MZ1

Reagent Authentication Certificate:

Chemical Probes Portal MZ1.pdf

Protein target name:

BRD4

Mechanism of action:

DegraderPROTAC

Recommended Concentration for use in cells:

100-250 nM

Probe Availability

Tocris (1797406-69-9)

Probe Information

Target and Probe Information

Target Details

Target class:

Epigenetics

Subclass:

Bromodomain

HUGO ID:

HGNC:13575

Entrez gene ID:

23476

UNIPROT ID:

O60885

Target alias:

BRD4, CAP, bromodomain containing 4, chromosome-associated protein, HUNK1, HUNKI, MCAP

Probe Details

Chemical Probes Portal ID:

CPP770

Probe alias:

PROTAC

PubChem CID:

122201421

ChEMBL ID:

CHEMBL4226570

SMILES string:

O[C@@H]1C[C@@H](C(NCC2=CC=C(C3=C(C)N=CS3)C=C2)=O)N(C([C@H](C(C)(C)C)NC(COCCOCCOCCNC(C[C@@H]4N=C(C5=CC=C(Cl)C=C5)C6=C(SC(C)=C6C)N7C4=NN=C7C)=O)=O)=O)C1

InChi Key:

PTAMRJLIOCHJMQ-PYNGZGNASA-N

Control compound:

cisMZ1

Orthogonal probe:

dBET1ARV-825dBET6

Physico-chemical properties

Solubility - preferred solvent:

DMSO

Solubility - concentration:

10 mM

Potential reactivities:

Unknown

NMR:

1H-NMR (CDCl3, 500 MHz) δ 0.97 (s, 9 H), 1.66 (s, 3 H), 2.12-2.17 (m, 1 H), 2.39 (s, 3 H), 2.43-2.49 (m, 1 H), 2.51 (s, 3 H), 2.61 (s, 3 H), 3.31-3.35 (m, 2 H), 3.47-3.74 (m, 13 H), 4.11-4.14 (m, 2 H), 4.27-4.33 (m, 2 H), 4.49-4.55 (m, 2 H), 4.65-4.69 (m, 3 H), 4.84 (t, 1 H, J(H,H)= 7.9 Hz), 7.23-7.25 (m, 1 H), 7.29-7.39 (m, 9 H), 7.91-7.94 (m, 1 H), 8.67 (s, 1 H); 13C-NMR (CDCl3, 101 MHz) δ 11.9, 13.2, 14.6, 16.2, 26.6, 35.6, 36.4, 38.2, 39.9, 43.3, 54.3, 56.9, 57.3, 59.0, 70.1-70.9, 71.7, 128.2, 128.9, 129.6, 130.1, 130.9, 131.0, 131.2, 131.8, 132.0, 136.7, 136.8, 138.4, 148.6, 149.9, 150.4, 156.0, 164.0, 171.0, 171.3, 171.4; HRMS m/z calc. for C49H61ClN9O8S2 [M+H+] 1002.3768, found 1002.3786.

Storage Recommendations

Storage (dry powder):

At least 6 months at room temperature, we recommend storage at -20 C for longer periods of time.

Storage (solution):

-20 oC for at least 3 months

Storage sensitivities:

Unknown

Validation in Cells and Model Organisms

In vitro validation

On target activity

Potency:

BRD4(BD1) 382 nM, (BD2) 120 nM

Potency assay:

Isothermal calorimetry (KD) Western blot (DC50=5 nM) DC50 refers to the half maximal degradation concentration.

Structure-activity relationship:

From Zengerle et al, ACS Chem Biol, 2015, 10, 1770 (DOI: 10.1021/acschembio.5b00216); MZ1 (PEG3), MZ2 (PEG4) and MZ3 (Phe-PEG3) were all evaluated for their mechanism and selectivity of degradation against BRD4 vs BRD2 and BRD3

Off target activity

Potency (off target):

Off target protein and potency:

BRD2(BD1) 307 nM, (BD2) 228 nM BRD3(BD1) 119 nM, (BD2) 115 nMVBC 149 nM

Potency assay (off target):

Isothermal calorimetry (KD) Western blot (DC50): DC50=70 nM (BRD2), DC50=70 nM (BRD3)

Probe Selectivity in Vitro:

Not available

In cell validation

On target activity in cells

Potency in cells:

DC50

5 nM

Potency assay (cells):

In HeLa cells, MZ1 led to time and concentration dependent degradation of BRD4. BRD4 was preferentially degraded over BRD2 and BRD3, especially at low concentration (0.1-1 uM). In U2OS cells, MZ1 led to the degradation of a BRD4-GFP reporter.

