This probe is available from Sigma and Cayman Chemical.
Its negative control (LLY-284) is available for purchase from Sigma.
| Probe | Negative control | |
 |
|  |
LLY-283 |
| LLY-284 |
A collaboration between Eli Lilly and the SGC has resulted in the identification of LLY-283, the first potent and selective SAM-competitive chemical probe for PRMT5. The in vitro activity of LLY-283 includes potent inhibition of PRMT5 enzyme activity with IC50 = 20 nM for methylation of an H4R3 derived peptide substrate, and greater than 100-fold selectivity over other histone methyltransferases and non-epigenetic targets. In cellular assays, LLY-283 inhibits the methylation of SmBB’ with IC50 = 25 nM (MCF7 cells; 3 days) and also affects MDM4 splicing with IC50-relative of 40 nM (A375 cells; 3 days).
A closely related compound, LLY-284, is less potent in the biochemical assay and is an ideal control compound for cellular studies.
| Probe | Negative control | |
|
| |
LLY-283 |
| LLY-284 |
| Physical and chemical properties for LLY-283 | |
| Molecular weight | 342.1 |
| Molecular formula | C17H18N4O4 |
| IUPAC name | 2-(5-amino-2,4,9-triaza-bicyclo[4.3.0]nona-1(6),2,4,7-tetraen-9-yl)-5-(hydroxy-phenyl-methyl)-tetrahydro-furan-3,4-diol |
| MollogP | -0.1422 |
| PSA | 98.65 |
| No. of chiral centres | 5 |
| No. of rotatable bonds | 3 |
| No. of hydrogen bond acceptors | 6 |
| No. of hydrogen bond donors | 5 |
| Physical and chemical properties for LLY-284 (Negative Control) | |
| Molecular weight | 342.1 |
| Molecular formula | C17H18N4O4 |
| IUPAC name | 2-(5-amino-2,4,9-triaza-bicyclo[4.3.0]nona-1(6),2,4,7-tetraen-9-yl)-5-(hydroxy-phenyl-methyl)-tetrahydro-furan-3,4-diol |
| MollogP | -0.1422 |
| PSA | 98.65 |
| No. of chiral centres | 5 |
| No. of rotatable bonds | 3 |
| No. of hydrogen bond acceptors | 6 |
| No. of hydrogen bond donors | 5 |
Enzyme inhibition assay for LLY-283 and its negative control (LLY-284). The IC50 values of 22 ± 3 nM (Hill Slope of 1) and 1074 ± 53 nM (Hill Slope of 1.2) were determined for LLY-283 () and LLY-284 (), respectively.
Zahid Q. Bonday, Guillermo S. Cortez, Michael J. Grogan, Stephen Antonysamy, Ken Weichert, Thomas J. Raub, Wayne P. Bocchinfuso, Fengling Li, Steven Kennedy, Binghui Li, Mary M. Mader, Cheryl H. Arrowsmith, Robert M. Campbell, Peter J. Brown, Mohammad S. Eram, Magdalena M. Szewczyk, Masoud Vedadi, Dalia Barsyte-Lovejoy. LLY-283, a Potent and Selective Inhibitor of Arginine Methyltransferase 5, PRMT5, with antitumor activity. ACS Med. Chem. Lett. (2018) DOI: 10.1021/acsmedchemlett.8b00014
PDB 6CKC
Main features
This probe is available from Tocris, Sigma and Cayman Chemical.
The inactive control is available from Sigma.
| Probe | Negative control | |
 |
|  |
BAY-299 |
| BAY-364 |
The BRPF (BRomodomain and PHD Finger) family consists of 3 members BRPF1, BRPF2 (also known as BRD1) and BRPF3 that act as scaffolding proteins to assemble HAT complexes of the MOZ/MORF family (MOZ, Ybf2/Sas3, Sas2, and Tip60). These MYST complexes have a tetrameric core containing BRPF, the tumour suppressor ING and Eaf6/EPC (enhancer of polycomb)-related scaffold subunits. MYST complexes play crucial roles in DNA repair, recombination, and replication as well as in transcription activation. MOZ is frequently translocated in acute myeloid leukemia and has an essential role in development and maintenance of hematopoietic stem cells[1].
Transcription initiation factor TFIID subunits 1 (TAF1) is a component of the STAGA complex, which contains TRRAP, GCN5, TFIID, CBP/P300, mediator and Sp1, TAF1 is susceptible to oncogenic activation by MYC. Moreover, TAF1 has been shown to block p53 activity and inactivation of TAF1 triggers a DNA damage response. Additionally, the TFIID complex is vital to stem cell reprogramming. There have been few reports of selective inhibitors of TAF1 which are needed to help elucidate its biological role and potentially function to inhibit cancer cell growth and survival.
