1. Cell Cycle/DNA Damage
    Metabolic Enzyme/Protease
  2. HSP
  3. ML346

ML346 

Cat. No.: HY-18669 Purity: >98.0%
Handling Instructions

ML346, a probe, has been extensively tested by the assay provider in several mechanism-of-action and protein folding assays.
ML346 (0-25μM; 24 hours) is not toxic to HeLa cells. ML346 increases the expression of Hsp70 mRNA by 2.4-fold. ML346 induces Hsp70, Hsp40, and Hsp27 expression.
ML346 activates transcription of the Hsp70 promoter.

ML346 is an activator of Hsp70 expression and HSF-1 activity, with an EC50 of 4.6 μM for Hsp70. ML346 restores protein folding in conformational disease models, without significant cytotoxicity or lack of specificity. ML346 induces specific increases in genes and protein effectors of the heat shock response (HSR), including chaperones such as Hsp70, Hsp40, and Hsp27.

For research use only. We do not sell to patients.

ML346 Chemical Structure

ML346 Chemical Structure

CAS No. : 100872-83-1

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Description

ML346 is an activator of Hsp70 expression and HSF-1 activity, with an EC50 of 4.6 μM for Hsp70. ML346 restores protein folding in conformational disease models, without significant cytotoxicity or lack of specificity. ML346 induces specific increases in genes and protein effectors of the heat shock response (HSR), including chaperones such as Hsp70, Hsp40, and Hsp27[1].

IC50 & Target[1]

HSP70

4.6 μM (EC50, HeLa cells)

In Vitro

ML346 is an activator of Hsp70, with an EC50 of 4600 nM in HeLa cells. ML346 (10 μM) restores proteostasis, restores CFTR-mediated iodide conductance, and enhances the correct folding of proteins expressed in two different cellular compartments[1]. ML346 (Compound F1) induces multiple responses and strongly induces Hsp70, the oxidative stress response genes (HO1 and GCLM), and a 2.5-fold upregulation of BiP in WT MEF cells. ML346 (0.5-25 μM) exhibits cytoprotective effects in cells after a 35 min severe heat shock, and also causes a two-fold protection from H2O2-induced apoptosis[2].

In Vivo

ML346 suppress the aggregation of polyQ35 in a C. elegans model, suggesting the probe has efficacy in modifying protein aggregation and associated toxicity[1].

Molecular Weight

272.26

Formula

C₁₄H₁₂N₂O₄

CAS No.

100872-83-1

SMILES

O=C1NC(/C(C(N1)=O)=C/C=C/C2=CC=C(OC)C=C2)=O

Shipping

Room temperature in continental US; may vary elsewhere.

Storage
Powder -20°C 3 years
  4°C 2 years
In solvent -80°C 6 months
  -20°C 1 month
Solvent & Solubility
In Vitro: 

DMSO : ≥ 29 mg/mL (106.52 mM)

*"≥" means soluble, but saturation unknown.

Preparing
Stock Solutions
Concentration Solvent Mass 1 mg 5 mg 10 mg
1 mM 3.6730 mL 18.3648 mL 36.7296 mL
5 mM 0.7346 mL 3.6730 mL 7.3459 mL
10 mM 0.3673 mL 1.8365 mL 3.6730 mL
*Please refer to the solubility information to select the appropriate solvent.
References
Kinase Assay
[2]

In brief, HeLa cells are incubated with either DMSO (negative control), the positive controls MG132 (10 μM) and lactacystin (6 μM) or the PRs A1, A3 and ML346 (F1) for 3 and 6 hours and then harvested. Cells are lysed in homogenization buffer (50 mM Tris-HCl, pH7.5, 250 mM sucrose, 5 mM MgCl2, 2 mM ATP, 1 mM DTT, 0.5 mM EDTA, 0.025% digitonin) for 5 min on ice, and total protein concentration of whole cell extract is determined. 3 μg of whole cell extracts are combined with assay buffer (50 mM Tris-HCl, pH 7.5, 40 mM KCl, 5 mM MgCl2, 0.5 mM ATP, 1 mM DTT, 0.05 mg/mL BSA) in a black 96-well plate and the reaction is initiated by the addition of a 2× (200 μM) fluorogenic peptide substrate Suc-LLVY-AMC. Fluorescence is measured every 10 min using a Synergy H4 multi-mode microplate reader[2].

MCE has not independently confirmed the accuracy of these methods. They are for reference only.

Cell Assay
[2]

HeLa cells are plated at a density of 10,000 cells per well in black 96-well plates in 100 μL of DMEM supplemented with 10% FBS and 1% Pen/Strep/Neo. Plates are incubated for 16 hours at 37°C, 5% CO2 and 95% relative humidity before compound addition. 1 μL of hit compounds (ML346) in DMSO or DMSO alone are added to the sample or control wells, respectively. Plates are then placed back in the incubator for 24 hours. After incubation, cells are washed 2× with 200 μL of PBS and 200 μL of a solution of 1 μg/mL of calcein AM is added to each well. Cells are then incubated for 45 min at 37°C, 5% CO2 before fluorescence measurement using an Analyst GT multimode reader. Percent cytotoxicity is expressed relative to wells containing cells treated with DMSO only (100%)[2].

MCE has not independently confirmed the accuracy of these methods. They are for reference only.

References
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Keywords:

ML346ML 346ML-346HSPHeat shock proteinsprobeheatshockresponseproteinconformationalcytotoxicitychaperonesInhibitorinhibitorinhibit

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