JNJ-8003
Probe criteria
| Inhibitor/agonist potency: goal is < 100 nM (IC50, Kd) | Surpasses criterion: Inhibition of RNA synthesis activity of the recombinant RSV L + P in a biotinylated primer extension Flashplate assay: IC50 = 0.67 nM (n = 4);
RSV L+P primer extension assay testing RNA-dependent RNA polymerase (RdRp) activity: IC50 = 0.73 nM (n = 18) Thermal shift assay (purified RSV L + P complex): deltaTm = 6 °C Surface plasmon resonance (SPR) using RSV L + P complex: Kd = 0.84 nM, on-rate = 2.16 μM−1 s−1, slow off-rate = 0.0000181 s−1 |
| Selectivity within target family: > 30-fold | Virus panel (reporter assay): Only minor activity observed against the human metapneumovirus (strain: CAN97-83a, cells: LLC-MK2) which belongs to the same genus: EC50 = 88 nM; PIV-1 (strain: Washington/20993/1964, cells: LLC-MK2, EC50 = 13 µM), PIV-3 (strains: JS, cells: LLC-MK2, EC50 = 11 µM), VSV (strains: Indiana, cells: A549, EC50 > 25 µM), Human: DNA Pol α (IC50 >100 µM), DNA Pol β (IC50 >100 µM), DNA Pol γ (IC50 > 100 µM), POLRMT (human mitochondrial RNA polymerase) (IC50 > 100 µM), RNA Pol II (IC50 > 100 µM) |
| Selectivity outside target family | Cerep panel (80 targets) with closest off-targets (IC50 [µM]): (h) ADORA3 (49), BZDp (h) (0.56), Cl- channel (GABA-gated) (7.4), PTGER4 (h) (1.3), TACR1(h) (8.4), TACR2 (h) (6.8) |
| On target cell activity for cell-based targets: goal is < 1 µM IC50/EC50 | Surpasses criterion:Inhibits the replication of RSV A (median (10 strains) EC50 = 0.18 nM) and RSV B (median (6 strains), EC50= 0.1 nM) clinical isolates and laboratory strains similarly by RT-qPCR (HeLa cells) Inhibition of an RSV sub-genomic replicon (APC-126 cells): EC50 = 0.15 nM, CC50 = 11 μM (n = 24) Inhibition of RSV A2 replication in a reporter assay (HeLa cell-based using rgRSV224 reporter strain): EC50 = 0.78 nM; CC50 = 27 μM, n = 23 |
| Control compound (100 times less potent than the probe) | JNJ-6643: Inhibition of RNA synthesis activity of the recombinant RSV L + P in a biotinylated primer extension Flashplate assay: IC50 = 530 nM (n = 1); Inhibition of RSV A2 replication in a reporter assay (HeLa cell-based using rgRSV224 reporter strain): EC50 = 150 nM |