TEAD

MYF-01-37 (10 μM; 24 hours) decreases the expression of the canonical YAP target gene CTGF in PC-9 cells and inhibits the direct YAP/TEAD response in HEK 293T cells [1]. , 1, 10, 100 μM) barely affects a few EGFR mutant NCSLC cell lines' capacity to survive [1]. In comparison to OT alone, MYF-01-37 (10 μM; 10 days) in conjunction with OT (osimertinib and trametinib combo) dramatically decreased quiescent cells [1].

TEAD2 protein was incubated with DMSO or a 20-fold molar excess of MYF-01-37 for 6 hours at 37 °C. Reactions were then analyzed by LC-MS using a Shimadzu autosampler and LC coupled to an LTQ ion trap mass spectrometer. Protein was injected onto a self packed column (0.5 mm I.D., packed 5 cm POROS 50R2), desalted for 4 minutes with 100% A (A=0.2 M acetic acid in water), eluted with a gradient (0–100% B in 1 minute; A=0.2 M acetic acid in water, B=0.2 M acetic acid in acetonitrile), and introduced to the mass spectrometer by electrospray ionization (spray voltage=4.5 kV). The mass spectrometer acquired full scan MS data (m/z 300–2000). Mass spectra were deconvoluted using MagTran version 1.03b2 (Zhang and Marshall, 1998).[1]

To identify the site of modification, labeled protein was diluted 1:1 with 100 mM ammonium bicarbonate, reduced with 10 mM DTT at 56 °C for 30 minutes, alkylated with 22.5 mM IAA for 30 minutes at room temperature, and then digested with trypsin overnight at 37 °C. Tryptic peptides were desalted by C18, dried by vacuum centrifugation, reconstituted in 5% MeCN, 0.1% trifluoroacetic acid, and analyzed by nanoLC-ion mobility MS/MS using a NanoAcquity UPLC system interfaced to a timsTOF Pro mass spectrometer. Peptides were injected onto a self-packed pre-column (4 cm POROS10R2), resolved on an analytical column (30 μm I.D. × 50 cm Monitor C18; 10–60% B in 40 minutes; A = 0.2 M acetic acid in water, B = 0.2 M acetic acid in acetonitrile) and introduced to the mass spectrometer by electrospray ionization using a captive spray ion source (spray voltage = 2 kV). The mass spectrometer collected ion mobility MS spectra over a mass range of m/z 100–1700 and 1/k0 of 0.6 to 1.6, and then performed 10 cycles of PASEF MS/MS with a target intensity of 20k and a threshold of 250. Active exclusion was enabled with a release time of 0.4 minutes. Raw data was converted to .mgf using the tdf to mgf converter, and searched using Mascot 2.6.1 against a forward reversed human refseq database (NCBI). Search parameters specified a precursor mass tolerance of 20 ppm, a product ion tolerance of 50 mmu, fixed carbamidomethylation of cysteine, and variable oxidation of methionine as well as variable MYF-01-37 modification of cysteine. Search results were downloaded and converted to xls using multiplierz software (Alexander et al., 2017) , and peptide fragment ions were assigned using mzStudio (Ficarro et al., 2017). Inhibitor related fragment ions were assigned as described (Ficarro et al., 2016).[1]

MYF-01-37 competition pulldown[1]
Cells were treated with test compounds for 6 h at the indicated doses. Total cell lysates were prepared using pulldown buffer (50 mM Tris, pH 7.4, 250 mM NaCl, 5 mM EDTA, 50 mM NaF, 1 mM Na3VO4, 1% NP40, 0.02% NaN2, 1 mM PMSF and protease inhibitor cocktail). To pull down TEADs, 1mg of total protein was combined with biotinylated MYF-01-37 at 1 μM and rotated at 4 °C for 6 hrs followed by RT for 1 h. Subsequently, streptavidin agarose resin (30 μl 50% slurry) was added and samples rotated for another 2 h at 4 °C. The resin was subsequently washed 3x with pulldown buffer, and TEADs released from the resin by boiling for 10 min in 2x gel loading buffer and resolved by western blotting. As a loading control, 25 μg of total protein was used.

[1]. Treatment-Induced Tumor Dormancy through YAP-Mediated Transcriptional Reprogramming of the Apoptotic Pathway. Cancer Cell. 2020 Jan 13;37(1):104-122.e12.

Eradicating tumor dormancy that develops following epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI) treatment of EGFR-mutant non-small cell lung cancer, is an attractive therapeutic strategy but the mechanisms governing this process are poorly understood. Blockade of ERK1/2 reactivation following EGFR TKI treatment by combined EGFR/MEK inhibition uncovers cells that survive by entering a senescence-like dormant state characterized by high YAP/TEAD activity. YAP/TEAD engage the epithelial-to-mesenchymal transition transcription factor SLUG to directly repress pro-apoptotic BMF, limiting drug-induced apoptosis. Pharmacological co-inhibition of YAP and TEAD, or genetic deletion of YAP1, all deplete dormant cells by enhancing EGFR/MEK inhibition-induced apoptosis. Enhancing the initial efficacy of targeted therapies could ultimately lead to prolonged treatment responses in cancer patients.[1]

C15H17F3N2O

298.3035

298.13

C, 60.40; H, 5.74; F, 19.11; N, 9.39; O, 5.36

2416417-65-5

146567795

White to off-white solid powder

3.3

1

5

3

21

410

0

MTBDEVWWJBIUOK-UHFFFAOYSA-N

InChI=1S/C15H17F3N2O/c1-3-13(21)20-8-7-14(2,10-20)19-12-6-4-5-11(9-12)15(16,17)18/h3-6,9,19H,1,7-8,10H2,2H3

1-(3-methyl-3-((3-(trifluoromethyl)phenyl)amino)pyrrolidin-1-yl)prop-2-en-1-one

MYF 0137; MYF-0137; MYF0137; MYF-01-37; 2416417-65-5; 1-(3-Methyl-3-((3-(trifluoromethyl)phenyl)amino)pyrrolidin-1-yl)prop-2-en-1-one; 1-[3-Methyl-3-[3-(trifluoromethyl)anilino]pyrrolidin-1-yl]prop-2-en-1-one; 1-(3-methyl-3-{[3-(trifluoromethyl)phenyl]amino}pyrrolidin-1-yl)prop-2-en-1-one; starbld0039701; SCHEMBL21958179; MTBDEVWWJBIUOK-UHFFFAOYSA-N; MYF 01-37; MYF-01-37; MYF01-37

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

DMSO : ~250 mg/mL (~838.08 mM)

Solubility in Formulation 1: ≥ 2.08 mg/mL (6.97 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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Solubility in Formulation 2: ≥ 2.08 mg/mL (6.97 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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 (Please use freshly prepared in vivo formulations for optimal results.)