Project Description
Mechanism of interaction of Arginase-1 inhibitors
Understanding the impact of different assay conditions
- Arginase-1 is a target for cancer immunotherapy
- N-terminal his-tagged Arginase was covalently immobilized on a Ni-NTA chip
Binding kinetics of ABH, nor-NOHA and CB-1158 were determined at pH 7.4 and pH 9.5 using SPR, while their potency and stabilizing effects were evaluated in an enzyme activity assay and a thermal shift assay
- At pH 9.5, ABH is the most potent inhibitor in the activity assay, and has the highest affinity for Arginase-1 as measured by SPR. Moreover, the IC50 of 22 nM and KD of 27 nM are highly comparable
- At pH 7.4, CB-1158 has the most favorable characteristics in all three assay formats
- Based on crystal structure analysis of Arginase-1 in complex with ABH at pH 7 and 9, a more symmetrical coordination structure presumably explains the increased potency of ABH at higher pH
Inhibition assay | Thermal shift assay | SPR assay | |||||
Inhibitor | IC50 (nM) | ΔTm (°C) | ka (M-1 s-1) | kd (s-1) | KD (nM) | τ (s) | |
pH 9.5 | ABH | 22 | 3.5 | 5.1 × 103 | 1.4 × 10-4 | 27 | 7200 |
nor-NOHA | 109 | 4.7 | 3.6 × 104 | 6.2 × 10-3 | 173 | 160 | |
CB-1158 | 132 | 5.6 | 1.3 × 103 | 9.2 × 10-5 | 72 | 11000 | |
pH 7.4 | ABH | 184 | 4.5 | 3.6 × 104 | 9.9 × 10-3 | 797 | 100 |
nor-NOHA | 59 | 4.8 | 3.6 × 104 | 1.9 × 10-2 | 1497 | 54 | |
CB-1158 | 8.6 | 8.1 | 4.8 × 103 | 1.8 × 10-4 | 38 | 5500 |