are scored. The number of colonies is determined from triplicate plates. The dose-response curves are generated using GraphPad Prism and analyzed by using CalcuSyn to calculate the combination index.
In Vivo
In vivo

ML323 is a selective USP1–UAF1 inhibitor that inhibit deubiquitinase activity of USP1–UAF1 complex.
Animal Research Animal Models Female C57BL/6J mice
Dosages 10 mg/kg
Administration i.p.

Chemical Information & Solubility

Molecular Weight 384.48 Formula

C23H24N6
CAS No. 1572414-83-5 SDF Download ML323 SDF
Smiles CC1=CN=C(N=C1NCC2=CC=C(C=C2)N3C=CN=N3)C4=CC=CC=C4C(C)C
Storage (From the date of receipt)

In vitro
Batch:

DMSO : 76 mg/mL ( (197.66 mM) Moisture-absorbing DMSO reduces solubility. Please use fresh DMSO.)

Ethanol : 38 mg/mL

Water : Insoluble


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In vivo
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Method for preparing DMSO master liquid: mg drug pre-dissolved in μL DMSO ( Master liquid concentration mg/mL, Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug. )

Method for preparing in vivo formulation: Take μL DMSO master liquid, next addμL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH2O, mix and clarify.

Method for preparing in vivo formulation: Take μL DMSO master liquid, next add μL Corn oil, mix and clarify.

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Tech Support

Answers to questions you may have can be found in the inhibitor handling instructions. Topics include how to prepare stock solutions, how to store inhibitors, and issues that need special attention for cell-based assays and animal experiments.

Handling Instructions

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