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ML385

Catalog No.: GC19254

ML385 is a specific nuclear factor erythroid 2-related factor 2 (NRF2) inhibitor.

ML385 Chemical Structure

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10mM (in 1mL DMSO)
$59.00
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5mg
$42.00
In stock
10mg
$63.00
In stock
25mg
$133.00
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50mg
$217.00
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100mg
$343.00
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Sample solution is provided at 25 µL, 10mM.

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Protocol

Cell experiment [1]:

Cell lines

A549 cells

Preparation Method

A549 cells were transfected with a firefly luciferase reporter (Fluc) construct driven by a minimal TATA promoter with upstream NRF2-specific antioxidant response element (ARE) enhancer sequence from human NQO1 promoter ARE and clones stably expressing ARE-FLuc7 were screened and validated.

Reaction Conditions

Cells were cultured in the presence of ML385 (5 μM) for 48h and 72 h and measured the changes in the expression levels of NRF2 and its target genes.

Applications

ML385 treatment to A549 cells demonstrated a reduction in glutathione synthesis and recycling enzymes, members of the thioredoxin family, and glucose metabolism-related genes with a time-dependent manner. ML385 treatment significantly attenuated NQO1 enzyme activity and reduced GSH levels along with cellular antioxidant capacity.

Animal experiment [2]:

Animal models

1-8-week-old C57B/6 male mice

Preparation Method

Mice received a daily intraperitoneal injection of ML385 (30 mg/kg) dissolved in PBS with 5% Dimethyl Sulfoxide (DMSO) for 7 d.

Dosage form

30 mg/kg

Applications

ML385 could inhibit Nrf2 and used to explore the mechanism of Nrf2-NF-κB signaling pathway involved in the inhibition of astrocyte activation. Mice treated with ML385 had a significant decrease in Keap1, HO-1, Nrf2, and p-P65/P-65 (P < 0.05) expression compared to Sham or Vehicle control groups.

References:

[1]. Singh A, et al. Small Molecule Inhibitor of NRF2 Selectively Intervenes Therapeutic Resistance in KEAP1-Deficient NSCLC Tumors. ACS Chem Biol. 2016 Nov 18;11(11):3214-3225.

[2]. Xian P, et al. Mesenchymal stem cell-derived exosomes as a nanotherapeutic agent for amelioration of inflammation-induced astrocyte alterations in mice. Theranostics. 2019 Aug 14;9(20):5956-5975.

Background

ML385 is a specific nuclear factor erythroid 2-related factor 2 (NRF2) inhibitor. ML385 binds to NRF2 and inhibits its downstream target gene expression as a probe molecule. Specifically, ML385 binds to the Neh1, the Cap ‘N’ Collar Basic Leucine Zipper (CNC-bZIP) domain of NRF2, and interferes with the binding of the V-Maf Avian Musculoaponeurotic Fibrosarcoma Oncogene Homolog G (MAFG)-NRF2 protein complex to regulatory DNA binding sequences. ML385 shows specificity and selectivity for NSCLC cells with KEAP1 mutation leading to gain of NRF2 function.[1][2]

In vitro study demonstrated that ML385 potentially inhibits NRF2 through direct interaction. The NRF2 signaling was decreased in a time-dependent manner and the maximum decline was at 72 h. A reduction in NRF2 mRNA levels was also observed. In addition, ML385 caused global inhibition of NRF2 signaling in lung cancer cells with KEAP1 mutations, and other target genes. ML385 treatment also significantly attenuated NQO1 enzyme activity and reduced GSH levels along with cellular antioxidant capacity. [2]

In vivo study of ML385 indicated that it inhibits NRF2 and showed promising anti-tumor activity. ML385 in combination with carboplatin showed a significant reduction in tumor growth compared to vehicle. Although the treatment with a single agent (either ML385 or carboplatin) led to a reduction in tumor growth, the magnitude of these effects was variable between cell lines and did not reach statistical significance. Moreover, tumor samples treated with ML385 showed a significant reduction in NRF2 protein level and its downstream target genes. Besides, the addition of ML385 decreased anisotropy in a dose-dependent manner, with an IC50 of 1.9 μM, suggesting that the NRF2-MAFG protein complex was dissociated from fluorescein-labeled ARE-DNA. [2]

References:
[1]. Xian P, et al. Mesenchymal stem cell-derived exosomes as a nanotherapeutic agent for amelioration of inflammation-induced astrocyte alterations in mice. Theranostics. 2019 Aug 14;9(20):5956-5975.
[2]. Singh A, et al. Small Molecule Inhibitor of NRF2 Selectively Intervenes Therapeutic Resistance in KEAP1-Deficient NSCLC Tumors. ACS Chem Biol. 2016 Nov 18;11(11):3214-3225.

Chemical Properties

Cas No. 846557-71-9 SDF
Synonyms N/A
Chemical Name N/A
Canonical SMILES O=C(NC1=NC(C2=CC=C(N(C(C3=C(C)C=CC=C3)=O)CC4)C4=C2)=C(C)S1)CC5=CC(OCO6)=C6C=C5
Formula C29H25N3O4S M.Wt 511.59
Solubility DMSO : ≥ 30 mg/mL (58.64 mM) 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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Research Update

Small Molecule Inhibitor of NRF2 Selectively Intervenes Therapeutic Resistance in KEAP1-Deficient NSCLC Tumors

ACS Chem Biol2016 Nov 18;11(11):3214-3225.PMID: 27552339DOI: 10.1021/acschembio.6b00651

