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. Author manuscript; available in PMC: 2019 Sep 21.
Published in final edited form as: ACS Chem Biol. 2018 Aug 31;13(9):2438–2448. doi: 10.1021/acschembio.7b00638

Table 2.

Activity profile of inhibitors with diazepinone scaffold

graphic file with name nihms-997138-t0006.jpg
Compound ID R1 R2 R3 X Biochemical IC50 (BRD4, μM)a Biochemical IC50 (BRD4, μM)b Enzymatic IC50 (ERK5, μM)c Enzymatic IC50 (LRRK2, μM)d Viability IC50(BRD4, μM)e Cellular EC50 (ERK5, μM)f Inhibitor classificationg Co-crystal (pdb number)
XMD11–50 (LRRK2-IN-1) Me Me Me N 0.703 ± 0.017 1.04 ± 0.18 0.114 ±0.011 0.0034 ±0.0017 1.989 ± 0.299 0.16 ± 0.04 BRD4/LRRK2 Roco4(4YZM)32 BRD4 (5WA5)
JWG-048 Et Me Me N 1.33 ± 0.05 2.41 ± 0.23 0.173 ±0.037 0.0044 ±0.0005 2.625 ± 0.540 0.022 ± 0.004 LRRK2 BRD4 (5W55)
JWG-046 i-Pr Me Me N 4.59 ± 0.14 5.14 ± 0.59 0.125 ±0.026 0.054 ± 0.004 ND 0.014 ± 0.004 ERK5/LRRK2 BRD4 (6CD4)
JWG-071 s-Bu Me Me N 5.42 ± 0.18 6.31 ± 1.93 0.088 ±0.005 0.109 ± 0.014 >10 0.020 ± 0.003 ERK5/LRRK2 BRD4 (6CJ1)
JWG-069 cyclobutyl Me Me N 0.201 ± 0.007 0.41 ± 0.01 0.075 ±0.009 0.021 ± 0.001 0.116 ± 0.001 0.019 ± 0.005 BRD4/ERK5/LRRK2 BRD4 (6CIY)
XMD17–26 cyclopentyl Me Me N 0.760 ± 0.012 0.75 ± 0.11 0.082 ±0.009 0.095 ± 0.010 0.664 ± 0.014 0.08 ± 0.02 BRD4/ERK5/LRRK2 ERK5 (4B99)21 BRD4 (5CD5)
JWG-112 s-Bu H Me N 25.7 ± 1.8 >40 >10 1.59 ± 0.37 ND ND No activity
JWG-049 i-Pr Me Et N 2.50 ± 0.07 3.53 ± 0.82 0.069 ±0.007 0.114 ± 0.011 ND 0.025 ± 0.007 ERK5/LRRK2
JWG-114 i-Pr Me i-Pr N 0.862 ± 0.065 1.08 ± 0.13 0.421 ±0.007 0.235 ± 0.034 ND ND BRD4/weak LRRK2
XMD17–109 (ERK5-IN-1) cyclopentyl Me Et N 0.217 ± 0.008 0.68 ± 0.03 0.162 ±0.006 0.171 ± 0.030 ND 0.09 ± 0.03 BRD4/ERK5/LRRK2
JWG-047 cyclopentyl Me i-Pr N 0.135 ± 0.004 0.30 ± 0.05 0.160 ±0.008 0.296 ± 0.031 0.465 ± 0.012 0.032 ± 0.009 BRD4/ERK5/LRRK2 BRD4 (6CIS)
AX15836 SO2Me Me Et N >100 >40 0.012 ±0.002 0.942 ± 0.569 ND 0.036 ± 0.004 ERK5
DFCI-2–208 Me Me Me C 5.21 ± 0.14 4.64 ± 0.65 >10 >10 ND ND No activity
JWG-115 cyclopentyl Me i-Pr C 1.08 ± 0.05 1.10 ± 0.05 >10 > 3.3 ND ND BRD4 (weak)
(+)-JQ-1 0.064 0.12 >10 >10 0.228 ND BRD4
a

IC50 values were measured by AlphaScreen™ binding assay and reported as the average of 2 replicates ± SE. Experiments were conducted in Dana-Farber Cancer Institute.

b

IC50 values were measured by AlphaScreen™ binding assay and reported as the average of 4 replicates ± SD. Experiments were conducted in University of Oxford.

c

IC50 values were measured by in vitro assay and reported as the average of 2 replicates ± SD.

d

IC50 values were measured using Adapta assay format (ThermoFisher Scientific) and reported as the average of 2 replicates ± SD.

e

IC50 were determined in 797 NUT-midline carcinoma cells by cell numbers using Cell Titer Glo and reported as the average of 3 replicates ± SD.

f

EC50 values were measured by EGF-stimulated autophosphorylation of ERK5 in Hela cells and reported as the average of 2 replicates ± SD.

g

Considering that weak BET bromodomain inhibition has been shown to result in phenotypic responses, a BRD4 activity within 20 fold of JQ1 was defined as BET active. 0.3 μM was chosen as the threshold for kinase activity. A 10-fold activity difference among kinases was considered selective.