L-685,458

Catalog No.S7673 Batch:S767301

Technical Data

Formula

C₃₉H₅₂N₄O₆

Molecular Weight

672.85

CAS No.

292632-98-5

Solubility (25°C)*

In vitro

DMSO

100 mg/mL (148.62 mM)

Ethanol

6 mg/mL (8.91 mM)

Water

Insoluble

In vivo (Add solvents to the product individually and in order)

Clear solution	5%DMSO 1 40%PEG300 2 5%Tween80 3 50%ddH2O	5.0mg/ml	Taking the 1 mL working solution as an example, add 50 μL of 100 mg/ml clarified DMSO stock solution to 400 μL of PEG300, mix evenly to clarify it; add 50 μL of Tween80 to the above system, mix evenly to clarify; then continue to add 500 μL of ddH2O to adjust the volume to 1 mL. The mixed solution should be used immediately for optimal results.
Clear solution	5% DMSO 1 95% Corn oil	5.0mg/ml	Taking the 1 mL working solution as an example, add 50 μL of 100 mg/ml clear DMSO stock solution to 950 μL of corn oil and mix evenly. The mixed solution should be used immediately for optimal results.

* <1 mg/ml means slightly soluble or insoluble.
* Please note that Selleck tests the solubility of all compounds in-house, and the actual solubility may differ slightly from published values. This is normal and is due to slight batch-to-batch variations.
* Room temperature shipping (Stability testing shows this product can be shipped without any cooling measures.)

Preparing Stock Solutions

Biological Activity

Description

L-685,458 is a specific and potent inhibitor of A beta PP gamma-secretase activity with K_i of 17 nM.

Targets

gamma-secretase ^[1]

17 nM(Ki)

In vitro

L-685,458 inhibits Aβ(40) formation in both Neuro2A and CHO cell lines overexpressing human AβPP695, and in SHSY5Y cells overexpressing the construct spβA4CTF with IC50 of 402 nM, 113 nM, and 48 nM, respectively, with the potency for reduction of Aβ(42) being about 2-fold lower. ^[1] In Tca8113 cells, L-685,458 inhibits cell growth by inducing G0–G1 cell cycle arrest and apoptosis. ^[2] In a T-cell acute lymphoblastic leukemia cell line, pre-treatment with L-685,458 enhances the anti-proliferative effect of imatinib. ^[3] L685,458 also significantly reduces HSV-1 replication in tissue culture by inhibiting signal peptide peptidase (SPP). ^[4]

Protocol (from reference)

Kinase Assay:^{^[1]}	HTRF Immunoassay for Aβ Quantitation Homogeneous time-resolved fluorescence (HTRF) immunoassay methodology uses fluorescence resonance energy transfer (FRET) between two fluorophores, a donor EuK and a modified allophycocyanine pigment acceptor molecule, XL-665. When in proximity, nonradiative FRET takes place from nitrogen laser-excited EuK to XL-665, resulting in the emission of an amplified long-lived fluorescence signal. Briefly, in a typical 96-well plate assay, each well contains 0.75 nM antibody-EuK, 1.0 nM antibody-biotin, 2.0 nM SA-XL665, and 0.1−0.2 M potassium fluoride. Samples of conditioned cell culture medium or synthetic peptide standards and culture medium alone are added to give a total assay volume of 200 μL/well. Blank values are determined by the inclusion of 1.0 nM nonbiotinylated antibody in place of the biotinylated antibody. Following mixing, the reaction mixture is left at 4 °C to reach equilibrium binding, and then read on the Discovery HTRF microplate analyzer using the manufacturer
Cell Assay:^{^[2]}	Cell lines Tca8113 cells Concentrations ~5.0 μM Incubation Time 48 h Method The in vitro growth rate of Tca8113 cells treated with L-685,458, is measured by MTT. Briefly, Tca8113 cells are seeded in 96-well plates. On the day of harvest, 100 μL of spent medium is replaced with an equal volume of fresh medium containing 10% MTT 5 mg ml−1 stock. Plates are incubated at 37°C for 4 h, then 100 μl of DMSO is added to each well, and then plates are shaken at room temperature for 10 min. The absorbance is measured at 570 nm.

Kinase Assay:^{^[1]}

HTRF Immunoassay for Aβ Quantitation
Homogeneous time-resolved fluorescence (HTRF) immunoassay methodology uses fluorescence resonance energy transfer (FRET) between two fluorophores, a donor EuK and a modified allophycocyanine pigment acceptor molecule, XL-665. When in proximity, nonradiative FRET takes place from nitrogen laser-excited EuK to XL-665, resulting in the emission of an amplified long-lived fluorescence signal. Briefly, in a typical 96-well plate assay, each well contains 0.75 nM antibody-EuK, 1.0 nM antibody-biotin, 2.0 nM SA-XL665, and 0.1−0.2 M potassium fluoride. Samples of conditioned cell culture medium or synthetic peptide standards and culture medium alone are added to give a total assay volume of 200 μL/well. Blank values are determined by the inclusion of 1.0 nM nonbiotinylated antibody in place of the biotinylated antibody. Following mixing, the reaction mixture is left at 4 °C to reach equilibrium binding, and then read on the Discovery HTRF microplate analyzer using the manufacturer

Cell Assay:^{^[2]}

Cell lines
Tca8113 cells
Concentrations
~5.0 μM
Incubation Time
48 h
Method
The in vitro growth rate of Tca8113 cells treated with L-685,458, is measured by MTT. Briefly, Tca8113 cells are seeded in 96-well plates. On the day of harvest, 100 μL of spent medium is replaced with an equal volume of fresh medium containing 10% MTT 5 mg ml−1 stock. Plates are incubated at 37°C for 4 h, then 100 μl of DMSO is added to each well, and then plates are shaken at room temperature for 10 min. The absorbance is measured at 570 nm.

References

[4] Allen SJ, et al. Exp Eye Res. 2014 Jun;123:8-15.

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Customer Product Validation

: Data from [Data independently produced by , , Sci Rep, 2017 , 7(1):4983]

: Data from [Data independently produced by , , PLoS One, 2017, 12(3):e0173240]

Selleck's L-685,458 has been cited by 4 publications

Comprehensive drug response profiling and pan-omic analysis identified therapeutic candidates and prognostic biomarkers for Asian cholangiocarcinoma [ iScience, 2022, 25(10):105182]	PubMed: 36248745
Carboxy-terminal fragment of amyloid precursor protein mediates lipid droplet accumulation upon γ-secretase inhibition [ Biochem Biophys Res Commun, 2021, 570:137-142]	PubMed: 34280617
A novel rhamnoside derivative PL402 up-regulates matrix metalloproteinase 3/9 to promote Aβ degradation and alleviates Alzheimer's-like pathology. [ Aging (Albany NY), 2020, 12(1):481-501]	PubMed: 31901901
A cellular complex of BACE1 and γ-secretase sequentially generates Aβ from its full-length precursor. [ J Cell Biol, 2019, 218(2):644-663]	PubMed: 30626721

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SHIPPING AND STORAGE
Selleck products are transported at room temperature. If you receive the product at room temperature, please rest assured, the Selleck Quality Inspection Department has conducted experiments to verify that the normal temperature placement of one month will not affect the biological activity of powder products. After collecting, please store the product according to the requirements described in the datasheet. Most Selleck products are stable under the recommended conditions.

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