Table 1. Potency and selectivity of cyclic peptide hits from the RaPID system.
|
IC50
(nM) |
|||||
|---|---|---|---|---|---|
|
DTyr-Library |
LTyr-Library |
||||
| CP1 | CP2 | CP3 | CP4 | CP5 | |
| KDM4A | >105 | 42 | >105 | 20 | 313 |
| KDM4B | — | 33 | — | 6 | 472 |
| KDM4C | — | 39 | >105 | 17 | 123 |
| KDM4D | — | 6,270 | — | 6,260 | >104 |
| KDM4E | — | 9,200 | — | 4,700 | 8,900 |
| KDM2A | — | >104 | — | >104 | >104 |
| KDM3A | — | >104 | — | 9,900 | >104 |
| KDM5C | — | >104 | — | >104 | >104 |
| KDM6B | — | 6,800 | — | 7,200 | >104 |
| KDM1A | — | >104 | — | >104 | >104 |
| PHD2 | — | >106 | — | >106 | >106 |
| FIH | — | >106 | — | >106 | >106 |
| KDM4A Binding | |||||
| Kd (nM) | 29.8 | 36.0 | 173 | ||
| kon (1/Ms) | 1.37 × 105 | 2.18 × 104 | 6.2 × 104 | ||
| kdiss (1/s) | 4.07 × 10−3 | 7.84 × 10−4 | 1.07 × 10−2 | ||
2OG, 2-oxoglutarate; FIH, factor inhibiting HIF; IC50, half-maximal inhibitory concentration; KDM, histone demethylase; LSD, lysine-specific demethylase; MALDI–TOF, matrix-assisted laser desorption/ionization–time of flight; MS, mass spectrometry; PHD2, prolyl hydroxylase domain 2; RaPID, Random nonstandard Peptides Integrated Discovery.
KDM IC50 values were determined using AlphaScreen, except for KDM1A/LSD1 where a fluorescence-based assay was used. MALDI–TOF MS assays were used for counterscreening against other 2OG oxygenases. Binding constants for KDM4A were measured using biolayer interferometry.