Loss of function mutations in Kelch-like ECH Associated Protein 1 (KEAP1), or gain-of-function mutations in nuclear factor erythroid 2-related factor 2 (NRF2), are common in non-small cell lung cancer (NSCLC) and associated with therapeutic resistance. To discover novel NRF2 inhibitors for targeted therapy, we conducted a quantitative high-throughput screen using a diverse set of ∼400 000 small molecules (Molecular Libraries Small Molecule Repository Library, MLSMR) at the National Center for Advancing Translational Sciences. We identified ML385 as a probe molecule that binds to NRF2 and inhibits its downstream target gene expression. Specifically, ML385 binds to Neh1, the Cap 'N' Collar Basic Leucine Zipper (CNC-bZIP) domain of NRF2, and interferes with the binding of the V-Maf Avian Musculoaponeurotic Fibrosarcoma Oncogene Homologue G (MAFG)-NRF2 protein complex to regulatory DNA binding sequences. In clonogenic assays, when used in combination with platinum-based drugs, doxorubicin or taxol, ML385 substantially enhances cytotoxicity in NSCLC cells, as compared to single agents. ML385 shows specificity and selectivity for NSCLC cells with KEAP1 mutation, leading to gain of NRF2 function. In preclinical models of NSCLC with gain of NRF2 function, ML385 in combination with carboplatin showed significant antitumor activity. We demonstrate the discovery and validation of ML385 as a novel and specific NRF2 inhibitor and conclude that targeting NRF2 may represent a promising strategy for the treatment of advanced NSCLC.

Nrf2 protects against seawater drowning-induced acute lung injury via inhibiting ferroptosis

Respir Res2020 Sep 9;21(1):232.PMID: 32907551DOI: 10.1186/s12931-020-01500-2

Background: Ferroptosis is a new type of nonapoptotic cell death model that was closely related to reactive oxygen species (ROS) accumulation. Seawater drowning-induced acute lung injury (ALI) which is caused by severe oxidative stress injury, has been a major cause of accidental death worldwide. The latest evidences indicate nuclear factor (erythroid-derived 2)-like 2 (Nrf2) suppress ferroptosis and maintain cellular redox balance. Here, we test the hypothesis that activation of Nrf2 pathway attenuates seawater drowning-induced ALI via inhibiting ferroptosis.
Methods: we performed studies using Nrf2-specific agonist (dimethyl fumarate), Nrf2 inhibitor (ML385), Nrf2-knockout mice and ferroptosis inhibitor (Ferrostatin-1) to investigate the potential roles of Nrf2 on seawater drowning-induced ALI and the underlying mechanisms.
Results: Our data shows that Nrf2 activator dimethyl fumarate could increase cell viability, reduced the levels of intracellular ROS and lipid ROS, prevented glutathione depletion and lipid peroxide accumulation, increased FTH1 and GPX4 mRNA expression, and maintained mitochondrial membrane potential in MLE-12 cells. However, ML385 promoted cell death and lipid ROS production in MLE-12 cells. Furthermore, the lung injury became more aggravated in the Nrf2-knockout mice than that in WT mice after seawater drowning.
Conclusions: These results suggested that Nrf2 can inhibit ferroptosis and therefore alleviate ALI induced by seawater drowning. The effectiveness of ferroptosis inhibition by Nrf2 provides a novel therapeutic target for seawater drowning-induced ALI.

Protective Effects of Chlorogenic Acid on Cerebral Ischemia/Reperfusion Injury Rats by Regulating Oxidative Stress-Related Nrf2 Pathway

Drug Des Devel Ther2020 Jan 7;14:51-60.PMID: 32021091DOI: 10.2147/DDDT.S228751

Introduction: Cerebral ischemia-reperfusion (CI/R) injury is caused by blood flow recovery after ischemic stroke. Chlorogenic acid (CGA, 5-O-caffeoylquinic acid) is a major polyphenol component of Coffea canephora, Coffea arabica L. and Mate (Ilex paraguariensis A. StHil.). Previous studies have shown that CGA has a significant neuroprotective effect and can improve global CI/R injury. However, the underlying molecular mechanism of CGA in CI/R injury has not been fully revealed.
Materials: In this study, CI/R rat model was constructed. The rats were randomly divided into nine groups with ten in each group: Control, CGA (500 mg·kg-1), CI/R, CI/R + CGA (20 mg·kg-1), CI/R + CGA (100 mg·kg-1), CI/R + CGA (500 mg·kg-1), ML385 (30 mg·kg-1), CI/R + ML385 (30 mg·kg-1), CI/R + CGA + ML385. Cerebral infarction volume was detected by TTC staining. Brain pathological damage was detected by H&E staining. Apoptosis of cortical cells was detected by TUNEL staining. The expression of related proteins was detected by RT-qPCR and Western blotting.
Results: Step-down test and Y maze test showed that CGA dose-dependently mitigated CI/R-induced brain damage and enhanced learning and spatial memory. Besides, CGA promoted the expression of BDNF and NGF in a dose-dependent manner and alleviated CI/R-induced nerve injury. Moreover, CGA increased the activity of SOD and the level of GSH, as well as decreased production of ROS and LDH and the accumulation of MDA. Notably, CGA attenuated oxidative stress-induced brain injury and apoptosis and inhibited the expression of apoptosis-related proteins (cleaved caspase 3 and caspase 9). Additionally, CGA reversed CI/R induced inactivation of Nrf2 pathway and promoted Nrf2, NQO-1 and HO-1 expression. Nrf2 pathway inhibitor ML385 destroyed this promotion.
Discussion: All the data indicated that CGA had a neuroprotective effect on the CI/R rats by regulating oxidative stress-related Nrf2 pathway.

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