TABLE 2

Molecular interaction of LMD-584, LMD-902, LMD-268, and LMD-174 with CCR8

The whole panel of CCR8 mutations was screened for the ability of the four additional nonpeptide agonists to activate every single mutation. The CCR8 receptors were expressed in transiently transfected COS-7 cells, and the activation was measured by means of accumulated inositol phosphates. The table shows the potency (both as Log EC50 and as EC50). The differences between the potency of each compound on a given mutation and the potency of each ligand on wt CCR8 are shown in the columns named “-Fold.” The number of experiments is given as n. The receptor residues are given according to the Schwartz (S) and Ballesteros/Weinstein (B-W) nomenclatures (see footnote 1 in the text).

Residue LMD-584 LMD-902 LMD-268 LMD-174
Position Number
S B-W EC50 EC50 -Fold n EC50 EC50 -Fold n EC50 EC50 -Fold n EC50 EC50 -Fold n
Log nM Log nM Log nM Log nM
wt wt −7.5 ± 0.07 29 1.0 15 −7.8 ± 0.10 15 1.0 8 −8.0 ± 0.05 9.0 1.0 4 −7.5 ± 0.05 30 1.0 15
I:07 1.39 Y42A −6.9 ± 0.04 131 4.6 3 −6.7 ± 0.18 178 12 3 −6.8 ± 0.22 172 19 3 −6.7 ± 0.14 199 6.6 3
I:11 1.43 F46A −6.9 ± 0.05 126 4.4 3 −7.3 ± 0.07 50 3.4 3 −7.4 ± 0.06 43 4.8 2 −7.3 ± 0.18 52 1.7 3
II:13 2.53 F84A −8.2 ± 0.25 7.0 0.24 4 −7.9 ± 0.03 12 0.85 3 −8.3 ± 0.06 4.8 0.54 4 −6.9 ± 0.28 133 4.4 4
II:17 2.57 F88A −6.6 ± 0.19 251 8.8 3 −6.7 ± 0.20 197 14 4 −7.5 ± 0.23 31 3.5 3 −7.0 ± 0.05 94 3.1 3
II:20 2.60 Q91W −6.4 ± 0.36 368 13 3 −6.7 ± 0.13 180 12 3 −6.8 ± 0.25 161 18 3 −6.7 ± 0.17 201 6.7 3
III:04 3.28 V109A −7.5 ± 0.26 34 1.2 3 −8.1 ± 0.05 8 0.52 3 −8.6 ± 0.11 2.6 0.29 3 −7.5 ± 0.18 33 1.1 3
III:05 3.29 S110A −8.2 ± 0.38 6.0 0.21 4 −8.7 ± 0.34 2 0.15 3 −8.8 ± 0.11 1.6 0.18 6 −8.4 ± 0.15 3.6 0.12 4
III:05 3.29 S110H <−5 >10,000 >323 3 <−5 ± 0.00 >10,000 >689 2 <−5 >10,000 >1100 2 <−5 >10,000 >323 3
III:05 3.29 S110W <−5 >10,000 >323 4 <−5 ± 0.00 >10,000 >689 2 <−5 >10,000 >1100 3 <−5 >10,000 >323 3
III:07 3.31 F112A −7.3 ± 0.08 51 1.8 4 −7.9 ± 0.09 13 0.88 4 −8.1 ± 0.08 7.1 0.79 6 −7.4 ± 0.08 37 1.2 4
III:08 3.32 Y113A <−5 ± 0.00 >10,000 >323 3 >−5 >10,000 >689 3 <−5 >10,000 >1100 3 <−5 ± >10,000 >332 3
III:09 3.33 Y114A −6.4 ± 0.32 430 15 3 −6.7 ± 0.11 191 13 3 −6.2 ± 0.06 579 64 3 −6.0 ± 0.11 945 31 3
III:18 3.42 F123A −7.7 ± 0.14 21 0.73 3 −7.9 ± 0.02 12 0.85 3 −8.3 ± 0.17 5.0 0.56 5 −7.7 ± 0.07 19 0.63 3
IV:24 4.64 Y172A −7.3 ± 0.19 56 1.9 4 −7.5 ± 0.15 31 2.1 3 −7.6 ± 0.20 28 3.1 3 −7.0 ± 0.04 93 3.1 5
V:−02 5.34 K193A −7.5 ± 0.18 31 1.1 4 −7.8 ± 0.52 17 1.2 2 −8.3 ± 0.17 4.7 0.52 3 −7.5 ± 0.06 30 1.0 3
V:−01 5.33 W194A −7.9 ± 0.15 12 0.44 4 −8.4 ± 0.40 4 0.29 3 −8.2 ± 0.18 6 0.71 3 −7.5 ± 0.11 32 1.1 3
V:01 5.35 K195A −7.7 ± 0.17 20 0.70 5 −8.2 ± 0.25 7 0.48 3 −9.7 ± 0.20 0.22 0.02 2 −7.5 ± 0.05 30 1.0 4
V:12 5.46 G206A −7.3 ± 0.21 47 1.6 3 −7.9 ± 0.04 12 0.83 3 −8.8 0.10 2 0.19 2 −7.20.5 62 2.1 3
V:12 5.46 G206W N.A. 3 N.A. 3 N.A. 3 3
VI:13 6.48 W251A −7.7 ± 0.38 20 0.70 3 −8.30.14 4.7 0.32 2 −7.4 ± 0.25 42 4.7 4 −7.5 ± 0.41 29 0.97 3
VI:13 6.48 W251Q −7.7 ± 0.41 22 0.77 3 −7.80.14 15 1.0 2 −8.0 ± 0.39 10 1.1 3 −7.5 ± 0.41 29 0.97 3
VI:16 6.51 F254A −6.6 ± 0.19 278 9.7 4 −7.1 ± 0.04 80 5.5 3 −8.0 ± 0.12 10 1.2 3 −7.4 ± 0.13 42 1.4 3
VI:20 6.55 L258A −6.5 ± 0.29 329 12 3 −7.5 ± 0.08 35 2.4 3 −7.2 ± 0.05 65 7.2 5 −7.2 ± 0.08 64 2.1 3
VI:24 6.59 S262A −7.2 ± 0.13 61 2.1 3 −7.9 ± 0.10 13 0.86 3 −8.1 ± 0.28 8.9 1.0 5 −7.4 ± 0.02 39 1.3 3
VII:02 7.35 T282A −7.1 ± 0.07 77 2.7 3 −7.5 ± 0.10 31 2.1 3 −7.4 ± 0.08 40 4.4 3 −7.0 ± 0.06 93 3.1 3
VII:03 7.36 H283A −7.5 ± 0.08 30 1.0 3 −7.5 ± 0.06 34 2.4 3 −7.8 ± 0.11 17 1.9 3 −7.3 ± 0.09 46 1.5 3
VII:06 7.39 E286A <−5 >10,000 >323 3 <−5 >10,000 >689 3 <−5 >10,000 >1100 5 <−5 >10,000 >332 3
VII:09 7.42 S289A −6.7 ± 0.27 211 7.4 3 −7.5 ± 0.04 34 2.3 3 −7.8 ± 0.08 17 1.9 3 −7.4 ± 0.02 36 1.2 3
VII:10 7.43 F290A −7.4 ± 0.24 41 1.4 3 −7.6 ± 0.31 26 1.8 3 −7.7 ± 0.34 21 2.3 4 −7.5 ± 0.46 29 1.0 3
  • N.A., no activation at all.