Target engagement assay (cells):

Not done

Off target activity in cells

Potency in cells, off target:

DC50

Off target protein and potency (cells):

BRD2 and BRD3, 70 nM

Potency assay, off target (cells):

Western blot

Probe Selectivity in Cell:

Not done

Toxicity

Cytoxicity assay:

Yes

Notes on cytotoxicity:

MZ1 is not cytotoxic to HeLa and U2OS cells at concentrations <10 uM.

Primary Reference

Reference (doi):

10.1021/acschembio.5b00216

Reference (PMID):

26035625

Endorsed probe

The Chemical Probes Portal endorses this compound as chemical probe for use in cells.

SAB Rating

Rating for use in Cells

4 review(s)

Rating for use in Model Organisms

no rating

SAB Members

SAB Comments

MZ1 is a PROTAC compound, made by conjugating the pan-BET inhibitor JQ1 via a PEG linker to a a potent ligand for the VHL E3 ligase. This compound preferentially removes a single member of the BET family, BRD4, over BRD2 and BRD3 at low concentrations. The downstream gene expression pattern resulting from treatment with MZ1 is similar to JQ1 inhibition for BRD4-dependent genes MYC, P21, and AREG but not for FAS, FGFR1, and TYRO3. Compound was not tested in an in vivo setting

Dec 26 2016 - 12:37pm

MZ1 is a compound based on the pan-BET inhibitor JQ1 linked to a ligand for the E3 ubiquitin ligase. In a narrow concentration and time range, MZ1 preferentially leads to degradation of BRD4 over the BET proteins BRD2 and BRD3. The differential effect to the parent compound JQ1 is reflected in the gene expression pattern upon treatment of HeLa cells with either compound, which reveals differences in the expression of some genes like TYRO3 or FGFR1.

Jan 24 2017 - 11:17am

A 3D crystal structure of MZ1 bound to both VHL and the second bromodomain of BRD4 was solved and published (Gadd et al. Nat Chem Biol, 13 March 2017, doi:10.1038/nchembio.2329) with thermodynamic binding data of ternary complex formation by isothermal titration calorimetry. The RSCB PDB entry code of the structure is 53T5.

Apr 7 2017 - 4:52pm

This is a very interesting PROTAC version of JQ1. Because of the preference for BRD4 at 100-250 nM, it would be great to compare cell toxicity between MZ1 and JQ1.

Apr 10 2017 - 11:47am

MZ1 exhibits potent cytotoxicity and profound anti-proliferative effects in AML cell lines such as Mv4;11 (pEC50 = 7.6), more potent and with greater Emax than (+)-JQ1 (pEC50 = 6.5).

Apr 28 2017 - 10:10pm

Further SAR data for MZ1 and other analogues with different PEG linkers (MZ4 = PEG2; MZ2 = PEG4) were published (Chan et al. J Med Chem. DOI: 10.1021/acs.jmedchem.6b01912). SAR data include BET protein degradation profiles in HeLa cells (pDC50s and Dmax); cellular cytotoxicity profiles in AML lines Mv4;11 and HL-60 (pEC50s, and maximal response to baseline level at the higher concentrations: Emax); and Brd4 and c-Myc percent depletion in AML cell lines. Application of MZ1 as a chemical probe to dissect individual distinct functions of BRD2 and BRD4 in pluripotent stem cells (PSCs) to coordinate pluripotent exit is reported (Fernandez-Alonso, EMBO Rep. DOI 10.15252/embr.201643534)

Jul 26 2017 - 11:42pm

Plasma stability ( In vitro Half life) of MZ1 > 180 min i.e. MZ1 is fully stable in plasma within the assay window (3 h).

Jan 26 2018 - 9:55pm

MZ1

Probe Availability

Probe Information

Target Details

BRD4, CAP, bromodomain containing 4, chromosome-associated protein, HUNK1, HUNKI, MCAP

Probe Details

Physico-chemical properties

In vitro validation

In cell validation

Toxicity

Primary Reference

Supporting Organizations

Interested in supporting the portal?

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MZ1

Probe Availability

Probe Information

Target Details

BRD4, CAP, bromodomain containing 4, chromosome-associated protein, HUNK1, HUNKI, MCAP

Probe Details

Physico-chemical properties

In vitro validation

In cell validation

Toxicity

Primary Reference

Supporting Organizations

Interested in supporting the portal?