In a collaborative effort Bayer and the SGC have identified and characterised BAY-299 as a potent and selective inhibitor of BRD1 (BROMOscan® [2] IC50 6nM) and the second bromodomain of TAF1 (BROMOscan® [2] IC50 13nM). BAY-299 is selective over other bromodomains; >30-fold selective over the other members of the BRPF family, >30-fold selective over close neighbours BRD9 and ATAD2 and >300-fold selective over BRD4. BAY-299 has <1uM activity in the BRD1 and TAF1 NanoBRET™ cell assays [3]. BAY-299 has a molecular weight of 429.5 and is soluble to 10mg/L in aqueous solutions. The structurally similar BAY-364 (MW 401.4) is inactive against BRD1 (>20uM), has moderate activity against TAF1 (3uM) and is suitable for use as a negative control.
| Probe | Negative control | |
|
| |
BAY-299 |
| BAY-364 |
| Physical and chemical properties for BAY-299 | |
| Molecular weight | 429.2 |
| Molecular formula | C25H23N3O4 |
| IUPAC name | 6-(3-Hydroxypropyl)- |
| MollogP | 4.33 |
| PSA | 62.28 |
| No. of chiral centres | 0 |
| No. of rotatable bonds | 4 |
| No. of hydrogen bond acceptors | 7 |
| No. of hydrogen bond donors | 1 |
| Physical and chemical properties for BAY-364 (Negative Control) | |
| Molecular weight | 401.1 |
| Molecular formula | C23H19N3O4 |
| MollogP | 3.844 |
| PSA | 71.76 |
| No. of chiral centres | 0 |
| No. of rotatable bonds | 4 |
| No. of hydrogen bond acceptors | 7 |
| No. of hydrogen bond donors | 2 |
Selectivity Against Non Target Family
BAY-299 and BAY-364 were screened in the Eurofins CEREP Diversity Profile to measure binding to a range of Receptors, Ion Channels and Enzymes @10 uM. The Inhibition as the mean % of control is shown below, results showing an inhibition or stimulation lower than 50% are considered to represent significant effects. BAY-299 showed binding to A1 receptor (48%), Cl- channel (GABA-gated) (23%), PDE2A1 (49%) and PDE5 (-2%). BAY-364 was clean.
1. Klein BJ, Lalonde ME, Côté J, Yang XJ, Kutateladze TG., Crosstalk between epigenetic readers regulates the MOZ/MORF HAT complexes. Epigenetics. 2014, 9(2), 186-93
2. DiscoverX https://www.discoverx.com/services/drug-discovery-development-services/epigenetic-profiling/bromoscan
3. Promega http://www.promega.com/a/forms/custom-assays/nanobret-protein-interaction.html
4. Benzo-isoquinoline-diones As Potent and Selective Inhibitors of BRPF2 and TAF1/TAF1L Bromodomains Léa Bouché, Clara D. Christ, Stephan Siegel, Amaury E. Fernández-Montalván, Simon J. Holton, Oleg Fedorov, Antonius ter Laak, Tatsuo Sugawara, Detlef Stöckigt, Cynthia Tallant, James M. Bennett, Octovia P Monteiro, Laura Díaz-Sáez, Paulina Siejka, Julia Meier, Vera Puetter, Joerg Weiske, Susanne Müller, Kilian Veit Marcus Huber, Ingo V. Hartung, and Bernard Haendler. J. Med. Chem .April 12, 2017
This probe is available from Tocris and Cayman Chemical.
The control may be requested by clicking here.
| Probe | Negative control | |
 |
|  |
TP-064 |
| TP-064N |
A collaboration between Takeda and the SGC has resulted in the discovery of TP-064, the first potent, selective and cell active chemical probe for PRMT4. The in vitro activity of TP-064 includes inhibition of PRMT4 with IC50 < 10nM for methylation of H3 (1-25) and greater than 100-fold selectivity over other histone methyltransferases and non-epigenetic targets. In cellular assays, TP-064 inhibits the methylation of MED12 with IC50 = 43 nM.
A closely related compound, TP-064N, exhibits no activity in the biochemical and cellular assays, and is an ideal control compound for cellular studies.
| Probe | Negative control | |
|
| |
TP-064 |
| TP-064N |
| Physical and chemical properties for TP-064 | |
| Molecular weight | 458.3 |
| Molecular formula | C28H34N4O2 |
| IUPAC name | (methyl-((2-(1-(2-methylamino-ethyl)-piperidin-4-yl)-pyridin-4-yl)-methyl)-amino)-(3-phenoxy-phenyl)-methanone |
| MollogP | 3.422 |
| PSA | 47.71 |
| No. of chiral centres | 0 |
| No. of rotatable bonds | 10 |
| No. of hydrogen bond acceptors | 6 |
| No. of hydrogen bond donors | 1 |
| Physical and chemical properties for TP-064N (Negative Control) | |
| Molecular weight | 459.3 |
| Molecular formula | C28H33N3O3 |
| IUPAC name | (((2-(1-(2-methoxy-ethyl)-piperidin-4-yl)-pyridin-4-yl)-methyl)-methyl-amino)-(3-phenoxy-phenyl)-methanone |
| MollogP | 4.051 |
| PSA | 44.24 |
| No. of chiral centres | 0 |
| No. of rotatable bonds | 10 |
| No. of hydrogen bond acceptors | 6 |
| No. of hydrogen bond donors | 0 |
Selectivity of TP-064N at 10 μM () and 1 μM () and of TP-064 at 10 μM () and 1 μM () for PRMT1, 3, 4, 5, 6, 7, 8, and 9 as well as for 24 histone and DNA methyltransferases was assessed. Dose response data are presented in the top panel as IC50s (μM).
TP-064 inhibits the dimethylation of PRMT4 substrates. HEK293 cells were treated with indicated concentrations of TP-064 for 3 days and dimethylation levels of BAF155 and MED12 in whole cell extracts were analyzed by western blotting. (B) Quantitation of data in (A). Graphs represent nonlinear curve fits of dimethyl-BAF155 and dimethyl-MED12 signal intensities normalized to total BAF155 or MED12, respectively. Data represent mean ± SEM of two independent experiments prepared in triplicate.
TP-064 (blue) inhibits PRMT4 activity with an IC50 value of < 10 nM under balanced conditions. TP-064N (red) has no effect on PRMT4 activity up to 100 nM. The binding of TP-064 to PRMT4 was confirmed by DSLS with stabilization at about 6°C. No binding was observed with TP-064N
SPR analysis of the TP-064 binding to PRMT4 in the presence of 50 μM SAM. A representative sensorgram (black dots) is shown with the kinetic fit (solid green). A Kd value of 7.1 ± 1.8 nM, with kon = 1.1 ± 0.1 × 105 M−1 s−1 and koff = 0.7 ± 0.1 × 10−3 s−1, was obtained from triplicate experiments. The steady state response (black circles) and 1:1 binding model fitting (red dashed line) is presented.
Mechanism of action of TP-064 was assessed by determining IC50 of both substrates values at various concentrations.
Kazuhide Nakayama, Magdalena M. Szewczyk, Carlo dela Sena, Hong Wu, Aiping Dong, Hong Zeng, Fengling Li, Renato Ferreira de Freitas, Mohammad S. Eram, Matthieu Schapira, Yuji Baba, Mihoko Kunitomo, Douglas R. Cary, Michiko Tawada, Akihiro Ohashi, Yasuhiro Imaeda, Kumar Singh Saikatendu, Charles E. Grimshaw, Masoud Vedadi, Cheryl H. Arrowsmith, Dalia Barsyte-Lovejoy, Atsushi Kiba, Daisuke Tomita and Peter J. Brown TP-064, a potent and selective small molecule inhibitor of PRMT4 for multiple myeloma. Oncotarget 9: 18480-93 (2018)
| Test compound | TP-064 |
| Animal | C.B-17/Icr-scid mouse, female, 7W, fed |
| Formulation | 10% DMSO/10% Cremophor EL /10% PropyleneGlycol (PG)/Distilled Water |
| Administration | i.p., 30 or 100 mg/kg as free body |
| Test items | Plasma |
| Sampling time | 0.25, 0.5, 1, 2, 4, 8, 24 hours after administration |
Main features
This probe is available from Tocris, Cayman Chemical and Sigma.
Its negative control (TP-472N) is available for purchase from Tocris and Sigma.
| Probe | Negative control | |
 |
|  |
TP-472 |
| TP-472N |
The bromodomain containing protein 9 (BRD9) has been reported as a component of the switch/sucrose non-fermentable (SWI/SNF) brahma-related gene 1-associated factor (BAF) complex, which plays a key role in chromatin remodelling and transcription control [1] although the precise biological role is unknown. The highly homologous bromodomain containing protein 7 (BRD7) is a component of the SWI/SNF polybromo-associated BAF (PBAF) complex and has been proposed as a tumor suppressor [2].
In a collaborative effort Takeda and the SGC have identified and characterised TP-472 as a BRD9/7 probe. This molecule is of a different chemotype to the BRD9/7 probes previously available, has a structurally similar negative control available (TP-472N), has a good PK profile and is suitable for in vivo applications and is synthetically tractable.
TP-472 has excellent potency (BRD9 KD 33nM; BRD7 340nM by ITC), selectivity >30 fold over all other bromodomain family members except BRD7 and is cell active (EC50 320 nM in a BRD9 NanoBRET assay). The negative control TP472N is inactive against other bromodomains (>20uM against BRD9)
| Probe | Negative control | |
|
| |
TP-472 |
| TP-472N |
| Physical and chemical properties for TP-472 | |
| Molecular weight | 333.1 |
| Molecular formula | C20H19N3O2 |
| IUPAC name | 1-(7-(5-(cyclopropylamino-formyl)-2-methyl-phenyl)-1,5-diaza-bicyclo[4.3.0]nona-2,4,6,8-tetraen-9-yl)-ethanone |
| MollogP | 3.102 |
| PSA | 46.85 |
| No. of chiral centres | 0 |
| No. of rotatable bonds | 5 |
| No. of hydrogen bond acceptors | 5 |
| No. of hydrogen bond donors | 1 |
| Physical and chemical properties for TP-472N (Negative Control) | |
| Molecular weight | 306.1 |
| Molecular formula | C19H18N2O2 |
| IUPAC name | 1-(7-(2-(cyclopropyl-methoxy)-phenyl)-1,5-diaza-bicyclo[4.3.0]nona-2,4,6,8-tetraen-9-yl)-ethanone |
| MollogP | 3.673 |
| PSA | 30.22 |
| No. of chiral centres | 0 |
| No. of rotatable bonds | 5 |
| No. of hydrogen bond acceptors | 4 |
| No. of hydrogen bond donors | 0 |
Selectivity beyond target family
TP-472 and TP-472N were screened in the Eurofins CEREP Diversity Profile to measure binding to a range of Receptors, Ion Channels and enzymes @10 uM. The Inhibition as the mean % of control is shown below, results showing an inhibition or stimulation lower than 50% are considered to represent significant effects. TP-472 showed binding to Adenosine A1 receptor (14%), Benzodiazepine receptor (47%), PDE2A1 (h) (25%), PDE3A (h) (48%) and PDE4D2 (h) (28%). TP-472N showed binding to A1 receptor (35%), A3 receptor (23%), Melatonin receptor MT1 (47%) and Cl- channel (GABA-gated) (20%).
For a second year, SGC Toronto hosted a team of senior law students from the University of Toronto's Faculty of Law as part of an externship clinic to help SGC draft new agreements protecting our open access ethos and promoting our open science.
Oxford, UK (April 23rd, 2016) -FOP patient groups have announced an extension of their support to the FOP research team in SGC Oxford led by Dr Alex Bullock since 2008. The team, currently comprising two postdoctoral fellows, is working towards developing a treatment for the rare monogenic disease Fibrodysplasia Ossificans Progressiva (FOP), a disorder that causes severely debilitating extraskeletal bone formation and impairs movement.
Farnborough, Hampshire UK (April 13th, 2016) -We are delighted to announce a groundbreaking partnership between The Brain Tumour Charity and the Structural Genomics Consortium (SGC), an international group of academic and industrial researchers with a strong track record in uncovering new drug targets and initiating clinical trials within unusually short timeframes.
Rachel Harding, a researcher at SGC Toronto, appears on Global News discussing her open science experiment. She's welcomed the world to view her research progress in real-time by placing her lab notes online and explaining her results on her blog Lab Scribbles.
Fremont, CA – February 29, 2016 – DiscoverX Corporation, the leading supplier of innovative cell-based assays and services for drug discovery and development, today announced its partnership with the Structural Genomics Consortium (SGC), a public-private organization established in 2003, to develop selective and potent chemical probes for the unexplored human kinome and to promote open sharing of these probes with the scientific community.
TORONTO, February 26, 2016 — University of Toronto researcher Rachel Harding will be the first known biomedical researcher to welcome the world to review her lab notes in real time. The post-doctoral fellow with U of T’s Structural Genomics Consortium (SGC) is also explaining her findings to the general public through her blog. She hopes her open approach will accelerate research into Huntington’s disease.
